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You are an expert at summarizing long articles. Proceed to summarize the following text:
branch retinal vein occlusion ( brvo ) is a common sight - threatening retinal vascular disorder , in which macular edema is the main cause of visual impairment . retinal ischemia after vascular occlusion can cause both vitreous and aqueous vascular endothelial growth factor ( vegf ) elevation [ 2 , 3 ] . increased vegf results in higher vascular permeability and associated macular edema in patients with brvo . intravitreal injections of anti - vegf , such as bevacizumab ( avastin , genentech inc . , south san francisco , ca , usa ) and aflibercept ( eylea , regeneron pharmaceuticals , inc . , tarrytown , ny , usa , and bayer pharma ag , berlin , germany ) , can effectively lower intraocular level of vegf and reduce vascular permeability related to macular edema in brvo [ 414 ] . herein , we performed a head - to - head comparison of efficacy after intravitreal bevacizumab and aflibercept treatments for macular edema following brvo . to our knowledge , this is the first paper associated with the issue after reviewing the literature . we conformed to the declaration of helsinki to accomplish this study . from august 2012 to september 2015 , 104 eyes of 104 patients with macular edema secondary to perfused brvo were retrospectively reviewed . the patients did not have prior trauma , intravitreal injections , retinal or macular laser , or ocular surgery except uneventful phacoemulsification . no patient had history of thromboembolic events , known coagulation abnormalities or current use of anticoagulative medication other than aspirin , or other major systemic diseases . all the patients did not have any intravitreal anti - vegf , macular laser , or other associated treatments . the patients with perfused brvo were diagnosed with retinal nonperfusion area less than 10 disc diameters by fundus fluorescein angiography ( trc - nw7sf , topcon , tokyo , japan ) . macular edema was defined as macular leakage on fundus fluorescein angiography and central foveal thickness ( cft ) more than 300 m detected by spectral - domain optical coherence tomography ( rtvue , optovue , san francisco , usa ) with macular pathologies including cystoid changes , diffuse thickening , and submacular fluid , using 6 radial line scans through the fovea in all patients . baseline best - corrected visual acuity ( bcva ) in snellen chart ( converting into logmar and edtrs letters for statistical comparison ) , intraocular pressure via pneumotonometer ( ct-80 , topcon , tokyo , japan ) , and biomicroscope of anterior segment were examined in all the patients . once the patients were diagnosed with macular edema secondary to brvo , intravitreal bevacizumab or aflibercept was administered within one week without accompanying macular laser or future laser rescue . the procedures were performed at far eastern memorial hospital by one surgeon ( wang jk ) . after detailed explanation of risks , benefits , and off - label use of these medications , all the participants signed the informed consents before the intravitreal injections . between august 2012 and february 2014 , 54 consecutive cases received prn intravitreal bevacizumab 1.25 mg/0.05 ml treatment . between march 2014 and september 2015 , 50 consecutive patients were treated by prn intravitreal aflibercept 2 mg/0.05 ml . following topical anesthesia and disinfection of eyelid and conjunctiva , aflibercept or bevacizumab was injected into the vitreous cavity using a 30-gauge needle inserted through the inferotemporal pars plana , 3.5 mm posterior to the limbus . the patient was instructed to instill one drop of 0.3% norfloxacin into the injected eye four times daily for one week . all the patients were followed up monthly for at least 12 months , with anterior segment and fundus examination and bcva , cft , and intraocular pressure measurement . the follow - up sd - oct scans used the baseline scan as a reference . retreatment was based on findings of optical coherence tomography including cft more than 300 m , or there were persistent or recurrent macular cysts or submacular fluid that affected the visual acuity even if cft is less than 300 m . the intragroup changes in cft and bcva were compared with wilcoxon signed rank test and the between - group numerical difference compared with wilcoxon rank sum test . one month after the first intravitreal aflibercept , mean bcva significantly improved from baseline 0.77 logmar ( equal to 46.1 letters ) to 0.43 logmar ( 62.3 letters ) ( p = 0.0007 ) ( figure 1 ) . the baseline mean cft was 470.2 m , which significantly decreases 1 month after the first aflibercept injection to 254.2 m ( p < 0.0001 ) ( figure 2 ) . from month 2 to month 11 , all time points in cft and bcva were significantly different from baseline data ( p < 0.01 ) ( figures 1 and 2 ) . at month 12 in aflibercept group , mean bcva significantly improved to 0.29 logmar ( 70.2 letters ) , significantly better than baseline bcva ( p < 0.0001 ) , with mean change of 0.48 logmar ( + 24.1 letters ) ( figure 1 ) . at month 12 in aflibercept group , mean cft significantly decreased to 241.9 m ( p < 0.0001 ) , with mean change of 228.3 m ( figure 2 ) . the baseline mean bcva was 0.72 logmar ( 48.9 letters ) , which significantly improved 1 month after the first bevacizumab injection to 0.50 logmar ( 60.1 letters ) ( p = 0.0002 ) ( figure 1 ) . the mean cft 1 month after the first intravitreal bevacizumab ( 268.3 m ) significantly improved compared with the mean cft at baseline ( 459.4 m ) ( p < 0.0001 ) ( figure 2 ) . from month 2 to month 11 , all time points in cft and bcva were significantly different from baseline data ( p < 0.01 ) ( figures 1 and 2 ) . at month 12 in bevacizumab group , mean bcva significantly improved to 0.31 logmar ( 69.5 letters ) , significantly better than baseline bcva ( p < 0.0001 ) , with mean change of 0.41 logmar ( + 21.1 letters ) ( figure 1 ) . at month 12 in bevacizumab group , mean cft significantly decreased to 252.7 m ( p < 0.0001 ) , with mean change of 212.8 m ( figure 2 ) . there was no statistical difference between two groups in baseline data , including age , gender , lens status , incidence of patients with diabetes , presenting bcva and cft , and duration of symptoms ( p > 0.05 ) ( table 1 ) . at all time points from month 2 to month 11 , cft and bcva were not significantly different between aflibercept and bevacizumab groups ( p > 0.05 ) . there was also no significant difference between two groups in final anatomical and functional outcome at month 12 , including final bcva , mean visual gains , final cft , and mean cft changes ( p > 0.05 ) ( table 2 ) . nearly two - thirds of the patients in both groups had final bcva equal to or more than 20/40 and visual gains more than 3 lines from baseline to month 12 ( table 2 ) . only less than 10% of the patients in both groups lost bcva equal to or more than one line 12 months after the first injection ( table 2 ) . mean injection number of aflibercept was 2.12 , comparable with that of bevacizumab ( 2.22 ) during 12-month period ( p = 0.11 ) . in aflibercept group , 22 ( 44% ) eyes required no additional injection except the baseline injections . in bevacizumab group , 22 ( 40.7% ) eyes required no additional injection except the baseline injections . there were no episodes of endophthalmitis , retinal detachment , vitreous hemorrhage , or elevated intraocular pressure . the most common side effect was local hyperemia or subconjunctival hemorrhage at the site of injection . macular grid laser photocoagulation was the standard of care for macular edema in perfused brvo according to results of the branch vein occlusion study . however , the visual improvement following macular laser was limited , with mean improvement of 1.33 lines of vision at the 3-year primary end point . anti - vegf intraocular injection has been shown to be a new promising treatment modality , which results in noticeable functional and anatomical improvement . bevacizumab is a full - length recombinant humanized monoclonal antibody directed against the vegf - a . intravitreal bevacizumab is employed to lower the intraocular vegf level , effectively reducing macular edema in patients with brvo . the pan american collaborative retina study group used intravitreal bevacizumab in prn regimen to treat 63 patients with macular edema following brvo . the 2-year results demonstrated that bevacizumab 1.25 mg injection resulted in a gain of 3.8 lines . decreased macular thickness was found , without accompanying serious ocular and systemic adverse events . hikichi and coauthors performed prn intravitreal bevacizumab for 89 eyes with macular edema in brvo . the 2-year results showed that injections resulted in visual gains of 0.30 logmar and subsided macular edema . ahn and colleagues performed prn intravitreal bevacizumab for 69 eyes and three monthly prn injections for 26 eyes with macular edema in brvo . the 6-month results revealed functional and anatomical improvement in both groups , and there was no significant difference between two groups in final visual acuity . injection number was far less in prn only regimen ( mean 1.8 ) than in three - loading and prn group ( mean 3.4 ) . intravitreal bevacizumab demonstrated superior ability than conventional macular grid laser for treating macular edema in brvo . russo and associates compared the efficacy of intravitreal bevacizumab and macular grid laser for patients with macular edema secondary to brvo in a randomized study . the one - year outcome demonstrated that bevacizumab injections had better visual gains and reduction of macular thickness than macular laser . leitritz and coauthors performed a prospective crossover study to compare the efficacy of intravitreal bevacizumab and macular grid laser for patients with macular edema associated with brvo . the one - year outcome demonstrated that bevacizumab injections had superior visual improvement than macular laser . hayashi and colleagues compared the efficacy of intravitreal bevacizumab alone and injections combined with macular grid laser for patients with macular edema in brvo . the one - year outcome demonstrated that macular laser did not have adding effect for bevacizumab injections , neither prolonging the bevacizumab effect nor bettering visual outcome in combined treatment . in this study , the patients with macular edema associated with brvo gained 4.1 lines after one - year prn intravitreal bevacizumab injections . the visual results were similar to prior studies , from 2.9- to 3.8-line visual improvement after one - year prn bevacizumab treatment [ 5 , 7 , 13 , 14 ] . there were 52.8% of patients with visual gain equal to or more than 3 lines at month 12 , comparable to the outcome of previous trials ( from 48% to 80% ) [ 5 , 7 , 10 , 13 , 14 ] . the percentage of bevacizumab - treated patients who had at least 20/40 in bcva was 66.6% at month 12 , similar to results of prior studies after one - year prn bevacizumab management ( from 55 to 67% ) [ 5 , 10 ] . the mean injection number was 2.22 in one year in this study , comparable to the results of preceding articles requiring 1.9 to 2.6 injections of prn bevacizumab within one year [ 5 , 7 , 13 ] . aflibercept is a decoy receptor fusion protein , composed of the second domain of human vegf receptor 1 and the third domain of vegf receptor 2 , which are fused to the fc domain of human igg1 . aflibercept can downregulate vegf - a , vegf - b , and placental growth factor , which are synergistic for pathologic angiogenesis . intravitreal aflibercept was noted to lower the intraocular vegf level in patients with neovascular age - related macular degeneration . the vibrant study , a randomized controlled trial , was conducted in north america and japan . the study demonstrated the efficacy of intravitreal aflibercept 2 mg over the macular grid laser for 183 patients with macular edema associated with brvo . the 6-month results showed that the aflibercept group gained mean of 17.0 letters , significantly better than the laser group having only mean 6.9-letter improvement . the proportion of eyes that gained more than 15 letters from baseline at week 24 was 52.7% . decrease of macular thickness was more prominent in the aflibercept group than in the laser group , without accompanying serious ocular and systemic adverse events . in this study , we had 19.2-letter visual gain in the first 6 months of prn aflibercept injections , compared to the results of vibrant study which had 17-letter improvement . there were 61.1% of patients having at least 15-letter gain after 6-month aflibercept treatment , similar to 52.7% in vibrant study . we only used mean 1.33 aflibercept injections in the first 6 months , compared to 6 injections in the vibrant study . the binding affinity of vegf for this drug is higher than for ranibizumab and bevacizumab . aflibercept displays a prolonged vegf inhibition in comparison with the other vegf - antagonists ranibizumab and bevacizumab in retinal pigment epithelium / choroid organ cultures . aflibercept also demonstrates better safety than bevacizumab in vitro associated with mitochondria exocytosis . in a randomized controlled trial , the authors concluded that aflibercept was more effective than bevacizumab for visual improvement in diabetic patients with poor baseline vision , but comparable efficacy in both pharmaceutical agents for better baseline vision . both medications showed similar safety profiles and injection number during one - year follow - up . to our knowledge , there is no publication comparing clinical outcome of aflibercept and bevacizumab for patients with macular edema secondary to brvo . in the present study , aflibercept had comparable efficacy , injection number , and safety as bevacizumab for patients with macular edema associated with brvo . a large prospective randomized study can further confirm the difference to treat macular edema in brvo in longer follow - up period . in summary , intravitreal aflibercept and bevacizumab had similar efficacy and duration of treatment for macular edema associated with brvo during 12-month period . | fifty - two eyes of 52 patients with treatment - nave macular edema associated with perfused branch retinal vein occlusion were retrospectively reviewed .
twenty - seven cases received prn intravitreal bevacizumab , and 25 cases were treated by prn intravitreal aflibercept with monthly follow - ups for 12 months .
both aflibercept and bevacizumab were effective in reduction of macular thickness and improvement of visual acuity for the participants .
both antivascular endothelial growth factor agents had similar efficacy and duration of treatment for these eyes with macular edema secondary to branch retinal vein occlusion during a 12-month period .
no serious systemic or ocular adverse events were reported . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
multiple sclerosis is a chronic progressive disease resulting in damage to the myelin sheath and nerves of the central nervous system ( brain , spinal cord , and optic nerves ) . it is thought to be an autoimmune disorder , is usually diagnosed during early to late adulthood , and affects 23 times more women than men.1 approximately 350,000 americans are diagnosed with multiple sclerosis,2 although the number may be higher because symptoms generally do not appear until the disease has progressed . symptoms and their severity and duration vary greatly among patients . symptoms may include extreme fatigue , difficulty with memory and concentration , vision loss , blurred or double vision , numbness , tingling , loss of balance and coordination , tremors , stiffness , slurred speech , and bladder / bowel dysfunction.1,2 multiple sclerosis is rarely fatal , and the majority of patients do not become severely disabled . the most common form of multiple sclerosis is relapsing - remitting ( 85% of patients ) , where patients experience discrete and acute exacerbations or flareups of symptoms , followed by partial or complete remission from symptoms or disease activity.2 current treatments for multiple sclerosis include disease - modifying drugs that reduce the frequency and intensity of exacerbations , and drugs that treat the symptoms or the exacerbation itself.1,2 the standard treatment for severe exacerbations is a 35-day course of intravenous methylprednisolone . if intravenous corticosteroids can not be administered , are unavailable , are not effective , or are not well tolerated , adrenocorticotropic hormone ( acth ) is recommended as an alternative option.2 acth has demonstrated efficacy in the treatment of acute exacerbations of multiple sclerosis in controlled clinical trials , and is approved by the us food and drug administration for treatment for acute exacerbations of multiple sclerosis.35 in all trials of acth in multiple sclerosis , the drug was administered as intramuscular injections for at least two weeks.35 based on these trials , the protocol currently approved by the food and drug administration prescribes daily doses of acth ranging from 80 to 120 u , given as intramuscular or subcutaneous injections for 1421 days to treat acute exacerbations of multiple sclerosis.6 however , a shorter protocol may be effective . one clinical trial of acth , for example , found symptoms significantly improved compared with placebo after seven days of treatment.4 similarly , methylprednisolone , the standard therapy for treatment of exacerbations , was originally given as a longer course of treatment before it was reported that a shorter five - day course was equally effective.7 in many instances , subcutaneous injections may offer a viable route of administration for drugs traditionally administered intramuscularly . it has recently been shown that subcutaneous and intramuscular injections of acth are bioequivalent in stimulating cortisol levels in healthy volunteers.8 the subcutaneous injection is easier to administer for the following reasons , ie , a larger injectable area , no need to identify anatomical landmarks , easier to administer in emaciated patients , and a superior safety profile.9 because the gel formulation of natural acth has been approved to be dosed subcutaneously for acute exacerbations of multiple sclerosis and for other indications , and shorter courses of treatment are suggested by the literature , we conducted a pilot study to test a shorter , self - administered five - day injection regimen and to compare the effectiveness , safety , and patient satisfaction of these two routes of acth administration in patients with an acute exacerbation of multiple sclerosis . study participants were men and women aged 18 years with a clinical diagnosis of relapsing - remitting , secondary progressive , or progressive relapsing multiple sclerosis , who presented at an outpatient clinic with an acute exacerbation within 24120 hours of symptom onset . while discussing treatment options , participants were given the option to be screened for the study by the investigator . patients who were on other investigative drugs , chemotherapeutic agents , the humanized monoclonal antibody , natalizumab ( tysabri , biogen idec , cambridge , ma ) , or had a history of a clinically significant infection within 30 days prior to study entry , were excluded . women who were nursing , pregnant , or planning to become pregnant , and those of childbearing potential ( ie , those who were not postmenopausal for at least one year ) who were unwilling to use effective contraception were excluded . concomitant use of certain medications ( acetylsalicylic acid , anticholinesterases , oral anticoagulants , chemotherapeutic agents , cholestyramine , systemic corticosteroids , cyclosporine , digitalis glycosides , ephedrine , ketoconazole , macrolide antibiotics , phenobarbital , phenytoin , rifampicin , thalidomide , or vaccines ) was not permitted during the study . informed consent was signed and collected by either the investigating physician or by staff after the patient had the opportunity to learn and ask questions about the study . this was a two - week prospective , randomized , open - label , single - center , pilot study with two treatment arms ( see figure 1 ) . a five - day treatment course was chosen because the standard corticosteroid intravenous treatment is 35 days . the dose of 80 u / day is the lower end of the approved dose range for acute exacerbations of multiple sclerosis . since this is a self - administration study , we chose the lower dose ( 80 u / day ) because each multiuse vial of acth holds 80 u / ml in 5 ml gel volume , and we believe this offers the most convenient and cost - effective dosing for the patient without the need to discard any unused drug at the end of treatment . after clinical evaluation and informed consent was obtained at the first visit on day 1 , subjects were randomized using a random - number generator in a 1:1 scheme to receive acth gel ( hp acthar gel [ repository corticotrophin injection ] , questcor pharmaceuticals , union city , ca)6 dosed at 80 u / day , given as either an intramuscular or subcutaneous injection on days 15 of the study . participants were trained to self - administer the injection by study personnel during the first visit . during the study period , both subjective and objective measures of drug response were used in the study ( table 1 ) . the primary outcome measure was the patient global impression of change ( pgi - c ) , a self - administered daily change scale , with scores ranging from 1 ( very much improved ) to 7 ( very much worse ) used to record a patient s impression about the effectiveness of the treatment.10 the measure of symptom alleviation was the final time point score of the pgi - c on day 14 ; patients reporting change scores of 1 and 2 were considered to be treatment responders . a patient visual analog scale was also used to quantify symptoms associated with their acute exacerbation of multiple sclerosis . each day during the course of the study , participants marked a linear scale that ranged from 1 ( no acute exacerbation of symptoms ) to 10 ( worst imaginable acute exacerbation of symptoms ) . in addition , we assessed patient satisfaction with the self - administration technique using a drug attitude questionnaire developed for this study . the clinical impression of treatment performance with intramuscular or subcutaneous acth gel was quantified using a clinical global impression of change ( cgi - c ) questionnaire.11 disability was quantified using the kurtzke expanded disability status scale ( edss ) at each study visit on days 1 , 7 , and 14.12 objective measures of drug performance included documentation of results of a timed walk13 and the nine - hole peg test for finger dexterity,14 which were measured pretreatment ( on day 1 ) and on days 7 and 14 after treatment . medical history was elicited , and physical examination , including vital signs , was performed on all participants at study entry ( day 1 ) and at the two follow - up visits ( days 7 and 14 ) . serious adverse events were classified as those that required inpatient hospitalization , were life - threatening , or resulted in significant disability or death . all participants receiving at least one dose of medication were included in the safety analysis ; subjects receiving at least one dose of study medication and completing one pgi - c scale were included in the treatment performance analysis . study participants were men and women aged 18 years with a clinical diagnosis of relapsing - remitting , secondary progressive , or progressive relapsing multiple sclerosis , who presented at an outpatient clinic with an acute exacerbation within 24120 hours of symptom onset . while discussing treatment options , participants were given the option to be screened for the study by the investigator . patients who were on other investigative drugs , chemotherapeutic agents , the humanized monoclonal antibody , natalizumab ( tysabri , biogen idec , cambridge , ma ) , or had a history of a clinically significant infection within 30 days prior to study entry , were excluded . women who were nursing , pregnant , or planning to become pregnant , and those of childbearing potential ( ie , those who were not postmenopausal for at least one year ) who were unwilling to use effective contraception were excluded . concomitant use of certain medications ( acetylsalicylic acid , anticholinesterases , oral anticoagulants , chemotherapeutic agents , cholestyramine , systemic corticosteroids , cyclosporine , digitalis glycosides , ephedrine , ketoconazole , macrolide antibiotics , phenobarbital , phenytoin , rifampicin , thalidomide , or vaccines ) was not permitted during the study . informed consent was signed and collected by either the investigating physician or by staff after the patient had the opportunity to learn and ask questions about the study . this was a two - week prospective , randomized , open - label , single - center , pilot study with two treatment arms ( see figure 1 ) . a five - day treatment course was chosen because the standard corticosteroid intravenous treatment is 35 days . the dose of 80 u / day is the lower end of the approved dose range for acute exacerbations of multiple sclerosis . since this is a self - administration study , we chose the lower dose ( 80 u / day ) because each multiuse vial of acth holds 80 u / ml in 5 ml gel volume , and we believe this offers the most convenient and cost - effective dosing for the patient without the need to discard any unused drug at the end of treatment . after clinical evaluation and informed consent was obtained at the first visit on day 1 , subjects were randomized using a random - number generator in a 1:1 scheme to receive acth gel ( hp acthar gel [ repository corticotrophin injection ] , questcor pharmaceuticals , union city , ca)6 dosed at 80 u / day , given as either an intramuscular or subcutaneous injection on days 15 of the study . participants were trained to self - administer the injection by study personnel during the first visit . during the study period , both subjective and objective measures of drug response were used in the study ( table 1 ) . the primary outcome measure was the patient global impression of change ( pgi - c ) , a self - administered daily change scale , with scores ranging from 1 ( very much improved ) to 7 ( very much worse ) used to record a patient s impression about the effectiveness of the treatment.10 the measure of symptom alleviation was the final time point score of the pgi - c on day 14 ; patients reporting change scores of 1 and 2 were considered to be treatment responders . a patient visual analog scale was also used to quantify symptoms associated with their acute exacerbation of multiple sclerosis . each day during the course of the study , participants marked a linear scale that ranged from 1 ( no acute exacerbation of symptoms ) to 10 ( worst imaginable acute exacerbation of symptoms ) . in addition , we assessed patient satisfaction with the self - administration technique using a drug attitude questionnaire developed for this study . the clinical impression of treatment performance with intramuscular or subcutaneous acth gel was quantified using a clinical global impression of change ( cgi - c ) questionnaire.11 disability was quantified using the kurtzke expanded disability status scale ( edss ) at each study visit on days 1 , 7 , and 14.12 objective measures of drug performance included documentation of results of a timed walk13 and the nine - hole peg test for finger dexterity,14 which were measured pretreatment ( on day 1 ) and on days 7 and 14 after treatment . medical history was elicited , and physical examination , including vital signs , was performed on all participants at study entry ( day 1 ) and at the two follow - up visits ( days 7 and 14 ) . serious adverse events were classified as those that required inpatient hospitalization , were life - threatening , or resulted in significant disability or death . both subjective and objective measures of drug response were used in the study ( table 1 ) . the primary outcome measure was the patient global impression of change ( pgi - c ) , a self - administered daily change scale , with scores ranging from 1 ( very much improved ) to 7 ( very much worse ) used to record a patient s impression about the effectiveness of the treatment.10 the measure of symptom alleviation was the final time point score of the pgi - c on day 14 ; patients reporting change scores of 1 and 2 were considered to be treatment responders . a patient visual analog scale was also used to quantify symptoms associated with their acute exacerbation of multiple sclerosis . each day during the course of the study , participants marked a linear scale that ranged from 1 ( no acute exacerbation of symptoms ) to 10 ( worst imaginable acute exacerbation of symptoms ) . in addition , we assessed patient satisfaction with the self - administration technique using a drug attitude questionnaire developed for this study . the clinical impression of treatment performance with intramuscular or subcutaneous acth gel was quantified using a clinical global impression of change ( cgi - c ) questionnaire.11 disability was quantified using the kurtzke expanded disability status scale ( edss ) at each study visit on days 1 , 7 , and 14.12 objective measures of drug performance included documentation of results of a timed walk13 and the nine - hole peg test for finger dexterity,14 which were measured pretreatment ( on day 1 ) and on days 7 and 14 after treatment . medical history was elicited , and physical examination , including vital signs , was performed on all participants at study entry ( day 1 ) and at the two follow - up visits ( days 7 and 14 ) . serious adverse events were classified as those that required inpatient hospitalization , were life - threatening , or resulted in significant disability or death . all participants receiving at least one dose of medication were included in the safety analysis ; subjects receiving at least one dose of study medication and completing one pgi - c scale were included in the treatment performance analysis . a total of 20 patients were enrolled in this pilot study , comprising 17 women ( 85% ) and three men ( 15% ) . all participants were caucasian , and aged 2559 ( mean 39.5 9.9 ) years . the characteristics of the two groups at baseline were similar and are detailed in table 2 . pgi - c scores were used to determine the percentage of responders versus nonresponders in the intramuscular and subcutaneous groups at days 7 and 14 ( figure 2 ) . in both groups , the percentage of responders increased between days 7 and 14 , with 77.7% ( 7/9 ) of the intramuscular group and 44.4% ( 4/9 ) of the subcutaneous group responding by day 14 . although there were more intramuscular responders at days 7 and 14 , the final time point scores for the two methods were not significantly different ( p = 0.335 , two - sided fisher s exact test , post hoc comparison ) . the cgi - c scores were consistent with the pgi - c scores ( figure 2 ) . in both the cgi - c and pgi - c assessments , interestingly , the clinician assessment scored more responders , indicating more symptom improvement than the patient assessments . the raw data scores for the pgi - c and cgi - c assessments are summarized in tables 3a and 3b . overall , the data indicate an improvement in symptom exacerbation . between days 7 and 14 the clinician assessment of very much improved or much improved increased to 100% for the intramuscular patients and 77.7% for the subcutaneous patients . these data show that patients responded to a shorter , five - day course of treatment , and that although the intramuscular route seemed more effective than the subcutaneous route , the difference was not statistically significant in this small sample . the visual analog scale data ( figure 3 ) support the pgi - c data showing symptom improvement by day 14 in both the intramuscular and subcutaneous treatment groups . the intramuscular treatment group showed a numerically greater response than the subcutaneous treatment group ( + 3.8 units versus + 1.4 units , respectively ) . the final improvement scores ( day 14 ) for the two groups were very similar , ie , 2.5 ( intramuscular ) and 2.9 ( subcutaneous ) , on a scale of 010 , where 0 = no exacerbation symptoms and 10 = worst exacerbation symptoms possible . the objective measures of function from a mean timed walk and the nine - hole peg test are presented in table 4 . there is some improvement seen in the mean timed walk and nine - hole peg test scores , although neither was statistically or clinically significant . this may be expected , because baseline scores ( day 1 ) were already in a highly functional range . the edss scores , measuring level of disability , are reported in table 5 . as for the other assessments , there was some ability improvement in both the intramuscular and subcutaneous treatment groups from visit 1 ( baseline ) to visit 3 ( day 14 ) . however , as in the objective measures of function , the baseline edss scores are already highly functional . the median scores at baseline in this patient population ranged from 2.3 to 2.8 , indicating mild to moderate disability . in all , the subjective assessments of a shorter five - day course of treatment showed good patient response and tolerability , regardless of route of administration . in the objective assessments of function , patients generally improved after treatment . however , the low initial baseline scores suggest that this cohort of patients retained a high level of ability at study enrollment . patients from both treatment groups rated the study treatment high for satisfaction , convenience , and tolerability on the drug attitude questionnaire on day 14 ( table 6 ) . of the seven patients ( 70% ) in the intramuscular treatment group who reported experiencing pain associated with injection , all but one patient described the pain as mild . in contrast , only 33.3% of patients in the subcutaneous group reported injection site pain , all of whom described the pain as mild . of note , 95% of study patients who completed the questionnaire ( n = 18 ) reported that an injection at home was significantly more convenient than infusion in the clinic . further , 90% of patients in the intramuscular group and 67% in the subcutaneous group said they would request the same intervention for any future acute exacerbations . a total of 20 patients were enrolled in this pilot study , comprising 17 women ( 85% ) and three men ( 15% ) . all participants were caucasian , and aged 2559 ( mean 39.5 9.9 ) years . the characteristics of the two groups at baseline were similar and are detailed in table 2 . pgi - c scores were used to determine the percentage of responders versus nonresponders in the intramuscular and subcutaneous groups at days 7 and 14 ( figure 2 ) . in both groups , the percentage of responders increased between days 7 and 14 , with 77.7% ( 7/9 ) of the intramuscular group and 44.4% ( 4/9 ) of the subcutaneous group responding by day 14 . although there were more intramuscular responders at days 7 and 14 , the final time point scores for the two methods were not significantly different ( p = 0.335 , two - sided fisher s exact test , post hoc comparison ) . the cgi - c scores were consistent with the pgi - c scores ( figure 2 ) . in both the cgi - c and pgi - c assessments , interestingly , the clinician assessment scored more responders , indicating more symptom improvement than the patient assessments . the raw data scores for the pgi - c and cgi - c assessments are summarized in tables 3a and 3b . overall , the data indicate an improvement in symptom exacerbation . between days 7 and 14 , the clinician assessment of very much improved or much improved increased to 100% for the intramuscular patients and 77.7% for the subcutaneous patients . these data show that patients responded to a shorter , five - day course of treatment , and that although the intramuscular route seemed more effective than the subcutaneous route , the difference was not statistically significant in this small sample . the visual analog scale data ( figure 3 ) support the pgi - c data showing symptom improvement by day 14 in both the intramuscular and subcutaneous treatment groups . the intramuscular treatment group showed a numerically greater response than the subcutaneous treatment group ( + 3.8 units versus + 1.4 units , respectively ) . the final improvement scores ( day 14 ) for the two groups were very similar , ie , 2.5 ( intramuscular ) and 2.9 ( subcutaneous ) , on a scale of 010 , where 0 = no exacerbation symptoms and 10 = worst exacerbation symptoms possible . the objective measures of function from a mean timed walk and the nine - hole peg test are presented in table 4 . there is some improvement seen in the mean timed walk and nine - hole peg test scores , although neither was statistically or clinically significant . this may be expected , because baseline scores ( day 1 ) were already in a highly functional range . the edss scores , measuring level of disability , are reported in table 5 . as for the other assessments , there was some ability improvement in both the intramuscular and subcutaneous treatment groups from visit 1 ( baseline ) to visit 3 ( day 14 ) . however , as in the objective measures of function , the baseline edss scores are already highly functional . the median scores at baseline in this patient population ranged from 2.3 to 2.8 , indicating mild to moderate disability . in all , the subjective assessments of a shorter five - day course of treatment showed good patient response and tolerability , regardless of route of administration . in the objective assessments of function , patients generally improved after treatment . however , the low initial baseline scores suggest that this cohort of patients retained a high level of ability at study enrollment . patients from both treatment groups rated the study treatment high for satisfaction , convenience , and tolerability on the drug attitude questionnaire on day 14 ( table 6 ) . of the seven patients ( 70% ) in the intramuscular treatment group who reported experiencing pain associated with injection , all but one patient described the pain as mild . in contrast , only 33.3% of patients in the subcutaneous group reported injection site pain , all of whom described the pain as mild . of note , 95% of study patients who completed the questionnaire ( n = 18 ) reported that an injection at home was significantly more convenient than infusion in the clinic . further , 90% of patients in the intramuscular group and 67% in the subcutaneous group said they would request the same intervention for any future acute exacerbations . nineteen patients completed the study , and one patient in the subcutaneous group withdrew , citing lack of improvement . severe adverse events were reported in one intramuscular subject who complained of localized pain and numbness of the left hand and leg on one occasion and of the back and leg on another occasion . a recent comprehensive review of steroid hormone therapy for exacerbations of multiple sclerosis suggested that a short duration ( five days ) of methylprednisolone and a long duration ( 14 days ) of acth appeared to have significant differences in treatment outcomes.7 we wanted to know if a short course of treatment ( five days ) using acth could still alleviate symptoms of an acute exacerbation of multiple sclerosis , and if patients liked the acth treatment protocol . in this small pilot study , we examined , for the first time , if a shorter , self - administered five - day course of acth therapy would show a treatment response . in the literature , clinical trial data on the use of acth in acute exacerbations of multiple sclerosis come from two placebo - controlled , double - blind studies and one double - blind comparator trial ( acth versus methylprednisolone ) involving a total of 298 subjects across all studies.35 in a study by miller et al , acth gel was dosed 60 u intramuscularly twice daily in the first week , 40 u twice daily in the second week , and then 60 u , 40 u , and 20 u on days 2 , 4 , and 6 of the third week.3 rose et al dosed acth at 40 u twice daily intramuscularly for seven days , tapered to 20 u twice daily for four days , and then 20 u daily for three days in their study.4 thompson et al compared a regimen of intravenous methylprednisolone dosed 1 g / day for three days , with a 14-day course of acth given intramuscularly as follows : 80 u / day for seven days , 40 u / day for four days , and 20 u / day for three days.5 a significant treatment effect of acth was demonstrated in the placebo - controlled studies , and no difference with methylprednisolone was reported in the small comparative study . mild side effects were reported with acth and not methylprednisolone in the comparative study ; it would be interesting to know if a shorter course of acth treatment would result in fewer side effects . no large studies have directly compared methylprednisolone with acth in the treatment of acute exacerbations of multiple sclerosis . the gel formulation of acth is approved by the food and drug administration for several different indications , given either intramuscularly or subcutaneously.6 there are limited data available on the duration , dosing , and route of administration of acth gel in the treatment of acute exacerbations of multiple sclerosis . this pilot study is the first attempt to compare acth gel therapy administered either intramuscularly or subcutaneously . we found that patients in both the intramuscular and subcutaneous treatment groups reported improvement in their condition after a five - day course of treatment with acth gel dosed at 80 u / day . improvement was also observed on the cgi - c scale , indicating that clinicians also observed meaningful improvements . because of the small sample size , random assignment to treatment arms may not have eliminated bias entirely . in addition , because there was no control arm , it is unknown what percentage of exacerbations would have resolved spontaneously . it was difficult to measure improvement objectively in this study , because the patients demonstrated good function at enrollment and the study period was short ( 14 days ) . lastly , while the drug attitude questionnaire showed general patient satisfaction with the acth treatment , the questionnaire failed to determine the sources of satisfaction , such as drug efficacy , tolerability , convenience , or cost . it will be important to address these issues in larger trials based upon the results of this small pilot study . if confirmed , a shorter protocol for treatment of acute exacerbations with acth could have implications for disease management and clinical practice as it moves treatment from the clinic and hospital setting to the home , giving patients more control over their disease and improving their quality of life . shortening the treatment time may lessen the cost of using acth while maintaining clinical efficacy and increasing convenience to the patient , and changing the injection route from intramuscular to subcutaneous may minimize the pain and difficulty of treatment . together , these elements of our modified protocol may decrease the burden of disease for the patient with multiple sclerosis , improve quality of life , and give physicians more patient - friendly options when treating acute exacerbations . more clinical trials are needed to examine new dosing protocols with our old drugs as well as exploring drug combinations with the newer drugs available . however , the expense and time required for large , placebo - controlled studies can make this impractical . small pilot studies may have value in such cases , where changes in drug protocol or special treatment protocols for patient subpopulations may be evaluated qualitatively and shared with the clinical community . in this way our study in 20 patients suggests that a five - day course of patient - administered acth gel therapy relieves symptoms of acute exacerbations of multiple sclerosis when administered either as intramuscular or subcutaneous injections . in most cases , the patient s lifestyle may be less disrupted by a self - administered injection compared with an infusion in the clinic , giving the patient more independence and control of their disease management . in addition , a shorter five - day course of treatment would reduce costs to the patient , in terms of drug cost , time , and the likelihood of adverse events . these results warrant further study of short - term , self - administered acth therapy for acute exacerbations of multiple sclerosis . larger , placebo - controlled studies are needed to determine the optimal dose of acth gel , duration of treatment , and route of administration , as well as its role compared with steroid therapy . | background : despite over 50 years of experience with adrenocorticotropic hormone ( acth ) as a treatment for acute exacerbations of multiple sclerosis , there have been no trials examining the options of the 23-week dosing regimen or intramuscular injection protocol used in the original trials . at our clinic , we performed a small , prospective , randomized pilot study to examine the efficacy and safety of , and patient satisfaction with , a short ( five - day ) self - administered acth dosing protocol for exacerbations of multiple sclerosis , and to compare the subcutaneous and intramuscular routes of administration.methods:patients for this study were recruited from an outpatient treatment clinic .
each patient self - administered natural acth gel 80 u / day by subcutaneous or intramuscular injection for five consecutive days and was evaluated at baseline and on days 7 and 14 .
patient feedback was collected using the patient global impression of change ( pgi - c , the primary efficacy measure ) , a patient global visual analog scale , the expanded disability status scale , a timed walk , the nine - hole peg test , and the clinical global impression of change.results:of the 20 enrolled patients ( mean age 39.5 years ) , 19 completed the study . on day 14
, 61.1% of patients ( 11 of 18 with day 14 scores ) were treatment responders , and rated their condition as very much improved or
much improved on the pgi - c .
the intramuscular group had numerically more responders , but there was no significant difference in the proportion of responders between the intramuscular and subcutaneous groups at day 14 ( p = 0.3 ) .
the intramuscular route of injection was associated with more injection site pain than the subcutaneous route.conclusion:a shorter five - day course of intramuscular or subcutaneous acth gel may improve symptoms associated with acute exacerbations of multiple sclerosis .
larger studies with standard of care controls are needed to confirm whether this shorter course of intramuscular or subcutaneous acth gel is effective and could potentially be substituted for the standard 14-day treatment . |
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high - frequency oscillation ventilators were initially designed for neonatal application . with the recognition of the role of ventilator - induced lung injury in the morbidity and mortality of patients with acute lung injury , there has also been increasing interest in the use of high - frequency oscillatory ventilation ( hfov ) in adult patients , since it theoretically offers , by design , an ideal mode for lung protection . the need to suppress patients ' spontaneous breathing activity with heavy sedation and muscle paralysis because of patient discomfort , due to a significantly increased level of imposed work of breathing ( wob ) , however , has so far limited the use of hfov in larger pediatric patients and in adult patients . in the present issue of critical care , van heerde and colleagues describe a new flow - demand system that significantly allows for reducing wob during hfov . this new concept , so far tested in a bench test , gives the potential for adult intensive care physicians to more often use hfov and to further investigate hfov . high - frequency oscillatory ventilators can be seen as continuous positive airway pressure ( cpap ) devices that allow generation of pressure oscillations around a continuous distending pressure , which will facilitate elimination co2 mainly by accelerating the molecular diffusion processes . accepting hfov as such a ' super - cpap ' allows one to realize that maintaining spontaneous breathing during hfov should be nothing other than natural . this maintenance is possible and well tolerated in newborns , and was probably a significant contributor to improved pulmonary outcome in this patient group . as previously shown by van heerde and colleagues , the imposed wob for a neonate or an infant ( up to a bodyweight of 10 kg ) on hfov is considerably low ( < 0.5 j / l or < 1.0 j / l , respectively ) during spontaneous tidal breathing with physiologic or smaller tidal volumes between 7 ml / kg to 5 ml / kg and this is independent of endotracheal tube size . with increasing patient size and weight , increasing the fresh gas flow rate allows one to reduce the imposed wob , but not to an acceptable level in the large child or in the adult necessitating heavy sedation , analgesia and often neuromuscular blockade . using the new flow - demand system , the imposed wob can be considerably reduced to a maximum of 0.5 j / l during shallow or normal breathing . on the basis of currently available data from the experience with airway pressure release ventilation or biphasic positive pressure both methods allowing for unrestricted spontaneous breathing at any phase of the ventilatory cycle because of an integrated high - flow or demand valve cpap system it might be postulated that a new high - frequency oscillation ventilator equipped with a flow - demand system allowing for unrestricted spontaneous breathing should allow for less sedation , and should therefore decrease the duration of mechanical support , decrease the length of stay in the intensive care unit , and , ultimately , decrease the overall costs of hospitalization . the application of hfov was mainly reported as a rescue ventilatory mode in adult patients with acute respiratory distress syndrome who were thought to have failed conventional ventilation . outcome results from such studies can not reflect the real potential of hfov as a lung protective ventilatory mode . with the possibility of maintaining spontaneous breathing , hfov could now be used in patients with mild and/or early forms of acute lung injury . with the neonatal experience demonstrating that a lung - protective effect with hfov requires an early initiation of hfov before the lung is damaged , continuing until the lung is no longer vulnerable to ventilator - induced injury , an early transition to hfov should now be considered in adult acute lung injury / acute respiratory distress syndrome patients , but this will need proper clinical testing . weaning concepts from hfov to any form of assisted ventilation or to extubation ( as is already possible in newborns and infants ) will also need re - evaluation . in patients with acute respiratory distress syndrome , perfusion matching , intrapulmonary shunting , and arterial oxygenation , indicating recruitment of previously nonventilated lung areas . hfov at high lung volumes ( i.e. recruited lung ) , which is classically achieved by a stepwise increase in continuous distending pressure to oxygenation and chest x - ray targets , has been shown superior to hfov at low lung volumes . this high - volume approach during hfov is often associated with relatively high airway pressures , which can cause hemodynamic compromise necessitating intravascular volume load ; the airway pressure might not , however , be sufficient to optimally expand the lungs . in the heavily sedated and paralyzed patient , the continuous distending pressure can be titrated up the inflation limb ( to recruit ) and down the deflation limb ( to find the least pressure required to keep the lungs open ) of the static pressure volume curve , which often allows substantial reduction of mean airway pressures while reducing hemo - dynamic side effects to a maximum . with the possibility of benefiting from spontaneous breathing for better recruitment of the dependent lung areas close to the diaphragm , it might become possible to use lower continuous distending pressures to achieve the same oxygenation goals . this may result , together with the effects of the periodic reduction of intrathoracic pressure resulting from spontaneous breathing , in better venous return to the heart , in improved ventricular filling , and therefore in increased cardiac output and oxygen delivery . converting hfov from a ventilation mode that often requires suppression of spontaneous breathing in larger children and adults to a ' super - cpap ' system that allows for unrestricted spontaneous breathing at any phase of the ventilatory cycle because of an integrated high - flow or demand valve system will give hfov the ultimate chance to prove its real potential for optimal lung protection . cpap = continuous positive airway pressure ; hfov = high - frequency oscillatory ventilation ; wob = work of breathing . | in the present issue of critical care , van heerde and colleagues describe a new technical development ( a flow - demand system during high - frequency oscillation ) that may have an important impact on the future use of high - frequency ventilation in children and adults .
flow compensation on patient demand seems to reduce the imposed work of breathing , may therefore increase patient comfort , and should theoretically allow for maintaining spontaneous breathing while heavy sedation and muscular paralysis could be avoided . with further technical development of this concept , high - frequency oscillation can finally be added to the techniques of mechanical ventilatory support that maintain , rather than suppress , spontaneous breathing efforts .
furthermore , this concept will give high - frequency oscillation the chance to prove its potential role as primary therapy in patients with acute lung injury / acute respiratory distress syndrome , the chance to reduce the incidence of high - frequency oscillation failure for patient or physician discomfort as reported in so many clinical trials in the past , the chance to most probably allow successful weaning from high - frequency oscillation to extubation , and , ultimately , in analogy to what has been reported from the experience with other ventilator modes that allow for maintaining spontaneous breathing , the chance to decrease ventilator days in patients with acute lung injury / acute respiratory distress syndrome . |
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an important conceptual advance in our understanding of the basic principles of cellular organization is that the peroxisome , an organelle known for its essential role in lipid metabolism , is derived from the er ( titorenko et al . , 1997 ; titorenko and rachubinski , 1998 ; mullen et al . , 1999 ; a growing body of evidence also supports the view that peroxisomes , similar to the secretory endomembrane system of vesicular flow , constitute a multicompartmental endomembrane system in which individual compartments undergo a stepwise , time - ordered conversion into mature , metabolically active peroxisomes ( titorenko et al . , 2000 ; all of these findings contradict the common textbook rendition of the peroxisome as a semiautonomous , static , and homogenous subcellular compartment whose assembly , as an organelle outside the secretory and endocytic pathways of vesicular flow , does not involve intercompartmental vesicular trafficking ( lazarow , 2003 ) . now , as this basic paradigm of cellular organization is about to be revised in cell biology textbooks ( kunau , 2005 ; schekman , 2005 ) , there is an urgent need to recapitulate numerous observations on the dynamic nature of the er - dependent process of peroxisome assembly . the scope of this paper is to summarize the growing evidence in support of a role for the er as the template for the formation and maintenance of peroxisomes . we also discuss our current knowledge of the multistep , bidirectional flow of membrane - enclosed protein carriers through the er - derived peroxisomal endomembrane system . in addition , we outline the most important unanswered questions and directions for future research in this vibrant and rapidly evolving field . the origin of peroxisomes has long been matter of debate , and partially underscoring this controversy has been the mode by which peroxisome - destined proteins are synthesized and targeted within the cell . for instance , a major tenant of the previous er - vesiculation model for peroxisome biogenesis was that all of the soluble and membrane bound protein constituents of the peroxisome were synthesized cotranslationally on the er ( beevers , 1979 ) . these nascent proteins were proposed to then be sequestered into an expanding vesicle that would eventually bud from the er to produce a mature , functional peroxisome ( beevers , 1979 ) . however , subsequent observations suggested that peroxisomal proteins were not synthesized on the er but on free polyribosomes in the cytosol . these and other data led to the growth and division model for peroxisome biogenesis wherein peroxisomes , like mitochondria and chloroplasts , were considered to increase in size by the posttranslational import of their protein constituents and proliferate only through the division of preexisting peroxisomes ( lazarow and fujiki , 1985 ; purdue and lazarow , 2001 ; lazarow , 2003 ) . notably , the er in the growth and division model was deemed only to be a source of membrane lipids for the enlargement of preexisting peroxisomes . although for most of the past two decades the growth and division model has generally been considered the paradigm for peroxisome biogenesis , the recent monitoring of the sorting of various pmps in evolutionarily diverse organisms has revealed that for at least a subset of these pmps , referred to as group i pmps ( titorenko and rachubinski , 2001 ) , the initial sorting site is the er rather than the peroxisome membrane . sorting of group i pmps to and within the er also appears to be mediated by several different mechanisms ( fig . 1 ) . for instance , in mammalian cells , the group i pmp pex16p is inserted cotranslationally into er membranes and seems to be localized throughout the entire er before its sorting to peroxisomes ( kim et al . , 2006 ) . in the yeast saccharomyces cerevisiae , yarrowia lipolytica , and hansenula polymorpha , group i pmps pex2p , -3p , and -16p are also initially targeted to the general er ( titorenko and rachubinski , 1998 ; hoepfner et al . , 2005 ; haan et al . , 2006 ) . however , unlike mammalian pex16p , the er targeting and insertion of these essential components of peroxisome assembly in s. cerevisiae does not require the sec61p - dependent machinery for co- and posttranslational import of secretory proteins ( south et al . , 2001 ) . furthermore , unlike mammalian pex16p that remains in the general er before its sorting to peroxisomes ( kim et al . , 2006 ) , at least one of the group i pmps in s. cerevisiae , namely , pex3p , is directed from the general er to a distinct subdomain of the er ( hoepfner et al . , 2005 ) . this er subdomain is referred to as the preperoxisomal template ( titorenko and rachubinski , 2001 ) and is considered to be the site where preperoxisomal carriers are formed . that is , after being segregated into the preperoxisomal template , pex3p serves as a docking factor for pex19p , a predominantly cytosolic protein ( hoepfner et al . , the pex3p - dependent recruitment of pex19p from the cytosol to the outer face of the preperoxisomal template in s. cerevisiae is mandatory for the budding of small preperoxisomal vesicles ( hoepfner et al . , 2005 ) . these er - derived carriers of pex2p , -3p , -16p , and -19p lack secretory cargo proteins generalized models for the flow of membrane - enclosed carriers through the peroxisomal endomembrane system in yeast , mammals , and plants . ppt , preperoxisomal template ; ppv , preperoxisomal vesicle ; sv , secretory vesicle ; tbsv p33 , tsbv 33-kd replicase protein . although the mechanism responsible for segregating group i pmps from secretory and er resident membrane proteins in yeast remains to be established , it is noteworthy that the membrane of the er - derived preperoxisomal vesicles in y. lipolytica has unusual ergosterol- and ceramide - rich lipid domains ( boukh - viner et al . , 2005 ) . these lipid domains are similar to detergent - resistant lipid domains in the membrane of s. cerevisiae er , where glycosylphosphatidylinositol - anchored secretory proteins cluster and thereby segregate from other secretory proteins ( mayor and riezman , 2004 ) . it is possible , therefore , that discrete lipid domains , perhaps ergosterol- and ceramide - rich lipid domains , in the membrane of yeast er serve also as a sorting platform for segregating group i pmps from secretory and er resident membrane proteins . the resulting partitioning of group i pmps into these membrane domains could also serve to generate an er template for the formation of preperoxisomal vesicles . in contrast to yeast pex2p , -3p , and -16p and mammalian pex16p , other group i pmps , such as ascorbate peroxidase ( apx ) in plant cells ( mullen et al . , 1999 ) and pex13p in mouse dendritic cells ( geuze et al . , 2003 ) , can only be detected in a distinct portion of the er , suggesting that they are targeted from the cytosol directly to a preexisting subdomain of the er membrane . the terms peroxisomal er ( per ) and lamellar er extension were coined for this er found in plants and mice , respectively ( mullen et al . , 2001 ; at least one notable difference between these two er subdomains is that per is considered to be a portion of rough er membrane ( lisenbee et al . , 2003 ) , whereas the lamellar er extension is a specialized domain in smooth er membrane ( geuze et al . , 2003 ) . in plant cells , the cytosol - to - per targeting of apx occurs posttranslationally and requires atp as well as at least three components of the hsp70 chaperone machinery ( mullen et al . , 1999 ) . collectively , the aforementioned findings suggest that by segregating a distinct set of membrane proteins and lipids into specialized er subdomains , plant and mouse dendritic cells have evolved a platform for the targeting of group i pmps from the cytosol to the er membrane . the existence of such a platform in the er membrane could increase the efficiency of the er - dependent , multistep process of peroxisome assembly in these cells . what structural features of group i pmps are crucial for their sorting to the er or to the peroxisomal membrane via either general er or an er subdomain remain to be determined . at present , it seems that the targeting of these pmps from the cytosol to the er membrane and their subsequent exit from the er are mediated by two partially overlapping sets of sorting signals . i pmps either co- or posttranslationally to the general er or an er subdomain , whereas the other set of signals act from within the er lumen to sort these pmps to the peroxisome ( baerends et al . , 1996 ; elgersma et al . , 1997 ; mullen and trelease , 2000 ; kim et al . , 2006 ) . although all group i pmps exit the er via distinct preperoxisomal carriers that do not enter the classical secretory pathway of vesicular flow ( titorenko and rachubinski , 1998 ; geuze et al . , 2003 ) , the morphology of these carriers in at least yeast and mammalian cells appears to differ ( fig . cerevisiae , y. lipolytica , and h. polymorpha , the er - derived preperoxisomal carriers are small vesicles ( titorenko and rachubinski , 1998 ; hoepfner et al . , 2005 ; in contrast , the preperoxisomal carriers in mouse dendritic cells arise through direct en block protrusion of the specialized er subdomain , the lamellar er extension ( geuze et al . , , the lamellar extension detaches from the er , giving rise to pleomorphic tubular - saccular carriers of group i pmps . this detachment of preperoxisomal tubular - saccular carriers from the er does not require coat protein complexes ( cops ) i and ii , which function in the formation of er - derived carriers for secretory proteins ( geuze et al . it is noteworthy that , akin to er - derived preperoxisomal carriers in yeast cells , all known types of transport carriers for secretory proteins are small vesicles in these cells ( lee et al . , 2004 ) . on the contrary , just like the preperoxisomal carriers in mammalian cells , at least a subset of er - to - golgi carriers for many secretory proteins in these cells are pleomorphic tubular - saccular structures that are formed through direct en block protrusion of specialized domains in the er membrane ( watson and stephens , 2005 ) this fundamental difference in the morphology of er - derived transport carriers is likely due to the difference in the spatial organization of transitional er ( ter ) , a specialized er subdomain at which proteins are packaged into membrane - enclosed carriers . in the traditionally used model yeast organism , s. cerevisiae , the entire er acts as ter , facilitating the budding of copii - coated vesicles ( rossanese et al . , 1999 ) . in contrast , the ter of mammalian cells is organized into discrete er export sites ( hammond and glick , 2000 ) . it is therefore possible that by segregating a distinct set of membrane proteins and lipids into a specialized er subdomain for the cytosol - to - er targeting of group i pmps ( see the previous section ) , higher eukaryotic organisms have not only separated these domains from the sites for the er targeting of secretory proteins but also developed a platform for the sculpturing of these per subdomains into pleomorphic tubular - saccular carriers of pmps . a critical evaluation of this hypothesis would require testing the spatial organization of the er subdomains for the cytosol - to - er targeting of group i pmps and examining the morphology of er - derived carriers for these pmps in the yeast pichia pastoris . unlike s. cerevisiae and similar to mammals , p. pastoris has discrete ter export sites that give rise to a conventional mammalian - type secretory apparatus ( rossanese et al . , 1999 ) . presently , no solid data exist for the nature of the preperoxisomal carriers in plant cells , although , similar to mammals , the ter in these cells is restricted to discrete sites in the er membrane ( hanton et al . , 2005 ) , suggesting that the organization of the per subdomain as well as the formation of preperoxisomal carriers in plants is similar to that in mammals . recent findings have provided strong evidence that , analogous to some organelles of the secretory endomembrane system , peroxisomes constitute a dynamic organelle population consisting of many structurally distinct compartments that differ in their import competency for various proteins . moreover , it appears that the individual compartments of this peroxisomal endomembrane system undergo a multistep conversion to mature peroxisomes in a time - ordered manner . the posttranslational sorting of two partially overlapping sets of pmps and a few matrix proteins converts two populations of er - derived preperoxisomal vesicular carriers into the small ( 75100 nm ) peroxisomal vesicles p1 and p2 ( titorenko et al . , 2000 ) . these vesicles then serve as the earliest intermediates in a multistep pathway that involves , at each step , the uptake of lipids and the selective import of matrix proteins , eventually resulting in the formation of a mature peroxisome referred to as p6 ( guo et al . , 2003 ) . overall , it seems that in y. lipolytica and perhaps in other yeast , import machineries specific for different peroxisomal matrix proteins undergo a temporally ordered assembly in distinct vesicular intermediates along the peroxisome maturation pathway ( titorenko and rachubinski , 2001 ) . the plasticity of these import machineries is further underscored by the observation that the efficiency with which they recognize nonoverlapping targeting signals present on some of their protein substrates varies under different metabolic conditions . in fact , peroxisomal subforms present in yeast cells growing under conditions that induce peroxisome proliferation differ from basal , nonproliferated subforms with respect to the targeting sequence motifs that are used to direct the same protein to these different subforms of peroxisomes ( wang et al . , 2004 ) . a quite different scenario orchestrates a multistep process of peroxisome assembly and maturation in mouse dendritic cells . herein , the extrusion of the lamellar er extensions is culminated by the detachment of pleomorphic tubular - saccular carriers of pex13p from the er ( geuze et al . , 2003 ) . only after their separation from the er are these preperoxisomal carriers able to recruit to their membranes the atp binding cassette transporter protein pmp70 and , perhaps , the membrane components of the import machinery for peroxisomal matrix proteins ( tabak et al . , this latter step of the peroxisome maturation pathway also results in the formation of the so - called peroxisomal reticulum . only the peroxisomal reticulum is capable of importing at least two peroxisomal matrix proteins , namely , thiolase and catalase , directly from the cytosol ( geuze et al . , 2003 ) instead , they are sorted exclusively into mature globular peroxisomes that , during the final step in the peroxisome maturation pathway in mouse cells , bud from the peroxisomal reticulum ( geuze et al . , 2003 ; tabak et al . , it remains to be established whether other peroxisomal matrix proteins , similar to thiolase and catalase , are imported into the domain of the peroxisomal reticulum that gives rise to mature globular peroxisomes or whether , alternatively , these other matrix proteins in mouse cells are sorted to globular ( mature ) peroxisomes only after their budding from the peroxisomal reticulum . in both models for the multistep assembly and maturation of peroxisomes , the targeting of pmps to the membrane of the early intermediates in a pathway precedes , and is mandatory for , the import of soluble peroxisomal proteins into the matrix of later intermediates . because this strategy for peroxisome biogenesis has been conserved in the course of evolution , it seemingly provides an important advantage for the efficient , stepwise assembly of mature , metabolically active peroxisomes . it remains to be established whether , similar to a stepwise assembly of import machineries specific for different peroxisomal matrix proteins in yeast cells ( titorenko and rachubinski , 2001 ) , the import machineries for such proteins in mammalian cells can undergo a temporally ordered assembly in distinct intermediates along the peroxisome maturation pathway . it is also unclear at the moment whether the peroxisome maturation pathway acting in mammalian cells , akin to the pathway that functions in yeast cells ( titorenko and rachubinski , 2000 ; titorenko et al . , 2000 ) , includes fusion of any early pathway intermediates . it is tempting to speculate that such fusion of early pathway intermediates in yeast results in the formation of an er peroxisome intermediate compartment ( erpic ) . such a compartment could ( 1 ) provide a template for the formation of downstream intermediates in the peroxisome assembly and maturation pathway and ( 2 ) function in the sorting of pmps from those escaped er resident proteins that are retrieved by retrograde vesicular transport between the erpic and the er . both of these tentative functions of the erpic share similarity with the functions that have been proposed for the er golgi intermediate compartment , also known as vesicular tubular clusters , which may regulate a bidirectional traffic of membrane - enclosed carriers through the classical secretory pathway ( lee et al . , 2004 ) . importantly , the resident proteins of the post - er compartments in both the peroxisomal endomembrane system and the classical secretory system return to the er in response to the treatment of yeast cells with brefeldin a , an inhibitor of copi formation ( salomons et al . , 1997 ) . thus , similar to its role in the secretory endomembrane system , yeast copi can function in the retrieval of those er resident proteins that had entered the peroxisomal endomembrane system by mistake . this is in contrast to copi in cultured human fibroblasts , in which peroxisome - to - er retrograde protein transport , if any , does not depend on copi ( south et al . , 2000 ; these findings further support the notion that yeast and higher eukaryotic organisms may use different strategies for the er - dependent formation and maintenance of their peroxisomal endomembrane systems . although it is not yet known whether , in plants , a multistep pathway for peroxisome assembly and maturation exits that is either similar or distinct from that in yeast and/or mammals , recent findings suggest that peroxisomes in plant cells can form large pleomorphic structures reminiscent of the mammalian peroxisomal reticulum ( mullen et al . , 2006 ) and are engaged in er - destined retrograde vesicular flow ( fig . 1 ) . evidence for this latter conclusion comes from observations that when the tomato bushy stunt virus ( tbsv ) replication protein p33 is expressed on its own in plant cells , it is sorted initially from the cytosol to peroxisomes and then , via peroxisome - derived vesicles and together with resident pmps , to the per ( mccartney et al . , 2005 ) . remarkably , several aspects of the peroxisome - to - per sorting of p33- and resident pmp for instance , both these processes depend on the adp - ribosylation factor ( arf ) 1 , which promotes the formation of copi - coated vesicles ( lee et al . , 2004 ; mccartney et al . , 2005 ) . in addition , the targeting signal of p33 that mediates the sorting of peroxisomal - derived vesicles to the per resembles an arginine - based motif responsible for the copi - dependent , vesicle - mediated retrieval of escaped er membrane proteins from the golgi ( mccartney et al . , , it has been suggested that the p33-promoted peroxisome - to - per retrograde transport of vesicles delivers to the per early peroxins ( membrane bound peroxins involved in the early stages of peroxisomal membrane assembly ) that stimulate the formation of membrane - enclosed carriers of pmps as an essential phase of the tbsv life cycle ( mccartney et al . , 2005 ; it is not clear at the moment whether the proposed reverse protein sorting pathway between peroxisomes and er can only be induced in tbsv - infected plant cells or if it can also function in uninfected plants , or in other organisms , as a mechanism for the retrieval of escaped er resident proteins . in addition to their proposed role in the peroxisome - to - er retrograde protein transport in virus - infected plant cells , both arf1 and copi can induce the proliferation of the peroxisomal endomembrane system in other evolutionarily diverse organisms by promoting the membrane scission event required for peroxisome division ( fig . , yeast mutants impaired in arf1 and copi , as well as mammalian cells deficient in copi assembly , accumulate a reduced number of elongated tubular peroxisomes , consistent with impairment in peroxisome vesiculation ( passreiter et al . , 1998 ; lay et al . , 2005 ) . incubation of highly purified rat liver peroxisomes with cytosol results in specific binding of both arf1 and copi to the peroxisomal membrane , further supporting the notion that their recruitment from the cytosol in living cells is an initial event in the proliferation of the peroxisomal endomembrane system ( anton et al . , 2000 ) . moreover , similar to arf1 , the subtype 3 of yeast arf also controls peroxisome division in vivo , although , in contrast to arf1 , in a negative fashion ( lay et al . , 2005 ) . collectively , these findings suggest that the peroxisomal endomembrane system and the classical secretory system of vesicular flow are served by a similar set of core protein components required for their communication with the er and for their proliferation . the proliferation of the individual compartments of the peroxisomal endomembrane system is also driven by a peroxisome - specific protein machinery , which includes a distinct set of the pmps and the dynamin - related proteins dlp1 ( dynamin - like protein 1 ) , drp3a ( dynamin - related protein 3a ) , and vps1p ( vacuolar protein sorting protein 1 ) , recruited from the cytosol to the peroxisomal surface by their receptor fis1p ( thoms and erdmann , 2005 ; yan et al . , 2005 ) . a challenge for the future will be to define how the interplay of all these protein components governs such proliferation under the different metabolic conditions in a given cell type or tissue . importantly , peroxisome biogenesis appears to occur by way of a collaborative effort between two equally important pathways . the first pathway operates through the er - dependent formation and maturation of the individual compartments of the peroxisomal endomembrane system , whereas the second pathway involves the precisely controlled division of these peroxisomal compartments . growing evidence supports the view that cells have evolved at least two strategies for the coordination of compartment assembly and division in the peroxisomal endomembrane system . in the first strategy , the multistep growth and maturation of the er - derived preperoxisomal carriers occurs before the completely assembled , mature peroxisomes undergo division ( guo et al . , a significant increase in the number of preperoxisomal carriers , either by their en masse formation from the er ( kim et al . , 2006 ) or by the proliferation of a few preexisting carriers ( veenhuis and goodman , 1990 ; 2003 ) , precedes the growth of these early peroxisomal precursors by membrane and matrix protein import and their conversion to mature , functional organelles containing a complete complement of peroxisomal proteins . determining the relative contribution of these different mechanisms in the formation of peroxisomes in any given organism should now be more feasible through the use of live - cell , photo / pulse - chase labeling methods similar to that reported recently for a study of peroxisome biogenesis in mammalian cells ( kim et al . , 2006 ) . regardless of the strategies that evolutionarily distant organisms use for coordinating the assembly and division of individual compartments of the peroxisomal endomembrane system , the tubulation , constriction , and scission of these compartments is regulated , depending on the cellular and/or environmental conditions of a particular cell type , either by signals emanating from within these compartments ( guo et al . , 2003 ) or by extraperoxisomal signals that are generated inside the cell in response to certain extracellular stimuli ( yan et al . , 2005 ) . these intracellular signals include a distinct group of transcriptional factors that induce the transcription of genes encoding several proteins of the pex11p family ( thoms and erdmann , 2005 ) . the peroxisome membrane bound pex11p - type proteins then directly promote the proliferation of peroxisomal endomembrane compartments or activate peroxisome division indirectly by recruiting the dynamin - related proteins from the cytosol ( yan et al . , 2005 ) . furthermore , the division of the individual compartments of the peroxisomal endomembrane system must be preceded by the expansion of their membranes because of the acquisition of lipids . the er , a principal site for the biosynthesis of phospholipids , is the most likely source of lipids for the growth of the peroxisomal membrane ( purdue and lazarow , 2001 ) , although oil bodies have been implicated also as a source of peroxisomal lipids in some organisms , e.g. , germinated oilseeds ( chapman and trelease , 1991 ) and y. lipolytica ( bascom et al . , 2003 ) . it seems that in y. lipolytica the bulk of phospholipids is transferred from the donor membrane of a specialized subcompartment of the er to the closely apposed acceptor membranes of the two early intermediates , p3 and p4 , in the peroxisome assembly pathway ( titorenko et al . , 1996 ) . although the mechanism responsible for such er - to - peroxisomal membrane transfer of phospholipids via membrane contact sites remains to be established , several working models for the role of er - associated lipid - transfer proteins in the establishment and functioning of such sites have recently been proposed ( levine , 2004 ) . these models should serve as a useful starting point for examining such events during peroxisome biogenesis . growing evidence supports the view that peroxisomes constitute a dynamic endomembrane system that originates from the er . a major challenge now is to identify the molecular players that coordinate the flow of membrane - enclosed carriers through the peroxisomal endomembrane system . future work will aim at understanding the spatiotemporal dynamics and molecular mechanisms underlying this multistep process in evolutionarily diverse organisms . it is conceivable that the analysis of a variety of model organisms , including tissue - cultured human cell lines and various yeast and plant species , will reveal as - yet - unknown strategies and mechanisms governing the biogenesis of the peroxisomal endomembrane system and its relationship with the er . | peroxisomes have long been viewed as semiautonomous , static , and homogenous organelles that exist outside the secretory and endocytic pathways of vesicular flow . however
, growing evidence supports the view that peroxisomes actually constitute a dynamic endomembrane system that originates from the endoplasmic reticulum .
this review highlights the various strategies used by evolutionarily diverse organisms for coordinating the flow of membrane - enclosed carriers through the peroxisomal endomembrane system and critically evaluates the dynamics and molecular mechanisms of this multistep process . |
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hepatic portal venous gas ( hpvg ) is a rare radiological sign often associated with intestinal ischaemia requiring emergency laparotomy . this case report will discuss a 77-year - old caucasian man with gastrointestinal symptoms , neutropenic sepsis and hpvg as seen on competent tomography . a review of the literature will also be conducted with regard to the pathophysiology , conditions associated with hpvg and the role of conservative management in non - ischaemic pathologies . a 77-year - old male was transferred from a rural hospital ; he initially presented with symptoms of left iliac fossa pain , diarrhoea , nausea , vomiting and fevers for 3 days duration . he has a past history of advanced prostate cancer with bony metastasis , previous chemotherapy and radiotherapy and type two diabetes mellitus requiring insulin . general examination revealed sinus tachycardia and an elevated temperature . on abdominal examination , there was diffuse tenderness worse in the left iliac fossa , but soft with positive bowel sounds . g / l ( 115165 g / l ) , low white cell count 1.56 10/l ( 4.010.0 10/l ) with normal coagulation , liver function , lipase and lactate parameters . competent tomography ( ct ) abdomen scan identified grossly thickened small bowel with a large amount of portal venous gas in liver ( figs 1 and 2 ) . like us , the radiologist was concerned these features were highly suggestive of small bowel ischaemia . figure 1:computerized tomography abdominal axial section showing air within the hepatic portal veins ( yellow arrows ) . figure 2:computerized tomography abdominal axial section showing air within the hepatic portal veins ( yellow arrows ) . computerized tomography abdominal axial section showing air within the hepatic portal veins ( yellow arrows ) . computerized tomography abdominal axial section showing air within the hepatic portal veins ( yellow arrows ) . however , with no peritoneal signs , he was treated conservatively and admitted to intensive care for further management of neutropenic sepsis . he was fluid resuscitated , commenced on intravenous tazocin , granulocyte - colony stimulating factor ( gcsf ) and nasogastric decompression . no source of infection was found on blood , stool and urine cultures . with a rising c - reactive protein of 560 mg / l ( 110 mg / l ) and his clinical condition , an exploratory laparotomy was performed . intra - operative findings were moderate bowel oedema , but no bowel or mesenteric necrosis . the patient was apprised of the likely diagnosis of chemotherapy / radiotherapy - related severe enterocolitis with adhesions . twenty - seven days post - admission and a conservatively managed complication of small bowel obstruction , he was transferred back to the rural hospital to be closer to his family . in libman 's and kinoshita 's studies of 64 cases and 182 cases , respectively [ 1 , 2 ] , hpvg was an ominous imaging finding necessitating urgent laparotomy . it had a 75% mortality rate in 43% of kinoshita 's study population that was associated with intestinal ischaemia . progression in imaging technology from plain abdominal radiography to ct scans has since altered the significance of this radiological finding , leading to a rise in clinically benign aetiologies . these include inflammatory bowel disease , abdominal trauma , post - chemotherapy and a rare complication of endoscopic procedures . in ischaemic pathologies , the sensitive and timely detection have allowed prompt treatment , significantly reducing overall mortality rates to 2939% [ 4 , 5 ] . the first states that factors such as mucosal barrier damage , bowel distension , increased intraluminal pressure and bacterial fermentation of carbohydrates in sepsis result in gas production . the second theory is that gas - forming organisms present in the intestinal wall or venous system causes hpvg . as seen in fig . 1 , hpvg is characteristically distributed as peripheral radiolucencies extending to within 2 cm of the liver capsule . with the increased incidence of benign pathologies , many authors in the literature now question the need for emergency surgery [ 35 ] . in the case of our elderly frail patient with a positive radiological sign of hpvg but no acute abdominal signs , the initial trial of conservative treatment was deemed appropriate . however , with minimal improvement and deterioration , an explorative laparotomy was warranted to definitively exclude the possibility of intra - abdominal sepsis with intestinal necrosis . with the negative laparotomy , we conclude that the hpvg was likely triggered by chemotherapy - induced enterocolitis , causing small bowel dilatation and oedema with resultant mucosal damage . . it could be neutropenic enterocolitis with his classic symptoms of fevers and neutropenia ; however the pathology did not occur 10 - 14 days after chemotherapy initiation and had no caecal involvement . a relatively newer type of enterocolitis due to taxane agents such as docetaxel- used in the treatment of metastatic prostate cancer also fits our patient 's presentation however no ischaemic bowel was found . also , his adhesions were likely due to radiotherapy . in other studies , the possibility that certain chemotherapy drugs resulting in intestinal mucosa damage leading to hpvg could not be excluded [ 79 ] . reports hpvg in a patient taking irinotecan and cisplatin who had normal bowel findings on subsequent laparotomy . concludes that when combined with cisplatin , it likely led to mucosal ulceration , bowel distension and anaerobic bacteria gas production . sakamoto et al . further describes a patient with non - small cell lung cancer who developed hpvg on a chemotherapy regimen of carboplatin and paclitaxel . he concludes the effects of chemotherapy agents and the increased intraluminal pressure due to severe constipation from opioid analgesics resulted in hpvg . thus with our patient , it is plausible that his previous chemotherapy treatment and long - term opioid analgesics could have caused his hpvg . in conclusion , the presence of hpvg alone is not a surgical indication , as it is no longer only associated with fatal aetiologies with a poor prognosis . however , more research needs to be conducted into the mechanism behind hpvg and its association with chemotherapy agents . to reduce the rate of negative laparotomies and post - operative complications , we suggest that in cases of hpvg where ischaemic abdominal pathologies have been excluded ; a conservative approach should be trialed . | hepatic portal venous gas ( hpvg ) is often viewed as an ominous imaging finding with a poor prognosis and a high mortality rate .
we recently encountered a case of hpvg in a patient with advanced metastatic prostate cancer previously treated with chemotherapy and radiotherapy .
a laparotomy was performed , which was negative .
although hpvg secondary to chemotherapy is extremely rare , we as clinicians need to consider this aetiology and other benign aetiologies . with the increased rate of benign aetiologies and their successful conservative management ,
the role of emergency laparotomies needs to be re - considered . |
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it may be required to ensure patency of the airway in the lateral position in certain circumstances . some patients who require airway support need to be tracheally intubated with an endotracheal tube to maintain airway patency as soon as possible . several studies have shown successful ventilation in lateral position with the laryngeal mask airway , ventilation and intubation in the lateral position with the intubating laryngeal mask airway with and without the aid of a lightwand , intubation with the lightwand , and fiberoptic intubation . however , newer videolaryngoscopes like c - mac have not been formally evaluated for tracheal intubation in laterally positioned patient . we compared c - mac video laryngoscope and macintosh laryngoscope for tracheal intubation in lateral position . the c - mac video laryngoscope is a new video laryngoscope using a modified macintosh blade , which may be a useful alternative both for routine and difficult airway management and for educational purposes . the c - mac videolaryngoscope has an original macintosh steel blade shape with a closed blade design with no edges and gaps for hygienic traps and is now available in four sizes ( 2 , 3 , and 4 and d the c - mac blade is flattened , resulting in a very slim blade profile ( maximum 14 mm ) , and the edges are slanted to avoid damage to the mouth and teeth . c - mac incorporates the smallest possible ( 2-mm ) digital camera and a high - power light - emitting diode , located laterally in the distal third of the blade . thus , reduced image quality due to damaged optical fibers , need for white color balance and focusing , and immobility due to external light source were eliminated . the view obtained , includes the tip of the blade and therefore , allows visual guidance of the tip of the blade into the vallecula . a color image is displayed on a lightweight , portable high - resolution liquid crystal display monitor . with approval of the institutional ethical committee and written informed consent , we enrolled 100 patients scheduled for various surgical procedures requiring ( flow diagram ) tracheal intubation . patients were aged 18 years or older and were american society of anesthesiologists physical status i , ii , or iii . patients were randomly allocated to c - mac or direct group by computer generated table of random numbers . exclusion criteria were an increased risk of pulmonary aspiration , cervical spine pathology , or anticipated airway difficulties ( i.e. , mallampati grade 4 or thyromental distance < 6 cm ) . patients were premedicated with tab alprozolam 0.5 mg and ranitidine 150 mg at bed time and 6.00 am morning , after they fulfilled inclusion criteria . anesthesia was induced with patients lying in the supine position with fentanyl 2 g / kg and thiopentone 5 mg / kg . head was supported with a firm pillow of 6 cm height . for patients in c - mac group , trachea was intubated using c - mac video laryngoscope by consultant anesthesiologist , who is well - versed with the use of c - mac laryngoscope . in direct group , flow diagram of patient distribution in each group , tracheal intubation was considered as a failure if not accomplished within three attempts . any single insertion of the laryngoscope past the patient 's lips was considered an intubation attempt . if intubation failed , the trachea was intubated after turning the patient to supine position . the investigator , who did intubation in this study , had previously performed about 50 intubations using c - mac video laryngoscope in supine position , but none in lateral position . the following outcomes were recorded by an unblinded observer :
overall intubation success rate;number of intubation attempts;modified cormack lehane score : visualization of the laryngeal inlet was assessed according to the classification of cormack lehane;intubation time ( defined as the time from picking up the laryngoscope to confirmation of tracheal intubation by capnography);frequency of esophageal intubation ; optimizing maneuvers were the external manipulation of the larynx , use of stylet.mucosal trauma ( i.e. , blood detected on the device);lip or dental injury ; anddesaturation ( spo2 < 95% ) .
overall intubation success rate ; number of intubation attempts ; modified cormack lehane score : visualization of the laryngeal inlet was assessed according to the classification of cormack lehane ; intubation time ( defined as the time from picking up the laryngoscope to confirmation of tracheal intubation by capnography ) ; frequency of esophageal intubation ; optimizing maneuvers were the external manipulation of the larynx , use of stylet . mucosal trauma ( i.e. , blood detected on the device ) ; lip or dental injury ; and desaturation ( spo2 < 95% ) . when > 1 intubation attempt was required , time from picking up the laryngoscope for the first intubation attempt until confirmation of successful intubation by capnography was considered to be the total intubation time . the randomized groups were descriptively compared for demographics and baseline airway assessments using summary statistics , such as mean and standard deviation , median and quartiles , or frequency , respectively , for symmetric continuous variables , skewed continuous variables , and categorical variables . number of attempts , esophageal intubation , dental injury , mucosal injury , use of stylet , and application of external laryngeal manipulation were analyzed using chi - square test . mean age was 36.92 15.1 years in direct group and 37.02 15.13 years in c - mac group . mean weight in direct group was 50.54 8.46 kg and 48.8 7.90 kg in c - mac group . modified mallampatti grade as assessed using chi - square test was comparable between the groups . mouth opening ( mean ) was 4.8 cm in direct group and 4.7 cm in c - mac group . the time taken for intubation time was 33.8 9.12 s ( mean ) in direct group and 24.8 8.56 s in c - mac group ( p = 0.001 ) . seven patients ( 14% ) in direct group required > 1 attempts at intubation whereas three patients ( 6% ) required second attempt in c - mac group [ table 2 ] . modified cormack lehane score for laryngeal view one ( cl - i ) was 62% in c - mac group and 36% in direct group . overall intubation success was similar in both the groups but 94% patients in c - mac group were intubated in first attempt whereas only 84% in direct group . external laryngeal manipulation was applied in 10 patients in direct group versus 5 in c - mac group . stylet was used in 10 patients in direct group versus 2 in c - mac group which was statistically significant . there was no failed intubation or dental injury in both the groups [ table 3 ] . mucosal injury was significant indirect group ( 8 patients vs. 1 in c - mac ) . endotracheal intubation in a patient positioned laterally is difficult for anesthesiologists probably because of unfamiliarity of this position . conventional technique of intubation using macintosh laryngoscope is more difficult because laryngeal view is compromised . we compared tracheal intubation with the c - mac video laryngoscope in patients placed in lateral position . from our study , we have shown that time taken for intubation using c - mac was significantly less compared to direct laryngoscope and modified cormack lehane score for visualization of the laryngeal inlet was better when c - mac was used . intubation time in our study was little more compared with previous study which is attributable to right - lateral position . relative difficulty in the right - lateral position was likely attributable to the positioning of the tongue , which ( influenced by gravity ) has a tendency to slip off the laryngoscope blade , while the blade is inserted from the right side of the tongue . besides the deteriorated laryngeal view , limited space between the laryngoscope handle and the tabletop in the right - lateral position would contribute to intubation difficulty if direct laryngoscopy were attempted conventionally , by inserting the tracheal tube from the right corner of the mouth . c - mac has advantage over conventional macintosh laryngoscope having shorter handle and video screen providing real time display . laryngeal view is better with c - mac videolaryngoscope and hence reducing intubation time and also no of attempts at intubation . in the study by kaplan video - assisted laryngoscopy provides an improved view of the larynx , when compared with direct visualization . this technique may be useful for cases of difficult intubation and reintubation as well as for teaching laryngoscopy and intubation . furthermore , external laryngeal manipulation can be applied effectively by the assistant as he can visualize on video screen . patients with anticipated airway difficulties were excluded from our study ; this explains why modified cormack in a recent multicenter randomized controlled trial evidenced no difference in performance of awake tracheal intubation between flexible fiberscope and video laryngoscope . in emergency situations , the patient 's blood or secretions in the airway often complicate intubation attempts , and pharmacological optimization of airway management with adequate sedation and muscle relaxation might not be available . in these situations , the c - mac may prove less effective than in the current study because the camera view could be compromised by fogging or pharyngeal blood . there is no difference in overall success in intubation between c - mac and macintosh laryngoscope . in summary , the c - mac video laryngoscope offers high success rates in lateral position and intubation can be accomplished in less time . the current study concludes that c - mac takes less time compared to macintosh for intubations in lateral position . the c - mac video laryngoscope thus seems to be an effective approach for endotracheal intubation in lateral position . | background and aims : endotracheal intubation is conventionally performed when the patient is in supine position .
it may be required to secure airway in laterally positioned patient .
tracheal intubation in lateral position seems to be difficult because the laryngeal view is compromised .
hence , c - mac video laryngoscope ( karl storz , germany ) , a newer device using a modified macintosh blade may be useful for intubation in lateral position.material and methods : a total of 100 american society of anesthesiologists grade i and ii patients , randomly allotted to c - mac or direct laryngoscopy group .
patients with difficult airway were excluded .
after induction of anesthesia , patient was put in right - lateral position and intubation was carried out by consultant who is well - versed in using c - mac laryngoscope .
time for intubation , number of attempts , modified cormack
lehane grade , mucosal injury , and external laryngeal manipulation applied were noted.statistical analysis : demographics and baseline airway assessments were analyzed using summary statistics .
unpaired t - test was used to assess intubation time .
number of attempts , esophageal intubation , dental injury , mucosal injury , use of stylet , and application of external laryngeal manipulation were analyzed using chi - square test.results:overall intubation success rate was 100% .
the time taken in c - mac group was 24.8 8.5 s and in direct group was 33.8 9.12 s. the number of intubation attempts was not significant .
cormack
lehane grade was better with c - mac laryngoscope .
mucosal injury and use of external laryngeal manipulation was more in direct group.conclusion:c-mac is better than macintosh laryngoscope for intubation in lateral position . |
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10%30% of people all over the world are influenced , while 526% of them are affected severely [ 15 ] . in contrast , pulsatile tinnitus ( pt ) is relatively rare and has been described in case reports or summaries with a relatively small sample [ 615 ] . . the neurobiological basis may be ongoing abnormal spontaneous neural activity , brain plasticity , or disturbed brain network [ 1627 ] , while in contrast , pt is an auditory percept by stimulation of the hair cells in the inner ear . complaints include a sound like water flow , wind blowing , or beat of the drum in the ear , and so forth . typically , symptomatic improvement could be achieved by external compression of the internal jugular vein in the neck on the symptomatic side . possible etiologies include sigmoid sinus diverticulum , atherosclerosis , abnormal vascular loops , aneurysm of internal carotid artery , mastoid emissary vein , dural arteriovenous shunts , paraganglioma , involuntary contraction of muscles in the middle ear , and so forth [ 611 , 2836 ] . the abnormal blood flow induced by a focal defect of mastoid bone shell in the region of the transverse - sigmoid junction is a common etiology according to our daily work . pt is more likely a vascular or muscular originating disorder rather than a true neural activity disorder induced disease . increasing attention is being given on the brain functional activity of the tinnitus patients [ 1627 ] . however , these studies were focused only on npt patients ; the pt patients were defined as exclusion criteria because of the totally different mechanisms of the two subtype tinnitus . thus , it is still not known whether long time pt stimulation can cause the brain abnormalities at present . besides , 2060% of tinnitus patients have been reported to be depressed [ 22 , 3741 ] . an investigation using resting - state functional magnetic resonance imaging ( rs - fmri ) technique focusing on brain activities of pt patients is an interesting and essential work . in this study these alterations may be considered as neural reorganization or brain plasticity in patients with pt . to the best of our knowledge forty two patients with persistent unilateral pulsatile tinnitus and 42 healthy controls , matched in gender , age , and right or left handedness , were enrolled in the study . eighteen patients suffered from left - side pulsatile tinnitus and 24 from right - side pulsatile tinnitus . all of the patients complained of a sound like low- to median - pitched beats of the drum in the ear . symptomatic improvement could be achieved by external compression of the internal jugular vein in the neck on the symptomatic side . ct angiography ( cta ) examinations [ 42 , 43 ] suspected the etiology of sigmoid sinus diverticulum caused by focal defect of mastoid bone shell in the region of the transverse - sigmoid junction . digital subtraction angiography thus , the etiology of all of the patients was confirmed as a focal defect of mastoid bone shell in the region of the transverse - sigmoid junction . all of the subjects had normal hearing threshold and magnetic resonance imaging findings of brain and had no history of neurological and psychiatric illness , alcohol or drug abuse , and severe visual impairment ( the hearing threshold was determined by puretone audiometry ( pta ) examination . participants had hearing thresholds < 25 db hl at the frequencies of 0.250 , 0.500 , 1 , 2 , 3 , 4 , 6 , and 8 khz . the severity of tinnitus and related distress were measured using the validated questionnaires such as tinnitus handicap inventory ( thi ) originally developed by kam et al . and newman et al . . this study was approved by medical research ethics committee and institutional review board of beijing tongren hospital , capital medical university , beijing , china , and written informed consent was obtained . imagines were acquired on a 3.0 t magnetic resonance scanner ( general electric medical systems , milwaukee , wi , usa ) . the matched eight - channel phased array coil was used with foam padding to reduce head motion and scanner noise . resting - state fmri was obtained using an epi ( echo planar imaging ) pulse sequence : tr ( repetition time)/te ( echo time ) = 2000/35 mm , flip angle = 90 , field of view = 24 cm 24 cm , matrix = 64 64 . twenty - eight axial slices were obtained with 4 mm thickness and a 1 mm gap . each fmri session lasted 400 seconds . a 3-dimensional brain volume ( 3d - bravo ) technique was used to acquire high - resolution structural images ( tr = 8.8 ms , te = 3.5 ms , ti = 450 ms , fov = 24 cm 24 cm , matrix = 256 256 , slice thickness = 1.0 mm without gap , 196 slices , 1 averages ) . during the scan , subjects were asked to remain motionless , not to think of anything particular during the functional scans . the preprocessing was carried out by using data processing assistant for resting - state fmri ( dparsf ) ( http://www.restfmri.net/ ) which is based on statistical parametric mapping ( spm8 ) ( http://www.fil.ion.ucl.ac.uk/spm/ ) and resting - state fmri data analysis toolkit ( rest ) ( http://www.restfmri.net/ ) , including removing the first 20 volumes for the signal equilibrium and participants ' adaptation to the scanning noise , slice timing , head motion correction , spatial normalization to the montreal neurological institute ( mni ) template ( resampling voxel size = 3 mm 3 mm 3 mm ) , linear trend removal , temporally bandpass filtering ( 0.010.08 hz ) , and spatially smoothed with a gaussian kernel of 4 mm full - width at half maximum . exclusion criteria include a head motion larger than 1.5 mm maximum displacement in any direction or an angular rotation greater than 1.5 throughout the scan . many studies have suggested that regional alff results could be influenced by gray matter ( gm ) volume [ 48 , 49 ] . we performed a voxel - based morphometry ( vbm ) analysis to investigate whether a changed functional result is related to changes of gm based on the comparison of volume and concentration of gm between pt patients and normal controls using the 3d - bravo sequence . the preprocessing protocols including normalization , optional modulation , segmentation , and smoothing are similar to previous studies described . gm intensity maps in the mni space data were spatially smoothed with 8 mm full width at half maximum gaussian kernel . two - sample t - tests were performed between the patient and normal control groups . the threshold was set to p < 0.01 corrected for multiple comparisons using monte carlo simulation . if there is any positive result , the voxel - wise gray matter volume will be taken as covariates in rest calculations . the threshold was also set to p < 0.01 corrected for multiple comparisons using monte carlo simulation . we applied rest to calculate the alff , which is similar to previous studies [ 5153 ] . briefly , the time courses were first converted to the frequency domain using an fft ( fast fourier transform ) . the square root of the power spectrum was computed and then averaged across 0.010.08 hz at each voxel . the alff of each voxel was divided by the global mean alff value for each subject , resulting in a relative alff . the global mean alff value was calculated for each participant within a group global mean mask . two - sample t - tests and fisher 's exact test were used to compare demographic data between two groups . two - sample t - tests were performed to calculate the gm volume and alff difference between groups . the alff values of all the pulsatile tinnitus patients and healthy controls will be compared by two - sample t - tests and the age was included as a covariate . , chicago , il ) to explore the relationships between the thi result , pt duration , and alff values of the peak voxels in the patient group . voxels with a p value < 0.01 ( corrected for multiple comparisons using monte carlo simulation ) and cluster size > 21 voxels were considered to show significant difference between the two groups . we performed a vbm analysis to reveal the gm changes and its relationship with the alff results . the result showed there 's no gm volume difference between the two groups after monte carlo simulation correction . two - sample t - tests were performed to assess differences between groups . as shown in figure 1 and table 2 , after statistically controlling for the age , the bilateral precuneus and inferior frontal gyrus ( ifg ) showed significantly increased alff in the pulsatile tinnitus patients than that in the controls . however , compared to the controls , the left cuneus , right precentral gyrus , and the bilateral middle - inferior occipital gyrus , lingual gyrus , and right superior parietal lobule significantly decreased alff in the pulsatile tinnitus patients . decreased alff values were found in left cuneus , bilateral middle - inferior occipital gyrus , lingual gyrus , right mpfc , right superior parietal lobule , and right precentral gyrus . a significant positive correlation was found between the thi score and alff value in the precuneus ( r = 0.549 , p < 0.001 ) ( figure 2(a ) ) . a correlation trend was also found between the pt duration and alff in the precuneus , but it was not statistically significant ( r = 0.290 , p = 0.062 ) ( figure 2(b ) ) . significant positive correlation was found between the pt duration and left and right ifg ( r = 0.314 , p = 0.043 ; r = 0.342 , p = 0.027 , resp . ) the gm atrophy may lead to artificial reduction in regional alff results . if there is any abnormality , all the functional results should be adjusted . but the vbm analysis performed in our study showed no significant change in the gm volume of pt patients . thus , on the basis of our result , the altered alff in pt patients in our group are believable . it implies that our results could reflect the changes in intrinsic brain functional activities in the pt patients . previously , husain et al . and mhlau et al apart from the totally different etiology between nonpulsatile and pulsatile tinnitus , there should be some other explanations . what is more , a large sample ( n = 257 ) makes it more possible to demonstrate statistically significant changed areas of gm in previous npt patients . however , our study contains relatively fewer patients ( 42 cases ) because of the relatively rare pt . the disease duration of the pt patients ( 660 months ) may not be long enough to exert changes in the volume of gray matter . another possible hypothesis for the diminished gray matter of those npt patients might be the hearing loss . they found that they were both gray and white matter changes around the auditory cortex for subjects with hearing loss alone relative to those with tinnitus and those with normal hearing . however , there was no significant gray matter volume changes in npt patients compared with normal controls . although they studied relatively fewer npt patients ( only 8 cases ) and hearing loss patients ( only 7 cases ) , they concluded that hearing loss , rather than tinnitus itself , had the greatest influence on gray and white matter alterations based on their vbm study . however , the pt patients enrolled in our study all confirmed normal hearing by hearing threshold examination . it is reasonable that no significant changes of brain volume were detected in pt patients . but we still need additional studies to make it clear whether there will be changes in the gray matter in pt patients with longer disease duration . abnormal baseline brain activity in several brain areas was found in the pt patients . increased alff brain areas include the bilateral precuneus and bilateral inferior frontal gyrus ( ifg ) . decreased alff brain areas include left cuneus , bilateral middle - inferior occipital gyrus , lingual gyrus , right mpfc , right superior parietal lobule , and right precentral gyrus . meanwhile , we also focused on the correlation between the alff values and clinical data such as thi score and pt duration . parts of the limbic system play a central role in the development of tinnitus [ 56 , 57 ] . the limbic network were believed to be closely associated with tinnitus distress [ 3841 , 56 , 58 , 59 ] . the precuneus is a highly integrated structure , supposed to be involved in self - consciousness , shifting of attention , auditory imagery , auditory memory retrieval , and memory - related aspects of the tinnitus percept [ 39 , 63 ] . for those who can cope with the npt and have only low distress , increased activities were presented in the pcc / precuneus area in previous studies [ 20 , 39 ] . but there is something we need to pay attention to ; these previous studies were based on continuous scalp eeg recordings and sloreta ( low resolution electromagnetic tomography ) , a tomographic inverse solution imaging technique . there is also some resting - state studies reporting that eeg is not directly linked to the changes in neural activity as measured by bold fmri [ 65 , 66 ] . on the other hand , logothetis et al . found that task - induced bold signal changes correlated better to local field potential ( lfp ) than to single unit spiking , indicating that the bold response reflects the integration of input and intraneuronal processing . such a combination is a good way of understanding the nature of lff anyway , one could presume that the fmri signal change should have a possible correlation with the eeg results but it may not always be accurate . resting - state pet / spect , on the other hand , directly measures the metabolism of different brain areas , reflecting the neural activity in a period of time ( while the alff measures the deviation of the bold signal ) . increased activity in precuneus / pcc regions was observed in subjects with major depression ( examined by pet ( positron emission tomography ) scan ) and unpleasant music perception ( examined by cbf ( cerebral blood flow ) changes ) . in our study , also we found that alff values in precuneus region have significant correlations ( r = 0.549 , p < 0.001 ) with thi score . thus , even though there was no similar study using alff analytic technique to study distressed patients or tinnitus ( npt or pt ) patients , the increased alff in precuneus of the pt patients is considered to be possibly related to pt awareness as well as the tinnitus related distress . a similar result was also reported in a connectivity analysis of npt patients using ica approach based on fmri ( significant correlation between the beta values of pcc / precuneus and thi score , r = 0.68 , p = 0.01 ) . note also the correlation trend between the activity of precuneus with the disease duration ( r = 0.290 , p = 0.062 ) . one can hypothesize that the pt sounds may integrate in the limbic system with time . the altered baseline brain activity in precuneus region should be considered as a kind of modulation secondary to pulsatile tinnitus . this brain system is comprised of precuneus / pcc , medial prefrontal cortex ( mpfc ) , inferior parietal lobule ( ipl ) , lateral temporal cortex ( ltc ) , parahippocampal gyrus ( phg ) , and hippocampus . dmn is active with high metabolic rates at rest [ 71 , 72 ] , during self - referential behavior , episodic memory processing , and so forth but shows divergent fluctuations in spontaneous activities during the task [ 72 , 75 ] . in our study , the patients maintained high activity in precuneus / pcc , indicating preservation of normal autobiographical reveries , despite the presence of persistent pulsatile tinnitus . also , there have been some researches indicating that a positive correlation between the strength of inflow to the temporal cortices and tinnitus - related distress was found in dmn , especially in precuneus / pcc areas . the role of ifg involved in tinnitus patients is still unclear . in a previous task - fmri study , significant increased signal intensity was found in bilateral ifg after trials with stimulation at the tinnitus frequency . these clusters also showed significant correlation between the tinnitus loudness ratings and the bold signal change in that experiment . even the result may be overestimated due to the task design , the correlation trend is still present . our results provide further support for abnormal alff in bilateral ifg in tinnitus patients ; even the loudness of pulsatile tinnitus can not be measured because of the insufficient measurement and standard . thus , the increased alff in bilateral ifg is considered to be vital areas to identify tinnitus awareness . the increased alff may reflect an inhibitory effort in patients with pt to suppress the sound which is in accordance with the heartbeat . meanwhile , there are also researches indicating that the brain connectivity is widely disturbed in the tinnitus patients over time [ 16 , 79 ] . positive correlations were also found between the alff values and disease duration in the left and right ifg ( r = 0.314 , p = 0.043 ; r = 0.342 , p = 0.027 , resp . ) . thus , we hypothesize that these results are possibly reflecting ongoing changes in neural networks of pt patients . early treatment might be essential in ensuring better prognosis . decreased alff in multiple occipital areas are interesting findings in our research . in a previous pet study , decreased occipital blood flow was reported during auditory tasks in the normal control group without temporal activation . the connections between auditory and visual cortex make it possible to alter the brain activity in the occipital areas [ 27 , 8186 ] . in blind subjects , the visual cortex was recruited in the context of auditory localization [ 80 , 81 ] . ( aoas ) , which may reflect the visual region processing soon - to - appear objects after sound source stimulation . contrary to the aoas , the decreased alff in occipital cortex may be caused by neural reorganization in this area . pt patients may need a downregulation adjustment of the aoas to avoid misinterpreting the sounds around . this could be understood as a kind of self - protect mechanism in pt patients . a previous study proved the negative correlations reciprocally characterizing the functional connectivity between auditory and occipital / visual cortex in npt patients . the possible reason is the totally different sensation and etiology of the pulsatile tinnitus from the nonpulsatile type . the pulsatile tinnitus usually sounds like wind blow , rushing water , or just rumbling in accordance with the heartbeat . pulsatile tinnitus is usually caused by vibrations from turbulent blood flow that reaches the cochlea , which often arises from sigmoid sinus diverticulum , atherosclerosis , aneurysm of internal carotid artery , abnormal mastoid emissary vein , dural arteriovenous shunts , paraganglioma , and so forth , [ 7 , 2830 ] , while the exact etiology of nonpulsatile tinnitus remains unclear . thus , it is this particular kind of disease that presents us with results different from the previous ones . we just discuss our findings and try to explore the clinical meanings of the pulsatile tinnitus patients . but the common etiology of pt , according to our daily work , was the abnormal blood flow induced by focal defect of mastoid bone shell in the region of the transverse - sigmoid junction . also , hundreds of patients with this etiology were cured in our hospital , while cured patients with other etiology were not quite common . thus , the underlying pathology of pulsatile tinnitus and the variability and the possibility that it might influence our results could not be discussed this time . data of the patients with the most common etiology are easier to acquire and more important to analyze . we firstly investigate the relatively common type and then , we could continue our investigation for more details . secondly , the neuropsychological tests , such as mini mental state exam ( mmse ) , auditory verbal learning test ( avlt ) , and so forth , were not administrated . but thi is suitable in reflecting the psychological status of the tinnitus patients in our study . thirdly , subjects in patient group with higher thi score and longer disease duration were not included . we only enrolled patients with disease duration between 6 and 60 months because previous results indicated that the activity and neural network of chronic tinnitus patients are quite different from that of patients with onset disease . further studies will enroll patients with extensively different disease duration to reveal the changes of brain activity or brain network over time . but the current study was the first to investigate the changes of baseline brain activity in patients with pulsatile tinnitus using resting - state fmri . our study is effective in offering information about understanding the changes in brain activity in pt patients . last but not least , functional connectivity analysis of the tinnitus network is an important part of tinnitus fmri research . we will apply functional connectivity analyses based on the alff results to study the altered brain network of pt patients . in summary , multiple altered baseline brain activity areas in patients with pulsatile tinnitus were focused on part of the limbic system , the ifg , and multiple occipital areas . results confirmed the disturbances in pt - related neural networks , which may be potentially helpful in understanding the pathophysiology of pt . | numerous investigations studying the brain functional activity of the tinnitus patients have indicated that neurological changes are important findings of this kind of disease .
however , the pulsatile tinnitus ( pt ) patients were excluded in previous studies because of the totally different mechanisms of the two subtype tinnitus .
the aim of this study is to investigate whether altered baseline brain activity presents in patients with pt using resting - state functional magnetic resonance imaging ( rs - fmri ) technique .
the present study used unilateral pt patients ( n = 42 ) and age- , sex- , and education - matched normal control subjects ( n = 42 ) to investigate the changes in structural and amplitude of low - frequency ( alff ) of the brain .
also , we analyzed the relationships between these changes with clinical data of the pt patients . compared with normal controls
, pt patients did not show any structural changes .
pt patients showed significant increased alff in the bilateral precuneus , and bilateral inferior frontal gyrus ( ifg ) and decreased alff in multiple occipital areas .
moreover , the increased thi score and pt duration was correlated with increased alff in precuneus and bilateral ifg .
the abnormalities of spontaneous brain activity reflected by alff measurements in the absence of structural changes may provide insights into the neural reorganization in pt patients . |
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this report depicts the feasibility of the concomitant repair of a large direct inguinal hernia with mesh by using the intraperitoneal onlay approach after extra - peritoneal laparoscopic radical prostatectomy . a 66-year - old man with localized adenocarcinoma of the prostate was referred for laparoscopic radical prostatectomy . the patient also had a 4-cm right , direct inguinal hernia , found on physical examination . to minimize the risk of infection of the mesh , an extraperitoneal laparoscopic prostatectomy was performed in the standard fashion after which transperitoneal access was obtained for the hernia repair . the hernia repair was completed by reduction of the hernia sac , followed by prosthetic mesh onlay . in this fashion , this can decrease the risk of potential infectious complications by separating the mesh from the space of retzius where the prostatectomy is performed and the lower urinary tract is opened . it is estimated that 5% to 10% of patients who are candidates for radical retropubic prostatectomy have a concomitant inguinal hernia . the anatomic view and simultaneous repair of inguinal hernias during radical retropubic prostatectomy has been well described . laparoscopic repair of inguinal hernias with mesh is also well described and can be performed safely and with a low recurrence rate . both laparoscopic inguinal hernia and laparoscopic radical prostatectomy ( lrp ) can be performed in an extraperitoneal or transperitoneal fashion . although rare , the possibility of exposure of the mesh to urinary extravasation and subsequent infection is a concern . herein , we describe the technique of concomitant repair of a large inguinal hernia defect with mesh utilizing the intraperitoneal onlay approach ( ipom ) in the same setting and immediately after extraperitoneal laparoscopic radical prostatectomy ( ep - lrp ) . a 66-year - old man with gleason 7 ( 3 + 4 ) adenocarcinoma of the prostate was referred for laparoscopic radical prostatectomy . he had no major co - morbidities but complained of a bulge in his groin , which had been present for 5 years . the prostate specific antigen ( psa ) was 9ng / dl , and digital rectal examination revealed an enlarged prostate with no nodules ( clinical stage t1c . ) the patient also had a 4-cm right direct inguinal hernia , found on physical examination . an extraperitoneal nerve - sparing laparoscopic prostatectomy and a bilateral pelvic lymphadenectomy were performed in the standard fashion with a 5-trocar approach . the space was developed with the oval - preperitoneal distention balloon ( united states surgical , norwalk , ct ) . after 5 minutes , the balloon was removed , and the blunt tip trocar with cuff ( united states surgical , norwalk , ct ) was introduced . a large hernia defect could be seen and contents could be easily reduced while the extraperitoneal insufflation pressure was maintained at 15 mm hg ( figure 1 ) . the prostate was enlarged with a prominent median lobe , necessitating a wide bladder neck resection and laparoscopic bladder neck reconstruction . to minimize the risk of mesh infection , we elected to perform the hernia repair intraperitoneally . the blunt tip trocar with cuff was inserted and secured in the peritoneal space by dividing the posterior rectus sheet and peritoneum under direct vision . all other trocars were reinserted through the same skin incisions into the peritoneal cavity under direct vision . note the peritoneum loosely hanging medially and anteriorly due to previous dissection of the space of retzius . the hernia sac was reduced . a 5-cm 10-cm gore - tex mesh was laid on the defect to cover it completely . the mesh onlay was secured with the laparoscopic tack device protac ( united states surgical , norwalk , ct ) to cooper 's ligament , the rectus sheath , and laterally to the iliopubic tract ( figure 3 ) . in this fashion , this included 225 minutes for the prostatectomy and 45 minutes for the hernioplasty . estimated blood loss was 450 ml . the pelvic drain was removed on postoperative day 1 , and the patient was discharged on postoperative day 3 . the catheter was removed on postoperative day 10 , as was routine for all our lrp patients at the time . the final pathology revealed pt2cn0m0 prostate cancer ( gleason score 4 + 4=8 ) with negative margins . postoperatively , the psa remained undetectable at < 0.04 ng / dl , and the patient had no evidence of hernia recurrence , infection , or bowel obstruction . the presence of inguinal hernias at the time of planned , radical retropubic prostatectomy is not uncommon . choi et al have described the technique of preperitoneal prosthetic mesh hernioplasty during radical retropubic prostatectomy and found it to be superior to the nonmesh technique with no significant complications . recently , the laparoscopic approach to inguinal hernia repair has emerged as an acceptable alternative to the open approach with minimal morbidity and faster recovery with comparable minimal recurrence rates of 0% to 4.5% with experience . recent meta - analysis of randomized clinical trials comparing open and laparoscopic inguinal hernia repair revealed equivalent efficacy and low recurrence rates combined with faster recovery and significantly fewer postoperative complications in the laparoscopic group . various techniques have been developed for the laparoscopic hernia repair including the totally extraperitoneal repair , the transabdominal preperitoneal repair ( tapp ) , and the intraperitoneal mesh onlay repair ( ipom ) . as with hernia repairs , laparoscopic prostatectomy can be performed in a transperitoneal or extraperitoneal approach . stolzenburg et al , who have described their technique of extraperitoneal radical prostatectomy , recently reported their technique of simultaneous extraperitoneal laparoscopic hernia repair with mesh in 10 patients . allaf et al have reported on a case where both operations were done transperitoneally with no complications . although reassuring , the theoretical risk of mesh infection resulting from a urine leak from the urethro - vesical anastomosis does exit . in addition , an external drain is routinely placed in the proximity of the anastomosis after a radical prostatectomy . even after a watertight anastomosis is ensured , the possibility of prolonged drainage due to lymphorrhea , resulting from pelvic lymph node dissection always exits . the possible prolonged presence of an external drain , in the same space where a synthetic mesh is used can raise the stakes for foreign body colonization and infection . the tapp procedure is generally regarded as the laparoscopic procedure of choice for inguinal hernia repairs and is associated with minimal morbidity and low recurrence rates . in this operation , the parietal peritoneum and preperitoneal contents are dissected to expose the myopectineal orifice . after placement of the appropriate size mesh prosthesis , the peritoneal defect is re - approximated over the mesh . however , during transperitoneal lrp and pelvic lymph node dissection , there is wide dissection of the parietal peritoneum to mobilize the bladder . attempts to retroperitonealize the mesh onlay are futile and not possible . this can leave the mesh in close proximity to the anastomosis and the pelvic drain . we elected to perform the laparoscopic intraperitoneal mesh onlay inguinal hernia repair ( ipom ) with gore - tex after an ep - lrp and lymphadenectomy in our patient . the ipom method is easy to perform and is a viable alternative in this clinical scenario that ensures the separation of the prostatectomy space ( space of retzius ) and the mesh by the posterior rectus sheet and peritoneum . the ipom method is a simple procedure that can be used to cover large defects and to treat both direct and indirect hernias successfully . the downside of an intraperitoneal mesh onlay inguinal hernioplasty has been suggested to be bowel adhesions to the mesh , resulting in bowel obstruction or subsequent fistula formation . we elected to use gore - tex , because the rate of adhesions has been found to be significantly less with its use while providing equivalent tensile strength compared with the strength of marlex ( polypropylene ) . furthermore , intraperitoneal mesh onlay techniques are routinely used in laparoscopic ventral or umbilical hernia repairs . the larger risk for mesh hernioplasties is not bowel obstruction or fistula formation but potential infection in the setting of a clean contaminated case where the urinary tract is entered . complications such as hematoma and seroma have been reported after open and laparoscopic inguinal hernia repairs . coupled with lymphatic drainage and possibly urinary extravasation in the same space , infection of the prosthetic mesh can occur that can necessitate removal of the mesh . choi et al do not report a case of mesh infection in 35 patients who underwent mesh hernioplasties during radical retropubic prostatectomy . when a chance exists of significant urinary leakage or lymphatic drainage , however , they suggest a nonmesh repair . current surgical practice favors the use of tension - free repairs with mesh for large inguinal defects . our patient had an obese pelvis and had a large prostate removed with significant laparoscopic bladder neck reconstruction and bilateral pelvic lymph node dissection . we deemed our patient at higher than average risk for urinary extravasation and lymphatic drainage . simultaneous ep - lrp and ipom hernioplasty is a unique combination in the surgeon 's armamentarium to treat localized prostate cancer patients with concomitant inguinal hernias . as such , both surgical objectives can be attained efficiently in the appropriate patient with minimal morbidity . | objective : this report depicts the feasibility of the concomitant repair of a large direct inguinal hernia with mesh by using the intraperitoneal onlay approach after extra - peritoneal laparoscopic radical prostatectomy.methods:a 66-year - old man with localized adenocarcinoma of the prostate was referred for laparoscopic radical prostatectomy .
the patient also had a 4-cm right , direct inguinal hernia , found on physical examination . to minimize the risk of infection of the mesh
, an extraperitoneal laparoscopic prostatectomy was performed in the standard fashion after which transperitoneal access was obtained for the hernia repair .
the hernia repair was completed by reduction of the hernia sac , followed by prosthetic mesh onlay . in this fashion
, the peritoneum separated the prostatectomy space from the mesh .
a single preoperative and postoperative dose of cefazolin was administered.results:the procedure was completed with no difficulty .
total operative time was 4.5 hours with an estimated blood loss of 450 ml .
the final pathology revealed pt2cn0m0 prostate cancer with negative margins .
no infectious or bowel complications occurred . at 10-month follow - up
, no evidence existed of recurrence of prostate cancer or the hernia.conclusion:concomitant intraperitoneal laparoscopic mesh hernia repair and extraperitoneal laparoscopic prostatectomy are feasible . this can decrease the risk of potential infectious complications by separating the mesh from the space of retzius where the prostatectomy is performed and the lower urinary tract is opened . |
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anterior segment optical coherence tomography ( asoct ) was used to categorize and provide insights into the etiology of capsular bag distension syndrome ( cbds ) . a prospective review was undertaken of 10 cases who presented with signs of late cbds 511 years after uneventful phacoemulsification with in - the - bag posterior chamber intraocular lens implantation . all 10 patients presented with a milky collection within the distended capsular bag without raised intraocular pressure or a shallow anterior chamber . asoct was used to confirm the diagnosis in all cases , and a hyperintense signal was seen in the space between the posterior chamber intraocular lens and the posteriorly bowed posterior capsule . uncomplicated neodymium - doped yttrium aluminium garnet ( nd : yag ) posterior capsulotomy was performed in eight patients , with no resulting change in the intraocular lens position ( measured by asoct ) or subjective refraction . our study showed that asoct is a useful modality to differentiate this condition clearly from posterior chamber intraocular lens opacification and to investigate its causation . nd : yag posterior capsulotomy proved to be a safe and successful treatment for late cbds with no change in biometric or refractive parameters . capsular bag distension syndrome ( cbds ) is a rare complication of phacoemulsification with an anterior continuous curvilinear capsulorhexis and in - the - bag intraocular lens implantation . it may also be associated with a can - opener type capsulorhexis and sulcus - implanted intraocular lens.1,2 in 1990 , davison first described cases of early postoperative cbds with a shallow anterior chamber , elevated intraocular pressure , and induced myopia from the anterior shift of the intraocular lens.3 miyake et al reclassified cbds into intraoperative , early postoperative , and late postoperative cbds based on the heterogeneity of the clinical features in these cases.4 late cbds has also been termed liquefied after - cataract5 or capsulorhexis - related lacteocrumenasia.6 we pooled 10 highly similar cases presenting with late cbds . uniquely in all our patients , anterior segment optical coherence tomography ( asoct , visante , carl zeiss meditec , dublin , ca ) was used to categorize correctly and elucidate the etiology of this subtype of cbds . all 10 patients had a history of uncomplicated phacoemulsification with in - the - bag intraocular pressure implantation . seven patients presented with insidious blurring of vision while three patients were asymptomatic and diagnosed incidentally during routine follow - up . in all patients , slit - lamp examination revealed a quiet anterior chamber with normal intraocular pressure and a characteristic milky fluid trapped within the capsular bag . figure 1a and b shows slit - lamp photographs of a case before and after nd - yag posterior capsulotomy , and figure 1c shows an asoct image demonstrating a hyperintense signal within the capsular bag . asoct was useful in confirming the diagnosis of cbds in all our cases , because a hyperintense signal was seen in the space between the intraocular lens and the posterior capsule . posterior capsular opacification was notably absent in our cases in contrast with the series by pinarci et al where cbds remained undiagnosed for a long time before posterior capsular opacification developed.7 the use of ultrasound biomicroscopy and scheimpflug imaging to aid in the diagnosis of cbds has also been reported.810 murat et al compared the use of ultrasound biomicroscopy and scheimpflug imaging in two patients and found that ultrasound biomicroscopy seemed to be superior to scheimpflug imaging in eyes with extremely distended capsular bags because the latter failed to visualize the posterior capsule and also had an error in estimation of the anterior chamber depth.8 lau et al reported asoct findings in a case of early cbds , demonstrating a shallow anterior chamber and distended capsular bag with a large distance between the intraocular pressure and posterior capsule.11 in our study , asoct was used to estimate the size of the continuous curvilinear capsulorhexis and the anterior chamber depth quantitatively ( pre - yag and post - yag capsulotomy ) . asoct is technically easier to perform because it is noncontact and done with the patient in the upright position , as compared with ultrasound biomicroscopy which requires immersion and the patient to be in a supine position . three cases had uncorrected visual acuity better than 6/12 and one retained good best - corrected visual acuity of 6/6 beyond one year of follow - up . in the presence of a distended capsular bag trapped with milky fluid , there could be technical difficulty in focusing of the nd : yag laser on the posterior capsule , hence leading to failure , which may then require surgical drainage of the trapped fluid.12 however , this problem was not encountered in our series , and nd - yag posterior capsulotomy was safely performed in eight of our patients with no complications , such as an intraocular pressure spike or excessive inflammation . kollias et al reported a case of propionibacterium acnes in late cbds,13 but in our series , the absence of vitreous inflammation post - yag posterior capsulotomy indicated that there was no deleterious infective or inflammatory etiology . of note , there were no cases of retinal detachment seen in our patients after yag posterior capsulotomy . whilst yag posterior capsulotomy remains a relatively simple treatment option hence , it may be prudent to observe the patient until the onset of visually significant symptoms . in four patients with pre - yag and post - yag capsulotomy refractions this finding is similar to that in the series by pinarci et al.7 in our series , we were also able to use asoct to measure the change in anterior chamber depth pre - yag and post - yag capsulotomy , and it similarly showed an insignificant change of less than 0.1 mm in anterior chamber depth measurements , indicating minimal biometric change post - yag capsulotomy ( table 1 ) . in most cases of cbds , be they early or late , the size of continuous curvilinear capsulorhexis tends to be small,9 and tight adherence to the anterior surface of the intraocular pressure prevents escape of intracapsular fluid . in our series , asoct measured the horizontal diameter of the continuous curvilinear capsulorhexis to be in the range of 3.184.70 mm , thus corroborating these observations . however , because there are no longitudinal measurements , one can not confirm whether there had been any capsular contraction . in conclusion , our study showed that asoct is a quick and easy method of imaging the anterior segment to differentiate cbds clearly from posterior chamber intraocular lens opacification . it can also aid in diagnosis of late cbds , especially when the milky fluid is not as obvious . common characteristics seen in our cases were the relatively small continuous curvilinear capsulorhexis in all cases and acrylic intraocular pressures ( ma60bm ) in seven of 10 cases . yag posterior capsulotomy was safe and yielded excellent visual results in all cases , with minimal biometric or refractive changes . | backgroundanterior segment optical coherence tomography ( asoct ) was used to categorize and provide insights into the etiology of capsular bag distension syndrome ( cbds).methodsa prospective review was undertaken of 10 cases who presented with signs of late cbds 511 years after uneventful phacoemulsification with in - the - bag posterior chamber intraocular lens implantation.resultsall 10 patients presented with a milky collection within the distended capsular bag without raised intraocular pressure or a shallow anterior chamber .
asoct was used to confirm the diagnosis in all cases , and a hyperintense signal was seen in the space between the posterior chamber intraocular lens and the posteriorly bowed posterior capsule .
the continuous curvilinear capsulorhexis was measured to be between 3.18 mm and 4.70 mm .
three cases had uncorrected visual acuity better than 6/12 .
uncomplicated neodymium - doped yttrium aluminium garnet ( nd : yag ) posterior capsulotomy was performed in eight patients , with no resulting change in the intraocular lens position ( measured by asoct ) or subjective refraction.conclusionour study showed that asoct is a useful modality to differentiate this condition clearly from posterior chamber intraocular lens opacification and to investigate its causation .
nd : yag posterior capsulotomy proved to be a safe and successful treatment for late cbds with no change in biometric or refractive parameters . |
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pseudomembranous colitis ( pmc ) is a severe form of clostridium difficile infection which is usually confined to rectum and sigmoid colon but sometimes involves ascending colon , making sigmoidoscopy sufficient to detect pmc.1 it is commonly associated with the antibiotics such as fluoroquinolones , clindamycin , cephalosporins , and penicillins.2 in addition , antitubercular ( anti - tb ) agent rifampin , which has broad spectrum of antimicrobial activity , is also known to be an uncommon cause of pmc.3,4 like most of the usual pmc , all of the reported cases of rifampin - induced pmc were also observed in rectosigmoid colon except for one.5 we present a case of rectosigmoid sparing pmc in a patient taking anti - tb agents . an 81-year - old man was admitted to our hospital with a 30 day history of watery diarrhea 10 times per day and general weakness . he was diagnosed as pulmonary tuberculosis 45 days before admission and was on standard anti - tb medications , rifampin ( 600 mg / day ) , isoniazid ( 300 mg / day ) , ethambutol ( 800 mg / day ) , and pyrazinamide ( 1,500 mg / day ) since then . on admission , he stopped taking all anti - tb medications for 2 weeks after 30 days on the regimen . vital signs were stable and abdominal physical examination did not show abnormal findings except for the hyperactive bowel sounds . laboratory findings showed increased leukocyte counts ( white blood cell , 21,200/mm ; neutrophil , 90.7% ) and increased inflammatory markers such as erythrocyte sediment rate ( 75 mm / hr ; normal , 0 - 9 mm / hr ) and c - reactive protein ( 5.14 mg / dl ; normal , 0.02 - 0.3 mg / dl ) . the sputum acid - fast bacteria stain was positive and the chest x - ray showed patchy consolidation in the right lung field . since watery diarrhea started after 15 days of anti - tb medications and continued for 30 days , anti - tb agents induced pmc was suspected before c. difficile toxin result was obtained . , the colonoscope was able to be advanced proximally into the terminal ileum , which showed multiple yellowish plaques only in the cecal mucosa ( fig . biopsy specimen of the cecum revealed disruption of surface epithelium with an overlying mushroom - shaped pseudomembrane which was compatible with the findings of pmc ( fig . , the stool assay for c. difficile toxin came out to be positive but stool culture for c.difficile was negative . oral metronidazole ( 1,500 mg / day ) was started and anti - tb agents excluding rifampin were readministerd 2 days after the initiation of metronidazole . his symptoms improved remarkably the day after taking the medications and did not recur since then . pmc is a characteristic manifestation of full blown c. difficile infection , which is acquired almost exclusively in association with the use of antimicrobials and the consequent disruption of the normal colonic flora . various antibiotics , including lincomycin , clindamycin , and -lactam compunds are well known causative agents of pmc . on the other hand , anti - tb agents are uncommon cause of pmc and a few cases of anti - tb agents - related pmc have been described since the 1980s.6 - 9 although it is still controversial whether rifampin can be a causative agent , rifampin has been postulated to be one of the causes because it has antibacterial activity on a broad spectrum of bacteria whereas isoniazid , pyrazinamide , and ethambutol do not have antibacterial activity to disturb the normal intestinal flora.10 besides the broad spectrum of antibacterial activity , frequent isolation of rifampin - resistant toxigenic c. difficile also implies that rifampin could be the causative agent.11,12 in the present case , given the temporal relationship between anti - tb medications and the onset of symptoms , it is suspected that pmc was associated with anti - tb agents . additionally , the patient 's symptoms did not recur after rechallenging the rest of anti - tb agents except for rifampin , which demonstrates that rifampin caused pmc in our case . the identified risk factors for pmc include advanced age , immunocompromised state , hospitalization , and procedures or medications that alter the intestinal motility or flora.13 the median age of rifampin - induced pmc was 59.5 years , which is younger than that of usual pmc patients.4 our patient was older than most of the reported cases , although he did not have other risk factors . rifampin - induced pmc was also associated with moderate - to - severe liver dysfunction.7,10,14 it is thought that liver dysfunction results in reduced biliary excretion of rifampin , which causes high concentration of serum levels and/or low intestinal rifampin levels.8,14 the clinical presentations of pmc include diarrhea , abdominal pain , fever , and leukocytosis . however , in rifampin - associated pmc , the median latency period was 4 weeks , longer than the commonly known causes of pmc,4 which reflects that rifampin may have less capacity to perturb the intestinal bacterial flora than other antibiotics such as -lactam compounds and clindamycin.10 the latency of our patient 's clinical symptom was about 2 weeks , which was shorter than that of previously reported cases.4,15 the confirmative diagnosis of pmc is either positive stool culture or positive stool toxin assay for c. difficile with clinical data . however , sigmoidoscopy , colonoscopy or abdominal computed tomography scan can provide useful diagnostic information . thus , sigmoidoscopy is usually sufficient to detect typical findings of pmc such as the pseudomembrane . however , there are reports of rectal or rectosigmoid sparing pmc occuring in as many as 23% to 69% of patients.16,17 normal mucosal appearance or nonspecific proctosigmoiditis may be the sigmoidoscopic finding but pseudomembrane can be found in ascending colon as occurred in our case . therefore , total colonoscopy should be performed to confirm the diagnosis of pmc , if sigmoidoscopy is inconclusive . the test for resistant strain of c. difficile is needed for the investigation of causative agent of pmc in a patient suspected to have rifampin - associated pmc . in our case , however , the test for rifampin resistance could not be performed due to negative stool culture of c. difficile . once pmc is diagnosed , it is usually treated with oral metronidazole 500 mg t.i.d . or 250 mg q.i.d . for 10 days.18 besides antibiotic therapy , restoration of beneficial intestinal flora with probiotics may suppress the abnormal overgrowth of c. difficile and thus ameliorate the course of the disease . in our case , watery diarrhea was disappeared with metronidazole therapy , although the discontinuation of all anti - tb medications did not affect his symptom . however , it was recently reported in korea that anti - tb agents induced pmc was treated with maintenance of the anti - tb medications and the addition of both oral metronidazole and probiotics.15 in conclusion , our case reinforces the concept that rifampin can be one of the causes of pmc and reveals that rifampin - induced pmc can be presented as a localized disease with rectosigmoid sparing . thus , total colonoscopy is needed for the differential diagnosis in patients with severe diarrhea developing during or after anti - tb treatment , if sigmoidoscopy is not confirmative . | pseudomembranous colitis ( pmc ) is known to be associated with antibiotic treatment , but is not commonly related to antitubercular ( anti - tb ) agent , rifampin .
pmc is frequently localized to rectum and sigmoid colon , which can be diagnosed with sigmoidoscopy .
we report a case of rifampin - induced pmc with rectosigmoid sparing in a pulmonary tuberculosis patient .
an 81-year - old man using anti - tb agents was admitted with a 30-day history of severe diarrhea and general weakness . on colonoscopy ,
nonspecific findings such as mucosal edema and erosion were found in sigmoid colon , whereas multiple yellowish plaques were confined to cecal mucosa only .
biopsy specimen of the cecum was compatible with pmc .
metronidazole was started orally , and the anti - tb medications excluding rifampin were readministerred .
his symptoms remarkably improved within a few days without recurrence .
awareness of rectosigmoid sparing pmc in patients who develop diarrhea during anti - tb treatment should encourage early total colonoscopy . |
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anencephaly is a devastating yet common neural tube defect which occurs due to the failure of closure of rostral pore of the neural tube that leads to exposed undifferentiated neural tissue . meroanencephaly is the classic form which presents as a lack of cranial vault bones and exposed dorsal undifferentiated dorsal neural tissue . usually , cerebral hemispheres fail to develop and only brainstem and some parts of midbrain may be present . craniorachischisis is the most severe variety whereby the defect extends into continuous spina bifida down to the thoracic vertebrae . here , we present a case of meronencephaly cared for and discharged from our neonatal intensive care unit . a male baby was born at term by vaginal delivery to a primigravida mother who did not have any antenatal visits or folic acid supplementation during pregnancy . he showed gross malformation of the cranium in the form of meroanencephaly and bilateral proptosis [ figure 1a and b ] . he was on intravenous fluid for 3 days after which he was shifted to oral feeds which he tolerated well . the vital need for folic acid supplementation prior to conception was advised and emphasized on the family . ( a - b ) the affected newborn . the flat cranial vault with defective fusion and protrusion of brain matter are notable . the cause may be multifactorial involving genes , gender , teratogen exposure , environment , diet as well as physiological abnormalities such as hyperglycemia or hyperthermia . several studies have shown a protective role of periconceptional intake of folic acid in reducing both the occurrence and recurrence of neural tube defects . without folic acid supplementation the empirical recurrence risk after one affected child is 34% and after two affected children it is 10% . exposure to valproic acid and other antimetabolites of folic acid and other toxins like lead etc . , during critical period of neural differentiation interfere with normal folate metabolism and increases the likelihood of anencephaly . neural tube closure is the result of upward bending and eventual fusing of neural tube to form the hollow tube that becomes the brain and the spinal cord . this takes place at around day 2126 days of gestation beginning at the hindbrain / cervical boundary ( closure 1 ) and spreading bi - directionally into the hindbrain and spinal region . separate closure initiation sites occur at the midbrain - forebrain boundary ( closure 2 ) and at the rostral extremity of the forebrain ( closure 3 ) . however , closure 2 found in other mammals like mice may be absent from human neurulation . failure of closure 1 leads to craniorachischisis involving midbrain , hindbrain , and spinal region . anencephaly develops if closure 1 is completed , but the closure of the cranial neural tube is incomplete , the resulting lesion may be either confined to the midbrain ( meroanencephaly ) or extend into the hindbrain ( holoanencephaly ) . anencephaly can be diagnosed antenatally by ultrasound examination and by elevated maternal alpha - fetoprotein level . anencephaly has to be differentiated from encephalocele which is a cystic extension of the brain from overlying scalp and skull defect . in addition , iniencephalocele is another differential characterized by a triad of the occipital bone defect , rachischisis , and fixed retroflexion of the head with severe cervicothoracic lordosis . the role of 0.4 mg of periconceptional folic acid in reducing the incidence of neural tube defects has been revealed by the medical research council . although the prevalence of neural tube defects has decreased after preconceptional folic acid supplementation , they continue to remain common birth defects for the obstetrician to screen , diagnose , and manage in the pregnant patient . the authors certify that they have obtained all appropriate patient consent forms . in the form the patient(s ) has / have given his / her / their consent for his / her / their images and other clinical information to be reported in the journal . the patients understand that their names and initials will not be published and due efforts will be made to conceal their identity , but anonymity can not be guaranteed . the authors certify that they have obtained all appropriate patient consent forms . in the form the patient(s ) has / have given his / her / their consent for his / her / their images and other clinical information to be reported in the journal . the patients understand that their names and initials will not be published and due efforts will be made to conceal their identity , but anonymity can not be guaranteed . | neural tube defects are a group of congenital anomalies of brain development that carry a significant risk of morbidity and mortality .
anencephaly is a serious form of this defect with a very poor prognosis .
it can present in three forms meroanencephaly , holoanencephaly , and craniorachischisis .
meroanencephaly is considered to be the classic form of anencephaly .
it manifests as a lack of cranial vault bones and exposed dorsal neural tissue resulting from defective neural tube closure in the very early period of gestation .
antenatally , the diagnosis may be suggested by ultrasound examination and by elevated maternal alpha - fetoprotein level . here
, we describe a case of meroanencephaly who was discharged from the hospital in fair condition despite the life - threatening anomaly . |
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extraskeletal or soft - tissue chondroma is a rare benign cartilaginous tumor that develops in the soft tissues without bone or joint involvement , occurring predominantly in the hands and feet of adults in their third and fourth decades of life.1 ) cli - nically , it manifests as a slowly growing subcutaneous nodule , and the diameter is less than 3 cm in most cases , with a development in the facial area reported very rarely . however , the presence of this tumor has been described in the head and neck region , mainly in the tongue , and less likely in the cheek , neck , parotid gland , parapharyngeal space , masticatory space , and masseter muscle,2 ) with the age of onset usually between 30 - 60 years of age . the case presented here is of a particular interest because of the exceptional location of the tumor , and a matter of reconstruction of a defect after the removal . we were unable to find any analogous report in the literature except for only two reports describing cases of extraskeletal chondroma in the auricle.2,3 ) in the presenting case , we report a rare case of chondroma that developed in the auricle , which were treated with wedge excision and reconstructed using a burow 's triangle . 52-year old male patient visited our clinic presenting with an unilateral mass of the auricle for several years . physical examination revealed a 1.51.5 cm sized painless , hard mass accompanying a nodular irregularity , located on the superior portion of the helix ( fig . a 1.00.8 cm sized firm , round , and well - encapsulated mass was completely removed by wedge excision ( fig . 2a ) , and the remaining defect was approximated with a creation of burow 's triangle in the scapha ( fig . 2b , c ) . histopathologic examination revealed a mass consisted of an acidophilic hyalinized matrix with cartilage cells scattered among them , and in the center , local calcification was observ - ed . tumor cells had cytoplasms with a distinct border , and the nuclei were of a circular or an oval form ; atypical or abnormal mitosis was not observed ( fig . histologically , extraskeletal or soft - tissue chondroma is a lobulated nodule surrounded by a fibrous capsule filled with mature hyalinized cartilage tissue , with no association of underlying bone.1,4 ) association with secondary degeneration within the matrix , such as cystic change , ossification , fibrosis , mucinous change , and calcification as in the presenting case , requires differentiation from other soft - tissue tumors . the diseases to be differentiated include tumoral calcinosis , calcifying aponeurotic fibroma , ectomesenchymal chondromyxoid tumor , and chondrosarcoma.5,6 ) the etiology of extraskeletal chondroma is uncertain . the following are 3 hypotheses that have been proposed.2 ) the first hypothesis suggests of the cartilage cells originated from the skeletal structure migrating to the adjacent connective tissue ; the second proposes a conversion of the precartilaginous tissues that are in the ligament attachment area at the activation period ; the last hypothesis proposes a metaplasia of synovial cells differentiating as cartilagi - nous cells . it may recur in 10% to 15% cases ; nevertheless , even in recurred cases , local resection is sufficient.1,2 ) however , excision of the tumor located in helix of the auricle requires a reconstruction of the defect , if it is bigger than 1 to 1.5 cm . many techniques have been described to treat a defect of the helix secondary to tumor excision , and the advancement of the helix by placing a burow 's triangle in the scapha is preferred whenever possible . in the presenting case , the defect measuring 1.5 cm in diameter , was successfully reconstructed with advancement of the helix using a burow 's triangle in the scapha . no irregularity in form and contour we report a case of a very rare extraskeletal chondroma that occurred in the auricle with a review of the literature . | extraskeletal chondroma is a rare benign tumor that develops in the soft tissues , and it manifests as a solitary subcutaneous nodule or a slowly growing tumor .
it occurs preferentially in the hand and foot area of adults , and the development in the auricle is very rare .
we report a rare case of chondroma that developed in the auricle , which were treated with wedge excision and reconstructed using a burow 's triangle , with a review of the literature . |
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. zone of depigmentation around primary cutaneous melanoma portends a bad prognosis whereas occurrence of vitiligo like lesions in patients with metastatic melanoma has been reported to have a better prognosis . most skin metastases of melanoma are located in the reticular dermis and are not difficult to differentiate from a primary melanoma , however epidermotropic metastasis poses a significant difficulty on histopathology to distinguish it from a new primary lesion . the concept of epidermotropic metastatic malignant melanoma ( emmm ) was developed in the 1970 's thereby which distinguishing the primary from epidermotropic metastasis became important , since recognition of this histopathologic variant catapults the disease from stage 1 to stage iv which has a bad prognosis . lesions of melanoma are known to show the occurrence of vitiligo like lesions at the site of regressing primary or at distant sites but rarely around epidermotropic secondaries . a 78 year old male presented to us with complaints of multiple , small , dark colored raised lesions over the left leg which were gradually increasing in size and number over a period of two months . a few of them occasionally ulcerated , bled , discharged pus and were associated with pain . the patient also complained of multiple asymptomatic swellings in left groin region since one month . on enquiry , he gave history of single , small , dark colored pea sized raised lesion on the sole of left foot which over a period of two years gradually increased in size to ulcerate and form a non healing ulcer which would bleed occasionally . complete excision with skin grafting of the lesion was done at his hometown six months prior and was reported as acral lentiginous malignant melanoma on histopathological examination . he gave no history of abdominal pain , bony pain , vomiting , chest pain , convulsions or breathlessness . he had been operated for varicosities of left leg three years back and was a known hypertensive on treatment since two years . clinical examination of the patient revealed multiple , soft , friable , well defined , jet black colored , firm papulonodular lesions ranging in size from 3 mm to 6 cm in dimension . a single well defined dark colored plaque of 5 6 cm was also seen on the heel of the left sole [ figure 2 ] . the left sided inguinal region showed three enlarged lymph nodes which were mobile , 2 - 3 cm sized , hard and non tender . multiple , well defined , jet black coloured , firm papulonodular lesions with surrounding rim of depigmentation extending from dorsa of left foot up to the knee single well defined dark coloured plaque seen on the heel of the left sole complete blood count showed mild anemia ( hb-12.3 gms / dl ) . platelet count , total leukocyte count and differential leukocyte count was normal except for mild eosinophilia . biopsy done from a jet back papule revealed an asymmetric melanocytic neoplasm with irregular distribution of melanocytes and nests of variable size . there was epidermal flattening overlying the dermal nests of atypical melanocytes extending up to mid reticular dermis [ figure 3 ] . neoplastic melanocytes showed mild to moderate atypia with occasional mitotic figures with haphazard pigment distribution in both normal and atypical melanocytes . several neoplastic melanocytes were present within some of the dilated capillaries in the deep dermis suggestive of angioinvasion . ( a ) biopsy done from a jet back papule on hematoxylin and eosin stain shows an asymmetric melanocytic neoplasm with irregular distribution of nests of variable size consisting of atypical melanocytes extending upto mid reticular dermis . ( h and e , 5 ) , ( b and c ) neoplastic melanocytes showed mild to moderate atypia with occasional mitotic figures with haphazard pigment distribution in both normal and atypical melanocytes . ( h and e , 10 ) , ( d ) neoplastic melanocytes are seen within the dilated capillary in the deep dermis . ( h and e , 20 ) biopsy from a papule showing peripheral rim of depigmentation revealed melanocytic neoplasm made up of irregular shaped and sized nests within the mid and upper dermis with numerous solitary melanocytes scattered throughout the surrounding epidermis including the stratum corneum with epidermal flattening . moderately dense superficial perivascular lymphocytic and plasma cellular infiltrate underlying the dermal nests were seen in addition which were responsible for the depigmented halo clinically [ figure 4 ] . ( a ) biopsy from a papule showing peripheral rim of de - pigmentation revealed melanocytic neoplasm with flattened epidermis made up of nests of neoplastic melanocytes in upper dermis and within the dilated capillaries in the deep dermis . ( h and e , 5 ) , ( b ) irregular shaped small nests of atypical melanocytes at dermo - epidermal junction with epidermal flattening overlying the dermal nests . ( h and e , 10 ) , ( c ) moderately dense superficial peri - vascular lymphocytic and plasma cellular infiltrate underlying the dermal nests . ( h and e , 10 ) , ( d ) irregular shaped small nests at dermo - epidermal junction with numerous solitary melanocytes scattered throughout the surrounding epidermis including the stratum corneum . ( h and e , 20 ) ultrasonography of abdomen and pelvis showed normal sized liver with multiple hypo echoic lesions involving both lobes , measuring 2.3 by 2.2 cm in size suggestive of secondary metastatic deposits . reports of positron emission tomography ( pet ) revealed presence of metastatic deposits in the lungs , liver , spleen , left para - aortic group of lymph nodes , left inguinal group of lymph nodes and in the subcutaneous tissue over left thigh region [ figure 5 ] . computed tomography of the brain was normal . pet shows metastatic deposits in the lungs and left para - aortic group of lymph nodes , left inguinal group of lymph nodes on co - relating the above findings we reached a diagnosis of epidermotropic metastatic melanoma stage iv ( t2n3m1c ) with peri - metastatic depigmentation . the patient , however refused further treatment and succumbed to death over a span of two months . one of the rare forms of melanoma metastasis is the epidermotropic metastasis which poses a significant difficulty on histopathology in its distinction not only from a new primary melanoma but also from residual incompletely excised primary melanoma . it may be seen on histopathology to include the dermo - epidermal junction , extend into the epidermis with intra - epidermal component not beyond the dermal component and as an epidermal - only pattern . this distinction has a great bearing on the staging and prognosis since staging of multiple primary melanomas is based on the primary lesion with the worst prognostic features , whereas a patient with a distant cutaneous metastasis catapults to stage iv disease with an approximate 10% five year survival rate . in 1978 , kornberg for the first time proposed histopathologic characteristics for differentiating epidermotropic metastatic melanoma from a primary melanoma . these features were : ( 1 ) thinning of the epidermis by the underlying aggregates of atypical melanocytes in the dermis , often in association with widened dermal papillae and an epidermal collarette at the periphery of the tumor ( 2 ) atypical melanocytes present within endothelial lined spaces , and ( 3 ) a zone of atypical melanocytes in the dermis equal to or broader than the epidermal component . however , over a period of time , multiple reports have described the varied histopathologic presentations of epidermotropic metastatic melanomas and proposed additional criteria . such as angiotropic nature of metastasis , maturation and other histologic features considered by some workers to be present in a metastatic lesion are localization of the metastatic atypical melanocyte nests in the superficial dermis , the presence of evident inflammatory infiltrate , presence of monomorphic , monoclonal malignant cells and occurrence of small sized lesions . studies on melanoma and its metastasis have shown that loss of heterozygosity ( loh ) on short arm of chromosome 9 and long arm of chromosome 10 i.e. , ( 9p and 10q ) is associated with early - stage primary disease , whereas loh on 6q , 1p , 8q , and 11q has been associated with its progression and metastasis . loh at 10q was found most frequently and in higher proportion in the epidermotropic metastatic melanomas . however , variable patterns within the same case indicate that , while some epidermotropic metastatic melanomas are clonally related to the primary , others are of independent origin . four patterns of hypomelanosis around melanomas have been observed such as i ) areas of depigmentation confined to primary lesion suggestive of a regressed lesion ii ) halos of depigmentation around the primary tumour iii ) a primary tumour with co - existent halo nevi i.e. , leukoderma around benign nevocytic nevi and iv ) wide spread hypomelanosis at sites distant from primary site . the postulated mechanism is the occurrence of immune response leading to circulating anti - melanocyte antibodies formation . these are capable of attracting natural killer cells which cause vitiligo by attacking and destroying normal melanocytes . becker et al . demonstrated that the cells infiltrating the depigmented skin were identical to the t lymphocytes infiltrating the adjacent tumor , indicating that the same cells responsible for tumor reduction were also responsible for depigmentation . in case of vitiligo and melanoma - related depigmentation , studies suggest cd8 + t cell reactivity to be directed against the melan - a / melanoma associated antigen recognized by t - cells ( mart-1 ) epitope 27 - 35 in hla - a2 patients . in mmm associated with widespread vitiligo the 5-year survival rate is 60% , and survival up to 29 years has been documented . thus , the presence of vitiligo may delay the inevitable outcome but does not eradicate the malignancy . the favourable influence of vitiligo on the prognosis of mmm does not seem to be related to whether the depigmentation develops before or after diagnosis of the malignant disease . the process of hemo - lymphatic spread of the atypical melanocytes of the primary lead to cryptic exposure of the tumor antigens eventuating in a t - cell mediated immune response of the body which manifested as peri - lesional depigmentary changes around the secondary metastatic deposits in our case . however , the occurrence of t - cell mediated immune response did not seem to confer any protection against metastatic spread of the disease . hence in cases with multiple lesions of melanoma , the clinician needs to be on a constant look out to distinguish multiple primary from the rare epidermotropic metastasis . the occurrence of vitilgo like changes around primary lesions have been reported to confer better survival chances in patients , however its occurrence around secondaries did not seem to confer any prognostic advantage in our case . t - cell mediated immune response against tumor antigens manifesting as peri - lesional depigmentation around metastatic deposits does not seem to confer favourable prognosis . | melanoma is a rare form of cutaneous malignancy encountered in the dark skin population .
epidermotropic metastatic melanoma is a rare form of cutaneous metastatic melanoma which can mimic primary melanoma on histopathology .
hence its differentiation is of immense prognostic importance .
the occurrence of rim of depigmentation around the primary cutaneous melanoma has previously been reported to portend a bad prognosis .
the occurrence of vitiligo like lesions in patients with metastatic melanoma in comparison has a better prognosis .
however the occurrence of depigmentation around the secondaries is rare and its importance is not well known .
hence we wish to report a case of epidermotropic metastatic melanoma with perilesional depigmentation in a 78 year old indian male . |
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purple urine bag syndrome ( pubs ) was first reported in 1978 ( barlow and dickson 1978 ) . the patient had a purple - colored urine bag following urinary catheterization for hours to days ( stott et al 1978 ; dealler et al 1988 ; al - jubouri and vardhan 2001 ; lin et al 2002 ) . although the definite chemical substrate of pubs is unknown , most authors believe it is a mixture of indigo ( blue ) and indirubin ( red ) that becomes purple ( dealler et al 1988 ; matsuo et al 1993 ; umeki 1993 ; nakayama and kanmatsuse 1997 ; ishiha et al 1999 ; al - jubouri and vardhan 2001 ; ribeiro et al 2004 ) . the chain reaction responsible for pubs began with tryptophan from the food chain being metabolized by gut bacteria . this metabolic process produces indole , which is absorbed into portal circulation and converted to indoxyl sulphate in the liver , after a series of detoxification transformations ( figure 1 ) ( chiou et al 2005 ) . in this study , we collected 10 elderly patients with pubs cared for by the senior registered nurses at two nursing homes in southern taiwan community hospitals ( table 1 ) . case 1 and case 2 who were identified by chart review in 1987 and 2003 , respectively , and the latter 8 cases ( 3 males , 5 females ) were collected between january 2007 and june 2007 . the two nursing home had 74 residents ( 52 males , 22 females ) , of which 19 ( 8 males , 11 females ) had undergone foley catheterization . the incidence of pubs was 42.1% among the 19 urinary catheterized nursing home residents in the follow - up half - year period . case 1 , a 72-year - old male with pubs , was found in 1987 . he was a patient with type 2 diabetes mellitus ( dm ) , diabetic nephropathy ( dn ) , and end - stage renal disease ( esrd ) , treated with hemodialysis for 3 years . he was lost to follow - up after being transferred to a medical hospital due to a medical illness in 1990 . case 2 , a 72-year - old male , was a patient with type 2 dm , diabetic nephropathy , and esrd , treated with hemodialysis . case 3 , an 83-year - old male , had had dm for about 15 years , alzheimer s dementia ( ad ) for 2 years , and hypertension treated with amlodipine besylate ( norvasc ) 5 mg per day for about 10 years . the patient had a purple - colored urine bag 23 days following urinary catheterization or changing of the foley off and on for more than 24 months ( figure 2 ) . case 4 , an 89-year - old male , had been a patient with dm and diabetic nephropathy for 40 years , esrd treated with hemodialysis for 20 years , alzheimer s dementia for about 5 years , and hypertension treated with furosemide ( lasix ) 40 mg per day for 20 years . he underwent foley catheterization due to benign prostate hypertrophy ( bph ) and had suffered from pubs off and on for more than 33 months , before being transferred to this nursing home ( figure 3 ) . case 5 , an 80-year - old male , had been a patient with vascular dementia ( vad ) with delirium since 2 years ago . he became long - term bedridden because of fracture of the left femoral neck about 3 years ago . he regularly took doxazosin mesylate ( doxaben ) 1 mg per day for hypertension and bph , and quetiapine fumarate ( seroquel ) 100 mg per day for vascular dementia with delirium . case 6 , an 80-year - old female , was a patient with alzheimer s dementia , anemia , hepatitis , hypercholestrolemia and pulmonary fibrosis . case 7 , a 76-year - old female , had been a patient with vad and anemia since 10 years ago . case 8 , a 66-year - old female , was a patient with hypertension , hypercholesterolemia , and vad with delirium . she took amlodipine besylate ( norvasc ) 10 mg per day for hypertension , and melitracen 10 mg / flupentixol 0.5 mg ( deanxit ) 1 tab per night for vad with delirium . she became long - term bedridden and received foley catheterization for 4 years after a stroke . case 9 , a 60-year - old female , was a patient with schizophrenia and poliomyelitis . she regularly took melitracen 10 mg / flupentixol 0.5 mg ( deanxit ) 2 tab per night and lorazepam ( ativan ) 0.5 mg per night . case 10 , a 75-year - old female , had been a patient with dm , hypertension , vad , and hypercholestrolemia for 6 years . she had taken glipizide ( minidiab ) 5 mg per day for dm , and nifedipine ( adalat oros ) 30 mg per day for hypertension . she had been long - term bedridden for 6 years after a stroke , and had received foley catheterization for 1 year . the mean age of the 10 elderly patients ( 5 males , 5 females ) was 75.30 years ( sd 8.39 , range 6089 ) . they were long - term bedridden for an average of 4.45 years ( sd 2.43 , range 1.510 ) , and received urinary catheterization for an average of 35.9 months ( sd 18.66 , range 1272 ) . only about 30.0% of the patients had had long - term constipation . their vital signs were all stable even though their urine routine showed urinary tract infection ( uti ) . among the ten cases , 9 ( 90.0% ) had dementia ; 80% of the male cases also had diagnoses of dm , diabetic nephropathy , and esrd with hemodialysis . therefore pubs had a higher incidence in patients with chronic disease such as dementia , diabetic nephropathy , a long - term bedridden state , and long - term foley catheterization . escherichia coli , klebsiella pneumoniae , providencia rettegeri , and proteus mirabilis were revealed in their urine culture . their blood routine showed white blood cells , 3800~8300/l , and hb , 8.6~14.1 g / dl . seven cases ( 87.5% ) had mild anemia . in the biochemistry examinations , seven cases ( 87.5% ) revealed normal renal functioning , except case 4 with diabetic nephropathy with esrd . all patients had stable vital signs , and had no infection sign such as leukocytosis , fever , or chill was found , even though pubs and bacteriuria persisted . we decreased their duration of catheterization , replaced their foley and urine bag regularly , and improved their quality of catheter care . pubs is rarely reported : only one larger survey study among the 23 papers regarding pubs published since 1987 was found in a pubmed search . although the exact prevalence of pubs is unknown , but it may not be rare because the prevalence of pubs was found to be 9.8% ( seven of 71 ) in the study of dealler and colleagues ( 1988 ) and 8.3% ( 13 of 157 ) at a long - term care service center in a taiwan community hospital ( su et al 2005 ) . in this report , the prevalence of pubs was as high as 42.1% ( eight of 19 ) in urinary catheterized nursing home residents . we need a larger study for further evaluation of pubs in taiwan nursing home in the future . a previous study showed that pubs occurred predominantly in chronically catheterized , constipated women ( su et al 2005 ) . no previous positive association study between pubs and medications , such as norvasc , lasix , doxaben , adalat oros , minidiab , seroquel , deanxit , or stilnox has been reported before . pubs was found to be associated with alkaline urine as well as uti induced by some species of bacteria with indoxyl sulphatase / phosphatase ( ribeiro et al 2004 ) . but in this report , the four cases at the first nursing home were all male and were nursing home residents since 1987 . this is similar to the viewpoint of lin and colleagues ( 2002 ) . in urine culture , several bacterial species have been reported in association with pubs including providencia stuartii , p. rettgeri , k. pneumoniae , p. mirabilis , e. coli , morganella morganii , and pseudomonas aeruginosa ( dealler et al 1988 ; matsuo et al 1993 ; umeki 1993 ; nobukuni et al 1995 ; ishiha et al 1999 ; al - jubouri et al 2001 ; lin et al 2002 ; zotti et al 2004 ) . our urine culture results were similar to those of previous reports , and revealing e. coli , k. pneumoniae , p. rettegeri , and p. mirabilis . p. mirabilis compromises the care of many patients undergoing long - term indwelling bladder catheterization . in a prevalence study of hospital - acquired infections ( hai ) , a significant correlation was found with major risk factors related to medical procedures , such as urinary catheterization ( zotti et al 2004 ) . a failure to decrease the unnecessary bladder catheter use and the duration of catheterization would result in poor quality of care ( landi et al 2004 ) , a greater mean length of stay in hospital , and a higher risk of nosocomial uti ( zotti et al 2004 ) . it was apparent that pubs affected not only the urine bags but also indwelling catheters . the urinary catheter drainage system changed color from red to purple and sometimes showed differently colored tubes and bags , as in case 3 with pubs ( figure 2 ) . we found that the longer the pvc bag stayed in use for a patient , the deeper the color purple the bag became , despite changing sterile pvc urine bags from 2 different companies , which was compatible with the findings of dealler and colleagues ( 1988 ) and su and colleagues ( 2005 ) . in conclusion , the pigments associated with pubs have been well shown to be due to indirubin and indigo . the pathogenesis of pubs is due to the metabolism of tryptophan by bacteria to indole and later converted to indicant in the liver ( dealler et al 1988 ) . this is excreted and in the urine broken down by bacteria possessing one or both enzymes , sulphatase and phosphatase that metabolize this pigment to indirubin and indigo in an alkaline environment ( urine ) . the indigo can also be precipitate in the catheter itself giving a blue discoloration ( harun et al 2007 ) . in this paper , we found that pubs seemed to have a higher incidence in patients with a bedridden status , alkaline urine , and species of bacteria with indoxyl sulphatase / phosphatase , but possibly not associated with gender , constipation , or type of pvc urine bag used . we also presented that pubs was a harmless , but alarming problem ( robinson 2003 ) . it is unnecessary to aggressively treat patients with pubs , but the control of urological sanitation is fundamental to treatment . in taiwan , geriatric medicine , nursing home care , and hai control have become very important because nosocomial catheter - associated utis comprise a huge silent reservoir of antibiotic - resistant bacteria and yeasts ( tambyah and maki 2000 ; robinson 2003 ; hampton 2004 ) . thus , we should decrease the duration of catheterization , improve catheter care , and deploy technological advances designed for prevention . | purple urine bag syndrome ( pubs ) is a rare occurrence , in which the patient has a purple - colored urine bag following urinary catheterization for hours to days .
most of authors believe it is a mixture of indigo ( blue ) and indirubin ( red ) that becomes purple .
previous study showed that pubs occurred predominantly in chronically catheterized , constipated women .
we collected 10 elderly patients with pubs in two nursing homes .
the first two cases were identified by chart review in 1987 and 2003 , and then later eight cases ( 42.1% ) were collected among 19 urinary catheterized elderly in the period between january 2007 and june 2007 . in the present report , pubs probably can occur in any patients with the right elements , namely urinary tract infection ( uti ) with bacteria possessing these enzymes , diet with enough tryptophan , and being catheterized .
associations with bed - bound state , alzheimer s , or dementia from other causes are reflections of the state of such patients who are at higher risk for uti , and hence pubs occurred .
although we presented pubs as a harmless problem , prevention and control of the nosocomial catheter - associated utis ( cautis ) has become very important in the new patient - centered medical era .
thus , we should decrease the duration of catheterization , improve catheter care , and deploy technological advances designed for prevention , especially in the elderly cared for in nursing homes . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
tb genotype clusters were defined as groups of > 3 tb case - patients whose isolates had matching spoligotyping and 12-locus mycobacterial interspersed repetitive unit variable number tandem repeat ( miru - vntr ) ( 9 ) genotyping results that were reported in the same county within washington . a log - likelihood ratio ( llr ) was calculated for each genotype cluster identified during each of the two 3-year periods ( figure ) . the larger the llr , the greater the possibility the cluster represented geographically concentrated tb cases , a proxy for recent tb transmission . the cutoff point for the llr was set to 5.0 , based on the value used by the national tb genotyping information management system ( 10 ) . formula used to calculate geospatial statistic ( a modified log - likelihood ratio [ llr ] ) on the basis of geographic distribution of mycobacterium tuberculosis genotype clusters , washington , usa . variables are classified as follows : a = number of tuberculosis ( tb ) cases with the genotype of interest in the selected county ; b = number of cases with the genotype of interest in the united states ; c = number of cases without the genotype of interest in the selected county ; d = number of cases without the genotype of interest in the united states ; n = total number of tb cases . qualitative analysis came from a 5-member expert panel of tb public health officials in washington . in 2008 , the panel participated in a discussion of all county - level tb clusters , ranking each as high or low priority for additional investigation . priority was determined on the basis of a review of patient characteristics , epidemiologic links from field investigations , and maps of genotype distributions . the panel also had information from enhanced contact investigations from local public health investigation teams that included the ability to order is6110 restriction fragment - length polymorphism ( is6110 rflp ) and 24-locus miru - vntr testing for clusters of concern , but results from these tests were not universally available . the ranking exercise with the same 5-member panel was repeated after period 1 for clusters from period 1 . llrs were compared with the expert opinion ranking to assess concordance . with expert opinion as the standard , negative and positive predictive values ( npv and ppv , respectively ) were calculated for period 1 using a cutoff point of llr > 5.0 . sensitivity and specificity of the > 5.0 llr cutoff point and exact binomial 95% cis were calculated for period 1 clusters . of 659 culture - positive cases , 642 ( 97.4% ) had genotyped isolates ; of these , 318 cases formed 21 clusters . five of these clusters had a high llr ; the expert panel ranked all 5 of these clusters high priority and identified them as clusters of concern . of the 16 clusters with llr < 5.0 , the expert panel ranked 12 ( 75.0% ) as low priority ( table ) . * llrs > 5.0 ( boldface ) indicate greater possibility that the cluster represents geographically concentrated tb cases , a proxy for recent tb transmission ) . variable number tandem repeat ; llr , log - likelihood ratio , a measure of geospatial concentration ; h , homelessness ; fb , foreign born ; sa , substance abuse . period 1 , january 1 , 2005december 31 , 2007 ; the expert panel reexamined the 21 clusters identified during period 1 and focused on new activity within those areas during period 2 . two clusters with a high llr during period 1 continued to have a high llr during period 2 ; the expert panel continued to rank these high priority . the 3 other clusters that had a high llr during period 1 had a low llr for period 2 ; one of those was still considered a high priority by the panel . of the remaining 16 clusters , which continued to have a llr < 5 , 14 ( 87.5% ) were still ranked low priority by the panel . two clusters in the same county , pcr00309 and pcr00803 , had low llrs but were considered high priority by the expert panel . for cluster pcr00309 , the panel cited high levels of homelessness among case - patients as reason to rank it high priority . for cluster pcr00803 , the panel cited a highly mobile population from a tb - endemic country that regularly traveled into and out of the united states as reason to rank it high priority . however , the travel history among case - patients in this cluster made it difficult for investigators to determine whether transmission was occurring within washington or abroad . the npv and ppv for a llr cutoff point of 5.0 were 75% and 100% , respectively . lowering the cutoff point to a llr > 2.0 increased the npv to 92.3% , but the ppv remained at 100% . for period 1 clusters only , a cutoff point of llr > 5.0 generated a sensitivity of 55.6% ( 95% ci 21.2%86.3% ) and specificity of 100% ( 95% ci 73.5%100.0% ) for identifying clusters for further investigation . decreasing the cutoff point to > 2.0 increased the sensitivity to 88.9% ( 95% ci 51.8%99.7% ) but did not change the specificity ( 100% ; 95% ci 73.5%100.0% ) . the geospatial statistic in this study was highly correlated with experts perceived need for public health action . this finding indicates that automated alerts generated on the basis of geospatial concentration of tb cases might help the state tb program identify clusters that would benefit from additional investigation . automated alerts can be generated by using routinely collected surveillance data and are currently part of the national tb genotyping information management system ( 10 ) . patient and contact characteristics , transmission venues , and temporality all contribute toward prioritization determination . for example , during period 1 , a total of 6 ( 66.6% ) clusters ranked high priority by the expert panel were characterized by homelessness or substance abuse among case - patients , and 8 ( 88.9% ) were characterized by us - born case - patients ( table ) . conversely , 11 ( 91.7% ) clusters ranked low priority were characterized by case - patients who were foreign - born , a known risk factor for latent tb infection ( 7 ) . none of the period 1 clusters with llr > 5 and only 1 of 9 clusters ranked as high priority by the expert panel were characterized by foreign - born case - patients . these results indicate the need for further study to identify the limitations of the llr score in detecting localized and recent tb transmission among foreign - born case - patients . the availability of is6110 rflp or 24-locus miru - vntr testing results to the expert panel is the current standard for fieldwork and could have introduced an information bias for the panel in this study . although this effect is unknown , lack of universal is6110 rflp and 24-locus miru - vntr test results is a limitation of this study . we found that geospatial statistics based on tb genotyping and surveillance data could help identify and prioritize likely recent disease transmission events in washington . in addition , llr values should be incorporated into ongoing evaluation by the expert panel ; in fact , llr is now included in routine genotype and cluster reviews . geospatial statistics are an attractive approach to prioritization , but additional field - based research is needed to assess whether factors such as epidemiologic characteristics could be used to further develop a prioritization algorithm . integrating these factors and determining ideal cutoff points in different settings will increase predictive value . | groups of tuberculosis cases with indistinguishable mycobacterium tuberculosis genotypes ( clusters ) might represent recent transmission .
we compared geospatial concentration of genotype clusters with independent priority rankings determined by local public health officials ; findings were highly correlated .
routine use of geospatial statistics could help health departments identify recent disease transmission . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
the 5 terminal structure of eukaryotic rna polymerase ii transcripts ( rna 5 cap ) plays a crucial role in gene expression and regulation . the cap is specifically bound to several cellular and viral proteins , including various isoforms of eukaryotic translation factor eif4e , nuclear cap - binding complex cbc , dcps scavenger enzyme , poly(a ) binding protein pabp , poly(a)-specific ribonuclease parn , pokeweed antiviral protein pap , cellular mrna cap ( guanine - n7 ) methyltransferase , human parneoplastic encephalomyeltis antigen hud , vaccinia virus 2-o - methyltransferase vp39 , influenza virus rna polymerase , and dimethyltransferase tgs1 . the eif4e factors from vertebrates and yeast were shown to be highly selective for 7-methylguanosine cap ( mmg - cap ) , mgpppn , n = g , a , u or c. in the nematodes caenorhabditis elegans and ascaris suum as well as in the parasitic flatworm schistosoma mansoni a high population of messenger ribonucleic acids ( mrnas ) contain a hypermethylated cap form , n , n,7-trimethylguanosine cap ( tmg - cap ) , m32,2,7gpppn , which is acquired along with a spliced leader during trans - splicing of pre - mrna . affinity chromatography [ 14 , 15 ] , and fluorescence titration [ 16 , 17 ] experiments showed that three out of five c. elegans eif4e isoforms , ife-1 , ife-2 , ife-5 , are capable of binding specifically to the mmg - cap and to the tmg - cap . two other isoforms , ife-3 , most similar to mammalian eif4e , and ife-4 , related to the mammalian 4e - homologous protein 4e - hp , bind only to the mmg - cap . the dual binding specificity was also observed for eif4es from s. mansoni and a. suum . values of the equilibrium dissociation constants , kd , for some selected complexes of the dual specificity eif4es with typical mmg - cap and tmg - cap analogues are shown in table 1 . the values reported by various groups and derived by various titration methods , mainly fluorescence and isothermal calorimetry , can differ between each other . nevertheless , the experimental data clearly show that both c. elegans ife-3 and murine eif4e strongly discriminate in favor of mmg - cap , ca . 11 kj mol and 16 kj mol , respectively , while in the case of c. elegans ife-5 the binding energies are similar . table 1experimentally derived dissociation constants , kd [ m ] , for the mmg - and tmg - cap analogues from c. elegans factors , ife-3 and ife-5 , and s. mansoni eif4e . nomgtpm32,2,7gtpeif4e murine0.016.99ife-3 c.elegans0.2016.70.363.5ife-5 c.elegans0.140.330.540.92eif4e s.mansoni0.5811.2 experimentally derived dissociation constants , kd [ m ] , for the mmg - and tmg - cap analogues from c. elegans factors , ife-3 and ife-5 , and s. mansoni eif4e . the kd values for murine eif4e are shown for comparison the tmg - cap occurs at the 5 terminus of small nuclear rna ( snrna ) , small nucleolar rna ( snorna ) and in telomerase rna tlc1 . it is specifically recognized by snurportin1 , a receptor for spliceosomal small nuclear particles ( snrnps ) . as shown by x - ray crystallography [ 9 , 12 , 2129 ] and multidimensional nmr most of the cap - binding proteins converged at a common mechanism of the cap recognition via stacking of the 7-methylguanine moiety in between two aromatic amino acid side chains . the 7-methylguanine base possesses a net positive charge , which seems indispensable for its proper recognition , i.e. , the 7-methylguanosine can not be replaced by guanosine in the cap structure . in the snurportin1 complex with the dinucleotide tmg - cap analogue the sandwich stacking involves one tryptophan , and two bases of the cap , the first one , trimethylated , and the second one , unmethylated . in dimethyltransferase tgs1 the 7-methylguanine moiety is stacked only on one tryptophan and a serine polar side chain limits the binding pocket on the opposite side . except for the cation- stacking , the cap is also stabilized by a network of hydrogen bonds , direct or water mediated salt bridges , as well as less specific van der waals and hydrophobic contacts . only a few exceptions have been found yet where the recognition specificity is entirely mediated through hydrogen bonds and van der waals contacts with 7-methylguanine , i.e. , cap methyltransferase , and reovirus polymerase 3 . in the mammalian , plant , and yeast eif4e - cap complexes , two tryptophan aromatic residues take part in the sandwich cation- stacking with the 7-methylguanine moiety . two hydrogen bonds involve n1h and nh atoms of 7-methylguanosine and the carboxyl group of a conserved glutamic acid , and one hydrogen bond is observed between o of mg and the backbone amide nitrogen . additional stabilizing interactions are between the phosphate chain of the cap and the arginine / lysine side chains of the protein . the first crystallographic structure of dual specificity eif4e from schistosoma mansoni in the complexes with two dinucleotide mmg - cap analogues showed a similar binding mode as for the single specificity eif4es . the only difference seems to be in the conformation of e90 carboxylate of s. mansoni eif4e that is rotated by 80 in comparison to the orientation of the equivalent e103 [ 12 , 23 ] in murine eif4e . a similar rotation was also observed in the human eif4e ternary complex with eif4gi peptide and a glycerol molecule located in the cap - binding center . still , the contribution of that conformational change to mmg - cap vs. tmg - cap binding specificity remains unclear . the nmr analysis of the mmg - cap and tmg - cap complexes with s. mansoni eif4e showed substantial chemical shift perturbation for ca . based on the crystallographic and nmr data the authors suggested that intrinsic and specific conformational flexibility of the s. mansoni eif4e plays a crucial role in the tmg - cap binding , analogous to an combined mutagenesis studies and molecular dynamics simulations of c. elegans dual specificity ife-5 led to a structural rather than a dynamic model . larger width and depth of the cap - binding pocket was postulated to be responsible for the tmg - cap binding specificity . / v65l decreased the size of the pocket and gave rise to discrimination against tmg - cap by steric hindrance . however , it was noted that dual specific a. suum eif4e does contain y64 and l65 residues . unfortunately , discrimination between tmg - cap and mmg - cap by snurportin1 is based on a mechanism that differs from that expected for the eif4e factors [ 20 , 33 ] . in order to get an insight into the mechanism of dual specificity in the cap recognition by some of highly homologic eif4es , we performed long - lasting molecular dynamics ( md ) simulations in water for three selected eif4e homologues , murine eif4e as well as c. elegans ife-3 and ife-5 , each of them in the apo form and in the complexes with mgdp or with m32,2,7gdp ( scheme 1 ) . the results point to a dynamic mechanism of discrimination between the mono - and hypermethylated cap structures . ( a ) mgdp , analogue of mmg - cap , ( b ) m32,2,7gdp , analogue of tmg - cap chemical structures of the cap analogues . ( a ) mgdp , analogue of mmg - cap , ( b ) m32,2,7gdp , analogue of tmg - cap the starting structure of the complex of truncated murine eif4e(28217 ) with mgdp for molecular dynamics simulations was taken from crystallography ( pdb code : 1ej1 ; ) . the missing atoms of some of the amino acid side chains were completed by scwrl . the starting structures of ife isoforms were obtained by homology modeling with murine eif4e(28217 ) bound to mgdp as a template , with 51% and 42% sequence identity to ife-3 and ife-5 , respectively . additional harmonic constraints were introduced for the distances between the protein and the cap atoms that were engaged in hydrogen bonds , salt bridges and van der waals contacts . subsequently , the resulting structures were subjected to detailed analysis regarding packing of the residues , steric hindrance , and loop conformations . due to high sequence homology the modeled ife structures were very similar to that of the eif4e template ( fig . 1 ) , especially regarding the main polypeptide chains . the structures of the complexes of three 4e factors with m32,2,7gdp , and of murine eif4e with gdp , were constructed by adding two methyl groups at n and by removing the methyl group from n7 in the protein - mgdp complexes , respectively . 1structural comparison of murine eif4e(28217 ) and its two c. elegans isoforms , ife-3 and ife-5 . ( b ) superposition of the three protein complexes with mgdp . the cap and the amino acids engaged in its stabilizing are marked in bold structural comparison of murine eif4e(28217 ) and its two c. elegans isoforms , ife-3 and ife-5 . the cap and the amino acids engaged in its stabilizing are marked in bold the esp charges of the isolated ligands were calculated at hf/631g(d , p ) level using gaussian 94 ( gaussian inc . , pittsburg pa , u.s.a . ) the md simulations were carried out by the program sigma using charmm22 force field . each protein or complex was subjected to energy minimization without electrostatic interaction and immersed in an equilibrated tip3p water box keeping at least 10 shell thickness from the protein surface . the simulation procedure consisted of several equilibration md runs preceded and followed by 500-step energy minimization , and a subsequent regular md run , as follows . first , energy minimization and 48 ps dynamics of water molecules was performed keeping the protein or the complex immobilized . second , energy minimization of the protein or of the complex with the water molecules kept fixed was followed by stepwise heating of the whole system from 50 k to 300 k for 82.56 ps . the equilibration md runs were performed in the nvt ensemble and the regular md simulations were performed in the npt ensemble at temperature t = 300 k and pressure p = 1 atm . the electrostatic interactions were calculated by multiple time step with a double cut - off at 6 and 10 . short- , middle- , and long range interactions , according to particle - mesh ewald method [ 43 , 44 ] , were calculated for an integration time - step of 2 , 4 , and 12 fs , respectively . the simulations were run for 5 ns in the case of the apo proteins or for 10 ns in the case of the complexes , in order to reach at least partial equilibrium according to a stability criterion for the fluctuations of root - mean - square - deviation ( rmsd ) of the proteins c atoms . the conformations of the solute on a simulated trajectory were written down every 0.96 ps and analyzed regarding interatomic distances and torsion angles . essential dynamics ( ed ) analysis of selected , equilibrium parts of the md trajectories was performed according to amadei et al . . the starting structure of the complex of truncated murine eif4e(28217 ) with mgdp for molecular dynamics simulations was taken from crystallography ( pdb code : 1ej1 ; ) . the missing atoms of some of the amino acid side chains were completed by scwrl . the starting structures of ife isoforms were obtained by homology modeling with murine eif4e(28217 ) bound to mgdp as a template , with 51% and 42% sequence identity to ife-3 and ife-5 , respectively . additional harmonic constraints were introduced for the distances between the protein and the cap atoms that were engaged in hydrogen bonds , salt bridges and van der waals contacts . subsequently , the resulting structures were subjected to detailed analysis regarding packing of the residues , steric hindrance , and loop conformations . due to high sequence homology the modeled ife structures were very similar to that of the eif4e template ( fig . 1 ) , especially regarding the main polypeptide chains . the structures of the complexes of three 4e factors with m32,2,7gdp , and of murine eif4e with gdp , were constructed by adding two methyl groups at n and by removing the methyl group from n7 in the protein - mgdp complexes , respectively . 1structural comparison of murine eif4e(28217 ) and its two c. elegans isoforms , ife-3 and ife-5 . ( b ) superposition of the three protein complexes with mgdp . the cap and the amino acids engaged in its stabilizing are marked in bold structural comparison of murine eif4e(28217 ) and its two c. elegans isoforms , ife-3 and ife-5 . the cap and the amino acids engaged in its stabilizing are marked in bold the esp charges of the isolated ligands were calculated at hf/631g(d , p ) level using gaussian 94 ( gaussian inc . , pittsburg pa , u.s.a . ) the md simulations were carried out by the program sigma using charmm22 force field . each protein or complex was subjected to energy minimization without electrostatic interaction and immersed in an equilibrated tip3p water box keeping at least 10 shell thickness from the protein surface . the simulation procedure consisted of several equilibration md runs preceded and followed by 500-step energy minimization , and a subsequent regular md run , as follows . first , energy minimization and 48 ps dynamics of water molecules was performed keeping the protein or the complex immobilized . second , energy minimization of the protein or of the complex with the water molecules kept fixed was followed by stepwise heating of the whole system from 50 k to 300 k for 82.56 ps . the equilibration md runs were performed in the nvt ensemble and the regular md simulations were performed in the npt ensemble at temperature t = 300 k and pressure p = 1 atm . the electrostatic interactions were calculated by multiple time step with a double cut - off at 6 and 10 . short- , middle- , and long range interactions , according to particle - mesh ewald method [ 43 , 44 ] , were calculated for an integration time - step of 2 , 4 , and 12 fs , respectively . the simulations were run for 5 ns in the case of the apo proteins or for 10 ns in the case of the complexes , in order to reach at least partial equilibrium according to a stability criterion for the fluctuations of root - mean - square - deviation ( rmsd ) of the proteins c atoms . the conformations of the solute on a simulated trajectory were written down every 0.96 ps and analyzed regarding interatomic distances and torsion angles . essential dynamics ( ed ) analysis of selected , equilibrium parts of the md trajectories was performed according to amadei et al . . an experimentally observed equilibrium association constant kas expressed in terms of the molar concentrations of the reactants in a protein - ligand association is related to the standard gibbs free energy g of the association process at temperature t , \documentclass[12pt]{minimal }
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\begin{document}$$ \delta { \hbox{g}}^\circ = { \hbox{rtln}}{{\hbox{k}}_{\rm{as } } } $ $ \end{document}. hence kas is a quantitative measure of the ligand affinity for the protein . comparison of the kas values for a series of structurally modified cap analogues enabled parsing of g into separate contributions from various stabilizing contacts inside the eif4e cap - binding pocket . bearing in mind an approximate character of the approach due to lack of additivity of the entropic terms [ 46 , 47 ] , combination of the crystallographic structure with such g analysis provided molecular mechanism of specific binding between the cap and eif4e . however , applying the procedure to detect the discrimination mechanism between mmg - and tmg - cap by some eif4es [ 17 , 18 ] has failed . the structures of ife isoforms derived by homology modeling were very similar to that of the eif4e template due to high sequence homology ( fig . 1 ) . the equilibrium association constant , \documentclass[12pt]{minimal }
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\begin{document}$$ { { \hbox{k}}_{\rm{as } } } = { { \hbox{k } } _ { + { 1}}}/{{\hbox{k } } _ { - { 1 } } } $ $ \end{document } , depends on the ligand ability to form and leave the complex , expressed by kinetic rate constants k+1 and k-1 , respectively . since it was impossible to calculate theoretically the rate constants from all - atom md simulations , we assumed that the md analysis of the apo proteins provided some information on k+1 that reflects accessibility of the mmg - cap analogue and of the tmg - cap analogue for the binding sites of the three eif4es . similarly , the md analysis of the three factors , each bound to either mmg - cap or tmg - cap , might provide some hints on the stability of the complexes that influences their dissociation kinetic constants k-1 . it should be emphasized that the simulated apo structures were generated by removing the ligand , and therefore they differ from those derived experimentally , at least in the case of the nmr structure of murine apo - eif4e . the simulated apo structures may be regarded as those , which approximate final conformations the ligands would dock into , especially if the cap phosphate chain anchores first as postulated by a two - state model of the cap - binding ( see below ) . it is also worth noting , that the large structural fluctuations observed in the nmr structure of apo - eif4e do not occur during our md simulations . the md trajectories of the apo eif4es display enhanced flexibility of the loops around the entrance to the cap - binding pocket ( fig . 2 ) , especially s1s2 and s7s8 loops , while the secondary structure elements remained unchanged . this general view of the dynamic behavior is confirmed by the experimental data derived for the apo form of human eif4e by multidimensional nmr , for the cap - free human eif4e in the complex with glycerol that seems to represent an intermediate state between the apo and the cap - bound form , and for the murine factor by hydrogen - deuterium exchange combined with electrospray mass spectrometry . the secondary elements were preserved in apo eif4e while the loops exhibited mobility on the ns - ps time scale that became abrogated upon the cap binding . the structural differences in the regions of loops s1s2 , s3s4 , s5s6 , and s7s8 ( fig . 2a ) resulted in the formation of the positively charged pocket to anchor the cap phosphate chain ( loops s1s2 and s7s8 ) , and the formation of the stacking triad and hydrogen bonding with the 7-methylguanine moiety via locking the w56 hinge ( loop s1s2 ) and rotating w102 ( loop s3s4 ) into the cap - binding site . the ability of w102 containing loop s3s4 to rotate from the open ( apo ) state into the closed form was confirmed by both crystallography and nmr studies . moreover , an alternative orientation of w102 indole ring was found by the diffraction studies of the eif4e complexes with 7-benzyl - gmp and 7-(p - fluorobenzyl)-gmp . the w102 residue flips through 180 in comparison with those complexes which contain the 7-methylguanosine caps [ 12 , 23 , 24 ] . all those observations agree with the two - state model of the cap - binding , in which anchoring of the phosphate chain was followed by a cooperative formation of the stacking triad and hydrogen bonds . this model , derived from analysis of the fluorescence titration of eif4e with structurally modified cap analogues , was confirmed by kinetic stopped flow experiments [ 51 , 52 ] . contrary to our analysis , slepenkov et al . performed and analyzed the stopped flow experiments differently , and postulated a one - step mechanism however , our results are confirmed not only by the standard analysis of the stop - flowed kinetic traces under pseudo first - order conditions but also by running the experiments under the second - order conditions combined with numerical integration of the suitable differential kinetic equations by the state - of - the - art program dynafit ( b ) time course of the distances between , ( upper ) s7s8 and s5s6 loops , measured for c atoms of s209 and k159 for apo - eif4e and apo - ife-3 , and for c atoms of q217 and k159 for apo - ife-5 , ( lower ) s5s6 and s1s2 loops , measured for c atoms of k159 and r52 . the lines are marked as follows : apo - eif4e dotted , apo - ife-3 dashed , apo - ife-5 solid analysis of the cap accessibility into the binding pockets . ( b ) time course of the distances between , ( upper ) s7s8 and s5s6 loops , measured for c atoms of s209 and k159 for apo - eif4e and apo - ife-3 , and for c atoms of q217 and k159 for apo - ife-5 , ( lower ) s5s6 and s1s2 loops , measured for c atoms of k159 and r52 . the lines are marked as follows : apo - eif4e dotted , apo - ife-3 dashed , apo - ife-5 solid the flexibility of the loops seem to be crucial for the discrimination between mgdp and m32,2,7gdp by murine eif4e and c. elegans ife-3 and ife-5 . the calculated distances between s7s8 and s5s6 , and between s5s6 and s1s2 , on the final parts of the md trajectories ( fig . 10 greater for ife-5 that binds the tmg - cap than for ife-3 and murine eif4e that are specific for the mmg - cap only . hence , the tmg - analogue with two additional methyl groups can easy penetrate the ife-5 binding pocket contrary to other factors . a similar analysis of the dynamics of the protein - cap complexes shows stable contacts between the cap phosphates and the arginine / lysine side chains , irrespective of the bound analogue . this is consistent with the anchoring character of the phosphate groups in the cap stabilization inside the eif4e binding slot . on the contrary , the mutual orientations of the rings in the cation- stacking triad undergo larger fluctuations in respect to the starting structure . as expected , the largest changes are observed in the eif4e - gdp complex that is stabilized by weaker - stacking , i.e. , a perpendicular orientation of the w56 and g rings and a shift of w102 deeper into the binding pocket . in the case of all the other complexes a temporary increase of the distance between w102 and mg rings are observed in the eif4e - m32,2,7gdp and ife-3-m32,2,7gdp , but not in ife-5-m32,2,7gdp , complexes . on the other hand , the hydrogen bond between the n - amino group of mg and the e103 carboxylate ( fig . 3 ) is being broken and reformed due to shifts of the e103 side chain into the bulk solvent . plasticity of that part of the cap - binding center , resulting in a rotation of e103 side chain and a movement of w102 side chain , was also observed in the cap - free eif4e ternary complex with eif4gi peptide and glycerol in the cap - binding pocket . therefore , the low affinity of the tmg - cap for murine eif4e can not be explained by lack of that hydrogen bond due to the hypermethylation . 3time dependence of the distance ( in ) between n of mgdp and glu103 side chain carboxyl in murine eif4e ( dotted ) , ife-3 ( dashed ) , and ife-5 ( solid ) time dependence of the distance ( in ) between n of mgdp and glu103 side chain carboxyl in murine eif4e ( dotted ) , ife-3 ( dashed ) , and ife-5 ( solid ) the distance between loops s7s8 and s5s6 in the eif4e - cap complexes ( fig . 4 ) is shown to correlate with the affinity of various cap types for the eif4e isoforms . binding of mgdp to murine eif4e results in bend of s7s8 toward s5s6 that might help to keep the ligand in the binding centre by water mediated interaction between k159 and , e.g. , s209 . closing the entrance to the cap - binding pocket by decreasing the distance between s209 and k159 was also observed in md simulations of phosphorylated eif4e . in the complexes with the low affinity ligands , m32,2,7gdp and gdp , the distance between the loops is larger , thus making it easier for those ligands to leave the complexes . similarly , the distance between the loops in the ife-3 complex with mgdp is much smaller than in the ife-3 complex with m32,2,7gdp , while for both ife-5 complexes it is kept fairly large , irrespective of the ligand . the analysis of the overall dynamics of the cap - bound eif4e isoforms was also carried out by ed analysis of the covariance matrix of the atomic displacements . the scalar products of the the vectors representing the normalized c displacements and the eigenvectors corresponding to the largest eigenvalue ( = 1 ) show non - gaussian distributions with 23 maxima . this can be interpreted as correlated , long - range movements , in which the loops oscillate around several mean positions ( fig . the histograms for the scalar products of the eigenvectors corresponding to lower eigenvalues ( = 10 ) are gaussian , as expected for equilibrated , independent and harmonic motions . time course of the distances between s7s8 and s5s6 loops , measured for c atoms of s209 and k159 ( ife-3 , eif4e ) , and for c atoms of q217 and k159 ( ife-5 ) . ( a ) murine eif4e bound to mgdp ( solid ) , to m32,2,7gdp ( dashed ) , and to gdp ( dotted ) . ( b ) ife-3 bound to mgdp ( solid bold ) and to m32,2,7gdp ( dashed ) , and ife-5 bound to mgdp ( solid ) and to m32,2,7gdp ( dotted)fig . motions along the first ( = 1 ) , fifth ( = 5 ) , and tenth ( = 10 ) eigenvectors obtained from the c coordinates covariance matrix , and the corresponding probability distribution for the displacements ( nm ) , ( a ) eif4e bound to mgdp , ( b ) eif4e bound to m32,2,7gdp , ( c ) ife-3 bound to mgdp , ( d ) ife-3 bound to m32,2,7gdp , ( e ) ife-5 bound to mgdp , ( f ) ife-5 bound to m32,2,7gdp analysis of the cap propensity to leave the cap - binding pocket . time course of the distances between s7s8 and s5s6 loops , measured for c atoms of s209 and k159 ( ife-3 , eif4e ) , and for c atoms of q217 and k159 ( ife-5 ) . ( a ) murine eif4e bound to mgdp ( solid ) , to m32,2,7gdp ( dashed ) , and to gdp ( dotted ) . ( b ) ife-3 bound to mgdp ( solid bold ) and to m32,2,7gdp ( dashed ) , and ife-5 bound to mgdp ( solid ) and to m32,2,7gdp ( dotted ) mobility of the apo - and cap - bound eif4es by essential dynamics analysis . motions along the first ( = 1 ) , fifth ( = 5 ) , and tenth ( = 10 ) eigenvectors obtained from the c coordinates covariance matrix , and the corresponding probability distribution for the displacements ( nm ) , ( a ) eif4e bound to mgdp , ( b ) eif4e bound to m32,2,7gdp , ( c ) ife-3 bound to mgdp , ( d ) ife-3 bound to m32,2,7gdp , ( e ) ife-5 bound to mgdp , ( f ) ife-5 bound to m32,2,7gdp the knowledge of the molecular basis of the rna 5 cap structure recognition by the cap - binding proteins is a prerequisite for understanding possible mechanisms of the cap functioning in various types of the gene expression processes in eukaryotes , such as translation initiation , mrna splicing , and export of rnas to the cytoplasm . it seems that various evolutionary unrelated cap - binding proteins converged on a similar general mechanism of the cap recognition . subtle modifications of the general recognition mechanism of the cap may lead to differences in the protein functions , e.g. , the diverse role of two aromatic amino acids that stack with the 7-methylguanosine moiety [ 5658 ] . the methyl group at n7 in the cap structure imparts a net positive charge to the guanine ring , and results in more efficient stacking compared with the unmethylated base . quantum mechanical calculations showed that a typical energy of the cation- stacking of the mg base in the complex with the tyrosine or tryptophan aromatic ring is in a range 11.4 kcal mol to16.23 kcal mol , while the g / y - stacking energy is ca.6 kcal mol . the stacking energy of m32,2,7g / w276 in snurportin 1 , 12.52 kcal mol , is close to typical values obtained for the mg / w complexes . the presence of two methyl substituents in the amino group of 7-methylguanosine brakes at least one stabilizing hydrogen bond in the cap - binding protein pockets and may lead to a substantial decrease of the association constants observed for the complexes with tmg - cap . on the other hand , the dual specificity cap - binding proteins possess high homology and structure similarity to those that discriminate between the mmg - cap and the tmg - cap . hence , it is a great challenge to conceive a molecular model of the discrimination vs. dual specificity for the protein - cap association . the explanations usually do not go beyond formulations like the differences in the size of the cap - binding pocket in the c. elegans isoforms of eif4e . our approach to elucidate the specificity of the caps recognition can be expressed in terms of the ligand ability to enter or leave the apo protein binding pocket , since the equilibrium association constant is determined by the ratio of the two rate constants . although we were not able to calculate the values of the rate constants by all - atom md simulations , the comparative analysis of the md trajectories of the apo- and cap - bound factors provides a more detailed explanation for the differences in the binding specificity of two c. elegans eif4e isoforms , ife-3 and ife-5 , than those published hitherto . the dynamic mechanism of the discrimination between two types of the cap may be ascribed to differences in mobility of the loops around the entrance to the protein cap - binding pockets , especially s7s8 loop . our results show also higher rigidity of the cation- stacking triad and of the stabilizing interactions ( salt bridges , hydrogen bonds ) involving the cap phosphate chain compared with a more flexible character of the hydrogen bonds . the results of our computer modeling are generally consistent with the experimental , structural and dynamic , data [ 32 , 48 , 49 ] . discrimination between two types of the cap , mmg - cap and tmg - cap , consist neither in the differences in the stacking energy nor in well - defined structural differences inside the cap - binding pocket . both 7-methylguanosine and its hypermethylated form were found to stack equally well in between two amino acid aromatic rings , and the structure of s. mansoni eif4e in complex with the mmg - cap analogues , showed a very similar mode of binding to that of the single specificity eif4es . md simulations based on the known structures of the cap - eif4e complexes provided means to evaluate the discrimination mechanism . net of stabilizing contacts inside the cap - binding pockets of highly homologic eif4es , we took into account the differences in the dynamics of the formation and dissociation of the eif4e - cap complexes . an exact specification of the role of particular amino acids in the proposed dynamical mechanism , e.g. , their mutual interactions and/or their interactions with various cap structures , needs further investigation . | specific recognition and binding of the ribonucleic acid 5 termini ( mrna 5 cap ) by the eukaryotic translation initiation factor 4e ( eif4e ) is a key , rate limiting step in translation initiation .
contrary to mammalian and yeast eif4es that discriminate in favor of 7-methylguanosine cap , three out of five eif4e isoforms from the nematode caenorhabditis elegans as well as eif4es from the parasites schistosome mansoni and ascaris suum , exhibit dual binding specificity for both 7-methylguanosine - and n2,n2,7-trimethylguanosine cap . to address the problem of the differences in the mechanism of the cap recognition by those highly homologic proteins , we carried out molecular dynamics simulations in water of three factors , ife-3 and ife-5 isoforms from c. elegans and murine eif4e , in the apo form as well as in the complexes with 7-methyl - gdp and n2,n2,7-trimethyl - gdp .
the results clearly pointed to a dynamical mechanism of discrimination between each type of the cap , viz .
differences in mobility of the loops located at the entrance into the protein binding pockets during the cap association and dissociation .
additionally , our data showed that the hydrogen bond involving the n2-amino group of 7-methylguanosine and the carboxylate of glutamic acid was not stable .
the dynamic mechanism proposed here differs from a typical , static one in that the differences in the protein - ligand binding specificity can not be ascribed to formation and/or disruption of well defined stabilizing contacts . |
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quantitative sensory testing ( qst ) to determine experimental pain sensitivity for healthy participants and participants with pain conditions appears to be gaining in popularity.1 recent reports have suggested standardized qst protocols for the assessment of individuals with chronic pain conditions.24 most qst clinical applications are for the assessment of threshold ( the time , or level of stimulation first perceived as painful ) , supra - threshold ( ratings of painful stimulation above threshold ) , temporal summation , or after - sensation.4 temporal summation refers to the increase in pain perception with repeated stimulation ( inter - stimulus interval three seconds or less ) that is believed to be related to c - fiber mediated dorsal horn central sensitization.5 this phenomenon has been reported to be related to the development of chronic pain conditions.5 after - sensation is the heightened level of pain that lingers following the cessation of pain stimulation.5 after - sensation has been shown to predict clinical pain ratings , and may be related to endogenous , pain inhibitory mechanisms of c - fiber activity.6 studies involving participants with pain conditions have suggested the importance of qst for supra - threshold testing and assessment of temporal summation.5,7,8 previous work has also shown that after - sensation is related to clinical pain report suggesting not only large group differences between patients and controls , but also a relationship between qst , clinical pain ratings , and psychological factors.5,6 however , few studies have investigated associations between different stimulus parameters and the influence of anatomical location of stimulus site for classic qst measures.9 the above - referenced evidence of the importance of the assessment of central sensitization - related phenomena indicates a need to investigate these parameters for supra - threshold response procedures , temporal summation , and after - sensation at multiple sites along the neural axis . in summary , studies employing qst methods have used a variety of body sites and stimulus parameters and rarely have multiple measures been compared to each other . this lack of direct comparison across site and test type makes different studies difficult to interpret . furthermore , the relative contribution of common psychological factors as they relate to supra - threshold response , temporal summation , and after - sensation also needs investigation . the purpose of the present study was to investigate the differences in pain report from three anatomical sites on the body , including the lower extremity , trunk , and upper extremity for supra - threshold response , temporal summation , and after - sensation . reporting the inter - relationships of the various qst methods of pain induction is important for the interpretation of data across studies , and may serve as a preliminary assessment of the construct validity of these measures . the analyses of these relationships were primarily for descriptive and exploratory purposes , although our default hypothesis was that the qst measures would all be significantly correlated with each other and that responses would be similar for each of the anatomical sites . we further hypothesized that the measures associated with c - fiber mediated processes ( temporal summation and after - sensation ) would be more strongly correlated in comparison to a - delta fiber mediated processes ( supra - threshold response ) . we also sought to provide novel information of the relationship of these particular tests with measures of fear of pain , anxiety , anxiety sensitivity , and pain catastrophizing . these particular psychological factors have been related to pain in both clinical and experimental settings10,11 and are relevant to fear avoidance models of musculoskeletal pain , which has an important influence on clinical pain conditions.12,13 we hypothesized that fear of pain , anxiety , and pain catastrophizing would be positively associated with the different qst measures . a convenience sample of 120 participants was pooled from two previously reported studies that focused on thermal pain responses.14,15 pooling these studies was appropriate for addressing our purposes because the studies had the same eligibility criteria and qst testing procedures . furthermore , the thermal pain sensitivity data have not been previously used to test the hypotheses reported in this planned secondary analysis . each study had institutional review board approval and all participants provided informed consent before being included in the studies . participants provided demographic information and completed validated psychological questionnaires that were consistent with fear - avoidance models of pain perception.12 participants also completed a visual analog scale ( vas ) for fear and anxiety ratings for the current stimuli they were about to experience . participants then underwent standardized qst for thermal pain sensitivity at the lower extremity , trunk , and upper extremity . the qst protocol as has been employed in previous studies8,15 and is further described below . the anxiety sensitivity index ( asi ) uses a 16-item , 4-point rating scale to assess anxiety sensitivity , which is the perception of whether experiencing symptoms of anxiety causes harm . the asi has been validated in community samples16 and demonstrated factor invariance across different sex and age groups.17 the state - trait anxiety questionnaire ( stai ) uses a 40-item , 4-point rating scale to assess dispositional ( trait ) and situational ( state ) anxiety symptoms.18 we reported the state portion of the stai as this construct better matched the purposes of this study . the coping strategies questionnaire ( csq ) uses a 27-item , 7-point rating scale to measures the frequency of use for common pain coping strategies.19 we used the catastrophizing subscale that measures helplessness and pessimistic cognitions related to pain perception . the validity of this particular subscale has been supported19,20 and the currently recommended scoring system was used in this study.20 the fear of pain questionnaire ( fpq - iii ) uses a 30-item , 5-point rating scale to measure fear about specific situations that would normally produce pain.21 the fpq - iii is a commonly used and well - validated instrument that is appropriate for use in nonclinical and clinical populations.2123 thermal stimuli were delivered with the medoc neurosensory analyzer ( tsa 2001 ; ramat yishai , israel ) . participants underwent a practice session in which threshold stimuli were rated with a numerical rating scale ( nrs ) ranging from 0 ( no pain ) to 100 ( worst pain imaginable ) . after the practice session , pain sensitivity was assessed at the lower - extremity , trunk , and upper - extremity using a counter - balanced testing schedule . supra - threshold responses are believed to be representative of a - delta fiber mediated pain sensitivity and these responses were assessed at the volar forearm , posterior superior iliac spine , and dorsal calf via a ramp and hold protocol . the ramp and hold protocol consisted of four heat pulses each lasting five seconds and delivered five seconds from the end of the preceding pulse . baseline temperature for each heat pulse was 35c and increased at a rate of 10c per second to a randomly determined end point of 45c , 47c , 49c , or 51c . the protocol was repeated twice and nrs ratings were recorded in response to these four temperatures . temporal summation and after - sensation are considered to be representative of c - fiber mediated pain sensitivity and these responses were assessed on the palmar surface of the hand , posterior superior iliac spine , and plantar surface of the foot . for temporal summation a train of ten consecutive heat pulses with baseline temperature of 41c and peak temperature of 51c was provided at an inter- stimulus frequently of 0.33 hz . participants rated each pulse using the nrs and the fifth pulse was used as the measure of temporal summation . the rationale for choosing the fifth pulse is that it has been demonstrated that first pain is either absent or nearly completely diminished after four heat pulses and the remaining pain is delayed by an extent that can only be accounted for by conduction of impulses in c - fibers.24,25 this delayed pain is also much greater in magnitude than that evoked by the first pulse in a train of 46 pulses or by a single pulse24,26,27 and therefore is considered to represent a simple measure of temporal summation.25 for after - sensations , nrs ( numeric rating scale ) ratings were collected at 15seconds following the last stimulus pulse , similar to our previous work.28 descriptive statistics were generated for the sample , for demographic , psychological , and qst measures . associations between supra - threshold response , temporal summation , and after - sensation were investigated by pearson correlation . the effect of the stimulation site was investigated by repeated measures anova , with separate models for supra - threshold response , temporal summation , and after - sensation . finally , psychological relationships to pain measures were investigated with pearson correlation and multiple regression . separate regression models included the fear of pain , anxiety , and pain catastrophizing measures as predictors and supra - threshold response , temporal summation , and after - sensations as the dependent variable . a convenience sample of 120 participants was pooled from two previously reported studies that focused on thermal pain responses.14,15 pooling these studies was appropriate for addressing our purposes because the studies had the same eligibility criteria and qst testing procedures . furthermore , the thermal pain sensitivity data have not been previously used to test the hypotheses reported in this planned secondary analysis . each study had institutional review board approval and all participants provided informed consent before being included in the studies . participants provided demographic information and completed validated psychological questionnaires that were consistent with fear - avoidance models of pain perception.12 participants also completed a visual analog scale ( vas ) for fear and anxiety ratings for the current stimuli they were about to experience . participants then underwent standardized qst for thermal pain sensitivity at the lower extremity , trunk , and upper extremity . the qst protocol as has been employed in previous studies8,15 and is further described below . the anxiety sensitivity index ( asi ) uses a 16-item , 4-point rating scale to assess anxiety sensitivity , which is the perception of whether experiencing symptoms of anxiety causes harm . the asi has been validated in community samples16 and demonstrated factor invariance across different sex and age groups.17 the state - trait anxiety questionnaire ( stai ) uses a 40-item , 4-point rating scale to assess dispositional ( trait ) and situational ( state ) anxiety symptoms.18 we reported the state portion of the stai as this construct better matched the purposes of this study . the coping strategies questionnaire ( csq ) uses a 27-item , 7-point rating scale to measures the frequency of use for common pain coping strategies.19 we used the catastrophizing subscale that measures helplessness and pessimistic cognitions related to pain perception . the validity of this particular subscale has been supported19,20 and the currently recommended scoring system was used in this study.20 the fear of pain questionnaire ( fpq - iii ) uses a 30-item , 5-point rating scale to measure fear about specific situations that would normally produce pain.21 the fpq - iii is a commonly used and well - validated instrument that is appropriate for use in nonclinical and clinical populations.2123 thermal stimuli were delivered with the medoc neurosensory analyzer ( tsa 2001 ; ramat yishai , israel ) . participants underwent a practice session in which threshold stimuli were rated with a numerical rating scale ( nrs ) ranging from 0 ( no pain ) to 100 ( worst pain imaginable ) . after the practice session , pain sensitivity was assessed at the lower - extremity , trunk , and upper - extremity using a counter - balanced testing schedule . supra - threshold responses are believed to be representative of a - delta fiber mediated pain sensitivity and these responses were assessed at the volar forearm , posterior superior iliac spine , and dorsal calf via a ramp and hold protocol . the ramp and hold protocol consisted of four heat pulses each lasting five seconds and delivered five seconds from the end of the preceding pulse . baseline temperature for each heat pulse was 35c and increased at a rate of 10c per second to a randomly determined end point of 45c , 47c , 49c , or 51c . the protocol was repeated twice and nrs ratings were recorded in response to these four temperatures . temporal summation and after - sensation are considered to be representative of c - fiber mediated pain sensitivity and these responses were assessed on the palmar surface of the hand , posterior superior iliac spine , and plantar surface of the foot . for temporal summation a train of ten consecutive heat pulses with baseline temperature of 41c and peak temperature of 51c was provided at an inter- stimulus frequently of 0.33 hz . participants rated each pulse using the nrs and the fifth pulse was used as the measure of temporal summation . the rationale for choosing the fifth pulse is that it has been demonstrated that first pain is either absent or nearly completely diminished after four heat pulses and the remaining pain is delayed by an extent that can only be accounted for by conduction of impulses in c - fibers.24,25 this delayed pain is also much greater in magnitude than that evoked by the first pulse in a train of 46 pulses or by a single pulse24,26,27 and therefore is considered to represent a simple measure of temporal summation.25 for after - sensations , nrs ( numeric rating scale ) ratings were collected at 15seconds following the last stimulus pulse , similar to our previous work.28 descriptive statistics were generated for the sample , for demographic , psychological , and qst measures . associations between supra - threshold response , temporal summation , and after - sensation were investigated by pearson correlation . the effect of the stimulation site was investigated by repeated measures anova , with separate models for supra - threshold response , temporal summation , and after - sensation . finally , psychological relationships to pain measures were investigated with pearson correlation and multiple regression . separate regression models included the fear of pain , anxiety , and pain catastrophizing measures as predictors and supra - threshold response , temporal summation , and after - sensations as the dependent variable . descriptive data for the participants is summarized in table 1 . to reduce the total number of analyses , ratings of the ramp and hold stimuli were averaged across the four stimulus temperatures to yield a mean rating for each participant and for each site of stimulation . this sample was 70% female , and sex differences in the relationships among pain and psychological measures were conducted with z - tests for differences among correlations . results of the repeated measures anova indicated a significant effect for site of stimulation and supra - threshold response ( f(2,238 ) = 20.3 , p = 0.001 , eta = 0.15 ) . simple contrasts indicated that pain ratings for the lower extremity were lower compared to trunk and upper extremity ratings ( p < 0.05 ) . upper extremity ratings were also significantly lower than those for the trunk ( p < 0.05 ) . results of the repeated measures anova indicated significantly greater temporal summation for the trunk than either lower or upper extremity sites ( f(2,118 ) = 63.2 , p < 0.001 , eta = 0.35 ) . results of the repeated measures anova indicated a significant effect for site of stimulation and after - sensation ( f(2,238 ) = 11.8 , p < 0.001 , eta = 0.09 ) . simple contrasts indicated that after - sensation in the trunk was significantly higher than in the lower and upper extremities ( p < 0.05 ) . however , after - sensation did not differ between lower and upper extremities ( p > 0.05 ) . pearson correlations for the psychological measures are presented in table 3 and correlations between psychological and qst measures are presented in table 4 . results of the regressions of fear and anxiety measures are summarized in table 5 . both the zero - order correlations and furthermore , all variables met criteria for normality ( skewness and kurtosis ) . for supra - threshold responses , the psychological measures were significant predictors of pain ratings at all anatomical sites with fear of pain scores as a unique predictor . all relationships were in the positive direction indicating greater scores on fear of pain and anxiety sensitivity were related to greater pain sensitivity . for temporal summation , only the regression for the trunk site was statistically reliable ( r = 0.29 , p = 0.004 ) . examination of the standardized beta weights indicated that anxiety sensitivity was the only unique contributor and indicated that increased anxiety sensitivity was associated with greater temporal summation . none of the psychological measures were significant predictors of lower extremity temporal summation . though the overall regression for upper extremity was not statistically reliable , the individual beta weight for anxiety sensitivity was significant and in the same direction as that for trunk site temporal summation . for after - sensations , psychological measures were statistically reliable predictors of pain sensitivity at the trunk . examination of standardized beta coefficients indicated that only fear of pain was a unique predictor of after - sensations at the trunk . neither lower nor upper extremity regressions yielded statistically reliable prediction equations for after - sensations . qst of individuals with and without pain conditions is increasingly common in the pain literature . there is a wide variety of types of stimulation , sites of stimulation , and stimulation methods employed across investigations . this study contributes to the experimental pain literature and suggests that some of the inconsistencies observed in the literature may be due to the differences related to site of stimulation , type of stimulus procedure , and psychological influence on pain sensitivity . the first aim of this study was to examine the inter - correlations among the qst measures of supra - threshold response , temporal summation , and after - sensation . these inter - correlations suggest that temporal summation , after - sensation and supra - threshold response , share moderate levels of variance within and across stimulation sites . there was no significant support that presumed c - fiber - related parameters would more closely correlate with each other versus presumed a - delta parameters . rather , the similar correlations among all psychophysical parameters suggest that an overall pain sensitivity is reflected in the pain responses used in this study . . it would be more consistent with a clinical sample that experienced a generalized central sensitization to somatic and painful stimulation . it is plausible that the findings in healthy controls simply reflect the lower range , or normal distribution of central influences on somatic perception . the second aim of the study was to examine the relationship of site of stimulation and pain responding within and between types of qst . supra - threshold , after - sensation , and temporal summation were influenced by site of stimulation with a similar pattern . specifically , all of the sensory parameters showed that the trunk region had higher pain sensitivity , in comparison to the extremities . our results suggest that the site of stimulation is important to consider when comparing or designing studies . specifically , ratings from the extremities and the trunk may not be comparable and could result in conclusions that are not robust and indeed may be misleading . the site of stimulation results may also have implications for neuraxis differences in processing standard pain stimuli for healthy individuals . first , given the high prevalence of chronic low back pain , it seems relevant to note that the trunk had the highest pain sensitivity across all parameters . in a nonpain sample , we did not expect the trunk to be more sensitive to thermal stimulation , but our data suggest the trunk may have elevated pain sensitivity for temporal summation and after - sensation , even in healthy individuals . furthermore , the measures of temporal summation , in particular , were expected to be more intense in the extremities because of the greater conduction distance compared to the trunk . this finding could indicate that the specific protocol used in the study did not provide sufficient discrimination between a - delta and c - fiber mediated pain responding and that from the trunk ; a more blended innervation was experienced . the final aim of the study was to examine the relationship of qst measures with common psychological constructs ( ie , fear , anxiety , and catastrophizing ) used in fear - avoidance models of pain . interestingly , the validated questionnaires measuring trait constructs had the stronger associations with the qst measures , in comparison to state specific vas ratings of fear and anxiety regarding the about to be experienced stimuli . when associations with fear and anxiety measures were examined , it was revealed that fear of pain and anxiety sensitivity measures were strong predictors of supra - threshold response for all anatomical areas . these data are consistent with some of our previous studies suggesting that fear is associated with experimental pain sensitivity during cold - pressor for healthy participants,29,30 and thermal pain for participants with low back pain.31 fear of pain and anxiety sensitivity measures were consistently associated with trunk pain perception for all three parameters ( ie , after sensation , temporal summation , and supra - threshold stimulation ) , but not with lower and upper extremity pain perception . this finding was counter to one of our earlier studies suggesting anxiety was associated with temporal summation in the upper extremity.32 previous studies have not investigated psychological influence on after - sensation , so there are few comparative studies available . collectively these data indicate that psychological influence might be more consistent across sites for a - delta fiber mediated processes ( supra - threshold response ) , with more site - specific ( trunk ) influence on purported c - fiber processes ( temporal summation and after - sensation ) . the use of asymptomatic control participants with presumably normally functioning nervous systems may mean that the results are not compatible with individuals whose chronic pain has altered their perception of normally , nonpainful and painful stimulation through central sensitization processes . furthermore , the experience of chronic pain may also alter the expectations and associations between psychological measures and the experience of pain in both quantitative and qualitative processes . the level of psychological measures is likely to be much lower in this asymptomatic sample than in a chronic pain sample . the generalizability of these findings to different psychophysical protocols , and different methods of inducing supra - threshold , temporal summation , and after - sensation pain may also be limited . finally , our psychological measures were not stimulation - site specific . we can only speculate about differences in anxiety sensitivity and fear related to trunk versus extremity associations to pain stimulation . perhaps stimulation to the trunk is more novel , and elicits greater anxiety and fear than extremity stimulation , resulting in greater associations between pain and psychological measures at the trunk . future studies might be improved by including site - specific measures of anxiety and fear . in summary , this was one of the first studies to directly compare different qst induction methods within the same individual and across body sites . results suggest that previous assumptions of a - delta and c - fiber mediated pain processes may not be adequately reflected in the described qst protocol . furthermore , site of stimulation may be an important consideration when designing and comparing studies using these methods . for both supra - threshold response and after - sensation pain ratings differed by site ; with trunk stimulation generally experienced as more intense . supra - threshold response was highly associated with fear and anxiety measures across all anatomical sites , while temporal summation and after - sensations were only correlated with anxiety sensitivity and fear at the trunk respectively . temporal summation ratings were higher for trunk stimulation and consistently associated with anxiety sensitivity at the same site . therefore , studies designed to examine pain - related fear and anxiety sensitivity are more likely to find associations if they employ supra - threshold response methods at most sites of stimulation , although temporal summation associations appeared stronger with trunk stimulation . | this study investigated the relationship of thermal pain testing from three types of quantitative sensory testing ( ie , supra - threshold stimulus response scaling , temporal summation , and after - sensation ) at three anatomical sites ( ie , upper extremity , lower extremity , and trunk ) .
pain ratings from these procedures were also compared with common psychological measures previously shown to be related to experimental pain responses and consistent with fear - avoidance models of pain .
results indicated that supra - threshold stimulus response scaling , temporal summation , and after - sensation , were significantly related to each other .
the site of stimulation was also an important factor , with the trunk site showing the highest sensitivity in all three quantitative sensory testing procedures .
supra - threshold response measures were highly related to measures of fear of pain and anxiety sensitivity for all stimulation sites . for temporal summation and
after - sensation , only the trunk site was significantly related to anxiety sensitivity , and fear of pain , respectively .
results suggest the importance of considering site of stimulation when designing and comparing studies . furthermore , psychological influence on quantitative sensory testing is also of importance when designing and comparing studies .
although there was some variation by site of stimulation , fear of pain and anxiety sensitivity had consistent influences on pain ratings . |
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tuberous sclerosis complex ( tsc ) is a rare autosomal dominant disorder first described by von recklinghausen in 1862 . in 1880 , bourneville coined the term sclerose tubereuse based on the pathologic features of the sclerotic tubers found in the postmortem investigation of patients with epilepsy and mental retardation , henceforth known as bourneville 's disease . tsc is classified as a phakomatosis ( neurocutaneous disorder ) and which is characterized by the presence of glial cell tumors arising in the cerebral hemispheres and retina . it is clinically characterized by the triad of epilepsy ( epi ) , intellectual disability ( loi ) , and adenoma sebaceum ( a ) , therefore , encompassing these features sherlock coined the term epiloia . the frequency of tsc has been estimated to be 1 in 10,000 live births and is about a third as common as neurofibromatosis type 1 . it is a multisystemic disorder which becomes perceptible only in late childhood , limiting the expediency for early diagnosis in infancy . here , we present a case report of a 26-year - old male patient with distinctive clinical , radiological , and histological features of tsc . a 26-year - old male patient presented with multiple growths in the upper and lower gums since 5 years . the patient noticed the growth for the past 5 years , which gradually increased in size to attain the present size . there was no history of pain or blood / purulent discharge associated with the growth . the patient was a known epileptic and hysteric since the age of 10 and under anticonvulsant and antidepressants . on general physical examination , multiple well - defined , reddish - brown sessile nodular growths were noted on the forehead , nose , and cheeks in a characteristic butterfly pattern and front and back trunk region - angiofibromas . similar sessile and firm nodular growths were noted in the upper and lower extremities of varying sizes , suggestive of periungual fibromas or kenen tumors . a well - defined roughened hypermelanotic patch was noted in the left shoulder girdle and the right lumbosacral region showing an orange peel appearance indicative of shagreen patch [ figure 1 ] . clinical photograph of the patient showing multiple well - defined , reddish - brown sessile nodular growths on the forehead , nose , and cheeks in a characteristic butterfly pattern , shagreen patch , and periungual fibromas on intraoral examination , similar well - defined , sessile , firm , and nodular growths were seen in the marginal and attached gingiva in the upper and lower anterior region of varying sizes . multiple hypoplastic enamel pits were noted in the occlusal aspect of posterior teeth [ figure 2 ] . based on the history and clinical findings , multiple gingival growths and hypoplastic enamel pits the patient was subjected to various radiological , hematological , and histopathological investigations . a radiopaque mass resembling tooth such as structure involving enamel , dentin , and pulp was seen in the periapical region of 21 , 22 regions suggestive of compound odontome . chest radiograph revealed inferior displacement of the left border of ventricle suggestive of left ventricular hypertrophy . axial computed tomographic sections of the brain showed hypodense areas measuring approximately 5 mm 5 mm in the subependymal regions of both ventricles indicating multiple calcified tuberous lesions ( subependymal nodules ) [ figure 3 ] . orthopantomogram , intraoral periapical radiographic view , chest radiograph , and axial computed tomography view hematological investigations were found to be satisfactory except for erythrocyte sedimentation rate which was found to be raised to 32 mm . henceforth , excisional biopsy of the gingival growth in 23 , 24 regions and cutaneous punch biopsy was performed . microscopically , hematoxylin and eosin - stained section of the intraoral tissue specimen showed a stretched out , hypoplastic stratified squamous epithelium overlying connective tissue stroma . the underlying connective stroma consisted of dense , thick collagenous bundles with hypocellularity comprising of few fibroblasts , mesenchymal cells , constricted blood vessels , and inflammatory cells . the epithelium appeared to be flattened and thinned out with the basal cells exhibiting an abundance of deeply stained melanocytes . underlying connective tissue was dense with collagen bundles , mesenchymal cells , and few nerve fiber bundles . there was no evidence of sebaceous glands or hair follicles [ figure 4a c ] . histopathological view ( a and b , 10 ; c , 40 ) based on all the clinical findings and investigations , a final diagnosis of tuberous sclerosis was confirmed . tsc is characterized by the development of unusual tumor - like growths ( hamartomas ) in brain , skin , retina , and viscera . the term tuberous sclerosis refers specifically to the presence of multiple sclerotic masses scattered throughout the cerebrum . the diagnosis of tsc is based on the identification of hamartomas in more than one organ system . the abnormal genes have been localized to one of two sites , the long arm of chromosome 9 ( 9q34 ) , designated as tsc 1 ( encoding hamartin ) , and the short arm of chromosome 16 ( 16p 13.3 ) designated as tsc 2 ( encoding tuberin ) . these gene products form a tumor suppressor complex which drives rheb ( ras homolog enriched in brain ) a member of ras superfamily into the inactive guanosine diphosphate - bound state . when rheb is in the guanosine triphosphate - bound active state , it stimulates the mammalian target of rapamycin ( mtor ) , an evolutionarily conserved protein kinase and a major effector of cell growth . the mutations in these genes result in constitutive mtor activation leading to the formation of various growths and hamartomas in various organs of the body . in a study conducted on 325 tsc individuals , 17% of the mutations were found in the tsc1gene , 50% in the tsc2gene , 4% unclassified variants , and 29% with no identifiable mutations . at the tuberous sclerosis consensus conference of 1998 , the clinical diagnostic criteria of tsc were revised and a new classification system based on major and minor findings was established . the presence of two major features or one major and two minor features was considered sufficient for a definitive diagnosis . the major features include ( a ) hypomelanotic macules ( 3 , at least 5-mm diameter ) , ( b ) angiofibromas ( 3 ) orfibrous cephalic plaque , ( c ) ungual fibromas ( 2 ) , and ( d ) shagreen patch . the minor features include ( 1 ) confetti skin lesions , ( 2 ) dental enamel pits ( 3 ) , and ( 3 ) intraoralfibromas ( 2 ) . the most common oral manifestations of tsc are fibromas , gingival hyperplasia , and enamel hypoplasia ( in almost 100% of these patients and is associated with an increased risk of caries ) . the other features include hamartomatous rectal polyps , nonrenal hamartomas , multiple renal and bone cysts , high arched palate , bifid uvula , cleft palate , delayed dental eruption , and presence of diastemas . our case presented with three major features facial angiofibromas , kenen tumors and subependymal nodules and two minor features the management of these patients is often multidisciplinary involving the neurosurgeon , neurologist , nephrologist , pulmonologist , cardiologist , ophthalmologist , and the genetic counselor . surgery including dermabrasion and laser treatment may be useful for the treatment of skin lesions . intervention programs including special schooling and occupational therapy may benefit individuals with special needs and developmental concern . drug therapy for some of the manifestations of tsc is currently in the developmental stage . the use of inhibitors of mtor in regression of various hamartomatous growths is a newer modality in the management of tsc . these patients must adopt measures for careful oral and dental hygiene with regular visits to the dentist in order to eliminate potential irritative factors and ensure the early diagnosis of any possible lesions . in the present case , the patient was advised to take his physician consultation for modification of the antiepileptic drug . at present , the daily dose of phenytoin is reduced from 400 to 300 mg as maintenance protocol . application of fluoride varnish in hypoplastic enamel pits was also performed and kept under periodic recall . the prognosis of tsc depends on the severity or multiplicity of organ involvement . about a quarter of severely affected infants however , in the case of individuals diagnosed late in life with few cutaneous signs , prognosis depends on the associated internal tumors and cerebral calcifications . the present case unambiguously exemplifies the systemic manifestations of tsc and its specific expression in the oral cavity . hence , in turn to establish an appropriate diagnosis , oral manifestations of the disorder must not be overlooked which implies the necessity for the level of awareness of tsc among oral practitioners . the authors certify that they have obtained all appropriate patient consent forms . in the form the patient(s ) has / have given his / her / their consent for his / her / their images and other clinical information to be reported in the journal . the patients understand that their names and initials will not be published and due efforts will be made to conceal their identity , but anonymity can not be guaranteed . the authors certify that they have obtained all appropriate patient consent forms . in the form the patient(s ) has / have given his / her / their consent for his / her / their images and other clinical information to be reported in the journal . the patients understand that their names and initials will not be published and due efforts will be made to conceal their identity , but anonymity can not be guaranteed . | tuberous sclerosis complex is an unusual autosomal dominant neurocutaneous syndrome characterized by the development of benign tumors affecting different body systems affecting the brain , skin , retina , and viscera .
it is characterized by cutaneous changes , neurologic conditions , and the formation of hamartomas in multiple organs leading to morbidity and mortality .
the most common oral manifestations are fibromas , gingival hyperplasia , and enamel hypoplasia .
the management of these patients is often multidisciplinary involving specialists from various fields . here ,
we present a case report of a 26-old - year male patient with characteristic clinical , radiological , and histological features of tuberous sclerosis complex . |
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the most common sites of metastatic thymic carcinomas ( masaoka - koga stage ivb ) are the lung , liver and brain . they can be found in the kidney , pancreas , pituitary , orbital , lymph nodes , peritoneum , and the ovaries . 1 ) . according to the french rythmic guidelines , the patient underwent preoperative chemo - radiotherapy followed by a complete resection . parietal and pleural tumors were detected by ct scan and [ f]fdg - pet / ct in 2008 . the patient was reoperated , and these tumors were confirmed to be metastases of mixed thymoma b3 and thymic carcinoma . a pelvic mass was detected in 2013 by [ f]fdg - pet / ct ( suvmax 8.4 ) and confirmed by mri ( fig . a laparotomy confirmed an isolated peritoneal 60-mm tumoral mass localized on the left part of the douglas 's pouch . peritoneal and/or ovarian metastasis from thymic malignancies are very rare ; only four cases have been reported in the literature [ 2 , 3 , 4 ] . these metastases can occur immediately or several years after primary thymic tumor ( 36 years ) . different thymic histological subtypes were observed ( carcinoma , b1 , neuroendocrine ) , and they were always associated with a loco - regional extension and/or distant metastasis . to our knowledge , this is the first report of an isolated pelvic metastasis from thymic carcinoma . this isolated pelvic metastasis probably had a systemic origin rather than a direct peritoneal extension below the diaphragm ( as described in previously reported cases ) . however , considering that this pelvic metastasis was isolated and completely resected , chemotherapy was not performed in our patient . efficacy of [ 18f]fdg - pet / ct for the detection and localization of mediastinal recurrence is well established . our case report emphasizes the utility of [ 18f]fdg - pet / ct for metastasis detection and long - term follow - up in patients with thymic epithelial tumors . | thymic epithelial tumors are rare and often occur somewhere local .
metastatic sites of thymic carcinomas ( masaoka - koga stage ivb ) are mostly seen in the lung , liver and brain .
we report a 64-year - old female with an initial diagnosis of thymoma b3 who first showed thoracic recurrences and then an asymptomatic isolated pelvic metastasis from her thymic carcinoma . |
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oral cavity harbors billions of microorganisms , some of these contribute to the development or progression of systemic diseases such as cardiovascular disease , diabetes mellitus , etc . antibiotic resistance , adverse effects and toxicity to modern medicines has prompted scientists to research on natural products . it came into lime light and popularity by dr f karach.3 , 4 the present article reviews the role of oil pulling in maintaining oral hygiene . oil pulling is mentioned in the ayurvedic text charak samhita and sushruta samhita as kavala graha or kavala gandoosha.4 , 7 in gandoosha mouth is completely filled with oil such that gargling is impossible whereas in kavala graha comfortable lesser quantities of oil is used such that gargling is possible.6 , 8 in oil pulling , a tablespoon full of oil is swished around the mouth in the early morning before breakfast and in empty stomach for about 20 min . in case of children greater than five years of age , pulled and forced in between all the teeth by swishing it all around the mouth . at the end of this activity if the procedure is done correctly , the viscous oil will become milky white and thinner . then it is spit out and mouth is thoroughly washed with clean warm saline water or tap water and teeth are cleaned with fingers or routine tooth brushing is performed . if the jaw aches , then the procedure can be done just for 510 min . the oil should not be spit into the sink as the oil can cause clogging of the pipes . instead , the oil should be spit into a trashcan or on a paper towel . oil pulling should be ideally performed daily morning on empty stomach before brushing teeth and care should be taken that oil is not swallowed.4 , 10 , 11 swallowing of oil during oil pulling should be avoided as the oil contains bacteria and toxins . it can be practiced thrice daily in empty stomach before meals to fasten the healing effects . it is contraindicated for children below 5 years due to risk of aspiration.14 , 15 the practitioner should take care not to aspirate the oil while performing rigorous oil pulling . in cases of oral ulcers , fever , vomiting tendency , asthma and in conditions where brushing is difficult and sometimes contraindicated , oil pulling can be advantageously used to maintain oral hygiene . ayurveda hypothesizes that tongue is connected to various organs such as kidneys , heart , lungs , small intestine , spine , etc . oil pulling is believed to help in the excretion of toxic heavy metals by saliva . oil pulling activates salivary enzymes which absorb toxins such as chemical toxins , bacterial toxins and environmental toxins from the blood and removed from the body through the tongue.9 , 16 , 18 , 19 thus oil pulling detoxifies and purifies the entire human body . however it is argued that since oral mucosa is not a semipermeable membrane , toxins of the body from the blood can not pass through it . organic oils such as sunflower oil , sesame oil , and coconut oil are of benefit especially if it is cold pressed , though refined oil also works in pulling the bacteria , viruses and protozoa from the oral cavity . since trans fats are absent in cold pressed oils when compared to commercial oils which are extracted from strong petroleum based solvents ; oil pulling is ideally performed with cold pressed oils . oil pulling using olive oil , milk , extracts of gooseberry and mangoes is also documented . also polyunsaturated fatty acids present in sesame oil reduces free radical injury occurring in oral cavity . these oils will attract the lipid layer of bacterial cell membranes , and cause it to stick or get attracted , and pulled to the oil . during oil pulling , the oil gets emulsified and surface area of the oil gets increased . this oil will coat the teeth and gingiva and inhibits bacterial co - aggregation and plaque formation . thus plaque building bacteria responsible for dental caries , gingivitis , periodontitis and bad breath are removed from the oral cavity . oil pulling is also of help to resolve symptoms of dry mouth / throat and chapped lips . also teeth become whiter ; breath becomes fresher ; oral cavity muscles and jaws become stronger with excellent achievement of oral hygiene.9 , 23 oil pulling prevents dental caries , gingivitis , oral candidiasis and periodontitis from occurring , helps to reduce tooth pain , fixes mobile teeth and achieves vigorous oral hygiene.5 , 9 , 13 , 24 oil pulling when practiced regularly is believed to freshen and stimulate the mind and strengthen the senses . it is also beneficial in sore throat , dry face , impaired vision , taste loss and anorexia . it contains lauric acid which can react with alkalis present in saliva such as sodium hydroxide and bicarbonates to form sodium laureate - soap like substance , which reduces plaque adhesion and accumulation , and possesses cleansing action.3 , 4 , 12 lauric acid has antimicrobial , anti - inflammatory properties , prevents dental caries and is beneficial to oral health . in addition to this it also has a pleasant taste.4 , 25 coconut oil has antimicrobial activity and is effective against streptococcus mutans and candida albicans in an in vitro biofilm model . coconut oil also has anti - septic properties and can be safely used as emollient and moisturizer . coconut oil does not have adverse effects produced by chlorhexidine such as brown staining and altered taste sensation . it also contains plant phenolic compounds , squalen , phytosterols ; vitamin a , e and k. these constituents have antimicrobial , immunomodulatory and antioxidative effect . almond oil containing mouth rinse is suggested to result in low gingival scores whereas olive oil based mouth rinses is believed to inhibit plaque formation and inhibition . sesame oil contains sesamin , sesamolin and sesaminol and has detoxification , antioxidant , and antibiotic actions . monolaurin in coconut oil is effective against microorganisms such as staphylococcus aureus , candida spp . it is hypothesized that monolaurin causes bacterial death by altering bacterial cell wall , penetrating and disrupting cell membranes and inhibiting enzymes related to energy production and nutrient transfer . monolaurin also has virucidal activity by dissolving lipids and phospholipids in the viral envelope ; leading to disintegration of virus . sucrose monolaurate of coconut has anti - caries properties due to reduction in glycolysis and sucrose oxidation by s. mutans and thus prevents the formation of dental plaque . however it should be kept in mind that oil pulling does not reverse the existing dental caries and hence regular visits to dentists are still required . in addition to maintaining oral hygiene it has been claimed to have systemic health benefits and cure systemic diseases . oil pulling is also supposed to increase the metabolism of the body , heal the cells , tissues and organs of the body and improve longevity in human beings . an improvement in oral hygiene is noticed within two weeks of practicing correct method of oil pulling . sunflower oil is observed to decrease both plaque and gingival indices after using it for oil pulling for period of forty five days . anand et al in their study observed 20% reduction in bacterial count upon 40 days of oil pulling using sesame oil . sesame oil was observed to possess moderate antimicrobial activity against s. mutans and l. acidophilus . they mentioned that toxins and bacteria from the body may be removed through the tongue and get trapped in oil and thrown out from the body . four researchers in their study involving 60 adolescents of age 1618 years with plaque induced gingivitis , observed statistically significant reduction of plaque and gingival indices upon oil pulling using coconut oil . subjects performed oil pulling in early morning at empty stomach in addition to their routine oral hygiene measures such as brushing and flossing . modified gingival index and plaque index by sillness and loe were measured at baseline and on days 1 , 7 , 15 and 30 . the study observed 50% decreases in gingival and plaque indices after four weeks which is comparatively similar to results produced by chlorhexidine . they concluded that oil pulling with coconut oil is helpful in decreasing plaque formation and plaque induced gingivitis . in a study conducted by jauhari d et al on children aged 612 years , the authors observed no significant reduction in s. mutans count using oratest and dentocult sm strip mutans kit after two weeks of oil pulling twice daily using sesame oil . however the result may probably be due to the fact that oil pulling may take atleast four weeks to show its effect . sesame oil is more palatable when compared to other refined edible oils . in an in vitro study on oral biofilm model , sesame oil was observed to possess antibacterial activity against s. mutans ; sunflower oil had antibacterial activity against c. albicans ; and coconut oil was active against both s. mutans and c. albicans . a group of researchers compared oil pulling method using sesame oil with chlorhexidine mouthwash for two weeks on twenty adolescent subjects . there was statistically significant reduction in the s. mutans count in the plaque samples of oil pulling group after one and two weeks . also there was reduction in the mean scores of salivary s. mutans count after two weeks . however the study noted that the reduction in s. mutans count is more in chlorhexidine group than oil pulling group . three authors assessed the effect of oil pulling on plaque , gingivitis and also its safety on oral hard and soft tissue . asokan et al in their in vitro study observed that benefits of sesame oil on oral health are due to saponification , emulsification and mechanical cleansing action . singla et al observed significant reduction in the mean s. mutans and lactobacillus count in saliva , and also in the plaque scores and gingival scores in 32 subjects upon gum massage using sesame oil , olive oil and coconut oil . the subjects massaged their gums around all the teeth with the 2 ml oil for 10 min every day after tooth brushing using index finger in circular motion for a period of three weeks . dani n et al assessed the antiplaque effect of oil pulling using sesame oil in a randomized controlled trial . also the effect of sesame oil pulling on plaque induced thereafter randomly 20 subjects were instructed to perform oil pulling for 14 days ; remaining 20 subjects were given chlorhexidine mouth wash for 14 days . plaque index scores , gingival index scores and total colony counts of aerobic bacteria were reduced in oil pulling group after 14 days . kuroyama et al reported two cases of exogenous lipoid pneumonia in patients who habitually practiced oil pulling with sesame oil . the symptoms of exogenous lipoid pneumonia are fever , weight loss , cough , dyspnea , chest pain , and hemoptysis . if the aspirated oil is microorganism rich , then it may result in manifestation of lipoid pneumonia . oral malodor is due to the proteolytic activity of three bacterial species porphyromonas gingivalis , tannerella forsythia and/or treponema denticola . volatile sulfur compounds like hydrogen sulphide , methyl mercaptan and dimethyl sulphide are responsible for oral malodor . sood et al in their three week randomized controlled trial involving sixty subjects observed that oil pulling with sesame oil was equally efficient when compared with chlorhexidine mouthwash in reducing oral malodor and the causative microorganisms . the researchers noted that sesame oil reduces volatile sulfur compounds and mean anaerobic bacterial count in the oral cavity and hence resulting in reduced mean objective and subjective organoleptic scores . one hindrance for oil pulling noted was the long duration of time required to perform the procedure . five researchers in a randomized controlled pilot trial involving 20 adolescent subjects concluded that oil pulling with sesame oil is as effective as chlorhexidine to reduce halitosis and microorganisms associated with it . the subjects performed oil pulling once daily for 1015 min before tooth brushing for 14 days . modified gingival index score , plaque index score , organoleptic breath assessment score , self assessment of breath score and bana test scores from tongue coating samples reduced in both chlorhexidine and oil pulling groups . in a randomized controlled triple blind study involving 20 age matched adolescents , the effect of sesame oil pulling on plaque induced gingivitis and its efficiency when compared to 0.12% chlorhexidine mouth wash was evaluated for a period of ten days . oil pulling was performed everyday for 1 min in the morning after tooth brushing . plaque index scores and modified gingival index scores were recorded at the baseline and after ten days . plaque samples were collected to evaluate the microorganisms present and to calculate the total colony count of aerobic microorganisms after ten days . the study found statistically significant reduction in the pre- and post - values of the plaque and modified gingival index scores in both groups . three authors carried out a microbiological study on the effect of sesame oil pulling on plaque , gingivitis and colony forming bacteria . twenty subjects of the study performed oil pulling for 1520 min daily for a period of forty five days . the study observed that oil pulling resulted in statistically significant decrease in plaque , gingival scores and bacterial counts . plaque scores , gingival scores and number of bacterial colonies increased after forty five days in the twenty subjects of the control group who practiced routine oral hygiene practices . the authors concluded that oil pulling with sesame oil is an effective preventive oral hygiene method when practiced daily . sesame oil does not have side effects like staining , lingering aftertaste , and allergy . oil pulling has also been claimed to cure several diseases such as arthritis , allergies , asthma , migraine headaches , nerve paralysis , kidney and heart disorders etc , though it is not scientifically proven.9 , 24 however it certainly has positive benefits on oral health . care should be taken that oil is not swallowed as it is heavily loaded with toxic microorganisms . however if accidently small amounts of oil is swallowed , there is nothing to worry as the same is removed from the body through feces . oil pulling is observed to bring improvement in oral hygiene when practiced correctly and regularly . limited available research on effect of oil pulling on oral hygiene shows promising benefits of oil pulling procedure on oral cavity . however oil pulling does not replace dental therapy and is currently not recommended by american dental association . extensive research on the role of this traditional , cheap and valuable remedy should be encouraged without bias . based on currently available research it can be concluded that oil pulling when performed as recommended , can be safely used as an adjunct to maintain good oral hygiene and health along with the routine tooth brushing and flossing with promising positive results . | oil pulling is a traditional folk remedy practiced in ancient india .
it is believed to cure more than thirty systemic diseases when practiced regularly and as directed . due to occurrence of side effects to modern medicines and oral hygiene products ,
people are increasingly attracted towards complementary and traditional practices .
oil pulling in addition to offering several oral health benefits has also beneficial effects on overall health .
the present article attempts to review and discuss this ancient practice . |
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asthma affects an
estimated 510% of children at ages 018 and is the
most common reason for
childhood hospitalizations . since 1980 , the national
prevalence of asthma has increased to 75% , with the greatest increase occurring
among children . in certain high - risk inner - city locations , asthma disproportionately affects minority
and low - income populations , with prevalence rates in african american and
latino children 2.5 times that of caucasians [ 4 , 5 ] . the implications of these data are
substantial , as children with asthma face increased risk for behavior problems ,
school dysfunction , and a variety of psychosocial problems which impact their
future development [ 68 ] . it has been 12 years
since the publication of the national asthma education and prevention program guidelines for
the diagnosis and treatment of asthma .
despite the widespread availability of these guidelines , there remains a
persistent and significant gap between what constitutes effective asthma care
for children and the actual care received , particularly for minority children
and those from low - income families . for example , minority and
low - income children are less likely than whites and children from households
with higher incomes to receive controller medications to prevent asthma
exacerbations . in one study of an
underserved urban area , researchers
found that 50% of children with asthma were not taking appropriate medications . extensive research
efforts have shown that appropriate management , including the routine use of
controller medications , can dramatically reduce asthma morbidity
in children [ 1113 ] . yet , translation of these
research findings into clinical practice has been slow , particularly for inner - city children from low - income families . several strategies have been devised to
increase the access to effective strategies for asthma care in underserved
areas . one of these strategies is the use of school - based asthma programs to bring
asthma care to a site where children are accessible on a daily basis , at least 180
days each year . evaluation of these programs has demonstrated increased student access
to needed care and improved students understanding of their health and
individual health outcomes ; [ 1416 ] however , the quality of care provided at these school - based clinics
has been inconsistent . a study of asthma care in inner - city school - based clinics found that the adherence to the national heart , lung , and blood
institute guidelines for asthma care was low among programs that did not have access
to expert asthma care and a targeted asthma intervention . other studies provide evidence that
asthma specialist involvement is a critical component of an effective asthma
program [ 1820 ] . although the evidence base is still growing ,
there are identifiable positive outcomes associated with telemedicine . telemedicine has been used in school settings
to improve access to care , treat otitis
media , and increase appropriate specialist referral and access to expert asthma
care for patients with asthma [ 10 , 22 , 23 ] . given these findings , we
hypothesized that we could improve access to effective asthma care for
underserved children by combining a school - based program and telemedicine to
increase access to asthma specialty care . we developed a
school - based program that identified children with asthma and used a
telemedicine link to elicit recommendations from an asthma specialist , which
were then communicated to the patient s primary care physician . we enrolled 96
subjects from 3 elementary schools in an inner - city neighborhood in san francisco . potential subjects were identified through
the use of school health records and/or a positive response on the
international survey of allergy and asthma in children ( isaac ) , which was
distributed to families of children in grades k-4 , indicating that they had asthma or had
experienced wheezing . children
between the ages of 5 and 12 who had a diagnosis of asthma on their school
health record or answered positively to the isaac survey were considered to be eligible
for study enrollment . children who were already under the care of an allergist or pulmonologist or spoke a language other than english or spanish were excluded from the study . this study was
approved by the stanford university school of medicine , institutional research board . each subject participated in 4
encounters at their respective schools occurring at baseline ( week 0 ) and weeks
8 , 16 , and 32 . at the baseline visit , an asthma specialist assessed each subject via a telemedicine link and developed an asthma action plan and recommendations based on the patient s
classification based on nhlbi guidelines . in this setting , telemedicine involved real - time video and audio conferencing between the patient / school nurse dyad on - site
at the school and the asthma specialist on site at san francisco general hospital . the asthma specialist directly interviewed the patient and family ( if present ) ,
observed an asthma - relevant examination by the on site nurse , and reviewed
spirometry data directly from the laptop computer used at the school site . at
the end of this visit , study personnel sent each subject s primary care physician a
letter informing them that their patient was a participant in the study and that
they would be receiving a comprehensive patient assessment with treatment
recommendations . we emphasized that we
would only provide recommendations and that implementation of any changes in
asthma management was the responsibility of the pcp . the results of the
assessment and the treatment recommendations were then communicated to the
primary care physician ( pcp ) using a web - based program called asthma
e - coordinator , fax , or mail , depending on their pcp s preference . the family was then told to schedule an appointment with their pcp and take their asthma action plan and treatment
recommendations to the visit . at the week-8 visit , the asthma specialist and school nurse assessed the need for changes in
therapy and communicated these recommendations to the pcp . at the week-16 visit each subject received formal ,
developmentally - appropriate asthma education using the american lung
association s open airways for schools curriculum . children in grades k-2 had two 30-minute sessions , and subjects in grades 35 received four 30-minute
sessions . parents were encouraged to attend all of these education sessions . finally , in the week-32 visit , all data collection tools were completed , and subjects graduated from the program . at the encounters of weaks 0 , 8 , and 32 , data collection included demographics , healthcare
utilization , and assessment of parental and child asthma knowledge . the parent satisfaction survey was given at
weeks 8 and 32 ; spirometry was conducted at baseline and week 8 . one parent or guardian completed the chsa at
baseline , 8 , and 32 weeks . the instrument
measures functional health status and is divided into 5 domains : physical
health of the child , social health of the child , social health of the family ,
emotional health of the child , and emotional health of the family . the test - retest
reliability , internal consistency , and construct validity of this instrument
have been reported previously [ 26 , 27 ] . one home visit , which
occurred between week-8 and week-32 visits , was made by the research nurse
who used a standard checklist to assess the environment relevant to potential
asthma triggers . the chsa also
provided data on the frequency of asthma symptoms as well as utilization data
including asthma - related inpatient stays , emergency department use , and
unscheduled outpatient visits . the
number of emergency department and unscheduled outpatient visits were summed up to
produce the total number of urgent care visits . the number of urgent care
visits , asthma - related inpatient stays , wheezing episodes , and asthma attacks
were included in the final analysis . pulmonary function tests were
conducted using a portable spirocard spirometer ( medgraphics , st . paul , minn , usa )
using percent predicted of normal based on height , sex , and race adjusted norms
using ats criteria . children were asked to perform a forced
expiratory maneuver after maximal inhalation to measure forced expiratory
volume in 1 second ( fev1 ) , forced vital capacity , and forced
expiratory flow in the middle half of the forced vital capacity and peak
expiratory flow rate ( pefr ) . a total of
3 maneuvers were recorded in accordance with american thoracic society
standards . we adapted a child
asthma knowledge survey from a previous study of the effectiveness of an asthma
nintendo game . similarly , we used
a 16-item survey developed for a previous study to assess parents evaluation
of their child s ability to manage their asthma . this survey was administered to one parent or
guardian at each relevant encounter after the baseline visit . in cases where
parents or guardians did not accompany the subject to the visit likewise , parental satisfaction was assessed
using a previously developed 8-item survey . to assess the
effect of the intervention , we compared outcomes between baseline and week 8
and baseline and week 32 . the outcomes
of interest included health care utilization and functional health status
( i.e. , five domains ) from the child health survey for asthma and child and
parental knowledge . because the study
did not collect spirometry data at week 32 , only baseline and week 8 are
compared . similarly , only week 8 and
week 32 values are compared for parental satisfaction . for those subjects missing follow - up data , we
used their baseline value to impute the outcome value at follow - up . because most of our outcomes were not
normally distributed , we used a nonparametric test , wilcoxons signed - rank
test , to perform statistical comparisons . this
research proposal was review by the institutional review boards of stanford
university school of medicine and the san francisco unified
school district . isaac surveys were
distributed to 680 students across the three schools ; 310 ( 45% ) of surveys were
returned . the prevalence rate for children with self - reported asthma or
wheezing on the isaac survey was 32% . twenty four percent of the
children did not have asthma indicated on their health cards and were identified only by their responses on the isaac
survey indicating that they had experienced an episode of wheezing or whistling
in the chest in the past 12 months . eighty three
( 86% ) of these identified asthmatic children agreed to participate in the study . seventy one
percent of the subjects were african - american , 14% were latino , and 14% were other . thirteen percent of children had spanish as their primary language spoken at
home , 78% of mothers had a high - school education or higher , and 92% of families
were medicaid eligible ( see table 1 ) . the study population reported mild to moderate levels of disease
activity based on nhlbi criteria . twenty four
percent of the study subjects did not have asthma identified on their school
health card ( see table 2 ) . sixty nine percent of
parents or guardians of children in the study were never told their children had
asthma . among children with identified
asthma , 96% stated that they used inhalers and 88% used spacers ; however , only
52% monitored their peak flow and 34% had asthma action plans . sixty percent of patients had persistent
asthma but only 23% of these persistent patients were taking anti - inflammatory
medications . twenty eight percent of
subjects had previously seen an asthma specialist but were included in the
study as they were not actively being followed by a specialist at the time of
enrollment . in
the home environment , 76% of subjects had carpeting on floor of the bedroom and
47% had feathered or down pillows . thirty six percent of
subjects had areas of mold or mildew in their home and 21% reported seeing cockroaches
in their home . it took
approximately 6 months to screen all eligible children , educate faculty , and
initiate the telemedicine encounters . all 83 children entered in the study were
seen by an asthma expert , underwent spirometry , and received an asthma action
and school emergency plan within three weeks of the baseline visit . the total
time commitment by the asthma specialist to see these children was 24 hours or
3.5 children / hr . ninety eight percent of the primary care physicians agreed to
participate in the study and received asthma action plans and treatment
recommendations . one hundred percent of
the 83 children completed the study . at the end of the study , 94% of parents
rated the asthma telemedicine program as excellent or good on a 5-point scale . scores on chsa physical domain and the social
activity of the family domain of the chsa were in the moderately high range
( 8090 ) , and scores in the emotional health of the child and family domains and
the social activity of the child domain were greater than 90 . similarly , mean spirometry
results showed forced expiratory volume at 1 second ( fev1 ) , forced
vital capacity ( fvc ) , fev1/fvc , and forced expiratory flow ( fef2575 )
that were all greater than 80% of predicted value . similarly , subjects had a low level of
utilization of hospital and emergency department visits as well as unscheduled
visits to their primary care physician . at the end of the
intervention , subjects demonstrated significant improvement in the physical and
social domains for child on the chsa ( table 3 ) . we were also able to demonstrate a significant improvement in child asthma knowledge and parent
asthma knowledge . in addition , we
observed a trend towards improvement in the number of asthma attacks in the
past 2 weeks ( table 4 ) . there were , however ,
no significant changes in spirometry , hospitalizations , emergency department
visits , or unscheduled visits to the primary care physician . this study
demonstrated both feasibility and acceptance of a school - based asthma program
that provided subspecialty access through a telemedicine link . we demonstrated that asthma specialists were
able to use their time efficiently to reach a large number of children . in fact , the project rate of 3.5 children per
hour was compared favorably with their experience in the asthma clinic where they
were able to see only 2 children per hour ( personal communication , shannon
thyne , february 2005 ) . we were also able
to demonstrate that children can easily undergo assessment and asthma education
during the school day with the assistance of a school nurse . the use of the school setting allowed for us
to complete the assessments and asthma education on 100% of our participants . we were able to enlist a high degree of participation among pcps as well as among
teachers . over 98% of the pcps were
willing to have their patients participating in the study . this mitigated any possible concerns on the part of the pcp that we may be taking their patients or fragmenting their care . this study also
demonstrated the feasibility of bringing subspecialty care to children with
asthma in the context of their school . several studies have documented improved outcomes when asthmatic
children have access to subspecialty care [ 20 , 30 , 31 ] . by using a telemedicine link , we were able to
increase the efficiency of the subspecialist s time and were able to ensure
that 100% of the study children , 92% of whom used california s version of medicaid ,
had access to subspecialty care . our study showed
that 24% of the students who screened positive for asthma symptoms were unaware
that they had asthma , which is higher than in a previous report that used more
rigorous spirometry criteria for the diagnosis of asthma . our study also found that a significant
proportion of students identified as having asthma were not receiving optimal
treatment with a low percentage having asthma action plans , measuring peak
expiratory flow rates and using spacers with their metered dose inhalers . these results are similar to other studies
that have demonstrated inadequate asthma care in urban
school - aged children [ 3 , 14 , 33 ] . though a significant proportion of our
subjects were classified as having mild and moderate persistent asthma according to
the child health survey of asthma , this degree of morbidity
was not reflected in the chsa . this may
be due to the fact that health status instruments reflect the degree of impact
on the child and family and may not be directly related to the disease
burden . because of the relatively high
scores on the preintervention subscales of the chsa , there was less
opportunity to demonstrate a more significant impact . previous work however has demonstrated that
the chsa is the most stable asthma outcome measure and best demonstrates
improvement over time . compared to baseline , health postintervention outcomes
were improved on two of the 5 subscales of the chsa . there are several possible reasons for a lack
of effect of the intervention on visits to the emergency department or the physician s
office . first , the number of subjects in
the study may have been too small to detect a significant difference in
relatively uncommon events . second , the
intervention did not ensure that the study subjects made a visit to their pcp
to have their medical regimen adjusted in light of recommendations from the
telemedicine encounter . in fact , many of
our families did not know the name or location of their pcp . this lack of an effective partnership between
our study families and their pcp made it difficult to leverage expert
recommendations and asthma action plans and ultimately decrease visits to the
emergency department and the physician office . lastly , the intervention for many of our children began in the spring
and with final outcomes assessed during the fall and winter . a lack of impact on utilization may have been
secondary to the increased incidence of respiratory infections during the fall
and winter months leading to increased symptom burden and utilization of health
care services . this study has
several important limitations . without a
control group , it is difficult to know whether or not our results may be
secondary to secular factors such as seasonality that are unrelated to the
intervention . second , we did not employ
the gold standard for measuring asthma outcomes , the number of days with
asthma symptoms . our previous work in
this community had raised serious doubts about the reliability of using diary
data to measure symptom days . it may be that the two - week recall window
used on the chsa was insufficiently sensitive to detect improvement in asthma
outcomes . third , the inclusion of
relatively low morbidity asthmatics , with low levels of utilization , relatively
high - pulmonary function , and high functional health status gave little
opportunity for statistically significant improvement with our intervention . the study was underpowered to show significant improvement in these mildly
effected outcomes , but did suggest several important trends in the positive
direction . another limitation
of this study was our inability to monitor and assess changes in the care
received from the primary care providers .
in an effort to maintain an appropriate link to the primary care
providers of our subjects , we decided to send the asthma specialist
recommendations to them for implementation rather than implementing these
recommendations ourselves . this model values the continuity and relationship
between primary care provider and subject , but did not ensure that the asthma
specialist recommendations were implemented . hence , the intervention tested was
actually the access to subspecialist recommendations rather than implementation
of these recommendations themselves . in their study of a school - based program
that prompted pediatric primary care physicians as to appropriate asthma care , halterman
and colleagues found poor compliance on the part of the primary care physicians
with the recommended care . thus , we suspect that inconsistent
implementation of the subspecialist recommendations by the primary care
provider negatively effected our outcomes . this study has demonstrated the
feasibility , efficiency , and acceptance of using a telemedicine link to bring
expert asthma care to underserved children in the school setting . the results also demonstrate the
effectiveness of the program in ensuring that children identified with asthma
receive a comprehensive assessment , asthma action plan , and asthma
education . the use of a telemedicine
link also allowed for a more efficient use of the asthma subspecialist s time
when contrasted with hospital - based asthma clinics . while the results suggest a positive
impact on asthma outcomes , a true assessment of program impact will require a randomized ,
controlled trial . | research has shown that access to an asthma specialist improves asthma outcomes .
we hypothesized
that we could improve access to expert asthma care through a telemedicine link between an asthma specialist and
a school - based asthma program .
we conducted a prospective cohort study in 3 urban schools to ascertain the
feasibility of using an asthma - focused telemedicine solution .
each subject was seen by an asthma expert at 0 , 8 ,
and 32 weeks .
the assessment and recommendations for care were sent to the primary care physician ( pcp ) and parents
were told to contact their physician for follow - up care .
eighty three subjects participated in the study .
subjects experienced
improvement ( p < .05 ) in family social activities and the number of asthma attacks .
ninety four percent of subjects rated the
program as good or excellent .
this study demonstrates the feasibility and acceptance of a school - based asthma program
using a telemedicine link to an asthma specialist . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
the prevalence of preterm birth , defined as delivery before 37 completed weeks ' gestation , varies from 6% to 15% , depending on geographical and demographic features of the population studied . preterm births account for 75% of perinatal deaths , with over two - thirds of these arising in preterm infants delivered before 32 weeks ' gestation . in 2002 , this was the most frequent cause of neonatal deaths in the usa . 65%75% of preterm births are defined as spontaneous preterm births caused by spontaneous preterm labor or preterm premature rupture of the membranes and 30%35% of preterm deliveries are medically indicated due to maternal or fetal complications in pregnancy . about one - quarter of preterm births occur in multiple pregnancies [ 15 ] . these encompass advanced age or teenage pregnancy , parity , race , history of cervical cone biopsy , low body mass index , tobacco use , assisted conception with in vitro fertilisation , and gamete intrafallopian transfer , especially for singleton gestations , systemic and genital - tract infection , and low parental socioeconomic status , this being the most important factor [ 14 , 6 ] . thanks to advances in neonatal medicine , the outcome for preterm infants born at or after 32 weeks ' gestation is similar to that of full - term infants . limits of viability have been lowered to a gestational age ( ga ) as low as 23 weeks at the expense of physical disabilities and long - term neurodevelopmental consequences [ 2 , 4 , 5 , 7 ] . thyroid hormones are essential for differentiation and maturation of the fetus and placenta and are especially important for the development of the fetal central nervous system . epidemiological studies have demonstrated that the intelligence quotient ( iq ) of the offspring correlates with maternal free thyroxine ( ft4 ) serum level . the higher maternal ft4 , the higher the iq of the offspring [ 813 ] . in a study with euthyroid thyroid peroxidase antibody ( tpoab ) positive pregnant women receiving substitution with l - thyroxine ( l - t4 ) it proved possible to decrease their preterm birth rate [ 1416 ] . the aim of this study is to investigate whether l - t4 replacement therapy by maintaining maternal ft4 in the high normal reference range prescribed for nonpregnant women can also lower the preterm birth rate in pregnant women who do not have thyroid autoimmune antibodies . for this purpose , a group of pregnant women , who had been treated with l - t4 with the goal of elevating maternal ft4 serum level to the high normal reference range prescribed for nonpregnant women to achieve an optimal fetal supply with thyroxine , were retrospectively examined . this clinical trial is a retrospective nonrandomized observational study with prospectively designed data . from april 2001 to march 2010 , all pregnant women below 12 weeks ' gestation presenting for their first medical consultation for prenatal care in a medical office were offered a serological thyroid scan . this included basal tsh , free triiodothyronine ( ft3 ) , free thyroxine ( ft4 ) , thyroglobulin antibody test ( tgab ) , thyroid peroxidase antibody test ( tpoab ) , and tsh receptor antibody test ( tsh - r - ab ) . the primary object was to elevate maternal ft4 to the high normal reference range for nonpregnant adults in order to optimise fetal brain development . for this purpose , and for better interpretation , the normal reference range for nonpregnant adults ( ft4 12.1419.62 the division was made as follows : the lower third ranged from 12.1414.72 pmol / l , the middle third ranged from 14.7217.17 pmol / l , and the upper third ranged from 17.1719.62 pmol / l . all women with an ft4 serum level in the lower and middle third of the reference range for nonpregnant adults at the first consultation were given a variable dose of l - t4 , usually starting with 25 g50 g l - t4 per day , after informed consent , to raise the ft4 serum level in the high normal reference range ( upper third ) for nonpregnant adults . all women with a physiological ft4 serum level in the high normal reference range ( upper third ) for nonpregnant adults at the first consultation were only serological assessed and l - t4 therapy was started later when serum ft4 declined to the middle or lower third . additionally , each woman received supplements containing 200 g iodide and 400 g folic acid . maternal ft4 serum levels were regularly assessed after an interval of 24 hours from the last l - t4 intake every 4 weeks during routinely performed serological scans for pregnancy care and if necessary , the l - t4 dose was adjusted . for adjustment , a variable dosage for example , between 2550 g if the increased dose was not tolerated , it was recommended to continue with the last tolerated dose and to increase it one week later . tsh was not further assessed because the goal was to avoid a low maternal ft4 serum level in pregnancy for optimal fetal brain development , and a decline of the ft4 serum level is not automatically accompanied by an increase of the tsh level . moreover , in pregnancy , tsh shows dependencies with other pregnancy - associated hormonal fluctuations and the interval between two blood samples of 4 weeks is too short for precise tsh readings . thyroid function control was handled by determining ft4 and not tsh as is the case when assessing central hypothyroidism [ 1820 ] . prenatal care included measurement of body weight , blood pressure , urinary test for protein , glucose , blood , and nitrite . check intervals were as follows : every 4 weeks until the 30 weeks ' gestation , thereafter every 3 weeks , weekly after the 35 weeks ' gestation , and after term every 2 days . beginning at 20 weeks ' gestation a digital vaginal examination was performed , and if necessary an ultrasound scan of the cervix length was done . if required , color doppler sonography and special ultrasound scan to rule out fetal malformation were undergone . a cardiotocographic survey of the fetus was routinely started at 28 weeks ' gestation and thereafter at each visit . women who developed preterm labor were treated with orally applied magnesium . in case of hospitalisation , treatment of preterm labor was performed according to hospital guidelines , for example , bed rest , intravenous fluids , tocolytic therapy , and steroid administration , if clinically indicated . all pregnant women were delivered in an obstetric unit in different hospitals in the vicinity . routinely , the newborns were medically examined by the gynaecologist who had assisted parturition immediately after birth . a further examination of the newborn was performed by a paediatrician between the 3rd7th day of life . after discharge , all children in germany have regular medical checkups performed by a paediatrician or family physician in private office in regular intervals beginning at 46 weeks after birth until adolescence . 68 weeks postpartum all women had a medical check for controlling uterine involution and after establishing normal serum ft4 levels , l - t4 substitution was discontinued . the study group included all women who fulfilled the following criteria : women with singleton pregnancies who had their first antenatal check before the 12 weeks ' gestation , determined by ultrasound ( crown - rump length ) . was designed as a retrospective nonrandomized observational study , we chose a surrogate as control group . the study was performed in the state of baden - wrttemberg , so we used as control group the preterm delivery rate from all 87.897 singleton pregnancies of the state of baden - wrttemberg , germany , in the year 2006 , collected in central database ( geqik ) with unknown thyroid status . the processed data , provided to us by the database geqik of the perinatal statistics of the state of baden - wrttemberg were related to preterm birth rate , maternal age and parity status . a further comparison of more obstetrical details and perinatal outcomes was not possible to realize . serum basal tsh and ft4 were measured using a third generation electrochemiluminescence immunoassay ( elecsys 1010/2010modular analytics e170 from roche diagnostics gmbh mannheim , germany ) . reference values for tsh were 0.274.2 iu / ml and for ft4 12.1419.62 pmol / l . intra- and interassay coefficients of variation were 3.0% and 7.2% for tsh , and 1.4% and 3.5% for ft4 . thyroid antibody titers were considered positive for tpo - ab titers above 34 u / ml , for tg - ab titers above 115 u / ml , and for the anti - tsh receptor - ab titers above 2.0 u / l . basis for data of the control group are population parameters statistical analysis took place by calculating the 99% confidence intervals ( 99%-ci ) of the values in the study group . between april 2001 and march 2010 , 771 pregnant women presented for medical care . among these , 96 ( 13% ) had a first trimester abortion , 12 ( 2% ) moved away to other regions , 18 ( 2% ) had multiple pregnancy , 87 ( 11% ) presented after 12 weeks ' gestation . 558 ( 72% ) women met the study inclusion criteria , being under 12 weeks ' gestation with singleton pregnancy . among these 558 women there were 108 ( 19% ) women with autoimmune thyroid antibodies , 43 primiparous , and 65 multiparous . by taking away these 108 women regarding the distribution of maternal age and parity status ( primiparous versus multiparous ) , the study group and control group were almost identical . in the study group , 39.8% were primiparous versus 39.2% in control group , and 60.2% were multiparous in study group versus 60.8% in control group . dictated by body mass index ( bmi ) , 61% of the primiparous group had normal weight at first consultation , and 39% were overweight or obese . of the multiparous group , 60% had normal weight at first consultation and 40% were overweight or obese . there was a similar mean weight gain of 1015 kg for primiparous and multiparous in pregnancy . peripartum outcome and fetal outcome resulted as follows : 68% of primiparous and 73% of multiparous had vaginal delivery within the study group . cesarean section occurred in 34% of the primiparous ( 15% were elective , and 19% were emergency cesarean section ) and in 28% of the multiparous ( 17% were elective , and 11% were emergency cesarean section ) . pregnancy - induced hypertension occurred in 9 ( 2% ) women of the study group ( 5 primiparous and 4 multiparous ) . data from 2 women ( 1 primiparous and 1 multiparous ) are missing . in the study group 22 ( 4.8% ) women had breech presentation at term ( 11 were primiparous and 11 were multiparous ) . data for apgar score and arterial umbilical cord ph were almost complete . an apgar score below 3 , as a sign of impaired vitality was not registered at all and only 2 ( 0.5% ) newborns had an arterial umbilical cord ph below 7.00 , demonstrating asphyxia . apart from preterm births that were all referred to a pediatric hospital , further 35 newborns were referred to a pediatric hospital , 18 ( 11% ) born by primiparous , and 17 ( 6% ) born by multiparous . only 16% of primiparous and 18% of multiparous had ft4 serum level in the high normal reference range prescribed for nonpregnant women at first consultation . preterm birth rate was first evaluated for the whole study group and separately depending on parity status , primiparous versus multiparous . in the study group there were 20 preterm births ( 4.4% ) , 13 in primiparous ( 7.3% ) and 7 in multiparous ( 2.6% ) . 70% of preterm births from the study group were spontaneous preterm births caused by spontaneous preterm labor or preterm premature rupture of the membranes and 30% of preterm births were medically indicated by pregnancy - induced hypertension , hellp syndrome , ovarian tumor , and acute pancreatitis concurring with obesity . preterm birth rate in the study group was 4.4% ( 99%-ci 1.9%6.9% ) versus 7.1% in the control group corresponding to a reduction of 38% . the subgroup analysis , according to parity status showed a preterm birth rate for primiparous in the study group of 7.3% versus 7.6% in the control group , and for multiparous a preterm birth rate of 2.6% in study group ( 99%-ci 0.1%5.1% ) versus 6.7% in the control group , respectively ( table 2 ) . thus , the reduction of preterm birth rate by maintaining maternal ft4 serum level in pregnancy in the high normal reference range prescribed for nonpregnant women is effective in multiparous but not in primiparous . the preterm birth rate dropped by approximately 61% in the multiparous of the study group . since there was no fixed dosage for l - t4 therapy and patients were advised to maintain the dosage unchanged in case of intolerance when dose augmentation had to occur , side effects were rarely noted . nevertheless , if patients experienced adverse reactions , the l - t4 dose was reduced . moreover , undesirable side effects for the fetus and newborn such as tachycardia or other signs of induced hyperthyroidism did not occur at all and were not recorded during cardiotocographic controls . due to the variable l - t4 dose and the intake regime , l - t4 was well tolerated by pregnant women and complaints such as palpitations , tachycardia , and other undesirable clinical signs of hyperthyroidism were very rare and transient . however , pregnancy - associated nausea was more pronounced , predominantly in the first trimester . to our knowledge this is the first study presenting an essential benefit for reducing preterm birth rate in euthyroid multiparous women without thyroid autoimmune antibodies by keeping maternal ft4 serum level in the high normal reference range prescribed for nonpregnant women with l - t4 substitution . the preterm birth rate for multiparous in the study group declined by 61% . possibly , the pathomechanism of preterm birth is essentially different among primiparous compared to multiparous , which would imply the necessity of different therapeutic strategies . casey et al . found no mentionable risk for pregnancy outcome if the ft4 serum level was found to be in the lowest third of the gestational age specific reference range . recently compared pregnancy outcome in women with subclinical hypothyroidism and in women with hypothyroxinaemia defined as ft4 serum level below the 2.5th percentile . they failed to determine a link between subclinical hypothyroidism and adverse pregnancy outcome and hypothyroxinemia was not associated with the majority of pregnancy complications [ 21 , 22 ] . in this study , a high maternal ft4 serum level decreased preterm birth rate in multiparous by yet unknown mechanisms . in some studies , it has been shown that human chorionic gonadotropin ( hcg ) plays a part in maintenance of uterine quiescence in the third trimester , and hence could be an endogenous tocolytic agent . hcg exerts a potent myometrial relaxant effect in human myometrium in the third trimester and inhibits preterm delivery in animals . thyroid hormone stimulates the synthesis of hcg and the level of serum thyroid hormone is a positive regulator of serum thyrotropin bioactivity . due to the fact that tsh and hcg share a certain similarity , being members of the same glucoprotein family , it appears possible that serum thyroid hormones can also positively regulate hcg bioactivity , which in turn increases the biological effects on the myometrium until parturition [ 1 , 4 , 14 , 23 ] . the study group received l - t4 substitution to avoid a low maternal ft4 serum level for an optimal fetal brain development . the prevalence of low maternal ft4 levels is probably 150200 times more common than congenital hypothyroidism [ 24 , 25 ] . it was demonstrated that if maternal serum ft4 is low , fetal t3 levels in the brain will be low even in the presence of normal maternal and fetal serum ft3 , suggesting that both t3 and t4 in the fetal brain are dependant on maternal ft4 . low maternal ft4 serum levels could have detrimental effects on fetal brain development . in animal experiments , it was demonstrated that even a modest and transient decrease in maternal ft4 resulted in altered brain histogenesis and cytoarchitecture of the fetal cerebral cortex [ 26 , 27 ] . various observational studies have shown that maternal subclinical hypothyroidism and low ft4 serum levels caused a significant decrease in iq scores of the progeny . in contrast to these findings , only one study failed to find an association between maternal thyroid function and cognitive test scores in children . handling thyroid function control by assessing only ft4 for a goal - oriented value as was done in this study and which is similar to controlling central hypothyroidism carries the risk to over dosage l - t4 and to provoke potentially metabolic hyperthyroidism . the usual assessment of an adequate l - t4 dose in replacement therapy is done by determining tsh and ft4 levels , preferably in a blood sample taken before ingestion of the subsequent l - t4 dose . in nonpregnant states it takes 69 weeks and more to normalize a suppressed tsh value typically found when initiating l - t4 administration . as a consequence , it has been shown that patients taking 100 to 150 g / d of l - t4 have a nearly 50% probability that serum tsh will be undetectable . determining tsh below an interval of 69 weeks would be likely to result in a suppressed tsh level but would not necessarily be accompanied by an ft4 serum level in the upper third of the reference range . furthermore , l - t4 replacement therapy is commonly associated with supraphysiologically high ft4 and low ft3 serum levels , in general without hyperthyroid symptoms . since ft3 serum level is maintained within normal limits by decreased peripheral conversion of the prohormone l - t4 , supraphysiological ft4 serum levels are not considered to be harmful . therefore , supraphysiological serum ft4 levels in patients taking l - t4 are not necessarily accompanied by clinical consequences , if there are no signs or symptoms of clinical toxicity . clinical experience with pregnant women on tsh suppressive thyroid therapy after thyroid cancer treatment silva and larson have shown that t4 is preferentially converted to t3 in the pituitary gland to a greater degree than in other tissues , so that its tsh suppressive effect is greater than its metabolic effect . in other words , exogenously administered l - t4 suppresses both the pituitary gland and the thyroid gland as well . in contrast to supraphysiological ft4 serum concentration seen in l - t4 replacement therapy , hyperthyroidism is defined as an excessive thyroid hormone production due to thyroid overactivity . the vast majority of cases of hyperthyroidism in pregnancy are induced by graves ' disease , toxic adenoma , or thyroid hormone resistance , where the negative feedback mechanism no longer functions . in these pathological conditions the overactive thyroid gland secretes both the metabolically inactive prohormone t4 and the metabolically highly active hormone t3 causing multiple symptoms . in graves ' disease , the intrathyroidal type - ii deiodinase ( d2 ) , which activates thyroid hormone , has a 50 to 150 fold higher activity than in placenta and contributes significantly to the intrathyroidal t3 production and secretion . hyperthyroidism caused by graves ' disease and toxic multinodular goiter , high t3 concentrations are the result of excessive production and release from the thyroid gland and not of peripheral deiodination . this explains why patients with graves ' disease or with toxic adenoma present symptoms of hyperthyroidism in contrast to patients taking l - t4 in a tsh suppressive dose . additionally , in replacement therapy with a tsh suppressive l - t4 dose , t3 is derived completely from peripheral monodeiodination in the liver , kidney , or muscle because the thyroid gland is also suppressed . to achieve physiological levels of t3 in humans treated with l - t4 , it is necessary to maintain ft4 levels at the higher end of the normal range [ 46 , 47 ] . only under pathological conditions , such as massive metastatic follicular thyroid cancer has t3 thyrotoxicosis by increased conversion of administered l - t4 been described . l - t4 replacement therapy is associated with supraphysiologically high ft4 and low ft3 serum levels without hyperthyroid symptoms , since the ft3 serum level is maintained within normal limits by decreased peripheral conversion of the prohormone l - t4 . this metabolic variance constitutes a fundamental difference between endogenously produced thyroid hormones t4 and especially t3 and exogenously administered thyroid hormone like l - t4 . l - t4 has a safe latitude in dosage and even after massive acute ingestion only minimal symptoms are seen as the peak t3 level does not exceed the upper reference range limit , while ft4 and the metabolically inactive rt3 show very high serum levels [ 41 , 43 , 45 , 47 , 4956 ] . disputable side effects are the risk of atrial fibrillation in patients with intrinsic heart disease and increased bone loss in postmenopausal women [ 57 , 58 ] . since there was no fixed dose regimen for l - t4 therapy in the study group , patients were advised to maintain their dosage unchanged in case of intolerance , and when dose augmentation proved necessary , side effects rarely occurred and in none l - t4 had to be discontinued . nevertheless , if patients experienced adverse reactions the l - t4 dose was reduced . moreover , undesirable side effects for the fetus and newborn such as tachycardia or other signs of induced hyperthyroidism did not occur at all and were not recorded during cardiotocographic controls . however , pregnancy - associated nausea was more pronounced , predominantly in the first trimester in only a few women . by reducing the l - t4 dosage , pregnancy - associated nausea resolved . in summary , thyroid hormone replacement therapy aiming at holding maternal ft4 serum levels in the upper third of the reference range prescribed for nonpregnant women and controlling this therapy by determining ft4 seems to be safe and to have beneficial effects for both mother and fetus . in all probability | preterm birth is the most common reason for perinatal morbidity and mortality in the western world .
it has been shown that in euthyreotic pregnant women with thyroid autoimmune antibodies , l - thyroxine replacement reduces preterm delivery rate in singleton pregnancies .
we investigated in a nonrandomized retrospective observational study whether l - thyroxine replacement , maintaining maternal free thyroxine serum level in the high normal reference range prescribed for nonpregnant women also influences the rate of preterm delivery in women without thyroid autoimmune antibodies .
as control group for preterm delivery rate , data from perinatal statistics of the state of baden - wrttemberg from 2006 were used .
the preterm delivery rate in the study group was significantly reduced .
the subgroup analysis shows no difference in primiparous but a decline in multiparous by approximately 61% with l - thyroxine replacement . maintaining free thyroxine serum level in the high normal reference range prescribed for nonpregnant women may reduce the preterm delivery rate . |
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some literatures have emphasized that occupational exposure , cigarette smoking and drinking are associated with infertility , while other studies have excluded these associations with infertility . some of lifestyle factors may be linked to varicocele in infertile men , such as smoking , alcohol consumption , and occupational exposure , these are associated with varicocele and increase the risk of infertility . therefore , further research is needed in order to elucidate the associations of smoking , occupation , and drinking with development of varicocele in male infertile . inclusion criteria for the study were men with infertility who presented to our center between april 2010 and february 2011 . a total of 816 patients were selected and reviewed based on inclusion criteria . demographic data , including age , education level , residence , occupation , type of infertility , age at marriage , duration of infertility and data about smoking habits and alcohol consumption was summarized in table 1 . chracteristics of male infertile in groups with and without varicocele all subjects were classified as presence or absence of varicocele . varicocele was diagnosed in the center during physical examination in the upright position , and confirmed using the scrotal ultrasonography . the ethical committee of medical faculty of babol university of medical sciences approved the study . spss software version 16.0 ( spss inc , chicago , usa ) was used for statistical analyses . the final multivariate model that included varicocele as dependent variables were related to the outcome ( p = 0.2 in bivariate analyses ) . adjusted regression analysis was used to test associations between categorical occupations , smoking , and drinking with varicocele . all independent variables that met the above criteria were included in the multiple logistic regression . spss software version 16.0 ( spss inc , chicago , usa ) was used for statistical analyses . descriptive statistics were used to describe baseline demographic . the final multivariate model that included varicocele as dependent variables were related to the outcome ( p = 0.2 in bivariate analyses ) . adjusted regression analysis was used to test associations between categorical occupations , smoking , and drinking with varicocele . all independent variables that met the above criteria were included in the multiple logistic regression . the overall prevalence of varicocele accounted for 32.0% ( ci 95% ; 28.1 , 36.0 ) among the infertile men . moreover , varicocele accounted for 32.2% ( ci 95% , 29.3 , 37.1 ) of patients with primary infertile , and 28.5% ( ci 95% ; 26.5 ; 30.5 ) with secondary infertile . the mean age was 31.6 6.4 years old ( 21 - 71 years ) . the mean age in the group with varicocele ( 30.9-years old ) was significantly younger than that without varicocele ( 31.9-years old ) ( p = 0.019 ) . the mean age at current marriage was 25.6 4.9 for varicocele and 26.4 5.7 for non - varicocele , there was a significant difference between them ( p = 0.02 ) . the mean body mass index ( bmi ) and infertility duration had no significantly difference in those with or without varicocele . chi - square test showed no significant relation between the varicocele group and education level . table 2 presents the estimated adjusted odds ratio ( with 95% ci ) for varicocele in relation of occupation , smoking habits , and drinking alcohol in male infertile . the adjusted or in smokers was higher than those in the non - cigarette - smoking individuals ( or = 2.42 , 95% ci = 1.04 , 5.61 ) . in no significant association . adjusted ratio ( or ) for varicocele according to occupation , smoking habits , and drinking of the subject ( n = 816 ) most of the studies have showed a higher prevalence of primary infertility compared with secondary infertility . however , men with the secondary infertile have an opportunity for future fertility , because varicocele is more frequently common in secondary infertile men . in this study , approximately 75% of infertile men had primary infertility and over 25% had secondary infertility ; around 32% of infertile men had varicocele . however , nielson reported 15%-20% of men with infertility had varicocele . a possible explanation for relatively higher prevalence of varicocele in infertile men in our study may be due to the various somatometrics parametric and environmental factors . in addition , those individuals with mild varicocele should not be missed in physical examination . despite a large number of researches regarding the effect of smoking on male infertility , it is suggested that smoking may have a greater adverse influence on the varicocele . in the present study cigarette smoking increase an accumulation of cadmium in testes , and may cause testicular damage in men with varicocele . men with varicocele seem more frequently to suffer from infertility than the men without varicocele . a large size of prospective study is , therefore , needed in order to clarify the importance of smoking in relation to varicocele risk factors among infertile men . some recent studies , which focused on the impact of occupation and alcohol on infertility , showed that alcohol intake and occupational factors influenced the reproductive system . although the role of alcohol intake and occupation on varicocele is unclear , our results do not support the hypothesis that the alcohol intake and occupational factors such as industrial and agriculture workers is closely related to varicocele in infertile men . therefore , a further prospective study is proposed to elucidate the association of alcohol and occupation with varicocele risk factor in male infertile . the present study examined a convenience sample , the studies in the future should use population - based random sample , which may provide stronger evidence with the associations between varicocele and infertility . in addition , the unavailability of some information , such as exact nature of men occupation , and amount of drinking and smoking caused an decrease in the number of variables in our present study . finally , this study did not assess the causality , whereas a case - control study could represent more appropriate in assessing the causes or risk factor associated with varicocele . the present study examined a convenience sample , the studies in the future should use population - based random sample , which may provide stronger evidence with the associations between varicocele and infertility . in addition , the unavailability of some information , such as exact nature of men occupation , and amount of drinking and smoking caused an decrease in the number of variables in our present study . finally , this study did not assess the causality , whereas a case - control study could represent more appropriate in assessing the causes or risk factor associated with varicocele . this percentage was nearly comparable with the findings from other studies . despite the limitation , therefore , further studies are needed to make a recommendation or guideline for a better prevention and management of varicocele . | background : existing data suggests that varicocele plays a central role in progressive infertility.aims:this study was designed to assess the occupational and lifestyle factors of development of varicocele among male infertile , for a better prevention and management of the varicocele.materials and methods : all males with infertility , who presented at fatemezahra infertility and reproductive health research center between april 2010 and february 2011 , were examined .
their occupations , smoking , and drinking alcohol , presence or absence of varicocele were evaluated .
the lifestyle factors associated with varicocele were analyzed.results:the data of 816 men , aged 21 - 71 years , were included in the study .
two hundred and sixty - one men ( 32% ) with varicocele and 555 ( 68% ) without varicocele were found .
percentage of varicocele was significantly higher in smokers compared with non - smokers ( p = 0.035 ) .
the adjusted or for varicocele in smokerswas significantly higher than that in non - smoker ( or = 2.420 ; 95% ci = 1.04 , 5.61 ) .
no significant differences were seen between varicocele and occupation or alcohol drinking for the prevalence of varicocele.conclusions:the findings of the present study indicated that a high frequency of varicocele was indentified among male infertile .
therefore , it is necessary for those male cigarette - smokers to evaluate their possibilities of infertility and varicocele . |
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metabolic syndrome ( mets ) is recognized as the clustering risk factors of obesity , insulin resistance , dyslipidemia and hypertension associated with the subsequent development of cardiovascular disease and type 2 diabetes . in addition to adults , the number of children and adolescents affected by mets is increasing . the prevalence of mets in children and adolescents is relatively low ( 4.2% ) when compared with adults ( 34% ) , but the overall prevalence of mets in moderate and severe obese subjects were 38.7% and 49.7% , respectively . the prevalence of mets in tehranian adolescents varied from 0.7% to 15.1% by different definitions , and on a larger scale , this prevalence was 14.1% for iranian adolescents defined based on criteria analogous to those of the adult treatment panel iii ( atpiii ) . some studies reveal that there is an independent and inverse association between pa and metabolic risk factors . but because of different definitions and prevalence of mets and the various methods used to determine levels of physical activity ( pa ) , it is difficult to compare the results of them . mets is increasing worldwide , and it has been shown that the prevalence of mets in overweight / obese children and adolescents is higher than in normal weight individuals ; therefore , obesity is strongly associated with the mets , and the beneficial effects of pa on mets could be a result of its influence on body mass index ( bmi ) and waist circumference ( wc ) . tehran lipid and glucose study ( tlgs ) is a population - based study , which was conducted to determine the prevalence of non - communicable diseases among tehran 's urban population and to develop population- based measures to decrease the prevalence or prevent the rising trend of diabetes mellitus and dyslipidemia . our study aimed to examine the association between pa and mets in addition to its individual components among normal weight and overweight / obese adolescents who participated in the tlgs . this cross - sectional study conducted within the framework of the tlgs , which occurs in the urban population of tehran . for tlgs , a multistage stratified cluster random sampling technique was used to select 15,005 people aged 3 years and over from district 13 of tehran ( latitude 354 ) , the capital of iran . the district is located in the center of tehran , and the age distribution of its population is representative of the overall population of tehran . the crude response rate was approximately 55.6% , and there was no significant difference between respondents and non - respondents in terms of age and gender distribution . for the current study , 1230 adolescents , aged 12 - 18 years , were selected by a multistage cluster random sampling from among this population , but after the exclusion of individuals who had thyroid diseases and diabetic mellitus or used drug for these diseases , 777 participants with complete relevant data / values for investigation remained . written informed consent was obtained from participants and approval for the study was granted by the ethics committee of the research institute for endocrine sciences , affiliated to the shahid beheshti university of medical sciences . information on age , physical activity status , education levels and medication usage for treatment diabetes , hypertension and lipid disorders was collected . weight was measured without shoes and heavy clothes , using digital scales was recorded to the nearest 100 g. standing height was measured without shoes , using a tape to the nearest 0.1 cm , while the shoulders were in a normal position . bmi was calculated as weight in kilograms divided by the square of height in meters ( kg / m ) . wc was measured at the umbilical site using an outstretched tape meter and without pressure to body surfaces and was recorded to the nearest 0.1 cm . systolic and diastolic blood pressures ( bp ) were measured with a qualified physician after 15 minutes of rest , while the subject was in a seated and relaxed position , two recordings were made at a 10-minute interval , measurements were taken in millimeters of mercury ( mm hg ) and the mean value of the 2 recordings ( not varying by more than 5 mm hg ) was calculated . fasting blood samples of 5 ml were taken by a trained laboratory technician . the day before the test , individuals were instructed to fast for 12 hours and their adherence to this protocol was confirmed on the morning of the examination before drawing blood . fasting blood glucose ( fbg ) was measured on the day of blood collection by the enzymatic colorimetric method using glucose oxides . triglyceride ( tg ) and total cholesterol ( chol ) concentrations were measured by commercially available enzymatic reagents ( pars azmoon , tehran , iran ) . high - density lipoprotein cholesterol ( hdl - c ) was measured after precipitation of the apob containing lipoprotein with phosphotungstic acid . individuals were asked to report the physical activities in which they had participated during the past 12 months , in addition specifying the frequency and duration for each activity identified . each activity was weighted by its relative intensity , referred to as metabolic equivalent of task ( met ) . one met represents the energy expenditure for an individual at rest ( 1 met = 3.5 ml.kg.min of o2 consumption ) . for all activity levels , obtained met was multiplied by the time spent at each level . met - time from each level was added to total 24 hour met- time , representing the average daily level of pa . in this study we categorized the pa levels as light ( met < 3 ) , moderate ( 3 met < 6 ) and vigorous ( met 6 ) intensity . for this study , subjects were classified as having mets according to the cook 's guidelines . this definition is based on criteria analogous to that of the national cholesterol education program expert panel on detection , evaluation and treatment of high blood cholesterol in adults treatment panel iii ; it defines mets as having three or more of the following : fasting tg 110 mg / dl ( 1.24 mmol / l ) ; hdl - c 40 mg / dl ( 1.04 mmol / l ) ; wc 90 percentile for age and sex , according to the national reference curve ; sbp and/or dbp 95 percentile for sex , age and height from the fourth report on the diagnosis , evaluation and treatment of high blood pressure in children and adolescents ; fbg 100 mg / dl ( 5.55 obesity was defined based on the standardized percentile curves of bmi suggested for iranian children and adolescents . overweight was defined as 85 percentile to < 95 percentile for age and sex , while normal weight was determined as < 85 percentile of bmi for age and sex . education levels for participants mothers were categorized as : elementary high school , diploma , and bachelor degree or higher . the kolmogorov - smirnov goodness - of - fit test was used to assess the normal distribution of continuous data . normally distributed continuous variables are reported as the mean standard deviation ( sd ) whereas categorical variables were summarized as frequencies and percentage . logarithmic transformation was performed to normalize the distribution of fbg and tg . to compare the levels of mets components among normal weight and overweight / obese individuals , independent sample t test for variables with normal distribution and mann - whitney u test for variables that was not normally distributed , multiple logistic regression was used to assess the association between pa as well as other covariates such as sex , age and maternal education with mets . for this study , subjects were classified as having mets according to the cook 's guidelines . this definition is based on criteria analogous to that of the national cholesterol education program expert panel on detection , evaluation and treatment of high blood cholesterol in adults treatment panel iii ; it defines mets as having three or more of the following : fasting tg 110 mg / dl ( 1.24 mmol / l ) ; hdl - c 40 mg / dl ( 1.04 mmol / l ) ; wc 90 percentile for age and sex , according to the national reference curve ; sbp and/or dbp 95 percentile for sex , age and height from the fourth report on the diagnosis , evaluation and treatment of high blood pressure in children and adolescents ; fbg 100 mg / dl ( 5.55 mmol / l ) . obesity was defined based on the standardized percentile curves of bmi suggested for iranian children and adolescents . overweight was defined as 85 percentile to < 95 percentile for age and sex , while normal weight was determined as < 85 percentile of bmi for age and sex . education levels for participants mothers were categorized as : elementary high school , diploma , and bachelor degree or higher . the kolmogorov - smirnov goodness - of - fit test was used to assess the normal distribution of continuous data . normally distributed continuous variables are reported as the mean standard deviation ( sd ) whereas categorical variables were summarized as frequencies and percentage . logarithmic transformation was performed to normalize the distribution of fbg and tg . to compare the levels of mets components among normal weight and overweight / obese individuals , independent sample t test for variables with normal distribution and mann - whitney u test for variables multiple logistic regression was used to assess the association between pa as well as other covariates such as sex , age and maternal education with mets . participants comprised 457 boys and 320 girls with a mean age of 15.2 1.9 years . in this study , 257 adolescents were overweight / obese ( bmi 85 percentile ) while 520 of them had normal weight ( bmi < 85 percentile ) . the overweight / obese group had significantly higher means for weight , wc , bmi , tg and dbp than normal weight group . the prevalence of mets was higher in overweight / obese groups ( 35% ) compared with normal weight ones ( 3% ) . individuals with normal weight were more physically active than overweight / obese ones ( 50.4% vs. 44% ) , as shown in table 1 . the mean sd daily time spent performing vigorous physical activities ( pa 6met of intensity ) was 0.7 0.3 h / day in the overall population ; this time was 0.8 0.4 and 0.6 0.2 h / day in the normal weight and overweight / obese group , respectively . figure 1 illustrates the components of mets based on the pa tertiles in bmi groups . in the normal weight group , dbp was significantly different between the pa tertiles ( p : 0.02 ) . comparing the components of mets among normal weight and overweight / obese groups in each tertiles of pa showed that the means sd of these variables were significantly higher in the overweight / obese group except for hdl - c and fbg . table 2 displays the odds ratio ( 95% confidence interval ) of the association between the mets and pa among the normal weight and overweight / obese group , for two models of logistic regression . in normal weight individuals , there was a significant association between the light pa and the risk of lower levels of hdl - c ( or = 1.61 ; ci 95% 1.11 , 2.35 ; p : 0.01 ) , an association which remained significant even after adjustment for age , sex and maternal education covariates ( or = 1.65 ; ci 95% 1.12 , 2.44 ; p : 0.01 ) . the overweight / obese subjects with light and moderate pa levels ( 1 and 2 tertiles ) had a higher level of wc than those who participated in vigorous pa only after adjustment for determined confounders ( or = 1.11 ; ci 95% 1.07 , 1.21 ; p : 0.01 ) ( or = 1.06 ; ci 95% 1.01 , 1.08 ; 0.02 , respectively ) . the results of the logistic regression analysis revealed no association between the higher risk of mets and pa . demographic and biochemical characteristics of tlgs adolescents in bmi groups means of mets components by physical activity levels among normal weight and overweight / obese adolescents association between physical activity levels and the metabolic syndrome components among tlgs adolescents the result of the current study shows that in the normal weight group , before and after adjustment for potential confounders , the risk of having lower levels of hdl - c was approximately 65% higher in adolescents with a sedentary lifestyle , compared to individuals who had vigorous pa level . furthermore , in the group of adolescents with overweight and obesity , those with light and moderate pa , had 11% and 6% more probability of having abdominal obesity ; however , there was no association between pa and other components of mets and its clustering . although there are several studies in various populations that have assessed the association between pa and mets , these studies differ from each other in terms of mets definition used , the questionnaire for pa , adjustment for confounders and continuum of age and sex , and in subjects studied , leading to some difficulties in their comparison . for instance , the results of conducted study on 4811 iranian individuals , aged 6 - 18 years using the atpiii criteria for mets definition and ipqa questionnaire for pa , showed that pa has an inverse association with mets ; the difference between results of this study and ours could be related to the different mets definitions and pa questionnaire used . in addition , this study was performed on children and adolescents , whereas our study was related to adolescents aged 12 - 18 years . another study from vietnam , which was done on 693 high school students , revealed that lifestyle with moderate or vigorous pa is associated with lower risk of mets ; the authors of this study believed that socio - economic status has an important effect on this association . it is important to note that , in our study , we did not take into account the role of socio - economic status . , who assessed the effect of overweight on the metabolic risk factors and the role of pa in 68 pre - pubertal children , showed that metabolic risk factors elevated independently of lower pa levels , among overweight children ; the lack of association could be explained by the inadequate number of subjects . in agreement with our results , there are studies showing the association between pa and each individual component of mets . the results of a study that assessed the association of leisure - time physical activity ( ltpa ) with mets and its components using self - reported ltpa on 542 adolescents , who were 13-years - old , showed that ltpa was associated with bmi , hdl - c and sbp as the mets components . similar to the above - mentioned study , in this study , we found that pa has significantly affected some components of mets such as hdl - c and wc . the limitations of the current study merit consideration : first , in addition to pa , diet and socio - economic status would be related to mets , therefore assessing the effect of these confounders on the association between pa and mets is useful , but we had no data on the diet and socio - economic status for the selected individuals . second , it was impossible for us to confirm individual 's pa reports with other techniques such as an accelerometer , which is used for recording activity count . third , we did not have data regarding puberty status , which would help us to assess the effects of puberty on mets . despite these limitations , we recruited our samples from among the participants of a large ongoing study in tehran , named tlgs ; hence the results of the present study could be representative for tehranian adolescents . the results of this study confirm the association between pa and individual components of mets as wc and hdl - c . | background : physical activity ( pa ) is associated with the metabolic syndrome ( mets ) and its components .
this study aimed to examine the association between pa and mets and its components among normal weight and overweight / obese adolescent in tehran lipid and glucose study ( tlgs).methods : this cross - sectional study includes 777 adolescents , aged 12 - 18 years , who were selected by multi - stage random cluster sampling from among tlgs participants .
subjects were classified as normal weight and overweight / obese based on the age- and sex - specific standardized percentile curves of bmi for iranian population .
levels of pa were assessed using a standardized and modifiable activity questionnaire ( maq ) , and categorized into tertiles .
mets was defined according to the cook 's criteria.results:prevalence of the mets was higher in overweight / obese than normal group ( 35% vs. 3% ; p : 0.02 ) .
normal groups were more physically active ( 50% vs. 44% ) ; however , difference was not significant .
there was a significant association between the light pa and risk of lower level of hdl - c before and after adjustment , in normal weight group ( or : 1.61 , ci 95% : 1.11 , 2.35 ; or : 1.65 , ci 95% : 1.12 , 2.44 , respectively ) .
the overweight / obese group with light and moderate pa had a higher risk of having abdominal obesity than those with vigorous pa , only after adjustment for determined covariates ( or : 1.11 , ci 95% : 1.07 , 1.21 ; or : 1.06 , ci 95% : 1.01 , 1.08 , respectively ) ; the association between mets and pa was not significant.conclusions:the results of this study confirm the association between pa and some individual components of mets such as waist and hdl - c . |
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synthetic
chemical dispersants are used for marine oil spills to
limit petroleum accumulation at the surface of the ocean and to accelerate biodegradation processes by
increasing the oil water interfacial area . oil
released from an underwater offshore source as in the deepwater horizon
spill can generate surface oil slicks containing large quantities
of hydrocarbons and introduce soluble organics into the water column . dispersants enhance the emulsification and dissolution
of oil into the water , which augments degradation and prevents slick
formation , reducing the amount of oil that reaches shorelines . however , chemical dispersants can increase the concentration of
petroleum hydrocarbons in the water column , enhancing their bioavailability
for marine organisms . corexit
9500 and 9527 , two dispersants used in the deepwater horizon oil spill
response , have low to moderate toxicity for most marine organisms but can enhance the toxicity of oil in others , including rotifers , coral larvae , and
copepods . the bioavailability of hydrocarbons
to marine organisms is variable and depends on the chemistry , structure ,
sedimentation , and binding to natural particulates . polycyclic aromatic hydrocarbons ( pahs ) are known to be more toxic
than the more abundant , nonaromatic compounds in petroleum . low molecular weight ( low - mw ) aromatic compounds , like the human
carcinogen benzene , have higher water solubility than other component
petroleum hydrocarbons and are of particular concern . these toxic compounds can volatilize into the atmosphere after a
surface spill or partition into deep water following subsurface release ,
as in the deepwater horizon spill . additionally ,
offshore oil drilling releases low - mw hydrocarbons ( namely benzene ,
toluene , ethylbenzene , xylene ) at levels 8 mg / l in produced
water . nanoparticles and fine microparticles
can assemble at water these particles are a potential alternative to chemical dispersants
such as corexit . over the past decade , the field of nanotechnology
has developed methods to precisely engineer the surface properties
and behaviors of fine particles , and this ability may be exploited
to fabricate a new class of high - efficiency , low - toxicity , particle - based
dispersants . these dispersants may act similarly to naturally occurring
oil - mineral aggregates ( omas ) formed from suspended mineral fines
in seawater agglomerating with spilled oil to form entities that are
stable for weeks in seawater without recoalescing . this is considered a key process in the natural biodegradation
of oil in coastal environments . this article focuses on evaluating the behavior and environmental
implications of novel surface - engineered carbon nanoparticle dispersants
under current development . studies have shown that surface - modified
carbon black ( cb ) particles are effective dispersants for stabilizing
oil - in - seawater emulsions . functionalized
cb particles are less likely than corexit 9500a to interact with crude
oil to form water - in - oil emulsions , which reduce bioremediation efficiency and make oil removal more difficult by increasing
volume and viscosity and reducing the
ability to burn . carbon surfaces are
well - known to adsorb organic compounds from aqueous phases and have
a particularly high affinity for aromatics . we therefore explored
the potential for these engineered particle dispersants , as a secondary
feature , to serve as adsorbents for dissolved aromatic compounds from
the water column and reduce their bioavailability and toxicity to
marine organisms . we evaluate the impact of engineered particle - based
dispersants
on biota in the water column using artemia franciscana ( brine shrimp ) , a filter - feeding microcrustacean previously used
as a laboratory model for marine zooplankton toxicity testing of petroleum
hydrocarbons and dispersants . surfactant - based dispersants like finasol , corexit 9500 , and corexit
9527 can cause changes in atpase activity , respiration , and death
in artemia , with the 48 h lc50 for corexit
9500 at 21 we assess acute toxicity of novel carbon
nanoparticle dispersants using larval mortality and a sublethal stress response characterized by a measurable
increase of heat shock protein 70 ( hsp70 ) levels . a key variable in this study is hydrophilicity or
hydrophobicity
of the carbon particle surfaces . successful pickering dispersants
must have surfaces of intermediate hydrophilicity / hydrophobicity , which some particles possess in the native state ,
but usually requires postsynthesis chemical surface modification . show that cb decorated with hydrophilic benzoic
acid functional groups was an effective emulsion stabilizer . surface functionalization to increase hydrophilicity
can also influence physical adsorption on carbon surfaces , potentially influencing the bioavailability and toxicity of the
particles . the surface engineering
of particle - based dispersants is thus a
complex design problem , which requires a type and degree of functionalization
that co - optimizes dispersant and adsorbent performance but minimizes
potential adverse effects of the particles on biological systems .
here we create three types of surface - engineered cb particles and
study their interactions with artemia franciscana and examine the three - way interactions between the particles , benzene
( used as a model low - mw aromatic ) , and the brine shrimp through coexposure
experiments . norit sx ultra powdered activated carbon
( pac ) ( norit , tx , usa ) was used as a reference sample in the commercially
available form without further modification . commercially available
cb ( regal 330 from cabot corporation ) was used as a baseline material
and then chemically modified to produce more - hydrophilic and less - hydrophilic
versions . this sample , called as - produced cb , was washed with 0.1
m hcl to remove potential metal impurities and then rinsed with deionized
water until reaching neutral ph . particles with increased hydrophobicity
were produced by annealing the acid - washed regal 330 in an environment
of 1% hydrogen in helium for 2 h at 800 c and are referred to
as annealed cb . para - amino benzoic acid ( paba ) was used to introduce
aryl - carboxylate groups using diazonium chemistry , producing particles with increased hydrophilicity that
are designated as functionalized cb . briefly , 250 mm paba in nanopure
water was added to cb precursor particles in a ratio of 80 ml of solution
per 100 mg of cb . , 3.3 ml of 600 mm sodium nitrite per 100 mg
of carbon were added to the mixture , which was then moved to a water
bath at 70 c . the reaction was mixed with a magnetic stir bar
and allowed to proceed for 30 min before being moved to a refrigerator
at 4 c to stop the reaction . the resulting particles were then
cleaned extensively using 3.5k molecular weight cutoff dialysis tubing . base titrations were performed with
a 907 titrando automatic titrator ( metrohm , switzerland ) . a sample
of 0.1 g were dispersed in 50 ml of 0.1 m nacl as background electrolyte . titration was carried out by dynamic addition of 1 n naoh or 1 n hcl
to the flask while the solution was stirred continuously . after each
addition of titrant , the system was allowed to equilibrate until a
stable ph value was obtained . thermogravimetric analyses were preformed
with a thermo cahn versa analyzer ( thermo scientific , ma , usa ) . each
material was heated from 25 to 800 c at a rate of 10 c / min ,
under n2 atmosphere . particle size
distributions for all three versions of cb particles were measured
by dynamic light scattering using a zetasizer nano s90 ( malvern instruments ,
worcesteshire , uk ) . acs reagent , sigma - aldrich ,
mo , usa ) were dissolved in simulated seawater ( instant ocean , va ,
usa ) with a known mass of carbon particles in 50 ml glass vials . the
vials were filled completely with no headspace and sealed tightly
with ptfe - lined caps . the vials were kept on lab rollers for 48 h
to ensure adsorption equilibrium was reached ( kinetics data shown
in figure s2 of the supporting information ) . the absorbance of at least 3 samples from each vial was measured
at 254 nm in a low - uv quartz cuvette ( thermo scientific , ma , usa )
using a jasco v-630 spectrophotometer ( jasco , ok , usa ) , and replicates
were used to calculate standard deviations . prior to spectrophotometric analysis , the cb was removed using a millex 0.2 um ptfe syringe filter
( millipore , ma , usa ) to avoid interference with the absorption spectra
of benzene . the absorbance was converted for a final concentration
of benzene using a previously constructed calibration curve . the adsorbed
amount , q , was determined from the following equationwhere v is the volume of
liquid in the vial , co is the initial
concentration of benzene in the simulated seawater , cf is the concentration of benzene after adsorption equilibrium ,
and mcarbon is the total mass of carbon
in the vial . data were then fit with various models ( freundlich , langmuir ,
linear ) and the best fit was determined by the coefficients of determination .
the mathematical model of best fit was then used to determine three - way
partitioning between air , simulated seawater , and carbon particles . a literature value for the partition coefficient for benzene between
ocean water and air was used after experimental
verification . artemia franciscana cysts from great salt lake ,
utah ( inve brand ) were stored at 20 c . cysts were hatched
under continuous light for 48 h at 28 c in aerated simulated
seawater ( instant ocean , va , usa ) at a ratio of 1 g cysts/1 l seawater .
based on anatomy and schrehardt staging system , exposure media prepared with seawater and particle dispersants or
mixtures of particles and benzene were placed in vials filled completely
with no headspace and sealed tightly with ptfe - lined caps and then
mixed on lab rollers for 24 h. benzene treatments were mixed immediately
before exposure . as - produced cb , functionalized cb , and pac were each
sonicated for at least 20 min in a water bath ( branson 2510 ) to facilitate
dispersion in simulated seawater . annealed cb was dispersed in simulated
seawater using a tight - fitting glass dounce homogenizer . for
mortality experiments , 25 ml glass scintillation vials received 15
ml of exposure medium and a minimum of 250 a. franciscana larvae . larvae were exposed for 24 h at 19 c and then counted
for total mortality . death was determined by motility after brief
( 23 s ) observation under a dissecting microscope . for observation
of particle agglomeration on surfaces , brine shrimp were fixed with
4% formaldehyde and visualized using a nikon eclipse e800 microscope
and brightfield microscopy . , 25 ml
glass scintillation vials
received 15 ml of exposure medium and a minimum of 500 larvae . larvae
were exposed for 24 h at 19 c , then collected into lysis buffer ,
and homogenized to isolate protein for western blot analysis ( protocol
available in the supporting information ) . samples were probed with primary antibodies for hsp70 ( abcam 136874 )
and alpha - tubulin ( abcam 52866 ) for the western blot analysis , and
the final chemiluminescent films were used for band density quantification
with nih imagej software . for brine shrimp mortality and hsp70
protein levels , statistical
significance of each treatment was determined using one - way anova
followed by tukey s multiple comparisons test . data were compared
to untreated for individual particle exposures and compared to benzene
alone at the given concentration for coexposure studies . to compare
trends in sublethal protein levels , the linear regression of the particle
coexposures was compared to benzene exposure alone using the extra
sum of squares f test . all hsp70 protein data were normalized to total
protein using alpha - tubulin protein expression and expressed relative
to the hsp70 protein levels induced by menadione sodium bisulfite
( msb ) in each experiment . msb , a strong inducer of oxidative stress
in brine shrimp , is used here as a positive control in all toxicity
studies . all statistical analysis were
performed by graphpad prism version 6 for mac ( graphpad software ,
la jolla , california usa , www.graphpad.com ) , and data were
considered significant at p < 0.05 . for the low
concentration coexposure experiments , particles were considered to
have a significant impact on hsp70 levels with a p < 0.01 for a given particle concentration . commercial cb was
used as a baseline sorbent and is denoted by the name as - produced
cb nanoparticles . particles with increased hydrophobicity were produced
by heating in 1% h2 in he and are called throughout the
paper annealed cb nanoparticles ( figure 1a ) . this process drives off many of the oxygen - containing hydrophilic
functional groups and caps the nascent carbon active sites with hydrogen
to minimize surface reoxidization in ambient air . particles with increased
hydrophilicity were produced by covalently grafting benzoic acid functional
groups through diazonium chemistry and are referred to as functionalized
cb nanoparticles ( figure 1a ) . the increase
in acidic surface functional groups was confirmed by measuring the
zeta potential ( figure 1b ) . the number and
nature of acidic and basic sites on the particle surface determine
the zeta potential vs ph curve shape . the values of the zeta potentials
shown in figure 1b indicate that the number
of acidic functional groups increase in the order annealed < as - produced
< functionalized , as expected . as - produced ) cb is washed with 0.1 m hcl to remove impurities and
then either annealed to a more hydrophobic version ( left ) or functionalized
using diazonium chemistry to a more hydrophilic form ( right ) . b ) change
in zeta potential as a function of ph , at an ionic strength of approximately
165 mm . c ) determination of the pka value
distribution by acid base titration of the functionalized cb
as compared to the precursor particles . d ) probable side reaction
is the self - polymerization of the diazonium intermediate leading to
greater than monolayer coverage of benzoic acid groups . base titrations ( figure 1c )
show a clear increase in the number of protonable functional groups
on the surface of the cb particle after the functionalization process . there are approximately 150 mmol / g of functional groups added at 4
< pka < 6 , which is characteristic
of a carboxylic acid moiety . using an estimated surface area of benzoic
acid , it is possible to show the total surface area of these molecules
is over 400% that of the original carbon particle surface . this greater - than - monolayer
coverage was initially surprising , but a known side reaction in the
diazonium functionalization process is grafting onto the aryl ring
of a paba molecule , which produces polymeric coatings on the carbon
surfaces ( figure 1d ) . it is likely that many of the carboxylic acid moieties seen in figure 1c are actually part of the polymer network and are
not directly attached to the carbon surface . figure 2 compares the ability of the different
engineered particle formulations to adsorb dissolved benzene as a
model low - mw aromatic hydrocarbon . the raw data are shown in figure 2a , while 2b is the fundamental equilibrium adsorption
isotherm . thermal annealing slightly increases benzene adsorption ,
which is consistent with the removal of native oxygen - containing hydrophilic
groups in the as - produced material . planar conjugated adsorbates ,
such as benzene , often adsorb through hydrophobic and
interactions with the carbon surface . the oxygen - containing functional groups cover portions of the pristine
hydrophobic surface and can also create h - bonded water clusters that
propagate from the groups to partially restrict access to the remaining
adjacent hydrophobic domains . the hydrophilic
formulation showed the highest benzene adsorption ,
which initially seemed inconsistent with the hydrophobic driving forces
cited above . we suggest that this is additional evidence for the side
reaction shown in figure 1d . the presence of
the aromatic polymer film creates an interfacial zone of finite thickness
that contains carboxylate groups for surface charge and hydrophilicity
( figure 1a - b ) but also numerous aryl groups
that provide sites for benzene uptake . this enhanced uptake of benzene
is likely caused by the favorable partitioning of benzene into that
organic polymer layer from the aqueous phase , rather than simple adsorption
onto a carbon surface . finally , pac is much more active as a
benzene adsorbent than any
of the engineered cb samples ( figure 2b ) , consistent
with its much higher surface area . the activated carbon adsorption
data are sufficient to define the full isotherm allowing extraction
of langmuir adsorption parameters , which are k =
2.7 10 l / mg ( 2.1 10 l / mol ) ,
maximum adsorption capacity ( qmax ) = 385
mg / g , and the free energy of adsorption , rt ln k , of 19 kj / mol . the ratio between the
highest observed amount of benzene adsorbed by the as - produced cb
( 32 mg / g at an equilibrium concentration of about 750 mg / l ) ,
and the qmax predicted by the langmuir
adsorption model for the pac ( 385 mg / g ) is roughly equivalent
to the ratio of their surface areas ( 92 m / g vs 1200 m / g , respectively ) . adsorption of benzene as a model low - mw aromatic
by various engineered
carbon materials . a ) raw adsorption data for three formulations of
carbon - black - based dispersants in simulated seawater . b ) adsorption
isotherm comparing the particle - based dispersants to powdered activated
carbon ( pac ) as an engineered sorbent reference material . inset shows
a detailed view of the adsorption isotherm for the particle - based
dispersants only . brine shrimp mortality
and particle accumulation after 24 h. error
bars indicate standard deviation . solid and dashed lines represent
linear regression and 95% confidence bands , respectively . * p < 0.05 , * * p < 0.01 compared to
untreated , # p < 0.05 , # # p <
0.01 compared to 200 mg / l benzene . b ) mortality after coexposure
to 200 mg / l benzene and particle dispersants . d ) calculated values for particle : effective concentration
for adsorption of 50% of 200 mg / l benzene onto the carbon surface ;
reduction of brine shrimp mortality following coexposure to benzene
at 200 mg / l by 50% ; and lc50 for particles alone . e ) the
untreated brine shrimp image to the left of the panel demonstrates
anatomical features at time of exposure . 1 : mouth ( underneath and
not visible ) , 2 : sensory antennule , 3 : locomotory antenna , 4 : nonfunctional
limb buds , 5 : gut lumen . the panel of images at right shows uptake
and accumulation of particles on the exoskeleton of whole brine shrimp
after 24 h at 100 mg / l ( i - iv ) and 500 mg / l ( v - viii ) for all three
cb forms and pac . red arrow indicates rod - like bundles of excreted
particles ; blue arrow indicates gut uptake of particles . brine shrimp larvae were exposed to the surface - engineered
cb nanoparticles and pac for 24 h and observed for mortality and particle
accumulation ( figure 3a ) . as - produced cb induces
significant mortality at concentrations of 100 mg / l and above . annealed
cb shows lower toxicity , with significant mortality only over 500
mg / l . using a linear regression model , the calculated median lethal
concentration ( lc50 ) for as - produced cb is 370
mg / l and for annealed is 1000 mg / l . functionalized cb and
pac are less toxic , with insignificant mortality above 750 mg / l . mg / l and thus outside the range of tested concentrations included
in this study . light microscopic images of brine shrimp exposed
to 100 and 500 mg / l of carbon particles or pac for 24 h reveal gut
uptake of all particle types and particle agglomerates adhering to
external surfaces ( figure 3e ) . agglomerates
of as - produced cb nanoparticles adhered to brine shrimp after exposure
to 5001000 mg / l . high concentrations ( 1000 mg / l ) of annealed
cb or pac formed larger surface agglomerates , while functionalized
cb particles remained well - dispersed with minimal surface adherence
at all concentrations . particles can be seen in the gut after all
particle exposures , indicating uptake by filter - feeding . also visible
in the seawater around the brine shrimp are rod - like bundles of particles ,
which most likely reflect gut uptake and excretion as described by
handy et al . these can be seen after
exposure to 100 mg / l as - produced and annealed cb and at all concentrations
of functionalized cb . after 24 h of benzene exposure , brine shrimp mortality is evident
at an initial concentration of 75 mg / l . mg / l , until reaching 100% mortality at a concentration of
500 mg / l benzene ( figure 3c ) . to test the ability of as - produced or functionalized
cb nanoparticles and pac to mitigate the toxicity of benzene , brine
shrimp were coexposed to 200 mg / l benzene ( the lc50 ) and
a range of particle concentrations . linear regression plots indicate
that coexposure to as - produced cb nanoparticles or pac induced a dose - dependent
decrease in mortality , but only pac caused a statistically significant
mitigation when compared to 200 mg / l benzene ( figure 3b ) . coexposure to 1000 mg / l as - produced cb reduced mortality
to 40% as compared to 10% at the same concentration
of pac . brine shrimp coexposed to benzene and functionalized nanoparticles
showed no attenuation of mortality , even at the highest concentration
( 51% at 1000 mg / l ) . coexposure to benzene reduced toxicity
of as - produced cb particles 50% over the concentration range
of 2501000 mg / l . imaging shows less particle aggregation and
accumulation on the surface of brine shrimp coexposed to benzene and
as - produced cb or pac when compared to particles alone ( figure s3 ) . to determine the impact of particle dispersants
on exposure of brine shrimp to low mortality ( 75 mg / l ) and sublethal
( 2550 mg / l ) concentrations of benzene , hsp70 protein levels
were assessed using western blot analysis ( figure 4 ) . hsp70 is a heat shock protein induced by a wide range of
physical and chemical stressors including heat , metals , and chemical
pollutants . menadione sodium bisulfite
( msb ) was used as a positive reference toxicant to induce high levels
of hsp70 . brine shrimp were exposed to particle concentrations of
25 and 50 mg / l alone ( figure 4a ) or in combination
with benzene ( 2575 mg / l ) ( figure 4c ) . protein levels after 24 h exposure . * p <
0.05 , compared to untreated . a ) western blot analysis of hsp70 and
alpha - tubulin protein levels induced by as - produced , annealed , and
functionalized cb , activated carbon or benzene , alone or in combination
with carbon particles . b ) relative quantitation of hsp70 protein levels
induced by particle dispersants , normalized to hsp70 induced by 20
mg / l msb . c ) relative quantitation of hsp70 levels induced by coexposure
to particle dispersants and benzene , normalized to hsp70 levels induced
by 20 mg / l msb . at 50 mg / l , as - produced
and annealed cb alone significantly increased
hsp70 levels , while exposure to functionalized cb or pac did not induce
this stress protein response ( figure 4b ) . using
linear regression to analyze protein expression , coexposure with as - produced
cb significantly increased hsp70 levels at 2550 mg / l compared
to benzene alone , while annealed cb increased levels at 50 mg / l ( figure 4c ) . coexposure to functionalized cb or to pac did
not significantly affect hsp70 protein levels induced by low concentrations
of benzene . this
study provides a preliminary safety assessment for particle - based
alternative dispersants currently under development for large - scale
petroleum spills . engineered hydrophilic cb particles that show good
dispersant performance demonstrate no
toxicity . at low concentrations , the functionalized hydrophilic samples
did not induce the stress response indicated by increased hsp70 protein
levels , and no significant mortality or particle agglomeration was
observed at high concentrations . 100 mg / l and were associated with
the accumulation of particle agglomerates on the brine shrimp exoskeleton . other studies have reported that hydrophilic surface treatments can
reduce the toxicity of carbon nanomaterials in mammalian cell cultures and aquatic organisms . at sublethal
concentrations , a mechanism proposed for brine
shrimp mortality at high concentrations
of hydrophobic carbon nanoparticles is physical adhesion to the exoskeleton
( figure 3 ) and locomotory appendages that decrease
motility . locomotor impairment has been previously associated with
external nanomaterial cb exposure in adult fruit flies , while dietary
exposure to the same material had no significant effect . for invertebrates , these engineered carbon nanomaterials
act as external physical or mechanical stressors . other studies using daphnia or artemia exposed to different
natural and engineered nanomaterials emphasized the contribution of
particle loading of the gut resulting in impaired food uptake and
increased mortality after prolonged time periods . in this study , both gut uptake and excretion of all tested carbon
particles were observed over 24 h similar to observations following
exposure to well - dispersed single wall carbon nanotubes . only the more hydrophobic carbon nanoparticles
caused mortality and only at high concentrations . it is noteworthy
that the hydrophilic surface treatment required to create effective
dispersants also reduces or eliminates
adverse effects of the nanoparticles on artemia . our study , along with previous work by others , demonstrates that functionalized cb particles are both
functional as dispersants and safe for marine organisms . we
also examined the potential for particle - based dispersants to
adsorb low - mw aromatics from the water column as a secondary useful
feature . using benzene as a model solute g / l . we would
also
like to note that these experiments were conducted in artificial seawater
and did not include any competitive effects from natural organic matter
or other compounds naturally present in seawater . an interesting finding
is that the functionalized , hydrophilic cb showed a higher affinity
for physical adsorption of benzene than the hydrophobic varieties . this is the opposite of the commonly observed trend , and we attribute it to benzene partitioning into an organic layer
formed by self - polymerization of the diazonium intermediate on the
cb surfaces . additionally , this study provides information on
the implications
of dissolved aromatic hydrocarbons in petroleum / water systems . at
high concentrations , inhalation of volatile or low - mw hydrocarbons
can cause irritation of the respiratory tract and central nervous
system depression . the lc50 of benzene for artemia was found to be 200 mg / l , and increased levels of the generalized
stress protein , hsp70 , were induced by sublethal concentrations ( 2550
mg may serve as a generalized
stress biomarker that is responsive to both physical and chemical
stressors , either alone or in combination . also of concern are
potential long - term human health consequences
of exposure to benzene , which can be inhaled by cleanup workers . inhalation
of benzene can induce hematotoxicity , reproductive and developmental
abnormalities , and leukemia . volatile
aromatic hydrocarbons , including benzene and alkylated monoaromatic
compounds , were detected at 68 g / l in subsurface water samples
following the gulf oil spill . a retrospective
study of exposed and unexposed gulf residents detected elevated levels
of phenol , a biomarker for benzene exposure , in the urine of exposed
workers . clinical laboratory findings included decreased platelet
counts , increased serum enzyme markers of liver injury , and reduced
kidney function in the exposed workers consistent with benzene - induced
toxicity . benzene and particle
coexposure experiments show that the cb can not
mitigate benzene toxicity , which is consistent with the small amounts
adsorbed at the particle concentrations used in the biological experiments
( < 1000 mg / l ) . pac , in contrast , can mitigate free benzene toxicity
when present at 250 mg / l , due to its much higher engineered
surface area . activated carbon , however , is not known to be active
as a pickering dispersant and would likely function primarily as a
sorbent in this system . the secondary role of cb - based pickering dispersants
as sorbents was not realized here , and further work is needed to see
if the adsorptive effects may be more pronounced for higher - mw polycyclic
aromatics . these preliminary studies suggest that functionalized
hydrophilic
cb particles under development as petroleum dispersants exhibit low
toxicity to the marine model organism , artemia franciscana . future studies are needed to evaluate a wider range of particulate
and polymeric dispersants under development as alternatives to existing
chemical dispersants . the low toxicity of these functionalized nanoparticles
demonstrates that co - optimization for dispersant performance and low
adverse environmental impacts can be integrated together for the surface
engineering of future dispersants . the brine shrimp model presented
here is suitable for evaluation of both lethal and sublethal acute
toxicity end points of novel , particle - based dispersants for marine
oil spills , individually or as coexposures with aromatic hydrocarbons . this model system can be used to assess potential inhibitory , additive ,
or synergistic effects of combined exposure to physical stressors
( particle - based dispersants ) plus chemical stressors ( petrogenic aromatic
hydrocarbons ) , which is a challenging problem in human risk assessment . continued work is required to identify and validate
more sensitive molecular biomarkers to assess chronic toxicity and
multigenerational impacts of particle - based dispersants with and without
adsorbed hydrocarbons at lower concentrations that are more relevant
for environmental exposures following marine oil spills | fine particles are under active consideration
as alternatives to
chemical dispersants for large - scale petroleum spills .
fine carbon
particles with engineered surface chemistry have been shown to stabilize
oil - in - water emulsions , but the environmental impacts of large - scale
particle introduction to the marine environment are unknown . here
we study the impact of surface - engineered carbon - black materials on
brine shrimp ( artemia franciscana ) as a model marine
microcrustacean .
mortality was characterized at 501000 mg / l ,
and levels of heat shock protein 70 ( hsp70 ) were characterized at
sublethal particle concentrations ( 2550
mg / l ) . functionalized
carbon black ( cb )
nanoparticles were found to be nontoxic at all concentrations ,
while hydrophobic ( annealed ) and as - produced cb induced adverse effects
at high concentrations .
cb was also shown to adsorb benzene , a model
hydrocarbon representing the more soluble and toxic low - molecular
weight aromatic fraction of petroleum , but the extent of adsorption
was insufficient to mitigate benzene toxicity to artemia in coexposure experiments . at lower benzene concentrations ( 2575
mg
/ l ) , coexposure with annealed and as - produced cb increased hsp70
protein levels .
this study suggests that surface functionalization
for increased hydrophilicity can not only improve the performance
of cb - based dispersants but also reduce their adverse environmental
impacts on marine organisms . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
dugorono i intenzivno fiziko naprezanje izaziva metabolike i biohemijske promene radi sportskih ali i nesportskih ciljeva . s obzirom na vanost aerobnog treninga za fudbalere , ova studija imala je za cilj odreivanje promena u aktivnostima : kreatinin kinaze ( ck ) , kreatin kinaze mb ( ckmb ) , laktat dehidrogenaze ( ldh ) , -hidroksibutirat dehidrogenaze ( hbdh ) , holinesteraze ( che ) i alkalne fosfataze ( alp ) u okviru odgovora na tranje napolju , na poludugoj stazi , u aerobnim uslovima , kod igraa fudbala mukog esnaest uesnika starosti 21,92 godine ( ene ) i 18,40,5 godina ( mukarci ) , dobrovoljno regrutovanih fudbalera , uestvovalo je u trci napolju , u kojoj su ene prelazile 7,40,3 km a mukarci razdaljinu od 10,71,0 km . nai rezultati pokazuju da bi ukupna aktivnost ldh mogla biti korisno sredstvo za procenjivanje fizike spremnosti kod sportista . istovremeno smo ustanovili da che ne moe biti koristan marker za odreivanje metabolikog odgovora na fiziko naprezanje kod sportista . pored toga , nai rezultati pokazuju da se promene u aktivnosti alp posle naprezanja mogu koristiti za otkrivanje ranih simptoma deficijencije nekih vitamina u ishrani sportista . potvrdili smo da odreivanje aktivnosti izabranih tradicionalnih dijagnostikih enzimskih markera donosi informacije o stanju miia posle fizikog naprezanja . long - term and intensive physical effort causes metabolic and biochemical adaptations for both athletic and non - athletic objectives . physical activity is one of the most important measures to enable an individual to control body weight , delay the occurrence of chronic disorders and prevent various diseases . hence , regular physical activity induces metabolic changes , yet can alter the serum concentrations of numerous laboratory parameters . these modifications , especially in terms of increases , can often lead to results outside the reference value range , leading to additional examinations for the athlete and/or discontinuation of training and competition ( 1 , 2 ) . various literature data emphasize that long - term training influences athletes cellular metabolism and can lead to muscle damage , and induce oxidative stress ( 1 , 3 , 4 ) that causes metabolic changes on a biochemical level in diagnostic parameters analysed in athletes blood ( 47 ) . there are some typical metabolic parameters described as sports markers which are used by trainers and athletes to follow and characterise their strength and efficiency e.g. creatine kinase activity , blood lactate level ( 8) . various intracellular enzymes are described in clinical enzymology as tissue or organ markers used as diagnostic markers in identification and differentiation of diseases , e.g. plasma aminotransferases activity ( aspartate aminotransferase , ast ; alanine aminotransferase , alt ) , gamma - glutamyl transferase ( ggt ) activity belonging to routinely analysed liver diagnostic markers or both mass and activity of creatine kinase isoenzyme mb ( ckmb ) combined with lactate dehydrogenase activity ( ldh ) as well as other non - enzymatic markers belonging to routinely analysed heart diagnostic markers ( 1 , 9 ) . however , most abnormalities in elite athletes biochemical parameters found during routine training monitoring have no clinical significance ( 2 ) . therefore , there is still a need to establish the influence of different types of effort ( in aerobic , anaerobic and aerobic - anaerobic conditions ) on changes in metabolic markers not only belonging to metabolites , but also those belonging to standardized diagnostic enzymatic tissue or organ markers . more importantly , there is a need to find pleiotropic markers of metabolic response to physical effort . consequently , it seems that changes in blood plasma activity of enzymes characterising different tissues and organs may provide a broader view of the influence of different types of physical effort among high level fitness athletes . it is well known that aerobic training in football plays an important role and is designed to improve cardiovascular health . it is imperative during soccer matches as well as training sessions that there is a good supply of oxygen to the active muscles , and that these tissues have the capability to use the oxygen provided by the circulatory system ( 10 ) . considering the importance of aerobic training in football players , the aim of this study was to evaluate the post - exertion changes in activity of selected intracellular enzymes belonging to traditional clinical markers of : muscles , liver , heart and bones . to better understand these phenomena , we determined the changes in the activity of : creatinine kinase ( ck , ec 2.7.3.2 ) , creatine kinase mb ( ckmb , ec 2.7.3.2 ) , lactate dehydrogenase ( ldh , ec 1.1.1.27 ) , -hydroxybutyrate dehydrogenase ( hbdh , ec 1.1.99.6 ) , cholinesterase ( che , ec 3.1.1.8 ) and alkaline phosphatase ( alp , ec 3.1.3.1 ) in response to a semi - long distance outdoor run under aerobic conditions in both female and male football players . the physical effort in football belongs to a mixed type ( both aerobic and anaerobic ) which is associated with the different activities of a player during the match . the high number of accelerations and decelerations associated with highly dynamic play create an additional burden on the muscles ( 1113 ) . due to the different types of exercise , the players must be adapted to generate energy using both anaerobic and aerobic metabolic pathways . various data in the literature identify the differences in biochemical parameters as a result of physical effort and the relation of those changes with the subject s fitness level . moreover , abnormalities in values of biochemical parameters very often have no clinical significance in elite athletes ( 2 ) . hence , it is important to establish the influence of different types of effort , especially in aerobic conditions , on changes in metabolic markers and of equal importance is the need to find pleiotropic markers of metabolic response to physical effort in athletes . therefore , our study aimed to assess post - effort changes in intracellular enzyme activity in football players blood . we determined ck , ckmb , ldh , hbdh , che and alp activity in response to a semi - long distance outdoor run under aerobic conditions among both female and male football players to explore these biochemical processes . participants ( n=16 ) were recruited among football players , belonging to the olimpia szczecin and pogon szczecin s.a . they had no history of any metabolic or cardiovascular diseases , were non - smokers and were cautioned against taking any medications or supplements known to affect metabolism . the baseline characteristics of both studied groups are presented in table i. the study was conducted in accordance with the ethical standards as described by kruk ( 14 ) . participants ( and their parents , when necessary ) were informed of experimental procedures and possible risks of the experiment before giving their written consent to participate . local ethics committee approval in accordance with the helsinki declaration was received before the beginning of the testing . the exercise test was performed on the last day of their training season in the afternoon of a warm , cloudless summer day . the exercise test consisted of a warm - up routine ( 10 minutes ) , the main run ( 60 minutes ) and stretching and breathing exercises ( 15 minutes ) . the aim of the exercise was to develop aerobic efficiency below the anaerobic threshold , calculated individually for each participant . to this end , participants ran to maintain a subliminal heart rate of 1584 beats / min . the female group of participants ran outdoors covering a distance of 7.40.3 km with a mean speed of 7.50.5 km / h . the male group of participants ran a semi - long distance of 10.71.0 km with a mean speed of 10.61.7 km / h . heart rate was analysed using a garmin forerunner 305 heart rate monitor ( garmin ( europe ) ltd . , additionally , in both studied groups the level of lactate was determined to ensure aerobic metabolism during the exercise test . blood samples were taken using a 4.9 ml s - monovette tube with ethyl - enediaminetetraacetic acid ( edta ) and separating gel . blood samples were centrifuged 500 g for 15 minutes at room temperature in order to receive blood plasma . the plasma was obtained and frozen at 80 c until later performed analyses ( 15 ) . plasma activities of ck , ckmb , ldh , hbdh , che and alp as well as lactate concentration were determined in samples obtained before exercise ( pre - exercise ) , immediately after the run ( post - exercise ) and 15 minutes after completion of the exercise test , at the beginning of recovery time ( recovery ) . plasma lactate level was determined using a diagnostic colorimetric enzymatic method ( liquick cor - lactate ) according to the manufacturer s protocol ( pz cormay s.a . , plasma activities of alp , ldh and ck ( u / l ) were determined using an appropriate kinetic assay kit according to the manufacturer s protocol ( biomaxima s.a . , lublin , poland ) . the kinetic assay kits were also used to determine the activity of hbdh and ckmb ( u / l ) ( pz cormay s.a . , omianki , poland ) and che ( u / l ) ( quimica clinica aplicada s.a . , all of the reactions were initiated by the addition of plasma to assay kit reaction mixtures and conducted at 37 c . a measurements were conducted at =340 nm in the case of ck , ckmb , ldh and hbdh and at =405 nm in the case of alp and che . all analysis procedures were validated with the use of multiparametric control serum ( biolabo s.a.s , maizy , france ) . absorption measurements were made on a semco s91e spectrophotometer ( emco , warszawa , poland ) . the results of the enzyme activity analyses were referred to reference values provided by appropriate kit manufacturers . all data are presented as mean standard deviation in cases of normal data distribution or median ( interquartile range ) in cases of non - normal data distribution . statistical analyses were performed using statistica ( data analysis software system ) , version 10 software ( statsoft , inc . significance level of differences observed between analysed time points ( pre - exercise vs. post - exercise vs. recovery ) for each participant was calculated according to the results of data distribution analysis . in cases of normal data distribution , we used analysis of variance ( anova ) with repeated measures test followed by contrast analyses . in cases of non - normal data distribution , we used friedman s two - way analysis of variance followed by appropriate post - hoc analyses . the physical effort in football belongs to a mixed type ( both aerobic and anaerobic ) which is associated with the different activities of a player during the match . the high number of accelerations and decelerations associated with highly dynamic play create an additional burden on the muscles ( 1113 ) . due to the different types of exercise , the players must be adapted to generate energy using both anaerobic and aerobic metabolic pathways . various data in the literature identify the differences in biochemical parameters as a result of physical effort and the relation of those changes with the subject s fitness level . moreover , abnormalities in values of biochemical parameters very often have no clinical significance in elite athletes ( 2 ) . hence , it is important to establish the influence of different types of effort , especially in aerobic conditions , on changes in metabolic markers and of equal importance is the need to find pleiotropic markers of metabolic response to physical effort in athletes . therefore , our study aimed to assess post - effort changes in intracellular enzyme activity in football players blood . we determined ck , ckmb , ldh , hbdh , che and alp activity in response to a semi - long distance outdoor run under aerobic conditions among both female and male football players to explore these biochemical processes . participants ( n=16 ) were recruited among football players , belonging to the olimpia szczecin and pogon szczecin s.a . football clubs , and divided into two groups according to sex . they had no history of any metabolic or cardiovascular diseases , were non - smokers and were cautioned against taking any medications or supplements known to affect metabolism . the baseline characteristics of both studied groups are presented in table i. the study was conducted in accordance with the ethical standards as described by kruk ( 14 ) . participants ( and their parents , when necessary ) were informed of experimental procedures and possible risks of the experiment before giving their written consent to participate . local ethics committee approval in accordance with the helsinki declaration was received before the beginning of the testing . the exercise test was performed on the last day of their training season in the afternoon of a warm , cloudless summer day . the exercise test consisted of a warm - up routine ( 10 minutes ) , the main run ( 60 minutes ) and stretching and breathing exercises ( 15 minutes ) . the aim of the exercise was to develop aerobic efficiency below the anaerobic threshold , calculated individually for each participant . to this end , participants ran to maintain a subliminal heart rate of 1584 beats / min . the female group of participants ran outdoors covering a distance of 7.40.3 km with a mean speed of 7.50.5 km / h . the male group of participants ran a semi - long distance of 10.71.0 km with a mean speed of 10.61.7 km / h . heart rate was analysed using a garmin forerunner 305 heart rate monitor ( garmin ( europe ) ltd . , romsey , uk ) . additionally , in both studied groups the level of lactate was determined to ensure aerobic metabolism during the exercise test . blood samples were taken using a 4.9 ml s - monovette tube with ethyl - enediaminetetraacetic acid ( edta ) and separating gel . blood samples were centrifuged 500 g for 15 minutes at room temperature in order to receive blood plasma . the plasma was obtained and frozen at 80 c until later performed analyses ( 15 ) . plasma activities of ck , ckmb , ldh , hbdh , che and alp as well as lactate concentration were determined in samples obtained before exercise ( pre - exercise ) , immediately after the run ( post - exercise ) and 15 minutes after completion of the exercise test , at the beginning of recovery time ( recovery ) . plasma lactate level was determined using a diagnostic colorimetric enzymatic method ( liquick cor - lactate ) according to the manufacturer s protocol ( pz cormay s.a . , plasma activities of alp , ldh and ck ( u / l ) were determined using an appropriate kinetic assay kit according to the manufacturer s protocol ( biomaxima s.a . , lublin , poland ) . the kinetic assay kits were also used to determine the activity of hbdh and ckmb ( u / l ) ( pz cormay s.a . , omianki , poland ) and che ( u / l ) ( quimica clinica aplicada s.a . , all of the reactions were initiated by the addition of plasma to assay kit reaction mixtures and conducted at 37 c . a measurements were conducted at =340 nm in the case of ck , ckmb , ldh and hbdh and at =405 nm in the case of alp and che . all analysis procedures were validated with the use of multiparametric control serum ( biolabo s.a.s , maizy , france ) . absorption measurements were made on a semco s91e spectrophotometer ( emco , warszawa , poland ) . the results of the enzyme activity analyses were referred to reference values provided by appropriate kit manufacturers . all data are presented as mean standard deviation in cases of normal data distribution or median ( interquartile range ) in cases of non - normal data distribution . statistical analyses were performed using statistica ( data analysis software system ) , version 10 software ( statsoft , inc . significance level of differences observed between analysed time points ( pre - exercise vs. post - exercise vs. recovery ) for each participant was calculated according to the results of data distribution analysis . in cases of normal data distribution , we used analysis of variance ( anova ) with repeated measures test followed by contrast analyses . in cases of non - normal data distribution , we used friedman s two - way analysis of variance followed by appropriate post - hoc analyses . each time , p0.05 was considered as a significant difference . all the participants were recruited from football clubs based on a similar period of training experience and divided into two groups according to sex . the baseline characteristics of both studied groups are presented in table i. it was found that the baseline values of ck activity in both studied groups were about 1.5-fold ( women ) and 1.2-fold ( men ) higher than the upper reference limits ( < 167 and < 190 , 60 minutes of outdoor running did not influence total ck activity values in the studied athletes blood ( figure 1 ) . on the other hand , statistically significant changes in ckmb activity and increases in these values in both studied groups immediately after physical effort ( figure 2 ) were observed , but it was only in women s blood that the increase was also found at the beginning of recovery time . the baseline values of ckmb activity in the studied athletes blood were almost equal to the upper limit of the reference range ( 24 u / l ) , yet in women s blood these values were slightly higher ( 26.24.4 u / l ) . our experiment indicated a statistically significant increase in ldh activity after aerobic effort ( figure 3 ) , which did not persist during recovery time . it is worth noting that activities of ldh and its isoenzyme hbdh before the exercise test in the female athletes were about 1.4 and 3.6-fold higher than the upper reference limit ( 240480 u / l and < 182 u / l for ldh and hbdh , respectively ) but in the male group , 2.6-fold higher values were found only in the case of hbdh . in the male group , as opposed to the female group , a statistically significant increase in hbdh after physical effort was found and it persisted at the beginning of recovery time ( figure 4 ) . interestingly , only among the female participants were statistically significant changes in che activity observed after the run and at the beginning of recovery time ( figure 5 ) . additionally , in the female group the baseline values of che activity were mostly within the range of reference values ( 433011500 u / l ) , yet in male athletes blood , the baseline values of che were slightly lower than the lower limit of the reference range ( 540013200 u / l ) . moreover , we found that the activity of alp in the examined participants plasma was statistically significantly higher after aerobic effort as compared to the baseline values ( figure 6 ) and this increase was also observed at the beginning of recovery time . the alp activity before the exercise test was about 1.4-fold higher than the upper reference limit ( < 105 and < 115 many functional and metabolic changes in the system are yielded with prolonged physical effort and training , which also leads to an increased physical efficiency and tolerance to exertion ( 1 , 47 ) . in football and other team sports , energy turnover of the working muscles requires mobilization of aerobic , anaerobic and aerobic - anaerobic metabolic pathways . furthermore , aerobic training in football plays an important role and is designed mostly to improve the oxygen transport system . it is imperative during soccer matches and training sessions that there is an adequate supply of oxygen to the active muscles , and that these tissues have the capability to use the oxygen that is provided by the circulatory system ( 10 ) . some literature data has evidenced that during a competitive match , e.g. a soccer referee may cover a distance of 913 km ( 16 ) , which is similar to the distance covered by the participants of our exercise experiment . investigating the impact of physical effort on the activity of intracellular enzyme profiles specific to certain tissues and organs provides additional information not only about the state of muscle , but also its biochemical adaptation to the training process of athletes . ck activity is an example of a biochemical marker frequently analysed by trainers and sport researchers . in athletes the investigation of ck activity provides information about muscle state after physical effort ( 17 , 18 ) . additional information is provided by measurement of the activity of ck isoenzymes : ckmm and ckmb . it was found in our study that a semi - long distance outdoor run among football players did not influence ck activity in either sex . on the other hand , it was found that the aerobic effort yielded a statistically significant increase in ckmb activity in the blood of both studied athlete groups . suggested that exercise may also influence markers of both skeletal and heart muscle damage including ckmb isoenzyme activity , especially when the physical effort is prolonged or strenuous ( 19 ) . in light of this , it seems that changes in ckmb activity may occur even earlier than those in ck activity , a marker of post - effort metabolic change in football players . additionally , increases in ckmb isoenzyme activity were found in the blood plasma of ultramarathon runners ( 20 ) . there is also an opinion that abnormal values of ckmb activity in serum or plasma should be treated with caution , because it may be connected with late cardiomyopathy ( 19 ) . it must be pointed out that the participants of our study had no history of cardiomyopathy or other heart disorders that rather exclude this phenomenon . the results of our study indicated that an increase in ckmb activity is caused by aerobic effort and , interestingly , this parameter decreases after physical effort to baseline values within 15 minutes . the higher than reference baseline values in athletes found in our study were also described by lucia et al . ldh activity is often considered as a marker of muscle cell damage because its blood activity is combined with disruption that occurs in sarcoma ( 20 , 21 , 23 ) . according to literature data , the activity of total ldh and its isoenzymes is different in muscle cells and it is combined with the endurance and strength of athletes ( 19 , 21 , 24 ) . interestingly , in muscle biopsies a lower total ldh activity was found at a prevalence of hbdh ( ldh1 ) and lhd2 isoenzyme activity ( 2426 ) . ldh and ck activities measured with the use of needle biopsy were also different before and after the training . it was observed that they changed due to different protocols , intensity and level of training ( 19 , 24 ) . these reports indicated that changes in athletes blood ldh and hbdh activities are related to the metabolic response to physical effort . in our results , post - effort changes in total ldh activity were found only in women s blood but this increase was observed only immediately after the exercise test . on the other hand , hbdh activity increased only in men s blood and it was observed not only after the run but also at the beginning of recovery time . the increase in total ldh activity after the physical effort in male athletes blood it is worth pointing out that post - effort changes in ldh activity in professional male cyclists were related to the length of distance covered and it was found that this activity increased only in cases of longer distances ( 27 ) . this may be an explanation of the gender discrepancies in total activity of ldh found in our experiment . we suggested that total ldh activity could be a useful tool to evaluate physical fitness in athletes . it must be emphasised that abnormally higher values of total ldh activity is a phenomenon observed very commonly in athletes blood ( 2 , 21 , 22 , 27 ) . che , also known as pseudocholinesterase , is a diagnostic marker of liver injuries ( 28 , 29 ) . to the best of our knowledge , there is a lack of information about che changes in athletes blood after physical effort . our study indicated that the baseline values of this enzyme in the case of studied participants were nearly within the range of reference values ( 433011500 and 540013200 u / interestingly , post - effort values were higher than baseline only in the female football players and these changes were not prolonged , since the activities of che at the beginning of recovery time were lower than the post - effort values . our results indicate that che is probably not a useful marker in the assessment of metabolic response to physical effort in athletes . the activity of blood alp is a well - known diagnostic marker of mineralization and pathological disorders in bones ( 2 , 30 , 31 ) . we found in our study that 60 minutes of outdoor running caused a slight increase in alp activity immediately after the exercise test in both groups of football players . ( 21 ) who did not observe post - effort changes in alp activity in studied athletes blood . on the other hand , our study was performed on the last day of a whole year of training as well as a competitive season ( before the summer holiday break ) and this could have influenced the results of alp activities . in this case , it may reflect the symptoms of bodily fatigue among the participants . diaz et al . ( 21 ) described a correlation between alp activity and daily vitamin b6 and niacin intake . the results of our study may indicate initial symptoms of vitamin b6 and/or niacin deficiency . it is possible but rather speculative , since the participants themselves reported to have a balanced diet . it would also explain a slightly higher baseline of alp activity in both women and men . therefore , our results suggest that post - effort changes in alp activity might be a useful tool to estimate early symptoms of some vitamin deficiencies in athletes diets , and help to prevent dangerous metabolic disorders associated with future hypovitaminosis . the establishment of the post - effort influence of different types of enzymatic tissues or organs metabolic markers would help in the understanding of a pleiotropic response to physical effort in athletes . to summarize , we confirmed that the assessment of the activity of selected traditional diagnostic enzymatic markers provides information about muscle state after physical effort . furthermore , our results indicate that total ldh activity could be a useful tool to evaluate physical fitness in athletes . we have also shown that che should not really be used as a marker in the assessment of metabolic response to physical effort in athletes . moreover , our results suggest that post - effort changes in alp activity might be useful in estimating early symptoms of some vitamin deficiencies in athletes diets . | summarybackgroundlong - term and intensive physical effort causes metabolic and biochemical adaptations for both athletic and non - athletic objectives . knowing the importance of aerobic training in football players , the aim of this study was to evaluate changes in the activity of : creatinine kinase ( ck ) , creatine kinase mb ( ckmb ) , lactate dehydrogenase ( ldh ) , -hydroxybutyrate dehydrogenase ( hbdh ) , cholinesterase ( che ) and alkaline phosphatase ( alp ) in response to a semi - long distance outdoor run under aerobic conditions among both female and male football players.methodssixteen participants aged 21.92 years ( women ) and 18.40.5 years ( men ) , all of them voluntarily recruited football players , took part in an outdoor run , the women covering a distance of 7.40.3 km while men covered a distance of 10.71.0 km .
plasma activities of the studied enzymes were determined using an appropriate diagnostic assay kit.resultsour results indicate that total ldh activity could be a useful tool in evaluating physical fitness among athletes .
we simultaneously established that che could not be a marker useful in assessing metabolic response to physical effort in athletes .
moreover , our results suggest that post - effort changes in alp activity might be used to estimate early symptoms of certain vitamin deficiencies in an athlete s diet.conclusionswe confirmed that the assessment of activity of selected traditional diagnostic enzymatic markers provides information about muscle state after physical effort . |
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formerly used as an ultimate ratio in end - stage pulmonary diseases , lung transplantation has become a commonly accepted therapy with an increasing number of procedures performed annually . despite the considerable advantages of perioperative medical care , two main problems remain unresolved : rejection of the graft and primary graft failure ( pgf ) of the allograft defined by a noncardiogenic pulmonary oedema appearing shortly after reperfusion of the transplanted organ . large clinical trials report an incidence of between 22% and 57% in patients receiving a lung transplant [ 1 , 2 ] . a central factor involved in the development of pgf seems to be the expression of adhesion molecules on the surface of pulmonary endothelial cells . they represent a group of different glycoproteins and carbohydrates expressed on the surface of a wide variety of cell types , including endothelial cells . by interfering with receptors on the leukocytes , firm adhesion and transendothelial migration follow , resulting in a sequestration of leukocytes at the endothelium and the later infiltration of the interstitial space . the release of proteolytic enzymes and oxygen - free radicals contributes to damage of the alveolar membrane , resulting in noncardiogenic pulmonary oedema . furthermore , some adhesion molecules seem to be involved in allograft rejection and the development of obliterative bronchiolitis . different investigators have demonstrated a positive correlation between the intensity of adhesion molecule expression on the endothelium of the allograft vasculature and clinical and histological signs of organ rejection [ 3 , 4 ] . therefore , suppression of adhesion molecule expression on the vascular endothelium ( thereby interrupting the leukocyte - endothelial interaction ) seems to be a promising tool to improve the quality of life of allograft recipients . e - selectin from the selectins group modulates the initial establishment of contact and causes rolling of the leukocytes along the vascular endothelium . intercellular adhesion molecule 1 and vascular adhesion molecule belong to the immunoglobulins group and mediate the fixed connection between leukocytes and endothelium . other groups of investigators have already demonstrated that adhesion molecules play a key role when it comes to recruitment of leukocytes in lungs , thus modulating the subsequent inflammatory reaction . using small interfering rna ( sirna ) to develop new therapies seems to show promise . originally identified as an intermediate of the rna interference pathway in the late 1980s , sirna has become a serious alternative in medical research and , therefore , therapeutic application . for practical use , analogous to the gene sequence , double - stranded sirna is manufactured by commercial providers and brought in the cytoplasm by transfection . one strand of sirna becomes part of a ribonucleoprotein complex called rna - induced silencing complex ( risc ) . risc can cleave rna sequences complementary to the sirna strand , thereby causing the rapid degradation of the target mrna . importantly , and in contrast to longer double - stranded rnas , sirnas in most cases do not induce an interferon response leading to a general cytotoxicity . the vision of our working group consists of using sirnas for effecting temporary suppression of endothelial adhesion molecules in pulmonary allografts in order to reduce the early and longer - lasting losses in functionality described above as well as the increase in patient morbidity and mortality resulting therefrom . the aim of the present study was to evaluate the potency of specific sirna to block the expression of intercellular adhesion molecule 1 ( icam1 , cd 54 ) , the vascular cell adhesion molecule 1 ( vcam1 , cd 106 ) , and e - selectin ( cd 62e ) in cultured human pulmonary microvasculature cells . stimulation of hlmecs with tnf resulted in a significant increase in receptor expression of all three adhesion proteins measured by facs . maximal expression was found 4 h and 12 h post - tnf- stimulation for esele and vcam-1 , respectively . icam-1 expression stayed high between 8 and 48 h ( see figure 1 ) . concerning all three types of adhesion molecules , the time span of twelve hours after stimulation ensured a high expression level of all adhesion molecules investigated . facs analysis showed a significant increase of the expression of all three adhesion proteins after hlmecs stimulation with tnf- ( figure 2 ) . for esele expression ( figure 2(a ) ) , the baseline value ( no tnf- and no sirna application ) was 7.02 4.36% compared to the tnf- group ( 100% ) . treatment of hlmecs with esele specific sirna resulted in a significant diminished receptor expression of 68.09 3.89% ( p < .001 ) , whereas the siscr control group remained at 97.68 0.89% . application of icam-1-specific sirna led to a significant reduction of the icam-1 expression to only 43.80 8.98% ( p < .001 ) , and treatment of hlmecs with vcam-1 sirna ( figure 2(c ) ) was 35.65 6.11% compared to tnf- stimulated cells ( p < .001 ) ( see figure 2 ) . hlmecs were transfected with the 3 sirnas in a similar manner as that described in the experiments shown above . however , this was not performed individually but by combining all 3 sirnas into a cocktail ( figure 3 ) . in addition to the determinations of the protein level of the adhesion receptor expressions , the effects of sirna transfection on esele , icam-1 and vcam-1 mrna transcript levels were analysed by qrt - pcr ( figures 4(a)4(c ) ) . normalized gene expression was calculated by the ct method ; gapdh was used as a reference , and group 1 ( nontreated and nonstimulated ) was set to the value of 1.00 . the effects of siesele , siicam-1 , and sivcam-1 on target protein expression were strongly reflected by changes in the corresponding transcript levels . all three sirnas ( siesele , siicam-1 and sivcam-1 ) significantly reduced the levels of their corresponding target mrna compared to the groups treated with tnf- only ( p < .05 ) . in contrast , the control groups treated with siscr showed no reduction in respective mrna levels ( figure 4 ) . additionally , hlmecs were transfected with the 3 different sirnas in a similar manner as that described in the experiments shown above . however , this was not performed individually , but by combining all 3 sirnas into a cocktail ( figure 5 ) . the application of this sirna cocktail targeting all three different adhesion receptors resulted in a high , significant reduction of all adhesion receptor mrnas ( figure 5 ) . two million pmn - neutrophils were added to each well containing confluent layers of hlmecs , and after 1 hour , the adherent neutrophils were quantified using a casy cell counter system ( figure 6 ) . on the nonactivated cell layer , the stimulation of the hlmecs with tnf- led to a more than 4-fold increase ( up to 7,0009,000 ) in the number of adherent granulocytes ( figures 6(a)6(d ) ) . groups transfected with specific sirna showed significantly reduced adhesion of neutrophils ( p < .05 ) . transfection with icam-1 sirna ( figure 6(b ) ) resulted in a diminished adhesion of 4,824 57 , while transfection with vcam-1 sirna ( figure 6(c ) ) resulted in an adhesion of 6,375 492 . finally , the cocktail transfection ( 3 different sirnas ) resulted in 4,325 113 adherent neutrophils ( figure 6(d ) ) . lung transplantation carries a higher perioperative morbidity than other transplant procedures including more frequent occurrence of primary graft failure . among a variety of other findings , histological examinations have shown increased infiltration of leukocytes into the parenchyma , causing damage to the graft and thus reducing graft function . in particular , the vascular endothelium seems to be the graft component that is most vulnerable to the stimulus exerted by ischemia . endothelial activation leads to expression of adhesion molecules , which , in turn , exert a considerable influence on the interaction between leukocytes and endothelium [ 1013 ] . have demonstrated a correlation between intercellular adhesion molecule 1 ( icam 1 ) and recruitment of leukocytes within pulmonary parenchyma . have shown that blocking of adhesion molecules exerts a positive influence on reperfusion - associated hyperpermeability of transplanted lungs . these findings indicate that adhesion molecules play an essential role in the modulation of acute as well as chronic graft rejection reactions [ 15 , 16 ] , thus exerting a significant influence on the further development of perioperative morbidity and mortality of transplanted patients . therefore , suppressing expression of adhesion molecules in pulmonary allografts in order to maintain a high level of graft function seems to be a very promising therapeutic approach . to gain a systematic insight into the expression patterns of adhesion molecules on pulmonary microvascular endothelial cells , tumour necrosis factor alpha , a well - known and highly potent cytokine , we have been able to show that various adhesion molecules from the selectin and immunoglobulin families can be stimulated to a significant degree but within different time windows ( see figure 1 ) , which partly differs from the findings published by other investigators . in analogy to the fact that different adhesion molecules exert different functions , we have been able to demonstrate , by means of our experiments , that expression of different adhesion molecules on human lung microvascular endothelial cells reaches a maximum after different latency periods following application of the stimulus . the fact that according to several authors , up to 40% of the pulmonary parenchymal mass might be composed of endothelial cells underlines the particular importance of these findings when it comes to protecting and preserving pulmonary allografts . furthermore , we were able to show that human lung microvascular endothelial cells can in fact be effectively transfected using a cationic liposomal transfection medium . we decided not to use a viral vector so as to work as closely as possible to actual clinical applicability while at the same time preventing the restriction of our method to a safe laboratory environment . in the course of practical application , however , it will not be sufficient to choose a single target or a single adhesion molecule . instead , it will be necessary to suppress a whole series of adhesion molecules in order to achieve a sustainable effect , and therefore , we have also performed what we call cocktail transfection . this means that we have performed transfection of cells with a protective medium containing a combination of the three specific sirna sequences against esele , icam , and vcam and subsequently examined the effect achieved with respect to the individual adhesion molecules . this resulted in a highly significant reduction of expression of the individual adhesion molecules , as shown by facs analysis , qrt - pcr , as well as by functional assays quantifying neutrophil adhesion . it remains to be shown , of course , whether the results achieved in a laboratory setting under static conditions will be confirmed under dynamic conditions in animal experiments . the present study has demonstrated that transfection with specific sirna in each case achieved a significant degree of suppression of the adhesion molecules examined . to the best of our knowledge , so far , this has not been described in the literature on lung microvascular cells . the silencing is sustained for a period of approximately 2 weeks only ( and internal datas ) , after this period , a normal vascular endothelium showing regular physiological reactions is once again available . on the basis of both the results described above and the knowledge of the consequences of pathological interaction between leukocytes and endothelium in transplanted organs , we believe that protecting pulmonary allografts by means of sirna transfection may represent a promising therapy for improving both short and long - term function of pulmonary grafts . human lung microvascular endothelial cells ( hlmecs ) and the mvecgm medium ( microvascular endothelial cell growth medium ) were purchased from sciencell ( carlsbad , calif , usa ) . these primary hlmecs are isolated from human lung tissue and are characterized by immunofluorescent methods with antibodies to vwf / factor viii and cd31 ( p - cam ) and by uptake of dii - ac - ldl . one day before sirna transfection , 150,000 hlmecs per well were cultured in gelatine - coated 12-well plates without antibiotics . after reaching 80% confluence , the cells were transfected with 100 nm specific sirna targeting e - selectin ( esele ) intercellular adhesion molecule-1 ( icam-1 ) or vascular adhesion molecule-1 ( vcam-1 ) or a nonspecific sirna ( negative control = scrambled sirna , siscr ) for 2 hours in a serum - free medium , followed by the replacement of the supernatants with ecm without antibiotics . transfections of sirna were carried out with cellfectin , 1 ng / ml ( invitrogen , karlsruhe , germany ) . twelve hours later , hlmecs were stimulated with tumour necrosis factor- ( tnf- ) ( immunotools , friesoythe , germany ) . keeping in mind the time dependent expression curves of the three adhesion molecules the time span of 12 hours after tnf stimulation ensured a high expression level of all three adhesion molecules . for esele and icam-1 , 2.5 ng / ml tnf- , and for vcam-1 , 5 ng / ml tnf- were used as described previously [ 2123 ] . using these concentrations experimental group setup
negative controls : hlmecs without sirna transfection and without tnf- stimulation.positive controls : hlmecs without sirna transfection , but with tnf- stimulation.sirna groups : hlmecs transfected with a receptor specific sirna ( or a cocktail out of three sirna ) and with tnf- stimulation.scrambled-sirna groups : hlmecs transfected with a nonspecific sirna and with tnf- stimulation . negative controls : hlmecs without sirna transfection and without tnf- stimulation.positive controls : hlmecs without sirna transfection , but with tnf- stimulation.sirna groups : hlmecs transfected with a receptor specific sirna ( or a cocktail out of three sirna ) and with tnf- stimulation.scrambled-sirna groups : hlmecs transfected with a nonspecific sirna and with tnf- stimulation . sirna groups : hlmecs transfected with a receptor specific sirna ( or a cocktail out of three sirna ) and with tnf- stimulation . scrambled - sirna groups : hlmecs transfected with a nonspecific sirna and with tnf- stimulation . three to five different sequences ( each targeting icam-1 , vcam-1 , or esele ) were designed by using various internet - based software tools . the sequences ( all synthesized by qiagen gmbh , hilden , germany ) showing the highest knockdown were chosen for all further experiments . the following sirna sequences and concentrations were used for transfection : sense : 5-gguugaaugcaccacucaadtdt-3 antisense : 5-uugaguggugcauucaaccdtdt-3 sense : 5-gccucagcacguaccucuadtdt-3 antisense : 5-uagagguacgugcugaggcdtdt-3 sense : 5-aaugcaacucucaccuuaadtdt-3 antisense : 5-uuaaggugagaguugcauudtdt-3 siscr ( scrambled sirna ) : sense : 5-uucuccgaacgugucacgudtdt-3 antisense : 5-acgugacacguucggagaadtdt-3 eight hours after stimulation with tnf- , the cells were stained for 1 hour at 4c with the respective antibodies in 1,000 l pbs containing 0.5% fcs in a 1 : 300 dilution for esele and icam-1 and 1 : 200 for vcam-1 . antibodies ( esele : pe - cy5 antihuman cd62e ( igg1 mouse , clone 68 - 5h11 ) , icam-1 : pe antihuman cd54 ( igg1 mouse , clone ha 58 ) , and vcam-1 : fitc antihuman cd106 ( igg1 mouse , clone 51 - 10c9 ) ) were all purchased from bd biosciences pharmingen ( heidelberg , germany ) . after the staining step , the cells were washed first with 1,000 l 0.5% fcs in pbs and then with 800 l hepes - bbs and detached . the cells were centrifuged at 220 g and fixed with 2.5% paraformaldehyde in pbs . flow cytometric analyses ( 5,000 cells / measurement ) were performed with a facscan ( becton dickinson gmbh ) and evaluated with cellquestpro software ( becton dickinson gmbh ) . eight hours after stimulation with tnf- , the cells were prepared for qrt - pcr . total rna from cultured hlmecs was purified by aurum total rna mini kit ( bio - rad laboratories , inc . 200 ng of each rna sample was reverse transcribed by iscript cdna synthesis kit ( bio - rad laboratories , inc . all qrt - pcrs contained iq sybr green supermix ( bio - rad laboratories , inc . , hercules , calif , usa ) , 400 nm forward and reverse primer and 2 ng of cdna in a total volume of 15 l . primer sequences
esele : sense : 5-gcccagagccttcagtgtacc-3antisense : 5-ggaatggctgcacctcacag-3icam-1:sense : 5-cttgagggcacctacctctgtc-3antisense : 5-cggctgctaccacagtgatg-3vcam-1:sense : 5-acactttatgtcaatgttgcccc-3antisense : 5-gaggctgtagctccccgt-3
esele : sense : 5-gcccagagccttcagtgtacc-3antisense : 5-ggaatggctgcacctcacag-3 sense : 5-gcccagagccttcagtgtacc-3 antisense : 5-ggaatggctgcacctcacag-3 icam-1:sense : 5-cttgagggcacctacctctgtc-3antisense : 5-cggctgctaccacagtgatg-3 sense : 5-cttgagggcacctacctctgtc-3 antisense : 5-cggctgctaccacagtgatg-3 vcam-1:sense : 5-acactttatgtcaatgttgcccc-3antisense : 5-gaggctgtagctccccgt-3 sense : 5-acactttatgtcaatgttgcccc-3 antisense : 5-gaggctgtagctccccgt-3 all qrt - pcrs were performed in triplicates . preliminary , not in this paper , demonstrated data showed a dose - dependent fashion of the different adhesion molecules , analogous to already published cell lines . they show a concentration of 2.5 g tnf for esele und icam and 5 g tnf for vcam to get a expression profile which was not increasable any more . for the sirna cocktail , we used a tnf concentration of 2.5 g . one hundred and fifty thousand hlmecs per well were cultured and stimulated with tnf-. every 2 hours for the next 24 ( or 48 ) hours , the expression of icam-1 , vcam-1 , or esele was measured by facs . healthcare bio - science ab , uppsala , sweden ) was formed by adding 0.18 g nacl to 20 ml percoll . the percoll solution was then diluted with 0.15 m nacl to final concentrations of 55% and 74% ; 3 ml of each solution was used to form the gradient . human blood was diluted 1 : 1 with pbs , and 3 ml of this solution was placed on the top of each gradient . the gradients were centrifuged at 350 g for 20 minutes ; cells were collected from the interface and washed with 6 ml of egm-2 . erythrocytes were lysed by incubating the cells with 5 ml of lysing buffer ( 8.29 g / l ammonium chloride , 1 g / l potassium hydrogen carbonate , 0.037 g / l na2-edta-2h2o ) . after stimulating the endothelial cells with tnf- , the medium was removed and 2 million granulocytes in 1 ml egm-2 were added to each well . after 1 hour , the granulocyte suspension was gently removed , and the wells were washed with medium twice . the adherent granulocytes were detached ( detachment kit , promocell gmbh , heidelberg , germany ) , resuspended in 100 l medium and quantified using a casy cell counter system ( schrfe system , reutlingen , germany ) . the results represent the average of three independent experiments ( each performed in triplicate ) . adhesion assay : the number of attached neutrophils on tnf- stimulated hlmecs was adjusted to 100% ( positive control ) , and the number of attached cells from the experiments with specific or scrambled sirna was calculated in relation to the 100% of the control . statistical analyses were performed using the statistics software package spss ( spss software inc . , differences between the groups were calculated by f - test followed by a post hoc test ( tukey hsd ) . | background .
adhesion molecules are known to influence postoperative organ function , they are hardly involved in the inflammatory response following the ischemia - reperfusion injury .
we sought to investigate the potency of small interfering rnas to suppress adhesion molecule expression in human pulmonary microvascular endothelial cells .
methods .
human lung microvascular endothelial cells were transfected with specific sirna followed by a stimulation of the cells with an inflammatory cytokine .
adhesion molecule expression was determined by facs - analysis , and reduction of intracellular mrna was determined by qrt - pcr .
furthermore , the attachment of isolated neutrophils on the endothelial layer was determined after sirna transfection .
results . in summary ,
sirna transfection significantly decreased the percentage positive cells in a single cocktail transfection of each adhesion molecule investigated .
adhering neutrophils were diminished as well .
conclusion .
sirna might be a promising tool for the effective suppression of adhesion molecule expression on pulmonary microvascular cells , potentially minimizing leukocyte - endothelial depending interactions of a pulmonary allograft . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
human saliva contains a large number of proteins and peptides that are easily accessible and may serve as a potential source of biomarkers to monitor changes that occur under pathological conditions . the value of saliva as a biological fluid for the detection of diagnostic and prognostic biomarkers has become increasingly well established . collection of human saliva is a simple , noninvasive , and cost - effective approach for screening large populations . it is easy to handle and may be repeated without inflicting much discomfort to the subjects . diabetes mellitus is a major global health problem and it has an increasing prevalence due to several factors , such as the population growth , aging , urbanization , and increasing prevalence of obesity or lack of physical exercise . the number of people diagnosed with diabetes is increasing at an alarming rate . it is estimated that by the year 2030 , 366 million people worldwide will have the disease . most of these deaths occur in smokers . a widely used approach for measuring exposure is determination of tobacco - derived biomarkers in biologic fluids . the oral cavity is the first organ in the human body to be exposed to the cigarette smoke . the tobacco smoke alters normal homeostasis of the oral cavity , including the saliva 's antioxidant and other protective systems . the mucosal changes in smokers may also arise from the drying effects of the mucosa , high intraoral temperatures , intraoral ph changes , local alteration of membrane barriers and immune responses , or altered resistance to bacteria , fungal , and viral infections . smoking - related cell damage may leave molecular footprints in the saliva , offering the potential for noninvasive early diagnosis of tobacco - related oral diseases . saliva , and not blood , was chosen as the sample used in the study , as many reports have suggested that saliva can be an alternative to blood . saliva contains a large number of proteins that have metabolic , immune response , transporting , and several other cellular functions . its collection is noninvasive compared to the collection of other body fluids , and hence has a great potential for use in the diagnosis of systemic and localized diseases . hence , we have made an attempt to analyze the levels of sodium , potassium , and total protein in diabetic smokers and nonsmokers in comparison with healthy controls . subjects of either sex aged 30 years and above attending the department of oral pathology and microbiology , bapuji dental college and hospital , davangere was considered for inclusion in the study . group i comprised of 25 known diabetic , nonsmoking patients , group ii comprised of 25 known diabetic smoking patients , and group iii comprised of 25 nondiabetic and nonsmoking controls . a thorough general and oral examination was carried out and blood samples were collected for random blood sugar . nonstimulated saliva samples were collected and centrifuged for 30 min at 3000 rpm to obtain a clear supernatant fluid . the clear supernatant saliva was analyzed for sodium , potassium , and total protein using a semiautoanalyzer . correlation between sialochemistry of na+ , k+ , and the total protein levels and glycemic status in diabetics was determined . salivary na+ , k+ , and total protein levels between the groups were compared for statistical significance . descriptive data that included mean , standard deviation , and percentages was calculated for each group . multiple group comparisons were made by one - way anova followed by unpaired t - test for pairwise comparisons . for all the tests , a p value of 0.05 or less was considered for statistical significance . total of 75 subjects were analyzed , out of which 25 were diabetic without smokers , 25 were diabetic smokers and 25 were healthy controls . group i comprised of 25 cases out of which 14 were males and 11 were females , age range varied from 32 - 86 years , mean and standard deviation of 51.3 12.2 . group ii comprised of 25 subjects were only males , age range varied from 30 - 76 years , mean and standard deviation of 50.2 11.4 . the age range varied from 33 - 86 years , mean and standard deviation of 50.0 13.1 . the mean age in all groups was found to be statistically nonsignificant [ table 1 ] . age and sex distribution between control and diabetics . in group i diabetic nonsmokers , the random blood sugar levels ranged from 143 - 428 mg% with mean and standard deviation of 278.9 91.6 mg% . salivary sodium levels ranged from 23.9 - 271.9 mmol / l with mean and standard deviation of 118.7 80.8 mmol / l . salivary potassium ranged from 7.2 - 24.5 mmol / l , mean and standard deviation of 13.6 4.8 mmol / l . similarly , salivary total protein ranged from 0.74 - 1.39 with mean and standard deviation of 1.00 0.19 mmol / l . in group ii diabetic smokers , the random blood sugar levels ranged from 149 - 522 mg% with mean and standard deviation of 296.7 91.1mg% . mmol / l with mean and standard deviation of 92.1 47.8 mmol / l . salivary potassium levels ranged from 11.4 - 27.4 mmol / l with mean and standard deviation of 20.5 5.7 mmol / l . similarly , salivary total protein ranged from 0.91 - 1.92 mmol / l with the mean and standard deviation of 1.37 0.35 mmol / l . in controls , the random blood sugar levels ranged from 65 - 116 mg% with the mean and standard deviation of 94.4 13.3 mg% . salivary sodium ranged from 11.5 - 217.3 mmol / l with mean and standard deviation of 80.0 51.1 mmol / l with mean and standard deviation of 8.9 4.2 mmol / l and salivary total protein ranged from 0.01 - 0.94 mmol / l with mean and standard deviation of 0.47 0.28 test , on comparison of salivary sodium [ graph 1 ] between groups i and iii , [ p but there was no significance between groups ii and iii , as well as groups i and ii . the value for the salivary potassium [ graph 2 ] showed high significance on comparison of group i and group iii , group ii and group iii , and group i and ii . total protein [ graph 3 ] also showed high significance between the groups , that is , on comparison of group i and iii , group groupwise comparison of salivary sodium groupwise comparison of salivary potassium groupwise comparison of salivary total protein the statistical analysis by anova factor showed a f value of 55.8 for random blood sugar levels . for salivary sodium , no significant difference of p = 0.08 with f = 2.57 was noted , but salivary potassium showed high significance p 0001 with f = 35.0 . obtaining saliva is advantageous for patients , especially children and diabetic subjects , since the procedure is noninvasive , stress - free , and allows multiple samplings . diabetes mellitus is one such disease affecting the salivary gland functioning and thus altering the salivary constituents . sharon et al . , ben - aryeh et al . , and yavuzyilmaz et al . analyzed whole saliva , and found significantly higher potassium levels in diabetic groups compared to the nondiabetic group . sharon et al . citing studies in animals have hypothesized that in diabetes mellitus an autonomic neuropathy exists that causes sympathetic this imbalance may perhaps exert a continuous stimulation on the salivary glands , bringing about increased potassium secretion into the saliva . marder et al . attempted to explain the higher concentration of potassium in whole saliva of diabetics by different factors such as peripheral vascular damage that had much higher potassium levels in diabetics than controls and elevated conductibility of the acinic cell membrane to potassium . the findings of this study that whole saliva potassium levels are significantly higher in diabetic patients when compared to the control group . laine et al . reported that smoking was associated with higher concentration of salivary potassium , sodium , and total protein . reported that potassium decreases with increase in the salivary flow rate of chronic tobacco users . our study showed significant increase in salivary potassium among diabetic smokers but the salivary sodium levels was not significant . total salivary protein levels in diabetic nonsmokers and diabetic smokers are increased on comparison with controls and our study is in general agreement with the findings of yavuzyilmaz et al . , tenovero et al . , harrison et al . , pal et al . , and lpez et al . the increased salivary total protein in diabetics could be attributed to the increase in basement membrane permeability , allowing easy and increased passage of serum proteins into the whole saliva via salivary gland and gingival crevices . streckfus et al . have demonstrated highly significant positive correlations in salivary total protein levels among uncontrolled and controlled diabetic groups . hence , the purpose of this study was to estimate and compare the levels of salivary potassium , sodium , and total protein in smoker diabetic patients and nondiabetic smokers and controls , and to explore potential of salivary electrolytes [ na , k ] and total proteins as markers . the estimated values of salivary constituents add to the data already recorded in indian population . however , further studies using large samples are required to evaluate the findings in our study . | aims : the aim of the study was to evaluate the difference in sodium , potassium , total protein in whole saliva in diabetic smokers , diabetic nonsmokers and healthy controls.materials and methods : nonstimulated saliva samples were collected from a group of diabetic smokers , diabetic nonsmokers , and controls .
supernatant after centrifugation was used to determine the levels of sodium , potassium , and total protein by using semiautomatic analyzer.results:there exists a statistical difference in the levels of potassium and total protein between diabetic smokers , nondiabetic smokers , and controls .
difference in the levels of sodium is only significant with nondiabetic smokers and controls.conclusion:diabetes mellitus is known to alter the composition of saliva .
the purpose of this study was to estimate and compare the levels of salivary potassium , sodium , and total protein in smoker diabetic patients and nondiabetic smokers and controls , and to explore potential of salivary electrolytes [ na+ , k+ ] and total proteins as markers .
the estimated values of salivary constituents add to the data already recorded in indian population .
however , further studies using large samples are required to evaluate the findings in our study . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
the estrogen receptor ( er ) is a ligand - inducible intracellular transcription factor that mediates most of the biological effects of estrogens at the level of gene regulation [ 13 ] . estrogen biology is exceedingly complex and important in the development and function of numerous tissues and physiological phenomena [ 46 ] . in the nucleus , the er up- or downregulates the expression of target genes by interacting through its site - specific dna and with other coregulatory proteins that include coactivators and corepressors [ 13 ] . the ligand - bound er binds as homodimer to specific dna sequences termed estrogen response elements ( eres ) and regulates transcription through interaction with transcription modulators and recruitment of the general transcription machinery . in recent years , new information regarding the er structures , intra- and intermolecular interactions , posttranslational modifications , and several other factors pertaining to the er actions has emerged [ 810 ] . like other members of the nuclear hormone receptor ( nhr ) family , the er is composed of several functional domains that serve specific roles . starting from nh2- to coo - terminus , the principal domains are ( 1 ) the n - terminal domain ( ntd ) ; ( 2 ) dna - binding domain ( dbd ) ; ( 3 ) ligand - binding domain ( lbd ) . two activation function ( af ) domains , af1 and af2 , located within the ntd and lbd , respectively , are responsible for regulating the transcriptional activity of er ( figure 1(a ) ) . full transcription activity of the er is thought to be achieved by synergism between the two afs , and their activities are promoter and cell specific . af1 functions as hormone independent , whereas af2 function requires the presence of hormone / steroid [ 12 , 17 ] . in this paper , we focus on the two isoforms of human er ( er ( nr3a1 ) and er ( nr3a2 ) ) , encoded by two different genes . both have been cloned and characterized . the physiological effects of estrogen are manifested through both er and er. the er and er receptor isoforms display distinct tissue distributions and signaling response [ 1921 ] . er and er have also been shown to form hetero dimers on eres . in terms of sequence homology , the er shows a high homology to er in the dbd ( more than 95% amino acid identity ) and in the lbd ( ~55% amino acid identity ) [ 19 , 22 ] . however , the ntd of er is shorter than that of er with a very poor sequence homology of only 15% compared to that of er. the three - dimensional structures of the independently expressed dbd and lbd have been solved and show overall folds that represent globular proteins with natively ordered conformations [ 13 , 2325 ] . to date , no three - dimensional natively folded structure for the ntd is available not only for the er but for the entire nuclear hormone receptor ( nhr ) superfamily . even though the full length structure of the peroxisome proliferator - activated receptor- ( ppar- ) has been solved , it failed to show any signature of structure formation in its ntd . warnmark et al . have previously provided insights about the structural and functional correlations regarding the er ntd . in this paper , we discuss the knowledge about the structural characteristics of the er and its role in gene regulation . the d domain which follows dbd is known as a hinge region ( figure 1(a ) ) . it contains nuclear localization signal which gets unmasked upon ligand binding and serves as a flexible region connecting dbd and lbd . its action was first characterized by montano et al . by single - point mutations in the domain as well as by whole domain deletion . f domain was found to modulate gene transcription in a ligand - specific manner . the ligand , promoter , and tissue - specific modulation capabilities of the f domain were recently studied in detail by koide et al . . like other nhrs , the e domain of er contains lbd ( figure 1(a ) ) . it consists of 12 helices , contains hormone binding pocket , and is responsible for the most part of functions activated by ligand binding , such as coregulator binding to af2 and dimerization interface . while er and er have both overlapping and unique functions , the overall homology between the er protein lbd and er protein lbd does not exceed 55% . however , the two proteins ( er and er ) display distinct regions of sequence homology [ 4 , 19 ] . the amino acid residues 223343 and 404457 in er and er show a significantly higher homology than that of the sequence encompassing 223457 and 344403 , respectively . interestingly , the stretch of the er lbd amino acid residues 465468 , with lowest homology to er , has been found to be most solvent accessible . on the other hand , the conserved regions with greater homology are protected against degradation and are in direct contact with the ligand . despite low sequence homology in lbds within the nhr superfamily , both isoforms of er - lbds have been shown to form dimers with agonist and antagonist ligands . the dimer interface is primarily encompassed by helices 10 and 11 . as a member of the nhr superfamily of transcription factors , er contains a globular lbd structure that harbors a hormone - binding site , a homo- or heterodimerization interface , and coregulator ( activator and repressor ) interaction sites [ 3538 ] . the er lbd structure contains 11 -helices ( h1h12 ) [ 24 , 39 ] ( figure 1(c ) ) . the first crystal structure of an er lbd bound to its natural ligand 17-estradiol ( e2 ) showed that in a compact ellipsoid cavity , e2 is buried in a highly hydrophobic environment . within this pocket ( formed by 22 residues ) , hydroxyl groups in estradiol at positions 3 and 17 play a crucial role in orienting the steroid / hormone ligand . these hydroxyl groups of the a and d rings are hydrogen bonded to glu353 from h3 , arg394 from h5 , and a water molecule and his524 from h11 . in an agonist - bound form , er is spatially organized in a three - layered structure with helices 4 , 5 , 6 , 8 , and 9 lining up on one side by h1 and h3 , and on the other side are helices 7 , 10 , and 11 . due to the central role of estrogen signaling in diverse diseases ranging from cancer to aging , the crystal structure of the complex of er lbd bound to the nonsteroidal ligand , diethylstilbestrol , also shows that the hydrophobic interactions primarily govern the accommodation of distinct lbd structures . the crystal structures of the human er bound to genistein , estradiol ( figure 1(c ) ) , and rat er to raloxifene assert the importance of hydrogen bond network on the opposite sides of the respective ligands . the bicyclic moiety of genistein orients in a position similar to the c- and d - ring of e2 , facilitating the formation of hydrogen bonds of hydroxyl moieties with histidine groups of the receptor . the specificity of the ligand association between the er and er may stem from the distinction in the residues lining the binding pocket . quite diverse family of compounds ( estrogens , some androgens , phytoestrogens , antiestrogens , and environmental estrogens ) have been shown in the past to have estrogenizing activity , and to interact with the er from rat uterus and human breast tumor cells . interactions of these structurally diverse ligands highlight the intrinsic er and er lbd plasticity [ 4749 ] . adjacent to the n - terminal transactivation region ( a / b domain ) , a conserved c domain encompasses the dna - binding sequence . this dna - binding domain associates with the response elements which can either reside proximally to the promoter regions or enhancer regions located distant from the transcription initiation site . er dna binding domain usually binds to the estrogen response element ( ere ) composed of a palindromic hexanucleotide 5aggtcannntgacct3 [ 5153 ] . the ere sequences play an important regulatory role [ 54 , 55 ] . not only does it dictate the binding affinity of the er , but also it has been shown to modulate the recruitment of coactivators [ 56 , 57 ] . the er dbd : ere structures have been studied extensively by several biophysical techniques [ 13 , 23 , 55 , 58 ] . three - dimensional structure of the er has been solved using nuclear magnetic resonance as well as x - ray crystallographic techniques both alone and in complex with dna ( figure 1(b ) ) [ 13 , 23 , 55 , 58 ] . the dbd : ere interactions and ere - facilitated dimerization are in part mediated through the p box and d box sequences in the zinc finger domains . these zn finger subdomains are comprised of 8 cysteine residues that coordinate with the two zn ions . while p box actively interacts with the ere nucleotides , the d box is present at the dimerization interface [ 29 , 30 , 54 ] . the specificity of er recognition by ere is exemplified by interesting studies describing its association with glucocorticoid response element ( gre ) . three amino acids in the first zn finger region or er dictate its interaction with ere and gre . substitution of these three amino acids with the corresponding amino acids from the glucocorticoid receptor 's dbd completely changes er dbd 's specificity for an ere , and it strongly binds to a gre sequence to initiate gre - mediated transcriptional activity [ 13 , 23 , 54 , 55 , 58 ] . transcriptional regulation at the ere can be mediated via two separate mechanisms of er action . liganded er can directly associate with specific response element sequences . in the other mode of action , the er may participate in a multiprotein , preinitiation complex and regulate gene transcription without a direct interaction with any dna sequence [ 5961 ] . together , these mechanisms highlight the complex role of coactivators and response elements in eliciting specificity in transcriptional output . to date relatively little information has been available on the structure of the n - terminal regions of the nhrs . even though the full - length structure of the peroxisome proliferator - activated receptor- ( ppar- ) has been solved it failed to show any signature of structure formation in its very short ntd . we and others have shown that the glucocorticoid receptor 's n - terminal transactivation af1 region and a shorter core fragment of af1 , the af1 core , are unstructured in aqueous solution [ 6266 ] . in other words , the ntd amino acid sequences possess an intrinsically disordered ( i d ) conformation , a feature of activation domains of many transcription factors [ 27 , 62 , 65 , 67 , 68 ] . similar results have been reported for the er and er , androgen- , and progesterone receptor [ 6971 ] . thus , activation domains of many signaling proteins including the er 's ntd / af1 are known to exist in an i d state . one of the reasons for their existence as an i d region seems to be to help them in promoting molecular recognition by providing surfaces capable of binding specific target molecules [ 7275 ] . the computational analyses have established that under physiological conditions , the combination of low mean hydrophobicity and relatively high net charge represent an important prerequisite for the lack of well - defined compact structure in proteins or protein regions / domains . the i d nature of the er ntd / af1 has been confirmed by circular dichroism method . we performed secondary structural analyses of the er and er ntd using network protein sequence analysis . the analytical results show that more than 67% of er ntd contains random coli conformation , whereas in case of er , the amount of random coil is found to be more than 80% with only a small proportion as helix and sheet in both the cases ( figure 2 ) . it has been proposed that the i d nature of an activation domain allows it to rapidly sample its environment until appropriate concentration and affinity of the binding partner proteins are found , meaning that they may not be structured until they have recruited and bound their proper interaction partners . then , either by induced - fit or selective binding of a particular conformer , a high - affinity activation domain : binding partner protein interaction occurs [ 65 , 73 ] . in case of nhrs ' i d ntd / af1 domains , it has been shown that they undergo a transition to a folded state upon interaction with either components of the general transcription machinery or with other comodulators . several coregulatory proteins are involved in the effect of the er on target gene transcription . the tata box - binding protein ( tbp ) has a central role in the basal transcription machinery and can directly bind to the ntd of the er but fails to bind to er ntd and to potentiate er - activated transcription . this difference in tbp binding could imply differential recruitment of target proteins by the ntds of er and er. the affinity of the er ntd : tbp interaction was determined to be in the micromolar range , as assessed by surface plasmon resonance spectroscopy . based on these results , it has been proposed that the interaction between er ntd and tbp may proceed in a two - step manner with initial very fast , low - affinity association , followed by a slow , folding event and tighter association . the initial association may be occurring by electrostatic interactions between the acidic residues of highly negatively charged er ntd and the positively charged tbp . however , this initial unstable protein complex subsequently may convert into a more stable form by the folding of the i d er ntd and the formation of specific contacts between the two proteins . in this study , the secondary structures of the independently expressed ntds of the er and er were analyzed using nmr and circular dichroism spectroscopy . further , when er ntd was bound to tbp , structural changes were induced in er ntd . these results support models of tbp as a target - protein for the n - terminal activation domain of er. further , the dissociation of this binding suggests a complex behavior , with a rapid dissociation for er ntd molecules that did not undergo proper folding and a slower dissociation for those molecules that did fold successfully upon physical interaction with the tbp . such a two - step binding mechanism is consistent with the change in protein conformation that accompanies the er ntd : tbp interaction . observed differences in binding of tbp to er ntd and er ntd supports a model where the two receptors may be utilizing different sets of target binding proteins . this is consistent with the reports of functional differences between er ntd and er ntd where it has been shown that the er af1 domain can function in an autonomous manner , whereas the af1 function of er can not . it has also been reported that under most conditions er possesses a weaker transactivational potency compared to er , and these differences appear to be cell and promoter specific . we have earlier shown that tbp binding induces secondary / tertiary structure formation in the i d af1 domain of the glucocorticoid receptor such that af1 's interaction with specific coregulatory proteins and subsequent af1-mediated transcriptional activity is significantly enhanced [ 77 , 78 ] . based on the binding of tbp and consequent folding of these i d activation domains , it can be hypothesized that the interaction between nhrs ' ntd / af1 and tbp may be a unified mechanism , through which these i d af1/ntd acquire a functionally active conformation under physiological conditions . in this conformation , the ntd / af1 may be able to create favorable protein interaction surfaces for its interaction with specific coregulatory proteins . of course , the exclusion of certain other binding partners can not be ruled out . it could thus be hypothesized that a complex and dynamic binding pattern for the n - terminal activation domains of the nhrs occurs to achieve transcriptional activation , where the ntd / af1 region must be able to obtain different conformations dependent on the binding partner(s ) . however , a clear picture will emerge only when the functionally folded three - dimensional structure of the ntd / af1 is solved . at least for now , the differential effects observed in case of two er isoforms ( er ntd and er ntd ) suggests that tbp may not be a common coregulator that must bind / fold all the nhrs ' ntd / af1 . thus , it is quite possible that other protein components from the basal transcription machinery may provide such interactions . in fact , we and others have observed that at least in case of the androgen receptor , its i d ntd / af1 undergoes disorder / order transition through its interaction with rap74 , a subunit of tfiif , an important component of basal transcription machinery [ 70 , 79 ] . recent observations have led to the conclusion that in cells , er and several other nhrs behave very dynamically such that their kinetic behavior in cells allows them to rapidly interact with various coregulatory proteins , and with chromatin and dna . further , the er moves to various sites in cells to function , and the local concentrations and various other constellations of potential coregulatory proteins are required to associate with the er to activate or repress the expression of target genes . the lbd crystal structures have clearly demonstrated that differing sets of coactivators / corepressors come together in response to agonist or antagonist ligand binding , such that agonist in one cell type can be an antagonist in another cell type . the overall picture is one of a complex , dynamic network controlled by the er . it is not yet clear whether unique tissue / cell - specific coregulatory protein interactions can fully explain the tissue / cell - specific actions of the er and other nhrs . when the clear picture will emerge , it is certain that other dynamic considerations will prove to be the dominant underlying mechanism . | the estrogen receptor ( er ) mediates most of the biological effects of estrogens at the level of gene regulation by interacting through its site - specific dna and with other coregulatory proteins . in recent years ,
new information regarding the dynamic structural nature of er has emerged .
the physiological effects of estrogen are manifested through er 's two isoforms , er and er. these two isoforms ( er and er ) display distinct regions of sequence homology .
the three - dimensional structures of the dna - binding domain ( dbd ) and ligand - binding domain ( lbd ) have been solved , whereas no three - dimensional natively folded structure for the er n - terminal domain ( ntd ) is available to date .
however , insights about the structural and functional correlations regarding the er ntd have recently emerged . in this paper , we discuss the knowledge about the structural characteristics of the er in general and how the structural features of the two isoforms differ , and its subsequent role in gene regulation . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
febuxostat , a nonpurine xanthine oxidase inhibitor , is approved as the first - line urate - lowering therapy in gout patients with normal renal function or mild to moderate renal impairment . the most common adverse effects of febuxostat are liver function test abnormalities , diarrhea , and skin rash . however , there is insufficient data in patients with severe renal impairment and end - stage renal disease ( esrd ) . we report the first case , to our knowledge , in which agranulocytosis developed after febuxostat treatment in an esrd patient . a 67-year - old woman with gout and esrd received febuxostat 40 mg a day for 2.5 months . after broad - spectrum antibiotics treatment and no more exposure to febuxostat for 17 days , her infection and neutrophil count recovered . as febuxostat use may become more common in the population of advanced renal failure , clinicians should be aware of this rare but potentially life - threatening adverse effect . based on our experience gout is the most common inflammatory arthritis , and the clinical presentation is characterized by acute episodic flares of intense joint pain , caused by the deposition of monosodium urate crystals . hyperuricemia is the central risk factor for gout and the pharmacologic therapeutic goal of gout is to lower serum uric acid level below 6 mg / dl , the saturation point for monosodium urate . in patients with chronic kidney disease ( ckd ) , their urinary uric acid secretion is decreased , and the prevalence of hyperuricemia is higher than those with normal renal function . on the other hand , elevated serum uric acid has been proposed to attribute to ckd progression . for ckd patients with gout , aggressive control of serum uric acid level is crucial for renal protection and gout therapy . febuxostat , a nonpurine xanthine oxidase inhibitor , has been approved by united states food and drug administration since 2009 for chronic management of hyperuricemia in patients with gout . when comparing with the traditional xanthine oxidase inhibitor , allopurinol , febuxostat has shown its superiority in greater urate - lowering efficacy . the most common adverse effects of febuxostat include liver function abnormalities , diarrhea , and skin rash . according to the american college of rheumatology guidelines , febuxostat is recommended as the first - line pharmacologic urate - lowering therapy approach in gout . the recommended starting dose is 40 mg once daily . in patients with mild to moderate renal impairment ( creatinine clearance , crcl 3089 ml / min ) , nevertheless , early clinical trials exclude patients with crcl less than 30 ml / min . there is still insufficient experience for febuxostat usage in patients with severe renal impairment and end - stage renal disease ( esrd ) , and the safety is still a concern . here , we report an esrd patient developing agranulocytosis after febuxostat treatment . a 67-year - old woman was sent to emergency department because of fever and sore throat . her medical history included esrd with regular hemodialysis 3 times per week for 15 years , hyperuricemia and gout . she also had comorbidity such as dyslipidemia , hypertension , secondary hyperparathyroidism , and diabetes . the hemogram revealed a white blood cell count ( wbc ) of 700/l , with 2% neutrophils , 94% lymphocytes and 2% monocytes , hemoglobin 11.1 g / dl , and platelet count , 131,000/l . under the impression of febrile neutropenia and acute pharyngitis , broad - spectrum antibiotics with piperacillin 2 g and tazobactam 0.25 gm i.v . q8h had been administered and her infection sign resolved gradually . upon admission , we reviewed her oral medication : glipizide 5 mg tid , saxagliptin 2.5 mg qd , fenofibrate 600 mg qd , aluminum hydroxide 324 mg tid , folic acid 5 mg qd , calcium carbonate 1000 mg tid , and febuxostat 40 mg qd . besides , she also received epoetin - beta 2000 iu i.v . febuxostat was administered 2 months before admission for inadequate serum uric acid control by allopurinol 50 mg qd . two weeks before febuxostat exposure , routine laboratory test revealed wbc 6000/l and serum uric acid level 9.8 mg / dl . febuxostat was discontinued thereafter due to the causal relationship of agranulocytosis can not be excluded . there were no clinical or laboratory evidence of epstein barr virus , cytomegalovirus , or human immunodeficiency virus infections ; antinuclear antibody ( ana ) and antiextractable nuclear antigen ( anti - ena ) were both negative . bone marrow examination during hospitalization showed hypocellular marrow with a marked decrease in myeloid component but no evidence of hematologic neoplasms . the patient denied history of radiation or chemicals exposure . after stopping febuxostat for 17 days , her neutropenia improved significantly ( wbc 2100/l , and neutrophil 66% ) , without any granulocyte colony - stimulating factor ( g - csf ) support . after discharge , her wbc and differential count was completely normal during follow - up ( fig . the time course of drug administration , total wbc count ( wbc 1000/l ) and absolute neutrophil count ( anc ) . agranulocytosis is characterized by a decrease in peripheral neutrophil count less than 500/l and most cases in adults result from adverse effects of drugs . except for chemotherapy , the clinical presentation is usually fever due to infection , so - called febrile neutropenia , and upper aero - digestive tract infection is the most common infection source just as our patient . the most important managements of drug - induced agranulocytosis are to identify the culprit drug , to withdraw this agent , and to give broad - spectrum antibiotics for those with infection . in our case , before the initiation of febuxostat , her wbc was normal and agranulocytosis developed 2.5 months after taking febuxostat . other medications that the patient had taken for her underlying chronic diseases were considered to be unlikely culprits , because she had been receiving them for several years and continued after this episode . we stopped febuxostat immediately when we suspected the causality of this offending drug to agranulocytosis . her neutrophil count recovered in 17 days without other intervention such as steroids or g - csf . due to the safety concern using naranjo probability scale , we determined the occurrence of agranulocytosis related to febuxostat treatment to be probable . febuxostat has been approved in the usa for the chronic management of hyperuricemia in patients with gout , and in eu for the treatment of chronic hyperuricemia in conditions where urate deposition has already occurred , and in japan for the treatment of hyperuricemia in patients with or without gout . the recommended starting dose range from 40 to 80 mg daily in different countries and no dose adjustment is required for those with mild to moderate renal impairment ( crcl > 30 ml / min ) . however , the efficacy and safety have not been fully evaluated in patients with severe renal impairment ( crcl < 30 ml / min ) and esrd , and febuxostat has not been approved in this population . in a small series of 17 hemodialysis patients with asymptomatic hyperuricemia , febuxostat 10 mg once daily has demonstrated significant urate - lowering efficacy and no specific adverse effect was observed . another series of 5 male cases with chronic hemodialysis , febuxostat 10 to 20 mg daily were administered without apparent adverse effects . regarding agranulocytosis induced by febuxostat , 2 patients with ckd have been reported . a 74-year - old woman with liver cirrhosis and ckd stage 3 eleven days after febuxostat administration , she developed neutropenia with absolute neutrophil count only 600/l . another 68-year - old man with ckd stage 5 due to diabetic nephropathy was administered with febuxostat 10 mg per day for inadequate uric acid control during admission . after 3 days of febuxostat treatment , his neutrophil count dropped from 5700 to 500/l . in these 2 patients , neutrophil count increased within 5 to 7 days after febuxostat discontinuation and with g - csf support . apparently , our patient had more advanced renal disease and exposed to higher dose of febuxostat for longer duration ( 40 mg qd for 2.5 months ) , and then had lower neutropenic nadir and exhibited longer neutropenic period of time . table 1 shows comparison of clinical presentation of reported cases of febuxostat - induced agranulocytosis . as for patients with esrd and hemodialysis , febuxostat could improve endothelial dysfunction and reduce serum uric acid levels and oxidative stress . with these study results , it is expected that more and more patients with crcl less than 30 ml / min will receive this drug and some other patients may experience this uncommon adverse effect , agranulocytosis , which is seldom reported in patients without advanced renal failure . immune - mediated destruction of circulating neutrophils suggests that a reactive metabolite of febuxostat may irreversibly bind to the membrane of mature neutrophils . we report an esrd patient experiencing febuxostat - induced agranulocytosis , a rare but severe complication . when prescribing febuxostat to patients with severe ckd or esrd , clinicians should be aware of this potentially life - threatening adverse effect and a regular hemogram monitoring during the initial treatment of febuxostat is recommended . | abstractintroduction : febuxostat , a nonpurine xanthine oxidase inhibitor , is approved as the first - line urate - lowering therapy in gout patients with normal renal function or mild to moderate renal impairment .
the most common adverse effects of febuxostat are liver function test abnormalities , diarrhea , and skin rash .
however , there is insufficient data in patients with severe renal impairment and end - stage renal disease ( esrd ) .
we report the first case , to our knowledge , in which agranulocytosis developed after febuxostat treatment in an esrd patient.clinical presentation : a 67-year - old woman with gout and esrd received febuxostat 40 mg a day for 2.5 months .
she subsequently complicated with febrile neutropenia and the absolute neutrophil count was only 14/l .
after broad - spectrum antibiotics treatment and no more exposure to febuxostat for 17 days , her infection and neutrophil count recovered .
bone marrow study during neutropenic period showed myeloid hypoplasia without evidence of hematologic neoplasms.conclusion:as febuxostat use may become more common in the population of advanced renal failure , clinicians should be aware of this rare but potentially life - threatening adverse effect . based on our experience , close monitoring hemogram and immediate discontinuation of this medication may prevent serious consequences . |
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the most frequent location for this tumor is the proximal phalanx , followed by the middle phalanx and the metacarpals . the tumor arises in the medullary cavity and grows into the cortex , forming a prominent endogenous mass in the bone . because this tumor type has no unique clinical symptoms , an enchondroma is always difficult to diagnose . no previous study has reviewed the clinical presentation , treatment , differential diagnosis , tumor features , rate of recurrence , or complications of enchondroma in the distal finger . the medullary cavity and the nail in the fingertip are small , and an enchondroma in the distal finger is distinct from enchondromas occurring in other parts of the body precisely because of these characteristics . the particular details of the clinical presentation ( clubbing or mallet finger deformity ) and the intraoperative treatment with curettage and/or bone grafting will be discussed in this paper . this was a retrospective study with 34 patients ( 19 women and 15 men ) presenting with an enchondroma in the distal phalanx of the finger who underwent an operation between may 2004 and september 2012 ( table 1 ) . the exclusion criteria were patients with type 2 diabetes , autoimmune diseases , or ollier disease ; those receiving steroids or immune suppressant medications ; and patients with a history of injury . we recorded the age , gender , tumor location , initial symptoms , types of surgery received , follow - up period , recurrence , and complications . the average patient age was 39.38 10.97 years ( range 1459 ) . a brachial tourniquet was used , and the procedure was carried out with surgical loupes . the surgical technique involved removal of tumors by opening a cortical window and curetting the cavity . after curettage , saline was used for medullary cavity brushing with a syringe needle , and dehydrated alcohol was instilled to kill the tumor cells . bony nonunion was diagnosed when radiographic evidence of grafting absorption was present 6 months after the surgery . radiological imaging , including anteroposterior ( ap ) and lateral plain radiographs of the finger , was used to assess for expansion of the bone and thinning of the cortex as a tumor recurrence standard . summary data of patients . written informed consent was obtained from each patient for publication of this article and any accompanying images . ethical approval was provided by the medical ethics committee of the first affiliated hospital , college of medicine , zhejiang university . the enchondroma was located roughly equally in the index finger ( 10 patients [ 29% of cases ] ) , middle finger ( 9 patients [ 26% of cases ] ) , and ring finger ( 8 patients [ 24% of cases ] ) and appeared less often in the thumb ( 5 patients [ 15% of cases ] ) and the little finger ( 2 patients [ 6% of cases ] ) . the presenting initial symptoms of patients included pain in 15 patients ( 44% of cases ) , a clubbing finger deformity in 9 patients ( 26% of cases ) , an infection in 6 patients ( 18% of cases ) , and a mallet finger deformity in 4 patients ( 12% of cases ) ( fig . a ct scan and 3-dimensional reconstruction were necessary if the patients had cortical bone defects ( fig . 2 ) . in total 15 patients ( 29% of cases ) had cortical thinning and enlargement , 12 patients ( 35% of cases ) had unilateral bone cortex defects , and 6 patients had bilateral bone cortex defects ( both volar and dorsal sides , 18% of cases ) . patients received 2 types of surgery , curettage ( 10 patients [ 29% of cases ] ) and curettage with bone grafting ( 24 patients [ 71% of cases ] ) . the mean follow - up period was 7.18 2.17 years ( range 311 ) . none of the patients were lost to follow - up or had a tumor recurrence . a delayed union occurred in 1 patient ( 3% of cases ) , but bone union was achieved 1 year later after the patient underwent conservative treatment with bracing . three patients ( 9% of cases ) had an infection treated by curettage without bone grafting . one patient who developed chronic osteomyelitis twice received surgical debridement and ultimately underwent an amputation . patient examples are presented with the different initial symptoms ( a ) clubbing finger deformity and ( b ) mallet finger deformity . an enchondroma in the distal phalanx has different clinical symptoms relative to other parts of the body . clubbing of the fingers , also known as hippocratic fingers , is part of a syndrome that includes hypertrophic osteoarthropathy and endocrine , pulmonary , neoplastic , and multisystem diseases . a neoplastic disease , such as an osteoid osteoma , mucoid cyst , or enchondroma , usually causes a bilateral clubbing deformity , but clubbing can rarely occur in a single finger . a cortical bone defect can lead to a local subcutaneous infection ; the tumor body was completely composed of liquefactive necrosis in some of our patients . the diagnosis should be differentiated from offending organisms , acute leukemic infiltration , cutaneous mycobacterium marinum infections , and metastases of various carcinomas . tearing off the attachment of the terminal extensor tendon can also cause a mallet finger deformity . the presence of a typical expansile lytic lesion with cortical thinning in ap and lateral plain radiographs and intra - operative bone necrosis are helpful to differentiate an enchondroma from other diseases . an mri may contribute to the differential diagnosis and treatment in patients with an infection ( fig . 3 ) . an mri can identify soft tissue infections , abscess formation , or osteomyelitis in the distal finger and can also provide guidance for clinical antibiotic therapy . ( a , b ) a t2-weighted mri image shows an enchondroma with hypersignal intensity , with the tumor invading the soft tissue leading to an infection of the whole finger . the surgical incision to treat an enchondroma can be palmar , dorsal , lateral , or fish mouth . in our experience , the advantage of a palmar incision is that this approach allows the tumor to be fully exposed , but a painful scar remains . based on the imaging , especially the results of ct 3-dimensional reconstruction , we prefer to choose the side with the defective bone cortex because this surgical approach is convenient for bone grafting . standardized management of an enchondroma in the distal finger is important , especially when dealing with such rare tumors . a high - speed burr or hypercator cauterization is not suitable for cavity curettage because the finger cavity is too small . we used saline for medullary cavity brushing and dehydrated alcohol instillation to kill the tumor cells . however , if patients have large osseous defects or a pathological fracture , bone grafting is helpful for their early exercises and functional recovery . considering the difficulty of a grafting operation in such a small cavity instead , we found that an injectable calcium phosphate cement was an acceptable alternative . for patients that had bilateral cortical bone defects , bone grafting will irritate the soft tissue and cause a postoperative infection . we selected bone grafting only for patients with a controlled infection and unilateral defects ( fig . 4 ) . these images show a 51-year - old male with an enchondroma in his distal ring finger . ( a , b ) preoperative radiography and ( c , d ) 1-year postoperative radiography after tumor curettage with a calcium phosphate cement graft shows bone union and no tumor recurrence . first , we had a relatively small group of patients available to assess due to the disease 's rarity . second , making a definitive diagnosis for the patients who present with pain or infection takes time , so those patients typically have more complications and a higher recurrence rate . in conclusion , if patients have large osseous defects and a controlled infection , we recommend a bone graft . based on the results of ct 3-dimensional reconstruction , we prefer a surgical approach involving the side with the defective bone cortex . if patients have an infection or bilateral bone cortex defects , bone grafting is challenging . | abstractthe goal of our study was to report the clinical presentation , treatment , and complications of enchondroma in the distal phalanx of the finger .
this was a retrospective study of 34 patients ( 19 women and 15 men ) who underwent surgery between may 2004 and september 2012 for enchondroma in the distal phalanx of the finger .
the average age of the patients was 39.38 10.97 years old ( range 1459 ) .
the presenting symptoms and imaging features were recorded .
the surgical procedure was performed under regional or general anesthesia .
the surgical technique involved removal of tumors by opening a cortical window and curetting the cavity .
the defects were filled with an injectable calcium phosphate cement .
all patients received follow - up in our outpatient clinic every 6 months .
expansion of bone or thinning of the cortex present in the radiological imaging , including anteroposterior and lateral plain radiographs of the fingers , was used to assess for tumor recurrence .
the observational end - point was reoperation.all tumors were confirmed as enchondromas by the pathological results .
none of the patients had a tumor recurrence .
three patients ( 9% of cases ) developed an infection .
after antibiotic treatment , 2 patients were cured , and 1 patient required an amputation .
enchondroma in the distal phalanx of the finger presents with a variety of clinical symptoms .
injectable calcium phosphate cement is adequate for bone grafting .
postoperative infection is more common than tumor recurrence .
if patients have an infection or bilateral bone cortex defects , bone grafting is challenging.level of evidence : therapeutic study , level iv |
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placental mesenchymal dysplasia ( pmd ) is a rare , benign placental , vascular anomaly . pmd is characterized by placentomegaly and grape - like vesicles resembling molar pregnancy on ultrasonography ( usg ) . the differential diagnosis of pmd includes partial molar pregnancy , complete mole with coexisting normal fetus , chorioangioma , subchorionic hematoma , and spontaneous abortion with hydropic changes . prenatal recognition of pmd during early and late gestation could prevent unnecessary termination of pregnancy . an 18-year - old female , gravida 1 , para 1 was referred for routine ultrasound at 15-week gestation . usg findings showed single live fetus , with mildly thickened , low lying posteriorly placed placenta . lower part of the placenta showed multiple anechoic cysts which were extending inferiorly and covering the internal os . on color doppler , low velocity blood flow was seen within the multicystic placental lesion [ figure 1 ] . a single live fetus of 15-week gestational age without any gross structural anomaly was seen [ figure 2 ] . based on usg findings , differential diagnoses of a partial mole or a complete mole with coexisting twin and pmd were considered . maternal serum levels of afp ( 181.37 ng / ml ) and - hcg ( 223846 miu / ml ) were raised . in an 18-year - old female at 15-week gestation , transabdominal ultrasonography ( tas ) of pelvis ( a - c ) shows mildly enlarged , low lying , posteriorly placed placenta . lower part of the placenta shows multiple anechoic cysts ( arrows ) which are extending inferiorly and covering the internal os . on color doppler imaging ( d ) , low velocity signals were seen in the multicystic placental lesion tas of the fetus showing no gross structural anomaly in view of the presence of normal fetus on ultrasound and low velocity color flow within the cystic placental mass on color doppler , the possibility of pmd was considered . she was kept on conservative management and informed about the increased risk of complications such as fetal growth restriction or intrauterine fetal demise and preterm delivery and association of preeclampsia . subsequently , dilatation and evacuation was done and placental tissue was sent for histopathological examination . on follow - up pathological examination suggested hydropic stem cell villi with central cistern containing myxoid stroma with interspersed normal - sized chorionic villi . few congested blood vessels were seen in the stroma of hydropic villi in the periphery . there was characteristic absence of trophoblastic proliferation around the periphery of the abnormal villi , absence of stromal cell inclusions , and scalloping of villous surface ( diagnostic hallmark of molar pregnancy ) [ figure 3 ] . based on usg and histopathological findings , histopathological examination ( higher magnification ) of placental lesion shows hydropic stem cell villi ( long white arrow ) , central cistern ( small white arrow ) containing myxoid stroma and few congested blood vessels ( small black arrows ) in the stroma of hydropic villi with interspersed normal villi ( long black arrow ) previous studies have described the characteristic placental pathology and the possible genetic basis for the placental abnormality in pmd . pmd cases with beckwith wiedemann syndrome ( bws ) are associated with paternal isodisomy of the 11p15.5 region . in addition to association with bws , there are multiple case reports in the literature describing the coexistence of pmd with fetal hepatic mesenchymal tumors , suggesting a common pathogenetic origin for the two anomalies . it is important to distinguish pmd from a partial mole with an abnormal triploid fetus because this diagnosis may result in termination of pregnancy . it is difficult to distinguish pmd from a complete mole with coexisting normal fetus , which carries significant morbidity to the mother ( persistent gtd ) . pmd is associated with intrauterine growth restriction and intrauterine fetal demise / neonatal death / without fetal development . the cause of fetal death in many cases is fetal vascular obstructive pathology causing longstanding , severe fetal hypoxia characterized by chorionic vessel thrombosis . pmd should be included in the differential diagnosis of cystic lesions of the placenta on usg . the placenta of a complete mole with coexisting normal fetus and partial molar pregnancy appears heterogeneous , with partially solid and cystic areas . on usg , a chorioangioma is a focal lesion and is hypoechoic compared to the rest of the placenta . pmd in the first trimester shows no blood flow in the cystic spaces of the placenta on color doppler . however , in the third trimester , large vascular areas with turbulent blood flow are observed ( either arterial or venous ) , which are located mainly under and at the level of the chorionic plate . these changes are due to the progressive dilatation of chorionic arteries and veins , which become aneurysmal . low or absent venous signals may be associated with pmd during the first two trimesters , as seen in our case also . on color doppler , high velocity and low resistance flow is seen in the molar mass and large feeding vessel or increased vascularity is seen in the mass of chorioangioma , although no blood flow is seen within the mass in hematoma . differentiation of pmd from a twin pregnancy with a complete mole and coexistent fetus is difficult . on the first trimester sonography , documentation of two gestation sacs , a lesion that constitute the entire thickness of the placenta , and the lack of blood flow signals suggest the diagnosis of complete mole with coexistent fetus . the diagnosis of pmd is only confirmed after evaluation of placental pathology . grossly , it is characterized by placentomegaly , dilated or aneurysmal chorionic vessels , and fibromuscular hyperplasia or cystic villi . microscopic findings include mesenchymal hyperplasia and edema of stem - cell villi , which contain thick - walled vessels . these findings were seen in the present case along with characteristic absence of trophoblastic hyperplasia , which is a diagnostic hallmark of gtd . there are no abnormal fetal vessels in the stem villi seen in cases of twin pregnancies with one complete mole , as in pmd . immunohistochemical tests using antibodies against p57 protein ( an imprinting gene only expressed in the maternal genome ) might prove helpful in distinguishing pmd from molar pregnancies . in a complete mole , invasive testing ( chorionic villus sampling or amniocentesis ) should be performed to confirm a normal karyotype and exclude partial molar pregnancies . triploidy associated with pmd is presumed to occur either as a result of maternal - derived triploidy or due to placental mosaicism . normal karyotype was seen in 78% of the cases , chromosomal abnormalities in 3 cases ( trisomy13 , klinefelter syndrome and triploidy ) , and bws in 15 cases ( 23% ) . although - hcg levels are always elevated in molar pregnancies , pmd may also present with increased - hcg levels . pmd is more likely to be associated with elevated maternal serum afp levels , as seen in our case . usg appearance suggestive of molar pregnancy along with increased maternal serum afp levels and normal or slightly elevated - hcg levels can suggest the diagnosis of pmd . the possibility for pmd should be evaluated when an abnormal fetus , or an abnormal karyotype , or both are found . a detailed anatomical evaluation should be performed to rule out associated fetal anomalies , mainly findings consistent with bws . because pmd is associated with adverse pregnancy outcome , surveillance with serial growth scans , genetic evaluation , and third - trimester assessment of wellbeing should be considered . | placental mesenchymal dysplasia ( pmd ) is an uncommon vascular anomaly of the placenta characterized by placentomegaly with multicystic placental lesion on ultrasonography and mesenchymal stem villous hyperplasia on histopathology .
placental mesenchymal dysplasia should be considered in the differential diagnosis of cases of multicystic placental lesion such as molar pregnancy , chorioangioma , subchorionic hematoma , and spontaneous abortion with hydropic placental changes
. however , lack of high - velocity signals inside the lesion and a normal karyotype favor a diagnosis of pmd .
pmd must be differentiated from gestational trophoblastic disease because management and outcomes differ .
we report the case of an 18-year - old female at 15 weeks of gestation with sonographic findings suggestive of placental mesenchymal dysplasia .
the diagnosis was confirmed on histopathology . |
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tobacco smoking is the second leading risk factor for disease globally,1 killing approximately six million people each year.2 the world health organization european region has one of the highest proportions of deaths caused by tobacco in the world.3 in the united kingdom ( uk ) , statistics from 2013 estimated that smoking caused around 80,000 deaths in adults aged > 35 years , accounting for 17% of all deaths.4 given the substantial harm caused by smoking , public health policies have focused on tobacco control measures.2 there have been large reductions in the prevalence of smoking on a global scale ; however , owing to population growth the number of smokers has increased.5 according to the world health organization , 21% of the global population aged 15 years smoked tobacco in 2012,6 amounting to 1.1 billion smokers in the world , more than at any time in history.7 in the uk , figures from 2014 reported that one in six adults were smokers , about 10 million , of whom 22% were males and 17% females.8 guidelines in the uk recommend that all smokers have their smoking status recorded at every medical consultation and are offered smoking cessation advice.9 the aim of nicotine replacement therapy ( nrt ) is to reduce both the motivation to smoke and withdrawal symptoms , by temporarily replacing the nicotine from cigarettes , thereby facilitating the transition from cigarettes toward abstinence.10 evidence supports the effectiveness of nrt for smoking cessation , and many guidelines recommend it as a first - line treatment for people seeking pharmacological treatment.1012 in 2011 , nrt was the most common smoking cessation intervention prescribed in england.13 the literature reports conflicting results as to the safety of nrt among high - risk patients . anecdotal evidence has highlighted the incidence of cardiovascular events in patients with unstable coronary syndromes.1417 conversely , a randomized controlled trial ( rct ) conducted by joseph et al,18 found no significant increase in cardiovascular events in two high - risk groups with cardiovascular disease ( cvd ) when nrt patch users were compared with placebo patch users . a systematic review and meta - analysis of adverse events associated with nrt , including 92 rcts and 28 observational studies , concluded that the use of nrt was associated with a variety of side effects , including chest pain and heart palpitations.19 a more recent meta - analysis of rcts reported an elevated risk of less serious cardiovascular events with nrt , but concluded that there was no clear evidence of harm with nrt.20 evidence on the relationship between nrt and cardiovascular events is largely derived from rcts , which frequently have strict eligibility criteria and a tendency to exclude patients at high risk of vascular events and vascular comorbidities . evidence from observational studies is more limited , but one study involving 33,247 patients prescribed nrt , concluded that the use of nrt was not associated with an increase in the risk of myocardial infarction ( mi ) , stroke , or death when used in a real - world routine care setting.21 further real - world effectiveness studies are needed to assess the safety profile of nrt in patients with , or at high risk of , cvd . we hypothesized that patients exposed to nrt ( nrt patients ) are at a higher risk of cvd compared with patients receiving smoking cessation advice only ( smoking cessation advice patients ) . the aim of this study was to compare cvd events , at 4 and 52 weeks respectively , in nrt patients ( individuals attempting smoking cessation with the aid of nrt as any , or a combination of , nasal spray , transdermal patches , inhaler or gum and tablets ) compared with those in smoking cessation advice patients , in a representative uk primary care population . this historical , matched cohort database study comprised a 1-year baseline characterization period and a 1-year outcome evaluation period on either side of an index date . the index date was defined as the time at which patients received either smoking cessation advice only or a first prescription of nrt . data were extracted from the clinical practice research datalink ( cprd ) , the world s largest validated computerized database of anonymized longitudinal medical records for primary care.22,23 at the time of this study , the cprd comprised of ~10.6 million patients from > 590 primary care practices throughout the uk . records are derived from a widely used general practice software system and contain prescribing and coded diagnostic and clinical information as well as information on tests requested , laboratory results , smoking cessation advice recorded by general practitioners ( gps ) using specific read codes,24 and referrals made at or following on from each consultation.25 the study protocol was developed in collaboration with an independent steering committee and approved by the cprd s independent scientific advisory committee ( protocol number 09_096r ) prior to data extraction . the study protocol was publicly registered with the european network of centers for pharmacoepidemiology and pharmacovigilance ( encepp ; registration number encepp / sdpp/4238 ) . eligible patients were current smokers during the baseline year , aged 1875 years , who sought smoking cessation advice from their gp and had at least 1 year of up - to - standard data ( as defined by the cprd ) prior to and following their quit date ( ie , index date ) or up to the time of death if death occurred within the outcome period . the outcome period is defined as up to 4 and/or 52 weeks post index date . end of observation occurred at the practice last collection date , patient transfer out date , outcome diagnosis date , end of the study period ( 4 and/or 52 weeks ) , or the study end date . patients were excluded if they were exposed to nrt or any other pharmacological smoking cessation interventions during the year preceding the index date . patients in the smoking cessation advice group , who received nrt or any other pharmacological smoking cessation interventions during the outcome period , were excluded , as were patients in the nrt group who switched to other ( non - nrt ) pharmacological smoking cessation interventions during the outcome period . switching between different nrt products , or use of multiple nrt products , was permitted . patients in the nrt group received a first prescription for nrt as any , or a combination of , transdermal patches , nasal spray , gum , tablets , or inhaler at the index date . patients who formed the group undertaking smoking cessation unaided by pharmacological interventions , only received smoking cessation advice . this group was defined to reflect , as closely as possible , the patients in the exposed group , with the main exception of note being the decision by their physician to provide smoking cessation advice / education only , rather than a pharmacological intervention , at the index date . the co - primary end points were 1 ) time to diagnosis of ischemic heart disease ( ihd ) and 2 ) time to diagnosis of cerebrovascular disease over a 4-week outcome period ( immediately post index date ) . the 52-week outcome period allowed us to assess cardiovascular events and mortality over a longer time period ; furthermore , it gave us the opportunity to detect any seasonal variations in the prevalence of cardiovascular events . additional secondary end points included the number of consultations for ihd or cerebrovascular disease ( gp consultations , inpatient admissions , and emergency department and outpatient attendances ) and survival time ( all - cause mortality , ihd - related death , cerebrovascular disease - related death ) during the 4-week and 52-week outcome periods . by investigating the number of consultations , we hoped to capture not only new diagnoses , but also a picture of the level of health care resource utilization , such as reviews , monitoring , and acute events , as existing disease potentially worsened . a consultation was taken as a date in the consultation table that was not inpatient , outpatient , or emergency department visit . specific consultation types were identified based on diagnostic ( read codes ) entered on the date corresponding with a code list for ihd / cerebrovascular disease . the cause was inferred on the basis of read codes recorded within a 7-day window of that event . the start of follow - up for end points of interest occurred from the index date of prescribed nrt and the smoking cessation advice date , respectively . time to analyses assessment ran until the earliest of the specific outcome of interest or end of the outcome period . to control potential confounding between comparator groups , patients in the smoking cessation advice group were matched to those in the nrt group on a 2:1 ratio based on sex , age ( 5 years ) , hypertension diagnosis ( on or at any time before the index date ) , cvd diagnosis ( on or at any time before the index date ) , cerebrovascular disease diagnosis ( on or at any time before the index date ) , ihd diagnosis ( on or at any time before the index date ) , diabetes diagnosis ever ( at any time in the records ) , and chronic obstructive pulmonary disease ( copd ) diagnosis ever . further information on the potential confounders evaluated in the study is available in the supplementary material . two - way comparisons between treatment groups using the reduced but matched datasets were carried out making minimal adjustments for other baseline confounders as necessary . the proportion of patients with ihd / cerebrovascular disease diagnosis , proportion of deaths ( all - cause mortality ) , and the number of primary and secondary care consultations due to ihd / cerebrovascular disease were compared using conditional logistic regression . the time to diagnosis of ihd was analyzed using cox proportional hazards model , with times censored at 4 or 52 weeks . the same method was used to analyze the time to diagnosis of cerebrovascular disease , but patients with a prior diagnosis of cerebrovascular disease were excluded from the analysis . the total number of consultations for ihd or cerebrovascular disease during the 4- and 52-week outcome periods was investigated using a conditional negative binomial regression model ( rate ratios ) to obtain estimates of consultation / hospitalization rates relative to the control group . where counts were low , a conditional logistic regression model ( odds ratios ) was used , with the outcome categorized as none versus any consultations . patients with a prior ihd and/or cerebrovascular disease diagnosis were analyzed separately from those with no prior diagnosis of ihd or cerebrovascular disease . all survival times , until death due to ihd , cerebrovascular disease , or any cause , were analyzed using cox proportional hazards regression ( hazard ratios ) with time censored at 4 or 52 weeks . all hazard ratios , odds ratios , and rate ratios were presented as nrt relative to cessation advice only . all analyses were carried out using ibm statistical package for the social sciences ( spss ) statistics version 21 ( ibm spss statistics , feltham , middlesex , uk ) , statistical analysis system version 9.3 ( sas institute , marlow , buckinghamshire , uk ) , and microsoft office excel 2007 ( microsoft corp . , redmond , wa , usa ) . this historical , matched cohort database study comprised a 1-year baseline characterization period and a 1-year outcome evaluation period on either side of an index date . the index date was defined as the time at which patients received either smoking cessation advice only or a first prescription of nrt . data were extracted from the clinical practice research datalink ( cprd ) , the world s largest validated computerized database of anonymized longitudinal medical records for primary care.22,23 at the time of this study , the cprd comprised of ~10.6 million patients from > 590 primary care practices throughout the uk . records are derived from a widely used general practice software system and contain prescribing and coded diagnostic and clinical information as well as information on tests requested , laboratory results , smoking cessation advice recorded by general practitioners ( gps ) using specific read codes,24 and referrals made at or following on from each consultation.25 the study protocol was developed in collaboration with an independent steering committee and approved by the cprd s independent scientific advisory committee ( protocol number 09_096r ) prior to data extraction . the study protocol was publicly registered with the european network of centers for pharmacoepidemiology and pharmacovigilance ( encepp ; registration number encepp / sdpp/4238 ) . eligible patients were current smokers during the baseline year , aged 1875 years , who sought smoking cessation advice from their gp and had at least 1 year of up - to - standard data ( as defined by the cprd ) prior to and following their quit date ( ie , index date ) or up to the time of death if death occurred within the outcome period . the outcome period is defined as up to 4 and/or 52 weeks post index date . end of observation occurred at the practice last collection date , patient transfer out date , outcome diagnosis date , end of the study period ( 4 and/or 52 weeks ) , or the study end date . patients were excluded if they were exposed to nrt or any other pharmacological smoking cessation interventions during the year preceding the index date . patients in the smoking cessation advice group , who received nrt or any other pharmacological smoking cessation interventions during the outcome period , were excluded , as were patients in the nrt group who switched to other ( non - nrt ) pharmacological smoking cessation interventions during the outcome period . switching between different nrt products , or use of multiple nrt products , was permitted . patients in the nrt group received a first prescription for nrt as any , or a combination of , transdermal patches , nasal spray , gum , tablets , or inhaler at the index date . patients who formed the group undertaking smoking cessation unaided by pharmacological interventions , only received smoking cessation advice . this group was defined to reflect , as closely as possible , the patients in the exposed group , with the main exception of note being the decision by their physician to provide smoking cessation advice / education only , rather than a pharmacological intervention , at the index date . the co - primary end points were 1 ) time to diagnosis of ischemic heart disease ( ihd ) and 2 ) time to diagnosis of cerebrovascular disease over a 4-week outcome period ( immediately post index date ) . the 52-week outcome period allowed us to assess cardiovascular events and mortality over a longer time period ; furthermore , it gave us the opportunity to detect any seasonal variations in the prevalence of cardiovascular events . additional secondary end points included the number of consultations for ihd or cerebrovascular disease ( gp consultations , inpatient admissions , and emergency department and outpatient attendances ) and survival time ( all - cause mortality , ihd - related death , cerebrovascular disease - related death ) during the 4-week and 52-week outcome periods . by investigating the number of consultations , we hoped to capture not only new diagnoses , but also a picture of the level of health care resource utilization , such as reviews , monitoring , and acute events , as existing disease potentially worsened . a consultation was taken as a date in the consultation table that was not inpatient , outpatient , or emergency department visit . specific consultation types were identified based on diagnostic ( read codes ) entered on the date corresponding with a code list for ihd / cerebrovascular disease . the cause was inferred on the basis of read codes recorded within a 7-day window of that event . the start of follow - up for end points of interest occurred from the index date of prescribed nrt and the smoking cessation advice date , respectively . the time to analyses assessment ran until the earliest of the specific outcome of interest or end of the outcome period . to control potential confounding between comparator groups , patients in the smoking cessation advice group were matched to those in the nrt group on a 2:1 ratio based on sex , age ( 5 years ) , hypertension diagnosis ( on or at any time before the index date ) , cvd diagnosis ( on or at any time before the index date ) , cerebrovascular disease diagnosis ( on or at any time before the index date ) , ihd diagnosis ( on or at any time before the index date ) , diabetes diagnosis ever ( at any time in the records ) , and chronic obstructive pulmonary disease ( copd ) diagnosis ever . further information on the potential confounders evaluated in the study is available in the supplementary material . two - way comparisons between treatment groups using the reduced but matched datasets were carried out making minimal adjustments for other baseline confounders as necessary . the proportion of patients with ihd / cerebrovascular disease diagnosis , proportion of deaths ( all - cause mortality ) , and the number of primary and secondary care consultations due to ihd / cerebrovascular disease were compared using conditional logistic regression . the time to diagnosis of ihd was analyzed using cox proportional hazards model , with times censored at 4 or 52 weeks . the same method was used to analyze the time to diagnosis of cerebrovascular disease , but patients with a prior diagnosis of cerebrovascular disease were excluded from the analysis . the total number of consultations for ihd or cerebrovascular disease during the 4- and 52-week outcome periods was investigated using a conditional negative binomial regression model ( rate ratios ) to obtain estimates of consultation / hospitalization rates relative to the control group . where counts were low , a conditional logistic regression model ( odds ratios ) was used , with the outcome categorized as none versus any consultations . patients with a prior ihd and/or cerebrovascular disease diagnosis were analyzed separately from those with no prior diagnosis of ihd or cerebrovascular disease . all survival times , until death due to ihd , cerebrovascular disease , or any cause , were analyzed using cox proportional hazards regression ( hazard ratios ) with time censored at 4 or 52 weeks . all hazard ratios , odds ratios , and rate ratios were presented as nrt relative to cessation advice only . all analyses were carried out using ibm statistical package for the social sciences ( spss ) statistics version 21 ( ibm spss statistics , feltham , middlesex , uk ) , statistical analysis system version 9.3 ( sas institute , marlow , buckinghamshire , uk ) , and microsoft office excel 2007 ( microsoft corp . , the unmatched cohort consisted of 57,920 patients , of whom 40,799 received smoking cessation advice , and 17,121 were prescribed nrt . the mean ( standard deviation [ sd ] ) age was 48 ( 11.5 ) years ; 48% were female ( figure s1 ) . after matching for sex , age ( 5 years ) , hypertension diagnosis , cvd diagnosis , cerebrovascular disease diagnosis , ihd diagnosis , diabetes diagnosis , and copd diagnosis , there were a total of 50,214 patients ( figure s2 ) the mean ( sd ) age of the matched cohort was 47 ( 11.2 ) years ; 51% were female ( tables 1 and 2 ) . at 4 weeks post index smoking cessation attempt , there was no difference between nrt patients and smoking cessation advice patients in terms of the primary outcomes of time to first ihd ( unadjusted hazard ratio [ hr ] 95% confidence interval [ ci ] : 1.08 [ 0.562.06 ] ) and cerebrovascular disease diagnosis ( unadjusted hr [ 95% ci ] : 1.00 [ 0.342.93 ] ) or the secondary outcomes survival time and odds of primary and secondary care consultations for ihd or cerebrovascular disease ( table 3 ) . by week 52 , the adjusted hr ( 95% ci ) for time to first diagnosis of ihd was higher for nrt patients compared with smoking cessation advice patients : 1.35 ( 1.031.77 ) . compared with smoking cessation advice patients , at 52 weeks there was also a shorter time to first diagnosis of cerebrovascular disease in nrt patients ( unadjusted hr [ 95% ci ] : 1.54 [ 1.082.19 ] , table 3 ; figure 1 ) and survival time ( adjusted hr [ 95% ci ] : 1.39 [ 1.091.76 ] , figure 1 ) . cerebrovascular disease - related mortality and ihd remained low for both groups throughout the 52-week secondary outcome period . among patients with a prior diagnosis of ihd or cerebrovascular disease , those on nrt therapy had a higher rate of primary or secondary care consultations for ihd or cerebrovascular disease over 52 weeks ( adjusted rate ratio [ rr ] : 1.50 , 95% ci : 1.141.99 ) compared with those receiving smoking cessation advice ( figure 1 ) . this study of a real - world population found no significant differences in the primary end points of time to first ihd and cerebrovascular disease diagnosis at 4 weeks between patients attempting to give up smoking with the assistance of prescribed nrt or via smoking cessation advice alone . however , prescription of nrt to assist smoking cessation ( compared with smoking cessation advice ) was associated with a higher risk of ihd and cerebrovascular disease after 1 year , perhaps owing to cumulative exposure . moreover , patients prescribed nrt to aid their quit attempt had a higher mortality in the following year than those patients supported with smoking cessation advice only . furthermore , we found an increased rate of primary and secondary care consultations for ihd and cerebrovascular disease for patients receiving nrt when a prior diagnosis of ihd or cerebrovascular disease was made . our results of increased ihd and cerebrovascular diagnoses by week 52 for patients taking nrt ( hr : 1.35 , 95% ci : 1.031.77 ) , and low instances of cardiovascular death , broadly support a recent meta - analysis.20 they reported an elevated risk of cardiovascular events with nrt that was driven by less serious events such as tachycardia , but concluded that nrt did not appear to be associated with more serious cardiovascular events . a study using the the health improvement network ( thin ) general practice database , included 33,247 patients taking nrt , and investigated acute mi , acute stroke , and death for each patient during exposed and unexposed time periods . the authors reported that although the incidence increased before exposure and decreased after exposure to nrt in the period of 56 days before and after first nrt prescription , nrt was not associated with an increase in risk of mi , stroke , or death.21 this is in agreement with our data that during short periods of evaluation of 4 weeks , there is no difference in cardiovascular incidence after exposure to nrt . however , our study examined the long - term effects of nrt and found a higher risk of ihd and cerebrovascular disease . nevertheless , gp practices may contribute patient data to both cprd and thin , resulting in overlap of some patients between databases . information about which practices contribute to either database is not publicly available ; therefore , specific methods to identify overlap need to be applied to the data to exclude these patients . furthermore , hubbard et al21 used a within - patient comparison , which could account for the differences observed compared with the current study , as well as looking only at outcomes up to 8 weeks of treatment . a second database study of 663 smokers with acute coronary syndrome that compared nrt versus no nrt reported no differences after 1 year for death , mi , repeat revascularization , or rehospitalization for angina , congestive heart failure , or arrhythmia.26 however , the authors acknowledge , as a limitation of the study , a lack in the number of patients needed to achieve 90% power , which could explain the difference in findings between that and the current study . this highlights the need for larger longer studies to evaluate the long - term effects of nrt . an explanation for the differences in all - cause mortality between treatment groups in the current study could be prescribing practices , with gps preferentially prescribing nrt in patients with other smoking - related diseases , such as lung cancer . these patients could be more likely to be prescribed nrt , as this intervention has been shown to increase the likelihood of quitting by 50%70%.10 an alternative explanation is that the nrt cohort consisted of heavier smokers with a greater illness burden , who therefore had higher all - cause mortality compared with those receiving advice only . however , this study was designed primarily to investigate the safety profile of nrt , and therefore treatment groups were not matched on other diseases . as such , we do not have cause of death information beyond that for ihd and cerebrovascular disease . patients with other diseases were not excluded in order to include as wide a population as possible who were undertaking smoking cessation . nrt is a widely used smoking cessation pharmacotherapy , owing to the low possibility of abuse and potential for dependence.27 bupropion sustained release was the first nonnicotine pharmacological treatment approved for smoking cessation.28 originally designed as an antidepressant in the us , it is the least prescribed smoking cessation treatment.11,13 there is evidence that bupropion is more effective than a placebo , but only weak evidence to suggest superiority over nrt.12 however , there is evidence of a potentially protective effect for the risk of major cardiovascular events with bupropion , although the reason for this is not well understood.20 varenicline , a competitive nicotine receptor antagonist , has been available as a smoking cessation treatment since 2006.29 it has been suggested that varenicline may be more effective than nrt.29 our study adds real - world data to the existing evidence from rcts on the safety profile of nrt in patients with , or at a higher risk of , cardiovascular events . our findings of increased cardiovascular events with nrt in high - risk patients indicate that further investigation is needed into both the safety of nrt and alternatives to nrt treatment in this group of smokers . strengths of the current study include the use of data on real - life patients from a large , high - quality source , the cprd , which is well described and has previously been used in respiratory research.23 furthermore , compared with many of the rcts investigating nrt as a smoking cessation treatment , this study has a larger patient population and longer follow - up period . firstly , there is lack of data on over - the - counter ( otc ) purchases , particularly in light of the fact that most nrt in the uk is used without advice , and purchased otc rather than by prescription.30 however , there is evidence that most patients who use otc nrt do not exceed the recommended time of 12 weeks,31 allowing us to infer that our secondary outcome period of 52 weeks is less likely to be affected by potential otc use . on the other hand , the results indicate that the potential excess use of otc nrt in the control group may be counteracting the observed difference between the groups . a further consideration is that patients taking nrt could potentially be exposed to elevated nicotine levels if they continue to smoke while taking nrt . however , owing to the flat dose cardiovascular response relation for nicotine , the effects of cigarette smoking in addition to nrt are likely to be similar to those of smoking alone.32 data on hospitalizations ( emergency department attendance , inpatient admissions , and outpatient attendance ) were limited because hospital episode statistics linked data were not available ; as a result , hospitalizations were ascertained using information from gp records . there was no matching or adjustment for the severity of the smoker ( cigarettes per day , pack years or fagerstrom test ) , and thus , there is the potential that those in the nrt group were heavier smokers than those in the advice group . we did not analyze the time period of nrt use , and patients were only required to initiate on nrt to be included in the nrt group of the study . our data did not provide information on the utilization of nrt , but only that it was prescribed , and we can only assume that nrt patients took the treatment as intended , making this an intention to treat analysis . we also lack information on details such as family history , a potentially influential factor as to whether a patient decides to seek cessation treatment with nrt . furthermore , we do not know the rates of smoking cessation in each group ; we can speculate that it was higher in the nrt group ; however , this might not be the case . compliance with nrt is likely to be higher in the prescription group as patients will have a clearer understanding of the benefit of the therapy owing to the advice of health care professionals . the charlson comorbidity index ( cci ) score was higher in the nrt group , which could lead us to assume that increased morbidity initially drove patients to seek treatment with nrt , and could therefore explain the worsened end points . confounding by indication may affect our results ; gps may assess patients with indications of poorer health , such as indicated by a high cci score , or those with a higher risk of future poor outcomes , and prescribe nrt rather than advice . if higher risk patients were more likely to be given nrt , we may have overestimated the effect of nrt on outcomes . confounding by indication is more of a concern in studies such as ours where initiators of treatment are compared with non - initiators as supported by schneeweiss et al.33 a comparison between similar treatments would minimize this bias . the more similar the compared treatments are , the less potential there is for unmeasured confounding . matched analysis was conducted to achieve balance of covariates between the cohorts . adjusted analysis was carried out to minimize confounding . however , as we did not randomly assign patients to either nrt or advice , some bias may remain . both smoking and nicotine treatment have been found to increase heart rate and blood pressure.3437 the hemodynamic effects of smoking have been linked to nicotine , with heart rate found to increase with intravenous nicotine , nicotine nasal sprays , and nicotine chewing gum.3840 nicotine was found to affect coronary artery constriction even at doses as low as 4 mg.41 these effects cause an increase in myocardial work and oxygen demand and result in impaired blood flow and oxygen supply to the heart . however , transdermal nicotine was found to have a lesser acute hemodynamic effect than smoking.42 although there are not much data available on the effect of transdermal nicotine on coronary blood flow , benowitz et al42 suggest that transdermal nicotine in smoking cessation treatment of patients with coronary heart disease is likely to be safer than cigarette smoking . further observational research is needed to provide insights into the effect of nrt on cardiovascular events . data on nicotine exposure , which could be extracted through linked pharmacy and gp data , would be insightful . extending the outcome period to longer than 1 year would increase the event numbers and allow a greater window to observe any potential associations . it would also be beneficial to obtain hospitalization data to ascertain whether increases in consultation rates are limited to gp consultations or also applicable to hospitalizations . although treatment with nrt during a short period ( 4 weeks ) does not appear to have an impact on cardiovascular risks , a longer follow - up period of 52 weeks resulted in an increase in cardiovascular events for patients prescribed nrt , compared with those receiving smoking cessation advice only . in view of the ongoing global public health risk of cigarette smoking , there is an urgent need to investigate the safest treatments available for patients attempting smoking cessation . owing to the potential for confounding demographic and comorbid factors , initial analysis identified key baseline confounders , and the outcome analyses utilized appropriate statistical methods ( logistical regression and matching ) , to minimize confounding . potential confounders examined at , or closest to , the index date were
agesexheightbody mass index ( bmi ) body mass index ( bmi ) potential confounders that were examined irrespective of when they occurred relative to the index date were confounding diagnoses including
diabetescopdrhinitishypertensioncardiovascular diseaseischemic heart disease ( ihd , subset of cardiovascular disease)cerebrovascular disease ( subset of cardiovascular disease)angina ( subset of ihd)myocardial infarction ( subset of ihd ) cardiovascular disease ischemic heart disease ( ihd , subset of cardiovascular disease ) cerebrovascular disease ( subset of cardiovascular disease ) angina ( subset of ihd ) myocardial infarction ( subset of ihd ) other important unrelated comorbidities were expressed using the cci . this was calculated on the basis of an algorithm of weighted read code diagnosis codes in the year prior to the index date . potential confounders that were examined the year prior to the index date included cholesterol measurements
blood pressuredrug therapiesgeneral practice consultationsoutpatient attendancesinpatient admissionsaccident and emergency admissions general practice consultations outpatient attendances accident and emergency admissions patient flow diagram for the unmatched nrt and smoking cessation advice cohorts . owing to the potential for confounding demographic and comorbid factors , initial analysis identified key baseline confounders , and the outcome analyses utilized appropriate statistical methods ( logistical regression and matching ) , to minimize confounding . potential confounders examined at , or closest to , the index date were
agesexheightbody mass index ( bmi ) body mass index ( bmi ) potential confounders that were examined irrespective of when they occurred relative to the index date were confounding diagnoses including
diabetescopdrhinitishypertensioncardiovascular diseaseischemic heart disease ( ihd , subset of cardiovascular disease)cerebrovascular disease ( subset of cardiovascular disease)angina ( subset of ihd)myocardial infarction ( subset of ihd ) cardiovascular disease ischemic heart disease ( ihd , subset of cardiovascular disease ) cerebrovascular disease ( subset of cardiovascular disease ) angina ( subset of ihd ) myocardial infarction ( subset of ihd ) other important unrelated comorbidities were expressed using the cci . this was calculated on the basis of an algorithm of weighted read code diagnosis codes in the year prior to the index date . potential confounders that were examined the year prior to the index date included cholesterol measurements
blood pressuredrug therapiesgeneral practice consultationsoutpatient attendancesinpatient admissionsaccident and emergency admissions general practice consultations outpatient attendances accident and emergency admissions patient flow diagram for the unmatched nrt and smoking cessation advice cohorts . | backgroundprevious research suggests exposure to nicotine replacement therapy ( nrt ) may be associated with an increased risk of cardiovascular disease ( cvd).methodsusing data from the united kingdom s clinical practice research datalink , this study aimed to evaluate cvd events and survival among individuals who attempted smoking cessation with the support of nrt compared with those aided by smoking cessation advice only .
we studied cvd outcomes over 4 and 52 weeks in 50,214 smokers attempting to quit
33,476 supported by smoking cessation advice and 16,738 with the support of nrt prescribed by their primary care physician .
patients were matched ( 2 smoking cessation advice patients:1 nrt patient ) on demographic and clinical characteristics during a baseline year preceding their quit attempt .
cox proportional hazard regression , conditional negative binomial regression model , and conditional logistic regression were used to analyze data.resultsmean ( standard deviation ) population age was 47 ( 11.2 ) years ; 51% were females .
time to first diagnosis of ischemic heart disease ( ihd ) among nrt and smoking cessation advice patients was similar within the first 4 weeks , but shorter for nrt patients over 52 weeks ( hazard ratio [ hr ] : 1.35 , 95% confidence interval [ ci ] : 1.031.77 ) .
a similar trend was observed for cerebrovascular disease ( hr : 1.54 , 95% ci : 1.082.19 ) .
nrt patients with a prior diagnosis of ihd or cerebrovascular disease had a higher rate of primary or secondary care consultations for ihd or cerebrovascular disease by 52 weeks ( rate ratio : 1.50 , 95% ci : 1.141.99 ) .
patients prescribed nrt had a shorter survival time over 52 weeks , compared with those receiving advice only ( hr : 1.39 , 95% ci : 1.091.76).conclusionour findings suggest that treatment with nrt over 4 weeks does not appear to have an impact on cardiovascular risks . however ,
a longer follow - up period of 52 weeks resulted in an increase in cardiovascular events for patients prescribed nrt , compared with those receiving smoking cessation advice only . |
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epidemiological studies show that higher levels of physical activity are associated with lower frequency of diabetes , coronary heart disease , stroke , colorectal cancer and depression . current recommendations recognise that there is a clear link between physical activity and chronic disease and advise that all adults should take at least 150 min of moderate physical activity per week with daily activity . health services increasingly emphasise healthy ageing , aiming to prevent disease and reduce the impact of long - term conditions in later life . a recent meta - analysis of randomised trials found that brief interventions in primary care could result in one additional person achieving recommended physical activity levels for each 12 persons exposed to the intervention , with the effect being maintained over 12 months . brief exercise advice has a small effect on increasing physical activity such a small effect could be important if carried out on a large population of patients . the potential long - term effects of interventions to promote physical activity in primary care are not known . existing reviews have included studies up to 2 years duration with behaviour change and physical fitness as outcomes . evidence that increased physical activity may reduce the incidence of diabetes is drawn from studies that included high risk individuals with impaired glucose tolerance or pre - diabetes . the present research therefore aimed to evaluate the potential long term health outcomes and cost effectiveness of a universal strategy to promote physical activity in primary care . the research specifically aimed to determine whether a low - cost intervention with a limited intervention effect size , such as that evidenced by the recent meta - analysis on the effectiveness of such interventions , would prove cost - effective for implementation across the general population through primary health care services . a markov model was employed to implement a cost - utility analysis of a universal strategy to promote physical activity in the general population registered in primary care , comparing a brief intervention to promote physical activity with standard care in which there is no systematic approach to promote physical activity . a simplified diagram of the markov model is shown in fig . 1 . healthy subjects , referred to as at risk , may develop one of the disease states of interest , including diabetes , coronary heart disease , stroke or colorectal cancer . participants in one of these disease states may develop a second , third or fourth disease , giving 16 single- or multi - disease states , consistent with the frequent occurrence of multimorbidity as a driver of health care utilisation in primary care populations . participants in each state were allowed to progress to depression , with each state divided into states representing not depressed and depressed. depression was associated with its own decrement in utility as well as its own rate of health care utilisation . depression was included because it occurs frequently in chronic illness and is associated with higher health care costs for a given chronic illness . there were therefore 32 states in the model that represented all potential combinations of the included diseases and depression . the perspective of the model is that of health care services and only health care costs were included . 1schematic diagram of markov model . in order to simplify the diagram , the 32 incidence transitions and 16 mortality transitions depressed and not depressed. chd coronary heart disease , dm type 2 diabetes mellitus schematic diagram of markov model . in order to simplify the diagram , the 32 incidence transitions and 16 mortality transitions are not represented in full . depressed and not depressed. chd coronary heart disease , dm type 2 diabetes mellitus data to populate the model were derived from a large cohort of participants drawn from the general population registered with the general practice research database ( gprd ) . the gprd includes electronic health records of participants registered with approximately 600 uk family practices . gprd data were used to estimate the incidence of each state included in the model ; the mortality in each state ; and the health care utilisation and costs associated with each state . the use of fully anonymised gprd data was approved by the mhra independent scientific advisory committee ( ref . the gprd cohort was drawn from all family practices that were continuously contributing data to the gprd between 1 january 2004 and 30 october 2010 . as colorectal cancer cases were less numerous , we made the assumptions that the incidence of colorectal cancer was the same in participants with and without cardiovascular comorbidity , and the incidence of cardiovascular comorbidity was the same in participants with and without colorectal cancer . incidence and mortality rates were similar to those reported from gprd previously [ 1315 ] . the prevalence of depression was estimated for each state in the model based on gprd data . in the model , we assumed that depression was not associated with mortality at any given level of physical disease . health care utilisation was estimated for each state from gprd records , including utilisation of primary care ( family practice consultations , telephone consultations , home visits and emergency and out - of - hours consultations ) , secondary care ( including hospital admissions , outpatient visits , day case visits and emergency visits ) and prescriptions . the annual costs associated with each state were estimated by multiplying the health care utilisation associated with the state by the costs of each unit of health care , which were obtained from standard reference sources for 2010 ( table 1 ) . prescription costs were obtained by linking the multilex drug code for each prescription record in gprd with the prescription cost . the empirical mean ( standard deviation ) of participant level costs were estimated by age group , gender , depression status and condition . as the number of colorectal cancer cases was too small to subdivide into multi - disease states , health care utilisation was estimated for all colorectal cancer cases together.table 1sources of data for modeldatanumber / valuesourcestratificationcommentsbase population299,912gprdgender , 1-year age groupparticipants with prevalent disease were excluded . age range 30100 yearsmodel states33 statesstratified by gender , 1-year age group.includes at risk , diabetes , chd , stroke , colorectal cancer , depression and all combinations , deadincidence of states32 incidence ratesgprdgender , 10-year age groupbeta - binomial distribution used to estimate transition probabilities ; incidence assumed independent of depression ; note that the same state may be reached by more than one transition ( e.g. chd in diabetes , diabetes in chd)mortality of states16 mortality ratesgprdgender , 10-year age groupbeta - binomial distribution used to estimate transition probabilities ; mortality assumed independent of depressiondepression prevalence16 depression prevalence ratesgprdgender , 10-year age groupbeta - binomial distribution used to estimate transition probabilitieshealth care utilizationutilisation rates estimated for 32 statesgprdgender , 10-year age group , depression statusutilisation included primary care consultations ( including at general practice , home , telephone and out of hours or emergency ) , secondary care ( including inpatient , outpatient , day case and emergency ) and prescription number and contentunit costs of health care utilization primary care consultation35pssru gamma distribution used to sample costs emergency / out of hours consultation35pssru home visit117pssru telephone consultation21pssru prescription unit costsvariablefdbe unit price linked by multilex code to gprd prescription . single pack price assumed secondary care inpatient episodes493pssru outpatient visits189pssru day case visits143pssru emergency visits110pssru utility valuesutility decrement age 43 years0.828sullivan et al . stratified by single year of agebeta function employed to sample utility values per year increase in age0.00029 diabetes0.0621 chd0.0557 stroke0.1009 colorectal cancer0.0378 depression0.1302 no . of chronic conditions 20.0615 no . of chronic conditions 30.0667 no . of chronic conditions 40.0433 no . of chronic conditions 50.0287
gprd general practice research database , pssru personal social services research unit , fdbe first databank europe sources of data for model
gprd general practice research database , pssru personal social services research unit , fdbe first databank europe the probabilistic markov model was estimated by cohort simulation , implemented through a program written in r software . after removing participants with prevalent disease , there were 262,704 healthy participants that entered the initial state of the model , based on the distribution observed in gprd , including 49 % men . there were 37 % aged < 45 years and 42 % were aged 4564 years . all simulations were stratified by single year of age with the initial population aging by 1 year per cycle . outcomes and costs were compared for intervention and standard care over 70 annual cycles , this allowed the entire cohort to progress either to death or to reach age 100 and exit the model . annual transition probabilities for the model were obtained by sampling from the beta - binomial distribution , using gprd data as inputs ( table 1 ) . utilities for each state were obtained from data published in a compendium of values ( table 1 ) . utility values for each state were stratified by single year of age but were the same for men and women . the costs of each state were sampled from the gamma distribution with the mean value from gprd , by 10-year age group , sex , condition and depression status , as the empirical input . total costs and quality adjusted life years ( qalys ) were obtained by summing across the 70 cycles of the model included in each simulation . mean costs , and the 95 % range , were obtained from the data for 2,000 simulations . incremental costs and qalys were obtained as the difference between intervention and standard care scenarios . costs and qalys were discounted using a rate of 3.5 % , but qalys were also discounted at a rate of 1.5 % as a sensitivity analysis . net health benefits ( nhb ) , at a threshold value of 30,000 per qaly , were calculated as the difference between the increment in qalys and the increment in costs divided by the threshold value of cost per qaly . the model was implemented with a half cycle correction for the estimation of qalys and costs . the intervention was assumed to modify only the incidence of disease in healthy participants at risk . the effect of intervention was estimated as a potential impact fraction ( pif ) , following cobiac et al . . the pif provides a means of estimating the extent to which a change in risk factor exposure is associated with a proportionate decline in the likelihood of an individual developing a disease outcome of interest . the pif was estimated from three sources of data : ( 1 ) the effect of brief interventions in primary care on physical activity levels . estimated that the number needed to treat for an additional sedentary subject to become active was approximately 12 with an odds ratio of 1.42 ( 95 % confidence interval 1.171.73 ) and an event rate in control participants of 26 % ( 507/1924 ) ; ( 2 ) data for the distribution of physical activity in the general population , by 10-year age group and sex , were obtained from the health survey for england 2008 ; ( 3 ) relative risks associating inactivity , or insufficient activity , with the four study disease outcomes ( diabetes , coronary heart disease , stroke and colorectal cancer ) were obtained from the who study comparative quantification of health risks . consistent with the who study we did not include a possible effect of intervention on depression prevalence because the evidence is inconsistent and disputed . , with estimates being derived , in each cycle , for single years of age and sex based on the empirical estimates from the three data sources . the intervention was modelled as being maintained for either 5 or 10 years . in the absence of evidence for the time course of intervention effects , the same estimates were used to model the intervention effect in each of the first five or ten cycles of the model as appropriate . the cost of the intervention was modelled as a fixed cost per person year depending on their physical activity level . the population at risk was divided into those that were physically active and those that were physically inactive or who took insufficient physical activity , based on the distribution observed in the health survey for england 2008 . in the population that was not sufficiently physically active the intervention cost , in the base case , was modelled as being equivalent to the cost of one family practice consultation per person year ( 35 ) . in the population that was physically active , the cost of screening questions to evaluate physical activity levels was 20 % of one family practice consultation per year . most [ brief physical activity interventions in primary care ] included written materials and two or more sessions of advice or counselling on physical activity , delivered face to face . as sensitivity analyses , therefore , we also considered costs of intervention in inactive individuals equivalent to two family practice consultations per year ( 70 ) and 20 % of a family practice consultation per year ( 7 ) . a markov model was employed to implement a cost - utility analysis of a universal strategy to promote physical activity in the general population registered in primary care , comparing a brief intervention to promote physical activity with standard care in which there is no systematic approach to promote physical activity . a simplified diagram of the markov model is shown in fig . 1 . healthy subjects , referred to as at risk , may develop one of the disease states of interest , including diabetes , coronary heart disease , stroke or colorectal cancer . participants in one of these disease states may develop a second , third or fourth disease , giving 16 single- or multi - disease states , consistent with the frequent occurrence of multimorbidity as a driver of health care utilisation in primary care populations . participants in each state were allowed to progress to depression , with each state divided into states representing not depressed and depressed. depression was associated with its own decrement in utility as well as its own rate of health care utilisation . depression was included because it occurs frequently in chronic illness and is associated with higher health care costs for a given chronic illness . there were therefore 32 states in the model that represented all potential combinations of the included diseases and depression . the perspective of the model is that of health care services and only health care costs were included . 1schematic diagram of markov model . in order to simplify the diagram , the 32 incidence transitions and 16 mortality transitions depressed and not depressed. chd coronary heart disease , dm type 2 diabetes mellitus schematic diagram of markov model . in order to simplify the diagram , the 32 incidence transitions and 16 mortality transitions are not represented in full . data to populate the model were derived from a large cohort of participants drawn from the general population registered with the general practice research database ( gprd ) . the gprd includes electronic health records of participants registered with approximately 600 uk family practices . gprd data were used to estimate the incidence of each state included in the model ; the mortality in each state ; and the health care utilisation and costs associated with each state . the use of fully anonymised gprd data was approved by the mhra independent scientific advisory committee ( ref . 09 - 085 ) . the gprd cohort was drawn from all family practices that were continuously contributing data to the gprd between 1 january 2004 and 30 october 2010 . as colorectal cancer cases were less numerous , we made the assumptions that the incidence of colorectal cancer was the same in participants with and without cardiovascular comorbidity , and the incidence of cardiovascular comorbidity was the same in participants with and without colorectal cancer . incidence and mortality rates were similar to those reported from gprd previously [ 1315 ] . the prevalence of depression was estimated for each state in the model based on gprd data . in the model , we assumed that depression was not associated with mortality at any given level of physical disease . health care utilisation was estimated for each state from gprd records , including utilisation of primary care ( family practice consultations , telephone consultations , home visits and emergency and out - of - hours consultations ) , secondary care ( including hospital admissions , outpatient visits , day case visits and emergency visits ) and prescriptions . the annual costs associated with each state were estimated by multiplying the health care utilisation associated with the state by the costs of each unit of health care , which were obtained from standard reference sources for 2010 ( table 1 ) . prescription costs were obtained by linking the multilex drug code for each prescription record in gprd with the prescription cost . the empirical mean ( standard deviation ) of participant level costs were estimated by age group , gender , depression status and condition . as the number of colorectal cancer cases was too small to subdivide into multi - disease states , health care utilisation was estimated for all colorectal cancer cases together.table 1sources of data for modeldatanumber / valuesourcestratificationcommentsbase population299,912gprdgender , 1-year age groupparticipants with prevalent disease were excluded . age range 30100 yearsmodel states33 statesstratified by gender , 1-year age group.includes at risk , diabetes , chd , stroke , colorectal cancer , depression and all combinations , deadincidence of states32 incidence ratesgprdgender , 10-year age groupbeta - binomial distribution used to estimate transition probabilities ; incidence assumed independent of depression ; note that the same state may be reached by more than one transition ( e.g. chd in diabetes , diabetes in chd)mortality of states16 mortality ratesgprdgender , 10-year age groupbeta - binomial distribution used to estimate transition probabilities ; mortality assumed independent of depressiondepression prevalence16 depression prevalence ratesgprdgender , 10-year age groupbeta - binomial distribution used to estimate transition probabilitieshealth care utilizationutilisation rates estimated for 32 statesgprdgender , 10-year age group , depression statusutilisation included primary care consultations ( including at general practice , home , telephone and out of hours or emergency ) , secondary care ( including inpatient , outpatient , day case and emergency ) and prescription number and contentunit costs of health care utilization primary care consultation35pssru gamma distribution used to sample costs emergency / out of hours consultation35pssru home visit117pssru telephone consultation21pssru prescription unit costsvariablefdbe unit price linked by multilex code to gprd prescription . single pack price assumed secondary care inpatient episodes493pssru outpatient visits189pssru day case visits143pssru emergency visits110pssru utility valuesutility decrement age 43 years0.828sullivan et al . stratified by single year of agebeta function employed to sample utility values per year increase in age0.00029 diabetes0.0621 chd0.0557 stroke0.1009 colorectal cancer0.0378 depression0.1302 no . of chronic conditions 20.0615 no . of chronic conditions 30.0667 no . of chronic conditions 40.0433 no . of chronic conditions 50.0287
gprd general practice research database , pssru personal social services research unit , fdbe first databank europe sources of data for model
gprd general practice research database , pssru personal social services research unit , fdbe first databank europe the probabilistic markov model was estimated by cohort simulation , implemented through a program written in r software . after removing participants with prevalent disease , there were 262,704 healthy participants that entered the initial state of the model , based on the distribution observed in gprd , including 49 % men . there were 37 % aged < 45 years and 42 % were aged 4564 years . all simulations were stratified by single year of age with the initial population aging by 1 year per cycle . outcomes and costs were compared for intervention and standard care over 70 annual cycles , this allowed the entire cohort to progress either to death or to reach age 100 and exit the model . annual transition probabilities for the model were obtained by sampling from the beta - binomial distribution , using gprd data as inputs ( table 1 ) . utilities for each state were obtained from data published in a compendium of values ( table 1 ) . utility values for each state were stratified by single year of age but were the same for men and women . the costs of each state were sampled from the gamma distribution with the mean value from gprd , by 10-year age group , sex , condition and depression status , as the empirical input . total costs and quality adjusted life years ( qalys ) were obtained by summing across the 70 cycles of the model included in each simulation . mean costs , and the 95 % range , were obtained from the data for 2,000 simulations . incremental costs and qalys were obtained as the difference between intervention and standard care scenarios . costs and qalys were discounted using a rate of 3.5 % , but qalys were also discounted at a rate of 1.5 % as a sensitivity analysis . net health benefits ( nhb ) , at a threshold value of 30,000 per qaly , were calculated as the difference between the increment in qalys and the increment in costs divided by the threshold value of cost per qaly . the model was implemented with a half cycle correction for the estimation of qalys and costs . the intervention was assumed to modify only the incidence of disease in healthy participants at risk . the effect of intervention was estimated as a potential impact fraction ( pif ) , following cobiac et al . . the pif provides a means of estimating the extent to which a change in risk factor exposure is associated with a proportionate decline in the likelihood of an individual developing a disease outcome of interest . the pif was estimated from three sources of data : ( 1 ) the effect of brief interventions in primary care on physical activity levels . estimated that the number needed to treat for an additional sedentary subject to become active was approximately 12 with an odds ratio of 1.42 ( 95 % confidence interval 1.171.73 ) and an event rate in control participants of 26 % ( 507/1924 ) ; ( 2 ) data for the distribution of physical activity in the general population , by 10-year age group and sex , were obtained from the health survey for england 2008 ; ( 3 ) relative risks associating inactivity , or insufficient activity , with the four study disease outcomes ( diabetes , coronary heart disease , stroke and colorectal cancer ) were obtained from the who study comparative quantification of health risks . consistent with the who study we did not include a possible effect of intervention on depression prevalence because the evidence is inconsistent and disputed . the pif was estimated as outlined by cobiac et al . , with estimates being derived , in each cycle , for single years of age and sex based on the empirical estimates from the three data sources . the intervention was modelled as being maintained for either 5 or 10 years . in the absence of evidence for the time course of intervention effects , the same estimates were used to model the intervention effect in each of the first five or ten cycles of the model as appropriate . the cost of the intervention was modelled as a fixed cost per person year depending on their physical activity level . the population at risk was divided into those that were physically active and those that were physically inactive or who took insufficient physical activity , based on the distribution observed in the health survey for england 2008 . in the population that was not sufficiently physically active the intervention cost , in the base case , was modelled as being equivalent to the cost of one family practice consultation per person year ( 35 ) . in the population that was physically active , the cost of screening questions to evaluate physical activity levels was 20 % of one family practice consultation per year . most [ brief physical activity interventions in primary care ] included written materials and two or more sessions of advice or counselling on physical activity , delivered face to face . as sensitivity analyses , therefore , we also considered costs of intervention in inactive individuals equivalent to two family practice consultations per year ( 70 ) and 20 % of a family practice consultation per year ( 7 ) . the estimated values for intervention effects , derived from the estimated pifs , are shown in table 2 . the figures are the mean , standard deviation and range of values for 2,000 simulations , with each value representing the mean across all ages in the first cycle of each simulation . an intervention effect of 0.95 indicates that the incidence of the condition of interest will , on average , be 5 % lower with intervention.table 2estimated values for intervention effects derived from potential impact fractions ( pifs)malefemalediabetes mellitus mean ( sd)0.966 ( 0.010)0.967 ( 0.010 ) range0.9190.9970.9340.998coronary heart disease mean ( sd)0.949 ( 0.015)0.951 ( 0.015 ) range0.8921.0030.9051.001stroke mean ( sd)0.968 ( 0.010)0.969 ( 0.010 ) range0.9321.0010.9301.001colorectal cancer mean ( sd)0.959 ( 0.012)0.961 ( 0.012 ) range0.9131.0020.9151.004figures are the mean ( sd ) and range for 2,000 simulations for values in the first cycle of the model . values may be interpreted as relative risks estimated values for intervention effects derived from potential impact fractions ( pifs ) figures are the mean ( sd ) and range for 2,000 simulations for values in the first cycle of the model . values may be interpreted as relative risks there were 262,704 healthy participants , with the same age and gender distribution as in gprd , who entered the model in each simulation ( table 3 , fig . 1 ) . for an intervention lasting 5 years , there was an increase in life years lived without physical disease of 52.1 ( 10.9 to 115.3 ) per 1,000 participants entering the model . figure 2 shows the time - course of changes in single- and multiple - morbidity following the start of intervention . as expected , single morbidities were reduced during the intervention period , but the reduction in single - morbidities persisted , while diminishing , following the end of intervention . there was greater than 95 % probability that single morbidities were reduced from the second to the 11th year following the start of intervention . a reduction in dual morbidities reached its maximum , approximately 20 years following the start of intervention , while a reduction in triple morbidity reached a maximum approximately 30 years following the start of intervention . there was an 87.6 % probability that life years lived with single morbidities was reduced overall ( table 3 ) . the equivalent figure for dual morbidities was 73.3 % ; triple morbidities , 58.7 % ; and quadruple morbidities , 49.6 % . although the intervention was modelled to have no direct effect on depression prevalence , overall life years with depression tended to be reduced because of the empirical observation that depression prevalence was higher in individuals with morbidity.table 3health outcomes and cost - effectiveness of a physical activity intervention in a population of 262,704 healthy participantsintervention duration5 years10 yearsdifference ( intervention - standard care)probability ( % ) difference ( intervention - standard care)probability ( % ) number entering intervention262,704262,704life years lived without disease ( per 1,000 )
52.1 ( 10.9 to 115.3)91.9
102.3 ( 42.3 to 163.7)99.7
life years lived with physical morbidity ( per 1,000 )
single condition34.1 ( 82.3 to 13.7)87.6
69.6 ( 119.3 to 21.6)98.7
dual conditions8.3 ( 29.1 to 12.6)73.3
16.1 ( 38.8 to 5.7)88.9
triple conditions0.96 ( 8.0 to 5.7)58.7
2.0 ( 9.0 to 5.0)69.8
quadruple conditions0.01 ( 1.4 to 1.4)49.6
0.1 ( 1.5 to 1.3)53.6
life years lived with depression ( per 1,000 )
2.8 ( 17.9 to 11.8)61.9
6.4 ( 20.4 to 7.9)76.6
total life years ( per 1,000 )
8.9 ( 35.6 to 52.3)62.7
14.6 ( 29.2 to 59.3)71.1
figures represent mean and 95 % range of 2,000 simulations
per 1,000 healthy participants entering model
probability measure is higher with intervention
probability measure is lower with interventionfig . 2changes over time in levels of single and multiple morbidity following an intervention of 5 years duration . data represent the mean difference between intervention and standard care by year ( color figure online ) health outcomes and cost - effectiveness of a physical activity intervention in a population of 262,704 healthy participants figures represent mean and 95 % range of 2,000 simulations
per 1,000 healthy participants entering model
probability measure is higher with intervention
probability measure is lower with intervention changes over time in levels of single and multiple morbidity following an intervention of 5 years duration . data represent the mean difference between intervention and standard care by year ( color figure online ) when the intervention was maintained for 10 years , life years lived free from disease were increased by 102.3 per 1,000 participants entering the intervention ( probability increased 99.7 % ) . life years lived with single morbidities were reduced by 69.6 per 1,000 , probability 98.7 % ; dual morbidities were reduced by 16.1 per 1,000 , probability , 88.9 % . there was a 69.8 % probability that life years with triple morbidity were reduced and 76.6 % probability that life years with depression were reduced . there was only weak evidence that total life years were increased after either 5 or 10 years intervention ( table 3 ) . table 4 shows the changes in costs and qalys associated with the intervention . with an intervention of 5 years duration , the discounted incremental costs of intervention were 97,572 per 1,000 participants entering intervention , with 86 % of the cost attributable to intervention in participants who were inactive or insufficiently active . approximately 14 % of the cost of intervention was attributable to confirming the physical activity status of participants that were already active . the costs of non - intervention health care utilisation tended to be reduced through intervention by 16,818 ( probability reduced 63.7 % ) . thus the overall total incremental costs under intervention were 80,744 ( probability increased 95.5 % ) .table 4health outcomes and cost - utility of a physical activity intervention in a population of 262,704 healthy participantsintervention duration5 years10 yearsdifference ( intervention - standard care)probability ( % ) difference ( intervention - standard care)probability ( % ) number entering intervention262,704262,704intervention costs in physically active ( per 1,000)13,995 ( 13,989 to 14,001)100.024,018 ( 24,003 to 24,033)100.0intervention costs in physically inactive ( per 1,000)83,567 ( 83,531 to 83,601)100.0152,210 ( 152,110 to 152,306)100.0total intervention costs ( per 1,000)97,572 ( 97,521 to 97,602)100.0176,228 ( 176,113 to 176,340)100.0incremental costs of non - intervention health care utilisation ( per 1,000)16,818 ( 94,269 to 60,747)63.7
31,760 ( 109,077 to 47,599)74.3
incremental total costs ( per 1,000 )
80,744 ( 3,326 to 158,251)95.5144,469 ( 67,103 to 223,843)99.9incremental qalys ( discounted 3.5 % ) ( per 1,000)5.9 ( 8.2 to 19.7)75.79.8 ( 4.6 to 23.6)87.3incremental qalys ( discounted 1.5 % ) ( per 1,000)8.6 ( 14.7 to 32.4)72.714.9 ( 8.7 to 38.5)85.1net health benefits ( qalys per 1,000)3.2 ( 11.1 to 16.9)64.75.0 ( 9.5 to 19.3)72.4probability cost effective at 30,000 per qaly ( % ) 64.772.4figures represent mean and 95 % range of 2,000 simulations . qaly quality - adjusted life year , chd coronary heart disease
per 1,000 healthy participants entering model
probability reduced health outcomes and cost - utility of a physical activity intervention in a population of 262,704 healthy participants figures represent mean and 95 % range of 2,000 simulations . qaly quality - adjusted life year , chd coronary heart disease
per 1,000 healthy participants entering model the discounted incremental qalys associated with intervention were 5.9 per 1,000 participants entering intervention ( table 4 ) . the probability that the intervention would be cost - effective at a threshold of 30,000 per qaly was 64.7 % . for an intervention maintained for 10 years the incremental qalys associated with intervention were 14.9 per 1,000 ( probability increased 85.1 % ) . the net health benefits at a threshold of 30,000 per qaly were 5.0 per 1,000 . the probability of the intervention being cost - effective at the same threshold was 72.4 % . figure 3 presents a cost - effectiveness plane , in which incremental costs are plotted against incremental qalys for each of the 2,000 simulations . figure 3 also presents a cost - effectiveness acceptability curve , in which the probability of the intervention being cost - effective was estimated at different thresholds values of cost per qaly . an intervention continuing for either 5 or 10 years did not achieve more than an 80 % probability of being cost - effective , except at longer intervention duration and high threshold values of cost per qaly , because there were appreciable numbers of simulations in which intervention was associated with no increase in qalys.fig . 3
left panel cost - effectiveness plane showing results of 2,000 simulations with 10 years intervention . right panel cost - effectiveness acceptability curves for 5 ( blue ) and 10 ( red ) years intervention ( color figure online )
left panel cost - effectiveness plane showing results of 2,000 simulations with 10 years intervention . right panel cost - effectiveness acceptability curves for 5 ( blue ) and 10 ( red ) years intervention ( color figure online ) sensitivity analyses were implemented to explore the effects of varying the unit costs of intervention and varying the discount rate . as expected , increasing the unit costs of intervention necessarily made the intervention less cost - effective . if the cost of a 5-year intervention was equivalent to two family practice consultations per year , then net health benefits were approximately zero ( 0.06 , 14.3 to 13.6 qalys per 1,000 ) . however , reducing the costs of intervention had only a modest effect because the proportion of simulations in which incremental qalys were zero or lower set a limit to the potential increase in cost - effectiveness . if the cost of a 5-year intervention was equivalent to 20 % of one family practice consultation per year , then net health benefits were 5.8 ( 8.5 to 19.5 qalys per 1,000 , probability cost effective 75.1 % ) . when qalys were discounted at 1.5 % rather than 3.5 % , there was only a small difference in the probability of the intervention proving cost effective at 5 years ( probability 66.0 % ) or 10 years ( 76.8 % ) , although estimated mean net health benefits were greater , being 6.0 qalys per 1,000 for an intervention lasting 5 years and 10.1 qalys per 1,000 for an intervention lasting 10 years . sensitivity analyses were implemented to explore the effects of varying the unit costs of intervention and varying the discount rate . as expected , increasing the unit costs of intervention necessarily made the intervention less cost - effective . if the cost of a 5-year intervention was equivalent to two family practice consultations per year , then net health benefits were approximately zero ( 0.06 , 14.3 to 13.6 qalys per 1,000 ) . however , reducing the costs of intervention had only a modest effect because the proportion of simulations in which incremental qalys were zero or lower set a limit to the potential increase in cost - effectiveness . if the cost of a 5-year intervention was equivalent to 20 % of one family practice consultation per year , then net health benefits were 5.8 ( 8.5 to 19.5 qalys per 1,000 , probability cost effective 75.1 % ) . when qalys were discounted at 1.5 % rather than 3.5 % , there was only a small difference in the probability of the intervention proving cost effective at 5 years ( probability 66.0 % ) or 10 years ( 76.8 % ) , although estimated mean net health benefits were greater , being 6.0 qalys per 1,000 for an intervention lasting 5 years and 10.1 qalys per 1,000 for an intervention lasting 10 years . this study modelled the health outcomes of a universal intervention , aimed at all healthy adults , to promote physical activity in primary care . the study employed an empirical population of adults registered with uk family practices ; the size of the population at risk was equivalent in size to that of a small town or a primary care organisation . the intervention effect was derived from a meta - analysis of randomised controlled trials , based on the distribution of physical activity observed in a representative population sample in england . the results show that an intervention with only a small effect on the risk of disease may yield appreciable health benefits including an increase in life years lived free from physical disease and a reduction in life years lived with either single or multiple comorbidity . an important finding from this study is that interventions , or intervention effects in terms of behavioural changes resulting from intervention , must be maintained over prolonged periods of time in order for substantial health benefits to accumulate . this long time scale for the emergence of health benefits presents a challenge in allocating primary care resources to delayed rather than immediate benefits . the present results suggest that a brief intervention in primary care represents a costly way of achieving population - wide outcomes . even when the intervention is delivered at very low unit cost , there is only weak evidence that the intervention could have acceptable cost - effectiveness . these results therefore offer only limited support to continued investigation of a universal intervention as part of a standard family practice visit . future research should evaluate whether interventions targeted at high - risk individuals may be more suitable for utilisation in primary care , with population strategies being delivered through multi - sectoral interventions . s review of interventions to promote physical activity included studies published up to 2004 . of the 29 studies , 15 were set in primary care , including a range of intervention and follow - up methods . the overall increase in physical activity through intervention amounted to 0.28 standard deviations ( sd ) for a physical activity measure , with a 0.52 sd increase in physical fitness in 11 studies . this review provides evidence that interventions in primary care to promote physical activity may be effective , at least in the short term . these conclusions are supported by more recent systematic reviews [ 6 , 24 ] . however , individual level behavioural interventions to promote physical activity may be more costly that we have estimated for this study . recent studies suggest that community - wide or environmental interventions aimed at increasing use of leisure facilities or promoting active travel may have more acceptable cost - effectiveness . this research was grounded in data from a very large cohort of participants from primary care , which provided strong empirical evidence to construct the model . nevertheless , some multi - disease states were less frequent , leading to imprecise estimates . there were numerous estimates for incidence , mortality , prevalence of depression , as well as health care utilisation contributing to stochastic error in the model inputs and consequently to uncertainty in the model outputs . utility values were drawn from a secondary source because it was not feasible to obtain primary data for multiple disease states within the context of this study . while our study is uk - based , this secondary data source provided consistent estimates covering the number of disease states in this research . we did not set an upper age limit to eligibility for intervention because physical activity is beneficial even in old age . the model included only a limited range of health conditions and it is possible that there are wider health benefits of intervention that were not included . the model assumed that mortality reductions would be achieved only through the conditions of interest and not by reductions in mortality from other causes . the model did not include secular trends in the measures of interest because the direction of future trends is unknown . we acknowledge that true long - term intervention effects are not known and the model requires an important assumption that short - term effects must be maintained if the intervention is continued . we used results from a meta - analysis of randomised trials to provide an estimate of the intervention effect . however , most of the included studies used self - reported measures of physical activity and a similar effect has not been demonstrated employing objective measures . in a probabilistic framework , we used the standard error to model random error in the point estimate of the intervention effect . the intervention time course is also unknown and we did not model scenarios in which the effect of intervention might outlast the intervention itself . we also did not assume any social multiplier effects in our modelling where the impact of one person taking on more physical exercise might influence others around him therefore possibly underestimating intervention effect size . intervention effects were not allowed to vary in different population groups but we intend to study this further in future studies . it is clear that an intervention of very low cost has only a limited probability of proving cost - effective , with a more costly intervention yielding little or no net benefit . however , more complex scenarios could be envisaged in which the major costs may be incurred at the start , with lower maintenance costs . the model included only health care costs ; conclusions might differ appreciably if other costs and productivity changes were to be included . . the data were obtained from primary care records and utilisation of secondary care may also have been underestimated as we could not follow patient use of secondary care resources beyond referral and admission . we used the average of costs over all stages of a disease but this is a simplification because , in reality , costs may be higher at the start of the illness , or at periods closer to death . we caution that most estimates included in the model derived from the british primary health care system . costs and outcomes may be different in other countries and settings where resource use , the costs of care , and levels of physical activity and barriers to physical activity may be different . this study modelled the health outcomes of a universal intervention , aimed at all healthy adults , to promote physical activity in primary care . the study employed an empirical population of adults registered with uk family practices ; the size of the population at risk was equivalent in size to that of a small town or a primary care organisation . the intervention effect was derived from a meta - analysis of randomised controlled trials , based on the distribution of physical activity observed in a representative population sample in england . the results show that an intervention with only a small effect on the risk of disease may yield appreciable health benefits including an increase in life years lived free from physical disease and a reduction in life years lived with either single or multiple comorbidity . an important finding from this study is that interventions , or intervention effects in terms of behavioural changes resulting from intervention , must be maintained over prolonged periods of time in order for substantial health benefits to accumulate . this long time scale for the emergence of health benefits presents a challenge in allocating primary care resources to delayed rather than immediate benefits . the present results suggest that a brief intervention in primary care represents a costly way of achieving population - wide outcomes . even when the intervention is delivered at very low unit cost , there is only weak evidence that the intervention could have acceptable cost - effectiveness . these results therefore offer only limited support to continued investigation of a universal intervention as part of a standard family practice visit . future research should evaluate whether interventions targeted at high - risk individuals may be more suitable for utilisation in primary care , with population strategies being delivered through multi - sectoral interventions . hillsdon s review of interventions to promote physical activity included studies published up to 2004 . of the 29 studies , 15 were set in primary care , including a range of intervention and follow - up methods . the overall increase in physical activity through intervention amounted to 0.28 standard deviations ( sd ) for a physical activity measure , with a 0.52 sd increase in physical fitness in 11 studies . this review provides evidence that interventions in primary care to promote physical activity may be effective , at least in the short term . these conclusions are supported by more recent systematic reviews [ 6 , 24 ] . however , individual level behavioural interventions to promote physical activity may be more costly that we have estimated for this study . recent studies suggest that community - wide or environmental interventions aimed at increasing use of leisure facilities or promoting active travel may have more acceptable cost - effectiveness . this research was grounded in data from a very large cohort of participants from primary care , which provided strong empirical evidence to construct the model . nevertheless , some multi - disease states were less frequent , leading to imprecise estimates . there were numerous estimates for incidence , mortality , prevalence of depression , as well as health care utilisation contributing to stochastic error in the model inputs and consequently to uncertainty in the model outputs . utility values were drawn from a secondary source because it was not feasible to obtain primary data for multiple disease states within the context of this study . while our study is uk - based , this secondary data source provided consistent estimates covering the number of disease states in this research . we did not set an upper age limit to eligibility for intervention because physical activity is beneficial even in old age . the model included only a limited range of health conditions and it is possible that there are wider health benefits of intervention that were not included . the model assumed that mortality reductions would be achieved only through the conditions of interest and not by reductions in mortality from other causes . the model did not include secular trends in the measures of interest because the direction of future trends is unknown . we acknowledge that true long - term intervention effects are not known and the model requires an important assumption that short - term effects must be maintained if the intervention is continued . we used results from a meta - analysis of randomised trials to provide an estimate of the intervention effect . however , most of the included studies used self - reported measures of physical activity and a similar effect has not been demonstrated employing objective measures . in a probabilistic framework , we used the standard error to model random error in the point estimate of the intervention effect . the intervention time course is also unknown and we did not model scenarios in which the effect of intervention might outlast the intervention itself . we also did not assume any social multiplier effects in our modelling where the impact of one person taking on more physical exercise might influence others around him therefore possibly underestimating intervention effect size . intervention effects were not allowed to vary in different population groups but we intend to study this further in future studies . it is clear that an intervention of very low cost has only a limited probability of proving cost - effective , with a more costly intervention yielding little or no net benefit . however , more complex scenarios could be envisaged in which the major costs may be incurred at the start , with lower maintenance costs . the model included only health care costs ; conclusions might differ appreciably if other costs and productivity changes were to be included . the data were obtained from primary care records and utilisation of secondary care may also have been underestimated as we could not follow patient use of secondary care resources beyond referral and admission . we used the average of costs over all stages of a disease but this is a simplification because , in reality , costs may be higher at the start of the illness , or at periods closer to death . we caution that most estimates included in the model derived from the british primary health care system . costs and outcomes may be different in other countries and settings where resource use , the costs of care , and levels of physical activity and barriers to physical activity may be different . the results contribute new information towards understanding the potential for a universal strategy for physical activity promotion in primary care . firstly , an important increase in time lived free from physical disease , and a reduction in the time lived with single or multiple morbidity , may result from even a modest increase in physical activity levels . secondly , intervention effects must be maintained over a prolonged period of time in order for substantial health benefits to be realised , though these may continue to accumulate after the end of the intervention . thirdly , even when interventions can be delivered over the long - term at low annual cost , such as the cost of an additional family practice consultation each year , there is only weak evidence that the intervention might have acceptable cost - effectiveness when employed in a universal strategy . the present results emphasise that physical activity is a determinant of health important to primary care professionals , but also show that implementation of a universal strategy within primary care faces several challenges . while the results indicate some potential for a universal strategy , an alternative approach , which will be evaluated in future research , is the delivery of a selective or targeted strategy to focus intervention efforts on those at higher risk of disease . the use of fully anonymised gprd data was approved by the mhra independent scientific advisory committee ( ref . 09 - 085 ) . | backgroundthis study aimed to estimate the cost - effectiveness of a universal strategy to promote physical activity in primary care.methodsdata were analysed for a cohort of participants from the general practice research database .
empirical estimates informed a markov model that included five long - term conditions ( diabetes , coronary heart disease , stroke , colorectal cancer and depression ) .
simulations compared an intervention promoting physical activity in healthy adults with standard care .
the intervention effect on physical activity was from a meta - analysis of randomised trials .
the annual cost of intervention , in the base case , was one family practice consultation per participant year .
the primary outcome was net health benefit in quality adjusted life years ( qalys).resultsa cohort of 262,704 healthy participants entered the model .
intervention was associated with an increase in life years lived free from physical disease . with 5 years
intervention the increase was 52 ( 95 % interval 11 to 115 ) per 1,000 participants entering the model ( probability increased 91.9 % ) ; with 10 years intervention the increase was 102 ( 42164 ) per 1,000 ( probability 99.7 % ) .
net health benefits at a threshold of 30,000 per qaly were 3.2 ( 11.1 to 16.9 ) qalys per 1,000 participants with 5 years intervention ( probability cost - effective 64.7 % ) and 5.0 ( 9.5 to 19.3 ) with 10 years intervention ( probability cost - effective 72.4 % ) .conclusionsa universal strategy to promote physical activity in primary care has the potential to increase life years lived free from physical disease .
there is only weak evidence that a universal intervention strategy might prove cost - effective . |
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tuberculosis ( tb ) is a major public health problem in developing countries like india . gastrointestinal tb accounts for 3% of extra pulmonary tb , the most common site of involvement being the ileocaecal region . however , involvement of the appendix , lying so close to the ileocaecal region is rare . primary tb of the appendix presenting as appendicular abscess is even rarer with incidence of 0.1 - 0.6% . because of its rarity and absence of any specific clinical and radiological finding , diagnosis is made only after histopathological examination of the appendectomy specimen . in our review of cases , for 3 years , we report our experience with tuberculous appendicitis in a tertiary center in a nation where tb is still endemic . a 20-year - old male patient presented to our outpatient department with complaints of colicky type of pain in right lower abdomen , associated with vomiting and fever since 5 days . on examination , the physical examination showed marked tenderness in right iliac fossa . on initial work up , patients leukocyte count was 17,600/mm . on laparotomy , a diffuse inflammatory mass and abscess of the appendix [ figure 1 ] was found and appendectomy was done . exploration of the bowel and mesentery , through the grid iron incision showed normal ileum , cecum , and mesentery . in post - operative period , he had wound infection , which was managed by regular dressing . histopathological examination of the appendix revealed caseating epitheliod granulomas and lumen filled with neutrophilic infiltrates [ figures 2 and 3 ] . patient was later evaluated for primary source of tb else where in the body . computed tomography ( ct ) abdomen and pelvis showed normal bowel loops and mesentery [ figure 4 ] . tuberculin skin test was negative and erythrocyte sedimentation rate ( esr ) was 80 mm / h . patient was started on standard anti - tb drugs , course similar to pulmonary tb . intraoperative photograph of inflamed appendix with mass histopathology showing epithelioid granuloma with langhans giant cells and neutrophilic infiltrate in the lumen of appendix ( 4 magnification ) histopathology showing epithelioid granuloma with langhans giant cells and neutrophilic infiltrate in the lumen of appendix ( 10 magnification ) ct photograph showing normal abdomen colonoscopy photo showing normal ileum colonoscopy photo showing normal caecum deaver reported 16 cases of tubercular appendicitis in his series of 7610 appendectomies , mayo in 1905 reported 29 ( 1888 appendectomies ) , allen reported 2 ( 89 appendectomies ) , and scott in 1917 , 1 case out of 179 appendectomies . in recent studies , shah et al . , reported 10 cases of tubercular appendicitis over a period of 10 years , dymock et al . , 2 cases in an analysis of 1000 appendectomy specimens . in a review of 2921 appendectomies carried out in a tertiary center in india , only 2.3% of cases were tubercular appendicitis . in our practice , for 4 years , we reported one case of tuberculous appendicitis out of 229 cases operated for appendicitis the most common forms of non - pulmonary tb are tb of bones and joints ( 30% ) , urinary system ( 24% ) , lymph nodes ( 13% ) , sexual organs ( 8% ) , cerebrospinal meninges ( 4% ) , and alimentary system ( 3% ) . appendicular tb can occur as a primary or secondary form : the first form is due to a primary infection of the intestinal mucosa by mycobacterium bovis ; the second form is usually a consequence and complication of primary pulmonary tb by m. tuberculosis . the infection of appendix by tuberculous bacillus can occur by local extension of ileocaecal or genital tb , hematogenous spread from a distant focus and contact with infected intestinal contents due to ingestion of food contaminated with tubercle bacilli . the disease can present either as a chronic disease with recurrent episodes of fever , weight loss , right iliac fossa pain or as acute appendicitis , a latent type that is detected incidentally . the acute presentation occurs due to severe pyogenic infection that is superimposed on the tubercular appendix . this type of presentation is seen during the quiescent phase of pulmonary tb , if present . the presence of chronic abdominal pain of long duration in young adults , pulmonary tb , poor nutritional status and loss of weight , and the presence of chronic diarrhea have been said to be indicative of tb of the appendix , but these symptoms are of doubtful value . since there are no clinical and radiological features that are pathognomonic of appendicular tb , diagnosis is usually made after histopathological examination of the appendectomy specimen . based on histopathology , tb appendix can be described as ulcerative ( commonest form ) , hyperplastic and ulcer - hyperplastic form . other causes of granulomatous appendicitis include parasite - related appendicitis , crohn 's disease , sarcoidosis and foreign body - induced inflammation . signs and symptoms are nonspecific and similar to those of several other chronic abdominal diseases or it may simulate an acute appendicitis such as the present case . the diagnosis of a secondary localization by a pulmonary infection is usually simpler since the radiological aspects of pulmonary tb are often characteristic . high levels of adenosine deaminase ( ada ) in the ascitic fluid in peritoneal tb have been shown to be compatible with the diagnosis of tb with high sensitivity ( 100% ) and specificity ( 97% ) . determination of serum ca-125 concentration can be used in tuberculous peritonitis , not only to make an accurate diagnosis and ascertain the activity of the disease but also to follow the response to treatment . the most accurate diagnostic alternative to surgery is endoscopic biopsy of the lesions , which depends on the localization of lesions . standard anti - tb treatment with four antituberculous drugs ( isoniazid 5 mg / kg / day , rifampicin 10 mg / kg / day , pyrazinamide 30 mg / kg / day , and ethambutal 20 mg / kg / day ) for two months followed by isoniazid and rifampicin for 4 months is advocated . | tuberculosis is still a common infection in india .
although the ileocecal region is the most affected part in intestinal tuberculosis , acute tuberculous appendicitis is quite a rare entity .
our case report highlights a rare presentation of tuberculosis and a brief review of literature . |
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dermoid cyst is rarely found congenital brain tumors , which takes part in only about 1% of all intracranial lesions1011 ) . moreover , intracranial dermoid cyst is frequently seen below the tentorium , and its development in the cavernous sinus has been rarely reported in the literature1467121416 ) . to the our best knowledge of english literatures review , two cavernous sinus cases with rupture particularly , the cavernous sinus dermoid cyst extending into the posterior fossa has been also extremely uncommon46 ) . a ruptured dermoid cyst of the cavernous sinus extending into posterior fossa has been only reported6 ) . with its mass effect or spillage of ruptured particles into the subarachnoid space and ventricles the computed tomography ( ct ) scans and magnetic resonance ( mr ) imaging have advantages how to make a diagnosis and manage the ruptured dermoid cyst1721 ) . in this report , we report a second case of a ruptured dermoid cyst of the cavernous sinus extending into the posterior fossa . a 32-year - old female presented with mild headache in the occipital area over 4 years . blurred vision in the right eye , right hemifacial clumsiness , tinnitus , and swallowing difficulty were developed with headache . the severity of symptoms had gradually increased over the last 3 months . in the cerebellar function test mr imaging showed a heterogenous - signaled huge mass ( 6.54.63.3 cm ) in the right middle fossa and cavernous sinus extending into the posterior fossa . we also observed multifocal hyperintense lesions in the subarachnoid space on t1-weighted ( t1w ) mr image suggesting the ruptured particle of dermoid cyst . this mass showed hyperintensity on t2-weighted ( t2w ) image , hypointense but focal hyperintensity on t1w images , and also hydrocephalus was observed by compression of the adjacent structures including the 4th ventricle , brain stem , and cerebellum ( fig . we underwent internal debulking of tumor following dural incision , and its content was friable and grayish yellow - colored . the cavernous sinus portion was meticulously removed with preservation of neural structures , and the cavernous segment of the internal carotid artery was seen medially . the part of the tumor in the posterior fossa was excised , however , it was hard to make total excision of tumor because of its severe adhesion to the cranial nerves and brain stem . in the histopathological examination , 2 ) . at a few days after the operation , dysesthesia in the right face and hearing were improved with intact tandem gait . we report a rare case of ruptured cavernous sinus dermoid cyst extending into the posterior fossa . intracranial dermoid cysts have the component of keratinized squamous epithelium originated from ectoderm composed with dermal materials like hair and sebaceous glands351011 ) . they prefer to be located near the subarachnoid space , sulci or fissures because of the space which has minimal resistance to grow easily , and the midline in the skull base1221 ) . supratentorial dermoid cysts were thought to be found more often in the areas of suprasellar , parasellar , temporal , and frontobasal compartment , but in recent studies , infratentorial dermoid cysts are frequently reported1112171821 ) . dermoid cysts are rarely developed in the cavernous sinus and a few cases have been reported as intracranial rather than extradural , interdural dermoid cysts of the cavernous sinus , or parasellar lesions127121421 ) . most reported cases are abutted to venous structures such as the cavernous sinus , and purely interdural dermoid cyst of the cavernous sinus was described by nakagawa et al.14 ) . in the radiologic images of our case , it was unclear to define the precise origin from the cavernous sinus or parasellar region . by intraoperative findings , our case seems to be originated in the cavernous sinus , and extended into the posterior fossa . pathogenesis of the dermoid cysts in the cavernous sinus is unclear , however , intracranial intradural dermoid cysts develop near or adherent to large venous structures such as , the cavernous sinus for supratentorial dermoid cysts , and the torcular herophili for infratentorial dermoid cysts12 ) . considering that dermal structures or venous channels derive from the mesoderm , this association suggests that these lesions derive from mesodermal nest cells adherent to primitive veins . in an embryo of about 5 mm , the venous drainage of the cranial region endothelial pseudopods and pial vessels in the adjacent glia then perforate and penetrate into the nervous tissue , allowing the formation of new intraneural vessels . we speculate that this vascular migration is likely for the development of dermoid cysts adjacent to primitive veins , such as cavernous sinus by the entrapment of mesodermal non - vascular cells12 ) . dermoid tumors of the cavernous region present clinical signs , particularly oculomotor nerve palsy15 ) . focal neurological deficits tend to develop lately in the clinical course due to slow growth of these lesions . cystic rupture like our case may suddenly present and deteriorate clinical symptoms . in our case , the patient presented right hemifacial clumsiness suggesting trigeminal neuropathy and symptoms related to increased intracranial pressure . in the radiologic imaging , ct scans show hypodense appearance by the fat content in the dermoid cysts on the non - enhanced images1320 ) . mr imaging is more accurate to diagnosis than ct scans , and usually reveals hyperintensity on t1w image and hypointense to mixed signals on t2w image depending on the contents of dermoid cysts2491213171920 ) . although tumor mass was too large to differentiate interdural or intradural tumor in our case , we suspected that tumor was originated from cavernous sinus , based on the findings of coronal mr image . ruptured particles from the dermoid cyst were detected in the lateral ventricles with hyperintense signals on t1w image . because most cases are developed in the young age , we consider that a surgical resection is desirable in majority . in our case , tumor was placed in the right temporal base with originating from the cavernous sinus , and extension into the posterior fossa as previously mentioned . so , we resected a tumor via the combined route by subtemporal transtentorial and transpetrosal retrosigmoid approach . when it extended to the cavernous sinus or hard adhesion to adjacent neurovascular structures , it is better to undertake subtotal removal for avoidance of neurovascular damages1221 ) . we report an uncommon case of the ruptured dermoid cyst of the cavernous sinus extending into the posterior fossa , and review with pertinent literatures . | supratentorial dermoid cysts are uncommon to develop in the cavernous sinus .
we present a ruptured dermoid cyst of the cavernous sinus extending into the posterior fossa .
the patient was a 32-year - old female who complained occipital headache , blurred vision , and tinnitus over 4 years .
brain magnetic resonance ( mr ) imaging revealed an enhanced tumor in the right cavernous sinus extending into the right temporal base and the posterior fossa with findings of ruptured cyst .
surgical resection was performed , and pathological findings were confirmed to be a dermoid cyst .
we report a second case with ruptured dermoid cyst of the cavernous sinus extending into the posterior fossa . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
nanopharmacology is an interdisciplinary research field , which was developed as an interaction between chemistry , engineering , biology , and medicine , and it is currently receiving growing interest in the clinic.1 progress in nanotechnology has gained attention in recent years by developing novel nanoparticle - based drugs or by discovering novel applications in early diagnostic or prognostic assays in cancer.2 multiple preclinical studies aim to improve the therapeutic index of a patient diagnosed with cancer using a wide range of nanostructures including carbon nanotubes , peptides , nanodiamonds , cyclodextrine , graphenes , liposomes , quantum dots , nanowires , and metal - based nanoparticles.3,4 the latest advances in nanotechnology have brought various options that could be used in the clinic by employing constructs for molecular diagnosis , disease detection , cytostatic drug delivery , and nanoscale immunotherapy.58 the united states food and drug administration has approved the use of liposome - encapsulated doxorubicin ( doxil ; janssen products , lp , johnson & johnson , new brunswick , nj , usa ) and paclitaxel attached to nanoparticles ( abraxane ; celgene corporation , summit , nj , usa)9,10 in cancer therapy . in this review , we present the latest investigation on nanostructure systems with applications in hematology and oncology . the latest advancements in nanopharmacology lead to heightened expectations concerning their application in diagnostics , therapy and imaging . in the last few years , our team has shed a new light on the field11,12 by different conjugation procedures for these therapeutics . overcoming this threshold bears major clinical significance in oncology and hematology , as developing nonviral gene delivery vehicles will bring new patient - tailored drugs within reach ( figure 1 ) . the transport of therapeutic nucleic acids through the cell membrane is inefficient mainly in experimental models and includes antisense or antigene oligonucleotides , short interference ribonucleic acid ( sirna ) or micro ribonucleic acids ( mirna).13,14 surgical resection in early tumor stages is the main therapeutic option for most solid malignancies , yet therapeutic benefits are frequently modest because of the high rate of tumor recurrence.15 chemotherapy and , more recently , molecular therapy , were proven to offer much more efficient therapeutic approaches for patients diagnosed with cancer.2 nevertheless , these options are most often accompanied by important systemic side effects associated with the active agent , making a direct delivery the most elegant and efficient therapeutic option . the direct delivery of chemotherapy drugs aims to achieve high concentrations of the cytostatic agents at the target site with minimized risk of systemic toxicity ( figure 2).16 in cancer chemotherapy , the clinician aims to achieve a good therapeutic index , which is the ratio of the lethal dose for 50% of the population to the minimum effective dose for 50% of the population.17 however , cancer is most often characterized by multidrug resistance ( mdr ) and thus scientists have developed new ways to target the mdr cells.18 mdr cells are known to be frequently located in hypoxic areas , distant from any blood supply , thus overcoming the natural barrier of drug efflux pump activity.18 such smart molecules may increase the drug s bioavailability and transform an active agent from a low therapeutic candidate into a highly efficient drug . a wide variety of both organic and inorganic substances are used for engineering nanostructures , such as liposomes , micelles , nanoemulsions , polymers , quantum dots , gold , iron oxide , and even dendrimers.1922 all these structures were developed in order to have a large surface area , making these particles suitable for suspension storage , as well as high drug encapsulation and extensive surface absorption capacity , which are pharmacokinetic features that are found in any current structure used in classical pharmacology.23 however , the most important aspect , especially for cancer , is that nanocarriers are able to bypass the extracellular efflux activity of the adenosine triphosphate - binding cassette transporters in order to be internalized via nonspecific endocytosis,2426 such as the case of immunoliposomes27,28 and poly ( butyl)-cyanoacrylate nanoparticles.29,30 at the same time , the nanocarriers use surface charge - switchable polymeric magnetic nanoparticles as a safe delivery system.3133 in this way , the active agent is released near the nucleus , far away from the membrane - bound p - glycoproteins , which is of paramount importance when trying to overcome the resistance to conventional chemotherapy of cancer stem - like cells.3436 ozeki et al37,38 have experimented with a new drug delivery model in malignant gliomas . brain barrier by using a unique thermo - reversible hydrogel , composed of drug / poly(lactic - co - glycolic acid ) ( plga ) microspheres . this thermo - reversible polymer is a gel at body temperature and a sol at room temperature conditions in which the drug / plga microspheres dispersed in the polymer are injected into the human body . following the procedure , a gel forms around the injection site ; this keeps a high concentration of the active substance in the tumor , preventing its dispersion in adjacent healthy tissues.39 devalapally et al40 used poly(epsilon caprolactone ) nanostructures whose surface has been modified with poly(ethylene glycol ) ( peg ) before being loaded with tamoxifen and paclitaxel for the treatment of multidrug - resistant cancer cells . the results were encouraging , as this combination resulted in a lower therapeutic dose of the cytostatic agent , with important clinical applications regarding chemotherapy - related side effects . the first groundbreaking drug was doxorubicin encapsulated in circulating liposomes ( doxil ) for the treatment of kaposi s sarcoma in patients diagnosed with acquired immunodeficiency syndrome ( aids).41 recently , this therapeutic option has been applied for other cancers such as breast cancer , since doxorubicin encapsulated in liposomes induces a twofold increase in intracellular drug levels when compared to standard doxorubicin treatment.42 doxil is a pegylated liposomal drug that has a 100 nm diameter in order to prevent the interaction with plasma proteins such as opsonins and high - density lipoproteins ( hdls ) and low - density lipoproteins ( ldls ) , or to avoid elimination by macrophages.4345 after the conjugation of a liposome with peg ( a process called pegylation ) , the new drug can stay in the systemic blood flow for longer periods of time due to the development of an aqueous layer on the surface of the liposome , leading to a lack of immune recognition and rejection.46,47 this results in the stabilization of the lipid bilayer and steric hindrance , with important consequences such as decreasing protein absorbance and recognition by the host s macrophages.48 many reports show the ability of various gold / silver nanoparticles or carbon nanostructures to enhance the antitumor effect of certain drugs.23 however , in hematology , hdl nanostructures target the scavenger receptors ( b1 ) and promote cholesterol efflux in lymphoma cells.49 indirectly , these exogenous lipoproteins inhibit lymphoma growth and invasion by starving the malignant cell,50,51 thereby aiding the classic chemotherapy regimen . pegylated liposomes loaded with docetaxel , and prepared using the thin film hydration method , showed enhanced in vitro cytotoxicity against a549 and b16f10 cells when compared to taxotere ( sanofi - aventis , bridgewater , nj , usa).52 the capacity of a self - nanoemulsifying drug delivery system was assessed in order to increase the bioavailability of docetaxel and , consequently , its therapeutic activity.53 this study showed that a self - nanoemulsifying drug delivery system exhibited superior efficacy with low associated toxicity when compared to the commercialized formulation of this bioactive agent ( taxotere).53 in recent years , gold nanoparticles have also emerged as therapeutic options for the targeted delivery of antineoplastic active substances , due to their special chemical and physical properties such as functional versatility , biocompatibility , and low toxicity.5456 apart from being of small size ( 3050 nm in diameter),57 naked gold nanostructures have a plasmon absorption in the near - infrared region and display strong photothermal ability . these structures lack a silica core , have a spherical shape , and have a strong and tunable absorption band between 550 nm and ~820 nm.5861 these properties make them highly efficient carriers of various drugs already used in the clinic . their effect has already been shown by our team in malignant gliomas and hepatocellular carcinoma for temozolomide , cisplatin , doxorubicin , and capecitabine.62 diamonds can provide a very efficient delivery system for some chemotherapy agents . in the last few months , nanodiamonds have emerged as potential carriers in neuro - oncology or hemato - oncology . xi et al63 have conjugated nanodiamonds with doxorubicin and used convection - enhanced delivery for supratentorial tumors in a murine model . man et al64 have also used nanodiamonds to deliver another type of anthracycline to multidrug - resistant malignant cells . they showed that acute myelogenous leukemia , often leading to patient death in the clinic because of resistance to chemotherapy , might be managed by a nanotechnology - based targeted delivery of daunorubicin to the hematological malignancies . camptothecin is a natural hydrophobic anticancer drug that could be potentially used for breast adenocarcinoma management if delivered correctly at the tumor site.65 delivery can be achieved using nanotechnology , as is the case of the self - assembling peptide amphiphile nanofibers . camptothecin was also confirmed by min et al67 to be efficient in breast chemotherapy when delivered to malignant cells , by encapsulating it in modified glycol chitosan nanoparticles , thereby achieving a high concentration with minimal side effects in healthy tissues , after having used subcutaneously implanted xenografts in immunocompromised mice . since monoclonal antibodies are increasingly used in clinical oncology , some investigators have tried to add the targeted effect of antibody - based drugs to a nanocarrier in order to obtain maximum anticancer effects with minimal side effects.2,23 thus , trastuzumab was conjugated with various nanostructures , including carbon nanotubes or nanospheres.68 the desired effect was achieved and , in the near future , we may expect newly described cytostatic agents in phase i or phase ii clinical trials . ni et al69 have applied this concept by developing a local vaccine after conjugating graphene oxide targeting interleukine-10 receptor . thus , the anti - inflammatory action of interleukin-10 is blocked and the suppressive tumor microenvironment becomes a target for the immune system . nanovectors can be used as carriers for drugs , but also for contrast substances , with a high applicability in diagnostic medicine.70 iron oxide , gold , gadolinium , or even quantum dots represent good alternatives for radiation oncology , photodynamic therapy , or hyperthermia.71,72 iron oxide has important superparamagnetic characteristics and is one of the most investigated nanostructures in diagnostics , including in lymph node imaging , the inhibition of cancer cell dissemination and stem cell trafficking , visualization of ribonucleic acid ( rna ) , interference and t - cell - specific labeling.7377 as a contrast agent , iron oxide is especially useful for magnetic resonance imaging ( mri ) , and it is very sensitive in detecting solid tumors , but it has little or no applicability in lymph node micrometastasis or hematological malignancies.78,79 while still in the early stages , the research in this field will more than likely improve in the near future . in prostate cancer , harisinghani et al78 have already proven that the use of iron nanostructures as a contrast agent in mris detects over 90% of all lymph node disseminations , which is in comparison with the detection rate of 35% in classic mris . even though chemotherapy remains the most widely used and effective treatment option for disseminated malignancies , acquired or intrinsic drug resistance accounts for almost 90% of treatment failure . mdr represents the simultaneous resistance to various medications that are different both structurally and functionally , most often as a result of the drug efflux pumps that reduce the intracellular levels and thus reduce the cytotoxic effect on the cancer cell.6 new nanotechnology - based theranostics are evolving and are expected to confer new strategies in overcoming the drug efflux transporters , which are findings that are presented further in the next section . the multifunctional characteristics of the nanocarriers make them very suitable for treating a heterogeneous tumor mass in comparison to classic approaches.15 nanocarriers have a preferential accumulation within the malignant cell due to the enhanced permeability and retention effect.23,80 thus , the drug concentration is increased in the malignancy and reduced in the surrounding , healthy tissue . this will result in an increased efficacy of systemic therapy , with decreased side effects.23,70 nanotechnology can be applied not only in chemotherapy , but also in radiation oncology , by combining radiobiology with experimental pharmacology . malignant cells are sensitive to ionizing radiation emitted by various radioactive metals.81 by delivering such substances to the primary tumor site , we may improve current radiotherapy or brachytherapy protocols . chanda et al82 have conjugated gum arabic glycoproteins to gold nanoparticles and tested this new assay on a murine model of prostate cancer . the administration of a single dose of beta - emitting irradiation increased the local administered dose up to 70 gy and induced the regression of prostate adenocarcinoma in nude mice . garrison et al83 also used an in vivo murine model of prostate cancer and demonstrated that beta radiation emitting bombesin could be used to specifically target cancer cells . the human body has natural barriers for preventing a wide range of diseases , whether considering the organism / body level , the tissue organ level , the cellular level , or even the molecular level . thus , the simple aim of achieving highly localized drug delivery with maximal anticancer effects and minimal side effects is very troublesome , as it can be expensive , time consuming , and it does not offer any guaranteed success.2,23 this emphasizes the need to design highly specific carriers that can deliver highly specific active agents in order to achieve maximum efficacy with minimal toxicity . in vivo , various mirs can be delivered either by viral or nonviral carriers , depending on transfection efficiency , the safety of the receiving host , immunogenicity , or side effects.84,85 nonviral carriers are nonetheless considered to be more suitable in the clinic , especially cationic transporters such as peg . this is because it has a strong buffering ability and it can release functional genetic material into the cytosol after having induced osmotic endosome breakage.8688 the main disadvantage is that peg is not cell - specific , and one would need very high doses in order to achieve the desired concentration , leading to potentially serious side effects.89,90 thus , the need to improve current knowledge in the field and to produce other ligand molecules for aptamers functionalization . aptamers are short , single - stranded oligonucleotides formed by 3050 bases and they express minimal or even no antigenicity and immunogenicity , making them more suitable for in vivo use in clinical hematology and oncology.9193 other nonviral vector systems may also include carbon nanomaterials , such as nanotubes or fullerenes . our studies used nanotubes because of their unique intrinsic physical and chemical properties in an attempt to deliver sirna in hepatocellular carcinoma cells.12 the molecular analysis of the experiments has proven that p53 , tnf- , and vegf levels were altered after sirna transfection . this proves that carboxylated carbon nanotubes may provide an alternative to the lipid transfection system - based therapy for liver malignancies . the successful functioning of the endosomal sirna system and followed by the release of the rna molecules into the cytoplasm are very important for the efficient use of oncogene silencing.1,2,13 in order for this process to be carried out with minimal side effects , tertiary complexes were developed out of nucleic acids , polycations , and a charge - reversal polymer that can ph - dependently alter its electric charge either into the positive or negative state.9496 when the vector arrives into an organelle such as an endosome or a lysosome , both are known to have a ph of 56 , the charge conversion facilitates endosomal escape through a membrane disruption process after having enhanced the so - called proton sponge.9799 apoptosomes represent other models of molecular self - assembly structures . in such wheel - like structures , an individual apaf-1 protein will form a complex with cytochrome - c100 before recruiting and activating procaspase-9.101 this will trigger a cascade of other events , which may lead to apoptosis . polymeric micelles are artificial structures that resemble apoptosomes and act as either drug solubilizers or carriers of antisense oligonucleotides and drug molecules.102 a single - stranded oligonucleotide can recognize a target molecule on a cancer cell both through watson crick base pairing with folic acid , and also through hydrophobic interactions and hydrogen bonding.103 such an oligonucleotide ligand is also known as an aptamer , which has very important properties that include its small size , a lack of immunogenicity , and ease of synthesis.104,105 exosomes are vesicles ranging from 3090 nm in diameter , and they are known to play a key role in intercellular communication.106,107 this communication is accomplished using various cytokines , interleukins , and a substantial amount of rna.108 rna carried by the exosomes is mostly a rna and mirna messenger , with very little 18s and 28s ribosomal rna.109,110 since exosomes are used in normal cell physiology in rna transport , researchers have attempted to use these nanostructures in gene therapy as a vector to deliver therapeutic nucleic acids to target cancer cells.108,111 gene therapy aims to provide a therapeutic solution for the cause of the disease , rather than for its symptoms . two types of vectors ( either viral or nonviral ) are currently available in the us , according to an online search of the national institutes of health database ( http://clinicaltrials.gov/ct2/home ) . most of the 262 ongoing trials use viral vectors , yet this approach is associated with a high toxicity and an important immunological response from the host organ . exosomes are far more efficient because they can target cancer cells and trigger little or no immune response since they are isolated from the patient s bodily fluids and are subsequently transferred back to the same patient after an insertion or deletion of the genetic material in vitro.112114 wahlgren et al115 used exosome - delivered sirna in order to achieve posttranscriptional gene silencing . they showed that the mapk-1 protein was downregulated in both monocytes and lymphocytes that were cocultured with particles , which were genetically modified to carry an anti - mapk-1 transcript . exosomes represent an important delivery system,116,117 which proved its efficacy in vitro for rna and protein transport.118 a good therapeutic effect with low immunogenicity was observed for sirna.1,2,13 in a study conducted by alvarez - erviti et al119 the capacity to downregulate the bace1 protein and messenger rna levels was demonstrated using exosome - mediated sirna delivery produced by dendritic cells . the same group119 has also engineered dendritic cells to express the exosome - specific protein , lamp2b , fused with the peptide , rabies virus glycoprotein , which is specific for neuronal lineage cells . the clinical implications are of great potential in the management of alzheimer s disease.120 gold is a noble metal used throughout the ages of human history in all aspects of civilization , including in medical science,121,122 and nanotechnology - based new approaches make no exception . polyvalent oligonucleotide - functionalized gold nanostructures have been designed to enter cancer cells without the use of a cationic cocarrier after having been functionalized with a synthetic mir sequence . the prototype of the mir mimic - gold nanoparticle construct consists of a 115 nm gold nanoparticle , which was functionalized with a monolayer of a double - stranded alkylthiol - modified rna molecule of around 30 duplexes.123 hao et al further proved that a gold nanostructure could carry the mimics of mir-205 , which are known to have a tumor suppressive effect , thus inhibiting cancer cell proliferation and migration.123 mirnas are able to modulate different pathways,124,125 taking into account that a single mirna is able to target multiple genes . various approaches were applied to assess the significance of a particular mirna or distinct representatives from a mirna family , while noting that mirnas from the same family could have antagonistic biological effects.126,127 there is increasing evidence that attempts to explain the mirna s observed correlation with drug sensitivity.128130 the practical implication of mirnas in the initiation , development , and progression of cancer has led to the buildup of novel therapeutic schemes . approaches include , among others , the inhibition of upregulated mirnas ( oncogenic role ) , as well as using mirna replacement therapy by restoring the normal levels of tumor supressors mirna . oncogenic mirnas are inhibited based on antisense oligonucleotides , antagomirs , sponges , or locked nucleic acid structures.118 additional approaches involve the restoration of tumor suppressor mirna expression using mirna mimics , based on viral or nonviral delivery systems . both approaches have showcased favorable outcomes in preclinical and clinical studies.118 considering the significance of mirnas in cancer and its capacity to modulate various biological pathways , mirna mimics / inhibition asserted a new and effective therapeutic strategy in cancer.1,2,118 specifically , mirnas or anti - mirna may be used individually or in combination with chemotherapy , leading to an enhanced therapeutic response and to an improved survival rate.131 in order to apply the vast potential of mirnas for therapy , the main obstacle for the successful translation of therapeutic strategies into the clinic remains the pathway of delivery.132 mirna expression patterns can be altered by various mechanisms , including genetic and epigenetic alterations.133 correlations between mirna expression and chromosomal abnormalities were shown to be involved in the pathogenesis of chronic lymphocytic leukemia , since mirnas are involved in the initiation , prognosis , and chemoresistance of chronic lymphocytic leukemia.134136 concomitantly , the inhibition of the oncogenic mir-21 with antisense oligonucleotides generates a proapoptotic and antiproliferative response in vitro in different cell models , reducing tumor development and metastatic potential in vivo.118 other examples are presented in table 1 . nanotechnology is of major interest in clinical hematology and oncology for both therapy and diagnosis because of their unique features . these include self - assembly or the ability to make use of the enhanced permeability and retention capacity that most malignancies have as a consequence of leaky neoangiogenesis and the absence of a functional lymphatic system.149 nanostructures ( table 2 ) can also be designed to carry useful payloads that include low molecular weight chemotherapy agents or contrast agents.150,151 moreover , the newly formed structures are able to rapidly detect cancer cells , load multiple anticancer agents on their surface , and deliver the drugs rapidly at the target cell,152154 while preventing their bioactive cargo degradation when the investigator chooses to use an rna - based approach . non - hodgkin s lymphomas are the most common lymphohematopoietic malignancies both in the us and in europe.166 one particular type is anaplastic lymphoma kinase ( alk)-positive anaplastic large cell lymphoma , which is a very aggressive t - cell lymphoma with an abnormal expression of both the alk oncogene , as well as the surface protein , cd30.167169 a nucleic acid - based knockdown of alk gene expression has been proven to promote cell death of the malignant t - cell.170,171 mori et al172 have developed an rna aptamer that specifically binds to the cd30 epitope . zhao et al173 have subsequently hypothesized that a lymphoma cell - selective delivery of a tumor gene - specific sirna could be achieved by assembling a functional rna nanocomplex comprising the cd30-specific aptamer and the alk - targeted sirna , all within a nanosized peg - based polymer carrier . peg - based structures are considered to be rather safe , as toxicity assays done using balb / c mice showed little or no side effects , except for 40% accumulation in the liver.174 this new approach proved that the nanocomplex could be cancer cell - selective and cancer gene - specific , with great potential in the clinic if hepatic damage can be avoided . another non - hodgkin s lymphoma with a very aggressive behavior and short - term survival is mantle cell lymphoma . this particular type of malignancy is resistant to most therapeutic approaches , including immunochemotherapy and stem cell transplantation , leading investigators to look for different salvage treatment options.175178 syk is a new target for the management of b - lineage leukemias and lymphomas,179 as it regulates apoptosis by controlling activation of the phosphoinositide 3-kinase / akt , nuclear factor - kappa b , and signal transducer and activator of transcription 3 pathways , which are all very important in the signaling of the stem cell lineage.180,181 cely et al182 reported a different approach by developing a nanotechnology - based platform that can be used to target a very selective syk inhibitor for the lymphoma cell . the designed liposomal nanoparticle was the pentapeptide mimic , 1,4-bis(9-o - dihydroquinidinyl)phthalazine / hydroquinidine 1,4-phathalazinediyl diether ( c16 ) . the liposomal nanoparticle of c16 was shown to induce apoptosis of the lymphoma cell after 24 hours , providing the scientific background for an alternative treatment for refractory mantle cell lymphoma.182 however , previous experience using liposomes shows that this treatment strategy is accompanied by several side effects . for patients with aids - related kaposi s sarcoma , 30% of those treated with doxil presented with low blood counts and palmar carbon nanotubes are tubes made out of graphic carbon that have very good mechanical strength , good flexibility , and excellent thermal and electrical conductivity,185187 qualities that initially made them suitable candidates for novel drug design . these tubes have been conjugated with monoclonal antibodies and plasmid deoxyribonucleic acid ( dna ) in order to achieve cancer cell inhibition,188191 and conjugates have also been made with paclitaxel and other cytostatics.192 liu et al inhibited the growth of breast cancer by conjugating carbon nanotubes with paclitaxel , and they showed that the intravenous administration of 10 mg / kg of the new com pound enhanced the therapeutic efficacy when compared with doxorubicin - free treated mice.104 still , because of their fiber shape and size , carbon nanotubes cause cytotoxicity , inflammation , and dna damage in vitro.193196 the animal models used to study the toxic effects of carbon nanotubes demonstrated that the high concentrations needed to induce the regression of the tumor may cause chronic lung inflammation , foreign - body granulomas , or interstitial fibrosis,197200 limiting their potential clinical use . other types of nanoparticles with biomedical application are metallic colloidal gold and silver.201206 these structures are used for photothermal ablation therapy , as well as for contrast enhancers in computed tomography or x - ray diagnostics.207,208 niidome et al202 have reported no toxicity in their studies in a mouse model of colon adenocarcinoma after having used intravenous peg - coated nanorods , in spite of the fact that gold may interact with intracellular proteins and modify their structure , causing autoimmune - related toxicity . silver nanostructures are commercially available for antimicrobial use,209,210 yet recent data show that silver oxide may also be used in cancer research , as the nanostructure cargo can induce the regression of cancer neoangiogenesis.211214 still , toxicity limits their use because silver nanoparticles can cause destruction of the blood brain barrier , brain degeneration , and edema,215217 as well as liver failure.218 diagnostics can also be aided based on diffident nanostructures types including quantum dots or metallic core - shell nanoparticles that usually contain cadmium telluride , cadmium selenide , and either indium arsenide or indium phosphide.1,2 this structure is then covered by a shell of zinc sulfide and is subsequently coated with peg in order to facilitate the attachment of various drugs , nucleic acids , or antibodies.219221 these structures are very good fluorophores because of their broad - spectrum fluorescence,222 and they can be used to properly identify cancer cells , as well as signal events such as peroxisome activity or the presence of certain membrane receptors.223226 toxicity in clinical use is not known in great detail , but it seems that following the removal of the coating after their exposure to oxidative environments such as the endosome,227229 quantum dots may be very toxic , which may limit their clinical use ( figure 3 ) . in recent years , important progress has been made in nanotechnology , with its ever - increasing applicability in basic and translational medicine , leading to the appearance of a new field known as nanomedicine . this new science deals with the engineering of various structures of nanoscale dimensions that can be properly conjugated with various highly specific targeting agents in order to be used in the clinic , for either early diagnostic purposes or for disease treatment.1,2 these endeavors are possible because nanoparticles have unique properties , such as a preferential accumulation in the neoplastic tissue in comparison with healthy cells.1,2 these particles hold great potential for possibly replacing current active agents , which have been shown to be highly inefficient , based on epigenetics and molecular pharmacology principles . this step in clinical oncology and hematology is , however , still far from being implemented in clinical practice . nevertheless , with each experimental report , we come closer to a patient - tailored approach in order to achieve maximum anticancer effects with minimal side effects . | nanoparticles have displayed considerable promise for safely delivering therapeutic agents with miscellaneous therapeutic properties .
current progress in nanotechnology has put forward , in the last few years , several therapeutic strategies that could be integrated into clinical use by using constructs for molecular diagnosis , disease detection , cytostatic drug delivery , and nanoscale immunotherapy . in the hope of bringing the concept of nanopharmacology toward a viable and feasible clinical reality in a cancer center , the present
report attempts to present the grounds for the use of cell - free nanoscale structures for molecular therapy in experimental hematology and oncology . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
over the past several decades , infant and child mortality remain one of the major public health challenges faced by the world . infant mortality rate may be an indicator of how a society meets the needs of its people indicator of deprivation , unmet health needs and unfavourable environmental factors and a sensitive indicator of country s health . there is consistent decline in infant mortality in india since 1970 s . from 127 deaths per thousand live births in 1971 , it has declined to 42 deaths per thousand live births in 2012 showing annual decline of 1.6 percent . although , infant mortality has declined over time , but at the current pace , india is unlikely to achieve the millennium development goal ( mdg-4 ) of reducing infant mortality to 27 per thousand live births by 2015 . the studies on infant mortality in india , has focused on the socio - economic and demographic factors that may have a direct or indirect effect on infant death . it is documented by different researchers that household environment , economic condition , place of residence , education of mother , and health care utilization ; are significant determinants of infant mortality.[6 - 8 ] recently , household headship as a factor of infant mortality has gained importance among the researchers,[9 - 12 ] but studies exploring the association between infant mortality and type of household headship are limited , at least in india . the changes in socio - economic conditions of the country has resulted in a continuous increase in female - headed households ( fhh).[9 , 13 ] according to the census of india , 2011 , about 27 million ( 11 percent ) of the total households in india is headed by women . earlier studies from developed and developing countries suggested both positive and negative associations between female headship and economic condition of household.[9 , 10 , 13 - 17 ] on the basis of the above discussion , this study examined the association between household headship and infant mortality in india . the study is based on the national family health survey ( nfhs-3 , 2005 - 06 ) covering 109,041 households with 11 percent fhh from all the states of india . nfhs is the most comprehensive survey that provides information on death at the household level , specifically , the exact age at death of infants for each month till the first year of life . this analysis is restricted to ever - married women who had at least one live birth for a period of five years preceding the survey , and recorded 51,555 live births and 2500 infant deaths during the same period . the variables included in the study are place of residence , religion , caste , wealth index , household size , age of the household head , number of eligible women in the household , number of children below five , age at first birth , antenatal care , education of mother , exposure to mass media , total number of children ever born , place of delivery , and preceding birth interval . in addition , kaplan meier survival curves are drawn to examine the pattern of survival at exact ages . chi - square test was used to examine the association of sex of the household head with some selected household and individual level variables . the results are presented in three sub - sections . the first section deals with the differences in socio - demographic characteristics according to household headship . the second section discusses the determinants of infant mortality , and the last section shows the survival pattern during infancy according to household headship . the differences observed in socio - demographic characteristics between mhh and fhh are presented in table 1 . it was observed that , in fhh , percentage of home deliveries was high ; and antenatal care and percentage of children ever breastfed was low than mhh . in addition , fhh had higher percentage of illiterate mothers ( 69% ) than mhh ( 63% ) . seventy percent of fhh had no exposure to mass media and only six percent had exposure to both radio and television . on the other hand , 13 percent of mhh had exposure to both radio and television . fhh also performed better in certain indicators ; for example , higher percentages of children were immunized and lower percentages of children were born with short birth interval . the study observed that the fhh with low economic status reported higher percentage of infant deaths . socio - demographic characteristics of households reporting infant death in india , nfhs-3 ( 2005 - 06 ) the results of chi - square test indicate significant association of household headship with infant mortality . group , number of children below five years of age , religion , caste , wealth index , exposure to mass media , preceding birth interval , place of delivery , antenatal care , age at first birth , education of mother also showed significant association with infant mortality . on the basis of the results obtained from chi square test , the study further examined the association of different factors with infant death according to household headship . the adjusted and unadjusted hazard of infant death from mhh and fhh is presented in table 2 and table 3 , respectively . unadjusted estimates indicate that household size , number of under - five children , age of the household head , wealth index , education of mother , age at first birth , length of preceding birth interval , antenatal care , and breastfeeding status all have independent effect on risk of infant death , in both mhh and fhh . cox proportional hazard model showing the odds of infant death in male headed households , nfhs-3 ( 2005 - 06 ) , india cox proportional hazard model showing odds ratio for infant death from female headed households , nfhs-3 ( 2005 - 06 ) , india however , after adjusting for different household level factors , the increase in household size , number of children below five and improvement in economic condition significantly reduced the risk of infant death in mhh , while household size had no significant association with infant death in fhh . similar results were observed from the adjusted effect of maternal characteristics for mhh and fhh . the results show that age of mother at first birth , education and number of children ever born are significant predictors for infant mortality . differences were observed in the association of child level factors with risk of infant death . none of the child - related factors had significant association with the risk of infant death in mhh , but antenatal care and breastfeeding status of the child were significantly associated with risk of infant death in fhh . when all the factors were considered simultaneously , household size , number of eligible women , number of children below five , education of mother , previous birth interval and breastfeeding status remained significant predictors of infant mortality in mhh . for fhh , only duration of preceding birth interval and breastfeeding status of the child had significant association with risk of infant death . urban children living in fhh has the highest probability of survival out of the four combinations of sex of the household head and place of residence ( see figure 1 ) . it may be postulated that female head of the household are more concerned about the type of care to be provided to the child during infancy , and the mother of the young child may feel comfortable in discussing different issues related to child care with an elderly female member of the household than an elderly male . as shown in figure 2 , female child in fhh enjoys some advantages in terms of survival during infancy . this may indicate that , along with the biological advantage of survival for a female over a male child , a newborn girl is not discriminated against in a fhh in providing necessary care during infancy . survival pattern during infancy by place of residence in mhh and fhh , nfhs-3 ( 2005 - 06 ) , india survival pattern according to sex of the child in mhh and fhh , nfhs-3 ( 2005 - 06 ) , india the differences observed in socio - demographic characteristics between mhh and fhh are presented in table 1 . it was observed that , in fhh , percentage of home deliveries was high ; and antenatal care and percentage of children ever breastfed was low than mhh . in addition , fhh had higher percentage of illiterate mothers ( 69% ) than mhh ( 63% ) . seventy percent of fhh had no exposure to mass media and only six percent had exposure to both radio and television . on the other hand , 13 percent of mhh had exposure to both radio and television . fhh also performed better in certain indicators ; for example , higher percentages of children were immunized and lower percentages of children were born with short birth interval . the study observed that the fhh with low economic status reported higher percentage of infant deaths . socio - demographic characteristics of households reporting infant death in india , nfhs-3 ( 2005 - 06 ) the results of chi - square test indicate significant association of household headship with infant mortality . group , number of children below five years of age , religion , caste , wealth index , exposure to mass media , preceding birth interval , place of delivery , antenatal care , age at first birth , education of mother also showed significant association with infant mortality . on the basis of the results obtained from chi square test , the study further examined the association of different factors with infant death according to household headship . the adjusted and unadjusted hazard of infant death from mhh and fhh is presented in table 2 and table 3 , respectively . unadjusted estimates indicate that household size , number of under - five children , age of the household head , wealth index , education of mother , age at first birth , length of preceding birth interval , antenatal care , and breastfeeding status all have independent effect on risk of infant death , in both mhh and fhh . on the other hand , rural residence increased the risk of infant death . cox proportional hazard model showing the odds of infant death in male headed households , nfhs-3 ( 2005 - 06 ) , india cox proportional hazard model showing odds ratio for infant death from female headed households , nfhs-3 ( 2005 - 06 ) , india however , after adjusting for different household level factors , the increase in household size , number of children below five and improvement in economic condition significantly reduced the risk of infant death in mhh , while household size had no significant association with infant death in fhh . similar results were observed from the adjusted effect of maternal characteristics for mhh and fhh . the results show that age of mother at first birth , education and number of children ever born are significant predictors for infant mortality . differences were observed in the association of child level factors with risk of infant death . none of the child - related factors had significant association with the risk of infant death in mhh , but antenatal care and breastfeeding status of the child were significantly associated with risk of infant death in fhh . when all the factors were considered simultaneously , household size , number of eligible women , number of children below five , education of mother , previous birth interval and breastfeeding status remained significant predictors of infant mortality in mhh . for fhh , only duration of preceding birth interval and breastfeeding status of the child had significant association with risk of infant death . urban children living in fhh has the highest probability of survival out of the four combinations of sex of the household head and place of residence ( see figure 1 ) . it may be postulated that female head of the household are more concerned about the type of care to be provided to the child during infancy , and the mother of the young child may feel comfortable in discussing different issues related to child care with an elderly female member of the household than an elderly male . as shown in figure 2 this may indicate that , along with the biological advantage of survival for a female over a male child , a newborn girl is not discriminated against in a fhh in providing necessary care during infancy . survival pattern during infancy by place of residence in mhh and fhh , nfhs-3 ( 2005 - 06 ) , india survival pattern according to sex of the child in mhh and fhh , nfhs-3 ( 2005 - 06 ) , india the study observed that each of the risk factors had significant association with infant death in mhh and fhh as independent predictors . the differences emerged when the risk factors were adjusted for maternal , child and other household characteristics . in mhh , household size , number of eligible women , number of children below five years and economic condition are significant risk factors , when the model is adjusted for household factors . among the maternal factors , increase in education of mother reduced risk of infant death while increase in average number of children born , elevated the hazard of infant death . when adjusted for all the study variables , household size , number of eligible women , number of children below five and child breastfeeding had significant association with infant death . in fhh , economic condition of the household was the only significant risk factor for infant death when adjusted for the household characteristics . again , education of mother and number of children ever born were significant risk factors when adjusted for maternal characteristics . among the child related characteristics , child ever breastfed and antenatal care were significant risk factors for infant death . after adjusting for all the risk factors , breastfeeding status and length of previous birth interval were the only significant risk factors of infant death . the study further observed that the pattern of survival also differ for fhh and mhh . children had higher survival probability at each age in fhh than mhh irrespective of place of residence and sex of the child . thus the paper concludes that the determinants of infant mortality should be examined according to household headship . government should promote fhh and appropriate policies should be formulated to improve economic condition of fhh . fhh report high percentage of home deliver - ies and illiterate women , less exposure to mass media , low economic condition and higher num - ber of children ever born than mhh.amidst all these disadvantages also , children have higher survival probability at each age in fhh than mhh.household size , number of eligible women in the household and number of children below five years of age have higher association with infant mortality in mhh than fhh . fhh report high percentage of home deliver - ies and illiterate women , less exposure to mass media , low economic condition and higher num - ber of children ever born than mhh . amidst all these disadvantages also , children have higher survival probability at each age in fhh than mhh . household size , number of eligible women in the household and number of children below five years of age have higher association with infant mortality in mhh than fhh . | background : there has been ample discussion on the levels and trends of infant mortality in india over time , but what remains less explored are , the differentials in infant mortality according to household headship .
this paper examined the differences in the determinants of infant mortality between male - headed households ( mhh ) and female - headed households ( fhh).methods : the study used cox proportional hazard model to examine the determinants of infant death , and kaplan - meier estimation technique to examine the survival pattern during infancy using data from indian national family health survey ( 2005 - 06 ) .
the analysis is restricted to women who had at least one live birth in the five years preceding the survey.results:the study observed that household size and number of children below five are significant risk factors of infant mortality in mhh while length of previous birth interval is the only significant risk factor of infant death in fhh.conclusions and global health implications : the results indicate that children from fhh have higher survival probability at each age than children from mhh irrespective of place of residence and sex of the child . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
cancer communication can be regarded as an attribute of good - quality care . and , a critically - important arena of cancer outcomes research is quality of care , including ways to disseminate the results of relevant outcomes research to users . increasingly , care is moving from hospitals to ambulatory care settings , to community facilities , and the home . such shifts in the locus of care can have major implications for those for whom access to institutional settings may be restricted , for reasons such as geographical location or socioeconomic status . the increasing private and public availability of internet - connected computers has created opportunities for overcoming some of these barriers to communication in general , and to the dissemination of relevant outcomes research in particular . for example , industry canada has supported programs designed to bring publicly available broadband access to canadian communities , with priority given to first nations , northern , remote and rural communities . these communities are currently either unserved or underserved . at least some forms of breast cancer are increasingly being viewed as a chronic illness , where an emphasis is placed on meeting the various ongoing needs of people living with cancer , their families and other members of their social support networks . models of care have evolved which place greater emphasis on meeting the needs of a particular situation , rather than on the roles of particular health professionals . examples of particular situations may be selected from a very wide range of cancer control activities , including ones designed to provide information and support to those living with cancer , or to those who are at an increased risk of cancer . an aspect of good - quality care that has received increasing recognition is health - related quality of life ( hqol ) . hqol is a dynamic phenomenon , which can change in response not only to various manifestations of a disease , but also to a variety of external influences . like other aspects of cancer outcomes research , research on hqol needs to involve the perspectives of the recipients of care , if only because of the importance of focusing on hqol outcomes not only at the level of populations , but also at the level of individuals . although reliable and valid indicators of hqol are a crucial aspect of cancer outcome studies , appropriate attention also needs to be paid to the transfer of research findings into practical applications . a conceptual framework for cancer control research , developed in canada , was designed to foster a wide range of cancer control research activities . these activities include research related to the development and implementation of interventions and research related to the development and evaluation of programs designed to deliver effective interventions in a well - organized manner . the evaluation of such interventions and programs is an important aspect of outcomes research on quality of care . a variety of psychosocial interventions have been designed to enhance the hqol of cancer patients [ 7 - 9 ] . in this commentary , the focus will be on the evaluation of cancer - oriented support groups , including both face - to - face groups and those designed to provide long - distance support via the internet . a particular emphasis will be placed on evaluation of the " navigator role " in meeting the informational , decisional and educational needs of women with breast cancer . navigation of the care system can be regarded as having four major components : coordination of care , education / information , decision - making and self - care . and , of course , the needs of women will be dependent on their current situation in relation to breast cancer ( e.g. pre - diagnosis but high - risk , post - diagnosis but recurrence - free , post - recurrence with metastases , etc . ) . so , assistance provided in relation to navigation of the various phases along the trajectory of the " cancer journey " also needs to be taken into account in efforts to evaluate the " navigator role " . to cope with their illness , many women participate in breast cancer support groups . in a review published in 2000 , davison et al . assessed participation in a wide variety of disease - related internet - based support groups . they reported that support groups for breast cancer were highly - ranked , as measured by prevalence - adjusted indices of overall online support activity , in comparison with other disease - related and cancer - related on - line support groups that were evaluated . they also reported similar results for disease - related and cancer - related face - to - face support groups in four major metropolitan areas in the usa . a research question is : do internet - based and face - to - face support groups involve similar probable benefits and possible harms ? usually , the support groups that have been studied have been face - to - face ones . one example is a study of 24 women in four community breast cancer self - help groups in ontario , canada . reported benefits of group involvement included both emotional support benefits and informational and practical support benefits . emotional support benefits included connecting with other breast cancer survivors , feeling understood , providing hope , and sharing experiences , including healing laughter . informational and practical support benefits included sharing of important information and " learning how to get what you want " . issues identified as problematic included how to deal with deaths of group members and how to balance the group 's primary purpose of providing mutual support with secondary goals of dealing with group business and engaging in meaningful advocacy . in a recent succinct review of the literature , it 's noted that although there is empiric evidence that community - based support groups are beneficial , not all studies have reported positive outcomes . an important issue is whether or not support groups of any kind might yield benefits not only for hqol , but also for survival . spiegel , in an editorial published in 2001 , has concluded that the literature is divided , and that " group therapy for patients with cancer can be prescribed for its psychological benefit , if not necessarily for any prolongation of survival " . goodwin et al . reviewed the results of trials of various psychosocial interventions , ranging from support groups to individual or couple counseling , and concluded that , rather than an effect on survival , it is more reasonable to expect an impact of such interventions on psychosocial functioning and/or hqol . in summary , research on face - to - face support groups has provided evidence that they usually , but not always , have beneficial impact on hqol . this research provides support for the view that self - help groups , whether online or face - to - face , are most likely to be beneficial when they are focused on their primary purpose of providing mutual support about navigating various phases of cancer . only a few studies of web - based breast cancer support groups have been reported [ 12,14 - 19 ] . , involved line - by - line analysis of postings on prostate , breast , and mixed internet - based cancer support groups . four categories of responses ( information giving / seeking , encouragement / support , personal opinion , and personal experience ) accounted for approximately 80% of responses across the groups . personal experience took priority in the breast group , while information giving / seeking was ranked first in the prostate group . women were more than twice as likely to give encouragement and support , and men were more than twice as likely to give information . some of the advantages and disadvantages of online support groups for breast cancer patients have recently been summarised . advantages include flexibility in the modality of delivery ( e.g. via discussion groups , chat rooms , etc . ) , a variety of facilitation options ( in comparison with face - to - face groups ) , and the comparatively fewer resources required . disadvantages include the need to be able to access and use computers and the internet and to be comfortable in the language which the online group is using . there is also a possibility that participants could become over - reliant on their internet - based relationships , resulting in increased social isolation . this trial is of particular interest , because it provides a good example of issues that are encountered when a rct , a " gold standard " of experimental design , is used for the evaluation of online support groups . in the trial , 72 women with primary breast carcinoma were randomly assigned to a 12-week , web - based , social support group . the support group was semistructured , moderated by a health care professional , and delivered in an asynchronous newsgroup format . it is noteworthy that the involvement of a health professional as the moderator ( or facilitator , or navigator ) , and some aspects of the format and the eligibility of participants , could be regarded as features of this study that were imposed by the rct design . this web - based program was found to be effective in reducing participants ' scores on depression , perceived stress , and cancer - related trauma measures . these authors concluded that web - based social support groups offer many advantages , but cautioned that this delivery mechanism presents some ethical issues that need to be addressed . one ethical issue they identified : how do the participants understand the limits of the moderator 's role ? to deal with this issue , participants were told , before joining the study , that the intervention was a psychoeducational support group , and was not meant to serve as a form of psychotherapy , nor as an alternative to psychotherapy . group members offered each other advice , but the group moderator refrained from doing so . a second ethical issue considered by these authors was the privacy of the participants . participants were able to read personal stories from survivors , share their own experiences , and keep a web - based personal journal . however , the personal journal was closed to review by other group members , and , in this intervention , participants were instructed not to disclose any information that they learned about group members to others , and not to allow family members or friends to access the website . participants were also warned that there are limits to the confidentiality of any web - based intervention , because no internet - connected system can be completely secure from attack by skilled " hackers " . but , the authors pointed out that the most likely threat to the privacy of group participants is their own security practices , such as allowing non - group members to view the website . participants in such groups should be reminded regularly of the limits of confidentiality of internet - based interventions , and should treat their postings as potentially public documents . because there is some ambiguity about the legal responsibilities of moderators of online support groups , these authors also made a conservative decision to restrict eligibility to participants who resided in the same state ( california ) in which the moderators were licensed to practice psychology . although in this rct , the moderator was a health professional ( a psychologist ) , many popular online support groups are peer - moderated , rather than professionally - moderated . in an evaluation of online support groups , different models of the role of the " navigator " need to be taken into account . , in their review of the literature , found that numerous terms were used to describe the role of various kinds of navigators in the health care system , such as case manager , clinical coordinator , support nurse , etc . the term " patient navigator " was rarely used . in the first of two examples that were identified , the patient navigator acted as an advocate for patients with abnormal breast cancer screening findings in underserved areas of the bronx , new york . in the second example , the patient navigator ( " the native sister " ) accompanied native american women to follow - up appointments and provided emotional support and advocacy . a subsequent publication by the latter group describes the native american cancer survivors ' support network , initiated in 1999 . this innovative public health program is designed to improve survival from cancer and the quality of life after a cancer diagnosis for american indians , alaska natives , and canadian aboriginal patients and their loved ones . farber et al . also performed an environmental scan of existing navigator roles in canada . key informants who might have information about the navigator role were identified , and 74/186 of these key informants were interviewed . on the basis of this study of the status of the navigator role for meeting the needs of women with breast cancer in canada , three conceptual models were identified . in the active coordination model , the navigator is actively involved in helping affected individuals to navigate a way through the care system . in the facilitating navigator model , the shared or tacit model involves several people providing navigation , either tacitly , or by design . approximately 38% of the key informants were located in rural or semi - rural communities . the only difference noted was that navigators in smaller communities might be more involved in transportation arrangements for patients ( e.g. if the needed facilities were n't located within the community ) . key informants were also asked what they believed , on the basis of their knowledge and experience , should be the key evaluation factors for the navigator role . generally , three types of outcome data were identified : workload indicators ( such as number of calls received ) , indicators of patient / client satisfaction ( such as meeting patient / client expectations ) and indicators of more systemic evaluation issues ( such as measures of cost - effectiveness ) . might these same conceptual models for the navigator role , and these same approaches to their evaluation , also be applied to internet - based support groups ? for example , perhaps the role of the moderator in the web - based support program described by winzelberg et al . could be regarded as an online version of the facilitating navigator model of farber et al . ? in the next section , another example of an internet - based support group ( the breast - cancer mailing list ) will be considered briefly . the bcml was established in early 1994 , and has operated continuously since that time . it 's an unmoderated english - speaking online ( via email ) discussion group , based at memorial university of newfoundland in st . although it is based in canada , the majority of members are from the usa , with the remainder from canada and other countries around the world . the volume of mail , often nearing a hundred messages a day , is sometimes a cause for complaint at least until new members learn ways of prioritising their email . on the other hand , the number of members , and their level of activity on the bcml , is sufficient to ensure that new subscribers are likely to be able to find other people who are , or have recently been , in a very similar situation . in addition , because different members of the bcml have already experienced different phases along the trajectory of the " cancer journey " , they can serve as " tacit peer - navigators " , and help new members to find their way through the various phases of the journey . also , a website for the bcml is available . statistical information about the frequency and types of visits to these archives have not been provided via the website , but it seems likely that present and former list members ( and perhaps others ) visit and use them frequently . so , messages found in such archives clearly should be regarded as public documents , even though this particular small segment of cyberspace is likely to be of much interest only to those directly involved in some way with breast cancer . the use of online support groups such as the bcml ( and their archives ) for research purposes raises ethical issues of privacy and informed consent that need to be considered carefully . the members of disease - related mailing lists of this kind can freely choose whether or not to participate in an online support group where their postings are potentially public documents . but , new subscribers may join and participate in a mailing list before they become fully informed about its various features . and , their particular personal situations may involve a vulnerability that is much greater than that of participants in online discussion groups that are not health - related . the more vulnerable the participants , the greater the need for careful attention to privacy and confidentiality . for example , the archives of a mailing list for patients with metastatic breast cancer are password - protected . for an unmoderated mailing list like the bcml , concerns about the limits of the moderator 's role are replaced by different issues . one example of such an issue : which members of an unmoderated list tacitly play " peer - navigator " roles ( as in the shared or tacit model of farber et al . ) ? the hypothesis is that good - quality unmoderated online support groups can be characterized as ones where respected long - time ( " veteran " ) members serve as " tacit peer - navigators " and become quite skilled in this role . an example is provided by musa mayer , a " veteran " of the bcml and a frequent contributor . she is the author of three books about breast cancer , and a respected breast cancer activist . her perspectives have been influenced by her experiences with counselling , with her own breast cancer , and with the bcml . to test this hypothesis , it should be feasible to collect examples of the three types of outcome data that were identified by farber et al . as appropriate for an evaluation of the navigator role . an example of a workload indicator might be the number of navigation - oriented messages posted ( e.g. per week ) by list members tentatively identified as " tacit peer - navigators " . an indicator of patient / client satisfaction might be the number of spontaneously - posted messages that express satisfaction with the mailing list in general , and with the contributions of " tacit peer - navigators " in particular . an example of an unsolicited quote from a posting to the bcml : " this group is at its best when they can help someone going through a rough time . you see , we have all been there and understand the fears , nervousness and what ever else plagues us at times like these " . it might also be feasible to develop indicators of more systemic evaluation issues , such as measures of cost - effectiveness . for example , because subscriptions to the bcml are free , and the " tacit peer - navigators " are volunteers , the navigator role is fulfilled at no extra cost to list members . so , the main systems evaluation issue remains one of assessing the effectiveness of online support groups , in comparison with each other , and with more conventional face - to - face support groups . it seems reasonable to assume that online support groups can be regarded as a complement to face - to - face support groups ( when the latter are available ) and as an alternative when they are not . whether or not this assumption is correct , another important systems evaluation issue is : how best to evaluate the effectiveness of the navigator role ? it seems likely that the greater the effectiveness of the navigator role , the greater the cost - effectiveness of the support group , whether or not it 's online . from this perspective , systemic evaluation outcome variables , and especially ones focused on the navigator role , may be the most crucial ones for which appropriate measures need to designed and implemented . to cope with their illness , many women participate in breast cancer support groups . in a review published in 2000 , davison et al . assessed participation in a wide variety of disease - related internet - based support groups . they reported that support groups for breast cancer were highly - ranked , as measured by prevalence - adjusted indices of overall online support activity , in comparison with other disease - related and cancer - related on - line support groups that were evaluated . they also reported similar results for disease - related and cancer - related face - to - face support groups in four major metropolitan areas in the usa . a research question is : do internet - based and face - to - face support groups involve similar probable benefits and possible harms ? usually , the support groups that have been studied have been face - to - face ones . one example is a study of 24 women in four community breast cancer self - help groups in ontario , canada . reported benefits of group involvement included both emotional support benefits and informational and practical support benefits . emotional support benefits included connecting with other breast cancer survivors , feeling understood , providing hope , and sharing experiences , including healing laughter . informational and practical support benefits included sharing of important information and " learning how to get what you want " . issues identified as problematic included how to deal with deaths of group members and how to balance the group 's primary purpose of providing mutual support with secondary goals of dealing with group business and engaging in meaningful advocacy . in a recent succinct review of the literature , it 's noted that although there is empiric evidence that community - based support groups are beneficial , not all studies have reported positive outcomes . an important issue is whether or not support groups of any kind might yield benefits not only for hqol , but also for survival . spiegel , in an editorial published in 2001 , has concluded that the literature is divided , and that " group therapy for patients with cancer can be prescribed for its psychological benefit , if not necessarily for any prolongation of survival " . goodwin et al . reviewed the results of trials of various psychosocial interventions , ranging from support groups to individual or couple counseling , and concluded that , rather than an effect on survival , it is more reasonable to expect an impact of such interventions on psychosocial functioning and/or hqol . in summary , research on face - to - face support groups has provided evidence that they usually , but not always , have beneficial impact on hqol . this research provides support for the view that self - help groups , whether online or face - to - face , are most likely to be beneficial when they are focused on their primary purpose of providing mutual support about navigating various phases of cancer . only a few studies of web - based breast cancer support groups have been reported [ 12,14 - 19 ] . , involved line - by - line analysis of postings on prostate , breast , and mixed internet - based cancer support groups . four categories of responses ( information giving / seeking , encouragement / support , personal opinion , and personal experience ) accounted for approximately 80% of responses across the groups . personal experience took priority in the breast group , while information giving / seeking was ranked first in the prostate group . women were more than twice as likely to give encouragement and support , and men were more than twice as likely to give information . some of the advantages and disadvantages of online support groups for breast cancer patients have recently been summarised . advantages include flexibility in the modality of delivery ( e.g. via discussion groups , chat rooms , etc . ) , a variety of facilitation options ( in comparison with face - to - face groups ) , and the comparatively fewer resources required . disadvantages include the need to be able to access and use computers and the internet and to be comfortable in the language which the online group is using . there is also a possibility that participants could become over - reliant on their internet - based relationships , resulting in increased social isolation . this trial is of particular interest , because it provides a good example of issues that are encountered when a rct , a " gold standard " of experimental design , is used for the evaluation of online support groups . in the trial , 72 women with primary breast carcinoma were randomly assigned to a 12-week , web - based , social support group . the support group was semistructured , moderated by a health care professional , and delivered in an asynchronous newsgroup format . it is noteworthy that the involvement of a health professional as the moderator ( or facilitator , or navigator ) , and some aspects of the format and the eligibility of participants , could be regarded as features of this study that were imposed by the rct design . this web - based program was found to be effective in reducing participants ' scores on depression , perceived stress , and cancer - related trauma measures . these authors concluded that web - based social support groups offer many advantages , but cautioned that this delivery mechanism presents some ethical issues that need to be addressed . one ethical issue they identified : how do the participants understand the limits of the moderator 's role ? to deal with this issue , participants were told , before joining the study , that the intervention was a psychoeducational support group , and was not meant to serve as a form of psychotherapy , nor as an alternative to psychotherapy . group members offered each other advice , but the group moderator refrained from doing so . a second ethical issue considered by these authors was the privacy of the participants . participants were able to read personal stories from survivors , share their own experiences , and keep a web - based personal journal . however , the personal journal was closed to review by other group members , and , in this intervention , participants were instructed not to disclose any information that they learned about group members to others , and not to allow family members or friends to access the website . participants were also warned that there are limits to the confidentiality of any web - based intervention , because no internet - connected system can be completely secure from attack by skilled " hackers " . but , the authors pointed out that the most likely threat to the privacy of group participants is their own security practices , such as allowing non - group members to view the website . participants in such groups should be reminded regularly of the limits of confidentiality of internet - based interventions , and should treat their postings as potentially public documents . because there is some ambiguity about the legal responsibilities of moderators of online support groups , these authors also made a conservative decision to restrict eligibility to participants who resided in the same state ( california ) in which the moderators were licensed to practice psychology . although in this rct , the moderator was a health professional ( a psychologist ) , many popular online support groups are peer - moderated , rather than professionally - moderated . in an evaluation of online support groups , different models of the role of the " navigator " need to be taken into account . farber at al . , in their review of the literature , found that numerous terms were used to describe the role of various kinds of navigators in the health care system , such as case manager , clinical coordinator , support nurse , etc . the term " patient navigator " was rarely used . in the first of two examples that were identified , the patient navigator acted as an advocate for patients with abnormal breast cancer screening findings in underserved areas of the bronx , new york . in the second example , the patient navigator ( " the native sister " ) accompanied native american women to follow - up appointments and provided emotional support and advocacy . a subsequent publication by the latter group describes the native american cancer survivors ' support network , initiated in 1999 . this innovative public health program is designed to improve survival from cancer and the quality of life after a cancer diagnosis for american indians , alaska natives , and canadian aboriginal patients and their loved ones . farber et al . also performed an environmental scan of existing navigator roles in canada . key informants who might have information about the navigator role were identified , and 74/186 of these key informants were interviewed . on the basis of this study of the status of the navigator role for meeting the needs of women with breast cancer in canada , three conceptual models were identified . in the active coordination model , the navigator is actively involved in helping affected individuals to navigate a way through the care system . in the facilitating navigator model , the shared or tacit model involves several people providing navigation , either tacitly , or by design . approximately 38% of the key informants were located in rural or semi - rural communities . the only difference noted was that navigators in smaller communities might be more involved in transportation arrangements for patients ( e.g. if the needed facilities were n't located within the community ) . key informants were also asked what they believed , on the basis of their knowledge and experience , should be the key evaluation factors for the navigator role . generally , three types of outcome data were identified : workload indicators ( such as number of calls received ) , indicators of patient / client satisfaction ( such as meeting patient / client expectations ) and indicators of more systemic evaluation issues ( such as measures of cost - effectiveness ) . might these same conceptual models for the navigator role , and these same approaches to their evaluation , also be applied to internet - based support groups ? for example , perhaps the role of the moderator in the web - based support program described by winzelberg et al . could be regarded as an online version of the facilitating navigator model of farber et al . ? in the next section , another example of an internet - based support group ( the breast - cancer mailing list ) will be considered briefly . the bcml was established in early 1994 , and has operated continuously since that time . it 's an unmoderated english - speaking online ( via email ) discussion group , based at memorial university of newfoundland in st . although it is based in canada , the majority of members are from the usa , with the remainder from canada and other countries around the world . the volume of mail , often nearing a hundred messages a day , is sometimes a cause for complaint at least until new members learn ways of prioritising their email . on the other hand , the number of members , and their level of activity on the bcml , is sufficient to ensure that new subscribers are likely to be able to find other people who are , or have recently been , in a very similar situation . in addition , because different members of the bcml have already experienced different phases along the trajectory of the " cancer journey " , they can serve as " tacit peer - navigators " , and help new members to find their way through the various phases of the journey . also , a website for the bcml is available . statistical information about the frequency and types of visits to these archives have not been provided via the website , but it seems likely that present and former list members ( and perhaps others ) visit and use them frequently . so , messages found in such archives clearly should be regarded as public documents , even though this particular small segment of cyberspace is likely to be of much interest only to those directly involved in some way with breast cancer . the use of online support groups such as the bcml ( and their archives ) for research purposes raises ethical issues of privacy and informed consent that need to be considered carefully . the members of disease - related mailing lists of this kind can freely choose whether or not to participate in an online support group where their postings are potentially public documents . but , new subscribers may join and participate in a mailing list before they become fully informed about its various features . and , their particular personal situations may involve a vulnerability that is much greater than that of participants in online discussion groups that are not health - related . the more vulnerable the participants , the greater the need for careful attention to privacy and confidentiality . for example , the archives of a mailing list for patients with metastatic breast cancer are password - protected . for an unmoderated mailing list like the bcml , concerns about the limits of the moderator 's role are replaced by different issues . one example of such an issue : which members of an unmoderated list tacitly play " peer - navigator " roles ( as in the shared or tacit model of farber et al . ) ? the hypothesis is that good - quality unmoderated online support groups can be characterized as ones where respected long - time ( " veteran " ) members serve as " tacit peer - navigators " and become quite skilled in this role . an example is provided by musa mayer , a " veteran " of the bcml and a frequent contributor . she is the author of three books about breast cancer , and a respected breast cancer activist . her perspectives have been influenced by her experiences with counselling , with her own breast cancer , and with the bcml . to test this hypothesis , it should be feasible to collect examples of the three types of outcome data that were identified by farber et al . as appropriate for an evaluation of the navigator role . an example of a workload indicator might be the number of navigation - oriented messages posted ( e.g. per week ) by list members tentatively identified as " tacit peer - navigators " . an indicator of patient / client satisfaction might be the number of spontaneously - posted messages that express satisfaction with the mailing list in general , and with the contributions of " tacit peer - navigators " in particular . an example of an unsolicited quote from a posting to the bcml : " this group is at its best when they can help someone going through a rough time . you see , we have all been there and understand the fears , nervousness and what ever else plagues us at times like these " . it might also be feasible to develop indicators of more systemic evaluation issues , such as measures of cost - effectiveness . for example , because subscriptions to the bcml are free , and the " tacit peer - navigators " are volunteers , the navigator role is fulfilled at no extra cost to list members . so , the main systems evaluation issue remains one of assessing the effectiveness of online support groups , in comparison with each other , and with more conventional face - to - face support groups . it seems reasonable to assume that online support groups can be regarded as a complement to face - to - face support groups ( when the latter are available ) and as an alternative when they are not . whether or not this assumption is correct , another important systems evaluation issue is : how best to evaluate the effectiveness of the navigator role ? it seems likely that the greater the effectiveness of the navigator role , the greater the cost - effectiveness of the support group , whether or not it 's online . from this perspective , systemic evaluation outcome variables , and especially ones focused on the navigator role , may be the most crucial ones for which appropriate measures need to designed and implemented . application of the concept of a " navigator role " to support groups in general , and to unmoderated online ones like the bcml in particular , has received little or no attention in the research literature . the navigator role should be taken into account in outcome - oriented research on this increasingly important aspect of cancer communication . bcml : breast - cancer mailing list hqol : health - related quality of life rct : randomized controlled trial partial support for this work was provided by a ncic / eli lilly award from the national cancer institute of canada ( ncic ) . | backgroundat least some forms of breast cancer are increasingly being viewed as a chronic illness , where an emphasis is placed on meeting the various ongoing needs of people living with cancer , their families and other members of their social support networks .
this commentary outlines some approaches to the evaluation of cancer - related support groups , with a particular emphasis on those designed to provide long - distance support , via the internet , for women with breast cancer.discussionthe literature on evaluations of community - based cancer support groups indicates that they offer a number of benefits , and that it is more reasonable to expect an impact of such interventions on psychosocial functioning and/or health - related quality of life than on survival .
the literature on both face - to - face and online social support groups suggests that they offer many advantages , although evaluation of the latter delivery mechanism presents some ethical issues that need to be addressed .
many popular online support groups are peer - moderated , rather than professionally - moderated . in an evaluation of online support groups , different models of the role of the " navigator " need to be taken into account .
some conceptual models are outlined for the evaluation of the " navigator role " in meeting the informational , decisional and educational needs of women with breast cancer .
the breast - cancer mailing list , an example of an unmoderated internet - based peer - support group , is considered within the context of a shared or tacit model of the navigator role.conclusionapplication of the concept of a " navigator role " to support groups in general , and to unmoderated online ones in particular , has received little or no attention in the research literature .
the navigator role should be taken into account in research on this increasingly important aspect of cancer communication . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
total joint arthroplasty is performed with objectives of providing pain relief and better quality of life to the patients suffering from arthritis of the hip and knee joints.1 rarely , serious complications including vascular injury can occur during the procedure , which may affect the outcome of surgery.123 the incidence of vascular injury in primary total hip arthroplasty ( tha ) is approximately 0.04%.4 we report an unusual case of vascular injury during preparation of the acetabulum in primary tha . a 72-year - old male presented with severe arthritis of left hip secondary to avascular necrosis of the femoral head . preoperative radiograph showed presence of a large osteophyte at the floor of the acetabulum [ figure 1a ] . a cementless total hip replacement with modified hardinge 's approach was performed using large head metal on metal articulation with articular surface replacement cup ( voluntary recall since august 2010 ) , unipolar metal head and corail femoral stem ( depuy orthopaedics , inc . , intraoperatively , a 3.2 mm drill was used to identify the floor of the acetabulum and the depth of reaming . two hours later , the patient developed unexplained hypotension with tachycardia , which was treated with colloids and 2 units of blood . abdomen and pelvis ultrasound examination conducted for investigating the cause of pain revealed a large ( 18 cm 10 cm 7 cm ) retroperitoneal hematoma . four units of fresh frozen plasma ( ffp ) and 6 units of platelets were administered for the management of hematoma . a lobulated pseudoaneurysm of 2.2 cm 1.2 cm size , arising from one of the branches of the left internal iliac artery [ figure 2a ] was reported in an emergency ct angiography . subsequently , the patient developed acute renal failure , which required hemodialysis and inotrope support for 10 days . on 19 day , the patient was discharged from the hospital with complete recovery and full weight bearing . a total of 16 units of blood , 4 units of ffp and 6 units of platelets were required for the treatment during his hospital stay . at similarly , no evidence of pseudoaneurysm was detected on repeat computed tomography ( ct ) angiography [ figure 2c ] . ( a ) preoperative radiograph of pelvis with both hips showing a large osteophyte at the floor of the acetabulum in the left hip ( b ) postoperative anteroposterior radiograph showing left hip arthroplasty ( a ) computed tomography angiography showing pseudoaneurysm of a tributary of left internal iliac artery ( b ) computed tomography angiography arrow showing post coil - embolisation of the pseudoaneurysm ( c ) six years followup of computed tomography angiography showing no pseudoaneurysm modern total joint arthroplasty is a successful and safe surgery ; however there are chances of limb or life threatening vascular complications.1 vascular injury is relatively uncommon during primary tha compared to total knee arthroplasty and revision tha . indirect vessel injury can be the result of various causes including mechanical stretching , maneuvering of the limb and thermal injury from bone cement.1567 the common causes of vascular injury in tha are inadvertent placement of acetabular screws,891011 thermal injury due to bone cement as well as fixation of intraoperative periprosthetic fractures , embolization or injury to the vessel wall during surgical approach or maneuvering of the limb.48 in our case , vascular injury was not caused by any of the above mentioned mechanisms , but occurred during preparation of the acetabulum by a depth assessment from a drill hole . medial acetabular osteophytes can result in lateralization of the cup which may affect abductor mechanics . these range from medial reaming to identify the pulvinar or using a power drill to make a small hole through the medial wall gradually and use a depth gauge to assess the bone thickness.12 we used a 3.2 mm drill bit to assist in placing the acetabular component in the optimum position . we believe that the vascular injury to the branch of internal iliac artery was probably caused by this technique which might explain the late presentation of symptoms of blood loss and formation of pseudoaneurysm [ figure 3 ] . the internal iliac artery is 4 cm long , begins at the common iliac bifurcation , level with the lumbosacral intervertebral disc and anterior to the sacroiliac joint . it descends posteriorly to the superior margin of the greater sciatic foramen where it divides into an anterior trunk , which continues in the same line towards the ischial spine and a posterior trunk , which passes back to the greater sciatic foramen . common iliac artery is most commonly involved ( 70% ) , next the internal iliac artery is involved ( 25% ) . iliac artery aneurysms are bilateral in approximately 30% cases.1314 line diagram of the iliac vessels showing how the drill bit may have injured the branch of the internal iliac artery in the previous case series on vascular injury during tha , the injury is reported to affect the external iliac vein and artery or profunda femoris artery , which was treated by open surgery and vascular repair.14 however , in our case , injury occurred to internal iliac artery . we advice surgeons to be extremely careful while drilling medial wall of the acetabulum or use some other technique to identify the true acetabular floor . similarly , close postoperative surveillance is required for diagnosing the vascular injury which can be successfully managed with aggressive multidisciplinary approach . | vascular injury is one of the rare complications of primary total hip arthroplasty ( tha ) .
we report an unusual case of lobulated pseudoaneurysm arising from one of the branches of the left internal iliac artery during acetabulum preparation in tha , which was successfully treated with coil embolization and multidisciplinary care .
after 6 years follow up , patient did not have any symptoms related to the hip replacement .
we recommend that surgeons should be extremely cautious while drilling medial wall of the acetabulum for depth assessment .
aggressive multidisciplinary approach , including possible support from an interventional radiologist is required for the treatment of such vascular injuries . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
improvements in hemodynamics parameters after the procedure have been demonstrated in several reports.5 , 6 , 7 , 8 , 9 however , few reports have investigated changes in ecg parameters along with the prevalence and type of arrhythmias postppvi.10 , 11
the goals of this study are to ( 1 ) describe the shortterm and mediumterm changes to the electrophysiologic substrate ( as identified by surface ecg ) in patients following ppvi and ( 2 ) determine the type and prevalence of arrhythmias ( including premature ventricular complexes [ pvcs ] , nonsustained ventricular tachycardia [ nsvt ] , and atrial fibrillation / atrial flutter [ af / afl ] ) in postppvi patients . louis school of medicine ( waiver of consent ) , we undertook a retrospective chart review of patients who underwent ppvi in the pulmonary valve position at st . the previously described inclusion criteria for ppvi include ( 1 ) rv systolic pressure greater than two thirds of systemic with symptoms or ( 2 ) rv systolic pressure greater than three fourths of systemic in absence of symptoms or ( 3 ) moderate / severe pulmonary regurgitation with either symptoms or severe rv dysfunction or severe rv dilatation or impaired exercise capacity.5 , 12 patients were excluded if the melody valve ( medtronic inc , minneapolis , mn ) was implanted in the branch pulmonary arteries or tricuspid position ( n=4 ) . patients were classified as having predominantly pulmonary insufficiency ( pi ) if they had at least moderate pulmonary regurgitation on echocardiography ; they were classified as having predominantly pulmonary stenosis ( ps ) if they had a mean rvot gradient 35 mm hg on echocardiogram . when the gradient could not be determined due to poor acoustic windows , a systolic gradient 35 mm hg during the catheterization was used as the inclusion criterion for the pulmonary stenosis group . patients with both stenosis and insufficiency were included in the mixedlesion group ( ps / pi ) . charts were then reviewed for demographic information , clinical and procedural characteristics , and electrophysiologic data ( including ecgs , 24hour holter monitors , electrophysiology studies [ eps ] , cardiac rhythm device reports and transmissions , and telemetry ) . ecgs preprocedure , 24 hours postprocedure , and at last followup visit were analyzed . postprocedural overnight telemetry alarms and holter monitors pre and postprocedure were reviewed for ecg changes and arrhythmia . right bundle branch block ( rbbb ) was defined as a qrs duration of 120 milliseconds . nsvt was defined as 3 or more consecutive pvcs with rates faster than 180 beats per minute . continuous data are not normally distributed and therefore are presented as median with interquartile range . the fisher exact test was used to compare nominal variables between pre and postppvi groups . all statistical analyses were performed using spss statistical software ( version 22.0 ; ibm corp , armonk , ny ) . continuous data are not normally distributed and therefore are presented as median with interquartile range . the fisher exact test was used to compare nominal variables between pre and postppvi groups . all statistical analyses were performed using spss statistical software ( version 22.0 ; ibm corp , armonk , ny ) . a total of 106 patients , 40 female ( 38% ) and 66 male ( 62% ) , were included in this study . median age at the time of the procedure was 20.7 years ( iqr 14.531.2 ) . the most common congenital anatomic diagnosis was tetralogy of fallot ( tof ) ( 55% ) , and the most common type of rvot dysfunction was pulmonary insufficiency ( 57% ) . prior to the procedure , 10 ( 9% ) patients had undergone eps , 3 of which included an ablation attempt . all the eps with ablation attempts occurred more than 3 years prior to the ppvi . implanted cardiac rhythm devices were present in 16 ( 15% ) patients ( table 1 ) . this patient did not have documented arrhythmia or st / t changes pre or postppvi . the second patient developed endocarditis of the melody valve 2 years postimplant that was being treated with intravenous antibiotics . patient clinical and demographic data ace indicates angiotensinconverting enzyme ; arb , angiotensin ii receptor blocker ; phtn , pulmonary hypertension ; rvot , right ventricular outflow tract . a total of 46 patients had pulmonary stenosis ( 26 ps patients and 20 ps / pi patients ) with a median preppvi rvot gradient of 42 mm hg ( iqr 3750 ) . median rvot gradient significantly lowered immediately postppvi to 11 mm hg ( iqr 815 ) ( p<0.001 ) . the median rvot gradient remained stable at 11.5 mm hg ( iqr 719 ) ( figure 1 ) . of the pi patients , only 1 patient had more than a mild degree of pulmonary regurgitation at last followup . sixteen patients with cardiac rhythm devices were excluded from this portion of the data analysis . there was no difference between the pre and postppvi median qrsd ( 141 [ iqr 110157 ] milliseconds vs ( 140 [ iqr 101157 ] milliseconds , p=0.07 ) . the baseline median qtc was 460 [ iqr 436486 ] milliseconds , which lengthened immediately postppvi ( 475 [ iqr 450504 ] milliseconds , p<0.01 ) . mediumterm followup ecgs were available in 58 patients with median followup time of 28 months ( 763 months ) . at followup time of at least 7 months postppvi , there was no statistical significance in the median qrsd ( preppvi=137 [ iqr 107155 ] milliseconds , postppvi=137 [ iqr 98154 ] milliseconds , followup=137 [ iqr 103157 ] milliseconds , p=0.8 ) . the median qtc shortened significantly to 442 [ iqr 420465 ] milliseconds at followup when compared to preppvi ( 463 [ iqr 435483 ] milliseconds , p<0.001 ) and immediately postppvi ( 476 [ iqr 452507 ] milliseconds , p<0.001 ) ( figure 2 ) . subgroup analysis by cardiac diagnosis or type of rvot dysfunction yielded similar results ( table 2 ) . interestingly , the median preppvi qtc of patients with pulmonary stenosis was shorter than that of patients with pulmonary regurgitation ( p=0.07 ) , whereas the median postppvi qtc of patients with pulmonary stenosis was longer than that of patients with pulmonary regurgitation ( p<0.001 ) . b , corrected qt interval ( qtc , calculated using the bazett formula ) compared preppvi , postppvi , and at followup . the qtc was prolonged immediately postppvi , but it significantly shortened at followup ( p<0.01 ) . ppvi indicates percutaneous pulmonary valve implantation ; qrsd , qrs duration ; qtc , corrected qt interval . ecg parameter changes by cardiac diagnosis and type of rvot dysfunction there was no statistical significance in the qrsd across time points regardless of cardiac diagnosis or type of rvot dysfunction . the qtc prolonged significantly immediately postppvi but shortened significantly at followup in all subgroups . ecg indicates electrocardiogram ; ppvi , percutaneous pulmonary valve implantation ; ps / pi pulmonary stenosis / pulmonary insufficiency ; rvot right ventricular outflow tract . at baseline , 23/90 ( 26% ) patients had st / t changes : 5 patients ( 6% ) had st depression , 7 patients ( 8% ) had twave inversion , and 11 patients ( 12% ) had nonspecific st / t changes . immediately postppvi , there were newonset st depression ( n=1 ) , twave inversion ( n=5 ) , and nonspecific st / t changes ( n=5 ) . overall , st / t changes resolved in 100% of cases with st depression , 88% of cases with twave inversion , and 80% of cases with st / t changes ( figures 3 , 4 through 5 ; figure s1 ) . immediately postprocedure , there was resolution of st depression in 2 ( 40% ) patients , while 3 ( 60% ) patients continued to have unchanged st depression . immediately postprocedure , there was resolution of twave inversion in 4 ( 57% ) patients , while 3 ( 43% ) patients continued to have unchanged twave inversion . patients who had resolved twave inversion postppvi did not have further ecgs at followup . additionally , 5 patients had a new finding of twave inversion on ecg . all newonset twave inversions and 2 of 3 persistent twave inversions postprocedure resolved at followup . immediately postprocedure , there was resolution of st / t changes in 3 ( 27% ) patients , while 8 ( 73% ) patients continued to have unchanged st / t changes . patients who had resolved st / t changes postppvi did not have further ecgs at followup . additionally , 5 patients had a new finding of st / t changes on ecg . all newonset st / t changes and 3 of 8 ( 38% ) persistent st / t changes postprocedure resolved at followup . two patients with persistent postprocedure st / t changes continued to have st / t changes at followup . finally , 1 patient had new st / t changes at followup . of note , nine patients ( 8% ) had nsvt , 6 patients ( 6% ) had frequent pvcs , 10 patients ( 10% ) had atrial af / afl , and 2 patients ( 2% ) had supraventricular tachycardia . only 2/8 ( 25% ) patients with preppvi vt , and no patient with preppvi af / afl was in the ps group . newonset arrhythmias immediately postppvi include 7 patients with nsvt , 8 patients with pvcs , and 1 patient with af / afl . once again , there were very few psonly patients ( 3/16 , 19% ) in this cohort . at followup , two out of 8 patients with newonset pvcs were started on blockers , which were discontinued at their 6month followup . one of these patients was taken off the amiodarone at 6 months of followup . one patient was started on a blocker and continued to be on this medication because he had persistent documented pvcs . overall , 12/16 ( 75% ) patients no longer had documented nsvt , and 12/14 ( 86% ) patients no longer had pvcs . finally , af / afl resolved in 6/11 ( 55% ) cases ( figures 6 , 7 through 8 ; figures s2 and s3 ) . arrhythmia burden was determined by holter monitoring , internal rhythm devices reports , and telemetry during subsequent hospitalizations . documented nonsustained ventricular tachycardia ( nsvt ) . prior to procedure , of 9 patients who previously had nsvt preprocedure continued to have nsvt , 1 of 9 patients no longer had nsvt but had pvcs , and 4 of 9 patients no longer had nsvt . among the patients with newonset nsvt postprocedure , 6 of 7 no longer had nonsustained vt , while 1 patient had pvcs . immediately postprocedure , there was resolution of pvcs in 4 ( 64% ) patients , while 1 ( 17% ) patient continued to have persistent pvcs , and 1 ( 17% ) patient had nonsustained ventricular tachycardia ( nsvt ) . seven out of 8 patients with newonset pvcs and both of the persistent pvcs postprocedure no longer had any arrhythmia at followup . one patient with newonset pvcs and the patient with persistent pvcs immediately postprocedure continued to have pvcs at followup . nsvt indicates nonsustained ventricular tachycardia ; ppvi , percutaneous pulmonary valve implantation ; pvcs , premature ventricular contractions . one patient had newonset af / afl immediately postprocedure . at followup , 5 patients continued to have af / afl ( 4 preprocedure and 1 new onset ) , while 6 patients no longer had af / afl . af / afl indicates atrial fibrillation / atrial flutter ; ppvi , percutaneous pulmonary valve implantation . forty patients had cardiac mri preppvi , but only 4 had postppvi studies for comparison ( 3 ps , 1 pi ) . this patient had nonspecific st changes both pre and postppvi , but these changes resolved at followup . the remaining 2 patients did not have ecg changes or appreciable structural changes on followup mri . of the 39 patients with ecg changes ( baseline + new onset postppvi + new onset at followup ) , 27 patients had ecg for analysis ( figures 5 , 6 through 7 ) . specifically , there were 19 resolved st / t changes , 3 persistent st / t changes , and 5 newonset st / t changes . although there was improvement in rv size by echocardiogram in 15/27 ( 56% ) patients , there was no change noted in 12/27 ( 44% ) patients . among patients with newonset / persistent ecg changes at followup ( n=8 ) , there were equal numbers of patients who had improved rv size versus no appreciable change by followup echo . in the 19 patients who had resolved ecg changes at followup , 10 patients had improved rv size , whereas 9 patients did show changes on followup echo . a total of 106 patients , 40 female ( 38% ) and 66 male ( 62% ) , were included in this study . median age at the time of the procedure was 20.7 years ( iqr 14.531.2 ) . the most common congenital anatomic diagnosis was tetralogy of fallot ( tof ) ( 55% ) , and the most common type of rvot dysfunction was pulmonary insufficiency ( 57% ) . prior to the procedure , 10 ( 9% ) patients had undergone eps , 3 of which included an ablation attempt . all the eps with ablation attempts occurred more than 3 years prior to the ppvi . implanted cardiac rhythm devices were present in 16 ( 15% ) patients ( table 1 ) . this patient did not have documented arrhythmia or st / t changes pre or postppvi . the second patient developed endocarditis of the melody valve 2 years postimplant that was being treated with intravenous antibiotics . patient clinical and demographic data ace indicates angiotensinconverting enzyme ; arb , angiotensin ii receptor blocker ; phtn , pulmonary hypertension ; rvot , right ventricular outflow tract . a total of 46 patients had pulmonary stenosis ( 26 ps patients and 20 ps / pi patients ) with a median preppvi rvot gradient of 42 mm hg ( iqr 3750 ) . median rvot gradient significantly lowered immediately postppvi to 11 mm hg ( iqr 815 ) ( p<0.001 ) . the median rvot gradient remained stable at 11.5 mm hg ( iqr 719 ) ( figure 1 ) . of the pi patients , only 1 patient had more than a mild degree of pulmonary regurgitation at last followup . sixteen patients with cardiac rhythm devices were excluded from this portion of the data analysis . there was no difference between the pre and postppvi median qrsd ( 141 [ iqr 110157 ] milliseconds vs ( 140 [ iqr 101157 ] milliseconds , p=0.07 ) . the baseline median qtc was 460 [ iqr 436486 ] milliseconds , which lengthened immediately postppvi ( 475 [ iqr 450504 ] milliseconds , p<0.01 ) . mediumterm followup ecgs were available in 58 patients with median followup time of 28 months ( 763 months ) . at followup time of at least 7 months postppvi , there was no statistical significance in the median qrsd ( preppvi=137 [ iqr 107155 ] milliseconds , postppvi=137 [ iqr 98154 ] milliseconds , followup=137 [ iqr 103157 ] milliseconds , p=0.8 ) . the median qtc shortened significantly to 442 [ iqr 420465 ] milliseconds at followup when compared to preppvi ( 463 [ iqr 435483 ] milliseconds , p<0.001 ) and immediately postppvi ( 476 [ iqr 452507 ] milliseconds , p<0.001 ) ( figure 2 ) . subgroup analysis by cardiac diagnosis or type of rvot dysfunction yielded similar results ( table 2 ) . interestingly , the median preppvi qtc of patients with pulmonary stenosis was shorter than that of patients with pulmonary regurgitation ( p=0.07 ) , whereas the median postppvi qtc of patients with pulmonary stenosis was longer than that of patients with pulmonary regurgitation ( p<0.001 ) . a , qrs duration ( qrsd ) is compared preppvi , postppvi , and at followup . b , corrected qt interval ( qtc , calculated using the bazett formula ) compared preppvi , postppvi , and at followup . the qtc was prolonged immediately postppvi , but it significantly shortened at followup ( p<0.01 ) . ppvi indicates percutaneous pulmonary valve implantation ; qrsd , qrs duration ; qtc , corrected qt interval . ecg parameter changes by cardiac diagnosis and type of rvot dysfunction there was no statistical significance in the qrsd across time points regardless of cardiac diagnosis or type of rvot dysfunction . ecg indicates electrocardiogram ; ppvi , percutaneous pulmonary valve implantation ; ps / pi pulmonary stenosis / pulmonary insufficiency ; rvot right ventricular outflow tract . at baseline , 23/90 ( 26% ) patients had st / t changes : 5 patients ( 6% ) had st depression , 7 patients ( 8% ) had twave inversion , and 11 patients ( 12% ) had nonspecific st / t changes . immediately postppvi , there were newonset st depression ( n=1 ) , twave inversion ( n=5 ) , and nonspecific st / t changes ( n=5 ) . overall , st / t changes resolved in 100% of cases with st depression , 88% of cases with twave inversion , and 80% of cases with st / t changes ( figures 3 , 4 through 5 ; figure s1 ) . immediately postprocedure , there was resolution of st depression in 2 ( 40% ) patients , while 3 ( 60% ) patients continued to have unchanged st depression . immediately postprocedure , there was resolution of twave inversion in 4 ( 57% ) patients , while 3 ( 43% ) patients continued to have unchanged twave inversion . patients who had resolved twave inversion postppvi did not have further ecgs at followup . additionally , 5 patients had a new finding of twave inversion on ecg . all newonset twave inversions and 2 of 3 persistent twave inversions postprocedure resolved at followup . one patient with persistent postprocedure twave inversion continued to have twave inversion at followup . finally , 4 patients had new twave inversions at followup . immediately postprocedure , there was resolution of st / t changes in 3 ( 27% ) patients , while 8 ( 73% ) patients continued to have unchanged st / t changes . patients who had resolved st / t changes postppvi did not have further ecgs at followup . additionally , 5 patients had a new finding of st / t changes on ecg . all newonset st / t changes and 3 of 8 ( 38% ) persistent st / t changes postprocedure resolved at followup . two patients with persistent postprocedure st / t changes continued to have st / t changes at followup . nine patients ( 8% ) had nsvt , 6 patients ( 6% ) had frequent pvcs , 10 patients ( 10% ) had atrial af / afl , and 2 patients ( 2% ) had supraventricular tachycardia . only 2/8 ( 25% ) patients with preppvi vt , and no patient with preppvi af / afl was in the ps group . newonset arrhythmias immediately postppvi include 7 patients with nsvt , 8 patients with pvcs , and 1 patient with af / afl . once again , there were very few psonly patients ( 3/16 , 19% ) in this cohort . at followup , two out of 8 patients with newonset pvcs were started on blockers , which were discontinued at their 6month followup . one of these patients was taken off the amiodarone at 6 months of followup . one patient was started on a blocker and continued to be on this medication because he had persistent documented pvcs . overall , 12/16 ( 75% ) patients no longer had documented nsvt , and 12/14 ( 86% ) patients no longer had pvcs . finally , af / afl resolved in 6/11 ( 55% ) cases ( figures 6 , 7 through 8 ; figures s2 and s3 ) . arrhythmia burden was determined by holter monitoring , internal rhythm devices reports , and telemetry during subsequent hospitalizations . documented nonsustained ventricular tachycardia ( nsvt ) . prior to procedure , 4 of 9 patients who previously had nsvt preprocedure continued to have nsvt , 1 of 9 patients no longer had nsvt but had pvcs , and 4 of 9 patients no longer had nsvt . among the patients with newonset nsvt postprocedure , immediately postprocedure , there was resolution of pvcs in 4 ( 64% ) patients , while 1 ( 17% ) patient continued to have persistent pvcs , and 1 ( 17% ) patient had nonsustained ventricular tachycardia ( nsvt ) . seven out of 8 patients with newonset pvcs and both of the persistent pvcs postprocedure no longer had any arrhythmia at followup . one patient with newonset pvcs and the patient with persistent pvcs immediately postprocedure continued to have pvcs at followup . nsvt indicates nonsustained ventricular tachycardia ; ppvi , percutaneous pulmonary valve implantation ; pvcs , premature ventricular contractions . , 5 patients continued to have af / afl ( 4 preprocedure and 1 new onset ) , while 6 patients no longer had af / afl . af / afl indicates atrial fibrillation / atrial flutter ; ppvi , percutaneous pulmonary valve implantation . forty patients had cardiac mri preppvi , but only 4 had postppvi studies for comparison ( 3 ps , 1 pi ) . this patient had nonspecific st changes both pre and postppvi , but these changes resolved at followup . the remaining 2 patients did not have ecg changes or appreciable structural changes on followup mri . of the 39 patients with ecg changes ( baseline + new onset postppvi + new onset at followup ) , 27 patients had ecg for analysis ( figures 5 , 6 through 7 ) . specifically , there were 19 resolved st / t changes , 3 persistent st / t changes , and 5 newonset st / t changes . although there was improvement in rv size by echocardiogram in 15/27 ( 56% ) patients , there was no change noted in 12/27 ( 44% ) patients . among patients with newonset / persistent ecg changes at followup ( n=8 ) , there were equal numbers of patients who had improved rv size versus no appreciable change by followup echo . in the 19 patients who had resolved ecg changes at followup , 10 patients had improved rv size , whereas 9 patients did show changes on followup echo . to our knowledge , this study is the first to evaluate changes in the electrophysiologic substrate and arrhythmia burden of patients undergoing ppvi . novel findings of this study include the observations of acute newly developing st / t changes and arrhythmias such as frequent isolated pvcs and nsvt immediately postppvi . moreover , the study was able to document improvement in the electrophysiologic substrate changes and arrhythmia burden at mediumterm followup . plymen et al10 studied conduction ( qrsd ) and repolarization ( qtc , qt , and jt dispersion ) parameters , and piotrowicz et al11 studied parameters of rv hypertrophy . similar to previous reports , our study did not find a significant change in qrsd postppvi in the overall cohort . however , plymen et al10 found significant shortening of qrsd in patients with pulmonary regurgitation . qrsd has served as a marker to assess the risk of arrhythmic death in congenital heart disease patients ( particularly tof ) with rvot dysfunction.2 , 13 patients with pi who do not undergo pulmonary valve replacement or ppvi have an increase in qrsd of 2 to 4 milliseconds per year that is associated with increasing right ventricular enddiastolic volume ( rvedv).1 , 14 , 15 because rvedv decreases postsurgical pulmonary valve replacement , the procedure has been postulated to improve the arrhythmia burden in patients with chronic pi.16 given the fact that ppvi serves as a better model to study the effects of improvements in hemodynamics on qrsd , plymen et al's10 particular findings could represent the true proportional relationship between improvement in rvedv and decreasing qrsd . unlike plymen et al,10 this study did not find a significant shortening of qrsd in the cohort of patients with purely pulmonary insufficiency . the reason for this discrepancy might reside in the fact that our sample size is too small to discern such difference . consistent with plymen et al , the qtc in our study uniformly increased immediately postppvi but shortened beyond baseline at followup , regardless of type of rvot dysfunction . qtc is a parameter of repolarization and a surrogate measurement of local action potential duration . it is recognized that myocardial stretch , via contractionexcitation feedback , increases monophasic action potential duration and causes afterdepolarizations , which in turn can cause triggered activity and generate the appropriate conditions for reentrant arrhythmias.17 contractionexcitation feedback has been demonstrated in both increased pressure and volume load conditions.18 , 19 , 20 for example , in the lamb , chronic pi led to inhomogeneity of right ventricular activation , decreased conduction velocity , and increased intracellular resistivity . these changes might contribute to the observed increased susceptibility to stretchinduced arrhythmias of the right ventricle.19 , 20 furthermore , when compared with the hypertrophied human myocardium , the dilated myocardium had longer steadystate action potential duration.21 consistent with this observation , our study showed that patients with pi had longer qtc than ps patients . contractionexcitation feedback and alterations in the local ventricular repolarization also explain the significant lengthening of qtc postppvi as previously demonstrated in the context of pulmonary valve valvuloplasty in ps patients.18 according to contractionexcitation theory , repolarization duration is inversely proportional to the mechanical stress on the myocardium . therefore , repolarization duration should be the longest after successful reduction in afterload seen with repetitive balloon occlusions of the rvot during placement of stents and the pulmonary valve in ppvi . consistent with this observation , our study showed that patients with ps had significantly longer qtc than pi patients immediately postppvi . it is important , therefore , to recognize that abnormalities in both depolarization and repolarization contribute to the increased risks of developing arrhythmia in patients with rvot dysfunction.21 the finding that qrsd and qtc stabilized and improved postppvi suggests that favorable changes in hemodynamics would translate into more stable myocardial depolarization and repolarization and reduce the risk of arrhythmias . other st / t changes ( such as nonspecific st / t changes , twave inversion , and st depression ) were also observed immediately postppvi . these changes most likely reflect transient ischemic injury that follows repeated total balloon occlusions of the rvot during rvot angioplasty , transient absent cardiac output during the check for coronary artery compression , and rapid ventricular pacing during 3dimensional rotational angiography in certain cases , or perhaps coronary artery spasm caused by selective coronary angiographies . these changes could also reflect the acute changes in hemodynamics with the newly placed valve , anesthesia effect , or possibly mechanical trauma of catheter manipulation during the procedure . interestingly , the majority of these st / t changes disappeared over time . despite favorable changes in measureable surface ecg parameters , the longterm effects of rv remodeling on the arrhythmias burden are unknown . our mean followup period is 31 months , which exceeds that of plymen et al10 and piotrowicz et al11 and is on a par with previous surgical pulmonary valve replacement series.16 , 22 , 23 , 24 , 25 , 26 previously , gengsakul et al25 and harrild et al26 studied matched pairs of patients with repaired tof with and without surgical pulmonary valve replacement and showed no significant change in qrsd and impact on arrhythmia burden . specifically , harrild et al26 reported no survival benefit in death or sustained ventricular tachycardia at a mean of 3 years of followup . however , the myocardial incision incurred with the surgical pulmonary valve replacement may represent another confounding factor in their analysis . therrien et al16 studied a cohort of 70 patients who underwent pulmonary valve replacement late after tof repair with some patients with preexisting arrhythmias undergoing intraoperative ep mapping and cryoablation . they found that the qrsd stabilized at a mean of 4.7 years of followup and that there was a significant decrease in the prevalence of monomorphic vt . overall , our study reported a substantial decrease in the prevalence of nsvt and pvcs postppvi . there was a significant incidence of new pvcs and nsvt immediately postppvi . some of the patients required shortterm antiarrhythmic therapy with a blocker or amiodarone that was discontinued at their 6month followup . we attribute these changes to the irritability caused by stents placed in the rvot prior to the melody valve placement and possibly due to contractionexcitation feedback caused by myocardial stretch associated with the repetitive balloon occlusion of the rvot during placement of stents and the pulmonary valve in ppvi . moreover , previous studies have shown that ventricular ectopy may be caused by afterdepolarizations associated with increase in mechanical stress.28 , 29 these arrhythmias would resolve once the stents completely endothelialized and hemodynamics reached a better steady state . unfortunately , many of the patients in our cohort received prestenting of the rvot , precluding any analysis of the effect of prestenting on arrhythmia burden . finally , pvcs and nsvt may be related to the underlying anatomical heart defect , focal scarring , fibrosis , or surgical suture lines rather than to right ventricular outflow tract dysfunction . this is a singlecenter retrospective study , with patient attrition over time , which limited the power of our analysis . additionally , our cohort was heterogeneous in their anatomical lesions , hemodynamics , and clinical status prior to the procedure . therefore , we predict that there is variability in the beneficial effects of improved hemodynamics on the electrophysiology substrate and measurable ecg parameters . an improved sample size would allow us to perform more detailed subgroup analyses to better tease out the different effects of ppvi . despite most patients with previously or newly diagnosed arrhythmias undergoing followup holter monitor evaluation at 6month followup , there was no uniform protocol for monitoring of pvcs and nsvt over the followup period , which may lead to an overestimation in the prevalence reduction of these arrhythmias . although our mean followup period is reasonable at 31 months , longterm followup is still needed to both assess the measurable ecg parameters and provide better correlation with the arrhythmia burden . last , a potential limitation is that changes to the qtc may be secondary to anesthesia , other medications , or electrolyte anomalies . patients in this study had ecgs obtained the morning following ppvi placement , and changes to the qtc were based on preprocedural ecg as a result , each patient was his or her own control . this is a singlecenter retrospective study , with patient attrition over time , which limited the power of our analysis . additionally , our cohort was heterogeneous in their anatomical lesions , hemodynamics , and clinical status prior to the procedure . therefore , we predict that there is variability in the beneficial effects of improved hemodynamics on the electrophysiology substrate and measurable ecg parameters . an improved sample size would allow us to perform more detailed subgroup analyses to better tease out the different effects of ppvi . despite most patients with previously or newly diagnosed arrhythmias undergoing followup holter monitor evaluation at 6month followup , there was no uniform protocol for monitoring of pvcs and nsvt over the followup period , which may lead to an overestimation in the prevalence reduction of these arrhythmias . although our mean followup period is reasonable at 31 months , longterm followup is still needed to both assess the measurable ecg parameters and provide better correlation with the arrhythmia burden . last , a potential limitation is that changes to the qtc may be secondary to anesthesia , other medications , or electrolyte anomalies . patients in this study had ecgs obtained the morning following ppvi placement , and changes to the qtc were based on preprocedural ecg as a result , each patient was his or her own control . our study demonstrates that ppvi stabilizes the qrsd and shortens the qtc in congenital heart disease patients with rvot dysfunction . furthermore , there is a substantial decrease in the prevalence of nsvt and pvcs postppvi . finally , acute changes to the electrophysiologic substrate and arrhythmia burden immediately postppvi improve over time at midterm followup . further prospective and longterm studies would be needed to further define and assess the causal effects of ppvi on electrophysiologic changes in these patients . st / t changes in patients without rbbb at baseline . nine patients had st / t changes preppvi . immediately postppvi , st / t changes resolved in 5 of 9 ( 55% ) patients while remaining persistent in 4 of 9 ( 45% ) patients . six patients had newonset st / t changes . at follow up , all newonset st / t changes resolved immediately postppvi . of the 4 persistent st / t changes immediately postppvi , 1 continued to have st / t changes , 1 resolved , and 2 were lost to followup . figure s3 . documented nonsustained ventricular tachycardia ( nsvt ) in patients without rbbb . | backgroundpercutaneous pulmonary valve implantation ( ppvi ) is firstline therapy for some congenital heart disease patients with right ventricular outflow tract dysfunction .
the hemodynamics improvements after ppvi are well documented , but little is known about its effects on the electrophysiologic substrate .
the objective of this study is to assess the short and mediumterm electrophysiologic substrate changes and elucidate postprocedure arrhythmias.methods and resultsa retrospective chart review of patients undergoing ppvi from may 2010 to april 2015 was performed .
a total of 106 patients underwent ppvi ; most commonly these patients had tetralogy of fallot ( n=59 , 55% ) and pulmonary insufficiency ( n=60 , 57% ) .
the median followup time was 28 months ( 763 months ) .
preppvi , 25 patients ( 24% ) had documented arrhythmias : nonsustained ventricular tachycardia ( nsvt ) ( n=9 , 8% ) , frequent premature ventricular contractions ( pvcs ) ( n=6 , 6% ) , and atrial fibrillation / flutter ( af / afl ) ( n=10 , 9% ) .
postppvi , arrhythmias resolved in 4 patients who had nsvt ( 44% ) and 5 patients who had pvcs ( 83% ) .
new arrhythmias were seen in 16 patients ( 15% ) : 7 nsvt , 8 pvcs , and 1 af / afl . there was resolution at mediumterm followup in 6 ( 86% ) patients with newonset nsvt and 7 ( 88% ) patients with newonset pvcs .
there was no difference in qrs duration preppvi , postppvi , and at mediumterm followup ( p=0.6 ) .
the median corrected qt lengthened immediately postppvi but shortened significantly at midterm followup ( p<0.01).conclusions
ppvi reduced the prevalence of nsvt .
the majority of postimplant arrhythmias resolve by 6 months of followup . |
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human and rabbit corneal epithelial cell lines used in various studies have been well characterized in previous studies . in the current study , two kinds of the corneal epithelial cells were used in the study including the primary human corneal epithelial cells and rabbit corneal epithelial cells . primary human and rabbit corneal epithelial cells were cultured in a serum - free keratinocyte medium ( defined keratinocyte - sfm ; invitrogen , carlsbad , ca , usa ) , and primary human corneal epithelial cells were grown in dulbecco 's modified eagle 's medium / f-12 ( 1:1 ; gibco , waltham , ma , usa ) containing 10% fetal bovine serum and 5-g / ml insulin . cells were cultured in an incubator supplied with 95% air and 5% co2 at 37c . the medium was replaced every 2 days , and cells were subcultured by treatment of cells with 0.05% trypsin - edta ( gibco ) . primary human corneal epithelial cells were obtained by passing fresh isolated limbal stem / progenitor cells into collagen / fibronectin ( 50%/50%)-coated surface and an airlift ( reduced medium level ) procedure to induce differentiation . the medium contains 10% fetal bovine serum , 5-mg / ml insulin , and 10,000 units / ml penicillin , and 10,000 g / ml streptomycin , and maintained in an incubator supplied with 95% air and 5% co2 at 37c . the medium was replaced every 2 days and cells were passed by treating cells with 0.05% trypsin - edta . for uvc irradiation experiments , corneal epithelial cells were exposed to uvc light at an intensity of 45 j / cm . in previous studies , we found that stimulation of corneal epithelial cells with uvc at 45 j / cm has a very similar or compatible effect as uvb at a dosage of 200 mj / cm . the reason that we used uvc is that a quick stimulation allowed us to record the fast response of cells under a real time microscope ( see below ) . the uvc source was from an 84-watt uvc lamp with a wavelength of 254 nm ( spectronic corp , westbury , ny , usa ) . the intensities of the uvc light were calibrated by using an optical radiometer ( ms-100 ; uvp llc , upland , ca , usa ) . hyperosmotic stress stimulation was performed by adding various concentrations of sorbitol to the culture media following a time course . osmotic pressures of hyperosmotic media were measured by using a vapor pressure osmometer ( vapro5520 ) . the experimental protocol was exempted by the university of california los angeles institutional review board ( los angeles , ca , usa ) . morphologic changes of corneal epithelial cells that were exposed or not exposed to uvc irradiation were measured in the absence and presence of 4-aminopyridine ( 4-ap ) by utilization of the nikon eclipse ti ( nikon instruments , inc . , melville , ny , usa ) inverted microscope with the following functions : ( 1 ) time - lapse videos of the phase - contrast / fluorescent live images , ( 2 ) built - in total internal reflection fluorescence ( tirf ) and frster resonance energy transfer ( fret ) ; ( 3 ) perfect focus system ( pfs ) , and ( 4 ) a digital charge - coupled device ( ccd ) camera in a time interval of 0.2 minutes for each photo . the system was equipped with a heated chamber at 37c and flushed with mixed 5% co2 that kept cells in a normal culture condition . cell motility was analyzed by tracking cell movements and distances ( m / h ) using nikon ti 's nis - elements software . changes in relative cell volume in the real time were estimated by measuring self - quenching of the fluorescent dye calcein . calcein - am stock solution was made at a concentration of 10 m in pbs with 1% dmso . corneal epithelial cells ( 70%80% confluence ) were loaded with calcein ( 200 l per sample ) and incubated at 37c for 60 minutes . unloaded calcein - am is removed by washing wells three times with 1-ml warm ( 37c ) 1 pbs . changes of calcein - am intensity were measured using the packard fusion microplate analyzer ( conquer scientific , san diego , ca , usa ) at 485- and 515-nm excitation and emission , respectively . calcein - am enters the cell and the am - group is cleaved and the intracellular calcein becomes impermeable to the cell membrane . thus , all the dye that has entered the cell is trapped within the cell . when the cell shrinks due to efflux of k ions followed by water , the dye is concentrated in the cellular compartment . after the process of self - quenching , intensity of the dye decrease within the cell . the relative cell volume in isotonic solution was initially recorded for 5 minutes before uvc or hyperosmotic exposure . after uvc or hyperosmotic exposure , cell volume was measured every 0.1 to 0.5 minutes . in the measurement of uvc - induced volume changes , primary rabbit corneal epithelial cells were used because these cells were easier to be loaded with the dyes and less of leakages . in addition , both human and rabbit corneal epithelial cells had similar effects in responses to the uvc irradiation . corneal epithelial cells ( 2 10 ) were lysed in sodium dodecyl sulfatepolyacrylamide ( sds ) sample buffer containing 62.5 mm tris - hcl ph 6.8 , 2% wt / vol sds , 10% glycerol , 50 mm dtt , 0.01% wt / vol bromophenol blue or phenol red . after denaturing , experimental samples in each testing tube are equally divided and loaded onto two identical protein gels that are running in the same gel apparatus , in the same condition and the same time . proteins were electrotransferred to two identical polyvinylidene difluoride ( pvdf ) membranes ( one of the membranes was for the loading control ) . they were exposed to blocking buffer containing 5% nonfat milk in tbs-0.1% tween 20 ( tbs - t ) for 1 hour at 22c , and then incubated with the primary antibodies at 4c overnight . horseradish peroxidase ( hrp)-conjugated secondary antibody was applied in tbs - t buffer for 1 hour at 22c . western blots were developed by an ecl plus system ( thermo fisher scientific , inc . , santa cruz , ca , usa ) and visualized by exposure of x - ray films . specific antibodies used in the study were phosphor - fak tyr-397 ( antibody 3283 ) , phosphor - fak tyr-925 ( antibody 3284 ) , and phospho - scr tyr-416 ( antibody 2101 ) from cell signaling ( danvers , ma , usa ) . for immunostaining experiments , the cells were fixed for 15 minutes in 4% paraformaldehyde , and then permeabilized with pbs-0.2% triton - x100 ( pbs - t ) for 30 minutes at 22c . the tissues were blocked by incubation with 10% normal horse serum ( nhs ) or 10% normal goat serum in pbs - t for 1 hour at 22c , followed by immunostaining with the corresponding antibodies , and/or incubated with fitc / cy3-conjugated goat anti - mouse / rabbit igg antibody ( cells signaling ) . elisa experiments were carried out using corresponding antibodies against phospho - fak and phospho - src according to the company 's instructions ( invitrogen ) . briefly , corneal epithelial cells were exposed to uvc irradiation with / without adding kv channel blockers in the indicated concentrations prior to lysis . the cell lysates were incubated in duplicate in the elisa system . for western analysis , signals in the films were scanned digitally and optical densities ( od ) were quantified by using the image calculator software ( photometrics , tucson , az , usa ) . the relative od was calculated by normalizing the signals from target proteins against intensities of loading controls . the data of elisa experiments were represented as the mean sd from three experiments independently performed in duplicates . all other experimental data were subjects to statistical analysis and plotted as mean se . significant differences between the control and treated groups were determined by 1-way anova and tukey 's tests ( f < 0.05 ) . student 's t - test was used to determine the significant difference for paired samples at p less than 0.05 . human and rabbit corneal epithelial cell lines used in various studies have been well characterized in previous studies . in the current study , two kinds of the corneal epithelial cells were used in the study including the primary human corneal epithelial cells and rabbit corneal epithelial cells . primary human and rabbit corneal epithelial cells were cultured in a serum - free keratinocyte medium ( defined keratinocyte - sfm ; invitrogen , carlsbad , ca , usa ) , and primary human corneal epithelial cells were grown in dulbecco 's modified eagle 's medium / f-12 ( 1:1 ; gibco , waltham , ma , usa ) containing 10% fetal bovine serum and 5-g / ml insulin . cells were cultured in an incubator supplied with 95% air and 5% co2 at 37c . the medium was replaced every 2 days , and cells were subcultured by treatment of cells with 0.05% trypsin - edta ( gibco ) . primary human corneal epithelial cells were obtained by passing fresh isolated limbal stem / progenitor cells into collagen / fibronectin ( 50%/50%)-coated surface and an airlift ( reduced medium level ) procedure to induce differentiation . the medium contains 10% fetal bovine serum , 5-mg / ml insulin , and 10,000 units / ml penicillin , and 10,000 g / ml streptomycin , and maintained in an incubator supplied with 95% air and 5% co2 at 37c . the medium was replaced every 2 days and cells were passed by treating cells with 0.05% trypsin - edta . for uvc irradiation experiments , corneal epithelial cells were exposed to uvc light at an intensity of 45 j / cm . in previous studies , we found that stimulation of corneal epithelial cells with uvc at 45 j / cm has a very similar or compatible effect as uvb at a dosage of 200 mj / cm . the reason that we used uvc is that a quick stimulation allowed us to record the fast response of cells under a real time microscope ( see below ) . the uvc source was from an 84-watt uvc lamp with a wavelength of 254 nm ( spectronic corp , westbury , ny , usa ) . the intensities of the uvc light were calibrated by using an optical radiometer ( ms-100 ; uvp llc , upland , ca , usa ) . hyperosmotic stress stimulation was performed by adding various concentrations of sorbitol to the culture media following a time course . osmotic pressures of hyperosmotic media were measured by using a vapor pressure osmometer ( vapro5520 ) . the experimental protocol was exempted by the university of california los angeles institutional review board ( los angeles , ca , usa ) . morphologic changes of corneal epithelial cells that were exposed or not exposed to uvc irradiation were measured in the absence and presence of 4-aminopyridine ( 4-ap ) by utilization of the nikon eclipse ti ( nikon instruments , inc . , melville , ny , usa ) inverted microscope with the following functions : ( 1 ) time - lapse videos of the phase - contrast / fluorescent live images , ( 2 ) built - in total internal reflection fluorescence ( tirf ) and frster resonance energy transfer ( fret ) ; ( 3 ) perfect focus system ( pfs ) , and ( 4 ) a digital charge - coupled device ( ccd ) camera in a time interval of 0.2 minutes for each photo . the system was equipped with a heated chamber at 37c and flushed with mixed 5% co2 that kept cells in a normal culture condition . cell motility was analyzed by tracking cell movements and distances ( m / h ) using nikon ti 's nis - elements software . changes in relative cell volume in the real time were estimated by measuring self - quenching of the fluorescent dye calcein . calcein - am stock solution was made at a concentration of 10 m in pbs with 1% dmso . corneal epithelial cells ( 70%80% confluence ) were loaded with calcein ( 200 l per sample ) and incubated at 37c for 60 minutes . unloaded calcein - am is removed by washing wells three times with 1-ml warm ( 37c ) 1 pbs . changes of calcein - am intensity were measured using the packard fusion microplate analyzer ( conquer scientific , san diego , ca , usa ) at 485- and 515-nm excitation and emission , respectively . calcein - am enters the cell and the am - group is cleaved and the intracellular calcein becomes impermeable to the cell membrane . thus , all the dye that has entered the cell is trapped within the cell . when the cell shrinks due to efflux of k ions followed by water , the dye is concentrated in the cellular compartment . after the process of self - quenching , intensity of the dye decrease within the cell . the relative cell volume in isotonic solution was initially recorded for 5 minutes before uvc or hyperosmotic exposure . after uvc or hyperosmotic exposure , cell volume was measured every 0.1 to 0.5 minutes . in the measurement of uvc - induced volume changes , primary rabbit corneal epithelial cells were used because these cells were easier to be loaded with the dyes and less of leakages . in addition , both human and rabbit corneal epithelial cells had similar effects in responses to the uvc irradiation . corneal epithelial cells ( 2 10 ) were lysed in sodium dodecyl sulfatepolyacrylamide ( sds ) sample buffer containing 62.5 mm tris - hcl ph 6.8 , 2% wt / vol sds , 10% glycerol , 50 mm dtt , 0.01% wt / vol bromophenol blue or phenol red . after denaturing , cell lysates were size - fractionated in 12% page gels . experimental samples in each testing tube are equally divided and loaded onto two identical protein gels that are running in the same gel apparatus , in the same condition and the same time . proteins were electrotransferred to two identical polyvinylidene difluoride ( pvdf ) membranes ( one of the membranes was for the loading control ) . they were exposed to blocking buffer containing 5% nonfat milk in tbs-0.1% tween 20 ( tbs - t ) for 1 hour at 22c , and then incubated with the primary antibodies at 4c overnight . horseradish peroxidase ( hrp)-conjugated secondary antibody was applied in tbs - t buffer for 1 hour at 22c . western blots were developed by an ecl plus system ( thermo fisher scientific , inc . , santa cruz , ca , usa ) and visualized by exposure of x - ray films . specific antibodies used in the study were phosphor - fak tyr-397 ( antibody 3283 ) , phosphor - fak tyr-925 ( antibody 3284 ) , and phospho - scr tyr-416 ( antibody 2101 ) from cell signaling ( danvers , ma , usa ) . for immunostaining experiments , corneal epithelial cells were grown on glass slides and treated as indicated in figures . the cells were fixed for 15 minutes in 4% paraformaldehyde , and then permeabilized with pbs-0.2% triton - x100 ( pbs - t ) for 30 minutes at 22c . the tissues were blocked by incubation with 10% normal horse serum ( nhs ) or 10% normal goat serum in pbs - t for 1 hour at 22c , followed by immunostaining with the corresponding antibodies , and/or incubated with fitc / cy3-conjugated goat anti - mouse / rabbit igg antibody ( cells signaling ) . elisa experiments were carried out using corresponding antibodies against phospho - fak and phospho - src according to the company 's instructions ( invitrogen ) . briefly , corneal epithelial cells were exposed to uvc irradiation with / without adding kv channel blockers in the indicated concentrations prior to lysis . for western analysis , signals in the films were scanned digitally and optical densities ( od ) were quantified by using the image calculator software ( photometrics , tucson , az , usa ) . the relative od was calculated by normalizing the signals from target proteins against intensities of loading controls . the data of elisa experiments were represented as the mean sd from three experiments independently performed in duplicates . all other experimental data were subjects to statistical analysis and plotted as mean se . significant differences between the control and treated groups were determined by 1-way anova and tukey 's tests ( f < 0.05 ) . student 's t - test was used to determine the significant difference for paired samples at p less than 0.05 . changes in cell membrane k channel activity can mediate functional adaptation to a variety of chemical and physical stresses through membrane voltage stabilization and maintenance of salt and water balance . previously , we reported data from theoretical calculation and modeling , demonstrating that uv irradiation induced hyperactivation of k channels results in cell volume changes . alterations of the cell sizes were recorded as cell attached areas by real - time microscopy with a computerized head stage and cell tracking system . there was a remarkable change in cell size measured by areas of the attached cells after exposure to uvc irradiation ( fig . statistical analysis showed that the measured cell areas were significantly changed before ( a0 ) and after ( a ) uvc irradiation ( fig . the effect of uvc irradiation on rabbit corneal epithelial cell volume decrease was measured in the absence ( v0 ) and presence ( vt ) of uvc irradiation following a time course ( fig . the relative cell volume changes upon exposure of primary cultured rabbit corneal epithelial cells to uvc irradiation were plotted following a real - time measurement ( fig . the cell volume fell to the lowest point within 1 minute followed by a slow volume recovery . ultraviolet c irradiation induced rabbit corneal epithelial cell shrinkage was markedly inhibited in the presence of 4-ap , a kv channel - specific blocker . in addition , the effect of suppressing kv channel on uvc irradiation evoked volume decrease was further examined by adding a group of kv channel blockers , including 4-ap ( 0.5 mm ) , -dendrotoxin ( -dtx , 200 nm ) , and blood depressing substance-1 ( bds-1 , 400 nm ) at different time points ( fig . induced cell size and volume decreases are resulted from hyperactivation of kv channels resulting in a fast loss of intracellular k ions in human and rabbit corneal epithelial cells , respectively . effects of uvc irradiation on human and rabbit corneal epithelial cell sizes and volume decreases . ( a ) ultraviolet c irradiation induced cell size decrease recorded by a real - time video in human corneal epithelial cells . ( c ) time course of uvc irradiation induced cell volume changes in rabbit corneal epithelial cells . rabbit corneal epithelial cells were exposed to uvc light in the absence and presence of 4-ap . ( e ) suppression of uvc induced cell volume decrease by various kv channel blockers . the effect of kv channel blockers on uvc - induced volume changes were measured at 0.5 , 1 , and 5 minutes after uvc exposure in the absence and presence of kv channel blockers , including 0.5-mm 4-ap , 200-m -dtx , and 400-m bds-1 . asterisks represent the significant difference , p < 0.01 . ultraviolet c irradiation induced fak and src activities were investigated in human corneal epithelial cells by measuring their phosphorylation levels using anti - phosphorylated src and fak antibodies , respectively . first , uvc irradiation induced increase in phosphorylation scr was detected by western analysis following a 60-minutes time course ( fig . change in fak phosphorylation levels was detected by elisa showing that the fak phosphorylation level was significantly increased within 5 minutes and kept to be increased for 60 minutes after uvc exposure ( fig . further studies were performed with western analysis by using anti-[tyr(p)416]-src , anti-[tyr(p)397]-fak , and anti-[tyr(p)925]-fak antibodies . ultraviolet c irradiation induced site - specific phosphorylations of active src and fak were detected and compared following a time course by using antibodies against phosphorylated forms of src , fak , and fak ( fig . statistical analysis showed that there were significant changes in phosphorylation levels of src , fak , and fak following uvc irradiation ( fig . the results suggest that there is an early effect of uvc irradiation on phosphorylation of src and fak and that the relationship of the time courses between uvc irradiation induced changes of the cell volume and phosphorylation of src and fak is closely correlated . effect of uvc irradiation on src and fak phosphorylation / activation in human corneal epithelial cells . ( a ) ultraviolet c induced changes of src phosphorylation following a time course were detected by western analysis . ( c ) western analysis of uvc irradiation induced site - specific phosphorylation of src , fak , and fak following a time course . ( d ) summarized time courses of site - specific phosphorylation of fak in uvc irradiation induced human corneal epithelial cells . these data demonstrate that uvc irradiation induced cell volume alteration resulted in activation of src and fak signaling events . we found that 4-ap effectively suppressed src phosphorylation at 1 and 5 minutes after exposure of human corneal epithelial cells to uvc irradiation ( fig . 3a ) . ultraviolet c irradiation induced increases in fak phosphorylation detected by elisa were also effectively suppressed by 4-ap in a dose - dependent manner , which is in the same dosage range for 4-ap to block the uvc irradiation induced volume decrease ( fig . the same site - specific fak antibody against phospho - fak was used in western analysis to demonstrate activation of fak induced by uvc irradiation in human corneal epithelial cells ( fig . 3c ) . ultraviolet c irradiation induced site - specific phosphorylation of fak was effectively suppressed by 4-ap ( 2 mm ) . further experiments were performed by testing the effect of 4-amino-5-(4-chlorophenyl)-7-(t- butyl)pyrazolo[3,4-d ] pyrimidine ( pp2 ) , a potent inhibitor of src and fak on site - specific phosphorylation of src , fak and fak , following a dose - dependent pattern ( fig . 3d ) . there was a rather weak effect for pp2 to inhibit uvc irradiation induced phosphorylation of fak . statistical analysis showed that pp2 significantly supressed uvc irradiation induced phosphorylation of both src and fak . these results indicate that uvc - induced a site - specific phosphorylation of src and fak , resulting in activation of src and fak in uvc - exposed human corneal epithelial cells . effect of kv channel blockade on src and fak phosphorylation / activation in human corneal epithelial cells . ( a ) effect of blocking of kv channel on uvc irradiation induced src phosphorylation / activation . ultraviolet c irradiation induced kv channel activity in human corneal epithelial cells was blocked by 0.5-mm 4-ap and src activation was detected by western analysis . ( b ) dose - response relationship of suppressing uvc irradiation induced fak activation by 4-ap . human corneal epithelial cells were exposed to uvc irradiation in the absence and presence of various concentrations of 4-ap . ultraviolet c irradiation induced changes of fak phosphorylation were determined by elisa . ( c ) effect of 4-ap on inhibition of uvc irradiation induced faky397 phosphorylation detected by western analysis . ( d ) dose - response relationship of inhibiting uvc irradiation induced src and fak site - specific phosphorylation by pp2 . ( e ) statistical analysis of various pp2 concentrations on site - specific inhibition of uvc irradiation - induced fak phosphorylation . effects of various concentrations of pp2 on phosphorylation levels of src , fak , and fak in uvc irradiation the effect of hyperosmotic pressure , established by adding 450-mm sorbitol in the medium , on human corneal epithelial cell volume decrease was also measured to compare with the uvc irradiation induced volume decrease ( fig . there were continuous decreases in cell volume when osmotic condition in the extracellular medium was increased from the control level of 300 to 510 mm by adding various concentrations of sorbitol within 1 minute ( fig . volume changes of the cells that were exposed to hypo - osmotic conditions were performed as the control measurements . upon exposure of cells to 225- and 150-mm extracellular solutions , cell volume after initial increase in the volume , the cell volume was gradually decreased toward the normal cell volume following a time course in the hypo - osmotic condition . in uvc irradiation induced and hyperosmotic stress ( 450 mm)stimulated cells , the peak volume changes were significant to show a volume shrinkage down to 67% and 65% in uvc irradiation induced and hyperosmotic pressure ( 450 mm)treated cells , respectively ( fig . 4c ) . however , applications of 4-ap had no significant effects on osmotic pressure induced initial volume changes ( fig . the results reveal that uvc irradiation induced a significant cell shrinkage comparable to the effect resulted from hyperosmotic pressure stimulation . effect of blocking kv channel on hyperosmotic pressure induced rabbit corneal epithelial cell volume changes . ( a ) real - time recording of hyperosmotic pressure induced changes in rabbit corneal epithelial cells . ( b ) effect of various osmotic pressures by changing sorbitol concentrations rabbit corneal epithelial cell volume . ( c ) comparison of cell volume changes in uvc irradiation and hyperosmotic pressure ( 450 mm of sorbitol)induced rabbit corneal epithelial cells . the time - points were selected for the peak time of cell volume changes for uvc exposure and hyperosmotic treatments . ( d ) comparison of 4-ap effect on uvc irradiation and hyperosmotic pressure induced rabbit epithelial cell volume changes . asterisks represent the significant difference ( n = 4 ) , p < 0.05 . the effect of hyperosmotic pressure on src and fak phosphorylation was measured in human corneal epithelial cells . exposure of human corneal epithelial cells to hyperosmotic pressure resulted in increases in phosphorylation of src and fak following a time course ( fig . 5a ) . statistical analysis showed the peak time of increased phosphorylation for fak and src at 1 and 15 minutes , respectively ( fig . time courses of hyperosmotic pressure induced activation of src and fak were similar to uvc irradiation stimulated cells . furthermore , application of k channel blocker 4ap at a dosage of 1 mm had no significant effects on hyperosmotic pressure induced src and fak phosphorylation levels ( figs . in hyperosmotic pressure - induced cells , both src and fak phosphorylation were increased as they have been observed in uvc irradiation induced cell , suggesting that src and fak phosphorylation are cell volume change sensitive . effect of kv channel blockade on hyperosmotic pressure induced src and fak phosphorylation / activation in human corneal epithelial cells . effect of blockade of kv channel on uvc irradiation induced src phosphorylation / activation . ( c ) effect of 4-ap on hyperosmotic pressure induced fak phosphorylation in human corneal epithelial cells . ( d ) effect of 4-ap on hyperosmotic pressure induced src phosphorylation in human corneal epithelial cells . we predict that integrins in the edge area of the attached human corneal epithelial cells are affected by uvc irradiation induced cell shrinkage . activation of integrin 5 in uvc - irradiated cells was detected by immunostaining using the anti - integrin 5 antibody in the absence and presence of 4-ap . further immunostaining experiments were performed to verify uvc irradiation induced increases in phospho - fak stained by using the antibody that specifically recognizes phospho - fak ( fig . ultraviolet c irradiation induced increases of the phospho - fak were effectively prevented by preincubation of cells with 4-ap . blockades of kv channel by 4-ap resulting in suppression of uvc irradiation induced activations of integrin 5 and fak provide further evidence that uvc irradiation induced cellular changes are mediated by initial activation of k channels in uvc - exposed human corneal epithelial cells . immunostaining detection of uvc irradiation induced alterations of integrin and fak activities in human corneal epithelial cells . ( a ) ultraviolet c irradiation induced integrin 5 activation in the absence and presence of 4-ap in different scales . induced fak phosphorylation stained by red color and nuclei were stained with dapi in the absence and presence of 0.5-mm 4-ap in different scales . changes in cell membrane k channel activity can mediate functional adaptation to a variety of chemical and physical stresses through membrane voltage stabilization and maintenance of salt and water balance . previously , we reported data from theoretical calculation and modeling , demonstrating that uv irradiation induced hyperactivation of k channels results in cell volume changes . alterations of the cell sizes were recorded as cell attached areas by real - time microscopy with a computerized head stage and cell tracking system . there was a remarkable change in cell size measured by areas of the attached cells after exposure to uvc irradiation ( fig . statistical analysis showed that the measured cell areas were significantly changed before ( a0 ) and after ( a ) uvc irradiation ( fig . the effect of uvc irradiation on rabbit corneal epithelial cell volume decrease was measured in the absence ( v0 ) and presence ( vt ) of uvc irradiation following a time course ( fig . the relative cell volume changes upon exposure of primary cultured rabbit corneal epithelial cells to uvc irradiation were plotted following a real - time measurement ( fig . the cell volume fell to the lowest point within 1 minute followed by a slow volume recovery . ultraviolet c irradiation induced rabbit corneal epithelial cell shrinkage was markedly inhibited in the presence of 4-ap , a kv channel - specific blocker . in addition , the effect of suppressing kv channel on uvc irradiation evoked volume decrease was further examined by adding a group of kv channel blockers , including 4-ap ( 0.5 mm ) , -dendrotoxin ( -dtx , 200 nm ) , and blood depressing substance-1 ( bds-1 , 400 nm ) at different time points ( fig . induced cell size and volume decreases are resulted from hyperactivation of kv channels resulting in a fast loss of intracellular k ions in human and rabbit corneal epithelial cells , respectively . effects of uvc irradiation on human and rabbit corneal epithelial cell sizes and volume decreases . ( a ) ultraviolet c irradiation induced cell size decrease recorded by a real - time video in human corneal epithelial cells . ( c ) time course of uvc irradiation induced cell volume changes in rabbit corneal epithelial cells . rabbit corneal epithelial cells were exposed to uvc light in the absence and presence of 4-ap . ( e ) suppression of uvc induced cell volume decrease by various kv channel blockers . the effect of kv channel blockers on uvc - induced volume changes were measured at 0.5 , 1 , and 5 minutes after uvc exposure in the absence and presence of kv channel blockers , including 0.5-mm 4-ap , 200-m -dtx , and 400-m bds-1 . ultraviolet c irradiation induced fak and src activities were investigated in human corneal epithelial cells by measuring their phosphorylation levels using anti - phosphorylated src and fak antibodies , respectively . first , uvc irradiation induced increase in phosphorylation scr was detected by western analysis following a 60-minutes time course ( fig . ultraviolet c irradiation induced change in fak phosphorylation levels was detected by elisa showing that the fak phosphorylation level was significantly increased within 5 minutes and kept to be increased for 60 minutes after uvc exposure ( fig . further studies were performed with western analysis by using anti-[tyr(p)416]-src , anti-[tyr(p)397]-fak , and anti-[tyr(p)925]-fak antibodies . ultraviolet c irradiation induced site - specific phosphorylations of active src and fak were detected and compared following a time course by using antibodies against phosphorylated forms of src , fak , and fak ( fig . statistical analysis showed that there were significant changes in phosphorylation levels of src , fak , and fak following uvc irradiation ( fig . the results suggest that there is an early effect of uvc irradiation on phosphorylation of src and fak and that the relationship of the time courses between uvc irradiation induced changes of the cell volume and phosphorylation of src and fak is closely correlated . effect of uvc irradiation on src and fak phosphorylation / activation in human corneal epithelial cells . ( a ) ultraviolet c induced changes of src phosphorylation following a time course were detected by western analysis . ( c ) western analysis of uvc irradiation induced site - specific phosphorylation of src , fak , and fak following a time course . ( d ) summarized time courses of site - specific phosphorylation of fak in uvc irradiation induced human corneal epithelial cells . these data demonstrate that uvc irradiation induced cell volume alteration resulted in activation of src and fak signaling events . we found that 4-ap effectively suppressed src phosphorylation at 1 and 5 minutes after exposure of human corneal epithelial cells to uvc irradiation ( fig . 3a ) . ultraviolet c irradiation induced increases in fak phosphorylation detected by elisa were also effectively suppressed by 4-ap in a dose - dependent manner , which is in the same dosage range for 4-ap to block the uvc irradiation induced volume decrease ( fig . the same site - specific fak antibody against phospho - fak was used in western analysis to demonstrate activation of fak induced by uvc irradiation in human corneal epithelial cells ( fig . 3c ) . ultraviolet c irradiation induced site - specific phosphorylation of fak was effectively suppressed by 4-ap ( 2 mm ) . further experiments were performed by testing the effect of 4-amino-5-(4-chlorophenyl)-7-(t- butyl)pyrazolo[3,4-d ] pyrimidine ( pp2 ) , a potent inhibitor of src and fak on site - specific phosphorylation of src , fak and fak , following a dose - dependent pattern ( fig . 3d ) . there was a rather weak effect for pp2 to inhibit uvc irradiation induced phosphorylation of fak . statistical analysis showed that pp2 significantly supressed uvc irradiation induced phosphorylation of both src and fak . these results indicate that uvc - induced a site - specific phosphorylation of src and fak , resulting in activation of src and fak in uvc - exposed human corneal epithelial cells . effect of kv channel blockade on src and fak phosphorylation / activation in human corneal epithelial cells . ( a ) effect of blocking of kv channel on uvc irradiation induced src phosphorylation / activation . ultraviolet c irradiation induced kv channel activity in human corneal epithelial cells was blocked by 0.5-mm 4-ap and src activation was detected by western analysis . ( b ) dose - response relationship of suppressing uvc irradiation induced fak activation by 4-ap . human corneal epithelial cells were exposed to uvc irradiation in the absence and presence of various concentrations of 4-ap . ultraviolet c irradiation induced changes of fak phosphorylation were determined by elisa . ( c ) effect of 4-ap on inhibition of uvc irradiation induced faky397 phosphorylation detected by western analysis . ( d ) dose - response relationship of inhibiting uvc irradiation induced src and fak site - specific phosphorylation by pp2 . ( e ) statistical analysis of various pp2 concentrations on site - specific inhibition of uvc irradiation - induced fak phosphorylation . effects of various concentrations of pp2 on phosphorylation levels of src , fak , and fak in uvc irradiation induced rabbit corneal epithelial cells were determined by western analysis . the effect of hyperosmotic pressure , established by adding 450-mm sorbitol in the medium , on human corneal epithelial cell volume decrease was also measured to compare with the uvc irradiation induced volume decrease ( fig . there were continuous decreases in cell volume when osmotic condition in the extracellular medium was increased from the control level of 300 to 510 mm by adding various concentrations of sorbitol within 1 minute ( fig . volume changes of the cells that were exposed to hypo - osmotic conditions were performed as the control measurements . upon exposure of cells to 225- and 150-mm extracellular solutions , cell volume after initial increase in the volume , the cell volume was gradually decreased toward the normal cell volume following a time course in the hypo - osmotic condition . in uvc irradiation induced and hyperosmotic stress ( 450 mm)stimulated cells , the peak volume changes were significant to show a volume shrinkage down to 67% and 65% in uvc irradiation induced and hyperosmotic pressure ( 450 mm)treated cells , respectively ( fig . however , applications of 4-ap had no significant effects on osmotic pressure induced initial volume changes ( fig . the results reveal that uvc irradiation induced a significant cell shrinkage comparable to the effect resulted from hyperosmotic pressure stimulation . effect of blocking kv channel on hyperosmotic pressure induced rabbit corneal epithelial cell volume changes . ( a ) real - time recording of hyperosmotic pressure induced changes in rabbit corneal epithelial cells . ( b ) effect of various osmotic pressures by changing sorbitol concentrations rabbit corneal epithelial cell volume . ( c ) comparison of cell volume changes in uvc irradiation and hyperosmotic pressure ( 450 mm of sorbitol)induced rabbit corneal epithelial cells . the time - points were selected for the peak time of cell volume changes for uvc exposure and hyperosmotic treatments . ( d ) comparison of 4-ap effect on uvc irradiation and hyperosmotic pressure induced rabbit epithelial cell volume changes . the effect of hyperosmotic pressure on src and fak phosphorylation was measured in human corneal epithelial cells . exposure of human corneal epithelial cells to hyperosmotic pressure resulted in increases in phosphorylation of src and fak following a time course ( fig . 5a ) . statistical analysis showed the peak time of increased phosphorylation for fak and src at 1 and 15 minutes , respectively ( fig . time courses of hyperosmotic pressure induced activation of src and fak were similar to uvc irradiation stimulated cells . furthermore , application of k channel blocker 4ap at a dosage of 1 mm had no significant effects on hyperosmotic pressure induced src and fak phosphorylation levels ( figs . 5c , 5d ) . in hyperosmotic pressure - induced cells , both src and fak phosphorylation were increased as they have been observed in uvc irradiation induced cell , suggesting that src and fak phosphorylation are cell volume change sensitive . effect of kv channel blockade on hyperosmotic pressure induced src and fak phosphorylation / activation in human corneal epithelial cells . effect of blockade of kv channel on uvc irradiation induced src phosphorylation / activation . ( c ) effect of 4-ap on hyperosmotic pressure induced fak phosphorylation in human corneal epithelial cells . ( d ) effect of 4-ap on hyperosmotic pressure induced src phosphorylation in human corneal epithelial cells . we predict that integrins in the edge area of the attached human corneal epithelial cells are affected by uvc irradiation induced cell shrinkage . activation of integrin 5 in uvc - irradiated cells was detected by immunostaining using the anti - integrin 5 antibody in the absence and presence of 4-ap . further immunostaining experiments were performed to verify uvc irradiation induced increases in phospho - fak stained by using the antibody that specifically recognizes phospho - fak ( fig . ultraviolet c irradiation induced increases of the phospho - fak were effectively prevented by preincubation of cells with 4-ap . blockades of kv channel by 4-ap resulting in suppression of uvc irradiation induced activations of integrin 5 and fak provide further evidence that uvc irradiation induced cellular changes are mediated by initial activation of k channels in uvc - exposed human corneal epithelial cells . immunostaining detection of uvc irradiation induced alterations of integrin and fak activities in human corneal epithelial cells . ( a ) ultraviolet c irradiation induced integrin 5 activation in the absence and presence of 4-ap in different scales . induced fak phosphorylation stained by red color and nuclei were stained with dapi in the absence and presence of 0.5-mm 4-ap in different scales . environmental stresses , such as uvc irradiation , hyperosmotic pressure , hypoxia / reoxygenation , and infection , cause damages of corneal epithelia cells and activate various signaling pathways that delay the process of corneal epithelial self - renewal and wound healing . we have previously studied a mechanism involving uvc irradiation induced hyperactivation of kv channels and activation of map kinase signaling pathways in corneal epithelial cells . studies in various cell types suggest that the cell volume can be affected by activations of membrane ion channels resulting in significant increases in k efflux occurs prior to the volume change . in addition , eliciting excessive k efflux or intracellular k depletion can result in apoptosis , which provides important evidence indicating that k channel activation plays a critical role to regulate cell fate . however , it still lacks detailed mechanism that can explain how the cell shrinkage occurs in response to uvc - stress stimulation , and how the linkage between the stress effect and cell shrinkage leads to activation of signaling pathways , resulting in alteration of cell fate . results obtained from previous patch clamp studies in corneal epithelial cells provide essential information about loss of intracellular k ions resulted from uvc irradiation induced hyper activation of k channels . theoretical calculation of cell volume based on our patch clamp data suggests that uvc irradiation induced hyper activation of k channels in corneal epithelial cells result in a loss of nearly 3.6% of the total intracellular k stock per second . if osmotic pressure change is considered , the change may be 10 mosm per second , which equals to 3% of the intracellular osmolarity . the predicted volume decreases by the theoretical calculation is verified by the real - time measurement of the volume changes in uvc - exposed corneal epithelial cells , suggesting that uvc irradiation induced k channel hyperactivation results in quick loss of intracellular k ions to decrease cell volume . in the present study , cell volume changes in corneal epithelial cells were measured in a real - time fashion with / without uvc irradiation by using a fluorescent dye calcein - am . it has been shown that calcein - am can be evenly distributed throughout the cell and used for the registration of volume alterations including epithelial cells . we observed cell volume changes in a real time measurement , indicating a more than 25% decrease in cell volume in the first a few seconds of the uvc exposure . induced cell shrinkage is similar to the volume decrease in the cells exposed to 50% increased extracellular osmotic pressure . induced cell volume alterations were effectively prevented by preincubation of cells with kv channel - specific blockers , such as 4-ap , -dtx , and bds-1 . shrinkages of uvc irradiation induced corneal epithelial cells undergo through a fast time course . thus , uvc irradiation induced activation of kv channels in the membrane plays a key role in triggering cell volume changes . our previous studies in corneal epithelial cells have provided strong evidence showing that exposure of corneal epithelial cells to uvc irradiation induces the complex signaling events finally leading to alterations of the cell fate and wound healing . the precise connections of uvc irradiation induced ion channel activation , volume change and cytoskeleton reorganization have not been completely revealed . here , we provide further evidence to demonstrate that exposure of corneal epithelial cells to uvc irradiation induces fast cell shrinkages . blockade of kv channel activity with specific blockers suppressed uvc irradiation induced : ( 1 ) cell shrinkage , ( 2 ) activation of scaffolding protein kinase cascades , and ( 3 ) structural alterations of cytoskeletal proteins in corneal epithelial cells . the result suggests that changes of cell volume due to uvc irradiation induced kv channel hyperactivity activate the fak - dependent integrin pathway that has been found to subsequently trigger cytoskeleton reorganization . because corneal epithelial cells are attached cells , it is very likely that cell shrinkage in response to uvc irradiation results in a partial ( or more precisely in the region of the cell edge ) detachment of corneal epithelial cells evidenced by formations of integrin 5 clusters in uvc irradiation induced cells . the leading edge of corneal epithelial cells is also the region where integrins first engage their ligands to activate the signaling pathways . further downstream , activation of the fak - dependent integrin signaling can also alter alignments of structural proteins , such as f - actin and -tubulin . structural changes of integrin and fak phosphorylation were detected in uvc irradiation induced cells by immunostaining using specific antibody against fak . induced integrin activation mainly in the edge of cells and formed a lot of clustering clouds in these areas , suggesting that uvc irradiation , it has been shown that the fak - dependent integrin - signaling pathway is activated during cell detachment and migration . induced cell size and volume changes activate src and fak signaling events that can determine cell survival . phosphorylations of fak and paxillin can be elicited in cultured cells by cyclical mechanical strain , and fak couples integrins and cytoskeletal proteins to multiple signaling pathways . in addition , several lines of evidence suggest that fak is required for stress stimulation to activate map kinase pathways including the jnk cascades . these studies suggest that stress - induced cellular changes including cell shrinkage may play key roles in the early event in the initiation of fak and src activation , and cytoskeleton remodeling . cell volume changes triggered activation of the fak - dependent integrin signaling and cytoskeleton reorganization , critically determining uvc irradiation induced early response in corneal epithelial cells . previous studies show that uvc irradiation induced hyperactivity of kv channels in these cells , blockade of kv channels with various kv channel blockers in uvc - exposed cells significantly suppressed uvc irradiation induced cell volume change , fak / src activation , and cytoskeleton remodeling . thus , a linkage between uvc irradiation induced cell volume alteration due to loss of intracellular k ions and activation of the fak - dependent integrin pathway can be established in corneal epithelial cells . it also helps to explain the observation of cytoskeleton reorganization in response to uvc stress . both scaffold kinases fak and src are important integrin receptor - linked tyrosine kinases to elicit cytoskeleton reorganization in many cell types . our data using pp2 to suppress src kinase activity is consistent with other lab 's results that src is responsible for further phosphorylation of fak at different tyrosine sites after the site of y397 is phosphorylated in these cells . we found that k channel blocker , 4-ap , effectively suppressed uvc irradiation induced increases in fak and src phosphorylation or activation at micromolar levels , indicating that activations of fak and src are downstream events resulted from uvc irradiation induced k channel hyperactivity and cell shrinkage because their activities were altered by suppression of kv channels in uvc irradiation induced cells . we believe that the connection between uvc irradiation induced activation of kv channel and scaffold protein kinase in stress - evoked pathways plays important roles in determining corneal epithelial function and fate . | purposethe purpose of the study is to understand how extracellular stresses , such as ultraviolet ( uv ) irradiation , affect corneal epithelial cells .
cell volume changes , damage to corneal epithelial integrity , and cellular responses were assessed after exposure to uvc stresses.methodsprimary human and rabbit corneal epithelial cells were exposed to uvc light in culture conditions . ultraviolet c irradiation induced changes in cell size and volume were measured by real - time microscopy and self - quenching of the fluorescent dye calcein , respectively .
the effects of uvc irradiation on src and focal adhesion kinase ( fak ) phosphorylation and fak - dependent integrin signaling were detected by elisa , immunoblotting , and immunostaining.resultsultraviolet c irradiation induced both size and volume shifts in human and rabbit corneal epithelial cells .
ultraviolet c irradiation - induced decrease of cell volume elicited activation of src and fak , characterized by increased phosphorylations of srcy416 , faky397 , and faky925 .
in addition , immunostaining studies showed uvc irradiation induced increases in phosphorylation of fak and formation of integrin 5 clustering .
application of kv channel blockers , including 4-aminopyridine ( 4-ap ) , -dtx , and depressing substance-1 ( bds-1 ) , effectively suppressed uvc irradiation induced cell volume changes , and subsequently inhibited uvc irradiation induced phosphorylation of src / fak , and formation of integrin 5 clustering , suggesting uvc irradiation induced volume changes and src / fak activation .
hyperosmotic pressure induced volume decreases were measured in comparison with effects of uvc irradiation on volume and src / fak activation .
however , kv channel blocker , 4-ap , had no effect on hyperosmotic pressure
induced responses.conclusionsthe present study demonstrates that uvc irradiation induced decreases in cell volume lead to src / fak activation due to a rapid loss of k ions through membrane kv channels . |
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spontaneous hypertensive cerebellar hemorrhage ( shch ) accounts for 5 - 10% of all cases of intracerebral hemorrhage ( ich).7 ) the mortality rate of shch is reportedly 20 - 75% higher than that of supratentorial ich.4 ) this high mortality rate results from direct compression of the hematoma by the brain stem and acute hydrocephalus due to hematoma extension into the fourth ventricle or direct compression of the fourth ventricle.17 ) in patients with a large volume shch , a large suboccipital craniectomy ( soc ) is frequently used for hematoma evacuation.9)14 ) due to the long operating time and operation - induced tissue damage , this procedure is particularly stressful for elderly patients . in addition , because of the large skin incision and durotomy required for the procedure , it can result in pseudomeningocele . because burr hole aspiration surgery is less invasive , we thought that it may result in improved clinical outcomes and lowered complication rates . the aim of this study is to investigate the clinical outcomes of patients treated with burr hole aspiration surgery using a stereotactic frame or navigation system . we conducted a retrospective study of the medical records of 26 patients with shch who visited our institution between jan 2002 and dec 2011 . all 26 patients who underwent stereotactic or navigation - guided burr hole aspiration surgery and catheter insertion were included in our study . on admission , all patients underwent standard neurological and clinical examinations , as well as routine laboratory tests . some patients underwent ct angiography or magnetic resonance imaging in order to exclude aneurysm rupture , cavernous angioma , arteriovenous malformation , tumor , or hemorrhagic infarction . in all patients , we evaluated variable factors , including the maximum diameter of the hematoma , the presence of hydrocephalus , brain stem compression , compression of the fourth ventricle , and the presence of intraventricular hemorrhage ( ivh ) . the volume of the hematoma was measured by calculating the area occupied by the hyperdense hematoma on each slice of the brain ct images and multiplying the area by the thickness of each slice . the indications for surgical treatment were provided by the american heart association / american stroke association ( aha / asa ) . in short , surgery is recommended if the patient showed neurological deterioration and the maximum diameter of the hematoma was greater than 3 cm.2 ) ( fig . 1a ) if the patient exhibited progressive mental deterioration associated with hydrocephalus caused by ivh or fourth ventricle obliteration , we performed external ventricular drainage ( evd ) . the glasgow outcome scale ( gos ) was used for assessment of patients ' outcomes and complications were checked after six months . based on findings on the brain ct , we attempted to find the shortest trajectory targeting the hematoma center , and avoiding the venous sinus and midline . under general anesthesia , then , using the leksell stereotactic system ( elekta instrument ab , stockholm , sweden ) or the vectorvision navigation system ( brainlab , feldkirchen , germany ) , we performed catheter insertion through the burr hole . an intradural catheter was placed in the center of the hematoma . using a 10 cc syringe , careful manual aspiration of hematoma the catheter was connected with a conventional cerebrospinal fluid ( csf ) collection system . if the patient was satisfied with our evd indication , we performed evd through kocher 's point before aspiration surgery or through the posterior parietal point after aspiration surgery . each day following surgery , we examined the color and amount of the drained hematoma . if the volume of the hematoma was effectively drained every day , we checked brain ct at three days after surgery ( fig . if a substantial amount of hematoma still remained on the ct images , it was resolved with 5,000 iu and fibrinolyzed twice daily for two or three days , and the remaining hematoma was drained each day . in cases where the hematoma was especially thick and difficult to drain , we also used urokinase similarly for three days following the day after surgery . when the hematoma was not almost removed on follow - up brain ct , we drained the catheter for two additional days . we conducted a retrospective study of the medical records of 26 patients with shch who visited our institution between jan 2002 and dec 2011 . all 26 patients who underwent stereotactic or navigation - guided burr hole aspiration surgery and catheter insertion were included in our study . on admission , all patients underwent standard neurological and clinical examinations , as well as routine laboratory tests . some patients underwent ct angiography or magnetic resonance imaging in order to exclude aneurysm rupture , cavernous angioma , arteriovenous malformation , tumor , or hemorrhagic infarction . in all patients , we evaluated variable factors , including the maximum diameter of the hematoma , the presence of hydrocephalus , brain stem compression , compression of the fourth ventricle , and the presence of intraventricular hemorrhage ( ivh ) . the volume of the hematoma was measured by calculating the area occupied by the hyperdense hematoma on each slice of the brain ct images and multiplying the area by the thickness of each slice . the indications for surgical treatment were provided by the american heart association / american stroke association ( aha / asa ) . in short , surgery is recommended if the patient showed neurological deterioration and the maximum diameter of the hematoma was greater than 3 cm.2 ) ( fig . 1a ) if the patient exhibited progressive mental deterioration associated with hydrocephalus caused by ivh or fourth ventricle obliteration , we performed external ventricular drainage ( evd ) . the glasgow outcome scale ( gos ) was used for assessment of patients ' outcomes and complications were checked after six months . based on findings on the brain ct , we attempted to find the shortest trajectory targeting the hematoma center , and avoiding the venous sinus and midline . under general anesthesia , then , using the leksell stereotactic system ( elekta instrument ab , stockholm , sweden ) or the vectorvision navigation system ( brainlab , feldkirchen , germany ) , we performed catheter insertion through the burr hole . an intradural catheter was placed in the center of the hematoma . using a 10 cc syringe , careful manual aspiration of hematoma if the patient was satisfied with our evd indication , we performed evd through kocher 's point before aspiration surgery or through the posterior parietal point after aspiration surgery . each day following surgery , we examined the color and amount of the drained hematoma . if the volume of the hematoma was effectively drained every day , we checked brain ct at three days after surgery ( fig . if a substantial amount of hematoma still remained on the ct images , it was resolved with 5,000 iu and fibrinolyzed twice daily for two or three days , and the remaining hematoma was drained each day . in cases where the hematoma was especially thick and difficult to drain , we also used urokinase similarly for three days following the day after surgery . when the hematoma was not almost removed on follow - up brain ct , we drained the catheter for two additional days . burr hole catheter insertion using the stereotactic and navigation - guided systems was performed in 10 and 16 patients . the mean hematoma volume on brain ct at admission was 21.8 5.8 cc , and the mean diameter of the hematomas on brain ct was 45.8 8.7 mm . seven of 26 patients had ivh and non - communicating hydrocephalus . the mean preoperative gcs score was 12.5 1.3 and the mean postoperative gcs score was 13.1 1.2 . in 19 cases , using only insertion of the catheter in the hematoma without evd , the mean operation time was 43.1 8.9 min . a mean the average volume of hematoma evacuated through the catheter during the operation was 9.3 1.3 ml . however , in 14 cases , because the residual hematoma volume indicated by the brain ct exceeded 10 cc , we performed urokinase irrigation . we performed an evd with placement of a catheter in seven cases and kept it in place for four days on average . on postoperative day seven , the mean follow - up duration was 23.7 15.2 months and the gos score was checked on all patients for an average of six months following surgery . twenty three of the 26 patients showed good recovery ( gr ) ; one patient was mildly disabled ( md ) , one patient was severely disabled ( sd ) , and one patient died ( d ) . the overall mortality rate was 3.8% and the mean gos score was 4.6 1.0 . the patient died of sudden myocardial infarction ( mi ) ; however , mental status had improved from deep drowsy to drowsy before occurrence of mi . among 25 patients with a high gcs score over 10 points at admission , there was no occurrence of surgery - related complications , including re - bleeding , infection , and pseudomeningocele . because it can compress the brainstem directly as well as generate acute hydrocephalus , shch is associated with high mortality rates . if certain diagnostic indications are satisfied , early operation for treatment of shch is strongly recommended.1)4)10 ) according to the 2007 aha / asa guidelines for management of ich in adults,3 ) the surgical indication for sch is patients with cerebellar hemorrhage > 3 cm who show neurological deterioration , or who have brain stem compression and/or hydrocephalus resulting from ventricular obstruction . in cases involving acute massive bleeding , soc is typically favorable for hematoma evacuation and brainstem decompression.3 ) soc has the advantage of assuring effective decompression and direct hematoma evacuation , however , it requires long operation times and incurs additional risk from operation - induced tissue damage . suboccipital craniectomy can also result in patient instability , additional brain injury , re - bleeding , pseudomeningocele due to durotomy , and wound infection in the postoperative period.15 ) important factors affecting the prognosis of patients with cerebellar hemorrhage include the mental status of the patient at admission , a decrease in consciousness , the amount of hematoma with or without ivh , the presence of obstructive hydrocephalus and brain stem compression or obliteration of quadrigeminal cistern.5)6)8)11)15)16 ) advantages of burr hole aspiration surgery include the short operation time , lowered risk of additional brain injury , small size of incision , lower complication rate , and the fact that it can be performed without durotomy . in our opinion , this operation is appropriate for patients who have mild brain stem compression , can not tolerate long periods of operation , or have a poor medical condition or coagulopathy , as well as those at risk for fourth ventricle obstruction due to progression of brain swelling but still have patency of csf flow . in 1980 , several institutions introduced stereotactic approaches for treatment of shch.12)13 ) according to mohsen et al.12 ) , who studied patients with good gcs scores ( above 7 ) , scores for all patients improved to above 11 . in our study , among 25 patients with gcs scores of over 10 , gcs score for all patients showed improvement to above 12 . in addition , the overall mortality of patients was only 3.8% , and there was no occurrence of surgery - related complications . even in the one case in which the patient passed away , the cause of death was a myocardial infarction rather than a complication of the shch or surgery . however , this technique is restricted in the acute stage because direct hemostasis and confirmation of the source of the bleeding can not be achieved.18 ) re - bleeding may occur if stereotactic aspiration is performed too soon after the onset of hemorrhage . the primary limitation of the current study is the lack of control groups and randomized multi - center case control series . conduct of further research is required in order to validate our findings against these controls . we suggest that , compared to conventional surgical approaches , aspiration surgery for treatment of shch can reduce the approaching route , operation time , and complication rates , such as injury of brain tissue and psudomeningocele . therefore , it can be regarded as an alternative surgical option for treatment of shch in selected patients . | objectivepatients with severe spontaneous cerebellar hemorrhage typically undergo treatment with suboccipital craniectomy and hematoma evacuation . however , this is a stressful procedure for patients due to the long operating time and operation - induced tissue damage .
in addition , the durotomy can result in pseudomeningocele .
we investigated the efficacy of stereotactic or navigation - guided burr hole aspiration surgery as a treatment for spontaneous hypertensive cerebellar hemorrhage ( shch).methodsbetween january 2002 and december 2011 , 26 patients with shch underwent surgery using the stereotactic or navigation - guided burr hole aspiration and catheter insertion technique in our institution.resultsmean hematoma volume was 21.8 5.8 cc at admission and 13.1 5.4 cc immediately following surgery .
preoperative glasgow coma scale ( gcs ) score was 12.5 1.3 and postoperative gcs score was 13.1 1.2 .
seven days after surgery , the mean hematoma volume was 4.3 5.6 cc , and there was no occurrence of surgery - related complications during the six - month follow - up period .
the mean operation time for catheter insertion was 43.1 8.9 min , and a mean 31.3 6.0 min was also added for extra - ventricular drainage .
the mean glasgow outcome scale ( gos ) score after six months was 4.6 1.0.conclusionstereotactic burr hole aspiration surgery for treatment of shch is less time - consuming and invasive than other interventions , and resulted in no surgery - related complications .
therefore , we suggest that this surgical method could be a safe and effective treatment option for selected patients with shch . |
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we encountered a woman with a preexisting large focal nodular hyperplasia ( fnh ) of the liver , persisting during two separate pregnancies . her elevated serum -glutamyltransferase and alkaline phosphatase levels before pregnancy were reduced during both pregnancies , but returned to prepregnancy levels after delivery . focal nodular hyperplasia ( fnh ) is a benign lesion of the liver that is not uncommon in adults and is frequently observed in healthy individuals . it is unclear , however , whether pregnancy in women with fnh increases the risks of fnh growth and complications or alters hepatic function in patients . we encountered a woman with a large fnh , who experienced two pregnancies and delivered healthy infants at term uneventfully . the courses of her pregnancies and the effects of pregnancy on fnh , including lesion size and hepatic function were evaluated at the same institution . the patient was a 25-year - old , gravida 0 , para 0 , female , who had been followed up since the age of 18 years at the internal medicine outpatient ward of our institution because of a large fnh . at the age of 13 years , a nodule measuring 8 cm was observed in the patient , at a medical institution near her residence . a biopsy was performed , and on histological analysis of the specimen , she was diagnosed with fnh . because the nodule was located near the hepatic hilus , surgical intervention was not performed . at the age of 25 years , the patient married and moved near our hospital . an mri taken before her first pregnancy showed that the nodule was 11 cm in diameter ( figs.1a and 2a ) . a high intensity area was observed at the center of the lesion , a finding compatible with central scar tissue , with a moderately high intensity area observed at the surface of the lesion . ( c ) during her second pregnancy at 34 weeks of gestation . before becoming pregnant , her liver function tests showed that her serum aspartate aminotransferase ( ast ) and alanine aminotransferase ( alt ) concentrations were in the normal ranges , but her -glutamyltransferase ( ggt ) and alkaline phosphatase ( alp ) concentrations were elevated , to 651 iu / l and 407 u / l , respectively . after conceiving , she was referred by her physician to our outpatient ward at 9 weeks of gestation . during her first pregnancy , her serum concentrations of liver enzymes decreased to the normal range without any medication ( fig.3 ) . magnetic resonance imaging performed at 37 weeks of gestation showed that the size of the fnh was unchanged , with no signs of bleeding or degeneration of the lesion ( figs.1b and 2b ) . the concentrations of aspartate aminotransferase ( ast ) , alanine aminotransferase ( alt ) , lactate dehydrogenase ( ldh ) , alkaline phosphatase ( alp ) , and -glutamyltransferase ( ggt ) are shown in u / l . , she delivered a normal , healthy female infant , weighing 3084 g and with 1 and 5 min apgar scores of 8 and 9 , respectively , by vaginal delivery . her postpartum recovery was normal , except that her serum ggt and alp concentrations became elevated , returning to their prepregnancy levels . this pregnancy was also uneventful ( figs.1c , 2c , and 3 ) and , at 40 weeks of gestation she delivered a normal , healthy male infant , weighing 3130 g and with 1 and 5 min apgar scores of 9 and 10 , respectively , by vaginal delivery . her postpartum course was again uneventful , and both mother and child were discharged after 5 days . the size of the fnh remained the same as before her first pregnancy , despite two pregnancies and deliveries . during both pregnancies , her serum concentrations of liver enzymes after conceiving , she was referred by her physician to our outpatient ward at 9 weeks of gestation . during her first pregnancy , her serum concentrations of liver enzymes decreased to the normal range without any medication ( fig.3 ) . magnetic resonance imaging performed at 37 weeks of gestation showed that the size of the fnh was unchanged , with no signs of bleeding or degeneration of the lesion ( figs.1b and 2b ) . the concentrations of aspartate aminotransferase ( ast ) , alanine aminotransferase ( alt ) , lactate dehydrogenase ( ldh ) , alkaline phosphatase ( alp ) , and -glutamyltransferase ( ggt ) are shown in u / l . her first course of pregnancy was uneventful . at 39 weeks , she delivered a normal , healthy female infant , weighing 3084 g and with 1 and 5 min apgar scores of 8 and 9 , respectively , by vaginal delivery . her postpartum recovery was normal , except that her serum ggt and alp concentrations became elevated , returning to their prepregnancy levels . this pregnancy was also uneventful ( figs.1c , 2c , and 3 ) and , at 40 weeks of gestation she delivered a normal , healthy male infant , weighing 3130 g and with 1 and 5 min apgar scores of 9 and 10 , respectively , by vaginal delivery . her postpartum course was again uneventful , and both mother and child were discharged after 5 days . the size of the fnh remained the same as before her first pregnancy , despite two pregnancies and deliveries . during both pregnancies , her serum concentrations of liver enzymes focal nodular hyperplasia is a common type of hepatic mass in adults , with an incidence of approximately 3% . we encountered a woman who had fnh prior to her first pregnancy and followed her throughout two courses of pregnancy . in contrast , the serum concentrations of liver enzymes were decreased during both pregnancies , and returned to their prepregnancy levels soon after the deliveries . a previous study described the effects of 41 pregnancies on fnh in 37 women , finding that the size of the fnh was not affected by pregnancy . another study evaluating the effects of pregnancy on fnh size in patients found that the mean tumor size before pregnancy was 58.5 22.7 mm . tumor size remained constant in seven of these 20 patients ( 35% ) and declined in 10 ( 50% ) . however , fnh size in multiparous women was assessed using a questionnaire , not by actual measurement . furthermore , that study evaluated only lesion size , not changes in serum enzyme concentrations . in our patient , the fnh was > 11 cm in diameter , the largest fnh reported to date , and its size was not affected by either of her two pregnancies . there have been no reports on changes in the serum concentrations of liver enzymes during pregnancy in women with fnh . in contrast , serum alp and ggt values , which were elevated before and after her pregnancies , decreased to normal levels during her pregnancies . alp and ggt are enzymes present in the cell membranes of the bile duct , gall bladder , pancreas , and kidneys . serum concentrations of alp elevate by placental origin and ggt are not affected in pregnancies in women without fnh , but were shown to be higher in few pregnant women ( < 1% ) with intrahepatic cholestasis . pregnancy may alter the results of liver function tests , with placental steroids playing a role . during pregnancy , the production of placental steroids increases the serum estrogen and progesterone concentrations compared with the nonpregnant state . for example , a 9-year study of 216 women in france showed that neither the size nor the numbers of fnh lesions are influenced by oral contraceptive use . although estrogen receptors were shown to be present in the cytosol and nuclei of hepatic cells including the tissue of fnh , few estrogen receptors are present on the cell surface . estrogen and progesterone affect the mrna expression and activity of cyps ( cytochrome p450 ) in primary human hepatic cells , as shown in a recent study . these findings potentially provide a basis to understand the mechanism of altered drug metabolism during pregnancy . however , further studies are needed to investigate the lowering of serum alp and ggt in fnh . in conclusion , we have described here a single patient with fnh of the liver complicating pregnancy and postpartum delivery twice over a 4-year observation period . fnh size was not affected by pregnancies , whereas liver function test results improved , allowing these enzymes to return to their prepregnancy levels after deliveries . lesion size and abnormally elevated serum liver enzyme concentrations are not indicated for surgery in pregnant women with asymptomatic fnh . our findings indicate that women with large fnh , even those with elevated serum liver enzymes , should not be discouraged from pregnancy . | key clinical messagewe encountered a woman with a preexisting large focal nodular hyperplasia ( fnh ) of the liver , persisting during two separate pregnancies .
fnh size was not affected by either pregnancy .
her elevated serum -glutamyltransferase and alkaline phosphatase levels before pregnancy were reduced during both pregnancies , but returned to prepregnancy levels after delivery . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
controlled - release ( cr ) formulations have been introduced into drug therapy with two main purposes : to reduce the number of single doses per day improving patient compliance of treatments and to decrease the fluctuations of plasma levels , in order to obtain better therapeutic efficacy and lower toxicity . there are many controlled - release pharmaceutical systems currently known , ranging from monolithic matrices , membrane reservoirs , erodible polymers , to the more technologically complex and sophisticated ph independent formulations , ion exchange resins , osmotically , and geometrically modified systems . many of these systems are not produced in a form that is amenable to large - scale manufacturing processes and usually do not exhibit the desirable zero - order release kinetics . in addition , the cost of formulation development , raw materials , and manufacture technology are among the principal factors in cr delivery systems formulation for oral dosing . an interesting approach to develop cr matrix formulations is based on melt granulation , which is a very short and easy one - step technique converting fine powders into granules . the powder agglomeration is promoted by the addition of a low melting point binder , which is solid at room temperature and melts at relatively low temperatures ( 5080c ) . the interest in melt granulation has increased due to the advantages of this technique over other cr delivery technologies . since it is a solvent - free process , the drying phase is eliminated , and thus it becomes less consuming in terms of time and energy [ 2 , 3 ] . zolpidem tartrate is a gaba agonist ( sedative and hypnotic ) used in the treatment of insomnia dosing ranging from 5 to 12.5 mg . the half life of the drug is about 1.9 to 3 hr , and oral bioavailability is 72 7% indicating its promising candidature for the controlled - release formulation . zolpidem was marketed as the immediate release product in the short - term treatment of insomnia . zolpidem is effective in reducing the time to sleep onset and increasing total sleep time ; however , its effect on sleep maintenance has not been consistently demonstrated . the hypnotic effects of zolpidem have been reported primarily in the first 3 hours postdose which can lead to subtherapeutic effects on sleep maintenance in the later portion of the night for some patients . so , it is desired to maintain plasma concentration of drug for 8 hr . moreover , melt granulation is one of the most widely applied processing techniques in the array of pharmaceutical manufacturing operations due to its simplicity and easy scaleup [ 68 ] . in recent years , melt granulation has also been successfully employed to improve the dissolution rate of poorly soluble compounds increasing the bioavailability of these kinds of drugs , [ 911 ] and in the development of cr formulations [ 1214 ] and masking the bitter taste of an active drug [ 15 , 16 ] . hence , the purpose of present investigation was to develope controlled - release tablet of zolpidem tartrate by using polyethylene glycol ( peg 6000 ) as melt binder , hydroxypropyl methylcellulose ( hpmc k4 m ) and polyvinylpyrrolidone ( pvp k30 ) as matrixing agent and filler , respectively , which would release the drug for prolonged period of time in view to maximize therapeutic effect of the drug and in an effort to expand the coverage of sleep complaints and overcome the lack of efficacy in sleep maintenance . lactose , magnesium stearate , and talc were purchased from shakti chemicals , mehsana , india . all other materials and chemicals used were of either pharmaceutical or analytical grade . accurately weigh peg 6000 was melted in a porcelain dish at 5560c on heating metal , and the accurate quantity of zolpidem was added to the melted mass of peg . previously prepared geometric mixture by tumbling method using spatula for 14 minutes of hpmc k4 m , pvp , and lactose was added to the molten zolpidem - peg 6000 mixture and stirred well to mix . then mass was removed from the hot plate and subjected to scrapping until it attained room temperature . the coherent mass was passed through 22 mesh , and the resulting granules were resifted over 44 meshes to separate granules and fines . the % loss of mass during melt granulation was found between 15 and 20% of total weight . the lubricated blend was compressed using 8 mm round flat punch on 10 station rimek - i rotary tablet machine ( karnavati engineering , kadi , india ) . compression was adjusted to obtain tablets with hardness in the range of 3 - 4 kg / cm . the fabricated tablets were characterized for weight variation ( n = 20 ) , hardness ( n = 6 ) pfizer hardness tester ( janki instrument ltd , ahmedabad , india ) , thickness using a screw - gauge micrometer ( campbell electronics , mumbai , india ) , and % friability ( n = 20 , roche friabilator , electrolab , mumbai , india ) . the in vitro dissolution study of zolpidem tablets ( n = 3 ) was performed as described in indian pharmacopoeia 2010 using usp apparatus ii ( model tdt-08 t , electrolab , mumbai , india ) fitted with paddle ( 50 rpm ) at 37c 0.5c using simulated gastric fluid ( ph 1.2 ; 900 ml ) as a dissolution medium for first 2 hours and followed by phosphate buffer ( ph 6.8 ; 900 ml ) for remaining hours . at the predetermined time intervals , 10-ml samples were withdrawn and analyzed at 238 nm using a shimadzu uv 1800 double - beam spectrophotometer ( shimadzu , kyoto , japan ) . cumulative percentage drug release was calculated using an equation obtained from a calibration curve which is developed in the range of 216 g / ml for 0.1 n hcl and ph-6.8 phosphate buffer ( see figure 1 ) . a 3 randomized full factorial design was employed in the present study . in this design , 2 factors were evaluated , each at 3 levels , and experimental trials were performed for all 9 possible combinations . the ratio of polymer ( hpmc k4 m : pvp ) ( x1 ) and concentration of melt binder ( peg 6000 ) ( x2 ) were chosen as independent variables in 3 full factorial design , while q1 , q4 , and q8 ( % drug release after 1 , 4 , and 8 hours , resp . ) , diffusion coefficient ( n ) , and release rate constant ( k ) were taken as dependent variables . the composition of factorial design batches ( f1 f9 ) is shown in table 1 . the prepared formulations were evaluated for assay , friability , and hardness and in vitro release study . statistical treatment was carried out to the factorial design batches using design expert dx8 software . the dissolution profile of all batches was fitted to various models such as zero order , first order , higuchi , hixon and crowell , and korsmeyer et al . the similarity factor ( f2 ) given by supac guidelines for a modified release dosage form was used as a basis to compare dissolution profiles . the dissolution profiles are considered to be similar when f2 is between 50 and 100 . the dissolution profile of products was compared using an f2 which is calculated from following formula :
( 1)f2=50log{[1+(1n)t=1nwt(rttt)2]0.5100 } ,
where n is the dissolution time , and rt and tt are the reference ( here , this is the theoretical dissolution profile of zolpidem ) and test dissolution value at time t . fourier transform infrared ( ftir ) spectra of zolpidem tartrate ( see figure 7 ) and granules of optimized batch were recorded using kbr mixing method on ftir ( ftir-1700 , shimadzu , kyoto , japan ) for drug excipients interaction study ( see figure 8) . from the in vitro dissolution study , it was found that hydrophobic binder mcc wax and bees wax have more sustaining effect on the release of drug than stearic acid and cetyl alcohol it is due to its hydrophobic nature . hydrophilic binder peg-6000 gave good drug release compared to all the other binders , which is due to its hydrophilic nature . hpmc k4 m ( hydrophilic ) was selected as a matrixing agent considering its widespread applicability and excellent gelling activity in controlled - release formulations . pvp was also selected in formulation because it helps in releasing loading dose from the formulation in the 1st hour which is required for the therapeutic effect of formulation . a statistical model incorporating interactive and polynominal terms was used to evaluate the responses
( 2)y = b0+b1x1+b2x2+b12x1x2+b11x12+b22x22 ,
where y is the dependent variable , b0 is the arithmetic mean response of the 9 runs , and b1 is the estimated coefficient for the factor xi . the main effects ( x1 and x2 ) represent the average result of changing 1 factor at a time from its low to high values . the interaction terms ( x1x2 ) show how the response changes when two factors are simultaneously changed . the dissolution profile for 9 batches showed a variation ( i.e. , initial 1 hr release ranging from 43.22% to 81.33% and drug release after 8 hr ranging from 96.82% to 100% ) . the fitted equations ( full and reduced ) relating the responses , q1 , q4 , and q8 , diffusion coefficient ( n ) , and release rate constant ( k ) to the transformed factor are shown in the table 3 . the polynomial equations can be used to draw conclusions after considering the magnitude of coefficient and the mathematical sign it carries ( i.e. , negative or positive ) . table 4 shows the results of analysis of variance ( anova ) , which was performed to identify insignificant factors . r
values for q1 , q4 , diffusion coefficient ( n ) , and release rate constant ( k ) are 0.7774 , 0.7122 , 0.8135 , and 0.7867 , respectively , indicating good correlation between dependent and independent variables . the low r value , 0.6055 for q8 , indicates poor correlation between dependent and independent variables showing that drug release at 8 hr is less dependent on selected variables . the reduced models were developed for response variables by omitting the insignificant terms with p > .1000 . the terms with p < .1000 were considered statistically significance and retained in the reduced model . the coefficients for full and reduced models for response variables are shown in table 4 . the significance levels of the coefficients in the q8 were found to be insignificant at p > .1000 and , hence , do not contribute significant information to the prediction of q8 . the significance levels of the coefficients b1 , b11 , b22 , and b12 were found to be p = .4583 , .9722 , .4441 , and .9534 , respectively , so they were omitted from the full model to generate a reduced model . the coefficients b0 and b2 were found to be significant at p < .1000 ; hence , they were retained in the reduced model . the reduced model was tested in proportion to determine whether the coefficients b1 , b11 , b12 , and b22 contribute significant information to the prediction of q1 . the critical value of f for = 0.1 is equal to 5.34 ( df = 4,3 ) . since the calculated value ( f = 0.414 ) is less than the critical value ( f = 5.34 ) , it may be concluded that the interaction terms b1 , b11 , b12 , and b22 do not contribute significantly to the prediction of q1 and can be omitted from the full model to generate the reduced model . the significance levels of the coefficients b1 , b11 , b22 , and b12 were found to be p = .6023 , .5762 , .390 , and .9344 , respectively , so they were omitted from the full model to generate a reduced model . the coefficients b0 , and b2 were found to be significant at p < .1000 ; hence , they were retained in the reduced model . the reduced model was tested in proportion to determine whether the coefficients b1 , b11 , b12 , and b22 contribute significant information to the prediction of q4 . the critical value of f for = 0.1 is equal to 5.34 ( df = 4,3 ) . since the calculated value ( f = 0.4172 ) is less than the critical value ( f = 5.34 ) , it may be concluded that the interaction terms b1 , b11 , b12 and b22 do not contribute significantly to the prediction of q4 and can be omitted from the full model to generate the reduced model . the significance levels of the coefficients b1 , b11 , b22 , and b12 were found to be p = .4318 , .8711 , .4198 , and 1.000 , respectively , so they were omitted from the full model to generate a reduced model . the coefficients b0 and b2 were found to be significant at p < .1000 ; hence , they were retained in the reduced model . the reduced model was tested in proportion to determine whether the coefficients b1 , b11 , b12 , and b22 contribute significant information to the prediction of diffusion coefficient ( n ) . the critical value of f for = 0.1 is equal to 5.34 ( df = 4,3 ) . since the calculated value ( f = 0.5717 ) is less than the critical value ( f = 5.34 ) , it may be concluded that the interaction terms b1 , b11 , b12 , and b22 do not contribute significantly to the prediction of diffusion coefficient ( n ) and can be omitted from the full model to generate the reduced model . the significance levels of the coefficients b1 , b11 , b22 , and b12 were found to be p = .4880 , .9164 , .4095 , and .9882 , respectively , so they were omitted from the full model to generate a reduced model . the coefficients b0 and b2 were found to be significant at p < .1000 ; hence , they were retained in the reduced model . the reduced model was tested in proportion to determine whether the coefficients b1 , b11 , b12 , and b22 contribute significant information to the prediction of release rate constant ( k ) . the critical value of f for = 0.1 is equal to 5.34 ( df = 4,3 ) . since the calculated value ( f = 0.5717 ) is less than the critical value ( f = 5.34 ) , it may be concluded that the interaction terms b1 , b11 , b12 , and b22 do not contribute significantly to the prediction of release rate constant ( k ) and can be omitted from the full model to generate the reduced model . to demonstrate graphically the effect of the ratio of polymer ( hpmc k4 m : pvp ) and concentration of melt binder ( peg 6000 ) , the response surface plots were generated by using design expert 8.0.2 trial version software for the dependent variables q1 , q4 , q8 ( % drug release after 1 , 4 , 8 hours , resp . ) , diffusion coefficient ( n ) , release rate constant ( k ) and shown in figures 2 , 3 , 4 , 5 , and 6 respectively . the kinetics of the dissolution data were well fitted to zero order , higuchi model , and krossmayer - peppas model as evident from regression coefficients in table 5 . in case of the controlled - release formulations , diffusion , swelling , and formulation containing swelling polymers show swelling as well as diffusion mechanism because the kinetic of swelling includes relaxation of polymer chains and imbibitions of water , causing the polymer to swell and changing it from a glassy to rubbery state . the value of diffusion exponent n for most factorial formulations is between 0.084 and 0.379 ( table 5 ) indicating fickian drug release from the formulations [ 23 , 24 ] . the values of similarity factor ( f2 ) for the batch f7 showed maximum 72.22 ( table 2 ) ; hence , it was selected as optimum batch . it was observed that there were no changes in these main peaks in the ftir spectra of a mixture of drug and polymers ( figure 3 ) ; hence , it was concluded that there were no physical or chemical interactions of zolpidem with peg 6000 , pvp , and hpmc k4 m . from the present investigation , it was concluded that the concentrations of peg 6000 as a melt binder have more pronounced effect than the ratio of hpmc k4 m and pvp k30 polymers on drug release from controlled - release tablet formulation and are useful to produced tablet dosage form with desirable drug release pattern . | the present investigation describes the influence of the concentration of peg 6000 as a melt binder and ratio of hpmc k4 m : pvp on zolpidem tartrate controlled - release tablet formulations using 32 full factorial design .
the ratio of hpmc k4 m and pvp k30 ( x1 ) and the concentration of melt binder ( x2 ) were selected as independent variables , and drug release at 1 hr ( q1 ) , 4 hr ( q4 ) , 8 hr ( q8 ) , diffusion coefficient ( n ) , and release rate constant ( k ) were selected as a dependent variable .
tablets were prepared by melt granulation technique and evaluated for various evaluation parameters .
it was observed that concentration of melt binder had significant effect on q1 , q4 , n , and k binder concentration 25% w / w was found optimum .
optimized formulation ( f7 ) showed good similarity with theoretical profile of drug .
the x2 variable had a significant effect on dependent variables , and the x1 variable had no significant effect on dependent variables . |
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interferon- ( alfaron , recombinant ifn-2b ; hualida biotechnology co. , tianjin , china ) had a specific activity of 3 10 units / ml and > 99% purity . the selective egfr - tki gefitinib ( gd760 , iressa ) , kindly provided by astrazeneca ( macclesfield , cheshire , uk ) , was dissolved in dmso to a working concentration of 20 mm . both drugs were stored at 20c in tightly sealed sterile tubes and diluted to the desired concentration in culture medium ( rpmi 1640 ; gibco , grand island , ny , usa ) before being added to the cells . four human nsclc cell lines were used : a549 , h1299 , and hcc827 ( cell bank of chinese academy of sciences , shanghai , china ) , which have different egfr and k - ras gene statuses ( exons 1821 of the egfr gene and exons 23 of the k - ras gene were sequenced , a549 cell line harbours mutation ( g12s ) of k - ras , hcc827 cell line carries egfr 19del , and the egfr and k - ras of h1299 cell line are all wild type , data not shown ) . the cell lines were grown in rpmi 1640 ( gibco ) supplemented with 10% foetal bovine serum and maintained at 37c in a humidified atmosphere containing 5% co2 and 95% air . four different experimental regimens were investigated to evaluate the impact of ifn- pretreatment on the antiproliferative effect of gefitinib : i , 72-hour incubation with gefitinib or ifn- alone;ii , 72-hour incubation with gefitinib ( at the indicated concentrations in figure 2 ) + 10 iu / ml ifn- ( at the concentration of 10 iu / ml , ifn- inhibits the proliferation of these lung cancer cells obviously);iii , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with phosphate - buffered saline ( pbs ) , followed by 72-hour incubation with gefitinib + 10 iu / ml ifn-;iv , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with pbs , followed by 72-hour incubation with gefitinib . i , 72-hour incubation with gefitinib or ifn- alone ; ii , 72-hour incubation with gefitinib ( at the indicated concentrations in figure 2 ) + 10 iu / ml ifn- ( at the concentration of 10 iu / ml , ifn- inhibits the proliferation of these lung cancer cells obviously ) ; iii , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with phosphate - buffered saline ( pbs ) , followed by 72-hour incubation with gefitinib + 10 iu / ml ifn- ; iv , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with pbs , followed by 72-hour incubation with gefitinib . control cells were processed in the same manner but with drug - free medium ( placebo ) instead of gefitinib . the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide ( mtt)-tetrazolium dye assay was used to evaluate cell viability . 100 l exponential growth phase cells ( about 3500 cells in total ) were seeded into 96-well plastic plates , incubated overnight , and added 100 l serial dilutions of gefitinib , ifn- , or gefitinib + ifn- into each well . after 72-hour incubation at 37c , 20 l 5 mg / ml mtt solution was added into each well and the cells were incubated for four hours . the media were removed and 150 l dmso was added into each well to dissolve the mtt completely , and the absorbance values were read at 570 nm using a microplate reader ( model 680 ; bio - rad , hercules , ca , usa ) . at least three replicate wells were used for each drug concentration , and the experiment was carried out independently at least three times . cell proliferation is expressed as the percentage of drug - treated surviving cells versus control cells ( in which viability was considered 100% ) . cells were incubated for 72 hours in drug - free medium ( control ) , 10 iu / ml ifn- , gefitinib alone , or gefitinib after 10 iu / ml ifn- pretreatment . according to the ic50 of gefitinib in these three cell lines , the concentration of gefitinib for hcc827 cell was 20 nm , and for other two cell lines it was 10 m . the cells were harvested by trypsinisation , washed twice with pbs , incubated in 500 l binding buffer and 10 l annexin v fitc at room temperature for 30 minutes , and then 5 l pi was added and incubated for five minutes . the cells were analysed using flow cytometry ( bd facscanto ii , bd biosciences , san jose , ca , usa ) , and the data were analysed with cellquest software ( becton dickinson , franklin lakes , nj , usa ) . h1299 and hcc827 cells were treated with ifn- ( 10 iu / ml ) or gefitinib ( 20 nm ) alone or in combination for 48 hours , washed with cooled pbs three times , and then lysed in 50100 l mammalian protein extraction reagent containing a protease inhibitor cocktail for 30 minutes . the lysates were normalised for total protein content using a bicinchoninic acid assay , and 60 g protein samples were loaded on sds - page gels and transferred to pvdf membranes . the membranes were probed with rabbit monoclonal antibodies against phosphorylated ( p)-egfr , egfr , p - stat3 ( signal transducers and activators of transcription 3 ) , stat3 , and -actin ( santa cruz , dallas , tx , usa ) at 4c overnight . after washing three times in tbs - t ( tris hydrochloride buffer containing 0.1% tween-20 ) , the membranes were incubated with the goat polyclonal secondary antibody to rabbit for two hours at room temperature . the membranes were washed with tbs - t three times for 10 minutes and exposed to x - ray film for 510 minutes . the relative expression of protein was determined from the optical density ratio of the corresponding protein bands using bandscan 5.0 software ( glyko inc . were calculated as previously described with minor modifications : cis = % ab / ( % a %b ) , where % a and % b are the effect of the individual drug and % ab is the effect of the combination . cis < 1 , = 1 , and > 1 indicates synergism , additive effect , and antagonism , respectively . all data are expressed as the mean standard error ( se ) of at least three experiments . a two - tailed , independent student 's t - test differences at p < 0.05 for the student t - test were considered statistically significant . interferon- ( alfaron , recombinant ifn-2b ; hualida biotechnology co. , tianjin , china ) had a specific activity of 3 10 units / ml and > 99% purity . the selective egfr - tki gefitinib ( gd760 , iressa ) , kindly provided by astrazeneca ( macclesfield , cheshire , uk ) , was dissolved in dmso to a working concentration of 20 mm . both drugs were stored at 20c in tightly sealed sterile tubes and diluted to the desired concentration in culture medium ( rpmi 1640 ; gibco , grand island , ny , usa ) before being added to the cells . four human nsclc cell lines were used : a549 , h1299 , and hcc827 ( cell bank of chinese academy of sciences , shanghai , china ) , which have different egfr and k - ras gene statuses ( exons 1821 of the egfr gene and exons 23 of the k - ras gene were sequenced , a549 cell line harbours mutation ( g12s ) of k - ras , hcc827 cell line carries egfr 19del , and the egfr and k - ras of h1299 cell line are all wild type , data not shown ) . the cell lines were grown in rpmi 1640 ( gibco ) supplemented with 10% foetal bovine serum and maintained at 37c in a humidified atmosphere containing 5% co2 and 95% air . four different experimental regimens were investigated to evaluate the impact of ifn- pretreatment on the antiproliferative effect of gefitinib : i , 72-hour incubation with gefitinib or ifn- alone;ii , 72-hour incubation with gefitinib ( at the indicated concentrations in figure 2 ) + 10 iu / ml ifn- ( at the concentration of 10 iu / ml , ifn- inhibits the proliferation of these lung cancer cells obviously);iii , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with phosphate - buffered saline ( pbs ) , followed by 72-hour incubation with gefitinib + 10 iu / ml ifn-;iv , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with pbs , followed by 72-hour incubation with gefitinib . i , 72-hour incubation with gefitinib or ifn- alone ; ii , 72-hour incubation with gefitinib ( at the indicated concentrations in figure 2 ) + 10 iu / ml ifn- ( at the concentration of 10 iu / ml , ifn- inhibits the proliferation of these lung cancer cells obviously ) ; iii , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with phosphate - buffered saline ( pbs ) , followed by 72-hour incubation with gefitinib + 10 iu / ml ifn- ; iv , 72-hour pretreatment with 10 iu / ml ifn- , following which the ifn- was removed by washing twice with pbs , followed by 72-hour incubation with gefitinib . control cells were processed in the same manner but with drug - free medium ( placebo ) instead of gefitinib . 100 l exponential growth phase cells ( about 3500 cells in total ) were seeded into 96-well plastic plates , incubated overnight , and added 100 l serial dilutions of gefitinib , ifn- , or gefitinib + ifn- into each well . after 72-hour incubation at 37c , 20 l 5 mg / ml mtt solution was added into each well and the cells were incubated for four hours . the media were removed and 150 l dmso was added into each well to dissolve the mtt completely , and the absorbance values were read at 570 nm using a microplate reader ( model 680 ; bio - rad , hercules , ca , usa ) . at least three replicate wells were used for each drug concentration , and the experiment was carried out independently at least three times . cell proliferation is expressed as the percentage of drug - treated surviving cells versus control cells ( in which viability was considered 100% ) . cells were incubated for 72 hours in drug - free medium ( control ) , 10 iu / ml ifn- , gefitinib alone , or gefitinib after 10 iu / ml ifn- pretreatment . according to the ic50 of gefitinib in these three cell lines , the concentration of gefitinib for hcc827 cell was 20 nm , and for other two cell lines it was 10 m . the cells were harvested by trypsinisation , washed twice with pbs , incubated in 500 l binding buffer and 10 l annexin v fitc at room temperature for 30 minutes , and then 5 l pi was added and incubated for five minutes . the cells were analysed using flow cytometry ( bd facscanto ii , bd biosciences , san jose , ca , usa ) , and the data were analysed with cellquest software ( becton dickinson , franklin lakes , nj , usa ) . h1299 and hcc827 cells were treated with ifn- ( 10 iu / ml ) or gefitinib ( 20 nm ) alone or in combination for 48 hours , washed with cooled pbs three times , and then lysed in 50100 l mammalian protein extraction reagent containing a protease inhibitor cocktail for 30 minutes . the lysates were normalised for total protein content using a bicinchoninic acid assay , and 60 g protein samples were loaded on sds - page gels and transferred to pvdf membranes . the membranes were probed with rabbit monoclonal antibodies against phosphorylated ( p)-egfr , egfr , p - stat3 ( signal transducers and activators of transcription 3 ) , stat3 , and -actin ( santa cruz , dallas , tx , usa ) at 4c overnight . after washing three times in tbs - t ( tris hydrochloride buffer containing 0.1% tween-20 ) , the membranes were incubated with the goat polyclonal secondary antibody to rabbit for two hours at room temperature . the membranes were washed with tbs - t three times for 10 minutes and exposed to x - ray film for 510 minutes . the relative expression of protein was determined from the optical density ratio of the corresponding protein bands using bandscan 5.0 software ( glyko inc . , upper heyford , uk ) . the combination indexes ( cis ) were calculated as previously described with minor modifications : cis = % ab / ( % a %b ) , where % a and % b are the effect of the individual drug and % ab is the effect of the combination . cis < 1 , = 1 , and > 1 indicates synergism , additive effect , and antagonism , respectively . all data are expressed as the mean standard error ( se ) of at least three experiments . a two - tailed , independent student 's t - test was used to determine the differences between the pretreatment and control groups . differences at p < 0.05 for the student t - test were considered statistically significant . first , the effects of gefitinib or ifn- on the growth of the a549 , h1299 , and hcc827 cells were evaluated . dose - dependent growth inhibition was observed in all cell lines following 72-hour incubation with ifn- ( fig . 2 ) alone . moreover , the sensitivity to the anti - proliferative effect of ifn- did not appear to be associated with the egfr and k - ras gene statuses . it is known that gefitinib sensitivity is associated with egfr mutation status [ 1013 ] . cell survival of the hcc827 cell line , containing the exon 19 deletion , was inhibited by up to 70% by 3.125 nm gefitinib , a concentration that was cytotoxic ( fig . p < 0.05 additive / antagonistic antiproliferative effect of ifn-/gefitinib combination on human nsclc cell lines . b ) the cis defining the growth inhibitory effects of the combination treatment of gefitinib and ifn-. combinations were antagonistic when cis > 1.0 , and additive when cis = 0.91.0 . points , mean of three individual treatments . mt , mutation type next , the antiproliferative effect of ifn- alone in the cell lines was evaluated . cell survivals were similar among three cell lines ( a549 : 71.8% , h1299 : 67.9% , and hcc827 : 63.2% ) . we investigated whether a synergistic effect on nsclc cells could be obtained by combining gefitinib with ifn-. however , it was found that when gefitinib was administered with ifn- , it resulted in an additive interaction in all four cell lines , as shown by the ci values ( ci = 1 ; fig . the mean ci values of gefitinib + ifn- for each cell line are listed in table 1 ( gefitinib + ifn- ) ; all of the ci values were approximately 1 . combination indexes for gefitinib and ifn-/combination each value represents the mean se of all combination index ( cis ) values for each cell line . the p - values were analysed by a two - tailed student 's t - test . p < 0.05 , statistically significant ; ns , p > 0.05 , statistically not significant . ci value : < 0.9 synergism , 0.91.0 additivity , > 1 antagonism . nevertheless , when the cells were pretreated with 10 iu / ml ifn- before administering gefitinib and ifn- , there were antagonistic interactions between gefitinib and ifn- in three cell lines . 2 ) , as demonstrated by the higher mean ci values of the gefitinib + ifn- combination after ifn- pretreatment ( table 1 , cisgefitinib + ifn- vs. cisifn-gefitinib + ifn- , p < 0.005 ) . in these cell lines , the cell survival rate according to each concentration of gefitinib increased after the ifn- pretreatment ( table 1 , fig . 2 ) . to gain insight into whether ifn- could influence gefitinib sensitivity , cell survival was evaluated after gefitinib treatment following ifn- pretreatment . after the pretreatment , cell survival rates were significantly increased in the a549 and h1299 cell lines as the gefitinib concentration increased ; a significant reduction was observed at each gefitinib concentration in the hcc827 cell line ( fig . 3a , cell survival rate ifn-gefitinib vs. cell survival rate placebogefitinib , p < 0.05 ) . a ) the antagonistic effect of ifn- pretreatment on the antiproliferative effect of gefitinib on human nsclc cell lines was evaluated by mtt assay . b ) apoptosis in the cell lines was analysed by flow cytometry for annexin v/ pi staining . the results ( top and bottom right panels ) are expressed as the percentage of the apoptotic cells . data are reported as the mean se of three determinations , p < 0.05 for gefitinib vs. ifn-gefitinib to determine whether ifn- could inhibit gefitinib - induced apoptosis , the outcome of the annexin v / pi detection demonstrated that there was a decrease in gefitinib - induced apoptosis after the ifn- pretreatment ( fig . previously , many studies reported that ifn- could activate egfr signalling by upregulating egfr expression [ 7 , 8 ] . two cell lines were examined : h1299 and hcc827 , where ifn- pretreatment inhibited the effect of gefitinib significantly . ifn- could not activate egfr phosphorylation but could inhibit stat3 phosphorylation in both cell lines ( fig . 4 ) . gefitinib inhibited egfr activation in both cell lines and inhibited stat3 activation in both cell lines ( fig . a ) p - egfr / egfr and p - stat3/stat3 were analysed by western blot . the whole protein from h1299 and hcc827 cells treated for 48 hours with ifn- ( 10 iu / ml ) and gefitinib ( 10 nm ) alone with ifn- was extracted . b ) the expression levels were analysed by the ratio of optical density with -actin . stat3 activation was inhibited by ifn- and gefitinib in both cell lines , p < 0.05 first , the effects of gefitinib or ifn- on the growth of the a549 , h1299 , and hcc827 cells were evaluated . dose - dependent growth inhibition was observed in all cell lines following 72-hour incubation with ifn- ( fig . 2 ) alone . moreover , the sensitivity to the anti - proliferative effect of ifn- did not appear to be associated with the egfr and k - ras gene statuses . it is known that gefitinib sensitivity is associated with egfr mutation status [ 1013 ] . cell survival of the hcc827 cell line , containing the exon 19 deletion , was inhibited by up to 70% by 3.125 nm gefitinib , a concentration that was cytotoxic ( fig . p < 0.05 additive / antagonistic antiproliferative effect of ifn-/gefitinib combination on human nsclc cell lines . b ) the cis defining the growth inhibitory effects of the combination treatment of gefitinib and ifn-. combinations were antagonistic when cis > 1.0 , and additive when cis = 0.91.0 . points , mean of three individual treatments . mt , mutation type next , the antiproliferative effect of ifn- alone in the cell lines was evaluated . cell survivals were similar among three cell lines ( a549 : 71.8% , h1299 : 67.9% , and hcc827 : 63.2% ) . we investigated whether a synergistic effect on nsclc cells could be obtained by combining gefitinib with ifn-. however , it was found that when gefitinib was administered with ifn- , it resulted in an additive interaction in all four cell lines , as shown by the ci values ( ci = 1 ; fig . the mean ci values of gefitinib + ifn- for each cell line are listed in table 1 ( gefitinib + ifn- ) ; all of the ci values were approximately 1 . combination indexes for gefitinib and ifn-/combination each value represents the mean se of all combination index ( cis ) values for each cell line . the p - values were analysed by a two - tailed student 's t - test . p < 0.05 , statistically significant ; ns , p > 0.05 , statistically not significant . ci value : < 0.9 synergism , 0.91.0 additivity , > 1 antagonism . nevertheless , when the cells were pretreated with 10 iu / ml ifn- before administering gefitinib and ifn- , there were antagonistic interactions between gefitinib and ifn- in three cell lines . 2 ) , as demonstrated by the higher mean ci values of the gefitinib + ifn- combination after ifn- pretreatment ( table 1 , cisgefitinib + ifn- vs. cisifn-gefitinib + ifn- , p < 0.005 ) . in these cell lines , the cell survival rate according to each concentration of gefitinib increased after the ifn- pretreatment ( table 1 , fig . to gain insight into whether ifn- could influence gefitinib sensitivity , cell survival was evaluated after gefitinib treatment following ifn- pretreatment . after the pretreatment , cell survival rates were significantly increased in the a549 and h1299 cell lines as the gefitinib concentration increased ; a significant reduction was observed at each gefitinib concentration in the hcc827 cell line ( fig . 3a , cell survival rate ifn-gefitinib vs. cell survival rate placebogefitinib , p < 0.05 ) . a ) the antagonistic effect of ifn- pretreatment on the antiproliferative effect of gefitinib on human nsclc cell lines was evaluated by mtt assay . b ) apoptosis in the cell lines was analysed by flow cytometry for annexin v/ pi staining . the results ( top and bottom right panels ) are expressed as the percentage of the apoptotic cells . data are reported as the mean se of three determinations , p < 0.05 for gefitinib vs. ifn-gefitinib to determine whether ifn- could inhibit gefitinib - induced apoptosis , the outcome of the annexin v / pi detection demonstrated that there was a decrease in gefitinib - induced apoptosis after the ifn- pretreatment ( fig . previously , many studies reported that ifn- could activate egfr signalling by upregulating egfr expression [ 7 , 8 ] . two cell lines were examined : h1299 and hcc827 , where ifn- pretreatment inhibited the effect of gefitinib significantly . ifn- could not activate egfr phosphorylation but could inhibit stat3 phosphorylation in both cell lines ( fig . 4 ) . gefitinib inhibited egfr activation in both cell lines and inhibited stat3 activation in both cell lines ( fig . a ) p - egfr / egfr and p - stat3/stat3 were analysed by western blot . the whole protein from h1299 and hcc827 cells treated for 48 hours with ifn- ( 10 iu / ml ) and gefitinib ( 10 nm ) alone with ifn- b ) the expression levels were analysed by the ratio of optical density with -actin . stat3 activation was inhibited by ifn- and gefitinib in both cell lines , p < 0.05 the present findings are not consistent with those of previous studies , which showed that ifn- could enhance the antiproliferation of egfr - tki in colon cancer cells , renal cell carcinoma , and hepatocellular carcinoma . in our study , ifn- reduced the gefitinib sensitivity of nsclc cells that contained egfr - tki sensitive mutations . we also noted that ifn- alone had a mild antiproliferative effect on the nsclc cell lines : the rate of cell survival inhibition was less than 50% even at 10 iu / ml ifn- ( fig . 3b ) . as expected , the three cell lines , carrying different egfr and k - ras gene types , responded differently to gefitinib alone : a549 and h1299 cells were less susceptible to gefitinib while the hcc827 cell line was sensitive to its effects ( fig . disappointingly , there was no interaction between gefitinib and ifn- when they were administered in combination in these cell lines . the cis values demonstrated that the gefitinib + ifn- combination effects were additive ( table 1 , fig . notably , the antiproliferation of gefitinib alone was reduced after ifn- pretreatment in the three cell lines ( fig . previous studies have suggested that ifn- can upregulate egfr expression in cancer cells [ 14 , 15 ] , and others have postulated that the increases in egfr expression tend to be accompanied by enhanced activity of egfr signalling . nevertheless , based on our observations , egfr activation was not associated with ifn- in the h1299 or hcc827 cells ( fig . stat3 is a stat family member that is involved in oncogenesis and tumour progression ; previous studies have shown that stat3 is activated in lung cancer , and its activation may stem from egfr mutations . similarly , it was found that stat3 was activated in the two cell lines ( fig . , ifn- can activate the janus kinase 1 ( jak1)/stat pathway and inhibit stat3 phosphorylation ; our research also showed that ifn- inhibited p - stat3 in the nsclc cell lines . it is worth mentioning that prior studies have shown that there is cross - talk between jak / stat and egfr signalling in the regulation of hyperproliferation , and that egfr is an upstream activator of stat3 that can mediate egfr signalling [ 16 , 17 , 21 ] ; therefore , egfr - tkis may be able to inhibit stat3 activation . 4 shows that gefitinib could inhibit p - stat3 expression in the h1299 and hcc827 cells in which ifn- could inhibit gefitinib - induced apoptosis to some extent . therefore , we hypothesised that egfr signalling may be dependent on stat3 activation , in other words , the inhibitory effect of gefitinib may depend on stat3 activation , and this point of view has been reported . the phosphorylation of stat3 was inhibited by ifn- pretreatment ; therefore , the egfr pathway , being somewhat dependent on stat3 activation , may not have been activated , thereby reducing the gefitinib sensitivity of these cell lines . although ifn- may increase the susceptibility of egfr - tki in other tumour cells , but in lung cancer there is no similar action . in contrast , the antiproliferation of egfr - tki can be inhibited by ifn- in lung cancer . our data suggests that egfr - tki should avoid sequential application with ifn-. in addition , as we know , finding the mechanisms of egfr - tki resistance is the current focus , ifn- inactivates gefitinib in nsclc , it may provide a new idea to explore and enrich the mechanisms of egfr - tki resistance . we hypothesised that the potential cause may be that ifn- inhibits phosphorylation of stat3 , which might be dependent on egfr signal activation . 320.6750.12241 ) , natural science foundation of china ( no . 81102012 ) , natural science foundation of china ( no . 91229104 ) , and zhejiang medicine scientific research fund projects ( no . | aim of the studymany studies have shown that interferon- ( ifn- ) enhances the antiproliferative effect of gefitinib in some solid tumours .
we aimed to determine the effect of combining ifn- with gefitinib in human non - small cell lung cancer ( nsclc ) cell lines ( a549 , h1299 , hcc827 ) with different egfr and k - ras gene statuses.material and methodsan mtt assay was used to assess cell proliferation .
apoptosis was detected by an annexin v / propidium iodide assay using flow cytometry , and western blotting was used to determine the expression of epidermal growth factor receptor / phosphorylated epidermal growth factor receptor ( egfr / p - egfr ) and signal transducers and activators of transcription 3/phosphorylated signal transducers and activators of transcription 3 ( stat3/p - stat3).resultsthere was an additive interaction when gefitinib was combined with ifn- in all cell lines ; however , there was antagonism when gefitinib followed ifn- pretreatment in three cell lines .
notably , ifn- pretreatment significantly reduced the gefitinib sensitivity of hcc827 cells . surprisingly , while ifn- inhibited stat3 phosphorylation in cell lines , gefitinib could do so.conclusionsthe results might confirm the hypothesis that ifn- induces gefitinib sensitivity of nsclc , and ifn- inhibits phosphorylation of stat3 , which may be dependent on egfr signal activation playing a role in the reduction of gefitinib sensitivity after ifn- treatment in nsclc cell lines . |
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in the usa , where more than one quarter of all adults smoke cigarettes,1 tobacco smoking is considered the leading preventable cause of disease , disability and premature death.23 it has been estimated that direct and indirect economic losses attributed to tobacco use in usa approach 85 billion us dollars annually.2 the risk of premature death in smokers was about double that of non - smokers and was directly proportional to the frequency of smoking.24 recent data showed that 15% of all cancer cases in the world were estimated to be due to cigarette smoking.5 moreover , slightly less than 20% of all deaths in industrialized countries were attributable to tobacco.67 tobacco contains about 4000 substances , more than 40 of which are carcinogenic and many others that are hazardous to cardiopulmonary systems.2 many studies have demonstrated an association between cigarette smoking and cancers of the oral cavity , pharynx , larynx , lungs , esophagus , stomach , colon , rectum , pancreas , kidney and urinary bladder,35,811 and a less strong association with leukemia and cervical cancer.8 a causal association was found to exist between smoking and coronary artery disease , peptic ulcer , chronic obstructive pulmonary disease and stroke.3101213 cigarette smoking is not only harmful to those using it deliberately , but also to involuntary smokers ( the so - called passive smokers ) . passive smoking is associated with coronary artery disease , chronic obstructive pulmonary disease , asthma , leukemia and cancers of the lung , sinuses , cervix , breast and brain.121416 in children , passive smoking is associated with higher incidence of asthma attacks , pneumonia , ear disease , sudden infant death syndrome and upper and lower respiratory tract infections.121417 fetuses are not exempted from hazards of smoking . it was found that pregnant mothers exposed to passive smoking were more prone to have low birth weight infants.18 it is doubtful that an ex - smoker will fully resume his pre - smoking health state . however , a substantial risk reduction with great improvement in life expectancy and quality is beyond doubt2111923 the benefit of quitting is documented even for old patients and those with an established tobacco - related disease.2325 more than half of americans are considered to have a sedentary life - style.3 this is considered the most preventable risk factor for coronary artery disease.32126 regular physical exercise decreases the risk for many health problems including non - insulin dependent diabetes mellitus , osteoporosis , osteoarthritis , cardiovascular disease , cancer , hypertension , obesity and hypercholestrolemia.32734 regular exer - cise was also found to be associated with better general well - being35 and life expectancy3637 in addition to lesser mortality rates .422 specific prescription of physical exercise should be given to almost all patients , bearing in mind the age and general condition of each individual .323 almost every patient including elderly people and those with severe coronary artery disease and patients with congestive heart failure , has the potential of benefiting from regular physical exercise.3242537 the physiologic effects of exercise depend on its type , intensity , duration and frequency.38 recreational physical exercise can be classified into the following two types:3839 this type is characterized by an increase in muscle tension without much change in fiber length , with substantial energy expenditure . examples are weight lifting , hand grip exercises and pushing or pulling against a resistance . anaerobic exercise increases total peripheral resistance and thus leads to considerable rise in blood pressure . this is the type of exercise which is recommended to the general population on a regular basis . the more frequent and prolonged the aerobic exercise , the better its effects , provided that the patient 's condition permits it.3340 this type is characterized by an increase in muscle tension without much change in fiber length , with substantial energy expenditure . examples are weight lifting , hand grip exercises and pushing or pulling against a resistance . anaerobic exercise increases total peripheral resistance and thus leads to considerable rise in blood pressure . this is the type of exercise which is recommended to the general population on a regular basis . the more frequent and prolonged the aerobic exercise , the better its effects , provided that the patient 's condition permits it.3340 the annual economic losses due to obesity are calculated in billions of dollars.41 the prevalence of obesity in uk ( 1986 ) approached 12% in males and 8% in females.42 the situation in usa is even worse , where about one quarter of all adult americans are overweight.4243 further- more , recent data indicated a linear increase in the average weight of american adults over the last few decades.41 obesity is defined by who as a body mass index ( bmi ) more than 30 kg per square metre.4244 the risk of overweight related problems , however , starts at body mass index of 25 kg per square metre.41 overweight adversely affects the general health of people.35 it is considered to be a risk factor for non insulin dependent diabetes mellitus , coronary artery disease , hypertension , hyperlipidemia , gallstones , low back pain , certain cancers ( colorectal and prostate in males , and cervical , breast and ovarian in females ) and premature death.4264348 distribution of regional , rather than total , body fat is increasingly gaining special importance . recent research has demonstrated an association between the amount of visceral fat and the disturbances in glucose tolerance and lipid concentrations.46 visceral fat , which can be measured by computerized tomography,46 was found to correlate highly significantly with waist - to - hip ratio.49 cut off points for waist - to - hip ratio , above which high risk begins , were generally agreed to be 0.95 for men and 0.80 for women.50 high waist - to - hip ratio ( upper body obesity ) is associated with high risk for hypertension , diabetes mellitus , hypercholestrolemia , hypertriglyceridemia , low hdl , hyperinsulinemia and mortality from cardiovascular disease.44465054 causes of obesity include a variety of medical , genetic and lifestyle factors . the easiest and most effective modality of prevention is to deal with the lifestyle factors , namely diet and physical activity.4255 prevention of obesity has been proved to be highly beneficial in preventing obesity - related health hazards , resulting in a better life expectancy and general health.112023325658 seat belts were introduced for the first time in the mid 1950s . by the late 1960s the introduction of interlock devices that prevent ignition if the driver 's safety belt was not fastened , resulted in a significant improvement in the use of seat belts . minimal improvement occurred with the introduction of warning lights and buzzers . in the usa ( 1984 ) , legislation for the use of seat belts took place and resulted in increased compliance from 15% to about 50% in a short time.59 studies from all over the world documented the benefits of seat belts and the hazards of non - use.1127356065 there was more than 60% reduction in the severity of injuries , hospital admissions and hospital charges for users of seat belts.66 in germany , morbidity and mortality due to road traffic accidents have shown substantial decrease when the rates of seat belts use increased.67 interestingly it was found that parents use of seat belts , influenced children to do the same68 and that non - use of seat belts was associated with other unhealthy behavior such as smoking , overweight and lack of exercise.6269 the airbag was recently introduced in many new motor vehicles as a supplement to seat belts . this new device has proved quite effective but its effect is still limited due to the relatively small number of cars equipped with them.70 there is a belief that during the last few decades some unhealthy lifestyles have been adopted by saudis . however , published national studies that quantify these important assumptions are few.71 most of the studies referred to below are apparently limited by being either retrospective , based on insufficient recording systems , clinic - based , hospital - based , or done on a small scale . these limitations do not rule out the importance of such studies in providing us with rough estimates of the extent of the problem in our country . available data point to cigarette smoking as an important health problem in saudi arabia.72 following the steady increase in the quantity of tobacco imported before 1987 , national statistical data7374 showed significant reduction in tobacco imports during the period from 1989 to 1994 ( table 1 ) . tobacco , however , is still considered a burden on the health and economy of saudi population . it was found that 33% of male medical students in riyadh smoke cigarettes.75 the prevalence among other university students was even higher ( 37%).76 in secondary schools , 22% of male students admitted being regular smokers.77 while smoking is considered socially unacceptable among females , up to 12% of university female students reported being current smokers.78 the weight ( in thousand tons ) and cost ( in million sr ) of tobacco imported during the period 1989 - 1994 in 1987 , a1-turki found that the smoking rate in the eastern province was 37% and 9% for adult males and females.79 in al - baha province , in 1989 , al - bedah reported a higher rate among males ; about 53% of adult males being smokers . the overall rate , however , was 29% because cigarette smoking was absolutely unacceptable among females in such a rural community.80 over the past two decades , the per capita calory consumption of saudis has shown a steady increase ( table 2).8182 considering bmi of 25 as a cut - off point , binhemd et a183 found 52% of males and 65% of females visiting the primary care clinic of king fahd hospital of the university at al - khobar , to be obese . despite the limitation of such a study , the high figures it showed were supported by findings of al - shammari et al.47 al - abbad ( 1995 ) demonstrated a prevalence of obesity ( defined as a bmi of 85th percentile ) approaching 29% among female students aged 11 - 21 years.84 the per capita daily dietary consumption ( kcal / day ) of saudis over the period 1974 - 1989 data about recreational aerobic physical activity of saudis are not yet available . however , if we consider the body weight as an indirect indicator , the physical activity of saudi population does not seem to be improving in relation to other healthy life styles . the number of cars imported by gulf countries increased by 1031% between 1971 and 1984 , and deaths due to road traffic accidents increased by 307% in the same period . in saudi arabia , the number of cars increased by 2400% within the 10 years ending 1986 and this was associated with 469% increase in trauma cases and a 561% increase in related deaths.85 it is not surprising , therefore , that road traffic accidents constitute the leading cause of death in the age group 16 - 36 years . in 1984 , mortality due to road traffic accidents approached 15 deaths per day . this number reflects only those who died at the scene of the accident . those who died en route to the hospital or following admission were almost double.86 in 1980 , khawashki estimated the economic loss in saudi arabia due to road traffic accidents as saudi rivals 4,776,836 per day.87 in asir region alone , which contains 2.5% of all cars in the country , about 7760 persons were involved in road traffic accidents in the period from 1989 - 1991 ; 62% of these were injured , and 12% died.88 fast driving , driving by children under 18 years of age and driving without a license were found to contribute enormously towards road traffic accidents.8890 the authorities have responded to this by taking different measures including the imposition of severe penalties to offenders . surprisingly , the mandatory use of seat belts which is probably easier to enforce , is still receiving little attention . in a study done by marwa et a1,91 none of the victims of road traffic accidents were found to have had the seat belts fastened . a few small studies attempted to measure the rate of seat belts use in certain localities in the country and ended up with the rates ranging from 7 - -19%.909293 unfortunately , to the best of the author 's knowledge , there have been no nation - wide studies on the rate of seat - belt use . indeed , there is a great need to enforce healthy behavior in the saudi community as means of promoting healthy lifestyle towards the eventual goal of positive health . | the negative effects on health by behavior such as cigarette smoking , lack of physical exercise , non - control of body weight and non - use of seat belts were empirically documented .
available findings of the various studies on lifestyle of the saudi arabian community were not encouraging .
if the general health status of the saudi population is to be improved , an enforcement of healthy lifestyles must be considered . |
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helicobacter pylori is a gram - negative bacteria with strong association with upper gastrointestinal diseases such as peptic ulcer diseases ( gastric ulcer , duodenal ulcer ) and malignancies ( gastric cancer , lymphoma ) , reported worldwide and in iran ( 1 - 3 ) . in developing countries , h. pylori is typically acquired in early childhood and may persist for a lifetime if left untreated ( 4 , 5 ) . in adults , the h. pylori infection rate increases with age ( 6 , 7 ) . oral - oral and fecal - oral routes are the most common modes of h. pylori transmission in developing countries . less common modes of transmission are zoonotic and iatrogenic ( such as endoscopic or through dental procedures ) transmissions ( 8 , 9 ) . in several previous studies , there were differences in the prevalence rate of h. pylori infection between countries ( 1 , 10 - 14 ) . it can be due to diverse contributing factors including socioeconomic status , geographical or living conditions , and ethnicity or location of each population ( 1 ) . dyspepsia is a medical condition characterized by chronic or recurrent pain in the upper abdomen , which is considered as one of the most common causes of patients referrals to gastroenterology centers ( 15 ) . the definition of dyspepsia has been elusive with variety of proposed descriptions ( 16 ) . dyspepsia is defined based on the recent asian consensus report as a symptom or set of symptoms that is ( are ) considered to originate from the gastroduodenal region . in this consensus report , the dyspepsia symptoms are epigastric pain , epigastric burning , postprandial fullness , early satiation , and others , including bloating in the upper abdomen , nausea , vomiting and belching . most of the consensus members agreed that symptom duration of equal or more than three months might be enough for diagnosis of dyspepsia . functional dyspepsia is defined as chronic dyspeptic symptoms in the absence of organic , systemic , or metabolic condition ( s ) , which is ( are ) likely to explain the symptoms ( 16 ) . in the western world , the reported prevalence of dyspepsia ranged 25 - 50% ( 17 , 18 ) . in the south - western united states , dyspepsia and h. pylori - related gastrointestinal diagnoses were remarkably common , with an annual incidence of 13 per 1000 individuals ( 15 ) . ( 12 ) demonstrated that the overall prevalence of patients with h. pylori infection and dyspepsia was 49.7% in kuwait . in a systematic review by amini et al . ( 19 ) , the prevalence of dyspepsia in iran varied between 2.2% to 29.9 % . understanding the latest prevalence of h. pylori infection in patients with dyspepsia and determining its associations with endoscopic findings and pathologic changes in gastric mucosa can be useful for health - related purposes in every region and country . some reports from asia and iran have demonstrated that the prevalence rate of h. pylori infection has declined rapidly in these regions ( 10 , 11 ) . this study was conducted to determine the prevalence rate of h. pylori infection among a population of adult patients with dyspepsia in shiraz , south of iran , using gram staining , rapid urease test , and culture . we also hoped to find the rate of h. pylori infection in different endoscopic findings . after obtaining the approval of the university ethical committee ( 91 - 01 - 36 - 4703 ) as well as written informed consent from the patients , all the patients with dyspepsia attending namazi hospital in shiraz ; as a major referral center for gastrointestinal diseases in south of iran , were recruited into the study between may and november 2012 . the exclusion criteria for all the patients were age less than 18 , history of h. pylori eradication , history of consumption of proton pump inhibitors ( ppis ) , h2-receptor antagonists , antacids or antibiotics within four weeks prior to the endoscopic evaluation , and history of gastric or esophageal surgery . we defined dyspepsia if the patient had one or more of these symptoms with duration of three months or longer : postprandial fullness , early satiation , epigastric pain , epigastric burning , bloating in the upper abdomen , nausea , vomiting , and belching ( 16 ) . upper gastrointestinal endoscopy was performed for all the patients with dyspepsia as an objective assessment for the diagnosis of causes was also performed . we categorized our patients based on the endoscopic finding into four groups : 1- normal , 2- abnormal nonulcerative ( any evidence of mucosal lesion without ulcer , e.g. erosion , erythema , nodularity , atrophy , white plaque , and petechiae ) , 3- ulcerative , and 4-combination of 2 and 3 . two antral biopsies were performed for all the included patients with normal and abnormal endoscopy results . after the endoscopy , the biopsy samples were transferred to the lab under an appropriate condition for pathology evaluation . all the samples were evaluated by rapid urease test and gram staining for the presence of h. pylori . if one of these test results was negative , the biopsy samples were gently cultured using standard methods ( 11 ) . we defined h. pylori as positive if two of these test results ( gram staining , rapid urease test , and culture ) were positive . chi square test was used and significance was defined as a p value < 0.05 . after obtaining the approval of the university ethical committee ( 91 - 01 - 36 - 4703 ) as well as written informed consent from the patients , all the patients with dyspepsia attending namazi hospital in shiraz ; as a major referral center for gastrointestinal diseases in south of iran , were recruited into the study between may and november 2012 . the exclusion criteria for all the patients were age less than 18 , history of h. pylori eradication , history of consumption of proton pump inhibitors ( ppis ) , h2-receptor antagonists , antacids or antibiotics within four weeks prior to the endoscopic evaluation , and history of gastric or esophageal surgery . we defined dyspepsia if the patient had one or more of these symptoms with duration of three months or longer : postprandial fullness , early satiation , epigastric pain , epigastric burning , bloating in the upper abdomen , nausea , vomiting , and belching ( 16 ) . upper gastrointestinal endoscopy was performed for all the patients with dyspepsia as an objective assessment for the diagnosis of causes was also performed . we categorized our patients based on the endoscopic finding into four groups : 1- normal , 2- abnormal nonulcerative ( any evidence of mucosal lesion without ulcer , e.g. erosion , erythema , nodularity , atrophy , white plaque , and petechiae ) , 3- ulcerative , and 4-combination of 2 and 3 . two antral biopsies were performed for all the included patients with normal and abnormal endoscopy results . after the endoscopy , the biopsy samples were transferred to the lab under an appropriate condition for pathology evaluation . all the samples were evaluated by rapid urease test and gram staining for the presence of h. pylori . if one of these test results was negative , the biopsy samples were gently cultured using standard methods ( 11 ) . we defined h. pylori as positive if two of these test results ( gram staining , rapid urease test , and culture ) were positive . chi square test was used and significance was defined as a p value < 0.05 . the endoscopic findings and the h. pylori test results of 631 patients with dyspepsia were evaluated retrospectively in this cross - sectional study . a total of 548 patients with a mean age of 40.38 15.39 years old were enrolled in the study and 83 patients were excluded . of the included patients , 238 ( 43.4% ) were male and 310 ( 56.6% ) were female . of the 548 patients tested , 170 ( 31% ) were positive and 378 ( 69% ) were negative for h. pylori infection . h. pylori infection was observed in 26.1% of male and 34.8% of female patients ( table 1 ) and its prevalence increased with age . the overall prevalence of h. pylori infection had ha sharp increase from the middle - aged patients ( 5.3% in 30 - 34-year - old and 5.8% in 35 - 44-year - old patients ) to 19.8% in those aged 45 or older ( table 1 ) . as the endoscopy results indicated , 83 ( 48.8% ) h. pylori - positive patients had normal or abnormal nonulcerative endoscopic findings ( groups 1 and 2 ) and 87 ( 51.2% ) had ulcer with or without concurrent abnormality ( groups 3 and 4 ) . h. pylori infection is a major cause of gastroduodenal diseases ; however , there have been different reports about the prevalence rate of this infection between countries ( 1 , 10 , 11 ) . several studies from asia and middle east including china , south korea , iran , and kuwait have shown that the prevalence of h. pylori has declined in these areas in the recent years ( 10 - 13 ) . in the present study , h. pylori positivity was 31% ( table 1 ) , which was surprisingly lower than other local reports ( 2 , 3 , 11 ) . in previous reports from shiraz it was shown that more than 85% of iranian adults carried igg against h. pylori ( 2 ) . in addition , histological evidence of h. pylori infection was recorded in 89.2% of biopsies in another report from north of iran ( 3 ) . recently , in a survey conducted by farshad et al . ( 11 ) in iran , the prevalence of h. pylori in a total of 522 patients who underwent endoscopic evaluation was 47.9% . interestingly , the results of this study showed a lower prevalence of h. pylori in comparison with other reports from asia and middle east . goh ( 14 ) reported that the overall prevalence rate of h. pylori in a malaysian population with dyspepsia was 49.0% . in this study , two antral biopsies were performed for diagnosis of h. pylori , using a rapid urease test . in alazmi ( 12 ) research in kuwait , an overall prevalence rate of 49.7% was reported in 362 patients with dyspepsia . in another report from india , poddar and yachha ( 6 ) demonstrated that almost 80% of the population in india was infected with h. pylori . ( 20 ) also showed that the overall prevalence of h. pylori in india was 80% . in this study , the diagnosis of h. pylori was established by polymerase chain reaction ( pcr ) amplification of the 16s rrna gene of h. pylori . as expected , similar to several other studies ( 10 - 13 ) our research showed that the prevalence of h. pylori was declined . according to the present study , the most common endoscopic abnormality in h. pylori - positive patients was ulcerative lesion , comprising groups 3 and 4 ( 51.2% ) ( table 2 ) . similar results were reported in 1990 by soll ( 21 ) . regarding the sex , this study showed that h. pylori positivity had predominance in females ; however , no significant association between sex and infection rate was shown . a vast variety of results have been reported regarding the predominance of either males or females ; but , it is generally concluded that there is no significant association between the infection rate of h. pylori and sex ( 1 , 12 , 20 ) . there was a significant association between the h. pylori infection prevalence and age in our study , which was similar to some other researches ( 1 , 12 , 20 ) . the findings of this cross - sectional study on dyspepsia showed a lower prevalence of h. pylori infection compared to other studies . abnormal nonulcerative and ulcerative changes constituted the dominant findings in h. pylori - positive patients . it seems that this lower prevalence of h. pylori may be due to an improvement in the socioeconomic status of the iranian population . | background : the prevalence rate of gastric infections caused by helicobacter pylori is different in between as well as within various countries , including iran.objectives:this study was aimed to evaluate the prevalence of h. pylori in iranian adult patients with dyspepsia.patients and methods : a total of 548 patients with dyspepsia referred to namazi hospital , a referral center for gastrointestinal diseases in shiraz , south of iran , were evaluated in a cross - sectional study .
the diagnosis of dyspepsia was based on clinical findings .
upper gastrointestinal endoscopy was performed for all the patients and two antral biopsies were taken from all the included ones .
the patients were categorized based on the endoscopic findings , to four groups : normal , abnormal nonulcerative , ulcerative , andcombination of 2 and 3.two positive test results ( gram staining , rapid urease test , and culture ) was considered as a positive - definite h. pylori infection for each patient.results:from the 548 included patients ( 238 males and 310 females ) , h. pylori was detected in 170 ( 31.2% ) .
the mean age was 40.38 15.39 years old .
h. pylori infection was detected in 26.1% of male and 34.8% of female patients and its prevalence increased with age .
eighty three ( 48.8% ) patients were positive for h. pylori infection , of which , 12.4% had normal and 36.4% had abnormal nonulcerative endoscopic findings ; 17.1% of patients were ulcerative and 34.1% had ulcer with or without concurrent abnormality.conclusions:findings from this study showed a lower prevalence of h. pylori infection than other studies and its prevalence increased with age .
there was no association between sex and infection .
the most common endoscopic abnormality in h. pylori - positive patients was ulcerative lesion . |
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the bioactivity of mta led to the interest at the development of new dental materials based on calcium silicates , including root canal sealers
. mta fillapex ( angelus industry dental products s / a , londrina , pr , brazil ) composition consists of mta associated with resins and pigments . mta plus ( avalon biomed inc . , sarasota , fl , usa ) is composed of calcium silicates in smaller particles than mta , according to the manufacturer . moreover , mta plus presents a water - soluble salt - free gel as a vehicle . this gel might improve the consistency , solubility and adhesiveness to dentin walls
, allowing its use as a sealer . studies evaluating mta fillapex biocompatibility and bioactivity in osteoblastic cells showed cytotoxicity effect of this sealer in the first periods of cells exposure
. however , for extended periods , cell viability and alp enzyme activity have substantially increased . the production of mineralized nodules by alizarin red staining was observed as well , demonstrating the cement biocompatibility and bioactivity after the setting time
. in other study , mta fillapex presented lower adhesion to dentin walls when compared to mta plus
. furthermore , the authors suggest that mta plus has a higher biomineralization potential than mta fillapex , being able to form apatite deposits in collagen fibrils inside the dentinal tubules , being similar to a tag between dentin - mta plus interface . when evaluating mta plus bioactive potential , a recent study showed that this cement is similar to proroot mta ( dentsply tulsa dental specialties , york , pa , usa ) , and releases calcium ions when in contact with water or hbss ( hank s saline based solution )
. although these sealers were launched in the market the last couple of years , literature still lacks information about their biological properties . the aim of this study was to evaluate the biocompatibility and bioactivity of mta plus and mta fillapex in primary culture of human dental pulp cells ( hdpcs ) . this cell line is also used to estimate the effects of mta and several calcium silicate - based cements in cell culture , demonstrating its cytocompatibility and bioactivity in vitro
. after approval from the institutional research ethics committee , patients with third molars at the stage of incomplete root formation , indicated to extraction for orthodontic reasons , were selected for this study . dental pulps were collected via apical opening using a hedstren file ( maillefer , dentsply , chemin du verger , ballaigues , switzerland ) and stored in a culture dish ( 35x10 mm ) containing alpha modification minimum essential eagle s medium ( -mem , sigma - aldrich , st . louis , mo , usa ) supplemented with 10% fetal bovine serum ( fbs , gibco , life technologies , grand island , ny , usa ) and 1% penicillin and streptomycin ( penstrep , gibco ) . the pulp tissues were sectioned into fragments of approximately 1 mm
and kept in supplemented -mem at 37c , 95% humidity and 5% co2 . the culture medium was changed 24 h after the pulp collection and every 48 h to obtain explants of human dental pulp cells ( hdpcs ) . the experiments were performed with cells from the fourth passage , and were repeated three times independently ( n=3/group ) . fillcanal ( fc ) was prepared according to grossman technique
, and mta plus ( mtap ) was prepared with the vehicle for root canal filling consistency . thereafter , 1 ml of each sealer was individually placed in wells of a 12-well plate ( tpp , techno plastic products , zollstrasse , trasadingen , sua ) and spread to fill the bottom completely . the plates with the samples were kept in an incubator at 37c , 95% humidity and 5% co2 during 24 h for the initial setting time of the materials . after this period , the specimens were exposed to ultraviolet light ( uv ) in a laminar flow for 30 min each side to disinfection
. the wells were filled with 5 ml of -mem supplemented with 1% penstrep and incubated at 37c , 95% humidity and 5% co2 for additional 24 h , in order to obtain the sealer extracts . figure 1endodontic sealers evaluated
cytotoxicity was assessed by mtt ( 3-(4,5-dimethyl - thiazoyl)-2,5-diphenyl - tetrazolium bromide ) and neutral red ( nr ) assays . hdpcs were plated at the concentration of 2x10 cells per well in 96-well plates ( tpp ) containing supplemented -mem culture medium and cultured for 24 h at 37c , 95% humidity and 5% co2 to obtain adherent cells prior to the material exposure . hdpcs were exposed to the sealers extracts in dilutions using culture medium without fbs of 1:2 , 1:3 and 1:4 for one day , according to iso 10993 - 5
. mtt is based on the conversion of tetrazolium salt to insoluble violet formazan crystals , determining the cytotoxicity by measuring cell viability rate . for this experiment , sealers extracts were replaced by 100 l of a 5 mg / ml mtt solution ( sigma - aldrich ) , followed by incubation at 37c , 95% humidity and 5% co2 for 3 h. thereafter , the content of the well was removed and the colorimetric product solubilized in 100 l of acidified isopropanol 0.04 n ( sigma - aldrich ) . the optical densities of the solutions were measured in a spectrophotometer ( elx800 ; instrumentos bio - tek , winooski , vt , usa ) at 570 nm wavelength
. the absorbance readouts were normalized with the absorbance of the group of cells exposed to -mem ( ct ) , and represented the activity of the viable cells . nr is a vital dye that is incorporated through endocytosis and accumulated on the lysosome of cells that have their membranes intact . for this experiment , sealers extracts were replaced by 0.1 ml -mem medium containing 50 g nr / ml ( sigma - aldrich ) followed by incubation at 37c , 95% humidity and 5% co2 for 3 h. then , the content of the well was removed and the colorimetric product solubilized in 100 l of an ethanol solution ( 50% ethanol and 1% acetic acid ) ( sigma - aldrich ) . the optical densities of the solutions were measured in a spectrophotometer ( elx800 ) at 570 nm . alp is an enzyme present in the plasma membrane that is associated with the mineralization process , and it is one of the major markers of osteoblast activity in the cells
. the bioactivity assay by alp enzyme activity was performed with hdpcs at the concentration of 2x10
cells per well in 12-well plates ( tpp ) , as previously described . alp activity was evaluated in accordance to the release of tymolphtaleine from tymolphtaleine monophosphate using alkaline phosphatase kit ( labtest diagnstica , lagoa santa , mg , brazil ) . hdpcs were lysed for 30 min at ambient temperature using sodium lauryl sulfate solution 1:1 ( sds , sigma - aldrich ) , and the assay was performed according to the manufacturer instructions . the data were expressed as alp activity ( mol tymophtaleine / min / l )
. therefore , we performed anova and tukey tests post hoc , considering a level of significance of 5% ( graphpad prism 5 software , graphpad software inc . , san diego , ca , usa ) . after approval from the institutional research ethics committee , patients with third molars at the stage of incomplete root formation , indicated to extraction for orthodontic reasons , were selected for this study . dental pulps were collected via apical opening using a hedstren file ( maillefer , dentsply , chemin du verger , ballaigues , switzerland ) and stored in a culture dish ( 35x10 mm ) containing alpha modification minimum essential eagle s medium ( -mem , sigma - aldrich , st . louis , mo , usa ) supplemented with 10% fetal bovine serum ( fbs , gibco , life technologies , grand island , ny , usa ) and 1% penicillin and streptomycin ( penstrep , gibco ) . the pulp tissues were sectioned into fragments of approximately 1 mm
and kept in supplemented -mem at 37c , 95% humidity and 5% co2 . the culture medium was changed 24 h after the pulp collection and every 48 h to obtain explants of human dental pulp cells ( hdpcs ) . the experiments were performed with cells from the fourth passage , and were repeated three times independently ( n=3/group ) . fillcanal ( fc ) was prepared according to grossman technique
, and mta plus ( mtap ) was prepared with the vehicle for root canal filling consistency . thereafter , 1 ml of each sealer was individually placed in wells of a 12-well plate ( tpp , techno plastic products , zollstrasse , trasadingen , sua ) and spread to fill the bottom completely . the plates with the samples were kept in an incubator at 37c , 95% humidity and 5% co2 during 24 h for the initial setting time of the materials . after this period , the specimens were exposed to ultraviolet light ( uv ) in a laminar flow for 30 min each side to disinfection
. the wells were filled with 5 ml of -mem supplemented with 1% penstrep and incubated at 37c , 95% humidity and 5% co2 for additional 24 h , in order to obtain the sealer extracts . cytotoxicity was assessed by mtt ( 3-(4,5-dimethyl - thiazoyl)-2,5-diphenyl - tetrazolium bromide ) and neutral red ( nr ) assays . hdpcs were plated at the concentration of 2x10 cells per well in 96-well plates ( tpp ) containing supplemented -mem culture medium and cultured for 24 h at 37c , 95% humidity and 5% co2 to obtain adherent cells prior to the material exposure . hdpcs were exposed to the sealers extracts in dilutions using culture medium without fbs of 1:2 , 1:3 and 1:4 for one day , according to iso 10993 - 5
. mtt is based on the conversion of tetrazolium salt to insoluble violet formazan crystals , determining the cytotoxicity by measuring cell viability rate . for this experiment , sealers extracts were replaced by 100 l of a 5 mg / ml mtt solution ( sigma - aldrich ) , followed by incubation at 37c , 95% humidity and 5% co2 for 3 h. thereafter , the content of the well was removed and the colorimetric product solubilized in 100 l of acidified isopropanol 0.04 n ( sigma - aldrich ) . the optical densities of the solutions were measured in a spectrophotometer ( elx800 ; instrumentos bio - tek , winooski , vt , usa ) at 570 nm wavelength
. the absorbance readouts were normalized with the absorbance of the group of cells exposed to -mem ( ct ) , and represented the activity of the viable cells . nr is a vital dye that is incorporated through endocytosis and accumulated on the lysosome of cells that have their membranes intact . for this experiment , sealers extracts were replaced by 0.1 ml -mem medium containing 50 g nr / ml ( sigma - aldrich ) followed by incubation at 37c , 95% humidity and 5% co2 for 3 h. then , the content of the well was removed and the colorimetric product solubilized in 100 l of an ethanol solution ( 50% ethanol and 1% acetic acid ) ( sigma - aldrich ) . the optical densities of the solutions were measured in a spectrophotometer ( elx800 ) at 570 nm . alp is an enzyme present in the plasma membrane that is associated with the mineralization process , and it is one of the major markers of osteoblast activity in the cells
. the bioactivity assay by alp enzyme activity was performed with hdpcs at the concentration of 2x10
cells per well in 12-well plates ( tpp ) , as previously described . alp activity was evaluated in accordance to the release of tymolphtaleine from tymolphtaleine monophosphate using alkaline phosphatase kit ( labtest diagnstica , lagoa santa , mg , brazil ) . hdpcs were lysed for 30 min at ambient temperature using sodium lauryl sulfate solution 1:1 ( sds , sigma - aldrich ) , and the assay was performed according to the manufacturer instructions . the data were expressed as alp activity ( mol tymophtaleine / min / l )
. therefore , we performed anova and tukey tests post hoc , considering a level of significance of 5% ( graphpad prism 5 software , graphpad software inc . , san diego , ca , usa ) . the mtt assay ( figure 2 ) showed 80 - 120% of viability to hdpcs exposed to mtap at all concentrations evaluated , being similar to the negative control of cytotoxicity ( p>0.05 ) . the mtap group viability rate was similar in the dilution of 1:4 to the other groups ( p>0.05 ) , although mtaf and fc groups viability rate showed significant difference in comparison to the ct . ( figure 3 ) , hdpcs exposed to mtap showed significant higher viability rate ( 130 - 135% ) than ct for all dilutions evaluated ( p<0.05 ) . mtaf showed the lowest viability of all groups in 1:2 dilution ( p<0.05 ) , however it increased in the dilutions of 1:3 and 1:4 ( p<0.05 ) . fc demonstrated good viability in all dilutions , being similar to mtaf in 1:4 ( p>0.05 ) . figure 2cell viability rate ( % ) according to mtt assay in human dental pulp cells ( hdpcs ) exposed to mta plus ( mtap ) , mta fillapex ( mtaf ) , fillcanal ( fc ) and culture medium used as control ( ct ) . bars with different letters represent significant differences between groups in each concentration of the material extracts ( p<0.05 )
figure 3cell viability rate ( % ) according to nr assay in human dental pulp cells ( hdpcs ) exposed to mta plus ( mtap ) , mta fillapex ( mtaf ) , fillcanal ( fc ) and culture medium used as control ( ct ) . bars with different letters represent significant differences between groups in each concentration of the material extracts ( p<0.05 )
in alp bioactivity assay ( figure 4 ) , mtap showed similar activity to the control group at both periods of time evaluated ( p>0.05 ) , while the other groups demonstrated significant lower alp activity when compared to ct ( p<0.05 ) . there was a tendency of alp activity increase in a time - dependent manner at all groups . figure 4alkaline phosphatase ( alp ) enzyme activity in human dental pulp cells ( hdpcs ) exposed to mta plus ( mtap ) , mta fillapex ( mtaf ) , fillcanal ( fc ) and culture medium used as control ( ct ) . bars with different letters represent significant differences between groups in each period of time ( p<0.05 )
hdpcs may represent a different population of differentiated and undifferentiated cells with great potential for pulp tissue and dentin regeneration . they can be successfully applied in studies to evaluate new biomaterials
, including endodontic sealers . studies in the literature reported that mta is capable to stimulate odontogenic differentiation in dental pulp mesenchymal cells
. in this study , human dental pulp was collected from third molars referred to extractions for orthodontic reasons , and obtained explants of hdpcs to investigate possible cytotoxic effects and the bioactivity of mtap , mtaf and fc . furthermore , mtt and nr assays demonstrated higher viability rate to mtap at all concentrations evaluated , and there was no evidence of cytotoxic effect . all materials presented similar viability in the higher dilution ( 1:4 ) on mtt assay . concentration - dependent results were also described by other authors when evaluating the viability of mta proroot and mtap in rat odontoblastic cells ( mdpc-23 ) , concluding that the cytotoxic risks become negligible after the dilution of the components
. the viability of mtaf has been evaluated in several cell types , such as fibroblasts
, osteoblasts
and human dental cell cultures
, demonstrating cytotoxic results in less than 7-day periods
. in this study , the cytotoxicity of mtaf may be considered moderate to low ( 50 - 80% , approximately ) and concentration - dependent , after 24 h setting - time . zinc oxide and eugenol cements as fc are considered cytotoxic both in vitro and in vivo , causing tissue inflammation and even bone resorption
. in addition , the literature demonstrated that these cements are capable of inducing the expression of cyclooxygenase-2 and nuclear factor kappa b , an immune mediator involved in cellular responses to stress , cytokines , free radicals , uv radiation , oxidation of ldl and bacterial and viral antigens
. mtaf cytotoxicity can be associated with its toxic components , such as salicylate resin , which can induce apoptosis
. besides the bioactivity , alp expression indicates the potential of the cells for differentiation
. therefore , the evaluation of this enzyme allows the determination of the material bioactivity and its potential to promote repair with formation of mineralized tissue . the results showed an increase in alp enzyme activity over time for cells exposed to all materials . mtap presented results similar to the control group in both evaluated periods , and higher than the other cements groups . other studies showed that after the initial period of cytotoxicity , mtaf sealer is able to promote bioactivity , stimulating the deposition of mineralized nodules and increasing alp activity
, corroborating the results of the present study . hdpcs may represent a different population of differentiated and undifferentiated cells with great potential for pulp tissue and dentin regeneration . they can be successfully applied in studies to evaluate new biomaterials
, including endodontic sealers . studies in the literature reported that mta is capable to stimulate odontogenic differentiation in dental pulp mesenchymal cells
. in this study , human dental pulp was collected from third molars referred to extractions for orthodontic reasons , and obtained explants of hdpcs to investigate possible cytotoxic effects and the bioactivity of mtap , mtaf and fc . furthermore , mtt and nr assays demonstrated higher viability rate to mtap at all concentrations evaluated , and there was no evidence of cytotoxic effect . all materials presented similar viability in the higher dilution ( 1:4 ) on mtt assay . concentration - dependent results were also described by other authors when evaluating the viability of mta proroot and mtap in rat odontoblastic cells ( mdpc-23 ) , concluding that the cytotoxic risks become negligible after the dilution of the components
. the viability of mtaf has been evaluated in several cell types , such as fibroblasts
, osteoblasts
and human dental cell cultures
, demonstrating cytotoxic results in less than 7-day periods
. in this study , the cytotoxicity of mtaf may be considered moderate to low ( 50 - 80% , approximately ) and concentration - dependent , after 24 h setting - time . zinc oxide and eugenol cements as fc are considered cytotoxic both in vitro and in vivo , causing tissue inflammation and even bone resorption
. in addition , the literature demonstrated that these cements are capable of inducing the expression of cyclooxygenase-2 and nuclear factor kappa b , an immune mediator involved in cellular responses to stress , cytokines , free radicals , uv radiation , oxidation of ldl and bacterial and viral antigens
. mtaf cytotoxicity can be associated with its toxic components , such as salicylate resin , which can induce apoptosis
. besides the bioactivity , alp expression indicates the potential of the cells for differentiation
. therefore , the evaluation of this enzyme allows the determination of the material bioactivity and its potential to promote repair with formation of mineralized tissue . the results showed an increase in alp enzyme activity over time for cells exposed to all materials . mtap presented results similar to the control group in both evaluated periods , and higher than the other cements groups . other studies showed that after the initial period of cytotoxicity , mtaf sealer is able to promote bioactivity , stimulating the deposition of mineralized nodules and increasing alp activity
, corroborating the results of the present study . this study shows that mtap presents biocompatibility and bioactivity in the primary culture of cells from human dental pulp . | mineral trioxide aggregate ( mta ) is a calcium silicate - based material .
new sealers have been developed based on calcium silicate as mta fillapex and mta plus .
objective the aim of this study was to evaluate biocompatibility and bioactivity of these two calcium silicate - based sealers in culture of human dental pulp cells ( hdpcs).material and methods the cells were isolated from third molars extracted from a 16-year - old patient .
pulp tissue was sectioned into fragments with approximately 1 mm3 and kept in supplemented medium to obtain hdpcs adherent cultures .
cell characterization assays were performed to prove the osteogenic potential .
the evaluated materials were : mta plus ( mtap ) ; mta fillapex ( mtaf ) and fillcanal ( fc ) .
biocompatibility was evaluated with mtt and neutral red ( nr ) assays , after hdpcs exposure for 24 h to different dilutions of each sealer extract ( 1:2 , 1:3 and 1:4 ) .
unexposed cells were the positive control ( ct ) .
bioactivity was assessed by alkaline phosphatase ( alp ) enzymatic assay in cells exposed for one and three days to sealer extracts ( 1:4 dilution ) .
all data were analyzed by anova and tukey post - test
( p0.05%).results mtt and nr results showed suitable cell viability rates for mtap at all dilutions ( 90 - 135% ) .
cells exposed to mtaf and fc ( 1:2 and 1:4 dilutions ) showed significant low viability rate when compared to ct in mtt .
the nr results demonstrated cell viability for all materials tested . in mtap group , the cells alp activity was similar to ct in one and three days of exposure to the material .
mtaf and fc groups demonstrated a decrease in alp activity when compared to ct at both periods of cell exposure.conclusions the hdpcs were suitable for the evaluation of new endodontic materials in vitro .
mtap may be considered a promising material for endodontic treatments . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
allergic rhinitis affects up to 30% of all children . some medications currently being used to treat this disorder , which significantly impacts learning , school performance , and attendance [ 13 ] , may adversely affect the central nervous system and contribute to learning impairment . in many cases , children with allergic rhinitis are also diagnosed with , and treated for , attention deficit / hyperactivity disorder ( adhd ) . while the etiology of adhd is unknown , it is most likely the result of interplay between various neuroanatomical / neurochemical systems and genetic and psychological factors . an association between adhd and allergy has long been touted but remains controversial [ 710 ] . it has also been suggested that children with adhd and those with allergic disease may share a common biological background [ 7 , 11 , 12 ] . previously , we suggested that neurotrophic protein nerve growth factor ( ngf ) might serve as a vehicle for cross talk between the immune and nervous systems . we found that ngf ( 1 ) regulates immunoglobulin ( ig ) production , particularly ige regulation , and ( 2 ) plays a major role in the control of allergic inflammatory response , serving as one of the critical proteins in the cross talk between the nervous and immune systems and to the endocrine system [ 1419 ] . we also demonstrated ( 1 ) the presence of functional ngf receptors on human b - lymphocytes and ( 2 ) that gp140trk - cells were the main active tyrosine kinase involved in ngf signaling . keeping in mind that adhd is a complex , multifaceted illness and that it is critical that we understand the role allergic response plays in the etiology of the subset of children with both diseases , we devised our study to ( 1 ) promote an understanding of the synergistic effects of treatment with both antihistamines and stimulants , ( 2 ) provide insight into the pathophysiology of this illness , and ( 3 ) determine the role ngf may play in children with both adhd and allergic rhinitis . ours was a randomized , 3-treatment group , double - blind , placebo - controlled , crossover study of 38 children with allergic rhinitis and adhd . approved by the local irb , it was conducted in colorado over a 4-year period during the tree and grass allergy seasons . we chose this period of time in order to capture results during the allergy season over an extended period of time . subject inclusion criteria included boys or girls aged 8 to 18 years with a diagnosis and history of seasonal allergic rhinitis to a prevalent allergen ( documented by a recognized skin test , skin prick , or intradermal [ i d ] ) , which required pharmacologic treatment at the time of enrollment and for the previous 2 years ( prick wheal > 3 mm over negative control ; intradermal wheal > 5 mm over negative control ) and a diagnosis of adhd and treatment regimen with stimulant medication . after obtaining written informed consent from a parent , legal guardian , or subjects of legal age , participants were randomized and screened using the multidimensional assessment of anxiety in children ( masc ) ; the children 's depression inventory ( cdi ) ; and the child symptom inventory ( csi ) [ 5 , 22 ] . qualified subjects were then enrolled into 1 of 2 order conditions and 1 of 3 treatment arms : cetirizine , 10 mg po qam for 2 weeks ( n = 12 ) ; methylphenidate , 20 mg if < 65 kg and 40 mg if 65 kg po qam for 2 weeks ( n = 12 ) ; or cetirizine 10 mg po qam + methylphenidate , 20 mg if < 65 kg and 40 mg if 65 kg po qam for 2 weeks ( n = 12 ) . all subjects crossed over to placebo for a period of 2 weeks during each treatment period . assessment included allergic rhinitis endpoint measures ( measuring symptoms of rhinoconjunctivitis such as sneezing , runny nose , itchy eyes , watery eyes , and stuffy nose on a scale of 04 ) ; the total symptom severity complex ( tssc ) ; and the rhinoconjunctivitis quality of life questionnaire ( rqlq ) [ 23 , 24 ] for allergy and the child behavior checklist ( cbcl ) , conners ' rating scale - revised ( crs - r ) , conners ' continuous performance task ( cpt ) , and the california verbal learning test for adhd symptoms . ngf levels in serum were measured using a 2-site immune - enzymatic assay ( elisa ) . nfg was detected in sandwich elisas according to the manufacturer 's instructions ( r&d systems , minneapolis , mn , usa ) . all assays were performed on f - bottom 96-well plates ( nunc , wiesbaden , germany ) . baseline scores and demographics for each of the 3 treatment groups were compared using manova tests , which are best suited to analyses involving several variables . within each group , mean scores for outcome variables were calculated for active treatment and placebo interventions . additional factors , including gender , age , methylphenidate dose , depression ( cdi ) , and anxiety ( masc ) , were assessed to determine any influence they may have had on scoring changes . thirty - eight individuals ( 9 girls , 29 boys , aged 8 to 18 years ) completed the study . for seasonal allergic - related symptoms , the 2-drug combination proved to be superior to either drug alone in reducing the tssc ( figure 1 ) . scores were reduced for the drug combination and for methylphenidate ; there was no significant reduction for cetirizine . when compared to cetirizine , methylphenidate had a superior effect on general scores and showed greater reduction of general symptoms ( methylphenidate : 8.1 to 5.2 points ; cetirizine : no improvement ) . for example , for nasal symptoms , methylphenidate reduced scores from 9.2 to 6.1 versus the combination of drugs ( 8.1 to 6.0 ) . for eye symptoms , combination treatment reduced scores from 7.8 to 5.2 , while methylphenidate alone demonstrated a 7.8 to 3.2 reduction . both methylphenidate and cetirizine displayed the same positive effect on emotional scores , showing reductions from 6.2 to 3.2 and 4.1 , respectively . generally , when a slow reaction time occurs with elevated omission ( missing ) and commission ( number of times the individual responded to a nontarget ) errors , there is evidence of inattention . however , the combination drug group elicited a significant decrease in the error rate compared with placebo . cetirizine lowered the commission rate from 27 to 18 and methylphenidate reduced it from 17 to 15 . is defined as the mean response time ( in milliseconds ) for all target responses over all 6 time blocks . the crs - r , in the hyperactivity and adhd index scores , was reduced from 70% ( placebo group ) to 62% ( methylphenidate / cetirizine group ) . in contrast , there was no improvement in scores when the drugs were administered individually . regarding inattention , the combination group also displayed the most improvement , with a reduction from 69% to 59% ; in the individual drug groups , the change was minor ( 70% to 68% ) . this trend continued with the oppositional scores : there was a reduction of 64% to 59% in the combination group , while the groups who received cetirizine or methylphenidate alone experienced only a 61% to 60% change , respectively . methylphenidate and cetirizine had almost identical effects on t - scores , reducing the totals from 78 to 72 and 48 to 43 points , respectively . when evaluating inattention and oppositional t - scores , we observed the same trend ; in particular , methylphenidate and cetirizine produced almost identical results in improving adhd endpoints . a large body of evidence shows that nerve growth factor ( ngf ) exerts biological activity on the immune system , thereby influencing allergic diseases and asthma [ 2931 ] . in our previous studies , we noted that enhanced ngf levels were found in patients with adhd ( compared to those in non - adhd patients ) [ 32 , 33 ] . when ngf serum levels were measured in both the treatment and placebo phases of this study , a significant reduction was noted in the treatment group versus the placebo group . in the methylphenidate group , the reduction in serum ngf levels was 21 5.1 ng / ml ; in the cetirizine group , it was 19.5 4.1 ng / ml . in the combined treatment , the greatest reduction was noted at 56.4 8.9 ng / ml ( figure 2 ) . overall , for the majority of measures for allergic rhinitis , adhd , and ngf analysis , the methylphenidate and cetirizine combination produced a greater impact than did either drug individually or placebo . a number of studies have suggested a link between allergies and adhd [ 3438 ] . two of these studies found a link between food allergies and neuropsychiatric conditions such as depression , anxiety , and adhd [ 34 , 38 ] , while others looked at immune dysregulation , asthma medications , and atopic eczema . our study supports the hypothesis that , in children diagnosed with adhd , there is a subset of children who also suffer from atopic disease . we propose that a cascade of events initiated by various stimulants ( i.e. , environmental factors , food , stress , or infection , in a person who is genetically predisposed to atopy ) may trigger an immune inflammatory cascade that may lead to the neuroimmune inflammation presented clinically as adhd . in this study , we demonstrated an improvement in adhd scores by using drugs known to regulate allergic inflammation ( e.g. , antihistamines such as cetirizine ) ; the benefit of stimulant medication on allergic inflammation ( e.g. , reduction in tssc scores by methylphenidate ) ; and the synergistic effect of methylphenidate and cetirizine on both allergic and neurological symptoms . based on these results , we suggest that there is some overlap in the mechanism of action of allergy and adhd in patients with a comorbid diagnosis . moreover , by using a synergistic approach , the individual effect of each drug on allergic or neurological endpoints is enhanced . this study also raised questions and concerns for which there are currently no obvious answers . for example , why did not we get the expected results in reduction of tssc with antihistamines in the comorbid population ? why did stimulant drugs have the same beneficial effect as antihistamines in reducing tssc scores ? and lastly , why did the combination drugs have a superior effect ? in regard to the first point , patients diagnosed with adhd have an unfavorable perception of life and , thus , antihistamine therapy may not be sufficient to control their tssc scores . additionally , in patients with specific comorbid disease , different cetirizine dosages may be required to achieve full control of allergy symptoms . regarding the second question , over the years we have observed that stimulants , specifically methylphenidate , have an antihistamine effect ( measured by suppressing wheal and flare ) . this antihistaminic effect may explain some of the improvements in tssc scores in the comorbid population . in response to the third question , antihistamines , specifically cetirizine , may have an anti - inflammatory effect [ 39 , 40 ] . this regulation of inflammatory processes can positively impact adhd endpoints in children with comorbid diseases . by adding to the antihistamine effect of cetirizine and the stimulant effect of methylphenidate the linkage between allergic and mental disorders may have a bidirectional mechanism : allergic triggers induce either a neuroimmune inflammation with a direct effect on the cns or cns stimuli ( e.g. , stress , anxiety , or other factors release neurostimulants that impact allergic inflammation ) . as we have seen , allergic disorders can lead to significant cognitive distortions , especially during the peak allergy season ( e.g. , ragweed season ) ; interestingly , this effect was found only in a subgroup of patients . these disorders have also been associated with such conditions as depression , anxiety , and panic disorder , thus raising the question as to whether these shared maladies are the result of shared genetic risks or some common mechanism , which may exist between allergic disorders and mental diseases . the mechanisms by which ngf may be involved in allergic disease or in comorbid disease of allergy and adhd are unknown , but a small number of theories have been postulated . one theory is that , upon mast activation and differentiation , ngf release is increased . the differentiation of human mast cell subtypes that could perpetuate human allergic reactions is dependent on ngf . ngf is increased in biologic fluids of allergy / immune - related diseases and has been evaluated as a th2 cytokine with a modulator role in allergic inflammation and tissue remodeling . in patients with allergic bronchial asthma , for example , elevated levels of neurotrophins have been reported . in a time course study of bronchoalveolar lavage fluid , levels of ngf correlated with the inflammatory response . it was shown that ngf , specifically , enhances inflammation in allergic early - phase response . another possibility is that , upon allergic response with a matching genetic underline , release of neurotrophins can induce brain inflammation . as part of neuroimmune healing , we need an intact apoptosis process . ngf belongs to the super family of cd40 , which plays a crucial role in this process . any failure in ngf signaling or regulation may lead to an abnormal apoptosis process and , as a result , may cause persistent inflammation . in turn our findings support the hypothesis that ( 1 ) atopy may , in some cases , contribute to the pathogenesis of adhd ; ( 2 ) ngf plays a role in the pathogenesis of comorbid diseases ; and ( 3 ) neurological triggers ( e.g. , anxiety , stress , or environmental factors , including food ) may trigger the comorbid condition . in this study , the role of ngf was demonstrated by the modulating effect of the antihistamines and stimulants on ngf levels in patients with the comorbid diseases . , our research clearly demonstrated that the combination of an antihistamine ( cetirizine ) and a stimulant ( methylphenidate ) produced superior results when treating children with comorbid allergic rhinitis and adhd , enhancing the efficacy of treatment for both allergic and adhd symptoms . our findings also suggest that ( 1 ) in patients with comorbid disease the treatment approach should include antihistamines and ( 2 ) a synergistic effect may be achieved by using both drugs , thus making it possible to reduce the dosage of the stimulant . ngf measurements followed the trends of allergic symptoms and adhd and could possibly be used as a marker for measuring the severity of either condition . | the association between adhd and allergy remains controversial .
our previous findings suggest that nerve growth factor may link the nervous and immune systems .
the primary objective of this study was to determine if a combination of cetirizine + methylphenidate is effective in children with comorbid adhd and allergic rhinitis .
we also examined the role of nerve growth factor in these comorbidities .
our randomized , double - blind , placebo - controlled , crossover study enrolled 38 children diagnosed with comorbid adhd and allergy using cetirizine ( n = 12 ) , sustained - release methylphenidate ( n = 12 ) , or cetirizine + methylphenidate ( n = 14 ) .
endpoints compared baseline to posttreatment evaluations for allergic rhinitis and adhd scores .
serum nerve growth factor levels were measured using elisa . for allergy endpoints ,
combination therapy produced results superior to individual therapy .
for adhd , similar scores were achieved for individual therapy ; however , combination therapy resulted in improved scores .
nerve growth factor levels were downregulated following this trend .
we conclude that adhd and allergic rhinitis may have common mechanism and represent a comorbid condition that links the nervous system to the immune system .
further studies are needed . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
colorectal cancer ( crc ) is a common malignant tumour and affects about 650,000 individuals worldwide . the patients are mainly of advanced age and cases before the age of 50 are infrequent , unless for a hereditary cause . the prognosis and the overall life expectancy are predominantly determined by the progression of metastatic lesions and not by the primary carcinoma . the liver constitutes the main host organ for colorectal metastases and despite the progress in diagnostic modalities , more than 25% of crc patients present with metastatic hepatic lesions at the time of initial diagnosis . surgery remains the best therapeutic approach , although only one third are potentially resectable metastases . curative resections may prolong survival up to 5 years in almost half of the patients . unfortunately , if colorectal liver metastases receive no treatment , life expectancy rarely exceeds 1 year [ 46 ] . malignant cells at the primary site initially migrate through the endothelium of the vasculature and enter the systemic circulation ( intravasation ) . then , they need to survive the mechanical pressure , collisions with other cells and attacks by immune cells ; some of them may reach the portal vein , which is the gateway to the hepatic sinusoids . the latter are specific capillaries which form a dense network , where important interactions among crc and hepatic resident cells occur , including apoptosis , angiogenesis , proteolysis and adhesion [ 79 ] . the maintenance , promotion or disruption of cell adhesion is critical for liver colonization by crc cells and numerous cell adhesion molecules ( cams ) are involved . selectins are present within the sinusoids , regulating crc cell arrest and extravasation in the liver . as they are expressed on non - parenchymal hepatic cells , such as sinusoidal endothelial cells ( secs ) and stellate cells , and their ligands on malignant cells , selectins stand at a molecular crossroads and play a pivotal role in crc liver metastases . the family of selectins includes three transmembrane cams : e - selectins , which are expressed exclusively by endothelial cells , l - selectins present in leucocytes and p - selectins in platelets and endothelial cells . selectin molecular structure consists of five domains : an n - terminal c - type lectin , a single epidermal growth factor like , two to nine complement binding domains , a single transmembrane and a short intracellular domain which forms a cytoplasmic tail ( fig . the domain termed transmembrane anchors the molecules to the cellular surface , whereas the cytoplasmic tail supports molecular signalling processes . notably , selectins require certain carbohydrates as connection mediators , including the p - selectin ligand glycoprotein 1 ( psgl-1 ) and the sialylated oligosaccharides sialyl lewis ( sle ) and x ( sle ) [ 1215 ] . moreover , it is well established that the fucose - generating fx enzyme is the main agent that interacts with selectin ligands , when the latter bind homologue molecules , functioning both as a reductase and an epimerase . consequently , fucosylation appears to control selectin - dependent adhesion [ 16 , 17 ] . selectins may be expressed and/or activated when certain mediators are present , such as numerous interleukins , tumour necrosis factor ( tnf ) or toxins . l - selectin ligands : glycosylation - dependent cam 1 ( glycam-1 ) , mucosal addressin cam 1 ( madcam-1 ) , leucocyte leukosialin ( cd34 ) , p - selectin - glycoprotein ligand 1 ( psgl-1 ) . e - selectin ligands : e - selectin ligand 1 ( esl-1 ) and psgl-1 . selectins bind their carbohydrate ligands via the c - type lectin domain in calcium - mediated manner [ 10 , 122 ] . selectins mediate tumour cell extravasation and metastasis in a similar way that they facilitate leucocyte arrest in the vasculature and migration to inflamed tissues . during the first steps of leucocyte recruitment , l - selectins expressed on t lymphocytes and other immune cells , in conjunction with p- and e - selectins on endothelial cells , interact with endothelial and leucocyte carbohydrate ligands respectively . when a primary adhesion occurs , immunoglobulin superfamily ( igsf ) members , such as intercellular adhesion molecules ( icams ) and vascular cam 1 ( vcam-1 ) , and integrins , are expressed to sustain a stable attachment and cellular signalling during transendothelial migration [ 1921 ] . following the methodology and investigating the results of leucocyte locomotion , multiple studies suggested that metastasizing crc cells originate and maintain their adhesion to the endothelium and migrate to foreign tissue through the expression of e- , p- and l - selectins , as well as their ligands , including sle , sle and cluster of differentiation 44 variants ( cd44v ) [ 2225 ] . when initial cellular bonds are successfully formed , 1 and 4 integrins in concert with igsfcams are activated to further reinforce the primary selectin bonds and support signalling toward normal tissue colonization [ 2628 ] . importantly , in vitro studies using static models [ 29 , 30 ] or flow shear stress conditions which better depict the metastatic environment , advocated that not only does e - selectin support primary malignant cell attachment to the endothelium , but also regulates diapedesis ( the transmigration of circulating cells through the vascular endothelium ) and crc cell invasion of the hepatic parenchyma . a dna microarray analysis revealed that e - selectin provoked gene expression alterations in metastatic crc cells , down - regulating seven genes ; the influence was 10-fold higher in comparison with primary non - metastatic cells . the cellular expression of high mobility group box 1 , a chromosomal protein involved in dna transcription and repair , was also decreased , but its free release was promoted , resulting in endothelial cell activation and e - selectin expression . it was also observed that while this cam is not present on colon cells , it is highly expressed on small blood vessels , close to metastatic colon cancer lesions . furthermore , measurements of soluble e - selectin in patients with crc demonstrated that high serum values significantly correlated with hepatic metastases [ 3436 ] . p - selectins play a crucial role in platelet - crc cell interactions , as they are expressed by the former and bind to fucosylated sialylated mucin ligands of the latter . accumulating data support that haematogenous metastasis involves platelet - malignant cell interactions . from a mechanistic point of view , platelets may form complexes with tumour cells and leucocytes and cause their arrest in the vascular wall ; these cellular masses of increased volume function as emboli and are prone to entrapment in the vasculature . subsequently , platelets appear to promote malignant cell extravasation . also , platelets may stimulate tumour proliferation , enhance interactions with the extracellular matrix ( ecm ) and induce tumour growth and angiogenesis mediating the production of molecules like the platelet - derived growth factor or vascular endothelial growth factor ( vegf ) [ 37 , 38 ] . experimental analysis of primary crc tissue specimens , in comparison with secondary hepatic lesions , indicated that liver metastases were virtually deprived from p - selectin expression and leucocyte infiltration . on the contrary , primary tumours presented significantly higher levels of this cam , as well as leucocyte intramural activity . it was concluded that p - selectin aids crc cells to evade inflammatory reaction , promoting the metastatic process . experiments under shear flow conditions revealed that p - selectin may form the initial bonds for metastasizing cells to adhere within the vasculature [ 4042 ] . similar flow models showed a favourable therapeutic action of heparin against colon metastasis through blockade of p - selectin bonds . venous thromboembolism appears to affect frequently patients who receive anti - cancer treatment , compromising the quality of their life and increasing mortality . although its clinical use is still under evaluation , it appears that this cam could be exploited in the identification of patients at high risk for venous thromboembolism , such as those under chemotherapy , who should receive thromboprophylaxis , including low molecular or unfractionated heparin [ 44 , 45 ] . the in vitro experimental investigation of crc cell kinetics in flow conditions indicated that these malignant cells interact with polymorphonuclear leucocytes and form complexes in the same order of magnitude , as they bind to platelets . notably , l - selectin - mediated bonds appeared to be the most stable against shear stress and permitted the formation of aggregates , facilitating crc cell arrest in the microvessels of distant organs . additionally , a synergy of p- and l - selectins during the metastatic process of colon carcinoma was observed , where p - selectins supported platelet and tumour cell interactions , while l - selectins acted in later stages of metastasis . the importance of adhesion molecules in liver metastasis , including the selectins , was also experimentally highlighted through intravital microscopy . fluorescence labelled crc cells were injected in rodents and their circulation was observed within the liver microvasculature . it was concluded that malignant cells interacted with the sinusoidal endothelium and adhered via e- and p - selectins , icam-1 and vcam-1 . the role of mechanical entrapment was underestimated in these experiments , because no tumour cell arrest was observed in capillary systems with smaller diameters , such as renal , mesenteric or muscular ones . moreover , tnf- , a cytokine produced by stimulated kupffer cells ( kcs ) , appeared to promote the expression of adhesion molecules early during the metastatic process [ 4850 ] . carcinoembryonic antigen ( cea ) may function as an auxiliary e- and l - selectin ligand and stabilize colon cells against fluid shear during their dissemination in the vasculature . also , it was observed that cea cooperates with cd44 variant isoforms ( cd44v ) , which are functional ligands for p - selectin , when fluid shear stress increases . these important findings could explain the high metastatic potential of cea - overexpressing crc cells , through selectin - mediated molecular pathways . kcs , the hepatic macrophages , present an 80 kda cea receptor ( cea - r ) , classified as -2 adrenergic , responsible for binding and subsequent degradation of cea [ 52 , 53 ] . experiments on murine livers , demonstrated that the stimulation of these macrophages with cea caused the production of cytokines , such as tnf- and il-1. culture of human umbilical cord endothelial cells ( huvecs ) in conditioned media from these stimulated kcs induced the expression of e - selectin by the endothelial cells and their adhesion with highly metastatic crc cell lines . the association of cea and e - selectin through kcs and vascular endothelium is another important clue in liver metastasis research , although it should be noted that the role of kcs is not limited to cell adhesion . these cells may also exert cytotoxic activities against tumour cells or release growth factors , such as hepatocyte growth factor ( hgf ) , and proteolytic enzymes , such as metalloproteinase 9 ( mmp-9 ) , promoting malignant cell proliferation and extravasation , as well as angiogenesis and ecm degradation [ 55 , 56 ] . selectins appear to be differentially expressed on the vascular endothelium of various tissues , as was observed for e - selectin on the rat central nervous system and the human ocular microvasculature [ 57 , 58 ] . furthermore , multiple experiments investigated e - selectin expression after tnf and il-1 stimulation on human dermal microvascular endothelial cells and human intestinal microvascular endothelial cells in comparison with huvecs [ 5961 ] or on human iliac venous- and arterial venous endothelial cells ; they all concluded that endothelial cells of different origin express e - selectin in a dissimilar way . on the other hand , colon cancer cell lines express different selectin ligands and consequently adhere to different selectin molecules . while colo320 cell line may link with p- and l- selectins , ht-29 only binds e - selectin and caco-2 shows no interaction with members of the selectin family . additionally , different colon cancer cell lines present alternative adhesion kinetics to e - selectin . the investigation and analysis of the preceding data concluded that selectins may play a pivotal role in the selection of the host organ for the development of distant metastases . this is an interesting proposal which may substantially contribute to the explanation of the high incidence of liver colonization by crc cells . the association of vegf with e - selectin was studied in murine models with sarcoma cells . it was reported that vegf is produced by tumour cells and enhances angiogenesis through a significant up - regulation of e - selectin on vascular endothelial cells . although this interrelationship appears crucial for the development of metastases , no research data have been published on crc . however , recent experiments on stellate cells , the liver sinusoidal fat - storing cells , showed that when activated , they express e - selectin ligand 1 ( esl-1 ) on their cellular membrane and that this expression could be associated with hypoxia . as stellate cells may function as oxygen - sensing cells and are involved in angiogenesis , their expression of the main e - selectin ligand reveals a new cell type potentially involved in crc liver metastasis [ 68 , 69 ] . selectins constitute a major therapeutic target for multiple maladies , such as asthma , psoriasis , endotoxemia and cancer , and various antagonists have been developed . 2 ) is a pan - selectin synthetic antagonist and the leading selectin inhibitor in clinical development . it has been tested in animal models and human beings for inflammatory diseases , such as asthma and chronic obstructive pulmonary disease , with favourable results and low toxicity [ 7072 ] . heparin also exerts anti - inflammatory as well as anti - metastatic effects , partly inhibiting l- and p - selectin binding . although , its animal origin and heterogeneous structure limit its value and new semi - synthetic glucan sulphates were produced and administered in murine models . phycarin sulphates were reported to block p - selectin effectively and their application as anti - inflammatory and anti - cancer drugs will be further evaluated [ 7375 ] . the pan - selectin antagonist bimosiamose ( [ hexane-1,6-diylbis [ 6-(-d - mannopyranosyloxy ) biphenyl-3,3-diyl ] ] diacetic acid ) http://www.who.int/druginformation ( who drug information vol.15 , no.3&4 , 2001 ) . a promising one appears to be huep5c7.g2 , a humanized antibody , which blocks leukaemia cell binding to e- and p - selectin positive cells and possesses favourable pharmacokinetic properties , with long circulation half - life [ 76 , 77 ] . moreover , synthetic ligands for selectin binding were in vitro tested and gold colloid particles presented high values of l- and p - selectin immobilization with no cytotoxic effect ; their further assessment on animal models is on the way . in general , while the inhibition of selectins appears a promising treatment for several diseases , it is still under evaluation mainly in animal models , a limited number of pharmaceutical compounds have reached small clinical trials and no reliable data exist for solid tumours , such as crc and its metastases . the lewis blood group includes multiple structurally similar carbohydrates present on erythrocytes , but also in other different tissues . it has been shown that certain sialylated lewis antigens , membranous cell glycoproteins which end in nine carbon molecules named sialic acids , are involved in cellular adhesions with the ecm and with endothelial cell - related ligands , such as the selectins , during tumour progression . it has been discovered that both these antigens are expressed in tumours located throughout its epithelium , while sle and sle are expressed only in neoplasms of the distal colon [ 7981 ] . glycosyltransferases constitute a wide category of enzymes that transfer monosaccharide units and include several families , such as sialyl- and fucosyltransferases . these two enzyme families are involved in neoplastic transformation and cancer development through their role in cell differentiation and adhesion . therefore , any modification in their function may alter the status of cell connection [ 8284 ] . chemical structure of sialyl lewis x ( sle ) and ( sle ) . in the primary crc with poor outcome the fucosyltransferases-3 and -4 ( fuc - tiii and fuc - tvi ) and sialyltransferase3galactosamine-4 ( st3gal - iv ) mediated sle antigen synthesis in the colon , but were not responsible for the increased expression of these antigens in crc . this increase should be attributed to a combinational up - regulation of multiple glycosyltransferases genes [ 85 , 86 ] . immunohistochemical studies revealed an augmented fuc - tvi expression in human colorectal carcinomas , which was associated with lymph node metastasis and tumour stage . notably , increased values of the enzyme correlated with the degree of tumour infiltration through the intestinal wall . murine experiments displayed that the expression of fuc - ti induces lower levels of sle in crc cells , inhibiting their dissemination and thus liver metastases , due to decreased adhesion capacity to e - selectin . moreover , in colorectal liver metastases , there was a decrease in sialyltransferase levels and increase in fucosyltransferase expression . the sle antigen is a tetrasaccharide , usually attached to o - glycans on the cellular surface . it is well studied in crc and appears to correlate closely with the bad prognosis of the disease . its levels are reversely associated with survival of operated patients for primary crc and analogous to crc metastatic ability . the same results were announced through an animal - based model of crc liver metastases . high levels of sle were associated with increased expression of e - selectin , cell adhesion and liver metastasis . a multivariate analysis of patient records with crc metastasis showed that sle is an independent prognostic factor for the histologic type and the recurrence of the disease , as well as the invasion depth , in contrast with sle that is not . sialyl lewis , also termed carbohydrate antigen 199 ( ca 199 ) , is most frequently linked with o - glycans on mucins of the cellular surface . on normal tissue it is restricted to ductal epithelium , but is widely expressed in multiple carcinomas including colon cancer [ 79 , 93 ] . this tetrasaccharide binds to e- and p - selectin and controls the extravasation and the attachment of crc cells to endothelium . it was also shown that down - regulation of this antigen on crc cells via genetic modifications , substantially reduced their extravasation [ 9496 ] . several studies on murine models assessed the involvement of sle in crc liver metastases either in vitro or in vivo . this action was also related to increased fuc - tiii enzymatic activity [ 9799 ] . dabrowska et al . studied the association of fuc - tiii with sle synthesis in five different colon cancer cell lines and concluded in favour of a strong positive regulatory role for the enzyme . furthermore , in vitro studies demonstrated that hypoxic conditions enhanced the expression of e - selectin ligands , such as sle and sle , by colon cancer cells and promoted selectin - mediated cell adhesion during metastasis . the clear role of sle and sle in crc metastasis caused therapeutically oriented research , which targeted these molecules . cimetidine , a histamine h2 receptor antagonist , was administered for 1 year to patients with crc , who had undergone a curative resection and were under 5-fluorouracil treatment ; significant beneficial effects were announced referring to 10-year survival . importantly , patients highly expressing sle and sle presented the most favourable results . it was claimed that cimetidine could block the expression of e - selectin on vascular endothelium and thus inhibit the adhesion of crc cell ligands . moreover , in vivo murine experiments , testing the effect of sle analogue gsc-150 in crc liver metastasis , reported that this agent reduced the number of metastatic nodules , inhibiting malignant cell adhesion ; this was also displayed via in vitro studies on huvecs . recently , sle conjugates were synthesized and used as vaccines in mice , against crc and lung cancer cells . the produced antibodies proved to be highly active and no reactivity with other sialyl lewis antigens was detected . cd44 or homing - associated cam ( h - cam ) is a family of transmembrane glycoproteins , including several isoforms expressed on epithelial , endothelial and tumour cells . these isoforms differ in the extracellular domain , where combinations of 10 variant exons may occur . cd44 serves as a hyaluronan receptor , a glycosaminoglycan , responsible for cell motility and proliferation . furthermore , through interactions with other molecules , such as osteopontin , collagen , matrix metalloproteinases and selectins , it regulates cell adhesion among endothelial and hematopoietic cells , fibroblasts and the ecm [ 25 , 105 , 106 ] . the wide expression of cd44 on neoplastic cells has been well studied and its predictive value has been proposed in tumour invasion , migration and angiogenesis . increased values of cd44 were associated with venous invasion in crc and correlated , along with e - cadherin , to poor overall survival , especially in stage ii . murine experiments demonstrated that it promotes tumour growth , anti - apoptosis and crc cell motility , and its genetic suppression induces malignant cell apoptosis and migration via akt kinase [ 108110 ] . various isoforms of cd44 are frequently present in advanced stages of colorectal carcinogenesis and its liver metastases . cd44 splice variant 6 ( cd44v6 ) appears to be associated with crc liver metastases , although there is no agreement if this glycoprotein is highly or lowly [ 112 , 113 ] expressed on tumour cells . clinical and experimental studies associated cd44v6 with colon cancer stage ( dukes classification ) , hepatic metastasis and 5-year survival [ 114 , 115 ] . additional confirmation of cd44 s value in the prognosis of colonic malignancies was provided by wang et al . who revealed that high levels of heat shock protein 72 , a molecular chaperone which regulates cancer cell growth and apoptosis , were connected with high cd44v6 levels in human colonic cancer , compared to normal colon . in contrast to the preceding conclusions , a clinical study on 56 patients with dukes c or d crc reported no difference of cd44v6 expression in the primary site and the metastatic lymph nodes . a recent analysis of crc metastatic lesions revealed that the complex of epithelial cam ( epcam ) , claudin-7 , cd44v6 and a tetraspanin member was strongly related to poor prognosis and low disease free survival , although the solitary expression of each of these proteins showed no similar outcome . moreover , the expression of cd44v8 to -10 revealed no significant correlation with the histology , lymphatic and venous invasion of the primary crc , but was significantly related with lymph node and haematogenous metastases . also , patients with increased values of these ligands presented higher recurrence and lower 5- and 10-year survival rates . consequently , there is mounting evidence which connects cd44v isoforms with crc prognosis and its migration to the liver , but this relationship is neither well defined , nor clearly demonstrated in the clinical field . in vitro studies on multiple colon carcinoma cell lines concluded that cd44v isoforms mediate tumour cell adhesion to platelets , leucocytes and endothelial cells , through links with p- , l- and e- selectins , and fibrin . additionally , heparin reduced cell adhesion to p- and l- selectins . as the selectin family and fibrin play a critical role in metastasis , these findings present cd44 as a potential target of future therapeutic applications [ 25 , 120 , 121 ] . the lewis blood group includes multiple structurally similar carbohydrates present on erythrocytes , but also in other different tissues . it has been shown that certain sialylated lewis antigens , membranous cell glycoproteins which end in nine carbon molecules named sialic acids , are involved in cellular adhesions with the ecm and with endothelial cell - related ligands , such as the selectins , during tumour progression . it has been discovered that both these antigens are expressed in tumours located throughout its epithelium , while sle and sle are expressed only in neoplasms of the distal colon [ 7981 ] . glycosyltransferases constitute a wide category of enzymes that transfer monosaccharide units and include several families , such as sialyl- and fucosyltransferases . these two enzyme families are involved in neoplastic transformation and cancer development through their role in cell differentiation and adhesion . therefore , any modification in their function may alter the status of cell connection [ 8284 ] . chemical structure of sialyl lewis x ( sle ) and ( sle ) . in the primary crc with poor outcome the fucosyltransferases-3 and -4 ( fuc - tiii and fuc - tvi ) and sialyltransferase3galactosamine-4 ( st3gal - iv ) mediated sle antigen synthesis in the colon , but were not responsible for the increased expression of these antigens in crc . this increase should be attributed to a combinational up - regulation of multiple glycosyltransferases genes [ 85 , 86 ] . immunohistochemical studies revealed an augmented fuc - tvi expression in human colorectal carcinomas , which was associated with lymph node metastasis and tumour stage . notably , increased values of the enzyme correlated with the degree of tumour infiltration through the intestinal wall . murine experiments displayed that the expression of fuc - ti induces lower levels of sle in crc cells , inhibiting their dissemination and thus liver metastases , due to decreased adhesion capacity to e - selectin . moreover , in colorectal liver metastases , there was a decrease in sialyltransferase levels and increase in fucosyltransferase expression . the sle antigen is a tetrasaccharide , usually attached to o - glycans on the cellular surface . it is well studied in crc and appears to correlate closely with the bad prognosis of the disease . its levels are reversely associated with survival of operated patients for primary crc and analogous to crc metastatic ability . the same results were announced through an animal - based model of crc liver metastases . high levels of sle were associated with increased expression of e - selectin , cell adhesion and liver metastasis . a multivariate analysis of patient records with crc metastasis showed that sle is an independent prognostic factor for the histologic type and the recurrence of the disease , as well as the invasion depth , in contrast with sle that is not . sialyl lewis , also termed carbohydrate antigen 199 ( ca 199 ) , is most frequently linked with o - glycans on mucins of the cellular surface . on normal tissue it is restricted to ductal epithelium , but is widely expressed in multiple carcinomas including colon cancer [ 79 , 93 ] . this tetrasaccharide binds to e- and p - selectin and controls the extravasation and the attachment of crc cells to endothelium . it was also shown that down - regulation of this antigen on crc cells via genetic modifications , substantially reduced their extravasation [ 9496 ] . several studies on murine models assessed the involvement of sle in crc liver metastases either in vitro or in vivo . this action was also related to increased fuc - tiii enzymatic activity [ 9799 ] . dabrowska et al . studied the association of fuc - tiii with sle synthesis in five different colon cancer cell lines and concluded in favour of a strong positive regulatory role for the enzyme . furthermore , in vitro studies demonstrated that hypoxic conditions enhanced the expression of e - selectin ligands , such as sle and sle , by colon cancer cells and promoted selectin - mediated cell adhesion during metastasis . the clear role of sle and sle in crc metastasis caused therapeutically oriented research , which targeted these molecules . cimetidine , a histamine h2 receptor antagonist , was administered for 1 year to patients with crc , who had undergone a curative resection and were under 5-fluorouracil treatment ; significant beneficial effects were announced referring to 10-year survival . importantly , patients highly expressing sle and sle presented the most favourable results . it was claimed that cimetidine could block the expression of e - selectin on vascular endothelium and thus inhibit the adhesion of crc cell ligands . moreover , in vivo murine experiments , testing the effect of sle analogue gsc-150 in crc liver metastasis , reported that this agent reduced the number of metastatic nodules , inhibiting malignant cell adhesion ; this was also displayed via in vitro studies on huvecs . recently , sle conjugates were synthesized and used as vaccines in mice , against crc and lung cancer cells . the produced antibodies proved to be highly active and no reactivity with other sialyl lewis antigens was detected . cd44 or homing - associated cam ( h - cam ) is a family of transmembrane glycoproteins , including several isoforms expressed on epithelial , endothelial and tumour cells . these isoforms differ in the extracellular domain , where combinations of 10 variant exons may occur . cd44 serves as a hyaluronan receptor , a glycosaminoglycan , responsible for cell motility and proliferation . furthermore , through interactions with other molecules , such as osteopontin , collagen , matrix metalloproteinases and selectins , it regulates cell adhesion among endothelial and hematopoietic cells , fibroblasts and the ecm [ 25 , 105 , 106 ] . the wide expression of cd44 on neoplastic cells has been well studied and its predictive value has been proposed in tumour invasion , migration and angiogenesis . increased values of cd44 were associated with venous invasion in crc and correlated , along with e - cadherin , to poor overall survival , especially in stage ii . murine experiments demonstrated that it promotes tumour growth , anti - apoptosis and crc cell motility , and its genetic suppression induces malignant cell apoptosis and migration via akt kinase [ 108110 ] . various isoforms of cd44 are frequently present in advanced stages of colorectal carcinogenesis and its liver metastases . cd44 splice variant 6 ( cd44v6 ) appears to be associated with crc liver metastases , although there is no agreement if this glycoprotein is highly or lowly [ 112 , 113 ] expressed on tumour cells . clinical and experimental studies associated cd44v6 with colon cancer stage ( dukes classification ) , hepatic metastasis and 5-year survival [ 114 , 115 ] . additional confirmation of cd44 s value in the prognosis of colonic malignancies was provided by wang et al . who revealed that high levels of heat shock protein 72 , a molecular chaperone which regulates cancer cell growth and apoptosis , were connected with high cd44v6 levels in human colonic cancer , compared to normal colon . in contrast to the preceding conclusions , a clinical study on 56 patients with dukes c or d crc reported no difference of cd44v6 expression in the primary site and the metastatic lymph nodes . a recent analysis of crc metastatic lesions revealed that the complex of epithelial cam ( epcam ) , claudin-7 , cd44v6 and a tetraspanin member was strongly related to poor prognosis and low disease free survival , although the solitary expression of each of these proteins showed no similar outcome . moreover , the expression of cd44v8 to -10 revealed no significant correlation with the histology , lymphatic and venous invasion of the primary crc , but was significantly related with lymph node and haematogenous metastases . also , patients with increased values of these ligands presented higher recurrence and lower 5- and 10-year survival rates . consequently , there is mounting evidence which connects cd44v isoforms with crc prognosis and its migration to the liver , but this relationship is neither well defined , nor clearly demonstrated in the clinical field . in vitro studies on multiple colon carcinoma cell lines concluded that cd44v isoforms mediate tumour cell adhesion to platelets , leucocytes and endothelial cells , through links with p- , l- and e- selectins , and fibrin . additionally , heparin reduced cell adhesion to p- and l- selectins . as the selectin family and fibrin play a critical role in metastasis , these findings present cd44 as a potential target of future therapeutic applications [ 25 , 120 , 121 ] . selectins and their ligands , such as sle , sle and cd44 isoforms , facilitate the metastatic process and promote crc cell extravasation from the sinusoids . while selectins are expressed in platelets , leucocytes and secs , their ligands are present on malignant cells and their connections mediate crc cell apoptosis , proliferation , motility and adhesion to different cells and the ecm . importantly , multiple cytokines released by kupffer , stellate and other cells , are also involved in the expression of selectins and thus the cellular intercommunication within the sinusoids develops and appears more complicated ( fig . kcs , presenting cea receptors , may be activated through cea molecules released by malignant cells , produce cytokines which stimulate endothelial cells to express e - selectin . metastasizing colorectal cells also present sle and cd44v ligands , which link with e- , p- and l - selectins . cd44v : cd44 variant , cea : carcinoembryonic antigen , cea - r : cea receptor , crc : colorectal cancer , il-1 : interleukin 1 , sle : sialyl lewis antigen , tnf- : tumour necrosis factor . effective anti - cancer treatment of the future will primarily rely on molecular analysis . in the field of crc liver metastasis several selectin inhibitors have been composed and tested via animal models and/or clinical trials , however progress was slow , because these cams are present in numerous physiological functions and their blockade could cause ample complications . cimetidine , a well - established h2 receptor antagonist , showed significant anti - cancer action in preliminary studies with crc patients and should be additionally studied , in conjunction with other medicines of this category . further research on human tissue should consolidate successful therapeutic results from animal models and bigger clinical trials should evaluate the potential of old and new selectin inhibitors in the treatment of crc liver metastases . | abstractthe colonization of the liver by colorectal cancer ( crc ) cells is a complicated process which includes many stages , until macrometastases occur .
the entrapment of malignant cells within the hepatic sinusoids and their interactions with resident non - parenchymal cells are considered very important for the whole metastatic sequence . in the sinusoids , cell connection and signalling
is mediated by multiple cell adhesion molecules , such as the selectins .
the three members of the selectin family , e- , p- and l - selectin , in conjunction with sialylated lewis ligands and cd44 variants , regulate colorectal cell communication and adhesion with platelets , leucocytes , sinusoidal endothelial cells and stellate cells .
their role in crc liver metastases has been investigated in animal models and human tissue , in vivo and in vitro , in static and shear flow conditions , and their key - function in several molecular pathways has been displayed .
therefore , trials have already commenced aiming to exploit selectins and their ligands in the treatment of benign and malignant diseases .
multiple pharmacological agents have been developed that are being tested for potential therapeutic applications . |
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besides their function as constituents of cellular membranes , sphingolipids play an important role as signalling molecules . they have essential functions for cell growth , cell differentiation , and cell death ( apoptosis ) , thus being indispensable for cell homeostasis and normal cell development . especially the equilibrium between ceramide and sphingosine-1-phosphate ( s1p ) seems to be decisive for a final cellular response [ 14 ] . sphingolipid rheostat is primarily regulated by three classes of enzymes : the ceramide generating sphingomyelinases , the ceramide degrading ceramidases , and finally the sphingosine kinases . further enzymes controlling ceramide levels are the sphingomyelin synthases [ 57 ] and the glucocerebroside synthase . ceramide often accumulates after stimulation of cells with stress factors like inflammatory cytokines , uv radiation , x - rays , heat shock , and oxidative stress . as a consequence to such primary triggers , neutral or acid sphingomyelinases ( nsmase or asmase ) are activated , in which both cleave sphingomyelin to form ceramide [ 9 , 10 ] . the exact mechanism , by which the respective smases are activated , is still not completely understood . small molecule inhibitors that specifically act on the various sphingolipid - degrading enzymes can help in clarifying the underlying mechanisms . recently , we developed a number of potent and specific inhibitors of the acid sphingomyelinase , which inhibited dexamethasone - induced apoptosis in hek-293 cells , respectively [ 1113 ] . the action of smases , however , controls only the influx of ceramide and thus only partially contributes to understanding cellular ceramide concentration . the contribution coming from the degradation of ceramide by ceramidases ( cdases ) is similarly important . several different cdases exist , which can be characterized by their different ph - optima ( e.g. , neutral and alkaline ) and subcellular localization . the acid ceramidase ( acdase ) is doubtlessly indispensable for the cellular ceramide degradation , and a deficiency in acid ceramidase leads to morbus farber , a recessively inherited lysosomal storage disease characterized by massive lysosomal ceramide accumulation . patients have a strongly limited life expectancy and depending on the residual enzyme activity die within a few years . the phenotype of the classical form ( type 1 ) includes progressive articular deformations , subcutaneous lumps , inflammatory granulomas , and progressive neurological dysfunction . the acdase was shown to be upregulated in prostate cancer and melanomas , which lead to the hypothesis that the acdase could be a tumour marker [ 22 , 23 ] . moreover , acdase in tumour cells obviously confers resistance to chemo- and radiotherapy thus making the inhibition of this enzyme a potential target for cancer therapy . at present , there are three different lead structures for acdase inhibitors ( figure 1 ) . the acdase is selectively inhibited by b-13 with an ic50 of about 10 m and less potently by n - oleoylethanolamine ( noe , ki ~ 500 m ) . d - mapp , which is structurally related to b-13 , has been reported to selectively inhibit alkaline ceramidase activity derived from hl-60 cell extracts with an ic50 of about 15 m . in another report , however , d - mapp is a moderate but selective inhibitor of acdase in human melanoma and hacat cells . the three lead structures , b-13 , noe , and d - mapp , can be obtained by a single synthetic step from commercially available chemicals , thus making them attractive starting points for combinatorial chemistry and sar studies . because of their potential as anticancer drugs , several novel ceramidase inhibitors have been developed recently . from a collection of noe analogues , two compounds not only inhibited acdase in cell culture ( ic50 ~ 15 m ) , but at the same time exhibited cytotoxicity to a549 cells ( ic50 ~ 40 m ) [ 29 , 30 ] . the screening of these potential cdase inhibitors has been facilitated by a fluorescent ceramidase probe allowing for high - throughput screening of cdase inhibitors in vitro and in cell culture . in an effort to improve the activity of b-13 and d - mapp in vitro and in cell culture , bielawska and colleagues recently conducted several very detailed and interesting studies . in an attempt to develop acdase inhibitors that accumulate in lysosomes , b-13 and d - mapp analogues carrying basic functional groups have been evaluated ( figure 2 ) [ 3234 ] . several of these compounds were synthesized before in the group of shimon gatt [ 35 , 36 ] . similar to the tricyclic antidepressants imipramine and desipramine , these molecules are in equilibrium between the protonated cationic form and the unprotonated neutral form . only the latter can permeate lysosomal membranes and enter the lysosomes . in the acidic environment of the lysosomes , these substances immediately get fully protonated . very rapidly , as a consequence of the steady - state equilibrium , nearly the whole portion of the substance can be found in the lysosomes , thereby increasing its effective in situ concentration . indeed , the substances lcl 204 , lcl 385 , and lcl 464 accumulated in the acidic compartments of the cells , while the neutral analogues showed no spatial preference . interestingly , the lysosomotropic acdase inhibitor lcl 204 was shown to induce apoptosis in prostate cancer cells and to enhance the cytotoxicity of the viral protein apoptin in prostate cancer . this compound is also active in head and neck squamous cell cancers ( hnsccs ) in vitro and in vivo in a xenograft model sensitizing cells to fas ligand - induced apoptosis . in addition , the closely related analogue lcl-385 , sensitized ppc-1 prostate cancer cells to radiation and significantly decreased tumor xenograft growth . a more detailed biochemical investigation , however , revealed that lcl 204 ( and probably also lcl 385 ) induce unwanted lysosomal permeabilization and degradation of acdase . these positively charged agents induce proteolytic degradation of acid sphingomyelinase and probably of acid ceramidase by blocking the negatively charged lipid bis(monoacylglycero)phosphate ( bmp ) , which is specific to the membranes of intralysosomal vesicles and has proven indispensable for the action of many sphingolipid hydrolases . obviously , bmp promotes the binding of sphingolipid hydrolases and sphingolipid activator proteins ( saps ) to the intralysosomal membranes thereby protecting these proteins from proteolytic degradation . recent investigation showed that bmp is essential for the binding of lysosomal hsp70 which promotes lysosomal integrity . thus , agents comprising basic groups that may be protonated in the lysosomes have the potential to act by perturbing lysosomal integrity . in fact , similar to the tricyclic antidepressants , lcl 204 hardly inhibits acdase from cell extracts ( 5% inhibition at 50 m ) . in contrast , b-13 ( 95% ) is much more potent in cell extracts but revealed being virtually inactive against acdase activity in intact mcf7 cells , probably due to its inability to accumulate in cellular lysosomes . the reason for the marked increase in cellular ceramide caused by b-13 is currently unknown . it has to be noted that in an earlier study , lcl 204 ( ad2646 ) showed an ic50 of 42 m , underscoring the importance of using purified enzymes for sar studies . a second generation basic b-13 analogue , lcl 464 , was shown to inhibit cdase activity in cell extracts ( 50% at 50 m ) . compared to lcl 204 this compound is not only more potent in cellular extracts , but also shows superior inhibition of a cdase in living cells , without reducing the amount of acdase at the protein level . these results show that the cellular inhibition of acid ceramidase depends on both , the in vitro activity and the ability to accumulate in cellular lysosomes . the molecular basis of the ceramidase inhibition by d - mapp and b-13 is still unknown . small libraries of compounds with systematically altered functional groups or stereochemistry can provide information about structural requirements for binding to ceramidases . preceding the investigation of the structure - activity relationship ( sar ) for b13-inhibiting acid ceramidase , we did a retrosynthetic analysis of the b-13/d - mapp scaffold . it is known that the base of d - mapp , norephedrine , can easily be obtained by nitroaldol reaction ( henry reaction ) of nitroethane with benzaldehyde and subsequent reduction ( see retrosynthetic analysis in figure 3 ) . the retrosynthetic analysis revealed three variable moieties , an aromatic part ( ar ) , a nitrogen - bound substituent r2 , and a nitrocomponent ( o2n ch2r1 ) . the simplest organic nitro component is nitromethane ( r1 = h ) , leading to bases of type 3 ( figure 4 ) . in our initial activity screening , the b-13 analogue dp24a displayed a remarkable inhibitory activity towards the recombinant human acdase ( figure 5 ) . in a more detailed study , the b-13 lead structure contains an aromatic nitro group which decreases electron density of the aromatic ring ( m effect ) . we wanted to test whether electron donors like methoxy groups ( + m effect ) would influence the inhibitory properties of our compounds . catalytic hydrogenation of the latter afforded 3b , which was treated with myristoyl chloride to give the final b-13 analogue dp24b . at a concentration of 2.5 m both , these results suggest that neither the nitro- nor the methoxygroup influence the potency of acdase inhibition . next we wanted to evaluate alternative possibilities for an introduction of basic amino groups for a facilitating lysosomal enrichment . lipid hydrolases obviously contain large hydrophobic pockets which harbour the fatty acid moieties of substrate lipids . recently we have shown for three different substance classes inhibiting acid sphingomyelinase that the inhibitory potency positively correlates with the length of hydrocarbon chains [ 1113 ] . we hypothesize that the remarkable drop in cell lysate acdase inhibition from b-13 ( 95% inhibition ) to lcl-464 ( 50% inhibition ) is a result of the partial disruption of the major hydrophobic interaction between the acdase and the positively charged fatty acid chain in lcl-464 . this problem might be circumvented by introduction of basic moieties at sites other than the fatty acid chain . thus we synthesized a pyridine analogue of dp24 by reaction of nicotinic aldehyde 1c with nitromethane . the resulting nitrocompound 2c was reduced to the respective amine 3c and reacted with myristoyl chloride to afford dp24c . although dp24c provides a basic moiety , it showed at least the same inhibition as dp24a or dp24b ( figure 5 ) . interestingly , we noticed that the enantiomer of b-13 ( ent - b-13 comprising s , s stereochemistry ) also showed a remarkable inhibition of a cdase , which is contradictory to the literature data obtained with cellular lysates . a possible explanation would be that ent - b-13 is readily cleaved by unknown enzymes from cellular lysates . although early investigations did not provide evidence for b-13 being an alternative substrate to acdase , we wanted to replace the biologically labile amide bond by the more stable urea group . evaluation in the acdase assay revealed only a slightly better inhibition , which is probably due to the slightly increased length of the hydrophobic tail of the b-13 analogues ( figure 5 ) . in our preliminary in vitro studies , we were able to identify a new b-13 analogue with increased potency versus the recombinant human acdase . this new simplified compound can easily be synthesized and offers the possibility for a more diverse series of b-13 analogues . moreover , we have proposed a new way for introducing weakly basic isosteres in the aromatic part of the molecule . in contrast , lcl 464 shows strongly reduced acdase inhibition in vitro when compared to b-13 meaning that the high potency in cells is to a large extent due to lysosomal accumulation . however , it remains to be clarified whether the pyridine group of dp24c ( pkb ~ 9 ) is as suitable as the tertiary amino group of lcl 464 ( pkb ~ 3 ) for lysosomal targeting . furthermore , it has to be noted that substances comprising a basic moiety for lysosomal enrichment may induce lysosomal permeabilization . comparative cellular studies will be performed in the near future . in the course of our initial investigation of the structure - activity relationship of b-13 analogues optical pure compounds , however , are possible via an enantioselective henry reaction [ 45 , 46 ] . general procedure for the synthesis of henry products 2b and 2cto a stirred solution of aldehyde 1b or 1c ( 1.5 mmol ) in 10 ml nitromethane was added triethylamine ( 1 ml ) as catalyst . the resulting solution was stirred at room temperature until the consumption of the aldehyde was indicated by tlc . it was diluted with ethyl acetate ( 200 ml ) , washed with sodium hydrogen sulfite , sat . sodium hydrogen carbonate , water , and brine , dried over anhydrous sodium sulfate , and concentrated in vacuum . the crude product was purified by column chromatography on silica gel ( cyclohexane / ethyl acetate 2 : 1 , v / v ) .
to a stirred solution of aldehyde 1b or 1c ( 1.5 mmol ) in 10 the resulting solution was stirred at room temperature until the consumption of the aldehyde was indicated by tlc . it was diluted with ethyl acetate ( 200 ml ) , washed with sodium hydrogen sulfite , sat . sodium hydrogen carbonate , water , and brine , dried over anhydrous sodium sulfate , and concentrated in vacuum . the crude product was purified by column chromatography on silica gel ( cyclohexane / ethyl acetate 2 : 1 , v / v ) . general procedure for the reduction of henry products to aminols 3b and 3cthe henry product ( 0.23 mmol ) was dissolved in 5 ml methanol and charged with a catalytic amount of palladium on activated charcoal . the reaction mixture was stirred at room temperature until the consumption of the henry products was indicated . the crude product was purified by column chromatography on silica gel ( cyclohexane / ethyl acetate 1 : 1 , v / v ) . the henry product ( 0.23 mmol ) was dissolved in 5 ml methanol and charged with a catalytic amount of palladium on activated charcoal . the reaction mixture was stirred at room temperature until the consumption of the henry products was indicated . the crude product was purified by column chromatography on silica gel ( cyclohexane / ethyl acetate 1 : 1 , v / v ) . general procedure for the preparation of amide inhibitors dp24 a ca solution of aminol ( 0.145 mmol ) in 5 ml pyridine was placed in an ice bath and treated with myristoyl chloride ( 0.143 mmol ) . the resulting solution was allowed to come to room temperature and stirred for 16 h. the reaction was evaporated to dryness . the crude product was dissolved and partitioned between dichloromethane and 1 m hcl , then washed with sat . sodium hydrogen carbonate , water , and brine , dried over anhydrous sodium sulfate , and concentrated in vacuum . the crude product was purified by column chromatography on silica gel ( cyclohexane / ethyl acetate 8 : 1 , v / v ) .
a solution of aminol ( 0.145 mmol ) in 5 ml pyridine was placed in an ice bath and treated with myristoyl chloride ( 0.143 mmol ) . the resulting solution was allowed to come to room temperature and stirred for 16 h. the reaction was evaporated to dryness . the crude product was dissolved and partitioned between dichloromethane and 1 m hcl , then washed with sat . sodium hydrogen carbonate , water , and brine , dried over anhydrous sodium sulfate , and concentrated in vacuum . the crude product was purified by column chromatography on silica gel ( cyclohexane / ethyl acetate 8 : 1 , v / v ) . general procedure for the preparation of urea inhibitors dp25 a , cto a stirred solution of aminol ( 0.188 mmol ) in 5 ml acetonitrile and 5 ml chloroform was added tetradecyl isocyanate ( 0.376 mmol ) . the mixture was stirred at room temperature for 12 h. the reaction mixture was evaporated to dryness . the crude product was purified by column chromatography on silica gel ( ch2cl2/methanol 20 : 1 , v / v ) .
to a stirred solution of aminol ( 0.188 mmol ) in 5 ml acetonitrile and 5 ml chloroform was added tetradecyl isocyanate ( 0.376 mmol ) . the mixture was stirred at room temperature for 12 h. the reaction mixture was evaporated to dryness . the crude product was purified by column chromatography on silica gel ( ch2cl2/methanol 20 : 1 , v / v ) . 1-(3-methoxyphenyl)-2-nitroethanol 2b300 mg ( 2.21 mmol ) aldehyde 1b gave 355 mg ( 1.99 mmol , 90% ) of a pale yellow liquid , c9h11no4 ( 197 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 2.87 ( bs , 1h , oh ) , 3.83 ( s , 3h , ch3 ) , 4.51 ( dd , j1 = 3.2 hz , j2 = 13.3 hz , 1h ; ch2 ) , 4.60 ( dd , j1 = 9.4 hz , j2 = 13.3 hz , 1h , ch2 ) , 5.43 ( dd , j1 = 3.2 hz , j2 = 9.4 hz , 1h ; ch ) , 6.90 ( ddd , j1 = 1.1 hz , j2 = 2.5 hz , j3 = 8.3 hz , 1h ; charom . ) , 6.97 ( m , 2h ; charom ) , 7.32 ( dd , j1 = 8.1 hz , j2 = 8.1 hz , 1h ; charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 55.4 , 70.9 , 81.2 , 111.5 , 114.4 , 118.1 , 130.1 , 139.8 , 160.0 ppm . hr - ms p esi calculated for c9h11no4 h m / z = 196.0609 , found : 196.0615 . 300 mg ( 2.21 mmol ) aldehyde 1b gave 355 mg ( 1.99 mmol , 90% ) of a pale yellow liquid , c9h11no4 ( 197 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 2.87 ( bs , 1h , oh ) , 3.83 ( s , 3h , ch3 ) , 4.51 ( dd , j1 = 3.2 hz , j2 = 13.3 hz , 1h ; ch2 ) , 4.60 ( dd , j1 = 9.4 hz , j2 = 13.3 hz , 1h , ch2 ) , 5.43 ( dd , j1 = 3.2 hz , j2 = 9.4 hz , 1h ; ch ) , 6.90 ( ddd , j1 = 1.1 hz , j2 = 2.5 hz , j3 = 8.3 hz , 1h ; charom . ) , 6.97 ( m , 2h ; charom ) , 7.32 ( dd , j1 = 8.1 hz , j2 = 8.1 hz , 1h ; charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 55.4 , 70.9 , 81.2 , 111.5 , 114.4 , 118.1 , 130.1 , 139.8 , 160.0 ppm . hr - ms p esi calculated for c9h11no4 h m / z = 196.0609 , found : 196.0615 . 2-nitro-1-(pyridin-3-yl)-ethanol 2c200 mg ( 1.86 mmol ) aldehyde 1c gave 250 mg ( 1.48 mmol , 79.6% ) of brown oil , c7h8n2o3 ( 170 g / mol ) . h - nmr ( cdcl3 ; 300,1 mhz ) : = 4.52 ( dd , j1 = 3.6 hz , j2 = 12.8 hz , 1h , ch2 ) , 4.61 ( dd , j1 = 9.3 hz , j2 = 12.8 hz , 1h , ch2 ) , 5.50 ( dd , j1 = 3.6 hz , j2 = 9.3 hz , 1h , ch ) , 5.67 ( bs , 1h , oh ) , 7.33 ( ddd , j1 = 0.6 hz , j2 = 4.9 hz , j3 = 7.9 hz , 1h , charom ) , 7.82 ( td , j1 = 1.7 hz , j2 = 1.7 hz , j3 = 7.9 hz , 1h , charom ) , 8.40 ( dd , j1 = 1.6 hz , j2 = 4.9 hz , 1h , charom ) , 8.48 ( d , j = 2.15 hz , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75,4 mhz ) : = 68.4 , 81.2 , 124.2 , 134.7 , 135.4 , 147.0 , 149.1 ppm . hr - ms p esi calculated for c7h7n2o3 h m / z = 169.0608 , found : 169.0615 . 200 mg ( 1.86 mmol ) aldehyde 1c gave 250 mg ( 1.48 mmol , 79.6% ) of brown oil , c7h8n2o3 ( 170 g / mol ) . h - nmr ( cdcl3 ; 300,1 mhz ) : = 4.52 ( dd , j1 = 3.6 hz , j2 = 12.8 hz , 1h , ch2 ) , 4.61 ( dd , j1 = 9.3 hz , j2 = 12.8 hz , 1h , ch2 ) , 5.50 ( dd , j1 = 3.6 hz , j2 = 9.3 hz , 1h , ch ) , 5.67 ( bs , 1h , oh ) , 7.33 ( ddd , j1 = 0.6 hz , j2 = 4.9 hz , j3 = 7.9 hz , 1h , charom ) , 7.82 ( td , j1 = 1.7 hz , j2 = 1.7 hz , j3 = 7.9 hz , 1h , charom ) , 8.40 ( dd , j1 = 1.6 hz , j2 = 4.9 hz , 1h , charom ) , 8.48 ( d , j = 2.15 hz , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75,4 mhz ) : = 68.4 , 81.2 , 124.2 , 134.7 , 135.4 , 147.0 , 149.1 ppm . hr - ms p esi calculated for c7h7n2o3 h m / z = 169.0608 , found : 169.0615 . 1-(3-methoxyphenyl)-2-aminoethanol 3b
170 mg ( 0.86 mmol ) 2b gave 124 mg ( 0.74 mmol , 86% ) of a brown solid , c9h13no2 ( 167 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 2.743.13 ( m , 2h , ch2 ) , 3.75 ( s , 3h , ch3 ) , 4.98 ( d , j = 7.4 hz , 1h ; ch ) , 6.78 ( dd , j1 = 2.1 hz , j2 = 8.1 hz , 1h ; charom . ) , 6.89 ( m , 2h ; charom ) , 7.21 ( dd , j1 = 7.9 hz , j2 = 7.9 hz , 1h ; charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 49.0 , 60.8 , 74.0 , 111.4 , 113.0 , 118.2 , 129.5 , 144.2 , 159.7 ppm . hr - ms p esi calculated for c9h13no2 h m / z = 168.1019 , found : 168.1015 . 170 mg ( 0.86 mmol ) 2b gave 124 mg ( 0.74 mmol , 86% ) of a brown solid , c9h13no2 ( 167 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 2.743.13 ( m , 2h , ch2 ) , 3.75 ( s , 3h , ch3 ) , 4.98 ( d , j = 7.4 hz , 1h ; ch ) , 6.78 ( dd , j1 = 2.1 hz , j2 = 8.1 hz , 1h ; charom . ) , 6.89 ( m , 2h ; charom ) , 7.21 ( dd , j1 = 7.9 hz , j2 = 7.9 hz , 1h ; charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 49.0 , 60.8 , 74.0 , 111.4 , 113.0 , 118.2 , 129.5 , 144.2 , 159.7 ppm . hr - ms p esi calculated for c9h13no2 h m / z = 168.1019 , found : 168.1015 . 2-amino-1-(pyridin-3-yl)-ethanol 3c132 mg ( 0.79 mmol ) gave 109 mg ( 0.79 mmol , quant . ) of a pale brown solid , which was used without purification , c7h7n2o3 ( 170 g / mol ) . 132 mg ( 0.79 mmol ) gave 109 mg ( 0.79 mmol , quant . ) of a pale brown solid , which was used without purification , c7h7n2o3 ( 170 g / mol ) . 1-(2-hydroxy-2-phenylethyl)-3-tetradecylurea dp25a100 mg ( 0.73 mmol ) 1-phenyl-2-aminoethanol gave 150 mg ( 0.40 mmol , 54.5% ) of a colorless solid , c23h40n2o2 ( 376 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.90 ( t , 3 h , ch3 ) , 1.29 ( s , 22h , ch2 ) , 3.13 ( dd , j1 = 7.1 hz , j2 = 7.1 hz , 2h , nch2 ) , 3.31 ( dd , j1 = 7.6 hz , j2 = 14.4 hz , 1h , ch ) , 5.67 ( bs , 1h , oh ) , 3.55 ( dd , j1 = 2.8 hz , j2 = 14.4 hz , j3 = 7.9 hz , 1h , charom ) , 4.83 ( dd , j1 = 2.7 hz , j2 = 7.5 hz , 1h , charom ) , 5.50 ( 1h , nhurea ) , 5.80 ( 1h , nhurea ) , 7.287.30 ( m , 5h , charom ) ppm . cnmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 26.9 , 29.4 , 29.6 , 29.7 , 30.1 , 31.9 , 40.8 , 48.7 , 74.7 , 125.9 , 127.6 , 128.4 , 142.2 ppm . hr - ms p esi calculated for c23h40n2o2 + h m / z = 377.3163 , found : 377.3163 . 100 mg ( 0.73 mmol ) 1-phenyl-2-aminoethanol gave 150 mg ( 0.40 mmol , 54.5% ) of a colorless solid , c23h40n2o2 ( 376 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.90 ( t , 3 h , ch3 ) , 1.29 ( s , 22h , ch2 ) , 3.13 ( dd , j1 = 7.1 hz , j2 = 7.1 hz , 2h , nch2 ) , 3.31 ( dd , j1 = 7.6 hz , j2 = 14.4 hz , 1h , ch ) , 5.67 ( bs , 1h , oh ) , 3.55 ( dd , j1 = 2.8 hz , j2 = 14.4 hz , j3 = 7.9 hz , 1h , charom ) , 4.83 ( dd , j1 = 2.7 hz , j2 = 7.5 hz , 1h , charom ) , 5.50 ( 1h , nhurea ) , 5.80 ( 1h , nhurea ) , 7.287.30 ( m , 5h , charom ) ppm . cnmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 26.9 , 29.4 , 29.6 , 29.7 , 30.1 , 31.9 , 40.8 , 48.7 , 74.7 , 125.9 , 127.6 , 128.4 , 142.2 ppm . hr - ms p esi calculated for c23h40n2o2 + h m / z = 377.3163 , found : 377.3163 . 1-(2-hydroxy-2-(pyridin-3-yl)ethyl)-3-tetradecylurea dp25c20 mg ( 0.14 mmol ) crude 1-(pyridine-3-yl)-2-aminoethanol gave 15 mg ( 0.040 mmol , 28.4% ) of a colorless solid , c22h39n3o2 ( 377 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.87 ( t , 3 h , ch3 ) , 1.24 ( s , 22h , ch2 ) , 1.45 ( br , 2h , ch2 ) , 2.85 ( br , 2h , nch2 ) , 3.12 ( br , 1h , oh ) 3.303.38 ( br , 1h , ch2 ) , 3.60 ( d , j = 15.4 hz , 1h , ch2 ) , 4.79 ( br , 1h , nhurea ) , 4.89 ( dd , j1 = 2.5 hz , j2 = 6.9 hz , 1h , ch ) , 5.07 ( br , 1h , nhurea ) , 7.31 ( dd , j1 = 3.7 hz , j2 = 8.3 hz , 1h , charom ) , 7.80 ( d , j = 8.5 hz , 1h , charom ) , 8.48 ( br , 1h , charom ) , 8.57 ( br , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.8 , 27.0 , 29.4 , 29.7 , 29.8 , 30.2 , 32.0 , 40.3 , 47.8 , 71.7 , 123.9 , 134.9 , 138.7 , 147.1 , 148.0 , 160.2 ppm . hr - ms p esi calculated for c22h39n3o2 + h m / z = 378.3115 , found : 378.3118 . 20 mg ( 0.14 mmol ) crude 1-(pyridine-3-yl)-2-aminoethanol gave 15 mg ( 0.040 mmol , 28.4% ) of a colorless solid , c22h39n3o2 ( 377 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.87 ( t , 3 h , ch3 ) , 1.24 ( s , 22h , ch2 ) , 1.45 ( br , 2h , ch2 ) , 2.85 ( br , 2h , nch2 ) , 3.12 ( br , 1h , oh ) 3.303.38 ( br , 1h , ch2 ) , 3.60 ( d , j = 15.4 hz , 1h , ch2 ) , 4.79 ( br , 1h , nhurea ) , 4.89 ( dd , j1 = 2.5 hz , j2 = 6.9 hz , 1h , ch ) , 5.07 ( br , 1h , nhurea ) , 7.31 ( dd , j1 = 3.7 hz , j2 = 8.3 hz , 1h , charom ) , 7.80 ( d , j = 8.5 hz , 1h , charom ) , 8.48 ( br , 1h , charom ) , 8.57 ( br , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.8 , 27.0 , 29.4 , 29.7 , 29.8 , 30.2 , 32.0 , 40.3 , 47.8 , 71.7 , 123.9 , 134.9 , 138.7 , 147.1 , 148.0 , 160.2 ppm . hr - ms p esi calculated for c22h39n3o2 + h m / z = 378.3115 , found : 378.3118 . n-(2-hydroxy-2-phenylethyl)-tetradecanamide dp24a100 mg ( 0.73 mmol ) 1-phenyl-2-aminoethanol gave 118 mg ( 0.34 mmol , 46.6% ) of a colorless solid , c22h37no2 ( 347 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.90 ( t , 3 h , ch3 ) , 1.28 ( s , 20h , ch2 ) , 1.57 ( br , 2h , ch2 ) , 2.11 ( t , 2h , nch2 ) , 3.27 ( m , 1h , ch2 ) , 3.61 ( ddd , j1 = 3.2 hz , j2 = 6.7 hz , j3 = 13.9 hz , 1h , ch2 ) , 4.77 ( dd , j1 = 3.1 hz , j2 = 7.9 hz , 1h , ch ) , 6.48 ( br , 1h , nhamide ) , 7.31 ( m , 5h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 25.7 , 29.3 , 29.4 , 29.5 , 29.7 , 31.9 , 36.6 , 47.8 , 73.5 , 125.8 , 127.7 , 128.4 , 141.9 , 174.9 ppm . hr - ms p esi calculated for c22h37no2 + h m / z = 348.2897 , found : 348.2897 . 100 mg ( 0.73 mmol ) 1-phenyl-2-aminoethanol gave 118 mg ( 0.34 mmol , 46.6% ) of a colorless solid , c22h37no2 ( 347 g / mol ) . rf ( ch2cl2/methanol 10% ) = 0.6 h - nmr ( cdcl3 ; 300 mhz ) : = 0.90 ( t , 3 h , ch3 ) , 1.28 ( s , 20h , ch2 ) , 1.57 ( br , 2h , ch2 ) , 2.11 ( t , 2h , nch2 ) , 3.27 ( m , 1h , ch2 ) , 3.61 ( ddd , j1 = 3.2 hz , j2 = 6.7 hz , j3 = 13.9 hz , 1h , ch2 ) , 4.77 ( dd , j1 = 3.1 hz , j2 = 7.9 hz , 1h , ch ) , 6.48 ( br , 1h , nhamide ) , 7.31 ( m , 5h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 25.7 , 29.3 , 29.4 , 29.5 , 29.7 , 31.9 , 36.6 , 47.8 , 73.5 , 125.8 , 127.7 , 128.4 , 141.9 , 174.9 ppm . hr - ms p esi calculated for c22h37no2 + h m / z = 348.2897 , found : 348.2897 . n-(2-hydroxy-2-(3-methoxyphenyl)ethyl)-tetradecanamide dp24b15 mg ( 0.090 mmol ) 1-(3-methoxyphenyl)-2-aminoethanol gave 20 mg ( 0.053 mmol , 58.9% d. th . ) of a white solid , c23h39no4 ( 377 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.90 ( t , 3 h , ch3 ) , 1.27 ( s , 20h , ch2 ) , 1.62 ( br , 2h , ch2 ) , 2.20 ( dd , j1 = 7.6 hz , j2 = 15.2 hz , 1h , nch2 ) , 2.34 ( m , 1h , nch2 ) , 3.82 ( s , 3h , och3 ) 3.87 ( d , j = 8.0 hz , 1h , ch ) , 6.00 ( br , 1h , nhamide ) , 6.857.00 ( m , 3h , charom ) 7.257.32 ( m , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 25.7 , 29.1 , 29.3 , 29.4 , 29.5 , 29.7 , 31.9 , 36.6 , 55.2 , 73.7 , 111.2 , 113.3 , 118.1 , 129.5 , 159.8 ppm . hr - ms p esi calculated for c23h39no3 + h m / z = 378.3003 , found : 378.3002 . 15 mg ( 0.090 mmol ) 1-(3-methoxyphenyl)-2-aminoethanol gave 20 mg ( 0.053 mmol , 58.9% d. th . ) of a white solid , c23h39no4 ( 377 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.90 ( t , 3 h , ch3 ) , 1.27 ( s , 20h , ch2 ) , 1.62 ( br , 2h , ch2 ) , 2.20 ( dd , j1 = 7.6 hz , j2 = 15.2 hz , 1h , nch2 ) , 2.34 ( m , 1h , nch2 ) , 3.82 ( s , 3h , och3 ) 3.87 ( d , j = 8.0 hz , 1h , ch ) , 6.00 ( br , 1h , nhamide ) , 6.857.00 ( m , 3h , charom ) 7.257.32 ( m , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 25.7 , 29.1 , 29.3 , 29.4 , 29.5 , 29.7 , 31.9 , 36.6 , 55.2 , 73.7 , 111.2 , 113.3 , 118.1 , 129.5 , 159.8 ppm . hr - ms p esi calculated for c23h39no3 + h m / z = 378.3003 , found : 378.3002 . n-(2-hydroxy-2-(pyridin-3-yl)-ethyl)tetradecanamide dp24c20.5 mg ( 0.15 mmol ) crude 1-(pyridine-3-yl)-2-aminoethanol gave 37 mg ( 0.106 mmol , 71.6% ) of a colorless solid , c21h36n2o2 ( 348 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.87 ( t , 3 h , ch3 ) , 1.27 ( s , 20h , ch2 ) , 1.58 ( m , 2h , ch2 ) , 2.20 ( m , 2h , coch2 ) , 3.12 ( br , 1h , oh ) 3.40 ( dd , j1 = 7.1 hz , j2 = 14.1 hz , 1h , ch2 ) , 3.69 ( ddd , j1 = 3.1 hz , j2 = 6.4 hz , j3 = 14.2 hz , 1h , ch2 ) , 4.91 ( dd , j1 = 2.6 hz , j2 = 7.0 hz , 1h , ch ) , 5.26 ( br , 1h , nhamide ) , 7.31 ( dd , j1 = 5.0 hz , j2 = 7.4 hz , 1h , charom ) , 7.78 ( d , j = 7.9 hz , 1h , charom ) , 8.48 ( br , 1h , charom ) , 8.54 ( br , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 25.7 , 29.3 , 29.4 , 29.5 , 29.7 , 31.9 , 36.5 , 47.6 , 71.8 , 123.7 , 134.4 , 147.2 , 148.4 , 175.4 ppm . hr - ms p esi calculated for c21h36n2o2 + h m / z = 349.2850 , found : 349.2852 . 20.5 mg ( 0.15 mmol ) crude 1-(pyridine-3-yl)-2-aminoethanol gave 37 mg ( 0.106 mmol , 71.6% ) of a colorless solid , c21h36n2o2 ( 348 g / mol ) . h - nmr ( cdcl3 ; 300 mhz ) : = 0.87 ( t , 3 h , ch3 ) , 1.27 ( s , 20h , ch2 ) , 1.58 ( m , 2h , ch2 ) , 2.20 ( m , 2h , coch2 ) , 3.12 ( br , 1h , oh ) 3.40 ( dd , j1 = 7.1 hz , j2 = 14.1 hz , 1h , ch2 ) , 3.69 ( ddd , j1 = 3.1 hz , j2 = 6.4 hz , j3 = 14.2 hz , 1h , ch2 ) , 4.91 ( dd , j1 = 2.6 hz , j2 = 7.0 hz , 1h , ch ) , 5.26 ( br , 1h , nhamide ) , 7.31 ( dd , j1 = 5.0 hz , j2 = 7.4 hz , 1h , charom ) , 7.78 ( d , j = 7.9 hz , 1h , charom ) , 8.48 ( br , 1h , charom ) , 8.54 ( br , 1h , charom ) ppm . c - nmr ( cdcl3 ; 75 mhz ) : = 14.1 , 22.7 , 25.7 , 29.3 , 29.4 , 29.5 , 29.7 , 31.9 , 36.5 , 47.6 , 71.8 , 123.7 , 134.4 , 147.2 , 148.4 , 175.4 ppm . hr - ms p esi calculated for c21h36n2o2 + h m / z = 349.2850 , found : 349.2852 . briefly , assays were performed in the presence of sodium acetate buffer ( 250 mm , ph 4.0 ) , 5 mm edta , 0.7% ( w / v ) triton x-100 , 0.4% ( w / v ) tween 20 , 0.4% ( w / v ) igepal ca-630 , and 17 nmol/50 l n - lauroyl - d - erythro - sphingosine . the activity was determined with 60 l of buffer , 35 l undiluted enzyme solution ( 43,7 g / ml ) , and 5 l of 50 m inhibitor solution in dmso which gave 100 l total volume with 2.5 m inhibitor concentration . tests were incubated for 30 min at 37c , the reactions terminated with 800 l chloroform / methanol 2 : 1 ( v / v ) and 200 l saturated aqueous ammonium carbonate solution , and c14- and c16-sphinganin in methanol ( 500 pm of each ) were added . afterwards the mixtures were centrifuged at 10000 g for 5 min , the chloroform phase was dried and redissolved in 50 l ethanol and incubated 10 min at 50c . samples were diluted with 0.9 ml hplc buffer ( acetonitrile/2 mm kh2po4 , ph 7.0 , 9/1 ( v / v ) ) and 0.1 ml 5 mm kh2po4 buffer ( ph 7.0 , 90 : 10 , v / v ) . aliquots were injected immediately onto an rp - c18 polaris a 5 m ( pn a 2000 - 250100)column by varian . lipids were eluted with mixtures out of water containing 1% acetonitrile and acetonitrile containing 1% water with a flow of 1 ml / min . excitation wavelength is = 355 nm , emission wavelength is = 435 nm . for ic50 determination , | the lipid - signalling molecule ceramide is known to induce apoptosis in a variety of cell types .
inhibition of the lysosomal acid ceramidase can increase cellular ceramide levels and thus induce apoptosis .
indeed , inhibitors of acid ceramidase have been reported to induce cell death and to display potentiating effects to classical radio- or chemo therapy in a number of in vitro and in vivo cancer models .
the most potent in vitro inhibitor of acid ceramidase , b-13 , recently revealed to be virtually inactive towards lysosomal acid ceramidase in living cells .
in contrast , a number of weakly basic b-13 analogues have been shown to accumulate in the acidic compartments of living cells and to efficiently inhibit lysosomal acid ceramidase .
however , introduction of weakly basic groups at the -position of the fatty acid moiety of b-13 led to a significant reduction of potency towards acid ceramidase from cellular extracts .
herein , we report a novel b-13-derived scaffold for more effective inhibitors of acid ceramidase .
furthermore , we provide hints for an introduction of basic functional groups at an alternative site of the b-13 scaffold that do not interfere with acid ceramidase inhibition in vitro . |
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metabolic syndrome comprises a cluster of components of impaired glucose metabolism , abdominal fat accumulation , dyslipidemia , and elevated blood pressure . each component has been shown to be an independent risk factor for cardiovascular diseases ( cvd ) in japanese community cohort studies : impaired fasting glucose ; abdominal obesity [ 3 , 4 ] ; low - density lipoprotein cholesterol [ 5 , 6 ] ; and high - normal blood pressure [ 79 ] . recently , a new streamlined definition of metabolic syndrome for global populations has been introduced . in this definition , however , in the japanese definition of metabolic syndrome , abdominal obesity is an essential component . the definition may mislead a prevention of cvd for non - obesity metabolic disorder , which is a modifiable risk factor for cvd . changes in lifestyle are the key to improving metabolic disorder , including diabetes mellitus ( dm ) and dyslipidemia , and thereby to reducing the risk of cardiovascular disease . in this review paper , the focus is on the relation of metabolic symptoms , namely impaired glucose metabolism and dyslipidemia to cvd in the japanese population . diabetes mellitus has become a major public health problem [ 12 , 13 ] as well as a risk factor for mortality and cvd [ 1416 ] . in japan , the frequencies of hyperglycemia for 1961 , 1974 , and 1989 show a trend of increasing incidence : 12.1% , 13.8% , and 31.9% in men and 4.8% , 8.1% , and 27.2% in women , respectively . previous meta - analysis studies have shown that diabetes is a risk factor for ischemic stroke and coronary artery disease [ 15 , 18 , 19 ] . the framingham offspring cohort study showed a positive relationship between impaired fasting glucose and coronary heart disease in women and between diabetes and coronary heart disease in men and women . these results are compatible with those of previous cohort studies in japan , as described below and summarized in table 1 . table 1association between blood glucose categories and cardiovascular diseases in japanese cohort studiesstudy namenumbersexfollow - upend pointresultsreferencethe suita study5321mf11.5strokedm , hr = 2.08chdifg , hr = 1.46 ; dm , hr = 2.28mstrokedm , hr = 1.78fstrokedm , hr = 2.66chdifg , hr = 1.83 ; dm , hr = 4.32the hisayama study2421m14cidm , hr = 2.15 ; 2hpg , hr = 2.71fcidm , hr = 2.10 ; 2hpg , hr = 2.19chddm , hr = 3.83 ; 2hpg , hr = 4.44five japanese communities4287m17cidm , hr = 1.8lacunar infarctiondm , hr = 2.16295f17cidm , hr = 2.2lacunar infarctiondm , hr = 2.410582mf17non - embolic ciborderline with non - hypertension , hr = 1.7 dm : with non - hypertension , hr = 1.7 dm with higher bmi , hr = 2.2jphc study31,192mf12.9chdborderline hr = 1.5 dm , hr = 2.38nippon data809444mf17.3all causes mortalitycbg>=11.1 mmol / l , hr=1.63cvd mortality5.22 mmol / l<=cbg<7.77 mmol / l , hr = 2.62the funagata diabetes study2534mf5.7all cause mortalityada 2007 : dm , hr = 2.11cvd mortalitywho 1985 : igt , hr = 2.3 ; dm , hr = 3.54ada 2007 : dm , hr = 3.17 m men ; f women ; chd coronary heart disease ; ci cerebral infarction ; cvd cardiovascular diseases ; dm diabetes mellitus ; ifg impaired fasting glucose ; 2hpg 2 h post - loaded glucose levels ; hr hazard ratio ; bmi body mass index ; cbg casual blood glucose ; ada american diabetes association ; who world health organization . association between blood glucose categories and cardiovascular diseases in japanese cohort studies m men ; f women ; chd coronary heart disease ; ci cerebral infarction ; cvd cardiovascular diseases ; dm diabetes mellitus ; ifg impaired fasting glucose ; 2hpg 2 h post - loaded glucose levels ; hr hazard ratio ; bmi body mass index ; cbg casual blood glucose ; ada american diabetes association ; who world health organization . the hisayama study indicated that diabetes as defined by the glucose tolerance test was a risk factor for ischemic stroke in men ( hazard ratio 2.54 ; 95% confidence intervals 1.40 to 4.63 ) and women ( hazard ratio 2.02 ; 95% confidence intervals 1.07 to 3.81 ) and coronary heart disease in women ( hazard ratio 3.46 ; 95% confidence intervals 1.59 to 7.54 ) ; however , impaired glucose tolerance and impaired fasting glucose were not risk factors for ischemic stroke or coronary heart disease . in a study of five communities in japan , diabetes defined by non - fasting glucose levels was a risk factor for non - embolic ischemic stroke in men ( hazard ratio 1.8 ; 95% confidence intervals 1.0 to 3.2 ) and women ( hazard ratio 2.2 ; 95% confidence intervals 1.2 to 4.0 ) . the positive association was particularly strong in hypertensive subjects with higher skin - fold thickness values ( hazard ratio 1.9 and 95% confidence intervals 1.0 to 3.7 for borderline diabetes ; hazard ratio 4.9 and 95% confidence intervals 2.5 to 9.5 for diabetes ) . a positive association between diabetes and cvd mortality the nippon data 80 study indicated that high and borderline - high casual blood glucose groups ( 11.1 mmol / l and 7.77 to 11.1 mmol / l , respectively ) had increased risks of coronary heart disease mortality . the funagata diabetes study showed that diabetes defined by both the who criteria ( 1985 ) and ada recommendations ( 1997 ) were risk factors for all - cause mortality and cardiovascular mortality . however , impaired fasting glucose was not a risk factor for all - cause mortality and cardiovascular mortality . these japanese cohort studies for mortality risks indicated that diabetes is a risk factor for all - cause and cvd mortality . however , impaired fasting glucose and impaired glucose tolerance may not be risk factors for all - cause or cvd mortality . recently , the suita study showed that impaired fasting glucose is a risk factor for the incidence of cardiovascular disease ( hazard ratio 1.49 ; 95% confidence intervals 1.02 to 2.16 ) or coronary heart disease ( hazard ratio 1.83 ; 95% confidence intervals 1.01 to 3.32 ) in women , and that diabetes is a risk factor for stroke in both men ( hazard ratio 1.78 ; 95% confidence intervals 1.00 to 3.12 ) and women ( hazard ratio 2.66 ; 95% confidence intervals 1.22 to 5.80 ) and for coronary heart disease in women ( hazard ratio 1.78 ; 95% confidence intervals 1.00 to 3.12 ) . in addition , compared with normoglycemic and optimal blood pressure japanese subjects , increased risks of cvd were observed in normoglycemic subjects with high - normal blood pressure or hypertension , impaired fasting glucose subjects with normal or higher blood pressure , and diabetic subjects regardless of blood pressure category ( fig . 1 : p - value for interaction=0.046 ) . 1multivariable - adjusted hazard ratios of cardiovascular diseases according to the combination of blood pressure and glucose categories . multivariable - adjusted hazard ratios were adjusting for age , sex , body mass index , smoking , drinking , and hyperlipidemia . bp blood pressure ; ngt normal glycemic tolerance ; ifg impaired fasting glucose ; dm diabetes mellitus ; * : p < 0.05 ( compared with the optimal blood pressure and normoglycemic group ) multivariable - adjusted hazard ratios of cardiovascular diseases according to the combination of blood pressure and glucose categories . multivariable - adjusted hazard ratios were adjusting for age , sex , body mass index , smoking , drinking , and hyperlipidemia . bp blood pressure ; ngt normal glycemic tolerance ; ifg impaired fasting glucose ; dm diabetes mellitus ; * : p < 0.05 ( compared with the optimal blood pressure and normoglycemic group ) in a collaborative meta - analysis of 102 prospective studies from around the world , increased risks of coronary heart disease were observed in unknown diabetic subjects ( i.e. , subjects with diabetes who are not aware of their disease ) with 5.6 to < 6.1 mmol / l and 6.1 to < 7 mmol / l ( hazard ratios [ 95% confidence intervals ] : 1.11 [ 1.04 to 1.18 ] and 1.17 [ 1.08 to 1.26 ] , respectively ) . quite recently , the japan public health center - based prospective ( jphc ) study consisting of a 12.9-year of follow - up for 31,192 individuals aged 4069 years was observed that diabetes mellitus and hyperglycemia carried an increased risk for coronary heart disease in a japanese general population . based on this study and the suita study [ 2 , 25 ] , both diabetes mellitus and impaired fasting glucose may be a risk factor for the incidence of coronary heart disease in japan . in the jphc study s systematic review of the evidence for pre - diabetes , impaired fasting glucose and impaired glucose tolerance were shown to be associated with modest increases in the risk for cvd . overall relative risks ( 95% confidence intervals ) for the association between two categories of impaired fasting glucose ( 100 to 125 mg / dl and 110 to 125 mg / dl ) and cvd were 1.18 ( 1.09 to 1.28 ) and 1.20 ( 1.12 to 1.28 ) , respectively . meanwhile , the risks of cvd for impaired fasting glucose and impaired glucose tolerance were 1.10 ( 0.98 to 1.23 ) and 1.20 ( 1.07 to 1.34 ) , respectively . in two japanese cohort studies using the oral glucose tolerance test , impaired fasting glucose was not found to pose a risk of cvd [ 21 , 24 ] . however , in one of these cohort studies , impaired glucose tolerance was observed as a risk of cvd mortality . larger japanese cohort studies are required to establish the association between oral glucose tolerance test results and cvd . in order to prevent cvd , a person with impaired glucose metabolism should eliminate or reduce other cardiovascular risk factors , such as high blood pressure , obesity , smoking , and excess drinking . the suita study showed that the combination of impaired fasting glucose and high blood pressure greatly increased the risk of cardiovascular disease . recently , the frequencies of obesity in men and elderly women have increased in japan . obesity is a risk factor for impaired glucose metabolism ; weight loss reduces the risk of both impaired glucose metabolism and cvd . the average smoking rate in men worldwide has rapidly decreased in recent years , and in 2005 had dropped by 45.5% since its peak . however , the rate in japan is still higher than that in western countries . and while , the smoking rate in women is generally lower than that in men . in japan , the rate for younger women aged 20 to 39 years has increased to around 17 to 20% . excess drinking is a risk factor for increasing blood pressure and stroke . in order to reduce the risk of cvd , subjects with a combination of two borderline risks , high - normal blood pressure and hyperglycemia , should modify their lifestyles , aiming at cessation of smoking and even moderate drinking , as well as weight reduction if there are overweight or obese . according to japan s national survey on circulatory disorders in 1980 , 1990 , and 2000 , mean levels of total cholesterol increased from 186 mg / dl and 191 mg / dl in 1980 to 200 mg / dl and 208 mg / dl in 2000 among adult men and women , respectively . frequencies of hypercholesterolemia ( total cholesterol levels of 220 mg / dl or more ) increased 15% and 19% in 1980 to 27% and 35% in 2000 among men and women , respectively . the difference in mean total cholesterol levels between japanese and american men was approximately 40 mg / dl in the 1980s . however , this difference had diminished to 15 mg / dl by 1990 and 2000 . no changes in mean total cholesterol levels in japanese or american men aged 60 years and over were observed from 1990 to 2000 . a previous meta - analytic study has shown that total cholesterol was positively associated with ischemic heart disease in both middle and old age and at all blood pressure categories . in contrast , the association between total cholesterol and the risk of stroke remains unclear . the multiple risk factor intervention trial has shown that the risk of death from intracerebral hemorrhage was three times higher in men with serum cholesterol levels under 160 mg / dl than in those with higher cholesterol levels , whereas a positive association was observed between the serum cholesterol level and death from ischemic stroke . a meta - analysis of 45 prospective cohort studies showed no association between blood cholesterol levels and stroke except in those under 45 years of age when screened . dyslipidemia may be important for some subtypes of stroke but not for others , because stroke is a heterogeneous disease of various etiologic origins . in american men , an inverse association was observed between the total cholesterol levels and the incidence of intracerebral hemorrhage , while a positive association was observed between total cholesterol levels and the incidence of cerebral infarction . japanese cohort studies have shown evidence of the positive association of total cholesterol with coronary heart disease similar to that in western studies . nippon data80 showed associations between total cholesterol and risk of all - cause mortality ( hazard ratio 1.19 and 1.36 ; 95% confidence intervals , 1.03 to 1.37 and 1.05 to 1.77 ) in both the lowest ( < 160 mg / dl ) and highest ( 260 mg / dl ) total cholesterol groups , respectively . in addition , the hazard ratios of coronary heart disease mortality were 1.4 in the 200 to 219 mg / dl total cholesterol group , 1.7 in the 220 to 239 mg / dl group , 1.8 in the 240 to 259 mg / dl groups , and 3.8 in the 260 mg / dl or above group , compared with the 160 to 179 mg / dl total cholesterol ( healthy control ) group . the suita study showed a positive association between serum low - density lipoprotein ( ldl ) cholesterol and non - high - density lipoprotein ( non - hdl ) cholesterol levels and increased incidence of myocardial infarction , but not with any type of stroke . the hazard ratio for myocardial infarction was highest in the top quintile of ldl cholesterol ( hazard ratio , 3.03 ; 95% confidence intervals , 1.32 to 6.96 ) when men and women were combined . the hazard ratio for myocardial infarction was also highest in the top quintile of non - hdl cholesterol ( hazard ratio , 2.97 ; 95% confidence intervals , 1.26 to 6.97 ) . there was no substantial difference in the predictive value for the incidence of myocardial infarction between ldl cholesterol and non - hdl cholesterol . ldl cholesterol can be calculated from fasting blood sample and measurement of total cholesterol , hdl cholesterol , and triglyceride levels from fasting blood samples , according to the friedewald formula . the formula is not applicable for serum triglyceride levels equal to or greater than 400 mg / dl . however , non - hdl cholesterol levels can be easily calculated by routine measured parameters , total and hdl cholesterol without the effect of non - fasting status or hypertriglyceridemia ( 400 mg / dl ) . the hisayama study showed that the positive association between ldl cholesterol levels and risk of atherothrombotic infarction ( p for trend=0.02 ) and coronary heart disease ( p for trend=0.03 ) remained significant after multivariable adjustment . in the ibaraki prefectural health study , which was a very large sample consisting of 30,802 men and 60,417 women , high ldl cholesterol levels were associated with an increased risk of mortality from coronary heart disease in men , but not in women . the same study showed that lower ldl cholesterol levels are associated with elevated risk of mortality from intracerebral hemorrhage , but are not associated with increasing risk of cerebral infarction . these japanese cohort studies suggest that higher levels of ldl cholesterol pose an increased risk of coronary heart disease and possibly atherothrombotic infarction , whereas lower levels of ldl cholesterol may increase the risk of intracerebral hemorrhage . recently , the lectin - like oxidized ldl receptor 1 ( lox-1 ) has been implicated in atherothrombotic diseases . activation of lox-1 in humans can be evaluated by use of the lox index , obtained by multiplying the circulating concentration of lox-1 ligands containing apolipoprotein b by that of the soluble form of lox-1 . in the suita study , lower hdl cholesterol predicts coronary heart disease mortality and occurrence of new coronary heart disease events . elevated total cholesterol was not found to be associated with coronary heart disease mortality in older men , but may be a risk factor for coronary heart disease in older women . the israeli ischemic heart disease study showed an independent negative association between hdl cholesterol and ischemic stroke mortality during 21 years of follow - up . among the japanese cohort studies , the oyabe study demonstrated that lower hdl cholesterol levels were related significantly and independently to an increased risk of all - stroke incidence and ischemic stroke incidence . and in a combined prospective cohort study of 13 urban industrial companies , coronary heart disease incidence was inversely related to hdl cholesterol in japanese men . based on combined data from prospective studies , a high serum triglyceride level is a risk factor for cvd for both men ( hazard ratio , 1.32 ; 95% confidence intervals , 1.26 to 1.39 ) and women ( hazard ratio , 1.76 ; 95% confidence intervals , 1.50 to 2.07 ) in the general population , independent of hdl cholesterol . in 26 prospective studies in asian and pacific populations , in 29 prospective meta - analytic studies in the west , triglyceride levels were moderately associated with coronary heart disease . in japanese cohort studies , two studies have provided evidence regarding the association between serum triglyceride levels and coronary heart disease and ischemic stroke . a cohort study of four rural communities showed that a high serum triglyceride level was a risk factor for coronary heart disease ; even after adjustment for hdl cholesterol levels the significant association remained , although 80% of the baseline participants were non - fasting . the suita study has also shown that the risk for ischemic stroke was highest in participants with high triglycerides alone , and that a combination of high serum levels of triglyceride and non - hdl cholesterol was associated with an increased risk of myocardial infarction . high serum levels of triglyceride and non - hdl cholesterol are both important targets for the prevention of cvd , which requires evidence - based guidelines for management in the primary care setting . a previous meta - analytic study has shown that total cholesterol was positively associated with ischemic heart disease in both middle and old age and at all blood pressure categories . in contrast , the association between total cholesterol and the risk of stroke remains unclear . the multiple risk factor intervention trial has shown that the risk of death from intracerebral hemorrhage was three times higher in men with serum cholesterol levels under 160 mg / dl than in those with higher cholesterol levels , whereas a positive association was observed between the serum cholesterol level and death from ischemic stroke . a meta - analysis of 45 prospective cohort studies showed no association between blood cholesterol levels and stroke except in those under 45 years of age when screened . dyslipidemia may be important for some subtypes of stroke but not for others , because stroke is a heterogeneous disease of various etiologic origins . in american men , an inverse association was observed between the total cholesterol levels and the incidence of intracerebral hemorrhage , while a positive association was observed between total cholesterol levels and the incidence of cerebral infarction . japanese cohort studies have shown evidence of the positive association of total cholesterol with coronary heart disease similar to that in western studies . nippon data80 showed associations between total cholesterol and risk of all - cause mortality ( hazard ratio 1.19 and 1.36 ; 95% confidence intervals , 1.03 to 1.37 and 1.05 to 1.77 ) in both the lowest ( < 160 mg / dl ) and highest ( 260 mg / dl ) total cholesterol groups , respectively . in addition , the hazard ratios of coronary heart disease mortality were 1.4 in the 200 to 219 mg / dl total cholesterol group , 1.7 in the 220 to 239 mg / dl group , 1.8 in the 240 to 259 mg / dl groups , and 3.8 in the 260 mg / dl or above group , compared with the 160 to 179 mg / dl total cholesterol ( healthy control ) group . the suita study showed a positive association between serum low - density lipoprotein ( ldl ) cholesterol and non - high - density lipoprotein ( non - hdl ) cholesterol levels and increased incidence of myocardial infarction , but not with any type of stroke . the hazard ratio for myocardial infarction was highest in the top quintile of ldl cholesterol ( hazard ratio , 3.03 ; 95% confidence intervals , 1.32 to 6.96 ) when men and women were combined . the hazard ratio for myocardial infarction was also highest in the top quintile of non - hdl cholesterol ( hazard ratio , 2.97 ; 95% confidence intervals , 1.26 to 6.97 ) . there was no substantial difference in the predictive value for the incidence of myocardial infarction between ldl cholesterol and non - hdl cholesterol . ldl cholesterol can be calculated from fasting blood sample and measurement of total cholesterol , hdl cholesterol , and triglyceride levels from fasting blood samples , according to the friedewald formula . the formula is not applicable for serum triglyceride levels equal to or greater than 400 mg / dl . however , non - hdl cholesterol levels can be easily calculated by routine measured parameters , total and hdl cholesterol without the effect of non - fasting status or hypertriglyceridemia ( 400 mg / dl ) . the hisayama study showed that the positive association between ldl cholesterol levels and risk of atherothrombotic infarction ( p for trend=0.02 ) and coronary heart disease ( p for trend=0.03 ) remained significant after multivariable adjustment . in the ibaraki prefectural health study , which was a very large sample consisting of 30,802 men and 60,417 women , high ldl cholesterol levels were associated with an increased risk of mortality from coronary heart disease in men , but not in women . the same study showed that lower ldl cholesterol levels are associated with elevated risk of mortality from intracerebral hemorrhage , but are not associated with increasing risk of cerebral infarction . these japanese cohort studies suggest that higher levels of ldl cholesterol pose an increased risk of coronary heart disease and possibly atherothrombotic infarction , whereas lower levels of ldl cholesterol may increase the risk of intracerebral hemorrhage . recently , the lectin - like oxidized ldl receptor 1 ( lox-1 ) has been implicated in atherothrombotic diseases . activation of lox-1 in humans can be evaluated by use of the lox index , obtained by multiplying the circulating concentration of lox-1 ligands containing apolipoprotein b by that of the soluble form of lox-1 . in the suita study , lower hdl cholesterol predicts coronary heart disease mortality and occurrence of new coronary heart disease events . elevated total cholesterol was not found to be associated with coronary heart disease mortality in older men , but may be a risk factor for coronary heart disease in older women . the israeli ischemic heart disease study showed an independent negative association between hdl cholesterol and ischemic stroke mortality during 21 years of follow - up . among the japanese cohort studies , the oyabe study demonstrated that lower hdl cholesterol levels were related significantly and independently to an increased risk of all - stroke incidence and ischemic stroke incidence . and in a combined prospective cohort study of 13 urban industrial companies , coronary heart disease incidence was inversely related to hdl cholesterol in japanese men . based on combined data from prospective studies , a high serum triglyceride level is a risk factor for cvd for both men ( hazard ratio , 1.32 ; 95% confidence intervals , 1.26 to 1.39 ) and women ( hazard ratio , 1.76 ; 95% confidence intervals , 1.50 to 2.07 ) in the general population , independent of hdl cholesterol . in 26 prospective studies in asian and pacific populations , in 29 prospective meta - analytic studies in the west , triglyceride levels were moderately associated with coronary heart disease . in japanese cohort studies , two studies have provided evidence regarding the association between serum triglyceride levels and coronary heart disease and ischemic stroke . a cohort study of four rural communities showed that a high serum triglyceride level was a risk factor for coronary heart disease ; even after adjustment for hdl cholesterol levels the significant association remained , although 80% of the baseline participants were non - fasting . the suita study has also shown that the risk for ischemic stroke was highest in participants with high triglycerides alone , and that a combination of high serum levels of triglyceride and non - hdl cholesterol was associated with an increased risk of myocardial infarction . high serum levels of triglyceride and non - hdl cholesterol are both important targets for the prevention of cvd , which requires evidence - based guidelines for management in the primary care setting . this paper reviews the associations of impaired glucose metabolism and dyslipidemia with cvd in japanese cohort studies . impaired fasting glucose and high - normal blood pressure were shown to be independent risk factors for cvd and coronary heart disease in an urban cohort . impaired glucose tolerance has not been observed as a risk factor for the incidence of cvd in japan . the japanese evidence for the positive association of total cholesterol with coronary heart disease is similar to that of previous western studies . associations with all - cause mortality were observed for both the lower and higher levels of cholesterol : higher levels of ldl cholesterol have been shown to increase the risk of coronary heart disease and atherothrombotic infarction , whereas lower levels of ldl cholesterol may increase the risk of intracerebral hemorrhage in japan , as elsewhere . positive associations between serum triglyceride levels and coronary heart disease and ischemic stroke have also been observed in japanese populations . the japanese diet has been rapidly changing in recent decades , as reflected in many of its health indicators such as cholesterol levels , and both impaired glucose metabolism and dyslipidemia are emerging as important risk factors for cvd in the japanese population . in order to reduce the risk of cvd , subjects with metabolic disorder should reduce other cardiovascular risk factors and improve their lifestyle . | metabolic disorder is a modifiable risk factor for cardiovascular diseases ( cvd ) , and lifestyle modification is the key to improving metabolic disorder .
diabetes mellitus has been shown to be a risk factor for coronary heart disease ( chd ) and ischemic stroke in both western and japanese populations .
an association between impaired fasting glucose and pre - hypertension found in an urban japanese population emphasized the combined risk of cvd .
mean total cholesterol levels in japan have been increasing in the last three decades .
the japanese evidence for the positive association of total cholesterol with chd is similar to that in the west .
higher low - density lipoprotein cholesterol ( ldl - c ) levels pose an increased risk of chd and atherothrombotic infarction , whereas lower ldl - c levels may pose an increased risk of intracerebral hemorrhage in japan .
overall , the studies reviewed here show that impaired glucose metabolism and dyslipidemia are emerging risk factors for cvd in the japanese population . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
anterior cruciate ligament ( acl ) injuries are one of the most common and devastating knee
injuries associated with sport participation1 . surgical repair of complete rupture of the acl is recommended to
prevent knee joint instability . while surgery corrects the structural defect , post - operative
impairments in knee strength , function and endurance are inevitable corollary problems
associated to the repair . many patients experience some degree of muscle weakness post - acl
reconstruction ( aclr ) , especially weakness of the quadriceps muscles , which can limit their
functional ability2 . palmieri - smith et
al.3 assessed the deficit in quadriceps
muscle strength between the reconstructed and non - injured knees in patients six months after
aclr . several studies have suggested use of an accelerated progression of knee strength ,
function and endurance following aclr to improve outcomes4 , 5.while most recent studies
have focused on the effects of postoperative rehabilitation protocols , only a few studies
exist that have studied about effects of preoperative exercise on post - aclr outcomes6,7,8 . keays et al.7 reported on the beneficial effects of a 5-week home - based exercise
program for improving quadriceps strength and knee function early post - aclr injury . based on
available research evidence , if a preoperative program of exercise can restore muscle
strength and function before surgery , it could be of benefit to improve outcomes
post - surgery . based on the study eitzen et al.9 the preoperative quadriceps strengthening may be a significant
predictor of knee function two year after aclr9 . preoperative deficits in quadriceps strength of greater than 20%
have been associated with significant persistent muscle weakness two years after aclr .
however , there is still little evidence regarding the benefits of preoperative exercise on
improving postoperative knee weakness . we hypothesized that a preoperative program of exercise would improve functional outcomes
post - aclr . therefore , the aim of the present study was to examine the effects of a 4 week ,
hospital - based preoperative program of exercise , focusing on quadriceps strengthening and
single - leg hop function , on after aclr outcomes . a prospective , randomized , controlled study was initiated to evaluate the effectiveness of
a 4 weeks preoperative program of exercises on after aclr knee strength and function . patients with acl rupture were recruited from samsung medical orthopedics centers from april
2012 to august 2014 . the inclusion criteria were men between the ages of 20 and 35 years ,
presenting with an isolated acl rupture in the knee . the exclusion criteria were previous
aclr or meniscus repair , injury to other ligaments in the same knee , and any associated
fractures . the 80 patients were randomly assigned into two groups , the preoperative exercise
group ( peg , n=40 ) and no preoperative exercise group ( npeg , n=40 ) . the mean height and body weight peg were 173.85.6 cm and 75.28.9 kg ; for the npeg , they were 172.84.4 cm and
74.319.77 kg . there were no significant between - group differences in height and weight
between groups . the peg participated in a 4-week exercise program preoperatively and in a 12 week
postoperative program , while the npeg only participated in the 12 weeks postoperative
program . knee muscle strength and function were measured at 4 weeks before surgery and 3
months after surgery . all patients were supervised at least three times a week in a sports
medicine center . the preoperative exercise program was based on known impairments associated with acl
injuries , focused mainly on strengthening , functional balance , muscle control , and
co - contraction7 . the exercise program mainly focused
on lower limb strengthening , with particular attention paid to strengthening of the
quadriceps muscles . prior to performing the main strength training program , after completion of the warm - up , subjects
proceeded to a series of lower - body strengthening exercises , short arc extension , wall
squat , leg press , isometric leg extension , and leg curls as well as other exercises ( table 1table 1.a summary of the preop exercise programexercise categoryexerciserepetitions and setscardio exercisestationary bike 20 minrange of motion exerciseseat flexion / extension10 minwall slidesstrengthening exerciseopen - chain strengtheningshort arc extension 102straight leg raises102leg curl ( isometric)102close - chain strengtheningleg extension ( rom 3080)102leg press152half squat152balance / proprioceptionsingle - leg standing30 sec3balance board30 sec3 ) . selected exercises reduced anterior shear force across the tibio - femoral
joint and , therefore , helped strengthen the muscles without risk of damaging intra - articular
structures of the knee . the postoperative exercise program was based on a combination of
evidence - based exercises identified from the scientific literature . immediately after aclr , the limb was immobilized in a postoperative functional brace , and
patients were instructed to perform straight leg raising and quadriceps setting exercises . patients were allowed to undergo partially weight bearing at 2 weeks after their surgery and
to move through full - range of knee joint motion and perform closed chain exercise at 4 weeks
after surgery . all patients underwent assessment of knee extensor strength and distance on
single - leg hop 4 weeks prior to surgery and again at 3 months after surgery . isokinetic
strength and endurance were measured using a csmi isokinetic dynamometer ( csmi medical
solution , stoughton , ma , usa ) . knee strength was measured through the range of 0 to 90 at
an angular speed of 60/sec , with four repetitions completed at an angular speed of
180/sec , with 20 repetitions completed to calculate average power10 . the highest peak torque value for each velocity was
compared with the uninjured side and described as percent of strength deficit . for the
single - leg hop for test , the mean average distance was quantified by limb symmetry index
( lsi ) using the following formula : ( distance for injured leg / distance for uninjured leg )
10011 . to investigate the change in knee extensor strength and single - leg hop distance between the
peg and npeg , a repeated measures analysis using the independent sample t - test . the knee extensor strength deficit was calculated as the percentage difference between the
injured and uninjured side . the knee extensor strength deficits was significantly different
between the groups , both at an angular velocity of 60/s ( p=0.018 ) and 180/s ( p=0.033 ) . patients in the peg showed a significantly greater improvement in post - operative strength
than patient in the npeg at 60/s , and180/s ( table
2table 2.change in knee extensor muscle strength and single - leg hop functionmain outcome measurementpeg ( n=40)npeg ( n=40)knee extensor 60/sec deficit ( % ) preop22.813.723.515.8post - op28.59.0 * 36.510.7knee extensor 180/sec deficit ( % ) preop16.610.617.511.9post - op23.39.0 * 27.912.6single - leg hop lsi ( % ) preop75.110.376.58.9post - op85.37.4 * peg : preop exercise group ; npeg : no pre - op exercise
group . * p<0.05).the peg also showed significant improvement in the single hop distance test
( p=0.029 ) . peg : preop exercise group ; npeg : no pre - op exercise
group . * p<0.05 the purpose of this study was to examine the effects of a 4 weeks preoperative program of
the knee extensor muscle strengthening on post - operative quadriceps muscle strength and knee
function . outcomes confirmed the benefits of the peg in significantly lowering
post - operative deficits in quadriceps strength and improving single - leg hop distance
compared to patients who did not participate in a preoperative exercise program . knee extensor strength deficit following aclr is a common problem which has been reported
in previous studies12 , 13 . in a recent review of the literature , 11 studies
including research by palmier - smith et al.3 , reported knee extensor muscle strength deficit , ranging between 24%
and 40.5% , 6 months after aclr , while another 10 studies reported a residual , deficit in
quadriceps strength of 10% to 27% , 12 months after aclr3 . furthermore , long - term negative outcomes of strength deficits on
knee function for sports activities and even during daily activities . thus , the recovery of
quadriceps muscles strength is one of the most important factors after aclr14 . recent studies have investigated the relationship between pre and postoperative quadriceps
muscle strength deficits and function . according eitzen et al.9 reported patients with greater than 20% deficit in quadriceps
strength to have a significantly greater impairment on functional knee scores and strength . further , shelbourne et al.8 reported that
the postoperative quadriceps muscle strength to be significantly higher in patients with
greater preoperative strength than in patients with poor preoperative strength . these
studies provide strong evidence that preoperative quadriceps muscle strength positively
affects postoperative muscle strength and function . therefore , adequate and suitable program
of exercise before surgery would have an important role of improving post - operative muscle
strength . in our study , there was a significant difference in postoperative knee extensor
strength deficit between the npeg and peg groups , with a mean strength deficit of 36.5% for
the npeg compared to 28.5% . restoration of muscle strength , which was measured by
performance on the single leg - hop distance test , is considered to be an important
determinant for a successful return to sports after aclr9 , 17 . the limb symmetry index
( lsi ) has been the most frequently reported criterion for comparing muscle strength and hop
performance with the uninjured side18 . our study demonstrated that patients in the showed peg significantly improved in the single
leg - hop distance compared to the npeg . the results indicate that the greater improvements in
quadriceps strength in the peg had a positive effect on knee function test , such as the
single - leg hop , compared to the npeg . the result indicated that preoperative exercises not
only prevented quadriceps weakness , but also accelerated the recovery of muscle strength ,
which helped patients to quickly adapt to the rehabilitation environment . it is also
anticipated that possible re - injury could be prevented by having better strength and
function . | [ purpose ] after an anterior cruciate ligament injury and subsequent reconstruction ,
quadriceps muscle weakness and disruption of proprioceptive function are common .
the
purpose of this study was to examine the effects of a 4 weeks preoperative exercise
intervention on knee strength power and function post - surgery .
[ subjects and methods ]
eighty male patients ( 27.85.7 age ) , scheduled for reconstruction surgery , were randomly
assigned to two groups , the preoperative exercise group ( n=40 ) and a no preoperative
exercise group ( n=40 ) .
the preoperative exercise group participated in a 4-week
preoperative and 12-week post - operative programs , while the no preoperative exercise group
participated only in the 12-week postoperative exercise program .
isokinetic measured of
quadriceps strength were obtained at 4 weeks before and 3 months after surgery .
[ results ]
the knee extensor strength deficits measured at 60/s and 180/s was significantly lower
in the preoperative exercise group compared with the no preoperative exercise group . at 3
months after surgery , the extensor strength deficit was 28.59.0% at 60/sec and 23.39.0%
at 180/sec in the preoperative exercise group , whereas the no preoperative exercise group
showed extensor strength deficits of 36.510.7% and 27.912.6% at 60/sec and 180/sec ,
respectively .
the preoperative exercise group demonstrated significant improvement the
single - leg hop distance .
[ conclusion ] four week preoperative exercise may produce many
positive effects post reconstruction surgery , including faster recovery of knee extensor
strength and function , as measured by single - leg hop ability . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
viral hepatitis b infection is a common , serious disease caused by the hepatitis b virus ( hbv ) ( 1 ) . hbv is one of the main causes of hepatic decompensation , cirrhosis , and hepatocellular carcinoma ( hcc ) ( 1 , 2 ) . worldwide , more than two billion people are infected with hbv and approximately 400 million people are hbv carriers ; of these , 75% are asian ( 3 , 4 ) . the prevalence of hbv carriers differs around the world and can be divided into regions of high , medium , or low endemicity . in low - prevalence areas , including the united states , northern europe , australia , and parts of south america , the incidence of hbv carriers is less than 2% . in moderate - prevalence areas , such as the middle east , some eastern european countries , and the mediterranean basin , finally , in high - endemic areas , such as the central asian republics , southeast asia , sub - saharan africa , and the amazon basin , the hbv carrier rate is over 8% ( 1 ) . approximately 45% of the world s population lives in hyper - endemic areas ( 5 ) . hbv infection is endemic in most asian countries , and over 80% of liver cancers that occur in asia are attributable to hbv ( 6 ) . hbv infection is still the main cause of chronic liver disease in iran , although its seroprevalence has decreased over the last two decades . iran has an intermediate - to - low endemicity for hbv infection ( 7 ) . a systematic review of hbv infections in iran revealed that the prevalence is approximately 2.14% ( range 1.3% - 6.3% ) across the provinces ( 8) , while the rate of hbv carriers varies from 1.7% to 5.4% , with an average of 3.5% ( 9 ) . so far , chronic hepatitis b remains the cause of more than 50% of cases of cirrhosis and hcc in iran ( 10 ) . a rise in general public awareness about hbv transmission , along with the mass vaccination program in place since 1993 for all neonates , healthcare workers , and others at risk of hbv infection , has influenced the outcome of this disease ( 10 ) . the results of epidemiological studies in iran showed that the predominant genotype and serotype of hbv are d and ayw2 , respectively ( 11 ) . the most common routes of hbv transmission in iran are perinatal transmission and intravenous drug abuse ; however , during recent years , the epidemiology of this infection is changing from a vertical to a horizontal route ( 12 ) . hcv infection also constitutes a major global health problem , and , like hbv , it is a leading cause of cirrhosis and hcc , which is considered as the third cancer - associated cause of deaths worldwide ( 13 ) . approximately 85% of individuals infected with hcv will develop chronic hcv infection ( 14 ) . this infection affects more than 170 million people worldwide and the prevalence ranges from 0.2% to 40% in different countries . the prevalence of hcv infection is less than 0.5% ( 0.1% in women and 1.0% in men ) in the general iranian population ( 15 ) . any modeling and assessment of burden associated with hbv and hcv infection requires prevalence estimates . so far , the hbv and hcv seroprevalence among high - risk jobs is limited , and comprehensive data are not available for many countries . therefore , knowledge about the rate of these infections and their related risk factors are necessary for the implementation of any preventive program . the aim of this study was to analyze the prevalence of hepatitis b and c infections in barbers ( as high - risk workers ) compared with barbers ( as the control group ) in tehran , and to evaluate their attitudes and knowledge about the occupational risk of hepatitis b and c in order to provide possible solutions for the preservation and promotion of health , the selection of appropriate preventive methods for the health authorities , and general public health . the ethical standards of this study were approved by the ethics committee of the avicenna research institute and the iranian ministry of health . this case - control , cross - sectional study was conducted in tehran , iran , during 2008 - 2009 . the target population was 611 barbers and 556 bakers who had been working in their professions for at least one year . the control group ( bakers ) was an occupational group with a lower occupational risk for hbv infection transmission . tehran is divided into 22 municipal districts , with almost 900 barbers working in each district . considering our sample size and the ability of the sampling team to cover almost 35 samples each day , barbers and bakers were selected from 15 different regions by simple random sampling . a minimum number of selected participants from each district was considered to be almost 35 ; however , more than 35 participants were selected from districts with more than 900 barbers of bakers . randomization was done using systematic sampling selection from each region based on an alphabetical list of the barbers and bakers provided by the environmental health office of tehran . after informed consent was obtained , 5 ml of blood was collected from each participant . the samples were centrifuged and the serum was separated and kept in the freezer until the main tests were conducted . hbv markers were detected using commercial elisa kits ( hbsag , hbcab , and hbsab , iema , radim , italy ; hcv ab , acon laboratories , inc . serum levels lower than 10 miu / ml were considered hbsab - negative . a questionnaire consisting of four sections ( sociodemographic characteristics , awareness , attitudes , and behavior ) was designed to measure the knowledge and attitudes of participants with regard to hepatitis b and c infections in the two study groups . the knowledge scores in this section were divided into five groups , with the lowest score considered < 10 and the highest score considered between 29 - 34 ; therefore , 18 was calculated as an average score . the attitude section checklist included 11 questions , which were classified and analyzed based on the likert scale . the attitude scores were divided into five groups , with the lowest score of 23 and the highest score of 49 - 55 therefore , 18 was calculated as an average score . in addition , the behavior and performance section of the checklist had 11 questions , which were analyzed based on the type of question . the content validity of the questionnaire was established by a panel of experts in infectious disease , public health , biostatistics , and immunology . the internal consistency of the final version of the checklists was measured by the coefficient of cronbach s alpha test value , which varied from 0.75 to 0.82 . based on the t - test , there was a significant difference ( p < 0.01 ) between the mean age of the bakers ( 30.7 10.7 years ) and the barbers ( 38.2 12 years ) . it was shown that age did not have a significant impact on other dependent variables , so it was excluded from the model . the collected data was analyzed with spss version 13 , using the chi - square test , fisher s exact test , the logistic regression model , and one - way analysis of variance ( anova ) . the ethical standards of this study were approved by the ethics committee of the avicenna research institute and the iranian ministry of health . this case - control , cross - sectional study was conducted in tehran , iran , during 2008 - 2009 . the target population was 611 barbers and 556 bakers who had been working in their professions for at least one year . the control group ( bakers ) was an occupational group with a lower occupational risk for hbv infection transmission . tehran is divided into 22 municipal districts , with almost 900 barbers working in each district . considering our sample size and the ability of the sampling team to cover almost 35 samples each day , barbers and bakers were selected from 15 different regions by simple random sampling . a minimum number of selected participants from each district was considered to be almost 35 ; however , more than 35 participants were selected from districts with more than 900 barbers of bakers . randomization was done using systematic sampling selection from each region based on an alphabetical list of the barbers and bakers provided by the environmental health office of tehran . after informed consent was obtained , 5 ml of blood was collected from each participant . the samples were centrifuged and the serum was separated and kept in the freezer until the main tests were conducted . hbv markers were detected using commercial elisa kits ( hbsag , hbcab , and hbsab , iema , radim , italy ; hcv ab , acon laboratories , inc . serum levels lower than 10 miu / ml were considered hbsab - negative . a questionnaire consisting of four sections ( sociodemographic characteristics , awareness , attitudes , and behavior ) was designed to measure the knowledge and attitudes of participants with regard to hepatitis b and c infections in the two study groups . the knowledge scores in this section were divided into five groups , with the lowest score considered < 10 and the highest score considered between 29 - 34 ; therefore , 18 was calculated as an average score . the attitude section checklist included 11 questions , which were classified and analyzed based on the likert scale . the attitude scores were divided into five groups , with the lowest score of 23 and the highest score of 49 - 55 therefore , 18 was calculated as an average score . in addition , the behavior and performance section of the checklist had 11 questions , which were analyzed based on the type of question . the content validity of the questionnaire was established by a panel of experts in infectious disease , public health , biostatistics , and immunology . the internal consistency of the final version of the checklists was measured by the coefficient of cronbach s alpha test value , which varied from 0.75 to 0.82 . based on the t - test , there was a significant difference ( p < 0.01 ) between the mean age of the bakers ( 30.7 10.7 years ) and the barbers ( 38.2 12 years ) . it was shown that age did not have a significant impact on other dependent variables , so it was excluded from the model . the collected data was analyzed with spss version 13 , using the chi - square test , fisher s exact test , the logistic regression model , and one - way analysis of variance ( anova ) . they were in the age range of 12 - 82 years , with an average age of 35 3 years . hbsag elisa test was positive in 1.1% of the barbers ( 7 of 611 ) and 1.4% of the bakers ( 8 of 556 ) , with an overall prevalence of 1.3% ( table 1 ) . although the prevalence rate of hbsag among the bakers was higher compared with the barbers ( 1.4% versus 1.1% ) , this difference was not significant . the prevalence of hbcab ( only the presence of antibody was determined ) among barbers was higher than that among bakers ( 9.2% versus 6.2% ) , but this was not significant ( p = 0.063 ) . besides , the prevalence of hbsab among barbers was also higher than that in bakers ( 31.9% versus 18% ) , which was statistically significant ( p < 0.001 ) . anti - hcv was detected in 13 individuals , with an overall prevalence of 1.1% . the prevalence rate of anti - hcv in barbers was higher than in bakers ( 2% versus 0.2% ) and this difference was statistically significant ( p = 0.004 ) . double infection was detected in only one barber , who was positive for both hbsag and anti - hcv . the majority of participants had a high level of knowledge about blood - borne hbv and hcv infections and their risk of transmission ( over 50% correct answers for most questions ) . the difference in awareness regarding the existence of disease risk between the two groups was significant ( p < 0.001 ) . the difference between the barbers and the bakers knowledge about hepatitis b and c infections and the use of special needles for tattooing , drug use , blood , blood products , and contaminated clips and about body fluids capable of transmitting hbv and hcv . table 3 reports the barbers and bakers knowledge levels about body fluids capable of transmitting hbv and hcv . a clear majority of barbers knew that these viruses can be transmitted by semen and saliva ( p < 0.001 ) . numbers before the parentheses represent the number of barbers or bakers who answered yes ; numbers in parentheses represent percentages ( valid percent ) of each group ; the total number represents the sum of numbers in each age group ; it is not equal to the sum of barbers and bakers because some participants did not give their age on the questionnaire . this table shows barbers and bakers knowledge about the possibility of transmitting hbv and hcv via bodily fluids ; no . represents the number of barbers or bakers who were aware of this information , and % represents the percentage ( valid percent ) of each group , by comparing the numbers in these groups with the total number of barbers or bakers . based on the information regarding the frequency of hbsag and the attitudes of participants about hepatitis b and c , the barbers with greater numbers of tattoos and finger - cuts with needles , blades , and scissors , were more likely to be positive for hbsag . there was a significant difference between the attitude of barbers and bakers regarding risk of the disease , their role in disease prevention , routes of disease transmission , self - protection against the disease , and staying in their occupation despite being disease carriers ( table 4 ) . analysis of the frequency of hbsag and the demographic information showed that the majority of barbers with a history of divorce ( 3.1% ) , barbers without a high school diploma ( 1.6% ) , and barbers in the age range of 40 - 49 years ( 2.1% ) were positive for hbsag . the level of hbsab was higher in barbers aged 50 - 59 years , those who were widows or widowers , those with 21 - 30 years of work experience , and those who had acquired skills through training courses . in addition , a greater number of barbers with positive hbsab lived in regions 1 and 8 , according to the tehran regional municipality . table 5 demonstrates the practical experiences of barbers and bakers with regard to hepatitis b and c infections . the results showed that there was a statistically significant difference in vaccination histories among the bakers and barbers , as a higher number of barbers had received the 3-dose hepatitis b vaccination ( p < 0.001 ) . however , there was no statistically significant difference in histories of blood transfusion and drug abuse between the bakers and barbers . represents the number of barbers or bakers who answered yes to these questions ; for example , in the first question , 116 barbers answered yes , which is 19.1% of the total barbers who participated in the survey . chi - square test was used to calculate p values ; the total number represents the sum of numbers in each age group ; it is not equal with the sum of barbers and bakers , because some participants did not provide their age on the questionnaire . they were in the age range of 12 - 82 years , with an average age of 35 3 years . hbsag elisa test was positive in 1.1% of the barbers ( 7 of 611 ) and 1.4% of the bakers ( 8 of 556 ) , with an overall prevalence of 1.3% ( table 1 ) . although the prevalence rate of hbsag among the bakers was higher compared with the barbers ( 1.4% versus 1.1% ) , this difference was not significant . the prevalence of hbcab ( only the presence of antibody was determined ) among barbers was higher than that among bakers ( 9.2% versus 6.2% ) , but this was not significant ( p = 0.063 ) . besides , the prevalence of hbsab among barbers was also higher than that in bakers ( 31.9% versus 18% ) , which was statistically significant ( p < 0.001 ) . anti - hcv was detected in 13 individuals , with an overall prevalence of 1.1% . the prevalence rate of anti - hcv in barbers was higher than in bakers ( 2% versus 0.2% ) and this difference was statistically significant ( p = 0.004 ) . double infection was detected in only one barber , who was positive for both hbsag and anti - hcv . the majority of participants had a high level of knowledge about blood - borne hbv and hcv infections and their risk of transmission ( over 50% correct answers for most questions ) . the difference in awareness regarding the existence of disease risk between the two groups was significant ( p < 0.001 ) . the difference between the barbers and the bakers knowledge about hepatitis b and c infections and the use of special needles for tattooing , drug use , blood , blood products , and contaminated clips and about body fluids capable of transmitting hbv and hcv . table 3 reports the barbers and bakers knowledge levels about body fluids capable of transmitting hbv and hcv . a clear majority of barbers knew that these viruses can be transmitted by semen and saliva ( p < 0.001 ) . numbers before the parentheses represent the number of barbers or bakers who answered yes ; numbers in parentheses represent percentages ( valid percent ) of each group ; the total number represents the sum of numbers in each age group ; it is not equal to the sum of barbers and bakers because some participants did not give their age on the questionnaire . this table shows barbers and bakers knowledge about the possibility of transmitting hbv and hcv via bodily fluids ; no . represents the number of barbers or bakers who were aware of this information , and % represents the percentage ( valid percent ) of each group , by comparing the numbers in these groups with the total number of barbers or bakers . based on the information regarding the frequency of hbsag and the attitudes of participants about hepatitis b and c , the barbers with greater numbers of tattoos and finger - cuts with needles , blades , and scissors , were more likely to be positive for hbsag . there was a significant difference between the attitude of barbers and bakers regarding risk of the disease , their role in disease prevention , routes of disease transmission , self - protection against the disease , and staying in their occupation despite being disease carriers ( table 4 ) . analysis of the frequency of hbsag and the demographic information showed that the majority of barbers with a history of divorce ( 3.1% ) , barbers without a high school diploma ( 1.6% ) , and barbers in the age range of 40 - 49 years ( 2.1% ) were positive for hbsag . the level of hbsab was higher in barbers aged 50 - 59 years , those who were widows or widowers , those with 21 - 30 years of work experience , and those who had acquired skills through training courses . in addition , a greater number of barbers with positive hbsab lived in regions 1 and 8 , according to the tehran regional municipality . table 5 demonstrates the practical experiences of barbers and bakers with regard to hepatitis b and c infections . the results showed that there was a statistically significant difference in vaccination histories among the bakers and barbers , as a higher number of barbers had received the 3-dose hepatitis b vaccination ( p < 0.001 ) . however , there was no statistically significant difference in histories of blood transfusion and drug abuse between the bakers and barbers . represents the number of barbers or bakers who answered yes to these questions ; for example , in the first question , 116 barbers answered yes , which is 19.1% of the total barbers who participated in the survey . chi - square test was used to calculate p values ; the total number represents the sum of numbers in each age group ; it is not equal with the sum of barbers and bakers , because some participants did not provide their age on the questionnaire . the results of our study showed that being a barber alone is not a potential risk factor for hbv infection . the results also showed barbers ' higher level of knowledge about hbv and hcv routes of transmission and being at risk of the diseases . barbers in iran can be considered at high risk for developing hcv infections , and hcv is still an occupational health hazard for these individuals . the assessment of hepatitis b and c prevalence in different communities , especially among high - risk occupations , has been considered a basic element of public health programs ( 8 , 16 ) . rates of hbv and hcv infections vary markedly from one country or region to another , and depend on host characteristics and environmental conditions ( 17 , 18 ) . in the last few decades the prevalence of hbv and hcv infections has been widely investigated in many occupational groups , but few data are available on the prevalence in barbers as a high - risk group . the present study would be considered a complementary study to previous ones in iran and other countries . the prevalence of hbv infection is approximately 2.14% ( range 1.3% - 6.3% ) across iran s provinces , while the prevalence of hcv infection is less than 0.5% in the iranian general population ( 8 , 15 ) . our results showed that the prevalence of hbsag among barbers was about 1.1% , almost the same as in the control group of bakers ( 1.4% ) . thus , we found that being a barber alone is not a potential risk factor for hbv infection , although the opposite has been shown in other countries ( 18 , 20 ) . based on similar studies in other countries , the prevalence of hbsag in egyptian barbers and their clients is 4.2% and 3.9% , respectively , while the rates are 8.5% and 1.9% among turkish and moroccan barbers , respectively ( 18 , 21 , 22 ) . we found that hbsab was present in 31.9% of barbers and only 18% of bakers , which is a significant difference . this demonstrates barbers greater knowledge about the benefits of hepatitis b vaccination . in our study , hcv seropositivity was 2% in barbers , which is significantly higher than that in the control group ( 0.2% ) . therefore , the prevalence of hcv was found to be higher in iranian barbers in comparison with the control group , while the rate was similar for hbv . thus , barbers could be considered at high risk of developing hcv infections in iran , and hcv is still an occupational health hazard for them . our results showed that 9.2% of the barbers were hbcab - positive compared with 6.2% of bakers . however , the rates of hbcab in barbers ( 12.3% ) and their clients ( 12.7% ) in some regions of egypt were almost the same ( 21 ) . the use of sharp tools in barbershops could be a potential risk factor for blood - borne disease transmission and infections such as hepatitis . previous studies have demonstrated that the use of shared and unsterile blades is a risk factor for hepatitis infection in developing countries , such as italy and turkey ( 19 , 23 , 24 ) . our results showed significant differences between barbers and bakers regarding knowledge and awareness of hepatitis and its mode of transmission ( table 2 ) . attitudes were also assessed in barbers and bakers , and the results showed that in special cases , barbers attitudes were better than those of bakers ( table 4 ) . all participants who were positive for hbsag and hbcab were asymptomatic , with a lack of awareness about their disease , which makes them high - risk candidates for chronic disease and disease transmission in society . in general , the concept of infectious risk associated with blood is well understood by many barbers in iran . the results of a study among egyptian barbers showed that the participants did not have sufficient knowledge , as only 40% were informed about hepatitis b and c protective vaccinations and drugs . egypt has the highest prevalence of hcv in the world , and the rate of this infection increases steadily with age ( 21 ) . awareness , especially about the risk of transmission of blood - borne infectious diseases , is even lower in pakistan , morocco , and ethiopia ( 21 , 22 , 24 ) . some scientists believe that the relatively high prevalence of hepatitis c in egypt is related to a weakened immune system following common infections such as schistosomiasis ; however , no significant correlation between the incidences of these two diseases in egypt or brazil has yet been documented ( 21 ) . interestingly , in some studies , researchers have not found any correlation between barbershops hair - cutting tools and an increased risk of viral infections , including hepatitis . shalaby et al . ( 21 ) believe that hepatitis b and c are not considered significant dangers for barbers in egypt , as there is no significant difference in the incidence of hepatitis between them and the general population . in 2007 , a study was conducted in pakistan to measure awareness of barbers regarding blood - borne diseases , such as hepatitis and hiv . it was found that approximately 70% of participants had not seen or heard in the media any educational program about these diseases . while nearly 90% of them did not agree on regular public screening programs against blood - borne infectious diseases , 97% were willing to be tested for these infections . the status of these professions becomes even more problematic because barbers in some rural communities in pakistan also provide other services , such as circumcision , incisions , and wound drainage ( 25 , 26 ) . hepatitis b vaccinations in morocco were begun in 1999 ; today , more than 90% of infants born in morocco receive the vaccination while adults remain at risk for both hepatitis b and c. the prevalence rates of hepatitis b and c among barbers in various regions of rabat in morocco are approximately 28% and 1.1% , respectively ( 22 ) . however , only 1.9% of barbers with the active form of hbv infection have been reported by measuring hbsag . the prevalence of hepatitis b in the age range of 45 - 70 years was significantly higher than that in younger age groups ( 22 ) . approximately 70% of participants in these studies had no education or dropped out of primary school , and approximately 94% suffered from low socioeconomic status . apparently , the risk of hepatitis b in moroccan barbers and general public was equal . however , similar studies in china , turkey , and italy showed that barbers are more likely to acquire active hbv infection ( 22 ) . ( 24 ) studied hepatitis rates and awareness among barbers in islamabad , pakistan , and showed that although the majority ( 89.7% ) showed an interest in screening for hepatitis b and c , only 1% were informed about the mode of transmission and prevention of the diseases . as mentioned above , based on candan s work , the seropositivity of hepatitis b in barbers in the sivas region of turkey was significantly different from the control group . it appears that scissor - cuts and needle - pricks confer a three - times - higher risk of hbv infection acquisition in barbers . they showed that 8.5% of barbers were hbsag - positive , while the prevalence of hbsag carriers in the turkish population is approximately 4% - 5% . they also showed that seropositivity of hepatitis c in the barber group was 2.8% , which was not significantly different from the control group ( 1.1% ) ( 18 , 27 ) . the use of blades for shaving in barbershops is a mode of hepatitis transmission throughout the world , and also the use of other unsterile tools could be considered an infection - transmission factor ( 28 ) . although the level of awareness about hepatitis and the risk of transmission were good among iranian barbers , there are still some unsafe practices that may lead to infections due to blood - borne viruses . therefore , extensive training programs and media campaigns are still essential for raising awareness throughout society . in conclusion , it is expected that educational classes could increase the awareness of high - risk groups about prevention and the signs and symptoms of hepatitis . according to the present results , it is necessary to conduct health - training courses and awareness programs for the general public , not just for high - risk groups . finally , the current study provides evidence and a reason for the initiation of larger studies in iran to obtain more useful information about the incidence , mode of transmission , and transmission patterns of infectious diseases and their associations with certain professional groups such as barbers . | backgroundhepatitis b virus ( hbv ) and hepatitis c virus ( hcv ) infections are major health problem in the world .
hairdressers ( barbers ) are in continuous contact with scissors and blades , and are considered a high - risk group for these infections.objectivesthe aim of this study was to analyze the prevalence of hepatitis b and c infections in barbers in tehran and to evaluate their attitudes and knowledge about the occupational risk of these infections.methodssix hundred eleven barbers were included in this study .
a group of 556 bakers were also selected from the same regions , as a low - risk control group .
serum levels of hepatitis b surface antigen ( hbsag ) , hbsag - specific antibody ( hbsab ) , hepatitis b core antigen - specific antibody ( hbcab ) , and hepatitis c virus - specific ( anti - hcv ) antibody markers were measured with the enzyme - linked immunosorbent assay ( elisa ) .
participants were interviewed using a questionnaire consisting of four sections : demographic information , awareness , behavior , and personal attitudes.resultsthere were no significant differences in the frequency of hbsag between the two groups . however , the frequency of hcv ab in barbers was significantly higher than that in bakers ( p < 0.005 ) .
in addition , the frequency of hbsab marker in barbers was significantly correlated with increased awareness ( p < 0.05 ) and number of tattoos ( p < 0.001 ) .
hbcab marker was significantly correlated with age ( p < 0.001 ) and duration of professional career ( p < 0.005 ) . with age ,
barbers attitudes improved significantly ( p < 0.05).conclusionsbeing a barber alone is not a potential risk factor for hbv infection , while hcv infection is still an occupational health hazards for barbers .
we suggest more extensive case - control studies with regard to rates of hepatitis b and c markers among barbers in other iranian cities to assess the incidence of hepatitis b and c infections among this population . |
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the classical strategies employed in the search for genes that predispose to complex phenotypes are linkage and association . linkage studies require recruitment , of family units of varying size , and are not , practically feasible in the context , of pharmacogenetics . family - based association studies require smaller family units , but they too are not practical for pharmacogenetics , except , for studies conducted in pediatric populations . their advantages are considerable , the most , striking being the possibility of recruiting large samples that have sufficient , statistical power to investigate genetic variants of relatively small effect . large samples are particularly important if the effect of more than one gene is being studied in the same sample , and interactions among genes and between genes and the environment , are being sought . on the other hand , case -control designs are notoriously susceptible to the effects of ethnic stratification , which can lead to spurious results . if this population is overrepresented in the case or control group , a spurious finding will result , in which the allele is erroneously associated with the phenotype . the nature of the problem is illustrated in figure 1 , which shows the frequency of the gly allele of the dopamine d3 receptor gene ( drd3 ) scr9gly polymorphism in samples from several populations . these samples were included in a pooled and meta - analytic study of the drd3 ser9gly polymorphism as a risk factor for td . great variability in the frequency of the 9gly allele in the different samples included in the study is immediately evident , ranging from 30% to 40% among caucasians from different countries , and reaching 80% among african - americans . if the association of the drd3 gly 9 allele with td were tested without controlling for ethnicity in this pooled sample , made up of 317 patients with td and 463 patients without td , spurious results could easily arise . therefore , a stepwise logistic regression was employed so as to allow the confounding effects of ethnicity and also age and gender to be taken into account . td was significantly associated with drd3
gly9 allele carrier status ( x=4.46 , df=1 , p=0.04 ) over and above the effect , of ethnicity . similar positive effects were observed when controlling for age and gender ( x=5.02 , df=1 , p=0.02 ) . a meta - analysis was performed , which included all the samples in the pooled analysis , as well as data from additional published studies . the mantel - haenszel pooled odds ratio ( or ) for drd3 gly9 allele carrier status increasing susceptibility to td was 1.33 ( 95% confidence interval [ ci ] 1.04 - 1.70 , p=0.02 ) ( figure 2 ) . meta - analyses are less sensitive to stratification and admixture ( unless these are present in the samples included in the meta - analysis ) , since original data are not pooled and the unit , of analysis is studies and not individuals . the findings of our studies of the drd3 ser9gly polymorphism and td indicate that , when pooled pharmacogenetic analyses arc conducted on well - characterized samples , it is possible to control for the potentially artifactual effects of ethnicity . this is important , because it is essential to gather large samples for pharmacogenetic studies so as to have sufficient statistical power . the same general strategy was implemented to examine the association of the serotonin 5-ht2a receptor gene and td , and also to examine association of the 5-ht2c receptor gene with unipolar and bipolar affective disorder . when using ostensibly homogeneous samples , the genomic control method may be implemented to rule out population influences on the markers being studied . this involves genotyping additional markers that are putatively unrelated to the phenotype , and these are used to determine the degree of stratification that is present in the sample . it is often not taken into account , that genetic variants may have differing functional importance depending on the stage of the life cycle . this is obvious in the case of genes that predispose to disorders of later life such as alzheimer 's disease ( although the pathophysiological effects of variations in these genes may be manifested long before the clinical threshold is crossed ) . similarly , genes that , influence neurodevelopment in utero may be associated with susceptibility to schizophrenia that only becomes clinically manifest , in late adolescence . we have observed an age - related association of two serotonergic genes , the 5-ht2c
( htr2c ) and the 5-ht2a receptor ( htr2a ) with susceptibility to td . our finding with the 5-ht2a receptor has been replicated in a large multicenter study .
in an earlier report
( figure 3 ) , we showed that scores on the abnormal involuntary movements scale ( aims , the hallmark of td ) were significantly related to the ser23 allele of the cys23ser polymorphism of the 5-htc receptor gene and to the 102c allele of the t102c polymorphism in the 5-ht2a receptor gene , in older but , not , younger patients . we sought , to replicate this finding the context of a multicenter study involving 635 patients , 256 of whom manifested td and 379 of whom did not . our initial analysis employed stepwise logistic regression as for our drd3 study and showed an overall association of the t102c polymorphism in the 5-ht1a receptor gene with td . we then analyzed the younger and older patients separately . in keeping with our finding in the jerusalem sample alone , the t102c polymorphism was significantly associated with td in the older patients only ( figure 4 ) . these replicated findings accentuate the importance of taking into account the possibility that pharmacogenetic effects may be age - related . thus , an association first , reported in an older sample may be missed if replication is sought , in a younger sample . this was the case with our initial report , of an association of the t102c polymorphism in the 5-ht2a receptor gene with td , which was initially not , replicated by basile et al who studied a sample more than two decades younger than ours . the age range of the pooled sample studied by lerer et al was wide enough to allow replication of the finding in the older subjects . pharmacogenetic phenotypes ( such as treatment outcome , onset , of therapeutic effect , rapidity of response , and adverse effects ) are complex traits to which background and demographic factors contribute , as well as factors related to the treatment . the genetic background is likely to be polygenic , with an as - yet unknown number of genes contributing a small proportion of the variance . one model is additive , the assumption being that , each gene contributes cumulatively to the phenotype , and that , the overall risk is a sum of these individual contributions . a second , more complex model postulates that the effects of certain genes are conditional on others , implying gene - gene interaction or epistasis . under this model , a specific allele of gene a will confer an effect , in the presence of a specific allele of gene b , but will not confer susceptibility in its absence . the situation is rendered more complex by the possibility that haplotypes made up of two or more variants may contribute additively or interactively to susceptibility . these considerations are applicable to td , which is a complex trait , related to drug treatment , to which several demographic and background factors contribute , most notably age , and with which several genes have been associated . as yet , there has been relatively little work in psychiatric pharmacogenetics on the combined effects of genes and on gene - gene interaction . these models have been tested on a limited basis in the context of susceptibility to td in samples that we and other investigators have studied ; the scope of such testing is constrained by the size of the samples . the additive effects of two genetic variants on susceptibility to td were first reported by scgman et al . patients who carried both the gly9 allele of the drd3 ser9gly polymorphism and the ser23 allele of the 5-ht2c cys23ser polymorphism showed evidence of an additive effect of the two risk alleles , both of which had been shown to separately increase risk for td . as shown in figure 5 , the highest , aims orofacial dyskinesia scores were observed in patients who carried both mutant risk alleles . studying gene - gene interaction and risk for td , segman et al examined the interactive contribution of a tc polymorphism in the promoter region of the cytochrome p450 17-hydroxylase gene ( cyp17 ) and the ser9gly polymorphism of drd3 that was previously shown to be associated with td . cyp17 variability could influence susceptibility to td because of effects on neuroprotective capacity or dopamine output through its role in the conversion of the neurosteroid , pregnanolone , to dehydroepiandrosterone ( dhea ) . a tc transition ( a2 allele ) in the 5 ' promoter region of the cyp17 gene creates an additional spl type ( ccacc box ) promoter site , resulting in a functionally relevant , increase in transcription rate . segman et al found that the tc polymorphism was not in itself associated with td . however , there was a significant excess of drd3 gly allele carriers among carriers of the cyp17 cc genotype with td compared with those without td ( 75.0% vs 14.3% ; x
= 6.0 , df=1 , p=0.01 ) . significant differences were not observed among carriers of the cyp17tt genotype or cyp17tc genotypes with td and carriers of these genotypes without td . two way analysis of variance ( anova ) with age at first , antipsychotic treatment as covariate showed a significant main effect of drd3 genotype on aims total score ( f=14.9 , df=1 , 106 , p=0.0002 ) . there was no main effect , of cyp17 genotype on the aims score . there was a significant , interaction between drd3 and cyp17 genotype on aims total score ( f=4.2 , df=2 , 106 , p=0.02 ) . figure 6 shows adjusted mean aims scores ( derived from analysis of covariance [ ancova ] ) for the patients grouped according to drd3 and cyp17 genotype . posthoc newman - keuls tests showed that the highest aims total ( p<0.03 ) was in patients carrying the drd3 gly9 allele and the cyp17 cc genotype . another gene - gene interaction reported in relation to td involved drd3 and cytochrome p450 1a2 ( cyp1a2 ) . basile et al further found that , those patients who exhibited the risk genotype at , both drd3 ( gly9/gly9 ) and at cyp1a2 ( cc ) had the most , severe td , whereas those who had only one risk genotype ( gly9/gly9 or cc ) demonstrated intermediate severity of td . those patients who did not , have any risk genotypes at either locus demonstrated the lowest mean td severity scores on the aim 's scale ( p<0.00007 ) . a more recent report of gene - gene interaction conferring susceptibility to td concerns the functional ala9val polymorphism in exon 2 of the magnesium superoxide dismutase gene ( mnsod ) . mnsod is one of three isoforms of superoxide dismutase ( sod ) that are important , as antioxidants in protecting cells from the deleterious effects of free radicals . the ala9 variant , hori et al found a small but , significant increase in the wild type val allele in japanese schizophrenia patients with td , suggesting that the ala allele may be protective against , free radical damage in td . this finding was not , replicated by zhang et al in chinese schizophrenia patients with td . however , in a subsequent report zhang et al found that the val - val genotype was in fact associated with td , but . only in those patients who were carriers of the gly allele of the drd3 scr9gly polymorphism . we examined association of the ala9val polymorphism of mnsod with td in israeli schizophrenia patients and also tested for an interaction between this polymorphism and the drd3 ser9gly polymorphism ( segman et al , unpublished data ) . patients were grouped according to whether they were carriers of the ala allele of mnsod , the gly allele of drd3 , both , or neither . this grouping takes into account an effect , of drd3 gly to increase susceptibility and a protective effect , of mnsod ala and anticipates that patients carrying drd3 gly and lacking mnsod - ala will be most , susceptible to td and will have the most severe abnormal involuntary movements . the mnsod - val drd3-gly genotype combination was the most , frequent , among patients with td ( 64% ) and the mnsod - ala drd3-ser genotype ( 24% ) was the least frequent . among patients without , the most frequent genotype combination was mn.sod - ala\ drd3-ser ( 76% ) and the least frequent , combination was mnsod - val drd3-gly ( 38% , x
= 10.5 , df=3 , p=0.01 ) . table i shows that patients with the mnsod - val\ drd3-gly genotype combination have the highest , aim 's total scores and patients with the mnsod - ala drd3-ser genotype combination have the lowest ( f=6.16 ; df=3 , 102 ; p=0.0007 ) . the aim of pharmacogenetic research is to develop clinically useful tests that will allow potential responders to a particular psychotropic agent to be identified prospectively , as well as those individuals likely to develop adverse effects . some of the reasons for this difficulty are applicable to drugs used to treat other complex disorders ; others are specific to psychotropic agents . the polygenic basis of pharmacogenetic traits is an issue of major importance . for most traits it , is unclear how many genes arc involved , and genes that have been implicated thus far in well - studied phenotypes such as td are of small effect . the or observed rarely exceed 2.0 , and for the most part , are less than 1.5 . it requires large samples to explore such small gene effects in the definitive fashion required for their inclusion in a pharmacogenetic test . large samples are also needed in order to tease apart , gene - gene and gene - environment interactions . recruitment , of large samples inevitably increases the likelihood of stratification , which can lead to spurious results and must be taken into account . a further consideration is that , background and demographic variables must be considered ( as pointed out in this paper for the role of age in the manifestation of certain gene effects in td ) as well as treatment - related variables such as medication dose , duration of treatment , and age at onset of treatment . furthermore , treatment outcome is frequently related to severity of illness , and this must , also be taken into consideration . thus , it , is clear that a model used to predict treatment outcome or susceptibility to adverse effects will be unavoidably complex , given the number of background and potential predicting variables that will need to be taken into account including gene - gene and gene - environment interactions . based on our previous work on the genetics of td , we have developed a preliminary model that takes into account the various , background , clinical and genetic factors that , we have studied ( scgman et al , unpublished data ) . we employed logistic regression and entered background and clinical variables known to influence susceptibility to td such as age , sex , cigarette - smoking , age at first , antipsychotic treatment , duration of antipsychotic treatment , antipsychotic dose in chlorpromazine units , and total score on the positive and negative syndrome scale ( panss ) . the genetic variables included were drd3 gly9 allele carrier status , mssod ala9 allele carrier status , htr2c ser23 allele carrier status , cyp17 genotype and htr2a genotype . interactions were also tested for inclusion , but none were retained in the final model . the overall model ( table ii ) was significant ( r=0.32 ; p=0.0005).the variables that , significantly predicted td within the model were age ( or=1.04 , p=0.047 ) , panss total score ( or=1.02 , p=0.014 ) , . drd3 gly9 allele carrier status ( or=4.39 , p=0.006 ) , and htr2a 102cc genotype ( or=4.18 , p=0.02 ) . mnsod ala9 allele carrier status and htr2c ser23 allele carrier status were retained in the overall model , but were not , significant . in this manner , 70.3% of cases could be correctly classified , compared with 60.2% prior to entry of the variables . . it is based on a single small sample of subjects , and may not , be generalizable to other samples and populations . while recognizing these limitations , the model does support the concept that a combination of background , clinical , and genetic variables could potentially be used to evaluate a priori the risk for td in patients treated with antipsychotic drugs that have the potential to induce this adverse effect . this approach could be extended to other pharmacogenetic phenotypes , and ultimately allow the development , of clinically viable pharmacogenetic tests that , will serve as the basis for a rational assessment of cost - benefit ratios in the choice of treatment with antipsychotics and other antipsychotic drugs . | the core hypothesis underlying pharmacogenetics is that genetic factors play a significant role in the well - recognized differences between individuals in response to medication and susceptibility to adverse effects .
if these genetic factors can be identified and understood , they may serve as predictors to guide clinicians in tailoring medication to the individual patient .
recent developments in the field of antipsychotic drug treatment suggest that pharmacogenetics could play an important role , permitting the use of first - generation antipsychotics ( fgas ) for patients in whom the use of second - generation antipsychotics ( sgas ) is limited by efficacy considerations or adverse effects , in this paper , key issues that need to be taken into consideration in designing and interpreting pharmacogenetic studies of antipsychotic drugs are discussed against the background of data emanaling from studies on the genetics of tardive dyskinesia ( td ) , an important adverse effect of fgas .
the issues considered include the advantages and potential pitfalls of case - control association studies of pharmacogenetic traits , the role of demographic factors such as age and gender , additive effects of genes , and gene - gene and gene - environment interaction .
the prospects for implementation of pharmacogenetic testing in the clinic are considered in the context of a preliminary model that has been tested for prediction of susceptibility to td . |
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an 84-year - old asian woman with 1 year history of type 2 diabetes on metformin presented to diabetes clinic with deranged serum glucose [ fasting : 140(70110 mg / dl ) and 2-hour postprandial : 147(<140 mg / dl ) ] . she complained of extreme weakness , fatigue , forgetfulness , somnolence , and a weight loss of 10 kg ( 22 lbs ) during 1 year duration . she denied increased sweating , heat intolerance , change in bowel habits , or any other symptoms suggestive of thyrotoxicosis . she had a thin built ( bmi : 16 kg / m ) , and exhibited listlessness , delayed reaction , apathy , tachycardia ( 108/minute ) and hypertension ( 180/90 mm hg ) . she also had diffuse , waxy thickening of skin involving pretibial area and dorsum of the foot with peau d'orange appearance ( fig . biochemical evaluation revealed thyrotoxicosis [ t3 : 338.59(60181 ng / dl ) , t4 : 19.5(3.212.6u / dl ) , tsh : < 0.01(0.355.50u / ml ) ] and hypocholesterolemia [ total cholesterol : 135 ( 150240 mg / dl ) , hdl : 30 ( 3070 mg / dl ) , ldl : 65 ( < 160 mg / dl ) ] . treatment with carbimazole and propranolol improved her symptoms , and her glucose levels returned to normal . metformin was gradually tapered and the result was normal glucose tolerance without any glucose - lowering therapy . application of topical corticosteroid for the pretibial myxedema produced gradual improvement in her skin texture . hyperthyroidism and diabetes can co - exist ; however , hyperthyroidism itself may cause hyperglycemia in such patients . excess thyroid hormone can induce glucose intolerance by increasing the hepatic glucose output via glycogenolysis and gluconeogenesis ( 3 ) , and by lowering both insulin secretion and peripheral insulin sensitivity ( 4 ) . diabetes and hyperthyroidism can have quite similar presentation , so thyrotoxicosis can be missed unless screened for specifically . in a patient with both the conditions concurrently , an oral glucose tolerance test should be done after correction of thyrotoxicosis , before considering a definite diagnosis of diabetes . after considering the weight loss attributable to hyperglycemia / diabetes , the first clinical impression in this patient was hypothyroidism . however , the picture of lethargy , slowness , weight loss , and apathy combined with low tsh and high t4 was compatible with apathetic hyperthyroidism . in this condition , the core presentation of apathy and depression is associated with severe muscle wasting and weight loss , dry skin , mild tachycardia , arrhythmias , and often congestive cardiac failure ( 5 ) . hand tremor and ophthalmopathy typical of graves disease are absent , and the thyroid gland if palpable is minimally enlarged ( 5 ) . although giving every appearance of apathy , the metabolic effect of the disease from excess thyroid hormone is severe as seen in our patient : hypertension ( increased receptor synthesis , positive inotropic , and chronotropic effect on heart ) , hyperglycemia , and hypocholesterolemia ( increased ldl receptor synthesis ) . pretibial myxedema due to dermal deposition of excess glycosaminoglycans may be seen on the lower legs , feet , and particularly the shins . the psychological aspects are probably related to increased levels of thyroxine and its effect on the nervous system . the elderly might have a reduced capacity to react to elevated thyroid hormones ( 2 ) . the disease may remain atypical throughout its course ; these patients under usual conditions do not appear extremely ill , but under stress may quietly sink into coma and die a relaxed death without activation , unlike the classic thyrotoxic patient who goes into crisis ( 1 , 6 ) . greater age , increased corrected calcium level , decreased tsh level , increased ft4 , decreased ft3/ft4 ratio , increased bone - specific alkaline phosphatase , and increased alt ( alanine aminotransferase ) may be significant in differentiating apathetic thyrotoxicosis from typical hyperthyroidism , which may aid in accurate diagnosis and treatment ( 7 ) . absence of hyperkinetic motor activity , hand tremor , and ocular signs of graves disease can sometimes mislead the diagnosis of hyperthyroidism . diabetes itself requires a screening for thyroid function , and when associated with apathy , weight loss and mild tachycardia in an elderly patient gives a strong indication for thyroid function testing to look for apathetic thyrotoxicosis . the authors have not received any funding or benefits from industry or elsewhere to conduct this study . | apathetic form of thyrotoxicosis occurs in the elderly who can present with features of hyperglycemia , hypothyroidism , depression , or an internal malignancy . a clinical suspicion and timely diagnosis of hyperthyroidism is needed to define the correct etiology of existing problems , and to prevent grave complications .
we discuss an 84-year - old woman who presented with fatigue and uncontrolled diabetes due to apathetic thyrotoxicosis . |
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cone beam computed tomography ( cbct ) is currently a valuable imaging modality with numerous applications in dental implant treatment , endodontic therapy , surgery , orthodontic assessment and evaluation of pathological lesions . the effects of factors such as system resolution and voxel size are undeniable on diagnostic accuracy of images , and there is the possibility of using a smaller voxel size to increase image resolution . however , use of a smaller voxel size to increase the resolution can create noise , and thus , significant improvement in the diagnostic value of images may not occur . also , the scanning time of objects becomes longer by increasing the resolution , the radiation dose increases and there would be higher risk of movement of patient during imaging . field of view ( fov ) refers to the scan volume of a particular cbct unit . a voxel describes the smallest distinguishable box - shaped part of a three - dimensional image . lowering the resolution may reduce the quality of images , increase noise and artifacts , and reduce the amount of anatomical information of the target areas . multiple factors such as fov , voxel size and the number of basis projections and image artifacts significantly affect the cbct image quality . in most cases , previous studies have reported inconsistencies in cbct measurements depending on the position of object within the fov . the accuracy of linear measurements is lower at the periphery compared to the center of the fov in cbct . because of the lack of any current protocol on selection of appropriate voxel size and also the proper location within the fov for cbct imaging in dentistry , the present study was conducted to assess the effects of voxel size ( resolution ) and the location of bone defects in different regions of the fov on diagnostic accuracy of cbct for their detection . four pieces of human mandible were selected after soft tissue removal and fixed in formalin . bone defects with 11 mm dimensions bone blocks were then fixed on a platform , which was parallel to the horizontal plane of the cbct unit ; cbct images of bone defects were obtained using alphard-3030 unit ( asahi roentgen ind . kyoto , japan ) operating at 4ma , 80kvp and 17s time in two different resolutions and five locations in the fov ( fig . bone defects were not created in the mandible of the negative ( healthy bone ) cases ( a total of eight cases ) and thus cbct images were obtained only from healthy bone under the same experimental conditions ( i.e. five locations in the fov at both high and low resolutions ) . locations in the fov were center , left , right , anterior and posterior , and resolutions used for cbct imaging included high resolution ( mode i : 0.2 mm voxel size , 1010 cm fov ) and low resolution ( mode p : 0.3 mm voxel size , 1515 cm fov ) ( figs . 2 and 3 ) . three observers evaluated the images using the same display monitor and romexis software ( planmeca , helsinki , finland ) , and then information was recorded in the relevant forms . the bone blocks were scanned at five different areas within the field of view of the cbct system a : anterior ; c : center ; l : left ; r : right ; p : posterior cbct scan obtained with 0.2 mm voxel size of a mandibular bone defect ; ( a ) center of the field of view ; ( b ) posterior region of the field of view cbct scan obtained with 0.3 mm voxel size of a mandibular bone defect ; ( a ) center of the field of view , ( b ) posterior region of the field of view in the first session , the observers were trained to record the information in the relevant forms . then , each sample was coded and the images were randomly presented to the observers . the ability to detect bone defects was assessed using a dichotomous scale ( seen and not seen ) . the images were coded again and randomly shown to the observers ; the related information was recorded . detection of bone defects ( seen and not seen ) was determined in positive cases ( bone defects ) and negative cases ( healthy bone ) based on the opinion of the observers ; the frequency of observations , high and low resolutions , and the location of object in the fov , and reported using number and percentage . intra- and inter - observer agreements for the detection of bone defects were calculated at different resolutions and locations in the fov using the kappa test . the sensitivity , specificity and 95% confidence interval values are presented in tables 1 to 3 . the highest sensitivity was found in the central region in both high and low resolutions , which was significantly different from other fov regions at 95% confidence level using the wilson formula . also , the least sensitivity was recorded in the posterior region of the fov at low resolution and significantly increased by increasing the resolution ( table 1 ) . resolution of images ( 0.3 mm and 0.2 mm ) and fov regions had insignificant effects on the specificity values ; because it was observed that the values were almost the same in all three observers in two resolutions and five different regions ( table 2 ) . the value of inter - observer agreement in different fov regions was 0.3 to 0.69 in high - resolution images and 0.46 to 0.69 in low - resolution images , and the lowest value was obtained in the posterior region in high resolution ( 0.3 ) . the highest value was obtained at the center of the fov in both resolutions ( 0.69 ) . the intra - observer agreement coefficients for all groups were adequately high ( kappa0.75 ) . sensitivity for detection of simulated bone defects at different object locations in the field of view ( fov ) and two different resolutions of cbct specificity for detection of simulated bone defects at different object locations in the field of view ( fov ) and two different resolutions of cbct system mean ( 95% confidence interval ) of simulated bone defects measured at different object locations in the field of view ( fov ) and two different resolutions of the cbct system according to the results of this study , some variations were observed in the diagnostic accuracy of cbct for bone defect detection in positive and negative cases during scanning in different regions of the fov . although the sensitivity significantly decreased in cbct images with the object in the posterior region of the fov in low resolution , it increased in high resolution , especially in the posterior region . high sensitivity was observed by increasing the resolution of cbct images from 0.3 mm to 0.2 mm ; and the values were significantly higher at the center of the fov compared to other regions in both high and low resolutions at 95% confidence interval using the wilson formula . in a study by ibrahim et al , in 2014 on the effects of different locations of object in the fov in two cbct systems , a significant difference in trabecular bone microstructure measurements in the central region was found relative to the peripheral areas in the fov in newtom system , but this difference was not significant in the accuitomo system , which may be due to differences between the two systems . the inherent artefacts of the system and the object region in a limited fov could also result in deviations of measurements . these factors increase the linear measurement values at the periphery compared to the central region of the fov of the cbct system . in the current study , defects were observed best at the center of the fov and had statistically significant difference with object in other regions . the non - uniformity of the beam intensity causes inconsistency in grey values within the cbct fov , in which it is relatively higher at the center than at the peripheral regions . a higher quality image can be obtained with the object at the center . in the current study , the lowest sensitivity was observed in the posterior region of the fov in low resolution , which could be due to under - sampling from the posterior region of the fov compared to other regions because of of device limitations . variations in diagnostic parameters with the object in different regions of the fov have also been observed in high - resolution peripheral ct and multislice ct . voxel size , contrast - to - noise ratio and image artifacts are among the factors associated with the fov . a small voxel size with a small fov is generally recommended to improve diagnostic accuracy . variations in the quality of images of bone defects in different fovs and resolutions may be attributed to increased image artefacts particularly in smaller fovs . also , an image with a bigger voxel and a higher contrast - to - noise ratio can have a higher resolution than an image with a smaller voxel and lower contrast - to - noise ratio . therefore , while the selection of voxel size depends on the nature of the diagnostic task , the clinician should be aware of the variations of resolution and selected fov regions . although cbct has been reported as a precise technique for assessment and detection of bone defects , some limitations of the systems should be well acknowledged . based on the results , the observer s performance with regard to cbct images taken at high resolution was estimated to be better than that with images with low resolution ; thus , the sensitivity values improved by increasing the resolution of cbct images from 0.3 mm to 0.2 mm ; and the frequency of correct diagnoses increased in both positive and negative cases . by rotation of x - ray beam around however , as patients vary in size , the fov parameter would be different as well . in addition , the selected fov remains the most important scanning parameter in limiting the radiation dose and image quality . theoretically , more uniform images can be obtained in the maxillofacial region due to high - density structures . substantial amount of scattered beams is produced in cbct technique that negatively affects the image quality . selected smaller fovs , anatomical structures located outside the defined area are imaged by the detector , and since only a small imaging range evaluates the tissue , the resulting images will not be uniform . kwong et al , in 2008 used 6-inch fov and showed that the resultant images had the highest resolution and the lowest voxel size ; although high resolution alone does not necessarily mean higher image quality . different fovs contain various voxel sizes and cbct devices also vary in voxel size for each fov . hassan et al , in 2010 showed that voxels larger than 0.6 , 0.9 , or 1.2 mm in the x and y plane significantly reduced the visibility of occlusal surfaces of the teeth , interproximal spaces between the teeth and alveolar bone ; whereas , the choice of larger voxels reduces the image noise . factors such as voxel size in imaging , exposure factors , various applied fovs , slice thickness , imaging systems and detectors are effective in diagnostic results . these parameters vary between the cbct units and different imaging protocols in the same unit . although short scanning time and low patient radiation dose are important factors in diagnostic imaging , risk of misdiagnosis and consequent complications should also be taken into account . in general , voxel size should be determined based on the severity of patient s condition and treatment plan . image quality variations can be reduced or increased by altering the fov , and the image reconstruction parameters and post - processing methods are studied based on the type of cbct system used . the selected fov for cbct images is directly related to voxel size and affects the resolution and contrast . extended fov creates lower contrast and resolution compared to smaller fov and thus directly affects the detection of anatomical structures on cbct images . in accordance with the results of the current study , fov position seems to have little impact on diagnostic specificity because the values were more or less the same in all three observers at five different positions . appropriate diagnostic accuracy of cbct images for bone defects in the fov and different resolutions in this study may be related to adequate ability and skills of the observers , good quality of cbct scans and lack of motion artifacts in images . in this study , bone defects caused by drilling had distinctive and appropriate form , enhancing their detection by the observers . also , kamburoglu et al , in 2014 reported the same performance of observers in detection of simulated buccal peri - implant defects using different parameters of fov and various resolutions . in another study , no difference was observed in diagnostic accuracy of cbct images for detection of simulated buccal peri - implant defects with and without pattern artifact reduction in planmeca promax system . pattern artifact reduction was not used in this study or in the study by kamburoglu et al , in 2014 . according to the results of the current study , the sensitivity for detecting bone defects by the observers and the frequency of detection was in the range of 0.251 in low resolution and 0.8751 in high resolution at different fov regions , showing that sensitivity was enhanced by increasing the resolution . on the other hand , the specificity value was between 0.75 and 1 for high - resolution images and 0.691 for low - resolution images . in the study by hedesiu et al , in 2012 , sensitivity for diagnosis of apical defects in cbct scans was estimated to be 0.720.8 and specificity was between 0.60.77 . however , higher levels ( a rate of 1 ) of sensitivity and specificity for cbct system were recorded in examining the human mandible by patel et al , in 2009 . the sensitivity parameter for cbct images in the study by patel et al , in 2014 was in the range of 79.2%91.7% . significant effects of resolution on sensitivity were not observed in the current study , which is different from the results of the above - mentioned studies . in total , the highest sensitivity was recorded in the central position in both high and low resolutions ; increased resolution generally led to improved sensitivity especially in the posterior region of the fov , but had no effect on specificity . the fov parameter in cbct should be adjusted to reduce patient radiation exposure for detection of bone defects . concerning some false - positive results in the negative cases , cbct should be used as an adjunct to clinical examination to make a correct diagnosis and for treatment planning . if the diagnostic purpose can be achieved by minimum radiation dose in the fov , there is no need to change this parameter or increase the absorbed dose of patient . improving the cbct image resolution from 0.3 mm to 0.2 mm increased the sensitivity values for detection of simulated bone defects . image resolution and position of the object in the fov seem to have little impact on specificity values . also , comparison of different fov regions revealed that the highest sensitivity rates were recorded in the central position in both high and low resolutions , and sensitivity significantly decreased in the posterior position at low resolution | objectives : this study aimed to assess the effect of voxel size and object location in the field of view ( fov ) on diagnostic accuracy of cone beam computed tomography ( cbct ) for detection of simulated bone defects.materials and methods : in this in vitro study , bone defects were drilled in four sections of a dry human mandible .
bone blocks were fixed on a platform parallel to the horizontal plane and cbct images were acquired using 0.2 mm and 0.3 mm resolutions and five locations of fov ( anterior , posterior , left , right and center ) .
three reviewers viewed the images twice and the presence or absence of simulated bone defects was determined in positive and negative cases.results:sensitivity in different locations of fov ranged between 0.251.0 and 0.751.0 in low and high resolutions , respectively .
these values were 0.6251.0 and 0.691.0 , respectively for specificity .
intra - observer agreements were in the range of 0.841.0 and 0.751.0 and inter - observer agreements were in the range of 0.30.61 and 0.460.69 in high and low resolutions , respectively .
the highest sensitivity was seen at the center of the fov and with an increase in resolution from 0.3 mm to 0.2 mm , the sensitivity increased specially in the posterior region of the fov while image resolutions and fov locations did not affect specificity.conclusions:the highest sensitivity values were obtained at the center of the fov and lowest values were seen in images acquired in the posterior region in low resolution . diagnostic accuracy improved with increased resolution . |
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stereotactic ablative radiotherapy ( sabr ) , which is also referred to as stereotactic body radiotherapy ( sbrt ) , is a form of high - precision radiotherapy ( rt ) . it is typically delivered in one to 8 intermittent fractions , taking up to 2.5 weeks to complete and commonly using fraction sizes of 7.5 - 20 gy or more . this compares with 20 or more fractions of 3 gy or less taking up to 7 weeks to complete that is typical of conventional rt ( fig . 1 ) . modern lung sabr is characterized by patient - specific planning techniques that account for individual tumor motion whilst delivering heterogeneous dose distributions to the tumor and sparing normal tissues ; accurate and reproducible imageguided patient setup prior to and during each fraction ; and ' beam - on ' radiation delivery times that can be 10 minutes or less with some techniques . sabr can be delivered using standard commercial equipment , and is readily incorporated into departmental workflows . local control rates of 90% or more are now possible with sabr for peripheral stage i non - small cell lung cancer ( nsclc ) measuring up to 5 cm in maximum diameter . the key to achieving this is believed to be a sufficiently high radiation dose delivered in only a few fractions , combined with accurate tumour targeting . these results contrast impressively with those of conventional rt using doses of at least 50 - 60 gy in fraction sizes of 3 gy or less , for which local failure rates of 40 - 50% and more can be expected . the combination of improved local control and an acceptable toxicity profile mean that in recent years , sabr has become the preferred treatment for patients with medically inoperable stage i nsclc in countries like the netherlands and japan . in addition to the institutional and phase ii studies already cited and a recent meta - analysis , there is now encouraging population - based data . in this latter study of patients aged 75-years or older , the introduction of sabr led to a 16% absolute increase in utilization of rt , a decline in the proportion of untreated elderly patients , and an improvement in overall survival . this was statistically significant only for the subgroup treated using rt , in which there was an estimated median increase in overall survival of 6 months , from 20 to 26 months , between cohorts of patients treated in 1999 - 2001 and 2005 - 2007 . the survival benefits observed in this population - based cancer registry are plausible as sabr has been shown to be well tolerated in elderly patients , and in those with severe chronic obstructive airways disease . both the baseline and post - sabr cohorts are from the period after which fluorodeoxyglucose ( fdg ) positron emission tomography ( pet ) scanning became widely available , increasing the likelihood that sabr may be responsible for the improved outcomes . although there have been suggestions that higher quality evidence is needed to confirm the improved therapeutic ratio for sabr over conventional rt before it enters routine use a randomized trial comparing these two treatment approaches seems unlikely ( table 1 ) [ 18 - 22 ] . indeed a convincing argument could now be made against such a trial on the grounds of a lack of uncertainty and clinical ( community ) equipoise . where sabr is not available or accepted , it should be noted that hypo - fractionated conventionally delivered rt is another option that might also deliver improvements in local control . for example , 48 - 60 gy in fractions of 4 gy has been associated with actuarial local control rates of 70% at 5 years . in the protracted debate over supportive evidence , it is appropriate to reflect on the seriousness of a diagnosis of lung cancer for today 's patients , who require access to the best currently available treatment . analysis of the surveillance , epidemiology and end results ( seer ) registry using data from elderly patients ( > 65 years of age ) diagnosed with stage i or ii nsclc between 1992 - 2002 who were not treated by surgery , found that if they also had no rt , their median survival was 7 months . this could be increased by 6 months , to 13 months , when they received rt , however only 59% of patients did so . a recent dutch analysis showed that 38% patients of elderly patients presenting with stage i nsclc in the period between 1999 - 2001 received no local therapy . for example , protracted courses of conventional rt may be perceived as ineffective , inconvenient and poorly tolerated by frail patients and their clinicians . furthermore , guidelines and training combined with advances in technology and quality assurance procedures may improve knowledge and increase confidence , facilitating the adoption of new techniques . a recent survey of sabr in the united states highlights other important drivers including research ( for academic radiation oncologists ) , and competitive reasons ( for those in private practice ) . however , despite such levers , introducing technology into some healthcare systems has been a major challenge . it is easy to overlook the fact that the process needs to be actively managed with advance planning , clear priorities and carefully considered implementation strategies . one concern expressed by some has been the wide range of dose - fractionation schemes reported in the lung sabr literature . however , there is now appreciable clinical data from a variety of sources to indicate that local control is compromised when a biological effective dose ( bed ) of less than 100 gy10 is delivered , calculated using an alpha / beta ratio of 10 gy [ 28 - 30 ] . data from our own center reports consistently high rates of initial tumor control across a range of ' riskadapted ' dose - fractionation schemes , all with a bed105 gy10 . caution is therefore required in advocating doses of less than 100 gy10 for peripheral lung sabr of targets up to 5 cm in diameter , especially when considering that such schedules are associated with low rates of significant toxicity . indeed , high - grade toxicity and mortality is uncommon following sabr for peripheral lung tumours , and rib fractures and radiation pneumonitis have generally occurred in less than 10% of patients ( although the former may depend on such factors as the proximity to the chest wall , dose - fractionation scheme and planning / delivery technique ) . damage to the brachial plexus has been reported in patients treated for apical tumors , particularly when plexus doses exceeded a bed of 100 gy3 ( i.e. , calculated using an alpha / beta ratio of 3 gy ) . it remains to be seen if the incidence of these complications will be reduced by the use of lower fraction sizes for selected tumors located closer to critical structures . such a risk - adapted strategy was adopted when our lung sabr program started in 2003 , taking in to account factors such as tumor size / treatment volume , the location of the target volume and the extent of contact with the chest wall . in some centers patients with tumors > 5 cm and also those that are centrally located are now being treated with risk - adapted sabr . follow up durations are currently limited , and it should be noted that the safety profile of sabr for central and to some extent larger tumors is likely to be different and so neither the clinical toxicity data , nor many of the treatment schedules for peripheral lung sabr should be directly extrapolated to these scenarios . one example concerns central airway toxicity , which has been reported after sabr for lung tumors extending into the mediastinum . the role of non - surgical treatment options in patients who are potentially fit to undergo surgery has increasingly been the subject of discussion . recent reports of sabr have included small numbers of patients who had declined to undergo surgery , and two prospective single - arm trials of sabr in patients who are fit to undergo surgery have completed accrual ( jcog 0403/nct00238875 and rtog 0618/nct00551369 ) . a markov model analysis of outcomes after either sabr or lobectomy for stage i nsclc for a 5-year time frame indicated that sabr might offer comparable overall survival and quality - adjusted life expectancy as compared with surgical resection . this is consistent with a matched propensity analysis from washington university of 57 high - risk surgical patients and 57 patients undergoing sabr for stage i nsclc , which found comparable local control and disease specific survival between the two groups at 3 years . importantly , both the 5- and 3-year time points used in these studies are considerably longer than the median time to loco - regional recurrence in patients who have had surgery for early - stage nsclc ( see below ) . current changes in demography mean that more than half of new patients with lung cancer are older than 70 years , and the absolute number of elderly patient is projected to increase faster than the increase in the overall population . extrapolating from a small sub - group analysis , sabr might be a particularly attractive treatment option for elderly patients as the benefits of adjuvant chemotherapy for patients with occult nodal metastases detected in stage i nsclc resection were not apparent . furthermore , even in a study where the median patient age was 64 years , only 66% of patients who were candidates for adjuvant chemotherapy post - surgery actually started chemotherapy . two issues that remain to be resolved in the coming years are techniques for improving methods for the diagnosis of peripheral lung tumours and for detecting local recurrence after sabr . obtaining a pre - treatment diagnosis in patients presenting with a peripheral lung nodule suspicious for a lung cancer can be challenging , particular as patients who are medically inoperable due to co - morbid disease , may be at higher risk for complications following a trans - thoracic needle biopsy . the likelihood of lung malignancy in this setting can be calculated using a combination of clinical , radiological and pet findings . applying this approach to sabr appears justified in a country such as the netherlands where a diagnosis of benign disease is typically made in less than 5% of patients undergoing surgery [ 46 - 48 ] . however this policy may not be well suited to populations with a higher likelihood of benign lung disease . indeed , in some populations the use of only a single computed tomography and pet scan without other supportive data can lead to nearly one third of resected pet - positive nodules turning out to be granulomas . actuarial distant failure - free survival rates of 85% and 77% were reported 1 and 2 years post - sabr . the rate of regional failure in patients staged with fdg - pet is generally around 10% . for patients who are fit enough for potential salvage options , however post - sabr pulmonary changes may be complex and can sometimes be hard to distinguish from tumor progression . this necessitates careful patient follow - up and access to specialist multi - disciplinary teams who are familiar with interpreting post treatment imaging . at the present time , one observation that may help to discriminate local recurrence from post - sabr change is that the former may be characterized by the rapid enlargement of a mass within a relatively short period of time after lung sabr . this was the case in recent reports describing experience with salvage surgery and is consistent with the timing of local - regional failure after surgery . for example , a retrospective study of 975 patients undergoing surgical resection for pathological stage i / ii nsclc between 1995 - 2005 found that the median time from surgery to loco - regional failure was 13.9 months . data such as these may help to guide optimal follow up strategies , however they need to be complemented by improvements in imaging and image analysis to discriminate between recurrent tumor and post - sabr changes . the feasibility of surgical salvage as a treatment option for recurrences post - sabr requires further study , particularly as it may increase the preference for sabr in some patients who are fit to undergo primary surgery . looking ahead , there are now a number of randomized clinical trials comparing sabr and surgery but the results will not be available for many years to come . the use of lung sabr for patients with medically operable disease generates discussions about the relative risks of the two interventions . in a recent population based analysis from the netherlands , lung sabr for medically inoperable patients was associated with a 1% 30-day mortality when it was largely delivered in only 2 regional centers . this is similar in magnitude to the post - operative 30-day mortality reported in the acosog z0030 study of pulmonary resection for early - stage lung cancer in medically operable patients . while results from individual centers can be expected to vary , a population - based study using seer data has linked an increased risk of operative mortality to several factors , including increased age and certain comorbidities . this emphasizes the importance of risk stratification and both expanding and individualizing the therapeutic options for all patients with lung cancer . the leading role played by the surgical community in studying the impact of practice organization and operating protocols on patient outcome is well recognized and as lung sabr diffuses into routine clinical practice throughout academic and community centers it will also become important to learn from this experience and consider similar issues in radiation oncology . as survival in patients with stage iv nsclc has historically been considered to be poor , low - dose palliative rt has been the standard of care in patients with symptomatic disease . justification for such an approach was found for example in the outcome of a large phase iii trial comparing 4 different third - generation platinum - based chemotherapy schemes . this eastern cooperative oncology group study reported a response rate for all 1,155 eligible patients of 19% , with a median survival of 7.9 months , a 1-year survival rate of 33% ( 95% confidence interval , 30 to 36% ) , and a 2-year survival rate of 11% ( 95% confidence interval , 8 to 12% ) . such results indicated that only a minority of patients would present with recurrent disease requiring re - irradiation , and if needed repeat low - dose palliative rt was the prevailing policy . a change in thinking regarding the management of metastatic nsclc lung cancer came about with several publications , including those dating from the 1990 's when long - term survival was noted selected patients who underwent resection ( or rarely rt ) for limited volume metastatic disease , in combination with radical management of the primary tumor . other studies have also suggested an advantage for ablative management strategies in selected patients with advanced nsclc . in patients who present with newly diagnosed nsclc and synchronous brain metastases , a retrospective study in 167 patients from our own institution has suggested that in a subgroup of selected patients aged 65 years or younger , who were also eligible to undergo surgery or radiosurgery for their brain metastases , radical treatment for their thoracic disease may be justified . this strategy is indirectly supported by a systematic review of randomized controlled trials of palliative thoracic rt which concluded that improvements in survival were more likely with higher - dose rt to the primary tumour ( bed35 gy10 ) . further support is also provided by the observation that the predominant pattern of failure in patients with advanced nsclc after first - line systemic therapy may be at the sites of known disease . in keeping with the view that cancer is a spectrum , it has been hypothesized that the most relevant issue may not be whether or not there are distant metastases , but instead what the ratio is between local and systemic disease . in this case , it is possible that the impact of chemotherapy may be fairly similar for tumours that are either primarily localized or systemic , whereas the benefit of adding chest rt could increase as the proportion of tumour cells that are localized increases . many institutional reports now show surprisingly long survival outcomes in ' definitively managed ' patients with metastatic nsclc , and recent guidelines , including those from the british thoracic society , support the need for clinical trials of radical treatment for patients with m1 disease . at present , several non - invasive treatment techniques , including sabr can be used to ablate metastases . reports evaluating sabr in the treatment of metastases to the liver , adrenal gland ( fig . 3 ) , have reported high rates of local control , sometimes in excess of 90% when ablative doses of rt have been administered . together with recent developments in sabr , this has lead to a resurgence of interest in the treatment of oligometastatic disease , a state characterized by a limited number of clinically apparent metastases with the possibility that these might genuinely represent the only distant sites of disease . such treatment paradigms support a move away from dichotomous definitions of treatments as curative or palliative , encouraging a more useful spectral interpretation of interventions and their potential for life - prolongation , as well as debates about how best to integrate an increasing range of possible therapies . one strategy , the identification of molecular signatures that can augment clinical features , is a priority . in recent randomized trials , treatment of nsclc with activated mutations of the epidermal growth factor receptor ( egfr ) has shown response rates in excess of 60% , and median survival exceeding 24 months . the identification of this and other molecular subtypes of stage iv disease , where prolonged disease control can be attained using targeted agents is now rapidly transforming the systemic treatment of stage iv nsclc . studies evaluating the additional role of ablative therapies in such patients with limited volume oligometastatic disease are a logical step ( table 2 ) , including in east asian populations who have a higher frequency of egfr ( and other ) mutations . ultimately , one aim of such treatment paradigms would be to achieve long - term survival , although a less - controversial first step may be to prolong life while also preserving quality and function . a number of issues need to be resolved before larger - scale trials can be initiated . phase i - ii data is required to establish the toxicity of both sequential and concurrent administration of molecularly targeted agents and high dose rt , particularly as unexpected toxicities have been encountered . in addition , innovative and careful trial design is likely to be necessary to increase the chance of successfully answering important clinical questions . simultaneously , it is also necessary to continue to refine treatment techniques and quality assurance strategies that support the safe , efficient and effective delivery of sabr to single and multiple targets [ 79 - 81 ] . in summary , at the present time the available data justify the use of ablative therapies such as sabr for treating selected patients with early - stage and advanced nsclc . while the history of surgical resection for colorectal cancer liver metastases suggests that efforts to radically change existing treatment paradigms in nsclc are likely to encounter scepticism and require time , for certain indications , in particular lung sabr for early stage peripheral nsclc in medically inoperable patients , the field may already have moved beyond the need for randomized controlled trials . while there is a legitimate debate to be had over the appropriate methodologies and standards of evidence with which to assess clinical interventions and advanced rt technologies , this should not overshadow the needs of today 's patients , nor should it deter immediate and pragmatic steps to responsibly integrate promising treatments and useful technologies into clinical practice . | a compelling body of non - randomized evidence has established stereotactic ablative lung radiotherapy ( sabr ) as a standard of care for medically inoperable patients with peripheral early - stage non - small cell lung cancer ( nsclc ) .
this convenient outpatient therapy , which is typically delivered in 3 - 8 fractions , is also well tolerated by elderly and frail patients , makes efficient use of resources and is feasible using standard commercial equipment .
the introduction of lung sabr into large populations has led to an increased utilization of radiotherapy , a reduction in the proportion of untreated patients and an increase in overall survival . in selected patients
, the same ablative technology can now achieve durable local control of nsclc metastases in a variety of common locations including the adrenal glands , bone , brain , and liver . at the same time as this , advances in prognostic molecular markers and targeted systemic therapies mean that there is now a subgroup of patients with stage iv nsclc and a median survival of around 2 years .
this creates opportunities for new trials that incorporate sabr and patient - specific systemic strategies .
this selective mini - review focuses on the emerging role of sabr in patients with early - stage and oligometastatic nsclc . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
there are two basic
strategies for constructing devices in which
the flow of electrons is mitigated by single molecules : top - down and
bottom - up . top - down methods rely on nanofabrication or mechanical
control to form the nanometer - scale junctions between top
and bottom electrodes necessary to contact molecules
end - to - end . bottom - up methods typically rely on the self - assembly
of molecules onto a bottom electrode in which the junction is defined
by the molecules themselves when a top electrode ( top - contact ) is
applied ; these are devices that , in part , fabricate themselves . nature provides a plethora of intricate molecular
complexes that form by and are prone to self - assembly ; however , these
complexes are subject to different constraints than their simpler ,
synthetic counterparts such as the well - studied alkanethiols . conventional techniques for forming top - contacts
for devices or for measuring electrical properties over large areas
of self - assembled monolayers ( sams ) of organic molecules have been
thus far limited to the direct deposition of metals by electron - beam or thermal evaporation or the addition
of an electrically conductive polymer layer between the sam and a
metallic top contact ; however , in these techniques ,
the high temperatures , vacuum processing and the need for acidic or
organic solvents to spin coat the polymers are not ideal for biomacromolecules . other , soft , nondamaging methods for forming top - contacts ,
such as the use of hanging hg drops ( hmds ) ,
are better suited to this task . ron et al . used hmds as a nondamaging
method for forming top - contacts to measure the electron transfer ( et )
through three protein ( complexes ) on doped - si substrates : azurin ( az ) ,
bacteriorhodopsin ( br ) , and bovine serum albumin ( bsa ) , establishing
that proteins can function as building blocks for bottom - up tunneling
junctions . although nondamaging , hg is better suited for si substrates
than metals , with which it tends to form amalgams ( au and ag are the
most common metals used in bottom - up tunneling junctions ) . these studies
on complete proteins were preceded by discussed models of mechanisms
of electron transport through peptides and proteins , for example ,
by giese et al . waleed shinawari et al . reviewed biomolecular charge transport , with
a particular focus on dna and protein molecules . a recent progress
report by amdursky et al . compares current
densities for different techniques , including cp - afm and scanning
tunneling microscopy ( stm ) and macroscopic , for junctions comprising
proteins and small molecules . the liquid metal eutectic , ga in
( which we abbreviate egain ;
75% ga , 25% in by weight , mp = 15.5 c ) , can function similarly to hg in the hmd method , but does not form
alloys with noble metals . it also exhibits
non - newtonian rheology , driven by a self - limiting oxide , and does not spontaneously reorganize to minimize
its surface energy . thus , it can be formed
into stable , nonspherical tips to form 100 m contacts and is stable in microfluidic channels , which facilitates
variable - temperature measurements . photosystem i ( psi ) is a complex that houses
one of the two reaction
centers used in the photosynthetic reactions in cyanobacteria , algae
and plants where the conversion of solar energy to electron hole pairs
takes place . isolated psi complexes from thermophilic unicellular
cyanobacterium thermosynechococcus elongatus bp-1 have evolutionally formed a trimer structure for improved
light absorption efficiency and stability at harsh conditions . the
monomer of psi has a polar stroma and lumen and an apolar backbone . its size is approximately 13 8 9 nm and contains 96
light sensitive chlorophyll a ( chla ) molecules that are densely packed in the protein scaffold to harvest light . in the complex , harvested
photons are converted into excited electrons by chlorophylls at p700
( via a special pair of chla molecules ) . this transfer
occurs from the primary electron donor complex to the primary electron
acceptor a0 ( chla ) , to the ferredoxine
docking region through the built - in electron transport chain a1 ( phylloquinone ) , fx , fa and fb ( fe4s4 clusters ) ( figure 1a ) . the complex has a photovoltage of 1 v under illumination
with an internal quantum efficiency close to unity.these features make psi a unique and interesting protein complex
to study . furthermore , the entire psi complex , as well as the reaction
center ( rci ) from plants and cyanobacteria have been extensively characterized
and the electrical properties have been observed by a variety of experimental
techniques . we studied the entire psi complex , not isolated rci , because it
contains the electron transport chain , the light - harvesting array
of chlorophylls and the protein scaffold that holds them in place . ( a ) schematic
representation of electron transfer chain in photosystem
i ( et chain ) . ( b ) possible orientations of psi on au surfaced induced
by chemical modification . orientation where the flow is in the opposite direction ( p700 adjacent
to substrate ) , a down orientation in which the natural
direction of the flow of electrons is toward the electrode surface
( fb down ) , or with its electron transport vector parallel to the substrate . we investigated the j v characteristics
of sams of psi in the dark on template - stripped au measured by forming top - contacts from tips of egain . egain
has been used to measure current density ( j ) versus
applied voltage ( v ) through sams of a variety of
molecules to form rectifiers , to observe quantum interference effects , to relate dipole moments to vacuum - level shifts , and it is sensitive enough to resolve the odd even
effect in sams of alkanethiolates . unlike sams
of small molecules , however , there are no data from egain tunneling
junctions of psi or any other protein complexes against which to compare .
thus , we characterized sams of psi trimers by interrogating the complexes
in the dark , one at a time , by cp - afm and compared those data to the
ensemble averages produced by egain contacts in order to relate our
findings to previous studies . we use the asymmetry of the conductance
data ( i.e. , rectification ) to elucidate the orientation of psi because
it is self - referencing and , therefore , is less sensitive than the
magnitude of j to other structural features of the
sam , but very sensitive to the specific geometry of the sam / egain
interface ( e.g. , orientation ) . asymmetry
observed in current transport is particularly useful for elucidating
the transport properties of protein complexes , which are vastly more
complex in structure , size , self - assembly and electrical properties
than small molecules , because it eliminates the uncertainty of area
calculations that arises from topology , packing density , etc . and while virtually all tunneling junctions
with egain top contacts show some asymmetry ( because the junction
itself is asymmetric ) , the rectifying behavior of the sam is readily
distinguishable . we are not suggesting
that sams of psi make particularly good rectifiers of current the
magnitude of the asymmetry of the current is quite small only
that small changes in asymmetry can provide information about the
orientation of psi complexes . orientation
in which the natural flow of photogenerated electrons is to the electrode
surface ( fb down ) , an up orientation where
electrons would flow in the opposite direction ( p700 adjacent to substrate ) ,
or with its electron transport vector parallel to the substrate ( figure 1b ) . this level of control over the orientation of
the electron transport chain provides an opportunity to determine
its role in the tunneling transport through psi through the self - assembly
process rather than by modifying the complexes themselves . different
methods have been used to control the orientation of psi and rci on
surfaces , ranging from surface modification with different functional
groups that interact electrostatically with different parts of the
protein complex , to direct covalent attachment via mutation and sams with different functional head groups . with egain , we can address the monolayers of psi complexes electrically
and from these electrical measurements we can probe the average orientation
of the complexes in the monolayer to compare against afm images and
cp - afm i v data . according to previous
reports on single complexes of rci ( i.e. ,
not sams or ensembles and not psi ) , asymmetric charge transport is
completely dependent on the orientation on the surface of the electrode . they platinized one end of the photosynthetic complex
and welded it to a au surface using sams of small
molecules as director monolayers . they used scanning tunneling spectroscopy ( sts ) to examine
the electronic properties of each rci , which elicited orientation - dependent
asymmetry . this phenomenon is easily conflated with light - driven processes , which involves hopping transport ( to move through
the transport chain as the electron changes in energy ) and therefore
should not play a role in tunneling measurements . yet , the absolute
orientation of rci and , by extension , psi is assigned from sts data
based on the assumption that the direction of asymmetry ( rectification )
follows the electron transport chain . by measuring
tunneling junctions comprising psi in the dark , we found that this
assignment of the orientation does not , in fact , map onto psi it
is backward and that tunneling transport , therefore , likely
dominates any hopping contribution from the electron transport chain . we exposed ultraflat
template stripped
au ( au ) substrates to a solution of 1 mm 2-mercaptoethanol
( 2me ) or sodium 3-mercapto-1-propanesulfonate ( mps ) to form director
sams to bias the orientation of psi trimers that self - assemble
on top of these sams . these director sams differ in length by 1.7 , which may affect
the magnitude of the tunneling current , but we do not rely on this
magnitude as a measure in this work and small changes in the thickness
of the director sam are unlikely to influence asymmetry . the psi in
cyanobacteria has an asymmetric distribution of surface charges in
the stroma and lumen ( see supporting information for details ) . two thirds of all the charged surface residues are
concentrated at the top of the complex ; the stromal ,
fab electron acceptor side ( figure 1a ) . this difference is likely what determines the preferred orientation
during self - assembly on modified surfaces . we immobilized psi on the
substrates by drop casting from aqueous buffer and incubating them
( i.e. , leaving them in contact ) for 2 h. we investigated each sam
topographically by afm and electrically using cp - afm and egain . the
cp - afm data are averages of i v curves from
100 complexes for each director sam . thus , they sample the distribution
of orientations similarly to the large - area egain measurements and
can be compared directly . we define the asymmetry of current transport
( rectification ratio ) , r , as the ratio of j or i at positive to negative bias ; r = |j()/j(+)|
for egain and r = |i()/i(+)| for cp - afm , both with respect to the wiring convention
for cp - afm ( see supporting information ) . measured afm height profile images show better psi coverage for
2me than for mps , with protein heights close to 6 nm on au . this value is less than the 9 nm thickness derived from the crystal
structure data because these sams are measured under ambient , anhydrous
conditions and contact with the afm tip can compress them somewhat
( for more details see supporting information ) . we imaged individual psi trimers within sams of psi formed on
both directing sams by afm to determine the density ( per m ) and measured their electrical properties by cp - afm to determine
the orientation of the complexes , assigning low values of r to psi that is oriented up and high values to down . values
of unity were assigned to psi that is oriented parallel to the substrate
( sideways ) . ( the average values of r for egain and cp - afm
are given in table 2 . ) individual i v traces were averaged over 100 psi complexes at different locations
within the sams . the results
were obtained by
measuring the asymmetry of the i v curves
by cp - afm . the average coverage on an area of 1 m was calculated from afm height images . the table summarizes the
statistical distributions of the i v curves
of over 100 points for both director sams . these values were calculated from
evaluating j v curves at
1 v. r = |j()/j(+)| for egain and r = |i()/i(+)| for cp - afm , both with respect to
the wiring convention for cp - afm ( see supporting
information ) . values refer to the calculated geometric mean
from curves at 1 v. for egain measurements , the sams of psi on both director sams were
contacted by lowering a syringe ( connected to an electrometer ) supporting
a tip of egain . recent reports on egain junctions comprising alkanethiolates
employ a slightly different method in which the tips are first flattened
against a si wafer and then pressed into the sam ; however , light contact with as - prepared tips yielded stable
and reproducible results on sams of psi . we observed four different
behaviors : ( i ) shorts , characterized by linear j v curves with currents in the a regime ; ( ii ) no - contact , characterized
by noisy currents in the pa regime ; ( iii ) poor - contact , which begins
with no - contact j v curves , but is followed
by shorts after further lowering of the egain tip ; and ( iv ) good contact ,
characterized by s - shaped j these data are summarized in table 3 and show higher rectification ratio when 2me is used as opposed
to mps , which we ascribe to the different distributions of orientations
of psi induced by the director sams . we analyzed the data identically
to the cp - afm data by averaging log j. see the supporting information for a detailed discussion
of the data analysis . the
values of j are the geometrical averages from measurements
with egain tips . the yield of working devices is based on devices with egain as top
electrode . as control experiments , we measured
the electrical properties of
denatured psi ( which are shown in the supporting
information ) . we boiled psi for 20 min at 99 c , which
is sufficient to denature it completely and then prepared sams of
deactivated psi by following the same procedures used to prepare sams
of active psi . we observed no rectifying behavior in egain junctions
comprising boiled psi , nor did we observe shorts ( table 3 ) . the lack of shorts implies that denatured psi still covers
the substrate , as sams of 2me and mps are fragile and give mostly
shorts . thus , the origin of the asymmetry is dependent on the presence
of intact psi structures ( i.e. , the overall asymmetry of the junction
depends on the presence of intact complexes of psi ) . we varied
the density ( surface coverage ) of the sams of psi by preparing psi
trimer solutions of different concentrations ( dilutions from 1:1 to
1:130 , ratio of concentration of 1 m psi to buffer ) while keeping
the incubation time fixed . from afm images , we determined the surface
coverage by comparing the psi - covered surface to the total surface
area ; psi= [ napsi / atotal]100% where n is the number density of psi , apsi is the area occupied by psi complexes ( 25 nm , determined from tem ) , atotal is the total area investigated . figure 2 shows the correlation between psi and the magnitude of j in egain junctions . the
exponential relationship between j and surface coverage
is further evidence that the charges are flowing through the psi complexes .
below 23% coverage the junctions become unstable , irreproducible ,
and yield mostly shorts ( as do bare director sams ) . presumably this
percentage is the cutoff value below which egain is able to penetrate
between the psi complexes and contact the director sam . this behavior
is not unlike defect - mediated transport in which the electrical properties
of highly conductive defects ( space between psi complexes ) become
dominant at a critical density . the saturation
of j at 23% implies that transport occurs exclusively
through psi and that there is no ( or constant ) leakage current , which
allows the direct comparison of the data from egain and cp - afm junctions . asymptotic
fit of j at 1 v vs percentage of surface
coverage for psi on 2me as directing sam . the threshold limit of coverage is 23% as determined
by imaging surfaces at different psi : buffer concentrations and characterizing
their behavior with an egain tips . inset images show psi complexes
on au at different concentrations which correspond to
the percent coverages shown . the devices were imaged on an afm at
2.5 m . to evaluate the influence
of contact with tips of egain , we marked
areas of the sams of psi and imaged them by afm before and after forming
egain junctions and acquiring j we observed no qualitative
damage to the monolayers and , by counting the number of complexes
in each junction before and after forming egain junctions , determined
that no complexes were extricated from the monolayer . from these experiments we conclude that the sams of psi are not damaged and that the individual
complexes do not move and are not removed during measurement . thus ,
egain is a demonstrably nondamaging method for investigating charge - transport
through sams of psi and is capable of forming reversible junctions ,
which may be useful for characterizing sams of psi ( or other protein
complexes ) as an intermediate step in the fabrication of devices to ,
for example , verify the orientation of the complexes . the rectifying behavior of rci
( not psi ) was previously observed by sts and cp - afm on individual protein complexes .
in our studies with egain , psi assembled on director sams exhibited
an asymmetric conductance between bias voltages with r = 5.0 with 2me and 2.0 with mps ( table 2 ) . the direction of rectification does not invert when the orientation
of the complexes is reversed because there is some built - in asymmetry
in these junctions ; the bottom electrode supports a covalently bound
director sam that supports the psi complexes , while the top - contact
is physisorbed either via contact with an afm tip or by supporting
an egain electrode . thus , one orientation works with the built - in
asymmetry and the other against it , but not sufficiently to overcome
it completely . we ascribe the asymmetry in both cases to the
ratio of the orientation of the complexes ( table 1 ) . an important distinction between single - complex studies
( e.g. , cp - afm and sts ) and large - area studies ( e.g. , egain and hmd )
is that the latter sample the average orientation . coupled with the
ability to measure many locations across many different substrates ,
large - area measurements are useful for characterizing the self - assembly
of psi ( or any protein with direction - dependent rectification ) . additionally ,
the nondamaging nature of egain makes it a useful tool for investigating
sams of psi during the fabrication of a thin - film device , for example ,
a photovoltaic device , the properties of which are dominated by the
average orientation of psi . single - complex measurements provide
details that are difficult or impossible to extract from large - area
measurements , thus the combination of the two gives a complete picture ,
capturing the details of transport through individual complexes and
the supramolecular structure of the sam . although it is clear
that the relative orientation of rci and psi
is influenced by the chemistry at the surface of the bottom electrode
( au in our case ) , the absolute orientation of either
has not been determined unambiguously . it was observed that rci preferentially
platinizes at one side , which was assumed to occur at the more polar
( electron accepting ) side of the complex . despite the lack of direct evidence , this one study has become the
reference point for the absolute orientation of rci and , by extension ,
psi . our data were acquired on sams of psi , which has a different
electrostatic profile than rci and therefore does not necessarily
orient identically to rci . the first
is that charges ( electrons or holes ) take advantage of the electron
transport chain or directly traverse it ; both mechanisms involve thermally
activated hopping processes . these mechanisms ( it is not clear specifically
which ) are used to infer the absolute orientation of rci and predict
higher values of r when psi is oriented up . the second
mechanism assumes that tunneling charges do not hop through electron
transport pathway and the rectification is instead driven by the large
dipole moment of the whole psi structure between the luminal and the
stromal surface ( i.e. , perpendicular to the substrate when psi is
in the up or down orientation ) . van haeringen et al . used linear dichroism
to elucidate the dipole moment within the structure of trimeric psi . the direction of the dipole is parallel to the
c3-symmetry axis of the protein trimer complex , with positive side
of the dipole moment on the luminal side and negative on the stromal
side . the dipole effect within psi trimers was also observed on solid - state
bulk heterojunction ( bhj ) solar cells with a work function shift . ( there , a monolayer of psi trimers was self - assembled
at a metal - oxide electrode rather than au . ) the electric field of
this dipole moment can either enhance or oppose the total field created
by biasing the electrodes as is shown in figure 3 . although we have no way to observe the absolute
up / down orientations of psi directly , we can assign the absolute orientation
by determining which mechanism ( the electron transport chain or the
internal electric field ) is more likely . first , however , we must establish
that cp - afm and egain are in agreement in order to relate our observations
to previous reports ( which exist for cp - afm , but not for egain ) . direction
of the electrical field ( dashed lines ) that arises from
the psi dipole moment within psi - egain devices , which are shown with
egain biased positively ( with respect to the normal wiring of egain ) . ( a ) when psi is oriented up , the electric
field from the applied bias opposes the internal electric field of
the psi complexes . the direction of the internal electric field is the same as the applied
bias . thus , this mechanism predicts that psi in the down orientation
will give higher values of r. although
the influence of the native oxide layer on egain has been thoroughly
studied on aliphatic sams and has been shown to have a negligible
influence on transport properties , it
is necessary to confirm that the same holds true for protein complexes .
due to the size of complexes of psi , we are able to isolate individual
complexes for cp - afm measurements , meaning that the calculation of
per - complex resistivity is unambiguous . for egain junctions , we know
the density of psi complexes and the measured contact area , ageo , but not the actual contact area , aeff , which is considerably smaller due to the
topology of the egain tip thus , we can
calculate the number of complexes in ageo , compute the resistance and determine the correction factor to relate ageo and aeff . if
the oxide is benign in this study , this correction should be comparable
to the values reported for aliphatic sams . for a given egain
junction , we considered every oriented psi complex as a resistor in
parallel . we calculated the resistance of individual complexes ( ri ) from average values of i from cp - afm measurements at 1 v ( range of 10 to 10 ohms depending on the orientation ) . the
resistance , robs , is the total resistance
of the circuit and , at v = 1 v , is the reciprocal
of the current calculated for an area with n number
of complexes , allowing the calculation of n from
eq 1 where iegain is the measured current of an egain / ga2o3//psi
junction.1 the values of n given by eq 1 are 4.5 and 2.2 10 for 2me and mps , respectively .
using these values of n , we calculated aeff from the densities of psi complexes shown in table 1 . from the measured value of ageo , eq 2 gives the overestimation of
the area , .2 from this calculation we find 10 ,
which is consistent with the values of 10 that
have been observed for aliphatic sams . thus , using
the per - complex conductivity determined by cp - afm and the magnitude
of the current obtained from large - area measurements with egain , we
arrive at a value of that is within a factor of 10 of previously
reported values . this result shows that the contact resistance associated
with the physisorbed electrode - psi interface is comparable for egain / ga2o3 and a pt / ir cp - afm tip and , therefore , that
the influence of the oxide layer on charge - transport is negligible
( i.e. , it does not contribute to robs more
than a cp - afm tip ) ; cp - afm and egain data are comparable . this results
is unsurprising in the context of existing studies on sams of alkanethiolates , but it is necessary to establish the benign
nature of the oxide specifically in junctions incorporating proteins . using our computed value of , we plotted the per - complex
current - densities from cp - afm and egain ( figure 4c , d ) . the values of r from egain are in agreement
with the values of r from cp - afm data , but figure 4c , d also shows a qualitative agreement between the
line shapes . v , j v curves with respect
to the original wiring for cp - afm and egain , respectively ( figure 4a , d ) . top : semilog plots of current and current density
versus voltage
for junctions measured using cp - afm ( a ) and egain ( b ) for sams of
psi on mps ( purple ) and 2me ( black ) the black arrows indicate the
orientation of the psi complexes on the surface . these data are plotted
according to the normal wiring of each technique ( see the supporting information for details ) . bottom : per - complex j v curves for sams of psi on 2me ( c ) and mps ( d ) measured by cp - afm
( black squares ) and egain ( green circles ) plotted with respect to
the standard wiring of cp - afm ( shown in the insets ) . per - complex values
of j for egain were calculated using number densities
of psi measured by afm and a correction factor for the difference
between the measured and effective area of the egain junctions . while egain junctions can not be
scanned past 1 v without
precipitating shorts ( from electrostatic pressure ) , cp - afm junctions
can be scanned further because the height of the tip is fixed by the
instrument rather than the sam of psi . thus , while the cp - afm and
egain data are in remarkably close agreement at 1 v , we can not
know for certain that they would not diverge at higher potentials . cahen
and co - workers have shown , using hmds , that the electrical properties
of junctions comprising metalloproteins are affected by the removal
of the metal centers . these studies show
that tunneling electrons can take advantage of the accessible states
of the metal centers . in studies of rci ( not psi ) where rectification
is observed , the mechanism is almost always ascribed to the electron
transport chain ; however , no evidence is offered to support or refute that hypothesis . previous studies have shown that , when the electron transport chain
is deliberately engaged , the rate of electron transfer in rci is higher
from p700 to fab ( forward ) than in the reverse direction , which could explain the diode - like behavior seen in the j v curves . the two
most straightforward experiments for establishing the mechanism are
removing the electron transport chain and variable - temperature studies . the former is not possible without substantially influencing the structure
of rci / psi , and the latter is not possible with the techniques that
have been used to study rci thus far . fortunately , the ( remarkable )
robustness of sams of psi in vacuo extends
to sufficiently low temperatures to enable variable - temperature studies
using microfluidic channels filled with egain in a crossbar configuration . one of the most robust rectifying tunneling
junctions comprises ferrocene - terminated sams on ag with egain top - contacts . the mechanism of rectification in these sams is the
( partial ) pinning of the homo of the ferrocene to egain , which pushes
it either into or out of resonance with a ag electrode
depending on the sign of the applied potential on egain ; when it moves
into resonance ( negative potential ) , charges tunneling onto the homo
and then hops onto the egain , shortening the effective tunneling distance . this mechanism was proven by arrhenius plots ( ln |j| vs 1/t ) , which clearly show the freezing
out of the hopping component , leading to the loss of rectification
at temperatures below the activation energy of the hopping process . if the mechanism of rectification in psi involves
thermally activated hopping processes in the electron transport chain ,
it should freeze out as well . thermally activated transport processes
have also been observed in cp - afm studies of long conjugated molecules and proteins such as azurin , ferritin , and cytochrome at
elevated temperatures . we did not collect transport data above room
temperature , but at low temperature we observed comparable results . thus , there may be temperature - dependent transport pathways at elevated
temperatures , but they do not contribute to the asymmetric transport
at room temperature . if the mechanism of rectification in our
psi junctions involves
the electron transport pathway then it must also involve a hopping
process , as the electrons change energy inside the complexes ; moving
with the electron transport chain ( downhill ) then leads to higher
values of j than against it ( uphill ) at a particular
value of |v| . if , however , the mechanism of rectification
is the interaction of the applied field with the built - in field of
the collective dipole moments of the psi complexes , the process is
entirely tunneling ; the rectification arises from
the different probabilities of tunneling from left to right and right
to left . the former mechanism , therefore , will show a loss of rectification
at low temperatures as the hopping processes are frozen out , while
the latter will be completely independent of temperature . we
fabricated microfluidic devices following literature procedures
and acquired j v traces at different temperatures . an arrhenius plot of ln |j| at 0.50 v for psi
on both directing sams is shown in figure 5 ( the raw data are shown in the supporting information ) . the magnitude of r , shown by the difference in
the magnitude of j at positive and negative bias ,
is invariant with temperature . the values of ln |j| are also nearly invariant , showing only a slight perturbation only
near room temperature . these data are unambiguous evidence that the
mechanism of charge transport through sams of psi on mps and 2me is
independent of temperature and , therefore , that the mechanism of rectification
does not involve hopping and/or the electron transport
chain . from this conclusion , we can ascribe an absolute orientation
of psi ; it is oriented down ( p700 adjacent to the
egain substrate ) on 2me and up on mps . thus , the direction
of rectification is exactly opposite to the natural flow of electrons
through the electron transport chain , which predicts higher currents
at positive bias ( with respect to egain ) when p700 is adjacent to
the egain electrode . an energy level diagram based on nakamura s
et al . it not only predicts that asymmetry will be more
pronounced in the up orientation , but that r <
1 . note that this diagram is drawn with respect to the normal wiring
of egain and is , therefore , backward from the wiring convention of
cp - afm . ln |j| at 0.50 v as a function of inverse
temperature for psi on directing sams of mps ( triangles ) and 2me ( squares ) . the solid symbols ( , ) represent the positive bias
( + 0.50 v ) and hollow ( , ) represent the negative bias
( 0.50 v ) . the linearity indicates that the mechanism of charge
transport is dominated by tunneling as no temperature
dependence was measured . the entirety of the curves for biases from
1 to 1 v can be found on the supporting
information . the barrier width
is defined by the thickness of one oriented psi complex , which is
depicted in the down orientation with respect to the
natural direction of electron flow . the green lines are the frontier
orbital energies of the chlorophyll molecules , which are distributed
evenly through the thickness of the psi complex . the black lines represent
the energies of the electron transport chain and their relative spatial
positions . on the basis of the orientation of the electron transport
chain , more current should flow when the egain electrode is biased
negatively than at the equivalent positive bias . that mechanism would
translate into higher values of r when the complexes
are oriented up ( because this figure is drawn with
respect to the wiring of egain junctions ; figure 4 shows the data with respect to the wiring diagram of cp - afm ) . however , in our experiments we observe more current at the positive
bias , which supports our dipole moment hypothesis . the distances between
cofactors were estimated with the software pymol from a crystal structure
of psi taken from the protein data bank ( 1jb0 ) . a common test for tunneling transport is to compare the decay
coefficient ,
, against literature values . this value is obtained from simmons
approximation , j = j0e , where d is distance between the electrodes and is obtained
by varying d. this study is not possible with psi
because d is fixed by the complex at 6 nm . estimated by comparing i in the presence
and absence of a protein , arriving at values of 0.18 , 0.12 , and 0.27
for three different proteins on si surfaces
modified with octadecyltrimethoxysilane . using the same analysis , we arrive at a value of 0.16 for psi on
2me and 0.08 for psi on mps with d = 60 and values of j taken from
table 3 . to gain more experimental insight
into the mechanism of rectification ,
we measured the transport properties of bovine and human serum albumin
( bsa and hsa ) , which have been studied extensively . we observed r 5.5 and 3.0 on 2me and mps , respectively ( see supporting information ) . this observation can
only be reconciled using the mechanism of rectification proposed in
this paper , given that sams of bsa and hsa show preferential ordering and that they contain only alpha helices , which
contribute to a collective dipole moment . it also explains the observation
by ron et al . that bsa rectifies on br - terminated si surfaces using
hmd electrodes . we have established a clear relationship between the average orientation
of psi ( not rci ) in sams and the asymmetry ( rectification ) of current
in metal / protein / metal junctions on template - stripped au surfaces
modified with directing sam . asymmetry is a useful observable for
capturing the complexities of assemblies of large , biological molecules ,
particularly in combination with single - complex measurements , which
are insensitive to collective properties , but which provide details
that are missed by large - area methods . the average orientation is
affected by the identity of the directing sam used to control the
surface chemistry of the substrate . through variable - temperature
measurements we have established that
the dominant mechanism of charge - transport through sams of psi on
au is likely tunneling and that , at the very least ,
it does not involve thermally activated transport . this observation
refutes the hypothesis that rectification is due to the natural direction
of the flow of electrons through the transport chain , which involve
thermally activated processes , and instead is likely the result of
the internal electric field that arises from the collective action
of dipoles ( and multipoles ) in the peptide backbone . it opposes the
natural direction of electron flow through the electron transport
pathway and therefore predicts that 2me preferentially orients psi
down and mps up with respect to the
direction of the flow of electrons through the transport chain in
vivo ; i.e. , with p700 adjacent to the egain substrate . this assignment
of the orientations of psi is in agreement with solid - state , thin - film
devices comprising psi ( not rci ) that is oriented by modifying the
surface of the bottom electrode . we thermally deposited
a 120 nm - thick layer of gold ( au 111 ) onto a technical - grade 3
silicon wafer supporting a native oxide layer ( si / sio2 ) . we fabricated the substrates by template stripping ( ts ) , where a drop of ultraviolet ( uv)-curable optical
adhesive ( oa ) was used to adhere a 1 1 cm piece of glass to
a preprepared gold coated wafer . the glass was then mechanically cleaved
exposing an ultraflat au surface with a root - mean - squared roughness
of 0.3 nm ( as measured using tapping - mode afm ) . next , we immersed
the substrates in a solution of 1 mm 2-mercaptoethanol ( 2me ) or sodium
3-mercapto-1-propanesulfonate ( mps ) to direct the psi complexes to
adopt a down or up orientation ( fb iron sulfur cluster
adjacent or away from substrate ) . the time of immersion was limited
to 2 h to avoid the formation of multilayers or aggregates . after
this step , were rinsed the substrates with mq water ( mps ) or ethanol
( 2me ) , dried them with nitrogen and incubated them in a previously
prepared psi solution . the psi solution
consisted of 1:1 in buffer a ( 20 mm hepes ( ph 7.5 ) ; 10 mm mgcl2 ; 10 mm cacl2 ; 500 mm mannitol with 0.05% ddm ( n - dodecyl--d - maltoside ) ) for 2 h. they were
then rinsed with mq water and dried with nitrogen . we analyzed each
substrate by afm after each fabrication step : on the template - stripped
substrate after cleavage , after surface modification , and after incubation
in a solution of psi . this was calculated by knowing the diameter
of the psi trimer from tem images and the coverage density ( calculated
manually ) for specific areas . we obtained the afm images with multimode
8 with scanasyst microscope in tapping mode with tesp probes ( bruker )
with spring constant k = 42 nm , resonance frequency f = 320410 khz and
tip radius of less than 10 nm . the scan rate and resolution were 1
hz and 640 lines / sample , respectively . we studied
the conductivity of the immobilized psi on the two orienting
monolayers with afm tunneling atomic force microscopy ( tuna ) contact
mode with a conducting probe . this mode was applied for electrical
characterization of single ( trimer ) psi complex with pt / ir coated
si n - type probe ( appnano ) , spring constant k = 0.020.8
nm , resonance frequency f = 525 khz and tip radius less than 30 nm with contact resistance
of 0.010.025 ohm / cm . the applied force
to cp - afm conducting probe on top of psi was started from low and
step - by - step increased to reach contacting for i this approach was used for each measurement point with
forces of less than 10 nn . we formed
the egain electrodes
by suspending a drop of egain from a 10 ml syringe on an adjustable
stage . using a piezo stepper ( open - loop , 5 nm resolution )
we brought the drop into contact with the sam of psi . the egain adhered
to both the needle and the au and by retracting the needle slowly
( ca . 50 ms , we produced conical tips of
egain with diameters of 25 m in diameter . we performed the electrical measurements in a custom - built
faraday cage using a keithley 6430 sub - femtoamp remote sourcemeter
smu instrument . the device was held in place with a spring - loaded
gold tip that was isolated from ground . data were obtained from an average of points by
sweeping the potential from 1.0 to 1.0 v at a rate of 0.2
v / s . to elucidate the transport mechanism of bacterial psi - based devices ,
we measured the dependence of the electric behavior on temperature .
in order to do this , we fabricated encapsulated and addressable devices
able to operate in a pressure and temperature controlled setup ( see supporting information ) . | recently , photoactive proteins have
gained a lot of attention due
to their incorporation into bioinspired ( photo)electrochemical and
solar cells .
this paper describes the measurement of the asymmetry
of current transport of self - assembled monolayers ( sams ) of the entire
photosystem i ( psi ) protein complex ( not the isolated reaction center ,
rci ) , on two different director sams supported by
ultraflat au substrates .
the director sams induce the preferential
orientation of psi , which manifest as asymmetry in tunneling charge - transport .
we measured the oriented sams of psi using eutectic ga in ( egain ) ,
a large - area technique , and conducting probe atomic force microscopy
( cp - afm ) , a single - complex technique , and determined that the transport
properties are comparable . by varying the temperatures
at which the
measurements were performed , we found that there is no measurable
dependence of the current on temperature from 0.1 to 1.0
v bias , and thus , we suggest tunneling as the mechanism for transport ;
there are no thermally activated ( e.g. , hopping )
processes .
therefore , it is likely that relaxation in the electron
transport chain is not responsible for the asymmetry in the conductance
of sams of psi complexes in these junctions , which we ascribe instead
to the presence of a large , net dipole moment present in psi . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
central retinal artery occlusion ( crao ) is the occlusion of the central retinal artery resulting in infarction of the retina and acute , painless vision loss . furthermore , atherosclerosis - related thrombus , embolism , vasospasm , and giant cell arteritis have been known as causes of crao . since the disease is analogous to an ischemic cerebral stroke , the associated atherosclerotic risk factors may place crao patients at risk of future cerebral stroke and ischemic heart disease . crao should be considered as an ocular emergency that should be treated without delay after symptom onset . immediate hyperbaric oxygen ( hbo2 ) therapy has been used to supply oxygen to the ischemic retina in order to maintain viability while other treatment modalities are applied to restore the central retinal artery blood flow . traditional treatment modalities include ocular massage , anterior chamber paracentesis , intraocular pressure - lowering medications , and vasodilators , all of which are relatively ineffective . in recent years , treatment modalities have been developed including delivery of thrombolytic agents by surgical approaches and the removal of embolism by the nd : yag laser . currently , digital subtraction angiography ( dsa)-guided superselective ophthalmic artery thrombolysis or selective carotid intervention remains the preferred treatment method for crao [ 1115 ] . during the procedure , the dsa - guided catheter is inserted through the femoral artery and extended successively through the aorta , common carotid , and internal carotid artery to directly apply ophthalmic or carotid thrombolysis . ophthalmic artery branch retrograde thrombolytic intervention , a novel surgical approach developed by our team , is achieved though the terminal branches of the ophthalmic artery . the approach does not involve large intracranial vessels and greatly simplifies the surgical procedure . in this study , the clinical efficacy and safety of the ophthalmic artery branch retrograde thrombolytic intervention and superselective ophthalmic artery thrombolysis / selective carotid intervention were compared in a group of crao patients in order to provide a basis for the wide application of the new therapeutic approach in clinical practice . this study included a total of 50 patients with monocular crao confirmed by fundus fluorescein angiography ( ffa ) who were hospitalized in the department of ophthalmology at the fourth hospital of xian between march 2000 and september 2015 . study subjects were selected based on the indications and contraindications of thrombolytic therapy in acute cerebral infarction . specifically , the following selection criteria were used : aged between 18 and 78 years , within 76 h after onset , blood pressure below 180/100 mm hg ( 1 mm hg=0.133 kpa ) , and normal prothrombin time , thrombin time , fibrinogen , and platelet counts . the exclusion criteria included computed tomography ( ct)-confirmed intracranial bleeding ; a history of intracranial hemorrhage , intracranial arteriovenous malformation , or aneurysm ; intracranial surgery ; head trauma or stroke with the past month ; active gastrointestinal or urinary bleeding with the past 21 d ; surgery within the past 14 d ; ultrasound - confirmed cardiogenic emboli and carotid atherosclerotic plaque ; and history of severe heart , lung , kidney , and liver disease . the 50 patients were randomly divided into two groups : group a ( n=26 ) , who underwent ophthalmic artery branch retrograde thrombolytic intervention ; and group b ( n=24 ) , who underwent superselective ophthalmic artery thrombolysis / selective carotid intervention . the blood pressure of all patients was controlled below 140/90 mm hg before thrombolytic intervention . a , the skin around the diseased eye was disinfected and the supraorbital nerve was blocked by an injection of 1% lidocaine . the skin below the medial canthus or above the supraorbital foramen was cut , and the supratrochlear or supraorbital artery was exposed by blunt dissection ( figure 1 ) . distal arterial blood flow was blocked , and the vascular wall was pierced by a sharp knife to insert a microcatheter with a guidewire ( figure 2 ) . the guidewire was removed and 4 ml of the contrast agent iohexol 300 ( national drug registration no . h10970325 , yangtze river pharmaceutical group ) was injected into the artery a speed of 1 ml / s through the microcatheter using a 1 ml injection syringe . the ophthalmic artery was examined by dsa to confirm the correct insertion of the microcatheter ( figure 3 ) . a total amount of 400,000 u of urokinase and 30 mg of papaverine was pumped into the artery through a coaxial microcatheter using a micropump . the patients in group b was subjected to total cerebral angiography using ge - innova 41001q in order to exclude intracranial aneurysms , arteriovenous malformations , and other diseases , and to determine subsequent intervention with superselective ophthalmic artery or selective carotid thrombolysis . briefly , the skin at the puncture site was disinfected , and the body part near the femoral artery was anesthetized by 2% lidocaine . a 2-mm incision was created , and the femoral artery was pierced by an arterial puncture needle . a guidewire was placed into the femoral artery in the right position , and a 5f arterial catheter sheath , cannula , and catheter were inserted sequentially through the aorta , common carotid , and internal carotid artery to reach the ophthalmic artery . a total amount of 8 ml and 6 ml of the contrast agent iohexol 300 was injected in the internal carotid and the external carotid , respectively , at a speed of 2 ml / s using a high - pressure syringe . a total amount of 400,000 u of urokinase and 30 mg of papaverine was pumped into the artery through a coaxial microcatheter using a micropump . the patients were required to rest in bed and closely monitored for bleeding at the puncture site and cardiovascular abnormities for 24 h after the surgery . the patients were followed up for 5 to 21 days ( mean follow - up period : 16 days ) to observe systemic and local adverse reactions , and to evaluate the clinical effects based on the following criteria : ( 1 ) an effective treatment was marked by a thickened ophthalmic artery and its branches , faster circulation , and a clear ring around the eye in postoperative dsa examination . ( 2 ) the mean arm - retina circulation time ( a - rct ) was defined as the time elapsed when the fluorescein traveled from the ulnar vein to the retinal artery , and the circulation time from the retinal artery to the posterior pole peripheral retinal artery filling was named retinal artery peripheral filling time . the a - rct and retinal artery peripheral filling time were measured by ffa using a topcon trc-50ex fundus camera ( topcon , tokyo , japan ) to assess the retinal blood circulation recovery based on the following criteria : markedly effective : a - rct 15 s , and retinal artery - peripheral filling time of all branches of the central retinal artery 3 s ; effective : a - rct was between 16 and 20 s , and retinal artery - peripheral filling time of all branches of the central retinal artery was between 4 and 8 s ; and invalid : a - rct 21 s , and retinal artery - peripheral filling time of all branches of the central retinal artery 9 s. ( 3 ) a visual acuity test was performed using the semiquantitative scale no light perception , light perception , counting fingers , and hand motion , and the quantitative scale 0.02 , 0.05 , 0.1 , 0.12 , 0.15 , 0.2 , 0.25 0.6 to evaluate the postoperative vision . the treatment was considered markedly effective when the postoperative vision was improved for 3 levels compared with the preoperative vision , effective in case of 2 levels of improvement , and invalid in case of 1 or 0 level of improvement . this study included a total of 50 patients with monocular crao confirmed by fundus fluorescein angiography ( ffa ) who were hospitalized in the department of ophthalmology at the fourth hospital of xian between march 2000 and september 2015 . study subjects were selected based on the indications and contraindications of thrombolytic therapy in acute cerebral infarction . specifically , the following selection criteria were used : aged between 18 and 78 years , within 76 h after onset , blood pressure below 180/100 mm hg ( 1 mm hg=0.133 kpa ) , and normal prothrombin time , thrombin time , fibrinogen , and platelet counts . the exclusion criteria included computed tomography ( ct)-confirmed intracranial bleeding ; a history of intracranial hemorrhage , intracranial arteriovenous malformation , or aneurysm ; intracranial surgery ; head trauma or stroke with the past month ; active gastrointestinal or urinary bleeding with the past 21 d ; surgery within the past 14 d ; ultrasound - confirmed cardiogenic emboli and carotid atherosclerotic plaque ; and history of severe heart , lung , kidney , and liver disease . the 50 patients were randomly divided into two groups : group a ( n=26 ) , who underwent ophthalmic artery branch retrograde thrombolytic intervention ; and group b ( n=24 ) , who underwent superselective ophthalmic artery thrombolysis / selective carotid intervention . the blood pressure of all patients was controlled below 140/90 mm hg before thrombolytic intervention . in group a , the skin around the diseased eye was disinfected and the supraorbital nerve was blocked by an injection of 1% lidocaine . the skin below the medial canthus or above the supraorbital foramen was cut , and the supratrochlear or supraorbital artery was exposed by blunt dissection ( figure 1 ) . distal arterial blood flow was blocked , and the vascular wall was pierced by a sharp knife to insert a microcatheter with a guidewire ( figure 2 ) . the guidewire was removed and 4 ml of the contrast agent iohexol 300 ( national drug registration no . h10970325 , yangtze river pharmaceutical group ) was injected into the artery a speed of 1 ml / s through the microcatheter using a 1 ml injection syringe . the ophthalmic artery was examined by dsa to confirm the correct insertion of the microcatheter ( figure 3 ) . a total amount of 400,000 u of urokinase and 30 mg of papaverine was pumped into the artery through a coaxial microcatheter using a micropump . the patients in group b was subjected to total cerebral angiography using ge - innova 41001q in order to exclude intracranial aneurysms , arteriovenous malformations , and other diseases , and to determine subsequent intervention with superselective ophthalmic artery or selective carotid thrombolysis . briefly , the skin at the puncture site was disinfected , and the body part near the femoral artery was anesthetized by 2% lidocaine . a 2-mm incision was created , and the femoral artery was pierced by an arterial puncture needle . a guidewire was placed into the femoral artery in the right position , and a 5f arterial catheter sheath , cannula , and catheter were inserted sequentially through the aorta , common carotid , and internal carotid artery to reach the ophthalmic artery . a total amount of 8 ml and 6 ml of the contrast agent iohexol 300 was injected in the internal carotid and the external carotid , respectively , at a speed of 2 ml / s using a high - pressure syringe . a total amount of 400,000 u of urokinase and 30 mg of papaverine was pumped into the artery through a coaxial microcatheter using a micropump . the patients were required to rest in bed and closely monitored for bleeding at the puncture site and cardiovascular abnormities for 24 h after the surgery . the patients were followed up for 5 to 21 days ( mean follow - up period : 16 days ) to observe systemic and local adverse reactions , and to evaluate the clinical effects based on the following criteria : ( 1 ) an effective treatment was marked by a thickened ophthalmic artery and its branches , faster circulation , and a clear ring around the eye in postoperative dsa examination . ( 2 ) the mean arm - retina circulation time ( a - rct ) was defined as the time elapsed when the fluorescein traveled from the ulnar vein to the retinal artery , and the circulation time from the retinal artery to the posterior pole peripheral retinal artery filling was named retinal artery peripheral filling time . trc-50ex fundus camera ( topcon , tokyo , japan ) to assess the retinal blood circulation recovery based on the following criteria : markedly effective : a - rct 15 s , and retinal artery - peripheral filling time of all branches of the central retinal artery 3 s ; effective : a - rct was between 16 and 20 s , and retinal artery - peripheral filling time of all branches of the central retinal artery was between 4 and 8 s ; and invalid : a - rct 21 s , and retinal artery - peripheral filling time of all branches of the central retinal artery 9 s. ( 3 ) a visual acuity test was performed using the semiquantitative scale no light perception , light perception , counting fingers , and hand motion , and the quantitative scale 0.02 , 0.05 , 0.1 , 0.12 , 0.15 , 0.2 , 0.25 0.6 to evaluate the postoperative vision . the treatment was considered markedly effective when the postoperative vision was improved for 3 levels compared with the preoperative vision , effective in case of 2 levels of improvement , and invalid in case of 1 or 0 level of improvement . a total of 50 crao patients were selected in this study including 28 males and 22 females . the mean age was 55.7 years , and the average time to onset was 22 h ( range : 876 h ) . concurrent hypertension ( 140180 mm hg/80100 mm hg ) and diabetes were reported in 37 cases and 1 case , respectively . none of the patients had any history of rheumatic disease , congenital heart disease , trauma , or infection . in the preoperative visual acuity test , there were 25 cases of no light perception , 18 of light perception , and 9 of hand motion . the intraocular pressure ( iop ) ranged between 14 and 20 mm hg with a mean iop of 19 mm hg . as shown in table 1 , there was no significant difference in age ( t=0.55 , p=0.58 ) , gender ratio ( =0.481 , p=0.49 ) , and time to onset ( t=0.001 , p=1.00 ) between the two groups . among the 26 cases in group a with the ophthalmic artery branch retrograde thrombolytic intervention , the catheter was successfully inserted through the supraorbital artery and the supratrochlear artery in 1 and 25 cases , respectively , as indicated by clear display of the ophthalmic artery in dsa examination . b , a patient developed intraoperative cerebral vasospasm and concurrent disturbance of consciousness , and recovered after prompt symptomatic treatment . follow - up examination , serious complications such as hematoma at the incision site , intracranial hemorrhage , and cerebral embolism occurred in none of the patients in both groups . treatment outcome of the two groups was compared by postoperative dsa , ffa , and visual acuity tests . effective treatment was indicated by the thickened ophthalmic artery and its branches , faster circulation , and a clear ring around the eye in postoperative dsa angiography . in group a , the ophthalmic artery branch retrograde thrombolytic intervention was effective in 92.3% of cases ( figure 4 ) , which was comparable to that in group b ( =2.08 , p=0.25 ) ( table 2 ) . the pre- and post - operative ffa images are shown in figure 5a5c and figure 6a6c ( group a ) , and in figure 7a7c and figure 8a8c ( group b ) . there was no significant difference in treatment efficacy between the two groups as indicated by ffa results ( =3.09 , p=0.21 ) ( table 2 ) . a , the percentage of markedly effective , effective , and invalid treatment was 42.3% , 50% , and 7.7% , respectively , which was similar to that in group b ( 41.7% , 50% , and 8.3% , respectively ) ( =0.25 , p=0.88 ) a total of 50 crao patients were selected in this study including 28 males and 22 females . the mean age was 55.7 years , and the average time to onset was 22 h ( range : 876 h ) . concurrent hypertension ( 140180 mm hg/80100 mm hg ) and diabetes were reported in 37 cases and 1 case , respectively . none of the patients had any history of rheumatic disease , congenital heart disease , trauma , or infection . in the preoperative visual acuity test , there were 25 cases of no light perception , 18 of light perception , and 9 of hand motion . the intraocular pressure ( iop ) ranged between 14 and 20 mm hg with a mean iop of 19 mm hg . as shown in table 1 , there was no significant difference in age ( t=0.55 , p=0.58 ) , gender ratio ( =0.481 , p=0.49 ) , and time to onset ( t=0.001 , p=1.00 ) between the two groups . among the 26 cases in group a with the ophthalmic artery branch retrograde thrombolytic intervention , the catheter was successfully inserted through the supraorbital artery and the supratrochlear artery in 1 and 25 cases , respectively , as indicated by clear display of the ophthalmic artery in dsa examination . b , a patient developed intraoperative cerebral vasospasm and concurrent disturbance of consciousness , and recovered after prompt symptomatic treatment . follow - up examination , serious complications such as hematoma at the incision site , intracranial hemorrhage , and cerebral embolism occurred in none of the patients in both groups . abnormal eye movement or vitreous hemorrhage treatment outcome of the two groups was compared by postoperative dsa , ffa , and visual acuity tests . effective treatment was indicated by the thickened ophthalmic artery and its branches , faster circulation , and a clear ring around the eye in postoperative dsa angiography . in group a , the ophthalmic artery branch retrograde thrombolytic intervention was effective in 92.3% of cases ( figure 4 ) , which was comparable to that in group b ( =2.08 , p=0.25 ) ( table 2 ) . the pre- and post - operative ffa images are shown in figure 5a5c and figure 6a6c ( group a ) , and in figure 7a7c and figure 8a8c ( group b ) . there was no significant difference in treatment efficacy between the two groups as indicated by ffa results ( =3.09 , p=0.21 ) ( table 2 ) . a , the percentage of markedly effective , effective , and invalid treatment was 42.3% , 50% , and 7.7% , respectively , which was similar to that in group b ( 41.7% , 50% , and 8.3% , respectively ) ( =0.25 , p=0.88 ) it has been known that the optimal therapeutic time window of arterial thrombolysis for acute stroke is 4.5 hours after onset . in clinical situations , the thrombolytic treatment for crao is often delayed due to various preoperative examinations , preparation , and communication with patients families . the mean time elapsed from crao onset to thrombolytic intervention is 12.78 hours , which is far beyond the optimal therapeutic time window . in the conventional dsa - guided superselective ophthalmic artery or selective carotid thrombolytic intervention , the catheter is sequentially extended through the aorta , common carotid , internal carotid artery , and ophthalmic artery , which might potentially cause arterial spasm , emboli shedding , postoperative aortic dissection , bleeding at the puncture site in the femoral artery , etc . moreover , the thrombolytic drugs might induce intracranial bleeding of closed blood vessels in past lesions . the incidence rate of serious complications ( such as transient ischemic attack , cerebral infarction , and cerebral hemorrhage ) and other adverse events is approximately 4% and 37.1% , respectively [ 1820 ] . in this study , a patient in group b developed intraoperative cerebral vasospasm and concurrent disturbance of consciousness , and recovered after prompt symptomatic treatment . preoperative brain ct has become an important step to detect abnormities such as intracranial hemorrhage , cerebromalacia , cerebral arteriovenous malformations , and intracranial aneurysms in order to prevent the occurrence of serious complications and adverse events . therefore , it is highly urgent to develop an intervention approach that involves neither large vessels in the body nor intracranial vessels to reduce the incidence rate of intra- and post - operative complications . furthermore , preoperative brain ct is not necessary before such approach , which may allow prompt treatment within the optimal therapeutic time window of crao . the ophthalmic artery branch retrograde intervention is a novel approach developed by our group , during which the catheter is extended from the supratrochlear or supraorbital artery to the ophthalmic artery for thrombolytic treatment . the path involved in this approach is less than 1/10 of that in conventional superselective ophthalmic artery or selective carotid intervention . as a result , the patients who undergo this approach are not at risk of complications associated with conventional intervention . more importantly , the intraoperative injection pressure of thrombolytic drug is slightly higher than the systolic pressure of the ophthalmic artery or internal carotid , which pushes the thrombolytic drug into the ophthalmic artery after the injection . neither the catheter nor the thrombolytic drugs enter any intracranial vessels other than the ophthalmic artery . in our study , the ophthalmic artery was clearly displayed during dsa examination , whereas the common carotid artery and internal carotid artery were not illustrated . the patients in group a were not required to rest in bed after the surgery , and no systemic and local adverse complications occurred . in this study , both treatments were effective as indicated by thickened ophthalmic artery , faster circulation , and a clear ring around the eye in postoperative dsa examination . intra- or post - operative complications occurred in none of the patients in group a despite the fact that preoperative brain ct was not performed in group a , suggesting the high safety of the developed ophthalmic artery branch retrograde approach . therefore , the approach does not require the time - consuming preoperative brain ct , and may greatly reduce the time elapsed from onset of the disease to surgical intervention . as a reliable method to directly display the conditions of retinal circulation no significant difference in the improvement of retinal blood circulation and visual function was detected between the two groups , revealing similar effectiveness of both therapies in the treatment of crao . during the ophthalmic artery branch nevertheless , the supratrochlear artery is the preferred vessel because the success rate of catheter insertion is much higher compared with the supraorbital artery due to its stable position and larger size . although special microcatheters and guidewires are used to deliver the thrombolytic drugs in both intervention approaches , the required length of microcatheters and guidewires in group a is much shorter compared with that in group b , which may largely reduce the treatment cost . in summary , the novel ophthalmic artery branch retrograde thrombolytic intervention developed by our team is highly effective and safe in the treatment of crao , and may be an alternative method for the delivery of therapeutic drugs in retinal vascular occlusive diseases . nevertheless , the surgeon should be well aware of the emergency nature of crao cases , because irreversible damage may occur in nerve fibers if the disease exceeds 240 minutes . | backgroundcentral retinal artery occlusion ( crao ) is the occlusion of the central retinal artery resulting in retinal infarction and acute vision loss .
digital subtraction angiography ( dsa)guided superselective ophthalmic artery or selective carotid thrombolysis remains the preferred treatment method for crao .
this study aimed to evaluate the safety and clinical efficacy of the novel ophthalmic artery branch retrograde thrombolytic intervention for crao.material/methodsfifty patients with monocular crao were enrolled , including 28 males and 22 females ( mean age : 55.72.3 years ) .
the patients were randomly divided into two groups for thrombolysis with urokinase ( 400,000 u ) and papaverine ( 30 mg ) by either ophthalmic artery branch retrograde intervention ( group a , n=26 ) or superselective ophthalmic artery / selective carotid intervention ( group b , n=24 ) .
there was no significant difference in age ( p=0.58 ) , gender ratio ( p=0.49 ) , and time to onset ( p=1.00 ) between the two groups .
the adverse reactions and clinical efficacy were evaluated by postoperative dsa , fundus fluorescein angiography ( ffa ) , and visual acuity tests.resultsno serious complications , abnormal eye movement , or vitreous hemorrhage occurred in either group .
dsa showed that group a had an effective rate ( 92.30% ) comparable to that of group b ( 100% , 2=2.08 , p=0.25 ) .
ffa suggested that both groups had similar treatment efficacy ( 2=3.09 , p=0.21 ) .
visual acuity tests also confirmed a similar efficacy of the two intervention approaches ( 2=0.25 , p=0.88).conclusionsthe developed novel ophthalmic artery branch retrograde intervention is highly effective and safe for crao , and may be a superior method compared with the conventional approach . |
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a 46-year - old female patient visited our hospital with pain in the right posterior neck area , which had continued since she had fallen from a chair six months previously . she said that she had fallen down from a chair with small wheels in her house at the time of the accident . the degree of pain was 8/10 on the visual analog scale ( vas ) , and the pain was localized mainly in the right posterior neck and occipital area . jackson 's compression test was positive and spurling 's test was positive on the right side . biceps , triceps , and brachioradialis tendon reflexes were normal . on lateral flexion - extension views of plain x - rays ( fig . 1 ) , there was narrowing of the disc spaces at the c4/5 and c5/6 levels . there was no sign of atlantoaxial instability , however , because the atlantodental interval ( adi ) was 2.86 mm . therefore , the patient 's pain was considered to be caused by a cervical herniated intervertebral disc and of myofascial origin , so a greater occipital nerve block , trigger point injection , and cervical epidural block were performed . however , these treatments were ineffective for her pain . considering the patient 's medical history of rheumatoid arthritis , we performed an open - mouth odontoid view on plain radiograph and a ct of the cervical spine for evaluation of the c1/2 joint , although there was no sign of instability on flexion / extension lateral images . the open - mouth view showed narrowing of the right atlantoaxial joint space , and the ct of the cervical spine showed an old fracture of the lateral mass at c1 and was suspicious for an avulsion injury of the right transverse ligament ( fig . atlantoaxial joint injection was performed , after which the patient 's pain was improved by 20% . mr imaging additionally revealed right facet subluxation at c1/2 with c2 nerve root compression ( fig . 3 ) . therefore , an additional right c2 drg block and c2 pulsed radiofrequency lesioning were performed . as a result , the patient experienced improvement in her pain , down to a vas score about 3/10 . she reported that the twinge of pain she felt was improved but that head flexion continued to be difficult . the patient 's current medication regimen is ultracet ( janssen korea , ltd . ) 3 tablets / day , and her pain intensity remains 3/10 on the vas . the stability of the craniocervical junction comes from bony structures and the ligaments of the facet capsules of occiput - c1 and c1-c2 . atlas fractures occur in 2 - 13% of cervical spine injuries and represent about 1 - 2% of all spinal fractures . since jefferson first described , in 1920 , an anterior and posterior arch fracture of the atlas , there has been no standard classification system of atlas fractures . the classification system modified by gehweiler distinguishes between isolated and combined arch fractures , lateral mass fractures , and transverse fractures of the anterior arch . type i fractures involve the posterior arch alone , whereas type ii fractures involve the anterior arch alone . type iii fractures involve bilateral posterior arch fractures related to unilateral or bilateral anterior arch fracture . type iv fractures involve the lateral mass , and type v fractures are transverse fractures of the anterior arch . according to landells and van peteghem 's classification , type iii fracture is a fracture of the lateral mass with or without a fracture of the arch . the present case would be classified as a gehweiler type iv , landells type iii fracture involving the lateral mass . the transverse ligament is the primary and crucial component of atlantoaxial stability , as the largest , thickest , and strongest of the cervical ligaments . the transverse ligament is a very firm , inelastic band of tissue , and its injury can occur suddenly when either rapid or slow loading forces are applied . it can rupture in the central portion of the transverse ligament , or it can cause an osteoperiosteal failure at the medial tubercle of the c1 lateral mass . they distinguished type i injuries involving the mid - portion or the insertion point of the transverse ligament and type ii injuries involving avulsion fractures of the tubercle for insertion of the transverse ligament . type i injuries are incapable of healing with external immobilization , but type ii injuries are capable of healing with only external immobilization . the present case was classified as a dickman 's type ii injury ; however , it was incapable of healing with external immobilization because it was an old injury that had happened six months ago . patients with fractures of the upper cervical spine mainly complain of neck pain , spasm of the cervical muscles , and limitation of neck motion . neuralgia and paresthesia also frequently result from compression or injuries of the c2 nerve , as in the present case . to evaluate atlas fractures , it is necessary to obtain lateral , flexion - extension , and open - mouth views on plain x - rays . on lateral flexion - extension views , an adi greater than 3 mm in adults and greater than 5 mm in children is highly suggestive of disruption of the transverse ligament . on the open - mouth view , displacement of the c1 lateral mass is suggestive of an atlas burst fracture , with displacement greater than 7 mm resulting in instability by transverse ligamentous disruption . however , if the adi is less than 3 mm with fractures of the c1 lateral mass , as inthis case , diagnosis by plain x - ray alone is of limited value . moreover , plain x - rays are relatively insensitive for detecting injuries of the transverse ligament . in this case , the cervical ct scan showed clearly the fracture of the c1 lateral mass missed by plain x - rays , known as the best modalities for evaluating osseous abnormalities . although ct scan is insufficient to demonstrate the anatomy of the transverse ligament , axial views on ct scans can detect ligament avulsion fractures , even if the displacement is less than 7 mm . on the other hand , mr imaging can be reliable to obtain visualization of the transverse ligament as well as to evaluate the spinal cord after trauma . mr imaging is highly sensitive fordetection of transverse ligamentous disruption , with mr imaging showing the anatomical continuity of the ligament , and abnormal signal intensity revealing ligamentous disruption . in the present case , the posterior neck pain was initially considered to be caused by a herniated cervical intervertebral disc ormyofascial origin , because plain radiographs showed only narrowing of the disc space without c1/2 instability . however , further evaluation by ct scan was performed on the basis of the patient 's history of old trauma and rheumatoid arthritis . a cervical ct scan revealed an unrecognized c1 lateral mass fracture and an avulsion fracture of the transverse ligament . moreover , compression of the c2 nerve root caused by c1/2 facet subluxation was discovered by mr imaging , and proper treatment was therefore possible . in these types of patients , detailed history taking and aggressive imaging assessment are necessary to allow accurate diagnosis and proper treatments . it is most important to assess atlantoaxial stability by ct scan and mr imaging , especially when apatient with a history of trauma complains of occipital neuralgia and has limited range of motion of the cervical spine . additionally , atlantoaxial joint injection and c2 drg block can be considered as conservative treatments for atlas fractures . | posterior neck pain is a common complaint of patients in the pain clinic .
the atlas ( c1 ) burst fracture is known to be a cause of posterior neck pain and instability .
although the atlas burst fracture and instability can be discovered by plain x - rays which show lateral mass displacement or widening of the atlantodental interval , assessment of an atlas burst fracture can be difficult if there is no instability in the imaging study . here
we report a case of a 46-year - old female patient who had complained of sustained posterior neck pain for 6 months .
plain x - rays showed only disc space narrowing at c4/5 and c5/6 , without any cervical instability .
however , an unrecognized c1 lateral mass fracture was detected by ct and mri .
the patient 's pain was then successfully treated after atlantoaxial joint injection with a c2 drg block . |
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gamma - aminobutyric acid ( gaba ) , the major inhibitory neurotransmitter in the mammalian brain , has two modes of release : phasic and tonic . while phasic release is mediated by synaptic vesicles in gabaergic neurons , tonic release is channel - mediated , a sustained form of release and originated from glial cells . unlike neurons , which convert glutamate to gaba by glutamate decarboxylases ( gads ) , glial cells utilize putrescine to produce gaba . among the enzymes involved in putrescine degradation pathway , monoamine oxidase b ( maob ) is a key enzyme that is responsible for glial gaba production in the cerebellum and striatum . treatment of maob inhibitor or gene silencing of maob effectively suppresses gaba production and release from glial cells . the content of glial gaba measured by immunohistochemistry varies in different regions of the brain and is known to positively correlate with a degree of tonic inhibition onto neurons . however , physiological relevance of glial gaba in various brain regions and specific neural circuits remains unknown . recently , we have shown a novel function of glial gaba in a pathological condition using two mouse models of alzheimer 's disease . in the molecular layer of dentate gyrus ( dg ) , diseased astrocytes produce abundant amount of gaba which is further released and inhibits excitatory synaptic transmission at the perforant path ( pp ) to dg granule neuron synapse . because the dg is the gateway of cortical input to the hippocampus and is essential for the formation and recall of memory , that gaba binds to neuronal gabaa and gabab receptors and results in impaired learning and memory in alzheimer 's disease . accordingly , suppressing astrocytic gaba production by oral administration of maob inhibitors rescues excitatory synaptic transmission and improves memory in alzheimer 's model . however , what proportion of gaba from reactive astrocyte targets gabaa and gabab has not been determined . although the content of astrocytic gaba is much lower under normal condition than that of diseased astrocytes , normal astrocytes show moderate levels of intracellular gaba and granule neurons show a small but significant level of tonic inhibition current in wild type mouse . interestingly , in wild - type mice , suppressing astrocytic gaba production was shown to decrease the amplitude of evoked excitatory postsynaptic currents ( eepscs ) at low intensity of pp stimulation , suggesting an excitatory action of astrocytic gaba . considering the presence of gabaergic interneurons in the molecular layer of dg , it is possible that the excitatory action of astrocytic gaba is by disinhibition . another possibility is that it could be due to the gabaa - mediated depolarizing effect if the intracellular chloride concentration is high . here , we investigated a physiological role of astrocytic gaba in the dg and compared its mechanism with a pathological condition by adopting app / ps1 mice , a popular model of alzheimer 's disease . all animal care and handling was performed according to the directives of the animal care and use committee of kaist ( daejeon , korea ) and the institutional animal care and use committee of kist ( seoul , korea ) . appswe / psen1de9 ( app / ps1 ; stock number 004462 , jackson laboratory , usa ) mice were maintained as hemizygotes by repeated backcrossing with b6c3 f1 mice . both sexes of 8- to 13-month - old transgenic mice and wild - type littermates were used for study . all experiments were performed with gender- and age - matched controls . for oral administration of selegiline , the mice were provided with water ( control ) or selegiline solution ad libitum , refreshed every two or three days . the selegiline solution was prepared by dissolving 10 mg of r-(-)-deprenyl hydrochloride ( sigma , usa ) in 150 ml drinking water . in this condition , the dose was calculated as 5~10 mg kg d. electrophysiological recordings were made from horizontal brain slices of 400-mm thickness . brain tissue was cut using a leica vt1000s microtome in oxygenated ice cold acsf composed of ( in mm ) : 130 nacl , 2.5 kcl , 26 nahco3 , 1.25 nah2po4 , 1.0 cacl2 , 3.0 mgcl2 , and 10 d - glucose . after slicing , the tissue was maintained at a room temperature of 20~22 for at least 60 min in acsf of the same composition and then transferred to the recording chamber . recordings were performed at room temperature of 20~22 in oxygenated acsf , contained ( in mm ) : 130 nacl , 2.5 kcl , 26 nahco3 , 1.25 nah2po4 , 1.5 cacl2 , 1.5 mgcl2 and 10 d - glucose . synaptic responses in dentate granule neurons were evoked by stimulation of lateral perforant path fibers applied at 0.1 hz ( 100 s duration ) via a constant current isolation unit . perforant path fibers were stimulated by placing a tungsten bipolar electrode in the outer half of the middle third dentate molecular layer . evoked epsps were recorded using glass pipette electrodes ( 6~8 m ) , filled with intracellular solution contained ( in mm ) : 140 cs - methanesulfonate , 8 nacl , 1 mgcl2 , 0.5 egta , 10 hepes , 7 phosphocreatine , di-(tris ) salt , 4 mg - atp , 0.3 na - gtp , 5 qx314 ( ph 7.3 was adjusted with nmdg ) . the epsc rise time was measured as the time from 10 to 90% peak amplitude . whole - cell patch - clamp recordings were performed from dg granule neurons , visually identified with infrared video microscopy and differential interference contrast optics . data were collected with a multi - clamp 700b amplifier ( molecular devices , usa ) using clampex10.2 acquisition software ( molecular devices , usa ) and digitized with digidata 1322a ( molecular devices , usa ) . raw data were low pass filtered at 4 khz and collected for off - line analysis at a sampling rate of 10 khz . all animal care and handling was performed according to the directives of the animal care and use committee of kaist ( daejeon , korea ) and the institutional animal care and use committee of kist ( seoul , korea ) . appswe / psen1de9 ( app / ps1 ; stock number 004462 , jackson laboratory , usa ) mice were maintained as hemizygotes by repeated backcrossing with b6c3 f1 mice . both sexes of 8- to 13-month - old transgenic mice and wild - type littermates were used for study . all experiments were performed with gender- and age - matched controls . for oral administration of selegiline , the mice were provided with water ( control ) or selegiline solution ad libitum , refreshed every two or three days . the selegiline solution was prepared by dissolving 10 mg of r-(-)-deprenyl hydrochloride ( sigma , usa ) in 150 ml drinking water . in this condition , the dose was calculated as 5~10 mg kg d. brain tissue was cut using a leica vt1000s microtome in oxygenated ice cold acsf composed of ( in mm ) : 130 nacl , 2.5 kcl , 26 nahco3 , 1.25 nah2po4 , 1.0 cacl2 , 3.0 mgcl2 , and 10 d - glucose . after slicing , the tissue was maintained at a room temperature of 20~22 for at least 60 min in acsf of the same composition and then transferred to the recording chamber . recordings were performed at room temperature of 20~22 in oxygenated acsf , contained ( in mm ) : 130 nacl , 2.5 kcl , 26 nahco3 , 1.25 nah2po4 , 1.5 cacl2 , 1.5 mgcl2 and 10 d - glucose . synaptic responses in dentate granule neurons were evoked by stimulation of lateral perforant path fibers applied at 0.1 hz ( 100 s duration ) via a constant current isolation unit . perforant path fibers were stimulated by placing a tungsten bipolar electrode in the outer half of the middle third dentate molecular layer . evoked epsps were recorded using glass pipette electrodes ( 6~8 m ) , filled with intracellular solution contained ( in mm ) : 140 cs - methanesulfonate , 8 nacl , 1 mgcl2 , 0.5 egta , 10 hepes , 7 phosphocreatine , di-(tris ) salt , 4 mg - atp , 0.3 na - gtp , 5 qx314 ( ph 7.3 was adjusted with nmdg ) . the epsc rise time was measured as the time from 10 to 90% peak amplitude . whole - cell patch - clamp recordings were performed from dg granule neurons , visually identified with infrared video microscopy and differential interference contrast optics . data were collected with a multi - clamp 700b amplifier ( molecular devices , usa ) using clampex10.2 acquisition software ( molecular devices , usa ) and digitized with digidata 1322a ( molecular devices , usa ) . raw data were low pass filtered at 4 khz and collected for off - line analysis at a sampling rate of 10 khz . to investigate the role of astrocytic gaba on the pp to dg granule neuron synaptic transmission , we assessed an effect of selegiline , an irreversible inhibitor that was demonstrated to block the production of gaba in astrocytes . consistently , when administered to mouse in drinking water or acutely applied to hippocampal slices , selegiline decreased the amplitude of evoked excitatory post - synaptic currents ( eepscs ) in dg granule neurons , without altering the resting membrane potential , somatic input resistance , amplitudes of the action potentials and the firing rate of granule neurons . mice were administered with selegiline for one week prior to electrophysiology experiments at a dosage sufficient to selectively inhibit maob over maoa activity ( see materials and methods ) . we have found that selegiline significantly reduced the amplitude of eepscs at 50 a or 100 a stimulation intensity in wild - type ( wt ) mice , whereas it significantly increased the eepsc amplitude in app / ps1 mice under the same stimulation conditions ( fig . these results imply an excitatory action of astrocytic gaba in wt mice and an inhibitory action in app / ps1 mice . the effects of astrocytic gaba on the amplitude of eepscs may result from a pre - synaptic action of gaba on the release probability at the pp terminals . to test an involvement of pre - synaptic mechanism to the observed phenomena , we estimated an effect of selegiline on paired pulse ratio ( ppr ) of two consecutively evoked epscs ( 50 ms apart ) in wt and app / ps1 mice . we found that selegiline did not change ppr in wt mice , whereas it significantly reduced ppr in app / ps1 mice by about 15% ( fig . these results suggest that astrocytic gaba does not change the release probability in wt mice , whereas it decreases the release probability in app / ps1 mice . in other words , the astrocytic gaba tonically decreased the amplitude of eepscs in app / ps1 mice via a pre - synaptic mechanism by inhibiting neurotransmitter release from pp terminals , whereas there was no involvement of pre - synaptic mechanism in wt mice . then how does astrocytic gaba enhance the amplitude of eepscs ? to address this question , we performed pharmacological experiments using selective antagonists for gabaa and gabab . the results revealed that bicuculline did not change the amplitude of eepscs in wt mice ( fig . 3a and 3b ) , whereas it significantly increased the amplitude of eepscs in app / ps1 mice ( fig . these results indicate that gabaa receptors are not involved in the excitatory action of astrocytic gaba in wt mice , whereas gabaa receptors mediate the inhibitory action of astrocytic gaba in app / ps1 mice . gaba released from interneurons attenuates excitation of post - synaptic neurons by reducing dendritic input resistance ( rin ) or by perisomatic inhibition . then , attenuation of the interneuronal activity would result in an increase in dendritic rin and subsequent increase in the rise time of eepscs in dg granule neurons . consistent with the hypothesis about an involvement of interneurons in the suppressing effect of selegiline on eepsc in wt mice , selegiline treatment significantly increased the rise time of eepsc from 2.840.17 ms to 4.350.26 ms ( fig . interestingly , the rise time of eepscs in dg granule neurons of app / ps1 mice was not significantly altered by selegiline ( fig . next we examined the contribution of gabab receptors in excitatory action of astrocytic gaba in wt mice . we estimated the effect of cgp55845 , a gabab receptor antagonist , on the amplitude of eepscs in the presence of bicuculline . we found that cgp55845 significantly attenuated the amplitude of eepsc by about 45% ( fig . 3c and 3d ) in the presence of bicuculline . the degree of reduction by cgp55845 was comparable to the degree of reduction by selegiline in the wt mice ( fig . 1i ) , indicating that gabab receptor accounts for the most of the excitatory effect shown by astrocytic gaba . the excitatory role of gabab receptor in wt mice and inhibitory role of gabaa in app / ps1 mice were confirmed in ppr experiment using bicuculline and cgp55845 . similar to selegiline treatment , we found that bicuculline did not change ppr in wt mice ( fig . 4a and 4b ) , whereas it significantly decreased ppr in app / ps1 mice by about 12% ( fig . 4e and 4f ) . 2c ) in app / ps1 mice indicate that presynaptic gabaa receptor at pp terminals is responsible for the inhibitory action of astrocytic gaba in app / ps1 mice . the observation that bicuculline did not alter ppr in wt mice ( fig . 4a and 4b ) is intriguing because it excludes a possibility of involvement of presynaptic gabaa receptors at pp terminals . more interestingly , ppr was not significantly changed by cgp55845 , again , eliminating the possibility of presynaptic gabab receptors at pp terminals ( fig . 4c and 4d ) . then where is the gabab receptor that is responsible for the excitatory action of astrocytic gaba ? the above results suggest that gabab receptor is present at the presynaptic terminals of local interneurons which form a feed - forward inhibitory circuit of dg and that astrocytic gaba targets these gabab receptors . thus , the action of astrocytic gaba in wt mice is a tonic disinhibitory action on gabab receptor of local interneurons of dg . attenuation of epsc in dg granule neurons by selegiline in the present study reveals an excitatory action of astrocytic gaba at weak stimulations of pp fibers . the finding that selegiline did not alter ppr at pp to granule neuron synapse calls for a non - presynaptic target of astrocytic gaba . in addition , the positive effect by gabab antagonist and negative effect by gabaa antagonist on eepsc suggest that the target of astrocytic gaba is gabab receptors , not gabaa . dg granule neurons are known to express gabab receptor sub - units on their dendrites in the outer molecular layer of dg . therefore , it may appear that astrocytic gaba directly inhibits activity of dg granule neurons by targeting gabab receptors on dg granule neurons . however , such a mechanism can not explain why the effect of astrocytic gaba was observed only at relatively weak stimulations of pp fibers ( fig . there is a subpopulation of hilar interneurons that project onto proximal dendrites and perisomatic regions of dg granule neurons . those interneurons receive direct input from pp to form feed - forward inhibitory circuit of dg , thus capable of indirectly modulating the pp to granule neuron transmission . those interneurons have strong immunoreactivity for gabab receptors in the inner molecular layer of dg . it has been also reported that all hilar interneurons that project to the inner molecular layer of dg have a low activation threshold for pp stimulation compared to that of dg granule neurons . based on these previous reports and our results demonstrating that gabab receptor antagonist attenuated the amplitude of eepscs in granule neurons , we conclude that astrocytic gaba targets gabab receptors on the axonal terminals of hilar interneurons projecting to the proximal dendrites and perisomatic region of dg granule neurons . the lower pp stimulation threshold for the activation of hilar interneurons could be the reason for the presence of effect by gabab antagonist only with a weak stimulation of the pp . astrocytic gaba is tonically and negatively modulating synaptic gaba release in interneuronal terminals , thus reducing the inhibitory drive on granule neurons in favor of excitatory drive from pp ( fig . 5 ) . with increasing stimulation strength of pp , the proportion of inhibitory drive on granule neurons is relatively decreased compared to the direct excitatory drive from pp ( fig . 5 ) . under this condition , the astrocytic gaba appears to be confined to the inner molecular layer of dg . future investigations are needed to explore the possible role of hilar interneurons in disinhibitory action of astrocytic gaba in wt mice . in contrast to astrocytes in wt , reactive astrocytes in the molecular layer of dg in app / ps1 mice have increased production and tonic release of gaba . this more abundant astrocytic gaba can extend to more outer molecular layer of dg to affect presynaptic terminals of pp . our results demonstrate that in app / ps1 mice , gaba from reactive astrocytes targets gabaa receptors at the pp terminals to powerfully reduce the release probability of pp to dg granule neuron synapse ( fig . this direct inhibitory action of astrocytic gaba targeting presynaptic gabaa receptors accounts for the majority of inhibitory action observed at this synapse , as evidenced by a similar degree of reduction of amplitude of eepscs and ppr between selegiline and bicuculline . these results should provide a useful target to alleviate memory impairment in alzheimer 's disease model . a precise molecular / cellular mechanisms underlying enhanced sensitivity of pp terminals in ad mice is not well understood . one possibility might be up - regulation of gabaa receptors in the perforant path terminals or alterations in expression of na - k - cl or k - cl cotransporters in pp terminals of app / ps1 mice . other possibilities may include morphological reorganization of astrocytes during gliosis and increased production and secretion of gaba by reactive astrocytes leading to altered spatial and temporal factors in glia - neuron communication in the dg of alzheimer 's brain that strengthens an influence of astrocytic gaba on pp - to - dg granule neuron synapse . in summary , we have found that at low intensity of pp fiber stimulation , astrocytic gaba exerts a disinhibitory effect on dg granule neurons by targeting presynaptic gabab receptors of feed - forward inhibitory circuit in dg of wt mice . in app / ps1 mice , astrocytic gaba extends its target to presynaptic gabaa receptors of pp terminals to powerfully reduce the release probability at the pp to dg granule neuron synapse . | like neurons , astrocytes produce and release gaba to influence neuronal signaling . at the perforant path to dentate gyrus granule neuron synapse , gaba from astrocyte
was found to be a strong inhibitory factor , which impairs synaptic transmission , synaptic plasticity and memory in alzheimer 's disease .
although astrocytic gaba is observed in many brain regions , its physiological role has not been clearly demonstrated yet . here
, we show that astrocytic gaba exerts disinhibitory action to dentate granule neurons by targeting gabab receptors of gabaergic interneurons in wild - type mice .
this disinhibitory effect is specific to a low intensity of electrical stimulation at perforant path fibers . inversely in alzheimer 's disease model mice , astrocytic gaba targets gabaa receptors and exerts inhibitory action by reducing release probability of glutamatergic perforant path terminals .
these results suggest that astrocytic gaba differentially modulates the signaling from cortical input to dentate gyrus under physiological and pathological conditions . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
one of the most popular athletic games , soccer , requires a basic element of strong physical
power almost at the threshold level1 ,
through which successful athletic performance is marked by technical and tactical
activities2 . physical power is based on
muscle strength and durability , quick reaction and agility , and flexibility , and it involves
complex components , such as the relationship between the strength of the quadriceps femoris
and anaerobic power3 . to maintain balance
and allow for quick reactions and agility during soccer , the iterative interaction among the
joints of the lower limb has a significant effect on the players athletic performance4 . the muscle strength of the knee joint plays a role in supporting the body s weight . during
a game , however , it also supports the center of body mass while performing such dynamic
movements as kicking the ball , jumping , and changing direction , which all rely on quick
reaction , agility , and coordination5 . functional abnormalities of the knee joint can include overuse , repetitive injuries , or
unstable proprioception due to repetition of limited movements within the joint s mobility
range6 . to maintain the stability of the
knee , it is important to evaluate the level of the muscle strength balance , to assess
endurance using isokinetic exercises , and then to facilitate muscle development
accordingly7 . in soccer , the foot and ankle joints have the highest rates of injury , and functional
movement impairment can be a cause of instability through insufficient muscle strength and
lack of proprioception8 . even when
structural instability is excluded , functional instability can cause the weakening of the
ankle joint when movement patterns are repetitive , direction is abruptly changed9 , or active movement is carried out
intensely10 . in competitive elite
sports like soccer , these problems can manifest themselves as chronic issues such as a
higher risk of injury or functional abnormalities11 . the coordinated activity of the joints and muscles involved in
movements of the ankle joint further enhances the functional stability by improving stable
weight support as well as motor control function12 . through the measurement of isokinetic variables , the aim of this study was to assess the
impacts of functional and chronic asymmetry on the muscle strength of the left and right
knee and ankle joints in soccer players at a range of athletic performance levels , and to
generate useful data for the development of effective functional exercise programs for
symmetrical muscle strength . the 49 study subjects were recruited from 3 different levels of athletic performance :
professional soccer players ( k league , n=15 ) , amateur players ( k3 league , n=16 ) , and college
soccer players ( u league , n=18 ) . all of the participants provided written consent to
participate in the study , after the study procedure and methods were explained to them . table 1table 1.general characteristics of participantsvariablesnpp ( k league)ap ( k3 league)up ( u league)age ( yrs)1526.94.421.11.920.31.5height ( cm)16179.47.3175.54.1180.94.6weight ( kg)1877.58.070.44.676.66.1values are meansd . pp : professional players , ap : amateur players , up : university players shows the general characteristics of the subjects . all the subjects understood
the purpose of this study and provided their written informed consent prior to
participation , in accordance with the ethical standards of the declaration of helsinki . pp : professional players , ap : amateur players , up : university players the isokinetic muscle functions of the knee and ankle joints were the main experimental
variables of the study , and they were measured using the humac norm testing and
rehabilitation system ( csmi medical & solution , usa ) . subjects were seated in the
measurement chair , and the torque of the knee joint was aligned with the rotating axis of
the dynamometer by adjusting the position with a table - tube cross clamp and a pedestal
column clamp . the thigh area and the upper body were tightly fixed using a strap and a belt
so that the quadriceps exercise would not be affected by external force during the flexion
and extension exercises of the knee joint . additionally , to isolate the muscle strength of
the area of interest , the ankle was fixed with a strap by adjusting the length of the lower
leg and the adjustment axis with an adapter , after which the flexion and extension exercises
of the knee were performed . the range of motion was determined by measuring the maximum
flexion from the position in which the joint was extended ( 0 ) in a sitting position . care
was taken so that the joint would not be either hyperextended more than 0 or hyperflexed
more than 135. measurements were taken by group . as a warm - up exercise , the subjects
performed flexion and extension exercises of the lower limb 3 times below maximum and 1 time
at maximum , at angle speeds of 60/sec and 180/sec . then , the measurements were collected 5
times at the angle speed of 60/sec and 15 times at the angle speed of 180/sec . after the
measurements were obtained at each speed , the subjects were told to repeat the flexion and
extension of the knee joint , while resting for approximately 2 minutes in a sitting
position . next , the subjects were seated in the measurement chair in the same manner described above
for the knee joint , and the strength of the plantar flexion and dorsiflexion muscles in the
ankle were measured at angle speeds of 30/sec and 120/sec . after completion of a warm - up
exercise of plantar flexion and dorsiflexion 3 times below maximum and 1 time at maximum , at
the angle speed of 30/sec and 120/sec , measurements were taken 5 times at 30/sec and 15
times at 120/sec . after the performance was measured at each speed , the subjects repeated
the flexion and extension exercises , while resting for approximately 2 minutes in a sitting
position . for each measurement , all of the data analysis was conducted using spss for windows version 20.0 ( spss inc . ,
usa ) . means and standard deviations were computed for all the measurement items . to test
group differences , one - way anova was performed , and when significant differences were found ,
post - hoc comparisons were performed using the scheffe method . tables 2table 2.comparison isokinetic peak torque of the knee jointppapupextensor ( nm)60/secright207.823.2205.318.8254.134.0left202.733.1204.920.3246.943.6180/secright123.021.1118.629.3148.121.3left128.027.9120.022.4147.320.8flexor ( nm)60/secright133.628.8129.120.0152.038.1left131.525.2128.117.5145.125.2180/secright90.125.3111.919.295.424.3left91.922.5104.411.997.621.2values are meansd , * p<0.05 , * * p<0.01 , * * * p<0.001 . pp : professional players , ap : amateur players , up : university players and 3table 3.comparison isokinetic ratio of the knee jointppapup60/sec ( % ) right64.110.964.111.560.013.2left64.97.064.910.559.27.6180/sec ( % ) right72.613.564.111.566.220.3left71.810.764.910.566.713.9values are meansd . pp : professional players , ap : amateur players , up : university players show the results of the one - way anova on peak isokinetic strength and
differences in muscle strength of the knee and ankle joints , and the level of muscle
strength on the left and the right , according to level of athletic performance . peak
extensor strength of the knee joint was significantly higher in the university players ( up )
than the professional players ( pp ) or amateur players ( ap ) , both in the right and left knee
at the angle speed of 60/sec ( both p<0.001 ) and the angle speed of 180/sec ( both
p<0.01 ) . the peak flexor strength of the knee joint was significantly higher in ap than
pp or up , at the angle speed of 180/sec in the right knee ( p<0.05 ) . values are meansd , * p<0.05 , * * p<0.01 , * * * p<0.001 . pp : professional players , ap : amateur players , up : university players values are meansd . pp : professional players , ap : amateur players , up : university players tables 4table 4.comparison isokinetic peak torque of the ankle jointppapupplantar flexion ( nm)30/secright106.828.3125.025.8109.128.1left101.820.9109.221.2105.423.9120/secright51.718.880.823.462.920.8left56.820.076.134.758.717.0dorsi flexion ( nm)30/secright30.38.734.95.822.35.1left29.97.433.65.535.45.1120/secright16.27.019.64.29.92.2left17.25.918.43.717.22.6values are meansd , * p<0.05 , * * p<0.01 , * * * p<0.001 . pp : professional players , ap : amateur players , up : university playersand 5table 5.comparison isokinetic ratio of the ankle jointppapup30/sec ( % ) right28.54.028.77.021.14.6left30.18.331.77.835.610.3120/sec ( % ) right32.69.726.29.417.76.3left32.311.627.09.530.29.1values are meansd , * * * p<0.001 . pp : professional players , ap : amateur players , up : university players show the results of the one - way anova on the peak strength of the isokinetic
function in the ankle joint , as well as the ratio of muscle strength on the left and right ,
according to participant s level of athletic performance . the peak plantar flexor strength
of the ankle joint was higher in ap than pp or up at the angle speed of 120/sec in the
right ankle ( p<0.01 ) . the peak dorsiflexor strength was significantly lower in up than pp
or ap in the right ankle at the angle speed of 30/sec , and at the angle speed of 120/sec
( p<0.001 ) , and significantly higher in the left ankle at the angle speed of 30/sec
( p<0.05 ) . up showed a significant asymmetry in muscle strength between the left and right
sides compared to pp and ap at the angle speeds of 30/sec and 120/sec ( p<0.001 ) . the
ratio of the muscle strength of the ankle joint in the left and the right was significantly
lower for up than pp or ap in the right ankle at the angle speed of 30/sec and at the angle
speed of 120/sec ( p<0.001 ) . values are meansd , * p<0.05 , * * p<0.01 , * * * p<0.001 . pp : professional players , ap : amateur players , up : university players values are meansd , * * * p<0.001 . reinforcing muscle strength to maintain the stability of the knee joint is essential for
soccer players13 . deficiencies in
quadriceps muscle strength have a direct link to the risk of injury and can cause
osteoarthritis in the knee joint14 . it
has been reported that injury frequency increases in athletes when there is a difference of
10% or more between the strength of the left and the right quadriceps femoris15 , 16 . one cause of instability of the main joints in the lower limbs is
instability of flexion and extension movements , accompanied by loss of muscle strength ; this
instability can be manifested in a variety of forms17 . in the present study , up had significantly higher isokinetic strength of the quadriceps
femoris than pp or ap , and no asymmetry was observed in the left and the right muscle
strength . however , a previous study on the isokinetic strength of elite players and ap
demonstrated that the muscle strength rate of the hamstring was higher in elite players18 . although in that study it was observed ,
as in this study , that up had significantly higher quadriceps femoris strength , pp showed
balance between the quadriceps femoris and the hamstring and a higher muscle strength rate
of the hamstring . it is thought that the muscle strength rate of the hamstring is higher
because it enhances the stability of the knee joint to increase speed over short distances
and to allow for precise foot techniques . the muscle strength in the ankles of soccer players also differs depending on players
athletic performance level . a study that compared the isokinetic muscle strength in the
ankle joint of up and pp found that it was greater in pp , who were at a higher level of
athletic performance than up19 . in this
study , the plantar flexor strength was higher in ap compared to pp and up , and no asymmetry
in muscle strength in the left and the right was observed , whereas the dorsiflexor strength
was significantly lower in up compared to pp and ap . our finding is similar to that of a
previous study that reported that muscle strength imbalance in the foot joints , deficiencies
in proprioception , mechanical instability , and weakening of the peroneal tendon , which can
all be caused by repetitive training , overuse , and damaged soft tissue , are
interrelated20 . thus , it is believed
that the tibialis anterior , which is involved in dorsiflexion and inversion , is relatively
weak in muscle strength expression , in comparison to the strength of the soleus ,
gastrocnemius , and peroneal tendon that are involved in plantar flexion and eversion . muscle balance is an indicator employed to judge the level of athletic performance , and the
extent of muscle imbalance due to exercise and incorrect positioning is one factor that
influences performance21,22,23 . soccer is a
sport in which a variety of connecting movements , including running , abrupt stopping ,
changing direction , jumping and landing , kicking and passing , repeatedly occur and these
movements require complex functioning of the knee and ankle joints . the hamstring and the
calf muscle work to maintain the stabilized center of body mass or to push the ground
surface using reaction force , and the quadriceps femoris and dorsiflexors are used when
moving forward , kicking , or adjusting a ball by flexion of the hip joint , extension of the
knee , and dorsiflexion of the ankle . it is not only the balance between the agonist and
antagonist muscles , but also the balance between the left and the right , and between the
proximal joint and distal joint in a weight - bearing position that are involved during
exercise24 , 25 . as seen in the up in this study , the weakening of dorsiflexors due
to muscle strength asymmetry in the right ankle joint is thought to cause the strengthening
of the extension muscles of the knee joint in order to maintain the center of body movement
and stability . to maintain stability and balance in lower limb movement , it is essential to maintain
muscle activation26 . when a movement is
made by a soccer the player , the muscles in the foot are already activated to function
before the foot touches the ground27 , and
when the knee joint sustains a large shock like landing or when the center of mass moves ,
the contraction greatly increases in the quadriceps femoris28 . it is thought that muscle strength asymmetry in the ankle joint
may strengthen the muscles of the knee joint as a counterbalance in order to maintain the
center of body mass . in the future , to investigate the extent of muscle activation due to
asymmetry between left and right , studies must be conducted on muscle contraction as the
center moves with the use of kinetic variables . | [ purpose ] the aim of this study was to collect basic data on the effect of asymmetry on
the muscle strength of the left and right knee and ankle joints of soccer players at
varying athletic performance levels , to guide the development of improved exercise
programs . [ subjects and methods ] forty - nine soccer players at three athletic performance
levels participated : 15 professional , 16 amateur , and 18 college .
knee extensor and flexor
strength were measured at 60/sec and 180/sec , and ankle plantar flexor and dorsiflexor
strength were measured at 30/sec and at 120/sec .
variables were analyzed by one - way
anova .
[ results ] college soccer players showed greater muscle strength at 60/sec and
180/sec in the knee extension muscles of both the right and the left sides , lower muscle
strength at 30/sec and 120/sec in the dorsiflexor of the right ankle , and similar levels
of asymmetry between left and right . the maximum muscle strength on the same side
significantly differed in the right ankle joint , with asymmetry between left and right at
30/sec and 120/sec . [
conclusion ] these findings suggest that muscle strength asymmetry
in the ankle joint may lead to counterbalancing muscle strengthening of the knee joint to
maintain the center of body mass . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
understanding the developmental patterns of vegetative and reproductive organs and factors that influence those patterns is of critical importance for conserving rare plants . we recently determined that leaf and leaflet expansion of the endangered cycas micronesica were influenced by spatial and temporal factors , but reproductive organ expansion was unaffected or only minimally influenced by the same factors . we pointed out that empirical approaches for fitting models to organ growth and development may be used to inform horticultural or conservation questions of other rare cycad taxa . here we argue that this approach may improve our understanding of the evolutionary and developmental biology of cycads . while slow to gain traction in botany , especially in studies of gymnosperms ( but see refs . 2 and 3 ) , evo - devo has become one of the most promising and productive perspectives in biology . this should not be surprising given that development describes processes within a generation and evolution describes processes among generations . in writing about fossil cycads , mamay first suggested that angiosperm carpels evolved from megasporophylls , possibly of cycads . in more modern terms , examining regulatory genes , frohlich suggested that angiosperm carpels evolved from microsporophylls . these two hypotheses are probably incommensurate given that female and male strobili of cycads are homologous , unlike strobili of all other extant gymnosperms . cycad megastrobili and microstrobili are effectively a telescoped flush of leaves ( fig . 1 ) . both female and male strobili are still highly modified , so we still expect different developmental pathways in vegetative leaves , megastrobili and microstrobili . development in cycads is undoubtedly modular , as in other gymnosperms , although maybe not as modular as in angiosperms . thus ontogeny of various cycad organs should progress differently from other organs in the same individual plant . we expect different developmental timing in vegetative vs. reproductive organs because cycad strobili are likely highly endoploid . thermogenesis probably requires extra mitochondria , as we see in several skunk cabbage ( symplocarpus ) species . while almost exclusively documented in angiosperms , plants have plenty of endoploidy , especially succulent plants , which might include stems and strobilus axes in cycads . furthermore , animal tissues with high metabolic demand are often highly endoploid , such as heart muscles , flight muscles and liver cells . we suspect that endoploidy provides a way around otherwise rate - limiting production of mrna . efforts to confirm endoploidy in cycad strobili are warranted because endoploidy alters developmental rates insofar as mitotic cycle rates are inversely proportional to ploidy levels and probably inversely proportional to chromosomal content per nucleus , i.e. , c - values . there are four modes of evolutionary response to changing environments : ( 1 ) environmental tracking , ( 2 ) phenotypic plasticity , ( 3 ) bet hedging , and ( 4 ) extinction . a priori , we expect greater environmental tracking and phenotypic plasticity in vegetative structures because the individual plant can not grow without leaves . however , we expect more bet hedging ( risk aversion ) in reproductive organ production for any perennial plant ; especially in cycas micronesica , which makes large investments in both female and male strobili . but these are all mechanisms by which organisms evolve via selection . by contrast , gorelick and olson argued that drift should play a disproportionate role in cycad evolution compared with selection because of the peculiar genomic architecture and small population sizes ( see also ref . similarly , lynch and conery hypothesized that large genome size , which we see in cycads , may be maladaptive . thus developmental rates of different plant parts may be nothing more that what gould and lewontin mistakenly called spandrels . maybe there are no adaptationist explanations nor fitness benefits to the developmental patterns we described . or , alternatively , maybe these developmental patterns are nothing more than a corollary of endoploidy levels , which we presume are greater in thermogenic cones than in other plant parts . cycads are a threatened group of plants worldwide for which recovery plans have already been proposed or implemented for some taxa ( e.g. , see ref . . a full understanding of evolutionary developmental trajectories of cycads may be a prerequisite for restoring habitats during recovery efforts . thus far , we have only examined phenology and evolutionary responses in the most basal genus of cycads , cycas , with its disaggregated female strobili . because of the putative recent ancestry of all extant cycads , we anticipate similarities among all living cycads , which may not be due to any selective advantage . does plasticity of cycad male cone development differ from that of other gymnosperms ? because cycad male cones are presumably not homologous with those of other extant gymnosperms , the relative roles of selection and drift may not be the same in evolution of cycads vs. other gymnosperms . given that cycads are the most basal of living seed plants , it is incumbent upon us to better understand these fascinating plants . measurement of effective population sizes , plasticity , heterochrony , and endoploidy will help immensely in answering these evolutionary questions about cycads and may well help in their conservation . figure 1 . a cycas seemannii megastrobilus ( or indeterminate strobilus sensu in ref . 28 ) emerges as a flush of megasporophylls bearing naked ovules , which are almost certainly homologous to the pinnately compound vegetative leaves . the genus cycas contain some species with pinnately compound megasporophylls , e.g. , c. revoluta and c. pectinata , while all members of the genus have new flushes of vegetative leaves that emerge from the center of the megastrobilus , as seen in this figure , further indicating homology between sporophylls and vegetative leaves in cycads . | we recently described lack of phenotypic plasticity in reproductive organ development and substantial plasticity in vegetative organ development for the cycad cycas micronesica .
is there an evo - devo explanation for the disparity in phenotypic plasticity of vegetative vs. reproductive organs ? despite modularity , might evolution of cycad phenology be controlled more by drift than selection ? |
You are an expert at summarizing long articles. Proceed to summarize the following text:
the burden of chronic respiratory diseases ( crds ) is increasing worldwide , and knowledge about the physical , mental , and social consequences of these diseases is extensive [ 14 ] . generally , hospital outpatient clinics play a prominent role in the nordic countries ' health care systems . in denmark , the number of outpatient visits to somatic hospital departments increased by 34% from 4,917,000 in 2002 to 6,612,000 in 2009 . the outpatient respiratory medical clinic ( ormc ) receives referrals after the patients ' discharge from medical wards or by their general practitioner for the purpose of processing diagnosis , rehabilitation , and counselling or drug treatment . many patients with crd suffering from severe or very severe crd are followed up regularly in the ormc , according to the chronic care model guidelines and recommendations . patients who suffer from crd may be vulnerable in many ways : crd is associated with increased risk of anxiety , insomnia , fatigue , anorexia , loneliness , guilt , isolation , and depression [ 713 ] . as crd progresses , it is characterized by severe daily symptoms such as shortness of breath , cough , and increased mucus production . thus , these diseases have not only physical but also existential implications in terms of their existence - threatening progressive and , over time , terminal character [ 1416 ] . at the same time , it is well documented that many patients suffering from crd may have limited economic and educational resources and , over time , a reduced and limited network [ 1719 ] . in light of the fact that crds are characterized by their chronic , progressive , and lethal nature , there is a growing awareness of the need for more palliative - directed treatment and care for these patients [ 20 , 21 ] . as suggested in other studies , the outpatient clinics are subject to many temporal , structural , and substantial requirements that may impact what may be talked about and how long the conversation can last [ 2224 ] . these factors may limit opportunities to recognize and listen to patients ' experiences during interactions . several studies suggest that patients ' stories may offer important knowledge of how patients overcome , adapt to , and understand life with crd [ 10 , 26 ] . furthermore , these stories may reveal strong emotions and existential suffering in patients [ 2729 ] . the aspects of limited time and the many demands on the content of the consultation seem to influence what the health care professional ( hcp ) can hope to accomplish during counselling . several studies point to external factors affecting the lack of patient involvement and hcps ' responsiveness to patients ' existential and emotional suffering , including aspects such as increasing work pressure , numerous changes in the structure of tasks , implementation of new or changed administrative tasks , and increasing demands for documentation [ 25 , 31 ] . furthermore , the demands of patients regarding empathy and the recognition of their suffering may exhaust hcps and cause burnout [ 25 , 31 ] . at the same time , many studies suggest that hcps have a large potential to be instrumental in facilitating patients ' coping and self - care management and optimizing their quality of life or , conversely , damaging these processes [ 8 , 21 , 32 , 33 ] . over the last several decades , the former authoritative approach of the hcp has been replaced by an ideological , institutional , and also vocational desire to include chronically ill patients in their own care and treatment [ 3436 ] . today , being patient - oriented is immensely popular with health authorities in many western countries , as it reflects a moral philosophical approach in which the patient is regarded as unique and the multidimensionality of the human experience of illness is recognized through a relation that builds on multiple understandings of the patient 's situation [ 37 , 38 ] . studies show that this paradigm change is not always implemented and present during the patient - hcp interaction [ 19 , 39 ] . the studies emphasized here , based primarily on hospitalized crd patients , suggest that the interaction with patients may be difficult for multiple reasons . the assumption was , consistent with other studies in related areas , that several specific characteristics affect the outpatient interaction . these characteristics arise from the typically short - term and very structured content and time frame for each consultation , which is of relevance to the extent and the content of the interaction since the demarcated time for counselling and treatment may create limitations for the interaction and hcps ' ability to meet the needs of patients . whether this is the case in the ormc has not been addressed in other studies exploring how the short time sessions affect the interaction in similar or particular ways . there seems to be a lack of studies exploring the clinical encounter in the ormc [ 23 , 24 , 40 ] and none exploring the processes of interaction between patients and hcps . targeted attention to the patient 's perspectives , reactions to actions , and behaviour could contribute to increased insight into patient 's concerns and expectations . this study aims to contribute with knowledge in this area concerning the needs and behaviour of the patients and patients ' reactions to the actions of the hcp 's during the short - term counselling in the ormc . the aim of the study was to develop a theoretical framework explaining patients ' behaviour and actions related to unmet needs during interactions with health care professionals in hospital - based outpatient respiratory medical clinics . this study includes 65 field observations and 30 individual interviews with patients with a sample of 43 patients , 11 doctors , and 11 nurses , in total 22 hcps , from three ormcs in denmark during a period of 18 months in 2012 - 2013 . the patients ' experiences with the ormc varied and spanned from initial contact to four years ' association with several annually recurring consultations . the first 13 patients , 7 women and 6 men , were included exclusively for purposeful sampling conducting the initial field observations during patient - hcp interactions in clinic 3 . nine patients suffered from copd and 2 from fibrosis , and 3 did not have a definitive diagnosis at the inclusion time . out of the subsequent 30 included patients , 17 had copd exclusively , 4 had copd and asthma , 3 had emphysema , 2 had fibrosis , and 3 had other interstitial lung diseases . a total of 22 hcps , 11 doctors and 11 nurses , were included from the same 3 ormcs as the patients in the field observations . the number and the distribution of field observations on patient / health care professional interactions are shown in table 2 . the hcps were included to the extent that they were communicating with participating patients during the field observations , as illustrated in table 2 . the nurses and doctors worked in separate premises and with different tasks related to the patients . doctor - patient conversation lasted 1520 minutes and typically referred to the status of the disease , the effect of medicine , and possible revision , referring to rehabilitation in the municipality or hospital for further investigation and assessment . nurse - patient conversation lasted 1545 minutes and covered disease status , lung function tests , mrc breathlessness scale , and body mass index , as well as guidance in lifestyle changes and advice and recommendations related to these subjects . all included clinics used a primary nurse and doctor system for all crd patients , and nurses and doctors held separate individual consultations in the included clinics . the patients were included and observed at different stages of the outpatient clinic course , including initial consultations , subsequent visits , diagnosis meetings , and conclusive meetings . the patients were included upon arrival to the outpatient clinic and invited to participate , thus giving the researcher permission to observe the interaction during the visit . the patients were followed during the waiting time before and in between the conversations , which often included conversations with a nurse and then a doctor and occasionally a follow - up conversation with the nurse concerning medication intake or a new appointment in the ormc . all patients were followed during their entire stay in the clinic . before the patient left , a home visit or a phone interview the researcher 's field observations were nonparticipatory in the ormc during the counselling and interactions between the patients and hcps . between examinations and interviews , the researcher participated in conversations with the patients , brought coffee , and listened to stories about , for example , everyday life , individual coping with illness and disease , and the patient 's individual experiences of interactions with hcps . while listening and talking , the interviewer wrote down important points , ideas , and statements as memos . these relaxed and unrecorded conversations gave important information about the patients ' experiences regarding the ormc , everyday life at home , and relations with close relatives . insofar as it was possible for the patient , the follow - up interview was performed within a week of the visit to strengthen both the patient 's and the researcher 's ability to remember important topics for conversation or other areas that the researchers wanted to cover . out of the 30 interviews , 19 took place in the patients ' own homes and 11 interviews were performed by phone , in accordance with the patient 's needs and preferences . the interview guide was initially loosely structured , exploring issues concerning the patient 's perspectives , expectations , experiences and outcome of the interaction , view on health and illness , and the personal and unique challenges and joys of everyday life . the focus of data collection of the included hcps was to understand and explain how the hcp actions during the interaction triggered counterreactions and actions in patients . the included field observations of the hcps had the purpose of exploring the actions of the hcps to increase the understanding of what patients reacted to and how patients reacted and identifying the meaning and significance attributed to the interaction when the needs of the patients were predominantly unmeet . all interviews and field observations have been carried out with respect to the hcps ' and patients ' experiences and actions . it has been a balancing act to be loyal to the participants ' understanding of their actions and interpretations while being able to say something more and something different than the respondents could say . the project was carried out in accordance with icn 's code of ethics for nursing research and approved by the danish data protection agency ( j.nr . oral and written information about the study was given to the participants , including information on anonymity , informed consent , confidentiality , and the right to end participation at any time without stating any reason . all personal identifiers have been removed or disguised , and the participants are not identifiable and can not be identified through the details of the stories . the study is exploratory and based on charmaz 's social constructionist interpretation of grounded theory [ 4345 ] . an important source of inspiration from symbolic interactionism is an approach to the research field in which social processes are regarded as not only structural but also situational and changeable over time . in line with this way of thinking , this study 's focus is on researching the meaning and action in specific situations as close to an inner perspective as possible , acknowledging that it is not possible to duplicate the participants ' experiences . the coding process led to identifying what was happening in the data , explaining the elements of the emergent theory generating categories that were made more and more abstract as data were gathered to refine the theory . during initial coding , fragments of data words , lines , segments , and incidents were closely studied for their analytic import . during focused coding , the most fruitful initial codes were selected and tested against extensive data to determine their analytical importance to categories in accordance with the constant comparative method in grounded theory studies . the process of data collection and initial and focused coding ended when the categories were saturated and when gathering fresh data no longer revealed new theoretical insights or new properties of the categories . during the data coding and analysis , the focus was directed towards the fact that many patients apparently felt a lack of hcps ' responsiveness to their problems and perspectives during interactions and was given limited possibilities to influence what the conversation could include , which was found to be a prominent frustration for patients in the follow - up interview . the criteria for validating the findings followed charmaz 's credibility , originality , resonance , and usefulness [ 45 , 47 ] . a clear and rigorous working process , as described in the data analysis , assured credibility . originality can be met by the fact that this study offers new knowledge regarding patients ' responses to unmet needs during interactions in an underexposed area where knowledge is limited . to achieve resonance , several of the participating patients pointed out the importance of developing better understanding and visibility of their needs in the ormc . at the same time , the participating hcps showed great interest in the study and recognized , among other things , the need of the patients to express feelings during their stories in the time - pressed ormc , and they confirmed and recognized the experience of not having the time to examine patients ' perspectives of life experiences with illness . subsequent studies should examine , adjust , and adapt the findings to further shape the theory and identify possible practical implications for hcps ' practice . during the coding and analysis process , it became clear that the unmet needs of the patient triggered counterreactions initially conceptualized as resistance and adaptation behaviour . based on the data , a decision was made to pursue the findings of the unmet needs of patients based on patients ' perspective . but a research decision based on the data led us to unfold the patients ' behaviour and experiences related to interactions in which they did not feel seen , heard , and recognized focusing on partially as well as predominantly failed interactions on the basis of the patients ' comprehension . to investigate and compare processes related to patients ' need to be seen and heard , further investigation during data collection verified or modified this first impression and led to the sample included in this study as listed in table 1 . the analysis generated a substantive theory explaining how a main concern of many patients was striving to share the significant story comprising the significant individual issues related to their coping of illness or existential aspects of their everyday life . regardless of the patients ' behaviour , they rarely had the opportunity to tell their stories during the interactions . patients ' efforts to share their story and their reactions when this demand was unmet by the hcp triggered an adaptation or resistance behaviour , conceptualized as a fitting in or fighting back behaviour , explaining the patients ' counterreactions when they were not seen , heard , and recognized during the medical encounter . the lack of shared understanding to the content of the interaction is illustrated in figure 1 . behaviour , the hcp apparently assumed as if the patients were responsive and adaptive to the organized agenda . the fitting in and fighting back behaviour could overlap , balance , and shift during the interaction , and both behaviours could appear in the same interaction with varying strength at various times . sometimes they would emerge during the interaction , and , in other situations , the patient had already chosen behaviour to the expected responses from the hcp . neither of these two reactions created a basis for mutual understanding of the interaction , whether the patient adapted to a fitting in or fighting back behaviour . the counterreactions of the patients to hcps ' responses were affected by individual negotiation and adaptation in interactions regarding patients ' efforts to play an active role during the interaction , their wanting of the hcp to respond to emotional statements , and their opportunities to modify their own expectations of content and counselling during the interaction . these recurring structural modes of patients were conceptualized as role negotiation , emotional resonance , and perspective modification modes and determinations of the meaning and significance attributed to the interactions when the needs of the patients were predominantly unmet . the process attributed to the interactions when the needs of the patients were predominantly unmet is explained in figure 2 . the outcome of role negotiation , perspective modification , and emotional resonance in patients determined the behaviour of either fitting in or fighting back and whether the patient adapted to or resisted the hcp - planned guidance for procedures and issues during counselling . the hcps displayed several actions to modify , maintain , and control the content of the conversation . this includes hcps ' resignation to let the patients talk when they finished practical tasks . in other cases , the hcp could elude the patient 's story by interruption , distraction , or an appeal to the fixed programme of the counselling session in order to win back control of the conversation . in other cases , the time for each session was used as an argument to prevent the patient from telling his or her story : we have to move on to ( hcp 14 ) , if we are to finish all of this in time , we need to ( hcp 20 ) , let 's stick with the lungs , shall we ? ( hcp 8) we have to move on to ( hcp 14 ) , if we are to finish all of this in time , we need to ( hcp 20 ) , let 's stick with the lungs , shall we ? ( hcp 8) the various modification mechanisms used to minimize or exclude patients ' stories included lack of reply , interruptions , listening in silence , lack of eye contact or physical contact , or referral to limited time . in some cases , the actions of the hcp resulted in the patient continuously insisting on telling the story , but mostly they closed down the patient 's stories . fitting in behaviour conceptualized a passive patient role , which maintained a mutually recognizable and , on the surface , smooth , evolving , and developing framework for interactions . fitting in consisted of resignation , adjustment , and surrender behaviour during interactions by handing over control and the power to distinguish between important and less important topics of conversation . patients gave up telling their significant story and adjusted to the hcp 's objectives for conversation related to treatment or illness subjects concerning crd recommendations , hospital standards , and guidelines . fitting in behaviour created a harmonic and consensual interaction concerning the content of the case and a mutual confirmation of the good atmosphere . despite the fact that fitting in behaviour could be met with recognition and attention , this did not necessarily mean that patients were satisfied with the conversation or considered the interaction meaningful or relevant . behaviour often caused the hcp to assume the patient understood and recognized the importance and relevance of the counselling and treatment . mode did not necessarily result in patients ' receptiveness to counselling and information , as many patients silently resisted guidance while appearing responsive during the interaction . below is an example showing a patient 's fitting in behaviour and at the same time a resistance to counselling in the patient . during a field observation , the patient silently nods and does not argue with the hcp , who strongly disapproves of the patient 's continued use of cough medicine . 15 ) the fitting in behaviour resulted in patients rarely expressing their concerns , strategies , and emotions to the hcp . several patients were preoccupied with symptoms of their other chronic diseases , which they found to be even more disabling , or troubled by a lack of understanding from relatives or feelings of loneliness or anxiety , which the interaction often rendered impossible to discuss . by contrast , the patient 's questions could be context - related and in accordance with the hcp 's predefined subjects , resulting in patient - directed and problem - based counselling . fighting back behaviour demonstrated resistance , retention , and frustration to the unmet needs of the patient . the role of the patient and of the hcp could be challenged or disturbed by the story . a patient 's fighting back to create opportunities for the story without obtaining the hcp 's responsiveness could furthermore create emotions such as anxiety , anger , despair , or resignation in the patient , as exemplified below : x ( hcp ) , who we went to see , was very nice so i got angry when we went to see y ( hcp ) who does she think she is talking about my back instead of talking about my lungs i know how to treat my back so that was not at all what i needed and this commanding tone in her voice do this and do that . 15 ) x ( hcp ) , who we went to see , was very nice so i got angry when we went to see y ( hcp ) who does she think she is talking about my back instead of talking about my lungs i know how to treat my back so that was not at all what i needed and this commanding tone in her voice do this and do that . 15 ) resignation could cause that patients gave up fighting back and reverted to fitting in behaviour , which was less energy demanding and made the interaction more predictable and less exhausting for them . the fighting back behaviour could relate to greater or lesser part of the interaction and consist of more or less important elements of patient 's stories . fighting back behaviour was often visible throughout the interaction . it was particularly evident in the many interactions where patients wanted to bring up other topics than planned or where patients considered the counselling dull , time - consuming , offensive , or invading.well , it was at the end , as he was becoming kind of offended and sulky . and my annoyance was particularly with the fact that i felt he started to become a little rude . , i do not care whether it is negative or positive , but you must have some answers as to what the hell is wrong , has it become better or worse , what do we do and why not . it was at the end , as he was becoming kind of offended and sulky . and my annoyance was particularly with the fact that i felt he started to become a little rude . , i do not care whether it is negative or positive , but you must have some answers as to what the hell is wrong , has it become better or worse , what do we do and why not . ( pt . 24 ) in other cases , patients ' fighting back behaviour seemed more veiled and expressed through a return to issues that the hcp had attempted to finish or through insistence on sharing a narrative or expressing their views on alternative treatment or the way they tackled common problems related to the illness . many patients considered the bodily learned knowledge as important and tried to share these stories , even though the hcp rarely called for or responded to their experiences . in figure 3 the overall grounded theory is presented . this explains how striving to share the significant story triggered a fitting in or fighting back behaviour based on a process of role negotiation , emotional resonance , and perspective modification leading to the meaning and significance in patients attributed to the interactions when the needs of the patients were predominantly unmet . the wavy arrow - line down the centre of the figure illustrates how a fitting in as well as a fighting back behaviour in patients undergoes the same process of identifying perceived opportunity to achieve recognition through the process of role negotiation , emotional resonance , and perspective modification . the text below the figure explains how these processes in patients are leading to either a pretended shared understanding or a visible or veiled resistance to counselling ( figure 3 ) . role negotiation describes the struggle for allocation of roles during the interaction and conceptualizes how the patient , silently or vocally , negotiates the patient role , including the right to define the content , the subject , and the framework for the interaction . patients could basically resist or adapt to the expected passive patient role the hcp expected them to undertake during interactions . at the same time , the hcp strived to uphold the dominant expert role and the right to define the content of the interaction , reacting with irritation , resignation , or frustration when challenged in the distribution of roles during the interaction . hcps displayed several actions to accommodate this , including disregard , interruption , or breakdown of the story . role negotiation can be explained as patient - driven breakdowns , surrender , or attempt to control the content of the interaction in the ormc . when the interaction caused breakdowns , it was often clear to both parties . many of the hcps experienced these situations as unpleasant and tried to prevent or alleviate the disharmony caused by it . breakdown situations clarified the patient 's anger or frustration when the hcp 's abortive assessment of the patient 's emotional state became visible to both sides . in one of the field observations , a dialogue between a patient and a nurse takes place illustrating the breakdown of the counselling . the patient suffers from posttraumatic stress syndrome , which , according to the patient , is a much greater challenge in his everyday life than his copd . early in the observed conversation , the hcp asks about the patient 's daily number of cigarettes . it 's just bullshit to quit smoking when you have ptss ( shakes his head).hcp.1 : how about nicotine patches?(pt . 22 ) : yes , but i ca n't light them up , can i ! ( laughs ) ( field observation , pt . 22 ) ( pt . it 's just bullshit to quit smoking when you have ptss ( shakes his head ) . 22 ) : yes , but i ca n't light them up , can i ! ( laughs ) ( field observation , pt . 22 ) the patient explained that he did not have the power to continue trying to make the hcp understand what his self - perceived biggest problems were : anxiety and water retention . in the follow - up interview , he pointed out that cigarettes were his only consolation and relief , the only thing that could alleviate his anxiety and ease his suffering . even though patients mostly were aware that the hcp was not receptive to their story , it did not necessarily mean a change of behaviour . patients ' fighting back against the expected patient role rarely created opportunities for their story but instead complicated and disturbed the maintenance of the planned content at the ormc . on the other hand , breaking or challenging the patient role created an opportunity to prevent the hcp from communicating information or recommendations to the patients they did not want to receive . this happened through irony , anger , humour , laughter , or bodily nonverbal rejection of the counselling . there was a connection between breaking or challenging the traditional patient role and the patient 's experience of anger directed at the hcp and , at the same time , an experience of the ormc as a place where the patient 's problems were neither met nor recognized . patients could mitigate any previous challenge of the patient role by surrender through recognizing the hcp 's advice or showing gratitude for the hcp 's efforts to help them . when accepting the passive role , the patients surrendered and gave up expressing their wishes for the conversation and their needs to share and explore hope , emotions , and strategies . instead , the patients followed the hcp 's agenda during interaction and counselling . during the follow - up interview , the patients often expressed feelings of disappointment and dwindling expectations for the ormc and the hcp . breakdowns often allowed the patient to take control of the interaction for a limited period of time . regardless of the outcome of the role distribution , the patients ' need and effort to share their significant story transformed their hope of creating opportunities during the interaction to a state of resignation , anger , or disappointment . the patients ' self - perception was often challenged and negotiated during interactions with the hcp in which patients tested their own attitudes and actions . through this , patients were able to adjust to the situation and modify their own reactions to the actions of the hcp . the hcp 's violation , indifference , or recognition was thereby turned into something meaningful or meaningless by the patient . through the story , patients modified and tested their own stances towards everyday life and the disease , strategies , self - perception , and the things perceived as important / not important for living with crd . a consensus emerged between the hcp and the patient that it was the hcp 's prerogative to comment on and reprimand the patient on issues such as weight , alcohol consumption , smoking , social network , moods , and strategies . these stories involved compromises on lifestyle changes that the hcp encouraged the patients to follow . the hcps displayed inertia in their response to these compromises and a lack of involvement in the patients ' own experiences of and tricks for coping with crd . a pattern emerged in which the hcp often displayed an inflexible approach to counselling on lifestyle changes , regardless of the prerequisites , interest , or motivation of the patient . the hcp rarely responded to patients ' own suggestions of strategies for living with breathlessness , which sometimes decreased patients ' adjustment to the hcp counselling.yes , i often see sulky attitudes at that outpatient clinic , actually i mean , i do n't think that they are very enthusiastic encouraging making people want to pitch in i think that they are a bunch of grumpy ladies and i think that i wasted my time by driving up here i 'd much rather go see xx , because he 's sympathetic and seems enthusiastic about the things that you tell him and what you accomplish on your own , right ? 4 ) yes , i often see sulky attitudes at that outpatient clinic , actually i mean , i do n't think that they are very enthusiastic encouraging making people want to pitch in i think that they are a bunch of grumpy ladies and i think that i wasted my time by driving up here i 'd much rather go see xx , because he 's sympathetic and seems enthusiastic about the things that you tell him and what you accomplish on your own , right ? ( pt . 4 ) some patients pointed to individual hcps whom they considered to be particularly skilled and motivating in their effort to create lifestyle changes by adjusting and finding new ways and strategies . these hcps displayed a recognizing approach to the patients ' own actions in relation to lifestyle changes and reacted to the patients ' emotional condition with advice , comfort , or encouragement . the presence or absence of emotional resonance was the key determinant of patients ' self - perceived satisfaction or dissatisfaction to interactions and was characterized by patients ' need to achieve emotional response and recognition during the interaction . when the patients experienced a lack of emotional response to their story 's emotional content , it resulted in additional suffering and discouragement in the patients . many patients were striving to share some kind of hope expressed by the story and , at the same time , a need to have the emotions related to hope in the story recognized . the hcp 's rejection , ignoring , or lack of recognition of the emotional part of the story contributed to extinguishing the hope of less suffering , distress , and worries of everyday life . many patients expressed the importance of emotional responsiveness during the interviews : i told her that i would like to quit the medicine and she told me to forget about it , but i told her that i had hoped , but she would n't listen and at that point in time , i just really needed the hope i mean that they understood that it was a hope of mine or a milestone to me , right ? 4 ) i told her that i would like to quit the medicine and she told me to forget about it , but i told her that i had hoped , but she would n't listen and at that point in time , i just really needed the hope i mean that they understood that it was a hope of mine or a milestone to me , right ? 4 ) the lack of emotional resonance negatively affected the patients ' self - perceived motivation and courage to create everyday lifestyle changes . furthermore , the hcps ' lack of emotional reflection on patients ' challenges tended to increase the patients ' fighting back mode to recommended lifestyle changes . the patients often reflected upon their own health and disease related choices and actions . patients were aware that emotions such as triumph , pride , and satisfaction from reducing cigarettes or reducing medicine would not give rise to recognition by the hcp . even if they wanted to share their enthusiasm for their own lifestyle changes , these stories were rarely considered good stories from the hcp 's point of view . apparently , there was a discrepancy between patients ' and hcps ' experience of what a good patient story might imply . in some cases , the patients reacted to the discrepancy with anger , frustration , or fighting back towards the hcp . other patients regarded the interaction as violating and did not feel encouraged to enter into a dialogue about lifestyle habits.no one bloody told me damn , you managed to go from 30 cigarettes per day to 5 , well done . 7 ) no one bloody told me damn , you managed to go from 30 cigarettes per day to 5 , well done . 7 ) when the emotions in the story remained unrecognized , counselling sometimes became more anxiety provoking , incomprehensible , or less relevant , resulting in increased disappointment , despair , or hopelessness in patients . when the hcp recognized and reflected the story about anxiety , despair , joy , victory , insecurity , resignation , or annoyance , it had a tremendous effect on patients . it was consistent across the data that patients responded positively and often with gratitude and appreciation when the hcp recognized and positively indicated an understanding of their difficult everyday life . it motivated them when the hcp recognized and really listened to the story and had great importance when they felt emotionally understood , when they were awarded a significant role , and when their effort to find strategies and perspectives was recognized . some of the hcps had the ability to reflect the patients ' emotions by acting interested and engaged in their life . it 's not easy for you and i understand your plight or leave - takings like keep up your spirit till we meet again had great value for patients , who mentioned these hcps as particularly talented , caring , and encouraging . the patients ' stories of hope could be expressed by and converted into , for example , striving for more good days with less breathlessness , reducing the number of cigarettes , or using less medicine . it was consistent across data that , to the extent the patients were emotionally reflected , they found counselling to be more relevant and were more willing to explore the perspectives regarding the hcps ' attitudes to an optimal life with crd . the original intention of this study was to explain interpretations and counterreactions to unmet needs in the patients during counselling with hcps in hospital - based outpatient respiratory medical clinics . patients ' efforts to share their story and the subsequent rejection or ignoring of this pursuit triggered predominantly an adaptation or resistance behaviour , conceptualized as fitting in and fighting back , explaining the patients ' counterreactions to unrecognized needs during the medical encounter , which turned out to be an exhausting but necessary behaviour to maintain perspectives and stories during the interactions . in addition , the patients ' counterreactions were based on whether it was possible to play an active role during the interaction , the extent to which hcps responded to the emotional intentions , and how the patients were able to change their own expectations of content during the interaction . these recurring structural patterns of interpretations of the patients , which were conceptualized as role negotiation , emotional resonance , and perspective modification conditions , turned out to be decisive of the meaning and significance patients attributed to the interaction and thus whether the patient adapted to a fitting in or fighting back behaviour . in accordance with the findings in this study , the patients ' capability of being open to guidance was related to the extent to which they felt recognized and experienced the opportunity to share their concerns , suffering , and hopes with the hcp . the findings do not provide any answers to what seems to be a significantly greater degree of willingness to adhere to disease related issues and , to a lesser extent , an attachment to the emotional needs of patients . in addition , other studies have suggested that crd patients lack knowledge of the disease and its long - term effects [ 48 , 49 ] and that patients experience limited access to specialized nurses and doctors within the field of lung diseases [ 48 , 50 ] . at the same time , it has been shown that lifestyle - related counselling and medicinal treatment at outpatient clinics can prevent readmittance , increase the level of patient satisfaction , and increase patients ' ability to cope with illness and optimize drug compliance [ 15 , 24 , 51 , 52 ] . however , before discussing the implication of these patterns of behaviour for clinical practice , it is essential to emphasize that although a general pattern of lacking commitment and responsiveness to patients ' suffering in the ormc was found , it may have complex causes and contexts , as suggested in other studies [ 25 , 30 ] . since this study maintains a patient - oriented perspective on the interaction , it is central to outline the limits and possibilities that affect the hcps ' actions in and attitudes to the interaction in the ormc . several studies suggest possible explanations for the lack of commitment to patients ' emotional needs . studies have shown that it is a difficult and complex task to maintain individual patient - oriented and emotionally responsive counselling , pointing to a high risk level of burnout and emotional exhaustion in many hcps caused by the emotional needs , expectations , and demands of responsiveness from patients [ 25 , 30 , 31 ] . in terms of emotional burnout , lack of reciprocity , and attached concern , many studies suggest that the lack of responsiveness to patients ' needs may be a way to resist emotional strain and stress [ 53 , 54 ] . it may be a difficult task to listen to patients ' stories of suffering , not knowing how to help . patients are often severely ill , treatment options poor , and healing possibilities nonexistent , and the existential suffering may be overwhelming to patients [ 1 , 34 ] . at the same time , demands to navigate a time - constrained and topic - bounded guidance session involving many compulsory subjects to be documented may leave limited time for individual and patient defined selection of topics for conversation . additionally , hcps are portrayed in a highly critical light in several studies , expressed particularly through the postmodern critique emerging from concepts like discipline , power , and individual control as problematic in the health care system [ 5658 ] . the criticism that this study to some extent supports can give a misleading picture of the knowledge that hcps possess and the practices associated with it . it needs to be taken into account that the hcps included in this study had extensive knowledge and experience with crd . mostly they played a central role for patients in treatment and disease control , and they expressed a desire to support and help patients who suffered from crd in the best possible way within the framework they had and the possibility they could recognize . at the same time , it is important to stress that the participating hcps had extensive practical and theoretical knowledge regarding the consequences of the lifestyle choices made by patients suffering from crd that they wish to introduce to patients , including knowledge on topics such as the benefit of exercise , smoking cessation , body weight , and medicine intake that may lead to extended life span and higher quality of everyday life in crd patients [ 1 , 34 ] . this means that there are many and often medically rational explanations to the hcps ' focus on control , monitoring , and treatment as found in this study . the vulnerability of patients suffering from crd , which is prominent in our study and confirmed in other studies , stresses the importance of a supporting , empathic , and encouraging approach in hcp counselling [ 19 , 39 ] . although several other studies examine patient behaviour and actions [ 28 , 39 , 59 ] , no other studies provide knowledge about the processes that constitute crd patients ' behaviour of fitting in or fighting back to counselling during interactions with hcps in the ormc . this study underlines the possibility of improving the experiences of being recognized in the ormc through susceptibility of hcps to listen to the stories of illness and suffering of the patients . the findings in this study suggest that an increasingly empathetic and extensive understanding of patients ' concerns may possibly lead to a lesser degree of patients ' resistance to advice and counselling during interactions . the findings suggest that patients rarely expected solutions to problems and concerns but rather anticipated an empathetic approach and willingness to listen to and understand their stories of suffering , hope , and ways of overcoming illness . this study offers new knowledge in an underexposed research field where the interaction is characterized by short meetings subject to a well - defined agenda during the clinical encounter . in accordance with other studies , our findings point to the importance of supplementing the disease - oriented perspective with a perspective increasingly inclusive of an empathetic and extensive understanding of patients ' concerns and an appreciative view of patients [ 56 , 60 ] . further research is required to explore patients ' counterreactions to unmet needs and further disclose the content and importance to patients with crd in the ormc . this study explores the patient 's behaviour and actions related to interactions with hcps in the ormc . this study is limited by the fact that the voice of the hcps is not explored in this study . further exploration of the hcp 's perspectives may provide further understanding of the knowledge of the ormc interaction , which this study can not provide . future research should explore the hcp 's reactions to the patient 's actions and behaviour in the ormc to provide an overall picture of reactions and counterreactions of both patients and hcps . the findings of this study allow for a better understanding of patients ' counterreactions in the time - pressured and , simultaneously , tight structured guidance program in the ormc . firstly , the findings show that patients ' efforts to share their story triggered predominantly an adaptation or resistance behaviour , conceptualized as fitting in and fighting back , explaining the patients ' counterreactions to unrecognized needs during the medical encounter . secondly , counterreactions of the patients were based on whether it was possible to play an active role during the interaction , the extent to which hcps responded to the emotional intentions , and how the patients were able to change their own expectations of content during the interaction . the patients ' capacity to receive help and guidance was related to the extent to which they felt recognized and experienced the opportunity to share their concerns , suffering , and hopes with the hcp . the study offers new knowledge in an underexposed research field where the interaction is characterized by short meetings subject to a well - defined agenda during the clinical encounter . the findings point to the importance of supplementing the disease - oriented perspective with a perspective increasingly inclusive of an empathetic and extensive understanding of patients ' concerns and an appreciative view of patients . an increasingly empathetic understanding of patients ' concerns may lead to less resistance in patients to hcp advice and counselling during interactions . further research is required to explore patients ' counterreactions to unmet needs and further disclose the content and importance to patients during counselling in the crd . the findings of patients ' counterreactions to unmet needs provide new knowledge to hcp practice regarding the interactional possibilities and limitations in the ormc . the findings offer practical and ethical implications as to how hcps ' attitudes towards patients can increase their ability to support emotional suffering and increase patient participation and guidance in the lifestyle changes that many hcps hope to achieve during counselling . throughout this paper , how the lack of emotional responsiveness and recognition triggered a fitting in or fighting back behaviour has been outlined and analysed , explaining the patients ' counterreactions to unmet needs during the interaction in the ormc . this paper emphasizes a lack of shared expectations between the patient and hcp as being a central problem in the interaction , proposing that hcp may involve utilising the already available time frame to conduct an open dialogue with patients and prioritising the tasks and issues that are most important for the patient quality of life and health at the time . the need to develop specific models or guidelines , while focusing on the time - dependent ormc interaction , is crucial in order to develop and strengthen an ormc based on empathy , ethics , and emotional responsiveness to patients ' needs and suffering during interaction in the ormc . a concrete proposal could be , for example , that future ormc visits included an initial exploration of patients ' experiences with the disease , hopes , ambitions , and expectations for the ormc interaction . furthermore , not only may a narrative approach be used in academic writing , but at the same time , it should be implemented as a way to forward the development of the ormc . actively using patient 's stories is a way of understanding how the patient experiences can be understood and what is perceived as meaningful to patients regarding support , treatment , and guidance . through this , the ormc could gain development of the ormc practice . narrative methods seem to embrace the possibility of a more individual and patient - directed guidance , focusing on patient experienced problems rather than on the prospects based on the problems that hcps perceive as typical , general , and specific to patient groups in the ormc . this approach would consequently be a shift in perspective and complement the hcp 's understanding of the ormc aims , tasks , and possibilities . | aim . developing a theoretical framework explaining patients ' behaviour and actions related to unmet needs during interactions with health care professionals in hospital - based outpatient respiratory medical clinics . background .
the outpatient respiratory medical clinic plays a prominent role in many patients ' lives regarding treatment and counselling increasing the need for a better understanding of patients ' perspective to the counselling of the health care professionals .
design .
the study is exploratory and based on charmaz 's interpretation of grounded theory
. methods .
the study included 65 field observations with a sample of 43 patients , 11 doctors , and 11 nurses , as well as 30 interviews with patients , conducted through theoretical sampling from three outpatient respiratory medical clinics in denmark .
findings .
the patients ' efforts to share their significant stories triggered predominantly an adaptation or resistance behaviour , conceptualized as fitting in and fighting back
behaviour , explaining the patients ' counterreactions to unrecognized needs during the medical encounter . conclusion .
firstly this study allows for a better understanding of patients ' counterreactions in the time - pressured and , simultaneously , tight structured guidance program in the outpatient clinic .
secondly the study offers practical and ethical implications as to how health care professionals ' attitudes towards patients can increase their ability to support emotional suffering and increase patient participation and responsiveness to guidance in the lifestyle changes . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
ipsilateral floating hip and floating knee are very rare injuries . these injuries so uncommon that only three cases of similar kind have been reported . management of such complex injuries is a challenging task even in experienced hands as there are no standard treatment guidelines for such fractures . we hereby report a 20 yr old male who sustained ipsilateral floating hip and ipsilateral floating knee injuries following motor vehicle accident . patient was treated with interlocking nail for femur and tibia in the same sitting whereas acetabulam fracture was managed conservatively . at five months all the fractures united well with restoration of good range of motion in both hip and knee ipsilateral floating knee and floating hip are very rare injuries seen following high velocity motor vehicle accidents . there are no standard guidelines for treatment of those fractures as only a few cases of similar kind have been reported in literature . ipsilateral floating hip and floating joint injury is a simultaneous skeletal disruption above and below a joint . this type of uncommon fractures results from high energy trauma with a complex mechanism of injury . to best of our knowledge only three cases of similar kind have been reported in literature [ 2 - 4 ] . we hereby report a twenty year old patient who sustained ipsilateral floating knee and hip injury on right side following a motor vehicle accident . a 20 year old male travelling in a motorcycle struck a car and sustained injuries to right lower limb . radiographs revealed undisplaced posterior column fracture and displaced femoral shaft fracture at distal third [ fig 1 ] . displaced tibia shaft fracture of mid third was seen on the same side [ fig 2 ] . ct scan pelvis confirmed undisplaced posterior column fracture and ruled out other injuries [ fig 3 - 4 ] . preoperative photograph showing tibia fracture preoperative photograph showing femur fracture preoperative radiograph of hip joint undisplaced posterior coloumn fracture seen on ct scan patient was taken up for surgery next day . posterior column fracture stability was checked after femur and tibia nailing under fluoroscope and was found to be stable . patient was advised bed rest for six weeks . at six weeks bone grafting of femur fracture all the fractures united by five months and patient returned daily activities [ fig 5 - 8 ] . at one year follow up patient is walking full weight bearing without aid and had good range of motion in both hip and knee joints [ fig 9 - 10 ] . floating knee and floating hip injuries render both hip and knee joint unstable both proximally and distally . the injury usually results from a collision between a pedestrian or a motorcyclist and a motor vehicle as seen in our case . there is enough literature for management floating knee and floating hip , but there is no sufficient literature in the management of simultaneous ipsilateral knee and hip floating injuries [ 6 - 8 ] . they concluded that operative stabilization of the femur and acetabulam gave the best clinical results , while non - operative treatment of acetabulam fractures had to be the treatment of choice in undisplaced or minimally displaced fractures [ 9 - 10 ] . described five patterns of ipsilateral tibial and femoral fractures and made treatment recommendations on the basis of these patterns . because of high prevalence of complications after closed treatment , operative stabilization of both fractures is recommended even for young children . veith r g et al treated fifty - seven consecutive ipsilateral fractures of the femur and tibia in fifty - four adults . twenty - one patients had concomitant life - threatening injuries , and in thirty - three extremities the fractures were open . local complications included below - the - knee amputation , three deep infections , and four ununited fractures . mortality rates from floating knees range from 5% to 15% and amputations are reported in approximately 25% of patients [ 11 - 16 ] . in our case there was only restriction of terminal 20 degree of terminal flexion in knee joint on the affected side . we opted not to nail both shaft fractures thorough same incision to avoid knee stiffness which is a notorious complications following such injuries . all the fractures were united by five months and patient was able to do routine daily activity . we want to emphasize the advantages of early fixation and aggressive mobilization of such complex fractures . modern anaesthetic and surgical advances have made early fixation of multiple fractures a realistic and often life saving alternative . we want highlight through this case report that such injuries are very very rare and only few cases of similar kind have been reported in literature . ipsilateral floating knee and floating hip are very rare injuries seen following high velocity motor vehicle accidents . there are no standard guidelines for treatment of those fractures as only a few cases of similar kind have been reported in literature . early fixation and aggressive mobilization ensures fracture union and fewer complications although ipsilateral floating knee and hip fractures are rare , the management depends on personality of individual fractures . rehabilitation should be aggressive and good results can be expected depending on severity of primary trauma . | introduction : ipsilateral floating hip and floating knee are very rare injuries .
these injuries so uncommon that only three cases of similar kind have been reported .
these injuries are due to high velocity injuries following motor vehicle accidents .
management of such complex injuries is a challenging task even in experienced hands as there are no standard treatment guidelines for such fractures.case report : we hereby report a 20 yr old male who sustained ipsilateral floating hip and ipsilateral floating knee injuries following motor vehicle accident .
patient was stabilized initially and later taken up for surgery .
patient was treated with interlocking nail for femur and tibia in the same sitting whereas acetabulam fracture was managed conservatively . at five months
all the fractures united well with restoration of good range of motion in both hip and kneeconclusion : ipsilateral floating knee and floating hip are very rare injuries seen following high velocity motor vehicle accidents .
there are no standard guidelines for treatment of those fractures as only a few cases of similar kind have been reported in literature .
early fixation and aggressive mobilization ensures fracture union and fewer complications . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
cancer cell metabolism is a direct result of the modulation of intracellular signaling pathways that are disrupted by mutated oncogenes and tumor - suppressor genes . mutated oncogenic genes metabolism comprises an energy - producing process of which cells have advantages for the maintenance of cell homeostasis as well as growth and proliferation . normal cells are equipped with complex signaling networks that are orchestrated by key control enzymes , sensing environmental cues and operating metabolic machineries to provide sufficient energy for survival in a perfectly controlled manner . during proliferation , normal cells activate metabolic pathways to accommodate increased adenosine triphosphate ( atp ) consumption for cell reproduction . along with this metabolic boost , unavoidable by - products of aerobic metabolism , such as reactive oxygen species , may unfortunately damage cells and promote dna mutations . mutations of oncogenes and tumor - suppressor genes can modify multiple intracellular signaling pathways and , in turn , alter cell metabolism to facilitate the tumorigenic process . altered signaling pathways not only enable cells to adapt to tumor cell metabolism , but several of these metabolic alterations are also essential for malignant transformation . the characteristic metabolic hallmark of tumor metabolism is aerobic glycolysis or the warburg effect ; named after otto warburg who first described it in 1926 . unlike normal cells that produce energy mostly through the oxidation of pyruvate in the mitochondria , cancer cells predominantly produce energy via enhanced glycolysis in the cytosol , even under aerobic conditions . most cancer cells use glycolysis as a means of energy production regardless of whether they are under normoxic or hypoxic condition . the correlation between glycolytic atp production and tumor malignancy it was originally hypothesized that these metabolic changes reflect damage to mitochondrial oxidative phosphorylation , implying that cancer cells could not respire properly to obtain sufficient atp . however , recent studies revealed that many cancer cells are capable of synthesizing atp through mitochondrial respiration . regardless of whether mitochondrial respiration is reduced , the fact remains that cancer cells exhibit high rates of glycolysis and lactate fermentation , and this dependence on glucose utilization may be exploited for therapeutic intervention . during the past decade , many studies focusing on mitochondrial function in cancer cells have suggested that the warburg effect is more closely related to alterations in signaling pathways that control the uptake and utilization of glucose than to mitochondrial defects . the warburg effect has recently gained attention again in the cancer research field , because researchers have begun to re - evaluate the significance of aerobic glycolysis in tumor cells . aerobic glycolysis in cancers is the combined result of oncogenes , tumor suppressors , a hypoxic microenvironment , mtdna mutations and others . understanding the complex cancer energy metabolism will help to develop new approaches in early diagnosis and cancer therapy . this review will discuss the metabolism of tumor cells , focusing on the analysis of altered enzymatic activities that are involved in aerobic glycolysis , and discuss current strategies of targeting metabolic pathways for cancer treatment . glucose is the primary source of energy and is the main fuel for cellular respiration . in glucose utilization in normal conditions , 70% of atp is known to be synthesized by oxidative phosphorylation and the rest by glycolysis . as the atp yield varies with cellular conditions , the ratio between glycolysis and oxidative phosphorylation also varies in different cells , growth states and microenvironments . in hypoxia , for example , enhanced glycolysis compensates the compromised oxidative phosphorylation to maintain the cellular energetic balance . in most solid tumor cells , a switch in metabolism towards glycolysis over respiration in spite of their functional oxidative phosphorylation machinery manifests the cancer - specific aerobic glycolysis . when the oxidative phosphorylation machinery is limited for any reason ( hypoxia , suppression of mitochondrial respiration and so on ) , cancer cells require a vast amount of energy in a short period of time in order to proliferate . muscle cells in hypoxic conditions and embryo cells during development can adapt to an altered metabolism when oxygen supply is limited or when growth rate exceeds the regular energy supply , respectively . these cells convert glucose to lactate for fast , but less efficient , atp generation , accompanying high glucose consumption . likewise , cancer cells exhibit altered metabolism to meet energy needs during tumor progression ; which is known as the warburg effect . the warburg effect involves a metabolic shift from oxidative phosphorylation to glycolysis or lactate fermentation , even in the presence of oxygen . otto warburg originally proposed that a mitochondrial defect disrupted mitochondrial respiration resulting in an increase in glycolysis to meet the energy demand of cancer cells . alternatively , hypoxia , which is common in cancerous tumors , can relieve oxygen inhibition of the glycolytic pathway ( pasteur 's effect ) , resulting in anaerobic glycolysis . however , it was later determined that cancer cells utilize aerobic glycolysis for energy generation instead of mitochondrial respiration , even in the presence of normally functioning mitochondria . converting glucose to lactate , rather than metabolizing it through mitochondrial oxidative phosphorylation , is far less efficient , as the former process generates fewer atp molecules per unit of glucose . therefore , a high rate of glucose uptake is required to meet the increased energy needs involved in supporting rapid tumor progression . the dependency on fermentative metabolism for atp generation is a long - term metabolic reprogramming in transformed cancer cells where glucose uptake and glycolytic activities are markedly increased . even under normoxia , an acute repressive signaling cascade imposed onto mitochondrial function is commonly observed . transformation of normal cells into cancer cells involves a long - lasting irreversible metabolic switch toward activation of glucose transport and utilization in addition to the suppression of mitochondrial respiration . if the function of respiratory machinery is normal , the glycolytic activity will be regulated via different pathways to maintain a balance of energy . although glutamine was not emphasized by warburg , glutamine is an essential bioenergetic and anabolic substrate for many cancer cell types . cancer cells exhibiting aerobic glycolysis rely on glutamine as well as glucose as the carbon source . glutamine is used to provide intermediates of the tricarboxylic acid ( tca ) cycle to feed other biosynthetic pathways as precursors . therefore , cancer cells are dependent on glutamine to maintain the tca cycle , as they rely on glucose during aerobic glycolysis . many studies have shown that metabolic alterations of glucose provide additional energy to support tumor growth . moreover , the inhibition of biosynthetic pathways that are related to glycolysis decreases tumor growth . these suggest that targeting glycolysis may be a useful strategy for preventing or stopping the development of cancer . mutations that activate oncogenes or inactivate tumor suppressors can significantly affect activities of metabolic enzymes and have a key role in aerobic glycolysis of cancer . among oncogenic mutations , alterations in phosphatidylinositol 3-kinase ( pi3k ) , phosphatase and tensin homolog ( pten ) , myc and p53 can all impact cellular metabolism ( figure 1 ) . the pi3k pathway has frequently been shown to be altered , and thus has a major role in tumor proliferation and survival for a wide variety of human cancers . activation of pi3k results in the downstream activation of akt and stabilization of hypoxia - inducible factor ( hif)-1 . the pi3k enzyme itself antagonizes the tumor - suppressor pten , and the loss of pten increases glycolysis by activation of akt and hif-1 . akt stimulates glycolysis by increasing the expression and membrane translocation of glucose transporters , and also by phosphorylation of glycolytic enzymes , such as hexokinase ( hk ) and phosphofructokinase ( pfk ) . moreover , akt activates mammalian target of rapamycin ( mtor ) , which indirectly affects other metabolic pathways by activating hif-1 , even under normoxic conditions . oncogene mutation of ras activates mtor via the pi3k - akt - mtor signaling pathway , and mtor promotes glycolysis through inducing hifs . the hifs are transcription factors that regulate gene expression in response to the cellular environment , specifically in the event of decreased oxygen or hypoxia . hif-1 regulates many different genes such as vascular endothelial growth factor , hepatocyte growth factor receptor ( c - met ) , erythropoietin , transforming growth factor- , platelet - derived growth factor- and glucose transporter glut1 and therefore influences such cellular activities as angiogenesis , glycolysis and cell survival . also , hif-1 activates pyruvate dehydrogenase kinases ( pdks ) , thereby inactivating pyruvate dehydrogenase and blocking the flow of pyruvate into the tca cycle . reduced transport of pyruvate into the mitochondria decreases the rate of oxidative phosphorylation and oxygen consumption . the oncogenic transcription factor , myc , also directly activates transcription of glycolytic enzymes that affect cell metabolism . myc was shown to regulate glycolysis in cells grown under normoxic conditions through the direct activation of lactate dehydrogenase a and virtually all other glycolytic genes . myc can also activate genes associated with mitochondrial biogenesis and function and cooperates with hif to activate certain glucose transporters and glycolytic enzymes , as well as lactate dehydrogenase a and pdk1 . in contrast to akt and hif , myc can regulate genes associated with glutamine metabolism . collectively , these alterations induce high levels of glycolysis and glutaminolysis in myc - induced liver cancer , which are associated with an aggressive tumor phenotype and histology . it is important to note that even though myc can enhance the glycolytic pathway , it also enhances mitochondrial respiration , all of which contribute to the overall increased metabolic function of cancer cells . the tumor - suppressor p53 is probably one of the single most important proteins mutated in many human cancers . in addition to its role in cell cycle control and cell death , p53 also shows an inhibitory effect on glycolysis . p53 upregulates the expression of tp53-induced glycolysis and apoptosis regulator ( tigar ) , which decreases fructose-2,6-bisphosphatase ( fru-2,6-p2 ) by dephosphorylation . increased expression of tigar results in a decreased level of fru-2,6-p2 and a decreased glycolytic rate . furthermore , p53 directly stimulates oxidative phosphorylation through upregulation of cytochrome c oxidase 2 ( sco2 ) , which is required for the assembly of the cytochrome c oxidase complex of the electron transport chain . although the warburg effect describes altered cancer metabolism , mutations in genes that encode metabolic enzymes can provide a direct genetic link to the altered metabolism ( figure 2 ) . mutations in two tca cycle - related genes that encode fumarate hydratase ( fh ) and succinate dehydrogenase ( sdh ) lead to accumulation of their respective substrates , fumarate and succinate , in a subset of human cancers . sdh also serves as respiratory complex ii in the electron transport chain ; therefore mutations in sdh may directly affect mitochondrial respiration . indeed , mutations in the genes encoding sdh subunits have been connected to hereditary parangangliomas and pheochromocytomas . additionally , fh gene encodes the enzyme that converts fumarate to malate in the mitochondria . thus , mutations in fh result in fumarate hydratase deficiency and have also been connected to uterine and skin leiomyomas and papillary renal cancer . connections between a mutant metabolic enzyme and tumorigenesis have been uncovered from cancer genome - sequencing efforts . nadp ( nicotinamide adenine dinucleotide phosphate)-dependent isocitrate dehydrogenase 1 ( idh1 ) and idh2 provide a fascinating example of a mutant metabolic enzyme driving tumorigenesis . idh1 and idh2 catalyze the conversion of isocitrate to -ketoglutarate ( kg ) in mammalian cells with the generation of one molecule of nadph . idh1 and idh2 are homodimeric enzymes that act in the cytoplasm and mitochondria , respectively . it has been shown that heterozygous point mutations in several residues in idh1 are common in gliomas and in acute myeloid leukemia . these mutations have been shown to act in a dominant - negative fashion to inhibit idh1 and idh2 activity , leading to a reduction in cytoplasmic kg concentration , an inhibition of prolyl hydroxylase activity and a stabilization of the hypoxia - induced transcription factor hif-1 . additionally , these mutations have been reported to provide idh1 and idh2 with novel enzymatic activity , causing conversion of kg to 2-hydroxyglutarate ( 2-hg ) , the effects of which are not well understood . although 2-hg is present at low concentrations in normal cells and tissues , mutations in idh1 and idh2 result in elevated 2-hg levels in glioma tissues and in acute myeloid leukemia cells . hence , it would be interesting to know if high concentrations of 2-hg are directly related to the tumorigenicity of idh1 and idh2 mutations . importantly , levels of kg , isocitrate and several other tca metabolites remain unchanged in cells or tissues with idh1 mutations , suggesting that other metabolic pathways can adjust and maintain normal levels of essential metabolites . a common feature associated with mutations in the metabolic enzymes sdh , fh and idh is an aberrant accumulation of metabolites with oncogenic potential . the metabolic products of mutant sdh , fh and idh inhibit the activity of enzymes ( for example , prolyl hydroxylases ) belonging to a class of kg - dependent enzymes . furthermore , mutant sdh , fh and idh have all been shown to induce hif activity . thus , oncogenic alterations in cellular metabolism can regulate pathways that are unrelated to metabolism and that contribute to the oncogenic process , such as altered hif activity . phosphoglycerate dehydrogenase ( phgdh ) and glycine decarboxylase are two enzymes showing a similar pattern of mutation that can both affect mitochondrial metabolism directly during oncogenesis . phgdh catalyzes the first step in serine synthesis , is amplified in estrogen receptor - negative breast cancers , and is suggested to be an oncogenic enzyme when overexpressed . phgdh also shuttles glycolytic intermediates into the one - carbon metabolic pathway and thus directs nucleotide biosynthesis . loss of phgdh decreases the level of a key tca intermediate , kg , but surprisingly not serine . glycine decarboxylase is also involved in glycine / serine metabolism and the one - carbon metabolic pathway , and its overexpression promotes tumorigenesis . metabolic targeting for cancer therapy is currently under investigation in an effort to identify small molecules that might specifically inhibit key metabolic steps associated with tumor growth ( table 1 ) . attenuation or inhibition of glycolysis has been found useful for preventing the development of cancer , demonstrating that glycolysis is essential for proliferation , invasion and metastasis of cancer . glycolysis can be blocked by inhibition of the glycolytic enzymes hk , pfk and pyruvate kinase ( pk ) , all of which regulate irreversible and rate - limiting steps in glycolysis . therefore , the strategy is to compromise or completely block the increased glycolysis in cancer cells by abating the enzymatic activity of these three proteins . hk facilitates the first step of glycolysis , where glucose is phosphorylated with the use of atp . inhibitors of hk such as 2-deoxyglucose ( 2-dg ) , 3-bromopyruvate ( 3-brpa ) and lonidamine ( lon , 1-[(2,4-dichlorophenyl)methyl]-1h - indazole-3-carboxylic acid ) are currently in pre - clinical and early phase clinical trials . 2-dg serves as a competitive inhibitor of hk , blocking access of glucose to the enzyme . 2-dg is taken up by glucose transporters and phosphorylated by hk to 2-dg - p and thereby trapped within the cell , because 2-dg - p is not recognized and metabolized by the next glycolytic enzyme , phosphoglucose isomerase . this leads to the accumulation of 2-dg - p within the cell and a depletion of cellular atp production . however , its effectiveness as a single agent is limited and combining 2-dg with radiotherapy or chemotherapy enhances the tumor - destroying effects . 3-brpa is known to inhibit cancer cell energy metabolism and is a key determinant of chemoresistance in certain cancer types . atp depletion caused by glycolysis inhibition with 3-brpa treatment suppresses atp - binding cassette ( abc ) transporter activity and subsequently drug efflux , enhancing drug retention . as a result consistently , it was previously reported that increased glycolysis is directly associated with glucocorticoid resistance that is associated with treatment failure in childhood acute lymphoblastic leukemia and inhibition of glycolysis by 2-dg , 3-brpa or lon increases prednisolone - induced toxicity in leukemia cells . lon , 1-[(2,4-dichlorophenyl)methyl]-1h - indazole-3-carboxylic acid , is a hk 2 inhibitor that has been shown to induce apoptosis and treat multidrug resistance in various cancer cell lines . however , phase ii clinical trials of lon as a treatment for benign prostatic hyperplasia have been suspended because of its liver toxicity . the combination of lon with other anticancer drugs like paclitaxel , a common chemotherapeutic agent that hyper - stabilizes microtubules and prevents cell division but shows high toxicity , is undergoing clinical trials for the treatment of cancers . pfk activity is extremely sensitive to small changes in ph , and its activity drops as ph decreases . the intracellular ph in cancer cells is typically high , activates the rate - limiting enzyme pfk and subsequently facilitates glycolysis . the na / h - exchanger nhe-1 is known to increase the intracellular ph in cancer cells . it has been reported that inhibitors of nhe-1 such as amiloride or 5,5-dimethylamiloride produce anticancer effects by decreasing intracellular ph , pfk activity and thus decreasing glycolysis . citrate is an archetypal feedback inhibitor of pfk and is normally converted into cytosolic acetyl - coa by the enzyme atp citrate lyase . however , use of an atp citrate lyase inhibitor , sb-204990 , can increase citrate levels that , in turn , inhibit glycolysis , thereby producing anticancer effects . the pk isozyme m2 ( pkm2 ) is present in very few types of normal proliferating cells but is highly expressed in cancer cells . pkm2 catalyzes the rate - limiting atp - generating step of glycolysis , controlling the conversion of phosphoenolpyruvate to pyruvate . by inhibiting pkm2 , the glycolytic flux regresses to limit the generation of atp in cancer cells . although such an effect might seem to be detrimental to tumor growth , the result is actually the opposite . similar to normal cells , cancer cells must generate macromolecules such as lipids , nucleic acids and proteins . without the biosynthesis of these macromolecules , rapid cell proliferation will cease even in the presence of atp . slowing glycolysis and shuttling of phosphoenolpyruvate through the pentose phosphate pathway generates macromolecule precursors that are necessary to support cell proliferation and hence counteract the effects of inhibited glycolysis . despite this setback , pkm2 is still of interest , because it is uniquely important to tumor metabolism . until recently , the importance of glutamine metabolism and its regulation has not been emphasized for tumor metabolism , because the dependency of cancer cells on glutamine was not fully appreciated by warburg . modulation of transamination , the transfer of an amine from glutamine during mitochondrial respiration , has been proposed to inhibit tumor growth . it consists of glutaminase that converts glutamine to glutamate and glutamate dehydrogenase that converts glutamate to kg . myc has been shown to regulate many steps in glutamine metabolism , which gives an insight on important enzymes in glutamine metabolism as therapeutic targets when metabolism is reprogrammed in cancer cells . it has been suggested that modulation of both transamination and pk can be a combined method for metabolic targeting of tumors . metabolites from transamination may also have a regulatory role on the dimer / tetramer structure of pk . a balanced presence of both pk structures is essential for enzymatic activity , and modulation of the dimer / tetramer ratio by transamination metabolites can be detrimental for tumor growth . this reduction step is absolutely necessary for the regeneration of nad , which is needed to continue glycolysis . it has been observed that decreases in ldh inhibits glycolysis and has antitumor effects in cancer cells . have synthesized dihydroxynaphthoic acids , which are potent inhibitors of ldh and inhibitors of glycolysis . the specific ldh inhibitor , oxamate , may also be useful in treating paclitaxel - resistant cancers . combinational treatment of paclitaxel and oxamate showed a synergistic inhibitory effect in the paclitaxel - resistant breast cancer cells by promoting apoptosis . lastly , inhibition of pdk reduces glycolysis and promotes mitochondrial function by activating pyruvate dehydrogenase , and reduction of glycolysis may lead to an antitumor response . many studies have shown that dichloroacetate , a pdk inhibitor , produced markedly positive outcomes for the treatment of lactic acidosis and mitochondrial diseases and may prove useful in the treatment of cancer . cancer cell metabolism utilizes aerobic glycolysis in which cancer cells use glucose for energy supply and glutamine to feed mitochondrial intermediates for biosynthetic precursor supply . altered metabolism is considered to be fundamental to the transformation of normal cells to cancer cells , and it is believed to be conserved in most tumors , including solid tumors , lymphoma and leukemia . certainly , cancers display extremely heterogeneous characteristics as tissue origin in every cancer type is different . also , cancer cells suffer from poor nutrient and oxygen supplies due to the imperfect vasculature formed during tumorigenesis . therefore , alterations of metabolic pathways in tumor cells may give rise to a selective advantage for rapid generation of atp and for sufficient biomolecule production to cancer cells under the unfavorable environment . hypoxic adaptation essential for survival and progression of a tumor is thought to be closely linked to metabolic changes in cancer cells . along with a reduced use of oxygen and the rapid energy production , there is a notable shift in mitochondrial function from an energy producer to a creator of biosynthetic intermediates . although the molecular mechanisms still remain largely unknown , mutations of oncogenes and tumor suppressors account for part of the metabolic reprogramming in cancer cells . cancer cell metabolism has recently become one of the most exciting and promising fields for the development of new anticancer agents . cancer research encompasses distinct fields of molecular , biological and metabolic research areas , and until recently , only signal transduction pathways have been recognized as a promising field for the development of therapeutic drugs against tumors . by better understanding the cancer - specific metabolic process , and by further comprehending the tumor cell , researchers in the field of metabolic research hope to find new drugs that may revolutionize cancer treatment . targeting cancer metabolism opens an opportunity to develop broadly applicable drugs that can treat multiple cancer types and hence may lead to a new class of anticancer drugs . numerous analogs of metabolites are being tested at present as potential drug candidates to target tumor metabolism . further studies on the role of the mitochondria in cancer cell metabolism may provide a selective delivery system for toxic chemicals specifically to cancer cells , thereby increasing the efficacy and reducing the toxicity of potentially powerful chemotherapeutic drugs . new understandings of cancer metabolic profiles give us a hope that a new class of therapeutic agents may be developed for cancer therapy . | cancer cell metabolism is characterized by an enhanced uptake and utilization of glucose , a phenomenon known as the warburg effect .
the persistent activation of aerobic glycolysis in cancer cells can be linked to activation of oncogenes or loss of tumor suppressors , thereby fundamentally advancing cancer progression . in this respect ,
inhibition of glycolytic capacity may contribute to an anticancer effect on malignant cells .
understanding the mechanisms of aerobic glycolysis may present a new basis for cancer treatment .
accordingly , interrupting lactate fermentation and/or other cancer - promoting metabolic sites may provide a promising strategy to halt tumor development . in this review
, we will discuss dysregulated and reprogrammed cancer metabolism followed by clinical relevance of the metabolic enzymes , such as hexokinase , phosphofructokinase , pyruvate kinase m2 , lactate dehydrogenase , pyruvate dehydrogenase kinase and glutaminase .
the proper intervention of these metabolic sites may provide a therapeutic advantage that can help overcome resistance to chemotherapy or radiotherapy . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
hepatitis e virus ( hev ) is a nonenveloped virus with a nonsegmented positive - sense rna genome containing three open reading frames ( orfs ) [ 1 , 2 ] . the mammalian hev , which infects humans and several mammalians , is classified into four major genotypes , namely , genotypes 1 , 2 , 3 , and 4 . hev causes large outbreaks of acute hepatitis in developing countries where sanitary conditions are inadequate and the infection is enterically transmitted by the fecal - oral route especially contaminated water [ 6 , 7 ] . the zoonotic spread of the disease is also possible especially in developed countries where the sporadic cases of the infection have been reported [ 2 , 4 ] . although the infection is usually asymptomatic or acute self - limiting in general population , but it can progress to lethal fulminant hepatitis in pregnant women and chronic hepatitis in immunosuppressed and organ transplant patients [ 5 , 8 ] . the general mortality rate of hev infection is between 1% and 15% , and this level may reach up to 30% due to fulminate hepatic failure during pregnancy , which may lead to high rate of spontaneous abortion and premature birth [ 5 , 9 ] . consequently , hev has become a serious public - health concern especially in endemic areas , and control and prevention of this infection are necessary . so far , no effective treatment or commercial vaccines for hev infection are available , and the only way to reduce incidence of the infection is prevention [ 2 , 10 ] . hev capsid protein has several immune - dominant conserved epitopes that can induce long - lived immunity ; therefore , it has been studied for vaccine development . however , the full - length capsid protein is not suitable for vaccine production . since it is hydrophobic and insoluble and its immunoreactive epitopes are masked , much attention has been drawn to the truncated or short forms of orf2 protein as protective vaccines [ 12 , 13 ] . the truncated orf2 protein , containing amino acids 112 to 660 of the orf2 protein , self assembles into virus - like particles and can induce strong immunity . moreover , 52 amino acids from the c terminus and 111 amino acids from the n terminus of the orf2 protein do not have any antigenic domain , and most antigenic domains have located within 268 amino acids of c terminal of the orf2 protein . also , the truncated orf2 protein containing amino acids 459 to 607 is needed to produce anti - hev antibodies ; therefore , it seems that antigenic epitopes are conformational . anti - hev antibodies from all genotypes cross - react with the orf2 protein of genotype 1 ; therefore , production of a broadly protective vaccine seems to be possible by expression the truncated orf2 protein of genotype 1 . in the present study , the cellular immune responses to the truncated orf2 protein were evaluated in the peripheral blood mononuclear cells ( pbmcs ) of individuals recovered from hepatitis e infection . this study ( with research project number d/574 ) was approved by the jundishapur university ethical committee , and informed consent was taken from all the participants . forty anti - hev igg positive subjects ( 24 , 60% males and 16 , 40% females ; mean age sd , 35.12 9.66 years ) and forty - eight anti - hev igg negative subjects ( 28 , 58.3% males and 20 , 41.7% females ; mean age sd , 33.06 10.70 years ) were randomly selected as hev - recovered and control groups to study the immunogenicity of the truncated orf2 protein . the presence of anti - hev igg antibody was detected by the commercial elisa kit ( hev igg elisa kit , dia.pro , milan , italy ) . all the participants were negative for anti - hev igm antibody , anti - hav igm antibody , anti - hcv antibodies , and hepatitis b surface antigen ( hev igm elisa kit , hav igm elisa kit , hcv ab elisa kit , hbs ag elisa kit , dia.pro , milan , italy ) and had normal alt level . ficoll - hypaque ( lymphoflot , biotest diagnostics , germany ) and density gradient centrifugation method were used to separate peripheral blood mononuclear cells ( pbmcs ) from 10 ml heparinized venous blood sample of the each individual . then the cells were washed twice by rpmi 1640 medium and resuspended in rpmi 1640 plus 10% fcs ( invitrogen , carlsbad , ca , usa ) . trypan blue staining was used to evaluate the cells viability . a truncated form of hev orf2 protein containing amino acids 112608 was produced as described previously . briefly , a truncated orf2 gene encoding amino acids 112660 of sar-55 strain from hev genotype 1 was optimized for expression in e. coli and synthesized and cloned into pbluescript ii sk(+ ) vector by biomatik company ( biomatik corporation , cambridge , canada ) . the truncated gene was subcloned into expression vector pet-30a(+ ) and the recombinant plasmid pet30a - orf2(aa 112660 ) was constructed . then , a 193-nucleotide fragment was removed from the nucleotide sequence of the truncated orf2 gene by digestion with nhei restriction enzyme and another truncated form of orf2 gene encoding amino acids 112608 was constructed . the recombinant plasmid pet-30a - orf2(aa 112608 ) was transformed into e. coli bl21 ( de3 ) using electroporation . expression of the truncated orf2 protein containing amino acids 112608 was induced by adding 1 mm iptg to the bacterial culture followed by 4 hr incubation at 37c . the expressed truncated orf2 protein was purified by ni2 + -chelate - affinity chromatography ( qiagen , hilden , germany ) . refolding of the purified protein was done by dialysis in pbs + 10% glycerol at 4c for 4 hr . amicon ultra-4 centrifugal filter unit ( emd millipore , billerica , ma , usa ) was used to improve the concentration and purity of the target protein . to remove endotoxin contamination that may be obtained during protein purification , endotoxin was extracted from the protein by toxin eraser endotoxin removal kit ( genscript , piscataway , nj , usa ) . the cell proliferation was carried out by the colorimetric assay using mtt ( 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide ) as described previously . for each sample , approximately 1 10 pbmcs / well in rpmi 1640 plus 10% fcs were added to 3 wells of 96-well plates in final volume of 180 l / well . the first well was stimulated with 10 g / ml of truncated orf2 protein in final volume 20 l . 20 l / well of phytohemagglutinin ( pha ) ( 5 g / ml ) ( sigma aldrich , st . louis , mo , usa ) was used as positive control in the second well and the third well remained unstimulated as negative control . blanking was done by putting rpmi 1640 ( 20 l ) alone in the well . the optimal concentrations of truncated orf2 protein and pha and the optimum stimulation duration were determined based on preliminary experiments . the plate was incubated in the presence of 5% co2 at 37c for 4 days , and then 10 l of mtt solution ( 5 mg / ml ) was added to each well . after incubation at 37c for 4 hr and centrifugation in 1000 g for 10 min , the supernatant was discarded and 100 l of dmso was added to each well and mixed by pipetting . the absorbance of the wells was measured using an elisa reader at 570 nm within an hour . the ifn elispot assay was performed by human ifn- elispot ready - set - go kit ( ebioscience , san diego , ca , usa ) to determine hev - specific ifn--producing cells . according to manufacturer 's structure , a pvdf bottomed elispot plate ( millipore , bedford , ma , usa ) was coated with capture antibody solution ( 100 l / well ) overnight at 4c . for each sample , approximately 1 10 pbmcs / well in rpmi 1640 plus 10% fcs were added to 3 wells of the 96-well elispot plate in final volume of 150 l / well . five g / ml of pha was used as positive control in the second well , and the third well remained unstimulated as negative control . the optimal count of pbmc in each well , the optimal concentrations of truncated orf2 protein and pha , and the optimum duration of stimulation were determined based on preliminary experiments . , the cells and medium were removed and the wells were washed 3 times with elispot wash buffer . the plate was incubated with biotinylated detection antibody ( 100 l / each well ) at 4c overnight and washed 4 times with elispot wash buffer . in the next step , the plate was incubated with 100 l / well of avidin - horseradish peroxidase reagent at room temperature for 45 min and washed with elispot wash buffer and 1x pbs . finally , the dark purple spots were developed by adding 100 l / well aec ( 3-amino-9-ethylcarbazole ) , followed by incubation at room temperature . the plate was washed 3 times with 200 l / well distilled water and dried at room temperature , and then a dissection stereoscope was used to count the number of spots . the results were shown as spot forming cells ( sfcs ) per 10 cells , while each spot was considered as one hev - specific ifn- secreting cell . pbmcs of each sample were stimulated with the truncated orf2 protein ( 10 g / ml ) at 37c . the optimal concentrations of truncated orf2 protein and pha and the optimum stimulation duration were determined based on preliminary experiments . culture supernatants were harvested after 1 - 2 days and used to measure amounts of four cytokines ( il-4 , il-12 p70 , ifn- , and il-10 ) . cytokine assay was performed by commercially available elisa kits ( ebioscience , san diego , ca , usa ) according to the manufacturer 's instructions . the limits of sensitivity of the assays were 1.3 pg / ml for il-4 , 1.0 pg / ml for il-10 , 2.1 pg / ml for il-12 p70 , and 0.99 pg / ml for ifn-. spss 17 package program ( spss inc . , chicago , usa ) was used for statistical analysis , and p values below 0.05 indicated statistical significance . the data were analyzed and compared by the mann - whitney u test or independent t - test . proliferation of pbmcs in response to the truncated orf2 protein was measured in the control and hev - recovered groups . the cell proliferative responses to truncated orf2 protein in the control and the corresponding figures following stimulation with pha were 2.91 0.37 and 2.81 0.33 in the control and recovered groups , respectively ( p = ns ) . overall , the recovered group showed significantly higher cell proliferation compared to the control group ( p < 0.001 ) ( table 1 , figure 1 ) . the frequencies of ifn--secreting cells in pbmcs of hev - recovered and control groups were assessed in response to truncated orf2 protein by elispot assay . ifn- elispot responses following stimulation with the truncated orf2 protein in the control and hev - recovered groups were 4.39 5.51 and 65.92 58.99 , respectively . elispot responses to pha in the control and recovered groups were 223.41 94.58 and 228.25 123.41 , respectively ( p = ns ) . overall , ifn- elispot responses were significantly higher in the recovered group compared to the control group ( p < 0.001 ) ( table 1 , figure 2 ) . significant amounts of ifn- and il-12 cytokines but low amounts of il-10 and il-4 cytokines were produced following stimulation with the truncated orf2 protein . comparison between the two groups revealed no significant differences in the levels of il-10 ( p = 0.100 ) and il-4 ( p = 0.067 ) , while ifn- and il-12 p70 were significantly higher in the recovered group compared to the control group ( p < 0.001 ) . stimulation with pha resulted in almost similar expression of all cytokines in the control and recovered groups , including 11.96 4.55 and 11.29 3.9 for il-4 ( p = ns ) , 199.45 108.43 and 204.97 103.61 for il-10 ( p = ns ) , 109.91 32.86 and 103.22 25.9 for il-12 ( p = ns ) , and 410.37 46.96 and 418.37 75.65 for ifn- ( p = ns ) , respectively ( table 1 , figure 3 ) . hev causes self - limited acute hepatic infection worldwide and especially in developing countries where the disease is endemic and 30% to 80% of the adult population has anti - hev antibody . the infection is more severe in pregnant women and patients with underlying liver problems and may progress to lethal fulminant hepatitis [ 5 , 8 ] . information about hev specific cellular immune responses could be useful in finding an effective way for development of hev vaccine , in particular as any protective vaccine for hev infection is not still licensed [ 2 , 9 ] . however , some studies have evaluated hev - specific cellular immune responses among different groups [ 6 , 1820 ] , but current information on cellular immune responses against hev is not sufficient to prevent hev infection . therefore , this study aimed to evaluate hev - specific cellular immune responses to the truncated orf2 protein in the recovered individuals from hev infection . in the present study , a significant increase in the cell proliferation and ifn- elispot responses following stimulation with the truncated orf2 protein were found in the hev - recovered subjects , while both groups showed good nonspecific stimulation with pha . therefore , the increase in responses induced by truncated orf2 protein was specific in the recovered group . reported a similar increase in the hev - specific t - cell responses by pbmcs obtained from recovered individuals from hev infection . the stimulating antigen in their study was overlapping peptide pools and not the whole hev orf2 protein . regarding the full length orf2 protein , husain et al . it has been shown that orf2 protein has several htl ( helper t - lymphocyte ) dominant immunogenic regions and therefore it is a good antigen to elicit high and long - lasting specific anti - hev immune responses . the hev - specific t - helper cell responses , especially against capsid protein , play a critical role in clearing and controlling viral infections , either directly through cytokines production or indirectly by activating cytotoxic t cells . production of t - helper type 1 ( th1 ) cytokines such as ifn- , tnf-/ , il-2 , il-3 , and il-12 have been implicated in viral clearance or resistance to infection , while production of th2 cytokines such as il-4 , il-5 , il-13 , il-6 , and il-10 cytokines is associated with progression of the infection or viral persistence [ 2328 ] . in the present study , the truncated orf2 protein could significantly stimulate pbmcs to produce ifn- and il-12 but little or no il-4 . the results of this study being consistent with those of others suggest that viral proteins that can shift immune responses to th1 , rather than th2 , may have a better chance of success as a vaccine candidate against hev infection . all participants in control and recovered groups showed almost similar expression of all cytokines following stimulation with pha . therefore , the increase in production of ifn- and il-12 cytokines following stimulation with truncated orf2 protein was specific in the recovered group . the results of the previous studies have shown that the production of th1 cytokines such as il-12 and ifn- during hev infection results in self - limiting hepatitis e , suggesting the role of these cytokines in the recovery and clearance of hepatitis e , while the th2 cytokines increase in fatal fulminant hepatitis e , especially during pregnancy [ 18 , 27 , 30 , 31 ] . in saravanabalaji study , levels of ifn- and il-12 cytokines following stimulation of pbmcs by recombinant orf2 protein were higher in recovered patients than those with fatal fulminant hepatitis e , while il-10 was higher in fatal patients . in pal et al . study , high levels of il-4 and il-10 were observed in pregnant women with acute hepatitis e following stimulation of pbmcs with different peptides of hev orfs , while the level of ifn- was low in this group . in srivastava study , the ifn- production following stimulation with orf2 protein was higher in patients with acute hev infection compared to control . in this study , antigenic properties of the expressed orf2 protein in e. coli bl21 cell were evaluated . among the various microorganisms and systems available for protein production , e. coli bl21 containing pet - derived plasmids has become the most commonly used expression system because of its high expression capabilities , protein production with high purity , low cost , simple and fast cultivation , its well - known genetics , and therefore ease of genetic manipulation . however , there are a variety of factors that affect the expression level as well as the solubility and especially biological function of overexpressed proteins . one of the ways to improve the expression of foreign protein in e. coli is codon optimization , which results in selection of host favorite codons and elimination of high gc contents [ 33 , 34 ] . by evaluating the antigenic properties and immunogenicity of the truncated orf2 protein , it was indicated that this protein has biological function and can stimulate specific anti - hev immune responses . in conclusion , the truncated orf2 protein is able to induce significant cellular immune responses and production of th1 cytokines in vitro and therefore can be introduced as a potential vaccine candidate for hev infection . | background .
the aim of this study was to evaluate hepatitis e virus ( hev ) specific cellular immune responses to truncated orf2 protein in iranian patients recovered from hev infection .
information about hev - specific immune responses could be useful in finding an effective way for development of hev vaccine . methods .
a truncated form of hev orf2 protein containing amino acids 112 - 608 was used to stimulate peripheral blood mononuclear cells ( pbmcs ) separated from hev - recovered and control groups .
finally , the levels of four cytokines , ifn- elispot , and cell proliferative responses following stimulation with the truncated orf2 protein were assessed in the both groups .
results .
the truncated orf2 protein was able to induce ifn- elispot and cell proliferation responses and to produce significant amounts of ifn- and il-12 cytokines , but low amounts of il-10 and il-4 cytokines in vitro .
these responses were significantly higher in the recovered group compared to the control group .
these results indicate the antigenic nature of the truncated orf2 protein and production of t helper type 1 cytokines . conclusion
. the truncated orf2 protein can effectively induce significant cellular immune responsesand can be introduced as a potential vaccine candidate .
however , further studies are required to evaluate this protein in vivo . |
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propylene glycol ( pg ) is a colorless , odorless organic chemical approved by the us food and drug administration for use as a solvent in food , drugs , and cosmetics . it is included in numerous intravenous and oral medications , some of them being broadly used in critically ill patients , like lorazepam or diazepam , barbiturates , etomidate , phenytoin , or trimethoprim / sulfamethoxazole ( tmp / smx ) . although pg is generally considered safe , there is an increasing body of evidence suggesting that large doses or prolonged treatments with intravenous medications containing pg can result in severe toxicity . the reported toxic effects of pg include hyperosmolality and anion gap metabolic acidosis , lactic acidosis , acute kidney injury , cns toxicity with seizures , and cardio - respiratory arrest . the precise mechanism of pg toxicity is presently unknown , however , the accumulation of pg and/or its byproducts may result in hyperosmolality and anion gap metabolic acidosis . furthermore , the findings of swelling and vacuolization of proximal tubular cells in a pg - exposed patient kidney biopsy as well as observed toxicity on cultured human proximal tubular cells suggest a mechanism by which pg - related cellular injury results in acute kidney injury . although the world health organization recommends a maximum consumption of 25 mg / kg / day of pg when used as a food additive , this does not address its use as a drug solvent . among the pg - containing drugs used in icu settings , intravenous lorazepam involves the largest amounts of pg with respect to the administered dosage ; as expected , lorazepam has been associated with most cases of pg toxicity which always involved large doses or prolonged treatments with large amounts of daily or total pg intakes . some authors have suggested risks factors for pg toxicity , including chronic renal or liver disease , alcohol abuse , and have recommended to limit daily intravenous pg intakes to 1 g / kg / d or 69 g / day . although tmp / smx contains pg as well , doses usually administered to treat pneumocystis jirovecii pneumonia ( 1520 mg / kg / d ) involve < 0.5 g / kg / d of pg , well below the reported levels of toxicity . even though 2 case reports of pg toxicity involved tmp / smx as a co - administered drug , both patients were receiving large doses of lorazepam which accounted for most of the pg intakes ( respectively 552 g over 12 days and 540 g over 5 days ) . we present a unique case of severe lactic acidosis after only a 3-day course of intravenous tmp / smx , involving amounts of pg well below the reported toxic levels , in a patient without known risk factor for pg toxicity . a 31-year - old caucasian female ( weight 106 kg ) with a past medical history of hypothyroidism , neurofibromatosis type 1 , and pilocytic astrocytoma ( figure 1 ) recently treated with metronomic temozolomide and steroids , presented with dyspnea at rest , a 2 week duration of nonproductive cough , and a 6 week history of progressive weakness , ataxia , anorexia , weight loss , and throat pain . her vital signs on admission to the emergency department were as follows : temperature 36.5c , heart rate 110/minute , respiratory rate 30/minute , blood pressure 114/83 mm hg , spo2 79% on room air , and then 88% on oxygen 6 l / min nasal cannula . initial assessment , including a chest radiograph and lung computed tomography ( figure 2 ) , revealed patchy bilateral parenchymal opacities with consolidation concerning for multifocal pneumonia , in addition to a small left pleural effusion . laboratory test revealed a white blood cell count of 8.75 10/mm ( normal range 4.010.4 10/mm ) with 92% neutrophils , 5% lymphocytes , 3% monocytes , 0.3% eosinophils , and no basophils or bands . mmol / l ) , potassium 2.8 mmol / l ( 3.55.1 mmol / l ) , chloride 100 mmol / l ( 96107 mmol / l ) , bicarbonate 31 mmol / l ( 2230 mmol / l ) , blood glucose 96 mg / dl ( 74106 mg / dl ) , and baseline blood urea nitrogen ( bun ) and serum creatinine were 7 mg / dl ( 720 mg / dl ) and 1.02 mg / dl ( 0.61.0 mg / dl ) , respectively . the patient 's initial acid base status was normal , with a venous ph of 7.4 . blood , urine , and sputum cultures were obtained , and the patient was treated empirically with intravenous cefepime , vancomycin , and azithromycin . given worsening hypoxia requiring oxygen via high flow nasal cannula ( fio2 65% ) , the patient was admitted to the intensive care unit ( icu ) and intubated shortly after for acute hypoxic respiratory failure requiring mechanical ventilation . brain magnetic resonance imaging with intravenous contrast showing the pilocytic astrocytoma as an enhancing cystic lesion ( 2.5 2.9 cm ) in the region of the hypothalamus and optic chiasm extending into the brain stem . chest computed tomography showing patchy bilateral parenchymal opacities with trace left pleural effusion , consistent with multifocal pneumonia . in the icu , 18 mg / kg / d of intravenous tpm / smx , corresponding to 0.47 g / kg / d of pg , was added for empiric coverage of pneumocystis jirovecii pneumonia , given the patient 's immunocompromised state . after intubation and mechanical ventilation , the patient 's hypoxia quickly improved and the fio2 was weaned down to 40% within 24 hours with spo2 99% . the patient was successfully extubated after 36 hours of mechanical ventilation . her vital signs after extubation were unremarkable with normal heart rate and blood pressure , respiratory rate at 18/minute , and spo2 97% on oxygen 2 twenty - four hours after extubation , the patient developed progressively worsening tachypnea ( respiratory rate 32/minute ) without worsening hypoxemia ( spo2 97% on oxygen 2 l with nasal cannula ) or change in her chest x - ray . this prompted an arterial blood gas analysis which revealed a severe metabolic acidosis with ph 7.2 , pco2 19 mm hg , po2 107 mm hg on 2 l oxygen , bicarbonate 8 mmol / l , an anion gap of 25 meq / l , and a lactate of 12.1 blood , urine , and sputum cultures remained negative and the patient 's vital signs ( temperature , heart rate , and blood pressure ) remained normal . as tmp / smx was suspected as the cause of her lactic acidosis and the p jirovecii , direct fluorescent - antibody test returned negative , tmp / smx was discontinued . the other medications received between admission and the occurrence of lactic acidosis included cefepime , vancomycin , azithromycin , enoxaparin , fentanyl , propofol , dexmedetomidine , hydrocortisone , levothyroxine , pantoprazole , and lorazepam ( 1 mg twice ) , among them only lorazepam included pg ( 0.8 g for 2 mg of lorazepam ) . within 24 hours of stopping the tmp / smx and without any additional intervention , the lactate decreased from 12.1 to 2.9 mmol / l and serum bicarbonate increased to 23 mmol / l . figures 3 and 4 display the course of relevant vital signs ( mean arterial pressure and spo2 ) and biological data ( serum bicarbonates , creatinine , and lactic acid ) during the initial 6-day course . in total , the patient received 54 g ( 0.5 g / kg ) of pg daily for 3 days while being treated with tmp / smx and an additional 0.8 g with lorazepam . no other cause for the patient 's lactic acidosis was identified , and her acid base status remained normal for the rest of her hospital admission . course of mean arterial pressure ( map , diamond markers with straight line on the left y - axis ) and spo2 ( x markers with dotted line on the right y - axis ) over the first 6 days of admission . tmp / smx was started on admission ( day 0 ) and stopped at day 4 . scales are different for the sake of legibility . except for initial hypoxemia ( spo2 79% on room air on admission ) and a brief ( 1 hour ) episode of desaturation at day 2 ( spo2 8687% on o2 2 liters ) , map and spo2 remained above 65 mmhg and 90% respectively . course of serum bicarbonates ( diamonds ) and lactates ( circles ) on the left y - axis and creatinine ( x markers ) on the right y - axis over the first 6 days of admission . tmp / smx was started on admission ( day 0 ) and stopped at day 4 . scales are different for the sake of legibility . while serum bicarbonates was normal on admission , a severe metabolic acidosis developed at day 3 , concomitant with an increased lactates . after tmp / smx cessation , serum bicarbonates and lactates levels normalized within 24 hours . the subsequent patient 's hospital course was complicated by persistent encephalopathy ascribed to her brainstem astrocytoma , with recurrent airway mucus plugging requiring re - intubation 10 days after extubation . the patient eventually underwent a tracheostomy for difficult ventilator weaning and was transferred to a rehabilitation facility after 3 weeks . there has been an increasing interest in pg toxicity in recent years , and multiple adverse reactions have been reported , including hyperosmolality , anion gap metabolic acidosis , lactic acidosis , acute kidney injury , cns toxicity with seizures , and a sepsis or systemic inflammatory - like syndrome . although drugs with high pg content , such as benzodiazepines ( lorazepam contains as high as 80% v / v of pg ) , barbiturates , etomidate , phenytoin , and tmp / smx , are broadly used in the icu setting , their sometimes life - threatening adverse reactions may be under - recognized . in a prospective observational study , arroliga and coworkers demonstrated that 6 out of 9 icu patients being treated with high dose ( 10 mg / h ) of intravenous lorazepam developed hyperosmolar high anion gap metabolic acidosis consistent with pg accumulation , the osmolar gap being the strongest predictor of serum pg concentrations . wilson and colleagues also observed that 4 out of 21 patients who had received pg - containing sedative medications ( either lorazepam or diazepam ) had metabolic abnormalities consistent with pg toxicity . these studies suggest that the incidence of pg toxicity is likely much higher than clinically recognized and underscore the need for a better knowledge by clinicians and icu staff of pg - containing drugs and for a prudent monitoring of patients receiving pg . although pg is generally considered safe , the reported cases of mild forms of lactic acidosis following the use of large doses of pg diluted medications led to recommendations to limit pg intakes to 69 g / d or 1 g / kg / d , a dose presumed safe in the absence of risk factors for pg toxicity . the precise mechanism by which pg causes toxicity is unclear , as no convincing evidence supports a direct cytotoxicity of either pg or its byproducts . after intravenous administration , pg has a elimination half - life of 2.3 0.7 hours and is excreted by two mechanisms : the liver metabolizes approximately half to lactate , acetate , and pyruvate via oxidation by alcohol dehydrogenase and aldehyde dehydrogenase , and the other half is excreted unchanged by the kidneys . the renal clearance of pg decreases as the dose administered increases ( from 390 ml / min/1.73 m for a pg dose of 5 g / d to 144 ml / min/1.73 m for a dose of 21 g / d ) ; this may explain why patients with chronic kidney diseases or acute renal failure may be predisposed to pg toxicity , but our patient 's renal function was normal . the present literature suggests a broad spectrum in the severity of clinical manifestations of pg toxicity , ranging from mild metabolic abnormalities to life - threatening clinical conditions . moreover , pg toxicity has been documented over a wide range of serum pg levels ( 12 13,080 mg / dl ) and significant overlap exists between levels resulting in only mild metabolic abnormalities as compared with critical clinical deterioration . for instance , in the study by wilson et al , pg plasma levels ranged from 58 to 127 mg / dl for patients presenting only metabolic abnormalities and from 104 to 144 mg / dl in patients with clinical deterioration because of pg toxicity . there is no clear correlation between cumulative lorazepam ( and therefore pg ) dose and serum pg concentration and pg pharmacokinetic studies have shown considerable intra- and interpatient variation in serum pg concentrations for a given dosage administered . a hypothetical critical pg concentration beyond which clinical deterioration may be observed is presently unknown and appears to be patient dependent , suggesting a patient - specific variability in the metabolism , excretion , and thus toxicity of pg . most , if not all of the case reports on pg toxicity involved benzodiazepines ( mostly lorazepam ) , which contain the largest amounts of pg among the drugs routinely used in icu setting . in 2 published cases , patients were receiving tmp / smx but the tmp / smx accounted for only a minor fraction of total pg intake as patients were also receiving large doses of lorazepam . in these cases , the total pg intake was well above the threshold generally considered as safe . for instance , in the case reported by hayman et al , the patient had received a total of 552 g of pg within 12 days , only 123 g ( 20% ) of which administered with tmp / smx during the last 6 days . the second case involved a pg load of 540 g within 5 days administered with lorazepam infusion in a patient who was also receiving tmp / smx at unknown dosages . here we report a unique case of severe lactic acidosis developing after only a 3-day course of intravenous tmp / smx , involving relatively low amounts of pg ( 162 g within 3 days ) , well below the levels generally considered as toxic . applying the naranjo adverse drug reaction probability scale , pg was determined to be the probable cause for her lactic acidosis with a score of 7 : we assigned 1 point for previous conclusive reports on this reaction , 2 points for the time between drug administration and the occurrence of lactic acidosis , 1 point for the improvement in lactic acidosis after discontinuation of tmp / smx , 2 points for the absence of alternative causes , and 1 point for the objective evidence confirming the reaction . unfortunately our patient 's serum osmolality was not measured and we were unable to obtain a serum pg level to add further evidence of pg toxicity . we believe that this case is clinically relevant and should raise awareness among clinicians , as ( 1 ) tmp / smx is broadly used in critically ill immunocompromised patients and the fact that it contains pg is unrecognized , ( 2 ) critically ill patients often have conditions ( shock , acute renal failure , and lactic acidosis ) that may mimic pg toxicity but also contribute to it , ( 3 ) immediate recognition and cessation of tmp / smx is crucial to prevent the development of life - threatening complications in patients already prone to multiorgan failure . intravenous tmp / smx is often used in the setting of immunosuppression , either to treat documented p jirovecii pneumonia or as an empiric treatment in immunocompromised patients presenting with acute respiratory failure and diffuse pulmonary infiltrates concerning for p jirovecii pneumonia . in icu settings , such patients may be easily exposed to pg toxicity as they are prone to develop acute renal or liver failure , reported as risk factors for pg toxicity . on the other hand , patients receiving tmp / smx often present with signs of sepsis , and any clinical or biological deterioration will easily be attributed to the ongoing sepsis , while potentially severe adverse drug reactions may be overlooked . despite a severe lactic acidosis , our patient 's clinical condition remained stable ( except for the acidosis - related tachypnea ) as well as her renal function . the normalization of her serum bicarbonates and lactate levels within 24 hours after tmp / smx cessation is consistent with pg half - life . without tmp / smx cessation , our patient did not present with acute renal or liver failure , and had no history of chronic kidney , liver disease , or alcohol abuse , described as risks factors for pg toxicity . although tmp / smx is a known pg - containing drug , pg intake for our patient were below toxic levels and she did not receive other pg - containing drugs except for 2 mg of lorazepam , which only accounted for 0.8 g of pg and likely played a minor role in her lactic acidosis . our patient had findings on chest imaging concerning for pneumonia ; however all of her cultures remained sterile and she never developed hypotension or shock ( her mean arterial pressure remained above 65 mm hg the whole time , as shown in figure 3 ) , making hypoperfusion and resulting lactic acidosis secondary to sepsis extremely unlikely . similarly , her severe hypoxemia on presentation ( spo2 79% on room air ) is unlikely to account for the delayed lactic acidosis , as it was rapidly corrected with additional oxygen and mechanical ventilation ( see figure 4 ) , and metabolic acidosis developed 48 hours later while her oxygenation had been normal on mechanical ventilation . to our knowledge , severe lactic acidosis after a 3-day course of tmp / smx , involving allegedly safe doses of pg , has never been described . although pg - related toxicity following large doses of lorazepam is now well acknowledged , physicians should bear in mind that tmp / smx also contains pg , and suspect pg toxicity in patients developing metabolic acidosis while receiving tmp / smx , even at the recommended dosage and without risk factors for toxicity . immediate cessation of the drug is crucial to prevent the development of worsening multiorgan failure . | abstractpropylene glycol ( pg ) is used as a solvent in numerous medications , including trimethoprim / sulfamethoxazole ( tmp / smx ) and lorazepam , and is metabolized in the liver to lactic acid .
cases of lactic acidosis related to pg toxicity have been described and always involved large doses of benzodiazepines and pg . we present the first case of severe lactic acidosis after a 3-day course of tmp / smx alone , involving allegedly safe amounts of pg.a 31-year - old female with neurofibromatosis and pilocytic astrocytoma , receiving temozolomide and steroids , was admitted to the intensive care unit for pneumonia and acute respiratory failure requiring intubation .
her initial hemodynamic and acid base statuses were normal .
she was treated with intravenous tmp / smx for possible pneumocystis jirovecii pneumonia and was successfully extubated on day 2 . on day 3 , she developed tachypnea and arterial blood gas analysis revealed a severe metabolic acidosis ( ph 7.2 , pco2 19 mm hg , bicarbonates 8
meq / l ) with anion gap of 25 meq / l and lactate of 12.1 mmol / l .
tmp / smx was discontinued and the lactate decreased to 2.9
mmol / l within 24 hours while her plasma bicarbonates normalized , without additional intervention .
the patient never developed hypotension or severe hypoxia , and her renal and liver functions were normal .
no other cause for lactic acidosis was identified and it resolved after tmp / smx cessation alone , suggesting pg toxicity.although pg - related lactic acidosis is well recognized after large doses of lorazepam , clinicians should bear in mind that tmp / smx contains pg as well and should suspect pg toxicity in patients developing unexplained metabolic acidosis while receiving tmp / smx . |
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scientists and public health officials have observed and mapped the geographical incidence of infectious diseases in relation to weather and climate for hundreds of years , and formally for at least a half century ( reisen , 2010 ) . in recent years , this work has accelerated significantly in the context of predictions for global climate change ( ipcc , 2007b ) . the development of and response to the ipcc 4 predictions of probable increases of vector - borne and diarrheal diseases during coming decades ( ipcc , 2007a ) has catalyzed a great deal of research , analysis , and speculation regarding the nature , magnitude , and extent of possible changes . numerous , and in some cases conflicting , predictions have been developed regarding the frequency , severity , and duration of epidemics that may emerge . with respect to the biogeographical focus of this issue , the central question is whether pathogens and parasites that are currently restricted to tropics and lower latitudes where the world s greatest biodiversity lies move toward poles ( mostly north ) and upward in altitude . perhaps the more controversial topic today is the corollary to this question how much will future ranges of diseases that that do move be constrained by socioeconomic conditions , including our capacity to control them ( hay et al . , 2002 ; until very recently , climate projections for the coming decades have been limited to very coarse scale projections using global circulation models ( gcms ) at the level of continents and gross latitudinal and altitudinal changes ( fowler et al . , 2007 ) . downscaling such climate models is still in its infancy and only beginning to be useful for ecological and public health research at the regional , national , and sub - national scale for which most species distribution studies are conducted and validated ( beaumont et al . , 2008 ) . as such , epidemiologists , ecologists , geographers , and others are grappling with the implications of global climate projections for infectious diseases using a variety of approaches . these include biological process models , statistical geospatial analysis of current and historical prevalence and incidence , and macroeconomic demographic models , all coupled with climate analyses and projections . although we are still at an early stage in our ability to make predictions for these extraordinarily complex phenomena , we are beginning to see some general patterns with regard to the important geophysical factors that govern biological basis for distribution change , the role of transport of disease , vectors and hosts , the biotic assemblages that influence establishment , and the socioeconomic conditions that constrain or enhance these dynamics . underlying most predictions for climate change effects on parasite and pathogen distribution are the physiological factors that regulate survivorship , reproduction , and transmission , and their interaction with extrinsic environmental changes associated with climate : precipitation , humidity , air and water temperature , principally . under moderate greenhouse gas emission scenarios , gcm models project during the next century approximately 24c average warming and greater precipitation at higher latitudes , with decreased precipitation at lower latitudes , and increases in heavy precipitation events in many regions ( ipcc , 2007b ) . biological models estimate the responsiveness and thresholds of parasites , pathogens , and vectors to temperature and precipitation . these models can be used to estimate survivorship , transmissibility , and reproduction , including estimates of r0the basic reproductive ratio under different climate scenarios ( see lafferty , 2009 for review ) . based largely on studies of vector and/or parasite development , warming and increases in humidity are predicted to open up new zones for malaria in africa ( epstein et al . , 1998 ; martens , 1999 ) , parasitic nematodes in the arctic ( kutz et al . , 2005 ) , west nile virus ( reisen et al . , 2006 ) , lyme disease in north america ( ogden et al . , 2008 ) , and schistosomiasis in china ( zhou et al . , 2008 ) . historical analyses of climate patterns coupled with these biological process models provide additional explanatory power . for example , observed altitudinal increases in falciparum malaria in the east african highlands during the past 30 years have been associated with increasing temperatures and are consistent with models of anopheline mosquito vector development ( pascual et al . , 2006 ) . process models also suggest , as one would expect , that in some cases , the same climate trends may reduce survivorship or transmission of pathogens , parasites , and their vectors at the warmer boundaries of their current ranges ( lafferty , 2009 ) . that is because future temperatures may exceed the biological thresholds for survival or transmission at these boundaries . more analysis of biological maxima and minima of pathogens , parasites , and their vectors , including their diurnal fluctuation thresholds , in concert with downscaled climate models or historical records will be helpful in this regard . ( 2009 ) analyzed development time of plasmodium falciparum parasites in relation to that of its vector under a range of diurnal temperature fluctuations . they found that at the higher mean temperatures ( > 26c ) associated with endemic malaria transmission in africa effects of global warming on malaria dynamics may be overestimated . however , at lower mean temperatures associated with epidemic outbreaks ( < 20c ) , such as those found in the kenyan highlands , real increases in r0 with a couple degrees of warming are biologically possible , and consistent with epidemiological trends during the past 20 years in that region . similarly , changes in seasonality , especially intensity of rainfall , has been shown to be important in several diseases ( koelle et al . , 2005 ; altizer et al . more frequent heavy rainfall events are predicted to occur in some areas , although the confidence levels and scaling of these future precipitation models may still be insufficient to support geographically specific predictions . whereas vector - borne diseases , and to a lesser extent diarrheal diseases , have been the principal focus of discussion , directly transmitted diseases , such as influenza and cold viruses , are likely to change as well . recent laboratory work ( shaman and kohn , 2009 ) on the persistence and transmission efficiency of influenza has shown the virus to be constrained markedly by absolute humidity . more recent epidemiological analysis support this finding ( shaman et al . , 2010 ) and suggest that the marked seasonality and generally predictable geographical trajectory of seasonal influenza likely reflect , at least in part , climate variables . analyses of biological processes in relation to climate - relevant geophysical parameters establish the potential for climate change effects . however , they are insufficient to explain the highly complex dynamics of infectious disease transmission and establishment , including biological , climatological and socioeconomic considerations . a frequently used alternative approach to predicting biogeographic spread is based on mapping the statistics of current and historical disease or vector prevalence or transmission coupled with climate projections , often based on gcms ( rogers and randolph , 2000 ; mabaso et al . , 2006 ; more recent statistical mapping ( also called niche modeling ) approaches have been able to use downscaled climate models ( de la rocque , 2008 ) in their projections . several have predicted northward shifting zones of infection with modest impacts on overall extent of distribution ( rogers and randolph , 2000 ; hay et al . although essential , many of these statistical mapping projects make their own set of assumptions . particularly problematic are those that assume that today s public health and socioeconomic conditions , including surveillance and control capabilities , populations affected , and other ecological and economic conditions , will be similar to future conditions at new locations . one significant and long - presumed constraint on long - range biogeographical spread of invasive species , including pathogens , parasites , vectors , and hosts , has been the role of spatial distance on the ability to arrive at a new hospitable site ( macarthur and wilson , 1967 ; reperant , 2009 ) . this is , of course , less of a constraint today and for the foreseeable future due to a diversity of globalized transport opportunities . specifically , agriculture trade ( arzt et al . , 2010 ) , livestock movement ( fevre et al . , 2006 ) , wildlife trade ( karesh et al . , 2005 ) , human travel and migration ( macpherson et al . , 2009 ) , ship ballast water ( aguirre - macedo et al . , 2008 ) , auto tire trade ( hawley et al . , 1987 ) , and the global transport of desert dust containing microbiota regularly move pathogens , vectors , and infected hosts all around the earth . at small and intermediate scales , distance is still a very important determinant of disease movement . for example , triatomine vector dispersal of chagas disease in argentina at a scale of meters to a few kilometers ( zu dohna et al . , 2009 ) or movement of african trypanosomiasis at a scale of tens to hundreds of kilometers via animal migrations ( batchelor et al . , 2009 ) of course , if pathogens establish in a new zone and the process is allowed to continue incrementally year after year , diseases can and have moved across countries in a decade without the aid of modern transportation . however , at continental and intercontinental scales , movement of many diseases in a warming world is unlikely to have a linear relationship to distance because of the numerous means of long distance transport operating today . because infectious diseases are components of species networks , simply arriving in a new location may be insufficient for establishment and creation of endemic lifecycles . a number of studies have shown that emerging and reemerging diseases , particularly those in the tropics , are predominantly zoonotic and/or vector - borne diseases ( daszak et al . , 2000 ; woolhouse and gowtage - sequeria , 2005 ; wolfe et al . , 2007 ) . the principal ecological determinant for successful establishment for zoonotic and vector - borne disease success in a decadal time frame is probably a broad host range ( woolhouse and gowtage - sequeria , 2005 ; daszak et al . , 2000 ) . in recent years , a number of vector - borne viral diseases have migrated across or between continents through their ability to acquire new vectors ( west nile , chikungunya ) or utilize diverse and widely dispersed domestic ( rift valley ) or native ( west nile ) animal hosts ( gould and higgs , 2009 ) . importantly , the capacity to acquire new vector and host species often is facilitated by the capacity of the pathogen to evolve mutant strains that are efficiently acquired and transmitted by this new species . parasitic and infectious diseases with long , specialized , and/or especially complex life cycles involving multiple host species would logically be more challenged to establish themselves in new habitats by the need to find appropriate hosts and vectors . for example , the highly pathogenic alveolar echinococcosis ( fox tapeworm ) has a northern distribution that is influenced by climate but constrained by landscape variables that determine host availability ( danson et al . , 2003 ; mas - coma et al . , egg survival is dependent on ground moisture levels , but intermediate stages survive in rodents , and its definitive hosts are typically foxes , coyotes , and dogs . at large spatial scales across china and inner mongolia , a. echinococcus distribution has become focal due to the interaction of the requirements for a physical environment that is moist and cool enough to allow for egg survival and for the grassland landscapes that support both high densities of small mammals and canine hosts ( giraudoux et al . , 2006 ) . high levels of forest cover or cleared agricultural landscapes reduce intermediate and/or definitive host densities . it follows that for long - lived , host - specialized parasites , such as echinococcus , it is likely that long time periods are required for the evolutionary changes required to enable dispersal to zones with insufficient host densities of its original species . however , some historical and paleontological studies of parasites suggest that host - switching behavior in parasites in response to ecological changes has been more common than generally thought and global climate changes are likely to produce broadening of parasite host ranges even for long - lived , nominally specialized parasites ( brooks and hoberg , 2007 ) . by contrast , water - borne diarrheal diseases often are predicted to increase with climate change . biogeographically restrictive interaction of diarrhea - causing pathogens with new hosts or other pathogens has received little attention to date in the context of climate change . an exception is a series of papers by colwell and workers on associations of vibrio cholera with copepods in zooplankton ( gil et al . , 2004 ; constantin de magny et al . , 2008 ) . for directly transmitted human diseases , the sorts of species network challenges may be even less of a constraint . we see rapid cross - national and continental movement and invasion by new viruses , such as seasonal influenza , sars , and other coronaviruses . although much of the climate change and disease discussion has focused on vector - borne diseases ( lafferty , 2009 ) , the biotic simplicity of directly transmitted diseases , in concert with the climate sensitivity of viruses as discussed earlier , might argue that we should pay more attention to them in climate change discussions . although there has been little serious discussion that climate change will not impact infectious disease distributions , a view expounded by a number of authors ( reiter , 2008 ; zell et al . , 2008 ; lafferty , 2009 ) discounts the relative role of climate change compared with other major influences on human disease . these factors include efficacy of control measures , such as vector control , sanitation , vaccination , and chemotherapy ( hay et al . , 2002 ) , as well as land use , urbanization , travel , wealth , and social behavior ( gubler et al . , 2001 ; sumilo et al . , statistical mapping analyses ( rogers and randolph , 2000 ; hay et al . , 2004 ) indicate that the spatial extent and number of countries reporting endemic malaria has shrunk toward the tropics from a much wider range during the twentieth century . the authors attribute this principally to better control of vectors and disease . further support can be seen in the rapid reemergence of tick - borne encephalitis in central and eastern european states after the collapse of the soviet union ( 19702005 ) ( sumilo et al . , 2008 ) . statistical comparisons of results of extensive socioeconomic surveys and public health data across five former soviet states indicate that various socioeconomic , land use , and public health programs can explain most of the variation across countries in the spread of tick - borne encephalitis during this period . in this analysis , climate variables were not assessed , but significant unemployment , reversion of agricultural lands , reduced pesticide use , reduced chemotherapy , and individual exposure through increased field collection of wild foods appear to have contributed to rapid increases during the early 1990s . the interaction of climate change , infectious diseases , socioeconomic conditions , and behavior is going to be increasingly important in the coming years and represents an exciting and complex area for research ( patz et al . , 2005 ) . for example , several authors have predicted mass migrations in response to climate change , particularly in response to worsening droughts in sub - saharan africa ( ramin and mcmichael , 2009 ) ( ipcc , 2007a ) . if borne out , it is likely that many diseases will travel with migrants , and in some cases will establish new infectious species or strains with significant effects on receiver populations ( macpherson et al . , 2009 ) . similarly , availability of clean water and hygienic behavior are principal determinants of diarrheal disease outbreaks , and clean water will be increasingly in short supply in much of africa and parts of southeast asia in the coming decades due to overexploitation and climate change ( ipcc , 2007a ) . in an analysis of frequency all cause diarrhea in children younger than age 5 years in low- and middle - income countries , lloyd et al . ( 2007 ) found that low mean monthly rainfall during the course of a year was significantly associated with disease . surprisingly , neither temperature nor simple measures of socioeconomic development ( gdp or gcp ) were significant in this analysis of a disease type mostly managed by traditional public health measures , such as sanitation and education . diarrheal diseases are produced by a diversity of bacterial , viral , and protozoan organisms . strain diversity within a species is high locally ( e.g. , jiang et al . , 2000 ) around the world , and migration to new zones has significant implications for biologically nave host populations . finally , given the importance and complexity of socioeconomic conditions on diseases , a variety of approaches and disciplines will be necessary to assess their effects on pathogen and parasite biogeography ( parkes et al . , 2005 ) . macroeconomic and demographic modeling may offer underexploited utility in this regard . a recent paper by tang et al . ( 2009 ) is instructive ; they set out to disentangle the causality of a climate - income trap model for human health by examining national life expectancy statistics ( 45 african , 113 non - african countries ) in a cross - sectional analysis . using a series of regression models and variation partitioning analyses to separate pure and combined effects of income and climate their findings support the hypotheses that : 1 ) income can moderate the adverse effect of climate on life expectancy ( le ) ; 2 ) as income grows climate will have less of an effect on le ; and 3 ) climate has a larger effect on le in african countries than in non - african countries , and further that for african countries , climate has had a larger effect on le than wealth debates over the impact of climate change on infectious disease emergence and migration are certain to continue in the years to come . these with statistical and mapping approaches help provide us greater clarity and ensure that we are asking the right questions . macroeconomic and demographic approaches may help us address some of the big questions related to the role of disease management capability . critical to a greater understanding of these dynamics and more predictive power is greater availability of downscaled general circulation models to scales that are more relevant to community and landscape biology and public health . i have not addressed time scales in any depth here , but they are extremely important aspects of biogeographic changes in relation to climate . longitudinal studies on the multi - decadal time frames that climate change presents are a major challenge . but the time required for evolution of host range , for population dynamics of hosts and parasites to stabilize , and for medical and public health systems to adapt will all have major roles in determining biogeographic patterns of disease , and will all operate relatively independently . debates about whether socioeconomic conditions or climate will be more important are probably more polarized than is productive . both of these complex suites of variables are interacting with each other , and may do so in different ways for different diseases in different places . however , understanding these disease forcing factors and their interactions with each other are crucial to our ability to predict and prevent the most severe health outcomes . the public health and medical community has made enormous strides in control of infectious diseases during the past half - century . widespread availability of antimicrobial drugs , vector - control systems , diagnostics , vaccines , and increasingly sophisticated predictive models represent a powerful set of tools to protect public health from emerging diseases . however , it would be unwise to discount the potential importance of climate change , land use impacts , and other ecological determinants of future disease . historically , ecological factors , including climate , have been extremely important in determining both the range and severity of diseases . these factors are now aggravated by the near disappearance of antimicrobial drug development in the industrial pharmaceutical sector , dramatic reductions in training and support for vector control scientists and implementers , persistent scientific and financial challenges in developing new drugs and vaccines for even the most widespread diseases ( e.g. , hiv , malaria , tb ) , and medical workforce shortages in the poorest parts of the world . as our scientific capabilities to describe and model diseases and disease processes expands rapidly , it will be important to ensure that we understand the changing nature of our planet and continually adapt our systems and invest in them to meet the challenges . | our ability to predict the effects of climate change on the spread of infectious diseases is in its infancy .
numerous , and in some cases conflicting , predictions have been developed , principally based on models of biological processes or mapping of current and historical disease statistics .
current debates on whether climate change , relative to socioeconomic determinants , will be a major influence on human disease distributions are useful to help identify research needs but are probably artificially polarized .
we have at least identified many of the critical geophysical constraints , transport opportunities , biotic requirements for some disease systems , and some of the socioeconomic factors that govern the process of migration and establishment of parasites and pathogens .
furthermore , we are beginning to develop a mechanistic understanding of many of these variables at specific sites .
better predictive understanding will emerge in the coming years from analyses regarding how these variables interact with each other . |
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diabetes mellitus ( dm ) with a prevalence of 5.1% of all world population is characterized by high blood glucose levels ( 1 - 3 ) . dm has several long - term complications including cerebrovascular , coronary , and peripheral vascular diseases , nephropathy , retinopathy and neuropathy ( 4 , 5 ) . diabetic neuropathy is one of the most frequent and severe complications of diabetes which could affect sensory , motor , and autonomic nerves ( 6 ) . despite numerous efforts to uncover the precise mechanism(s ) of diabetic neuropathy hyperglycemia plays a key role in the development of diabetic complications ( 7 ) ; on the other hand a significant improvement in neuropathy following reduction of glucose level has been reported ( 8) . however , it is suggested that insufficiency of arterial blood flow due to a high glucose - induced atherosclerosis ( vascular aspect ) may be involved ( 8 , 9 ) . in addition , intracellular alteration of biochemical reactions induced by direct effect of high glucose on neuronal cells ( non - vascular aspect ) may also contribute ( 6 , 8 - 10 ) . some of this biochemical alterations including increased formation of advanced glycation end products , activation of protein kinase c ( pkc ) through increased diacylglycerol production , enhanced polyol pathway flux and increased hexosamine pathway flux have been reported as underlying mechanisms ( 9 ) . increased glucose concentrations activate the reactive oxygen species ( ros ) production through two main sources , the first is nicotinamide adenine dinucleotide phosphate ( nadph ) oxidase located on plasma membrane , and the second are mitochondria through a dysfunction of electron transport chain ( 2 , 11 - 13 ) . a large growing body of evidence reports an increased oxidative injuries induced by hyperglycemia in many diabetic tissues such as neural tissue ( 14 - 16 ) . in fact , oxidative stress is an upstream crucial trigger which promotes numerous detrimental pathways ending to apoptosis and cell death ( 9 , 13 ) . several studies have reported that apoptosis is the other mechanism involved in diabetic neural injury ( 2 , 17 , 18 ) . aside from classical renin - angiotensin system ( ras ) , a hormonal one maintaining blood pressure , electrolytes and water balance in the body , there are local or even intracellular renin - angiotensin systems in different tissues ( 5 , 19 - 21 ) . researchers refer many physiological and pathophysiological functions to local ras including inflammation , fibrosis , proliferation , differentiation or even apoptosis ( 22 ) . the local ras physiologically has been found in almost all body parts including : adrenal gland ( 5 , 21 ) , pancreas ( 23 ) , heart ( 24 ) , blood vessels ( 25 ) , kidney ( 26 ) , nervous system ( 19 , 22 ) , reproductive system ( 27 ) , and many tissues such as adipose tissue ( 28 ) . several evidences have been reported an over - activation of ras in diabetic complications , as it has been well - documented that the renin - angiotensin system ( ras ) plays a major role in the pathophysiology of diabetic nephropathy ( 26 ) . angiotensin ii has two main receptors named angiotensin receptor type 1 ( at1 ) and type 2 ( at2 ) ( 9 , 29 ) . actually the most known actions of angiotensin such as vasoconstriction , renal sodium reabsorption and also aldosterone and vasopressin secretion are mediated by at1 but the roles of at2 receptor remain mysterious and controversial ( 29 ) . at2 receptor has been abundantly found in fetus organs , uterus and neural tissues ( 29 ) . surprisingly , functions such as cellular proliferation , differentiation , and apoptosis that have been attributed to at2 receptors are corresponded well to the local ras ( 9 , 29 ) . there is several evidence of activation of local ras in most of diabetic tissues as well as neural tissue ( 8 , 9 , 26 ) . recently angiotensin converting enzyme ( ace ) inhibitors associated with angiotensin ii receptor antagonist are recognized as first line treatment for nephropathy(9 , 30 ) . in addition , it seems that ras contributes to almost all biochemical events which are promoted by hyperglycemia and finally result in neural cell death ( 9 ) . not only hyperglycemia , but also angiotensin ii , the main effector of ras , is among strong inducers of oxidative stress ( 9 , 31 , 32 ) . ( ang ii ) could add to high glucose - induced ros production and cause even more damages . interestingly , in a study on proximal tubule cells in rat , it was found that the effect of hyperglycemia on stimulation of angiotensinogen gene expression was through increasing ros production ( 33 ) . it was suggested that besides locally ang ii elevation following hyperglycemia , increased ros formation may in turn act as a second messenger for local ang ii production ( 13 ) . as previously mentioned , one possible mechanism to develop diabetic neuropathy is reduction of blood supply due to vasoconstriction of microvasculature ; thus , ace inhibitors such as captopril or at1 antagonists such as telmisartan and losartan as vasodilator agents can be curative in this situation ( 11 , 30 ) . although , some studies have shown a vascular effect of these drugs , but nonvascular actions of these agents in high glucose - induced neuropathy is still unclear ( 9 ) . the required components of an intrinsic ras have been found in rat 's adrenal medulla as well as pc12 cells ( 21 , 34 ) . the pc12 cells , pheochromocytoma cells are derived from rat s adrenal medulla , have a neural crest origin and abundantly express at2 angiotensin receptor ( 34 - 37 ) . in the present study , the role of local ras on high glucose - induced cell toxicity , apoptosis and ros production was investigated in pc12 cells . 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium ( mtt ) , 2,7-dichlorofluorescein diacetate ( dcf - da ) , captopril , telmisartan , losartan , pd-123319 , dimethyl sulfoxide , and d - glucose were purchased from sigma - aldrich company ( st . dmem , fetal bovine serum , trypsin , penicillin , and streptomycin were obtained from gibco brl life technologies ( grand island , ny , usa ) and annexin v - fitc & pi kit was purchased from biovision ( mountain view , ca , usa ) . pc12 cells were routinely cultured in dulbecco s modified eagle s medium , containing 4.5 mm - d-(+ ) -glucose , 10% heat - inactivated fetal bovine serum , 100u / ml penicillin , and , 100 u / ml streptomycin , incubated at a temperature of 37c in a humidified atmosphere of 95% air and 5% co2 , and were passaged when reached to 70% confluency . pc12 cells were pretreated for 3 hr prior to and after high glucose - induced toxicity with a glucose concentration of 27 mg / ml by following drugs : captopril ( 10 m ) , telmisartan ( 10 m ) , losartan ( 10 m ) , pd-123319 ( 10 m ) , and two combination groups include telmisartan plus pd-123319 ( each 10 m ) , and , losartan plus pd-123319 ( each 10 m ) . to evaluate cell viability , pc12 cells were seeded ( at 1x10 cells per well ) in a 96-well culture plate and cell viability was measured by mtt colorimetric assay . thus , after treatment , 11 l mtt solution ( 5mg / ml ) was added into each well and cells were incubated at 37c for 4h in a humidified atmosphere containing of 5% co2 and 95% air . after removing the medium , formazan crystals the absorption was measured at 570 nm ( 620 nm as a reference wavelength ) by means of an elisa reader . apoptosis was evaluated by annexin v - fitc and propidium iodide ( pi ) staining followed by flow cytometry . cells were detached and annexin v&pi double staining was performed according to manufacturer 's instructions and then measured by flow cytometry . after treatment , the pc12 cells once washed with prewarmed pbs , then they were incubated with 10 m dcfh - da at 37c in the dark . 30 min later , the cells were rewashed and resuspended in pbs for analysis by facscalibur flow cytometer ( bd , usa ) with excitation and emission wave lengths of 475 and 525 nm , respectively . for each analysis all results derived from at least three independent experiments are expressed as mean sem . the significance of difference was analyzed by a one - way anova followed by tukey as post - test using graphpad prism 5 and p<0.05 was considered to be statistically significant . 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium ( mtt ) , 2,7-dichlorofluorescein diacetate ( dcf - da ) , captopril , telmisartan , losartan , pd-123319 , dimethyl sulfoxide , and d - glucose were purchased from sigma - aldrich company ( st . dmem , fetal bovine serum , trypsin , penicillin , and streptomycin were obtained from gibco brl life technologies ( grand island , ny , usa ) and annexin v - fitc & pi kit was purchased from biovision ( mountain view , ca , usa ) . pc12 cells were routinely cultured in dulbecco s modified eagle s medium , containing 4.5 mm - d-(+ ) -glucose , 10% heat - inactivated fetal bovine serum , 100u / ml penicillin , and , 100 u / ml streptomycin , incubated at a temperature of 37c in a humidified atmosphere of 95% air and 5% co2 , and were passaged when reached to 70% confluency . pc12 cells were pretreated for 3 hr prior to and after high glucose - induced toxicity with a glucose concentration of 27 mg / ml by following drugs : captopril ( 10 m ) , telmisartan ( 10 m ) , losartan ( 10 m ) , pd-123319 ( 10 m ) , and two combination groups include telmisartan plus pd-123319 ( each 10 m ) , and , losartan plus pd-123319 ( each 10 m ) . to evaluate cell viability , pc12 cells were seeded ( at 1x10 cells per well ) in a 96-well culture plate and cell viability was measured by mtt colorimetric assay . thus , after treatment , 11 l mtt solution ( 5mg / ml ) was added into each well and cells were incubated at 37c for 4h in a humidified atmosphere containing of 5% co2 and 95% air . after removing the medium the absorption was measured at 570 nm ( 620 nm as a reference wavelength ) by means of an elisa reader . apoptosis was evaluated by annexin v - fitc and propidium iodide ( pi ) staining followed by flow cytometry . cells were detached and annexin v&pi double staining was performed according to manufacturer 's instructions and then measured by flow cytometry . after treatment , the pc12 cells once washed with prewarmed pbs , then they were incubated with 10 m dcfh - da at 37c in the dark . 30 min later , the cells were rewashed and resuspended in pbs for analysis by facscalibur flow cytometer ( bd , usa ) with excitation and emission wave lengths of 475 and 525 nm , respectively . for each analysis all results derived from at least three independent experiments are expressed as mean sem . the significance of difference was analyzed by a one - way anova followed by tukey as post - test using graphpad prism 5 and p<0.05 was considered to be statistically significant . mtt assay showed that treatment with high glucose concentrations decreased cell viability in a time- and concentration - dependent manner . concentrations of 13.5 , 18 , 22.5 and 27 mg / ml showed significant difference ( p<0.05-p<0.001 ) . figure 2 shows that 27 mg / ml of glucose had cytotoxic effects in a time dependent manner and it decreased cell viability at 96 , 120 and 144 hrs ( p<0.05-p<0.001 ) . the cell viability at 27 mg / ml of glucose ( high glucose ; hg ) was significantly lower than normal glucose ( ng ) concentration after 144 hrs ( p<0.001 ) . viability was increased in experimental groups : captopril ( p<0.05 ) , telmisartan ( p<0.001 ) , pd123319 ( p<0.05 ) , telmisartan / pd123319 ( p<0.001 ) , and losartan+pd123319 ( p<0.01 ) , respectively ( figure 3 ) . the flow cytometric results of annexin v - fitc and propidium iodide staining assay showed that apoptotic and necrotic cells were increased in high glucose condition ( 27 mg / ml ) by ( p<0.001 ) and ( p<0.01 ) respectively . pretreatment followed by treatment with captopril ( p0.05 ) , telmisartan ( p<0.01 ) , pd123319 ( p0.01 ) , telmisartan / pd123319 ( p<0.001 ) and losartan / pd123319 ( p<0.01 ) signify - cantly decreased apoptotic cells in high glucose conditions . as shown in figure 4 , telmisartan and telmisartan / pd groups had significant reduction ( p<0.05 ) in necrotic cells . dcf - da used as a probe to evaluate the intracellular ros alterations by flow cytometery . it was revealed that high glucose concentration increased the ros production after 1 h ( p<0.001 ) . as the figure 5 shows it was increased to the most high level after 2 hrs ( p<0.001 ) and then it was returned to the base up to 5 hrs . the figure 6 shows the effects of the drugs on decreasing ros production at 27 mg / ml of glucose after 2 hrs . the ros production in high glucose ( hg ) condition was increased compared to ng condition ( p<0.001 ) . treatment by quercetin ( rutin ) as an antioxidant decreased the ros production ( p<0.001 ) . after 3hs pretreatment and then 2hs treatment with the drugs in high glucose medium ( 27 mg / ml ) , tel / pd ( p<0.001 ) , telmisartan ( p<0.001 ) , los / pd ( p<0.01 ) , captopril ( p<0.01 ) and losartan ( p<0.05 ) , all treatments reduced ros production as amount of 69 , 66 , 40 , 34 and 32% , respectively . telmisartan in comparison with losartan had a better effect in lowering ros which statistically significant ( figures 6 , 7 , p<0.01 ) . mtt assay showed that treatment with high glucose concentrations decreased cell viability in a time- and concentration - dependent manner . concentrations of 13.5 , 18 , 22.5 and 27 mg / ml showed significant difference ( p<0.05-p<0.001 ) . figure 2 shows that 27 mg / ml of glucose had cytotoxic effects in a time dependent manner and it decreased cell viability at 96 , 120 and 144 hrs ( p<0.05-p<0.001 ) . the cell viability at 27 mg / ml of glucose ( high glucose ; hg ) was significantly lower than normal glucose ( ng ) concentration after 144 hrs ( p<0.001 ) . viability was increased in experimental groups : captopril ( p<0.05 ) , telmisartan ( p<0.001 ) , pd123319 ( p<0.05 ) , telmisartan / pd123319 ( p<0.001 ) , and losartan+pd123319 ( p<0.01 ) , respectively ( figure 3 ) . the flow cytometric results of annexin v - fitc and propidium iodide staining assay showed that apoptotic and necrotic cells were increased in high glucose condition ( 27 mg / ml ) by ( p<0.001 ) and ( p<0.01 ) respectively . pretreatment followed by treatment with captopril ( p0.05 ) , telmisartan ( p<0.01 ) , pd123319 ( p0.01 ) , telmisartan / pd123319 ( p<0.001 ) and losartan / pd123319 ( p<0.01 ) signify - cantly decreased apoptotic cells in high glucose conditions . as shown in figure 4 , telmisartan and telmisartan / pd groups had significant reduction ( p<0.05 ) in necrotic cells . dcf - da used as a probe to evaluate the intracellular ros alterations by flow cytometery . it was revealed that high glucose concentration increased the ros production after 1 h ( p<0.001 ) . as the figure 5 shows it was increased to the most high level after 2 hrs ( p<0.001 ) and then it was returned to the base up to 5 hrs . the figure 6 shows the effects of the drugs on decreasing ros production at 27 mg / ml of glucose after 2 hrs . the ros production in high glucose ( hg ) condition was increased compared to ng condition ( p<0.001 ) . treatment by quercetin ( rutin ) as an antioxidant decreased the ros production ( p<0.001 ) . after 3hs pretreatment and then 2hs treatment with the drugs in high glucose medium ( 27 mg / ml ) , tel / pd ( p<0.001 ) , telmisartan ( p<0.001 ) , los / pd ( p<0.01 ) , captopril ( p<0.01 ) and losartan ( p<0.05 ) , all treatments reduced ros production as amount of 69 , 66 , 40 , 34 and 32% , respectively . telmisartan in comparison with losartan had a better effect in lowering ros which statistically significant ( figures 6 , 7 , p<0.01 ) . the results of present study showed that high levels of glucose had both time and concentration dependent toxicity effects in pc12 cells . the results also showed that the cytotoxic effects of glucose were accompanied with increasing of ros production , the later was also attenuated by ras inhibitor drugs . hyperglycemia clearly plays an important role in the development and progression of diabetic neuropathy which characterized by an increasing loss of nerve fiber functions ( 1 , 2 ) . pc 12 cells or pheochromocytoma cells have neural crest origin and thus frequently have been used as a cell model for studying of high glucose toxicity ; the condition which mimics the effects of hyperglycemia to induce neuropathy ( 2 , 3 , 6 , 12 , 18 ) . despite a long time needed for diabetic neuropathy to occur in human , diabetic neuropathy symptoms appear in stz - induced diabetic rats just after 8 - 10 weeks ( 38 , 39 ) ; also , direct exposure of high glucose concentrations to susceptible cells such as pc12 cells can accelerate this deleterious process . it has been well - documented in pc12 cells that 4 - 7 days exposure to high glucose concentrations could decrease cell viability significantly ; thereby , this high glucose cytotoxicity is considered as an acceptable model to simulate diabetic neuropathy in many publications ( 2 , 3 , 6 , 12 , 18 ) . it has been well documented that hyperglycemia in diabetes mellitus as well as high glucose toxicity lead to neuronal damage , apoptosis and cell death ( 1 , 2 ) . the results of the present study also showed that the cell viability in high glucose concentrations was time and concentration dependently lower than normal concentration of glucose . the flow cytometry results of annexin v - fitc and propidium iodide staining also showed that apoptotic pc12 cells were increased in high glucose concentrations . several mechanisms including mitochondrial - dependent and -independent mechanisms are involved in diabetic neuropathy ( 40 ) . it was shown that mitogen - activated protein kinases ( mapk ) and p53-mediated pathways have important roles in apoptosis and cell death due to hyperglycemia ( 40 ) . an increased formation of advanced glycation end products and activation of protein kinase c ( pkc ) through increased diacylglycerol production have been reported to contribute in high glucose toxicity ( 41 , 42 ) . enhanced polyol pathway flux , and increased hexosamine pathway flux have also been suggested to be involved in neuronal damage due to high glucose levels ( 40 - 42 ) . it has also been well documented that oxidative stress , directly , or , as a second messenger , is one of the main mechanism involved in development of diabetic neuropathy(2,13 , 43 ) . the results of the present study added another evidence to confirm that free radicals have an important role in neural damage and cell death as well as apoptosis in hyperglycemia . it has been previously reported that hyperglycemia promotes oxidative stress through cell - membrane enzyme , nicotinamide adenine dinucleotide phosphate ( nadph ) oxidase , and mitochondrial pathways in diabetic neurons resulting in neuronal damage and dysfunction ( 8 , 17 ) . it has also been reported that hyperglycemia can induce apoptosis in neural cell such as pc12 cells by increasing production of reactive oxygen species such as hydrogen peroxide ( 3,6 , 12 ) . in addition to classic renin angiotensin system that is well known for its hemodynamic and cardiovascular functions , the presence of local renin angiotensin systems and their physiological roles within different tissues and organs including nervous system have been reported ( 5 , 19 , 22 - 28 ) . it has been well documented that local rass are involved in several cell processes such as growth , differentiation , and even apoptosis at cellular levels ( 9 , 22 ) . it was suggested that the local ras contributes to virtually all biochemical events including ros production and apoptosis which are triggered by hyperglycemia ( 9,24 , 26 ) . in the present study , our finding showed that all ras inhibitors including captopril , telmisartan and pd123319 increased the cell viability in high glucose toxicity . the neuroprotective effects of ace inhibitor as well as the angiotensin receptor antagonists ( 11,30 , 44 ) confirm the results observed in the present study . these drugs have been shown to treat diabetic neuropathy mostly with vascular aspect ( 30 ) . coppy and his colleagues showed that administration of enalapril , as an ace inhibitor , or l-158809 , as an angiotensin ii receptor blocker , reduced neurovascular deficits such as blood flow and motor nerve conduction velocity in diabetic conditions ( 11 ) . malik et al assessed the effect of trandolapril , an ace blocker , on human diabetic neuropathy and found a significant increase in peroneal motor nerve conduction velocity , m - wave amplitude and sural nerve action potential amplitude ; also a significant decrease in f - wave latency ( 44 ) . several mechanisms have been suggested for the protective effects of ras inhibitors against hyperglycemic conditions ( 30,45 , 46 ) . it has been suggested that , not only hyperglycemia , but also ang ii is an inducer of oxidative stress ( 9 , 31 ) . it has also been reported that oxidative stress can in turn induce expression of ang ii and its receptors , in a positive feedback manner ( 9 , 47 ) . in fact , ros as a second messenger may serve as a link between ras , hyperglycemia and apoptosis in pathophysiological events of diabetic neuropathy . reduction of ros production by ras inhibitors which were seen in the present study confirmed this idea . on the other hand , the insignificant effects of losartan on improving cell viability and reducing apoptosis which was observed here might be attributed to the presence of a limited number of at1 receptors on pc12 cells ( 36 ) . the results of ros production showed a decrease in high glucose - induced ros formation after treating by all drugs except pd123319 . several experiments have suggested a receptor - independent intracellular radical scavenging activity for ace inhibitors and at1 receptor blockers ( 48 ) . according to some studies , thus , such at1 independent antioxidant activity was observed in this study at least in case of losartan is in agreement with above reports ; although , it needs further investigation in future . in addition to its at1 blocker activity , telmisartan also serve as partial agonist of peroxisome proliferator - activated receptor gamma ( ppar- ) ( 50 , 51 ) . furthermore , the effectiveness of telmisartan seems to be through activation of different receptors , such as ppar- , which has been suggested as a possible mechanism for its neuroprotective effects ( 52 , 53 ) . it has been previously reported that the anti - inflammatory and antioxidant activities of telmisartan is mainly mediated activating of the ppar- receptor ( 50,52,53 ) . thiazolidinediones as ppar- agonists for many years have been used in diabetic treatments ( 54 , 55 ) . these drugs were even proposed as neuroprotective factors in several conditions such as ischemia and glucose cytotoxicity ( 56 , 57 ) . the presence of ppar- receptor on pc12 cells ( 58 ) and the stronger free radical scavenging effects of telmisartan in comparison with losartan which is seen in the present study might be another evidence for the effects of telmisartan by activating of ppar- receptors . overall , the results of the present study showed that telmisartan , pd123319 and captopril antagonized high glucose - induced cell death and apoptosis but only telmisartan and captopril performed these effects through reducing ros - induced cell damage . the results of the present study showed that telmisartan , pd123319 and captopril antagonized high glucose - induced cell death and apoptosis . so , it seems that at2 receptor is important in neuroprotection against high glucose toxicity by a mechanism other than protection against oxidative stress . as previously has been reported , nadph oxidase has a pivotal role in ang ii - induced ros production ( 13 ) ; this mechanism and/or other mechanism(s ) may contribute in ros generation . it is also suggested that both losartan and telmisartan have a non at1 receptor mediated antioxidant effects . the better antioxidant and cell protective effects of telmisartan observed in the present study are suggested to be through other pathways such as ppar- receptors . | objective(s):hyperglycemia , oxidative stress and apoptosis have key roles in pathogenesis of
diabetic neuropathy .
there are local renin - angiotensin systems ( rass ) in different tissues such
as neural tissue .
local rass are involved in physiological and pathophysiological processes such
as inflammation , proliferation and apoptosis .
this study aimed to investigate the role of local
renin - angiotensin system on high glucose - induced cell toxicity , apoptosis and reactive oxygen
species ( ros ) production in pc12 cells , as a cell model of diabetic neuropathy.materials and methods : pc12 cells were exposed to a high glucose concentration ( 27 mg / ml ) ,
captopril ( ace inhibitor ) , telmisartan and losartan ( at1 antagonists ) , and also pd123319 ( at2
antagonist ) were administered before and after induction of high glucose toxicity .
then cell
viability was assessed by mtt assay and apoptotic cells and intracellular ros production were
detected by annexin v - propidium iodide and dcfda , respectively , using flow cytometry.results:high glucose concentration decreased cell viability , and increased apoptotic cells .
intracellular ros production was also increased . in pc12 cells pretreatment and treatment by
the drugs showed a significant improvement in cell viability and reduced apoptosis in captopril ,
telmisartan and pd123319 but only captopril and telmisartan were able to reduce ros
production .
losrtan significantly lowered ros but did n't show any improvements in cell viability
and apoptotic cells.conclusion:the results of the present study showed that ras inhibitors reduced cell toxicity
and apoptosis and ros production was induced by high glucose .
it may be suggested that local
ras has a role in high glucose toxicity . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
human immunodeficiency virus ( hiv)-associated immune reconstitution inflammatory syndrome ( iris ) has emerged as an important early complication of antiretroviral therapy ( art ) initiation , associated with considerable morbidity and mortality . in this condition , immune recovery following the art initiation associates with a pathological inflammatory response , usually directed toward the microbial antigens . this was first noted following the introduction of zidovudine monotherapy in the early 1990s , when the localized forms of mycobacterium avium - intracellulare infection were observed in association with the recovery rather than the failure of cellular immune responses . here , we present a 7-year - old hiv positive girl who developed iris as a complication of the initiation of art . a 7-year - old hiv infected girl presented with chronic diarrhea and hypocalcemic tetany ( serum calcium = 5.6 mg / dl [ normal = 8.5 to 11 mg / dl ] ) . both the parents were also hiv infected but not on art . on examination , her weight was 12 kg , height was 103 cm , had generalized as nonsignificant lymphadenopathy with edema feet and pallor . on systemic examination , she had hepatosplenomegaly . her mantoux test was negative ; barium meal follow through was normal , and stool showed 2025 pus cells / hpf . she was treated with intravenous ( iv ) ceftriaxone , metronidazole , nitazoxanide , and fluconazole to which she responded along with calcium supplements . she was treated with iv cefotaxime , amikacin , tmp - smx , and required inotropes in the form of dopamine and dobutamine , and also prednisolone to which she responded . diarrhea in patients with acquired immune deficiency syndrome ( aids ) is significantly caused by intestinal parasites . studies in adults have demonstrated that enteritis due to g. intestinalis is a frequent event among the aids patients , especially in the most advanced stage of the disease . in a study of 75 hiv - infected adults in india , g. intestinalis was the most commonly isolated parasite , and patients with the lower cd4 cell counts presented with significantly more enteric disease and chronic diarrhea . these include tuberculosis , herpes zoster ( shingles ) , cryptococcus neoformans , kaposi 's sarcoma , pneumocystis pneumonia , hepatitis b virus , hepatitis c virus , herpes simplex virus , histoplasma capsulatum , human papillomavirus , and cytomegalovirus . however , the immune reconstitution syndrome ( irs ) has never been documented with a giardiasis or its treatment . our patient was treated empirically with anti - bacterials such as ceftriaxone ; anti - protozoal such as metronidazole and nitazoxanide ; and anti - fungal agents such as fluconazole because the routine stool examination did not detect any particular organism and hiv - infected children are known to have polymicrobial infections . however , the post - art , the preexisting infections which might have been partially treated or may have been subclinical was later unmasked by the patient 's regained capacity to mount an inflammatory response . giardiasis was then diagnosed from the stool sample where it showed up as the causative organism . the patient 's symptoms , this time were more severe , and she went into sepsis for which she was given in inotropic support . a low baseline cd4 + t cell count has also been documented as a major risk factor for the development of iris . in a study conducted at a hospital in mumbai , india , to evaluate incidence and risk factors for iris in 37 hiv - infected children , it was found that the patients with opportunistic infections before the antiretroviral treatment would have increased the incidence of iris . however , there was no correlation between the degree of immunosuppression and development of irs . there may also be a genetic predisposition and certain genes have been associated with an increased susceptibility to the development of iris in the presence of mycobacteria and herpes viruses which might have been the case with our patient who had a prior history of herpes zoster infection . therefore , this case highlights that giardiasis may also be considered as a cause of iris in patients with hiv infection . learning points from the case in family medicine and the primary care :
always screen for iris in patients who present with infection post - artiris may be associated with unusual infections . always screen for iris in patients who present with infection post - art iris may be associated with unusual infections . | human immunodeficiency virus ( hiv)-associated immune reconstitution inflammatory syndrome has been reported in association with tuberculosis , herpes zoster ( shingles ) , cryptococcus neoformans , kaposi 's sarcoma , pneumocystis pneumonia , hepatitis b virus , hepatitis c virus , herpes simplex virus , histoplasma capsulatum , human papillomavirus , and cytomegalovirus . however , it has never been documented with giardiasis .
we present a 7-year - old hiv infected girl who developed diarrhea and shock following the initiation of antiretroviral therapy , and her stool showed the presence of giardiasis . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
lifestyle - related diseases such as hazardous drinking of alcohol are gaining an increasing actuality within the public health . changing the lifestyle of risk groups is an important part of health - related prevention activities . hereby , the role of doctors as ideal examples to the population with regard to their lifestyles and as experts to prevent lifestyle - related diseases stay in focus . how much effort a doctor spends on prevention actions against hazardous drinking depends on several factors - one of them is his own individual practice . several studies have shown that practising a healthful behaviour individually is the most consistent and powerful predictor of how doctors are counselling patients about related prevention issues , . furthermore , doctors ' health behaviours appear to affect the patient 's attitude and their motivation to make lifestyle changes . however , previous studies have shown that doctors have high levels of alcohol consumption , , and run an increased risk of alcohol - related diseases such as alcohol - related mortality , cancer , accident and suicide , . a clear sign for problem - related drinking is a suicide rate , that was found to be higher among doctors than among the general population or other academics in several countries , . if we wish to improve the personal health status of doctors and their preventive interventions in their daily clinical work with patients , we need information on the group of doctors with problem - related drinking . until today therefore , the purpose of this article is to examine and review representative data of the prevalence of problem - related drinking among doctors undertaken in international studies and to discuss their results . a search of computer literature databases - pubmed and etoh - was performed to locate articles reporting problem - related drinking of doctors . in order to strengthen the comparison among doctors , selection criteria were established to minimise the limitation and potential threats to validity inherent to all studies . first , comparison was based only on studies where population - based samples of doctors were used . these studies used probability sampling techniques that meant every medical speciality had a chance of being selected , thus enhancing the generalising of the results . second , to get more reliable results in the present surveys on alcohol - related problems within the medical profession , only published data within the past two decades were selected . 1 ) reviewing the literature was created for the medical profession based on representative data , organising the publications by methodology , the type of samples studied , and population survey based samples . in addition to summarising the characteristics of each of the samples , the outcome variables and a summary of the findings are provided . because of the methodological differences , this review does not entirely rely on statistical significance as a basis for comparison , but rather on the ranking and prevalence findings . most of the studies have examined the alcohol consumption of doctors in a single speciality , gender group , geographic region , or done by medical students . tabulated summaries and direct comparisons across the selected studies are difficult , because the definitions of actual drinking and problem - related drinking - alcohol misuse , hazardous drinking , heavy drinking - are classified in various ways . the results show that most of the doctors belong to the group of actual drinkers : about 94% of the us doctors drank alcohol within the past year , , 80%-85% of the norwegian doctors drank more than once monthly within the past year , and 93% of the finnish doctors drank more than 1 g alcohol during the past week . with regard to problem - related drinking , a wide range of findings is shown : in finland , a high alcohol consumption of > 200 g alcohol a week was found among 16% of doctors . in norway , the proportion of doctors with hazardous drinking according to the audit - score of 6 or more , increased from 12.2% to 16.5% from 1993 to 2000 . in the new england states of the us , the percentage of combined abusers and potential abusers was 7.8% , and in the us study the percentage of doctors with alcohol abuse problems or dependence on alcohol was 6% . high alcohol consumption in finland and hazardous drinking in norway were positively related to gender and to age , e.g. more male than female doctors , and more older than younger doctors were apt to qualify as actual drinkers and risk group . a strength of all selected studies is that they collected data by mailed self - administrated questionnaires , and so , in comparison with other methods , have a relatively high reliable validity rate concerning self - reported alcohol consumption . however , if we consider the general underreporting of alcohol consumption , which is about 40%-60% without much variation between nationals , then the volume of alcohol consumption could be even greater . in the present review , selected studies showed alcohol consumption as an integral part of the daily life of doctors . in regard to the prevalence of problem - related drinking among doctors , a wide variation in the reported number of doctors affected with this problem is shown , ca . the exact incidence is difficult to ascertain because statistical sources for estimates on doctors are conducted in a number of ways , including the definition of problem - related drinking : both nordic studies have focused on the measure of the early stage of problem - related drinking as harmful or hazardous alcohol use , and the us studies on the measure of the later stage of problem - related drinking such as abuse of or dependence on alcohol . therefore , the differences between nordic and us studies could be even smaller than those found here . however , drinking habits are connected to cultural factors . for instance , in a pilot project of the author , , concerning attitudes , smoking habits and drinking patterns of doctors it was shown that doctors in aarhus ( denmark ) - correlating with a generally more liberal attitude to alcohol - had a significantly higher prevalence of heavy drinkers than doctors in mainz . this favourable situation seen from an epidemiological point - of - view does , of course , not mean that doctors in mainz do not have any alcohol problems . among doctors in mainz , 17% drank hazardously at least once a month and 6% at least once a week . this pilot project suggests rates of hazardous drinking for doctors which are similar to the 6%-16% rates of problem - related drinking in the representative samples . applying the prevalence rates taken from the representative studies - apart from the methodological differences - to german conditions , a considerable number of 30,000 to 40,000 practising doctors with a problematical alcohol consumption will emerge . the recent representative studies do show distinctive differences in the prevalence rates for problematical alcohol consumption among doctors . both nordic studies have found some demographic factors for problem - related drinking : an increased risk was positively related to male doctors and doctors over the age of 40 years in finland and the age of 45 years in norway . this seems surprising when it is shown that within the general population ( at least in finland , norway and also in germany ) older people drink less than the younger ones . the age - effect among doctors was explained by the fact that younger doctors put an increased emphasis on the dangers of alcohol which will hopefully contribute to an increased risk - awareness concerning personal consumption . a similar precedent has been set by the change in doctors ' smoking habits in the us and many other european countries . a support for this explanation could be the conclusion of a norwegian study among doctors that younger doctors are becoming more aware of the danger of alcohol use as a public health problem . independent of the age - effect , expected gender differences in drinking - females are less likely to drink alcohol and less likely to be heavier drinkers , - were also present in the norwegian and finnish study . yet , their alcohol consumption and prevalence of problem - related drinking can vary , e.g. between medical specialities . a pilot project of the author in aarhus and mainz and a study in norway showed that female doctors within surgical specialities were more likely to engage in detrimental drinking than other female doctors . here , the culture of surgery as a masculine culture may play a special role . of course , there are several reasons why doctors may be susceptive to problem - related drinking . causal factors may include individual traits , drinking patterns during their time at medical school , social conditions and the highly responsible nature of their profession , , , . in this respect , it may be of interest to mention mcauliffe and his colleagues ' observation of sensation - seeking as a prominent factor in recreational alcohol drinking among physicians - in - training . it is also important to note the presence of stressful conditions within the medical profession . however , there is only a limited empirical support for a relationship between job stress and alcohol consumption . it is discussed that such a relationship exists if one believes in the efficacy of alcohol to relieve stress . on the other hand it is also mentioned that work stress factors can result in job dissatisfaction , or burnout which can relate with heavy drinking , which was also shown in the finnish study . special attention and help for the medical profession are generally needed : firstly , doctors with a regular alcohol abuse pose a problem for patients , fellow - workers , and last , but not least for their own health . secondly , doctors find it on the whole difficult to cope with their own physical and psychological ailments . thirdly , even within their own ranks , fellow - doctors do not consider alcohol - related problems as serious . fourthly , the attitude of doctors towards their own health has a direct impact upon the population , i.e. doctors play an important part as role - models for a healthy way - of - life , and as experts for finding solutions concerning health - problems , . for the future it would be important to pay more attention to the drinking patterns of doctors in germany . if one wants to give doctors with problem - related drinking in germany the benefit of professional help , and to strengthen their engagement in the field of prevention , one can not do otherwise but to strive for data compilations in a more systematic way , and to reflect on them both conceptually and methodically . at first , we need to have representative data to identify the risk - factors . understanding the factors related to drinking would be greatly aided by closely studying not only gender and age differences , but also other important variables such as speciality differences , work - related conditions and health status in a large scale survey . moreover , because of the increasing number of women in medicine , more attention should be paid to alcohol consumption patterns of female doctors . in addition , since data from other countries suggest problem - related drinking among doctors , it would be of interest to use comparative data from other countries for future analysis . with the help of data we can find better ways of planning preventive and interventional strategies for this group , e.g. initiate projects to assess the changing working - conditions for doctors in order to improve the situation and to offer help to physicians who experience such problems in their professional work . proceeding on the assumption that it is possible to influence attitudes and to change ways of behaviour with regards to drinking by training , for the future our hope may lie in promoting more intensive education concerning alcohol issues among doctors . we have a good chance to prevent harmful drinking in this group of professionals , because in the light of u.s . studies , doctors as a group probably respond favourably to prevention , even more favourably to alcohol addiction programmes , than do members of the general population . | aims : this paper is a review of the literature on problem - related drinking of alcohol among medical doctors , and it deals with the epidemiology and results .
methods : a search of computer literature databases - pubmed and etoh - was performed to locate articles reporting problem - related drinking among doctors , using population - based samples of doctors within the last two decades.results : in the light of different definitions of problem - related drinking , there was found a breadth of prevalence of problem - related drinking - from heavy drinking and hazardous drinking ( 12%-16% ) to misuse and dependence ( 6%-8% ) - within the population - based samples of doctors .
an increased risk was positively related to male doctors and doctors of the age of 40 - 45 years and older , and to some factors of work , lifestyle and health .
conclusion : for the future , it seems necessary to sensitise the research for problem - related drinking of doctors in germany , e.g. initiating a representative survey , analysing the drinking of alcohol in the context of health , life - style and work - related factors . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
heparina , a highly sulfated glycosaminoglycan , has the highest negative charge density of any known natural biological molecule . heparin has been extensively used as an injectable anticoagulant in many clinical procedures for the prevention of blood clotting , especially during open heart surgery [ 2 , 3 ] . pharmaceutical - grade heparin is derived from mucosal tissues of slaughtered meat animals such as porcine intestine and bovine lung . native heparin is a polymer with a molecular weight ranging from 3,000 to 50,000 , and the average molecular weight of most commercially prepared heparin is in the range of 12,000 to 15,000 . under physiological conditions , the sulfate groups are deprotonated and attract positively charged counterions ( usually sodium ) to form a heparin salt . it is in this form that heparin is usually administered as an anticoagulant [ 5 , 6 ] . at the end of the medical procedure , the heparin has to be rendered inactive to prevent possible bleeding problems . therefore , clinicians need to know the exact concentration of heparin remaining when they attempt to neutralize the anticoagulant activity of heparin via precise doses of protamine . the current heparin detection method used in hospital is based on the blood clotting time [ 7 , 8 ] and varies widely due to variation in the blood content of the different individuals . thus , the discovery of accurate , rapid , reliable safe , and low - cost methods of heparin detection remains an important area of sensor research . for example , electrochemical sensors [ 9 , 10 ] and optical sensors [ 11 , 12 ] were intensively studies in the 1990s , and more recent studies have focused on fluorescent spectroscopy [ 13 , 14 ] and uv / vis spectroscopy using gold nanoparticles . luminescence spectroscopy is an analytical method that studies the luminescence emission from a chromophore after their electrons are excited from the ground state to a higher energy level . one of the most attractive features of luminescence spectroscopy as an analytical method is its inherent high sensitivity , wide linear response range , and fast detection speed . however , the luminescence intensity of the chromophore ( or excited state complex ) can be seriously reduced by collision with other chemical species or enhanced by chemical interactions . through studying the effect of these chemical species on the luminescence intensity of the chromophore , the quantitative determinations of a variety of important inorganic and organic species in trace amounts have been realized by photoluminescence methods . recently , a series of os(ii ) carbonyl ( co ) complexes with two bipyridine or phenanthroline ligands and one pyridyl or triphenylphosphine ligand as the five other ligands have been synthesized in our lab . the osmium centers in these complexes are all in the + 2 oxidation state with an overall + 2 charge of the complex ion , and exhibit high emission intensity in the visible region [ 17 , 18 ] . this research focuses on studying the effect of the heparin polyanion on the photoluminescence of these polypyridyl osmium(ii ) complexes , for the purpose of finding potential luminescence markers for the detection of the optically nonactive polyanion heparin . to our surprise , polyanions show unique influences on the luminescence spectra and intensity of these osmium complexes . the 1,10-phenanthroline ( phen ) complexes were compared with the 2,2-bipyridine ( bpy ) complexes . for each of these chromophoric ligands , three analogous sets were compared using pyridine ( py ) , triphenylphosphine ( pph3 ) , or 4-phenylpyridine ( 4-phpy ) as the sixth ligand . these ligands showed a significant impact on the luminescence characteristics of the osmium complex in the absence as well as in the presence of polyanions . the results of the studies could lead to a new , potential effective detection method for heparin monitoring . the six new os(ii ) complexes are [ os(phen)2co(pph3)](pf6)2 , [ os(phen)2co(py)](pf6)2 , [ os(phen)2co(4-phpy)](pf6)2 , [ os(bpy)2co(pph3)](pf6)2 , [ os(bpy)2co(py)](pf6)2 , and [ os(bpy)2co(4-phpy)](pf6)2 . the structure of each complex is shown in figure 1 . the synthesis of six osmium complexes was described in [ 17 , 18 ] . heparin sodium sulfate ( > 180 usp units / mg ) , ethanol ( 99.5+% ) , reagent or analytical grade naclo4 , mgso4 , ch3coona , nacl , kbr , nano3 , naoh , nano2 , kscn , and ki were all obtained from sigma - aldrich . injectable heparin solutions , 100 units / ml and 40 units / ml with 5% dextrose , were purchased from b. braun medical inc . luminescence measurements were made on a perkin elmer luminescence spectrometer ls 50 b equipped with a high - intensity xenon flash lamp . a branson 1210 sonicator was used to help dissolve the osmium complex in solvent when needed . thermo scientific digital pipets were used for the addition of all solutions into the quartz cuvette . the stock solutions of the osmium complexes were prepared by dissolving 0.62.8 mg of each osmium complex in ethanol ( ethanol solution ) or distilled water ( aqueous solution ) in a 100 ml volumetric flask . the 1.0 mg / ml heparin aqueous solution was prepared by dissolving 5.0 mg of heparin in 5.00 ml of distilled water . the 0.10 mg / ml and 0.010 mg / ml heparin aqueous solutions were diluted from the 1.0 mg / ml stock solution . for small anions solutions , a 0.100 m solution of each salts was first made and then diluted to 0.0100 m and 0.00100 m. for the luminescence measurements of each osmium complex solution ( ethanol , aqueous ) , 2 ml stock solution was transferred to a quartz cuvette ( 1 cm 1 cm 4.4 the emission and excitation slits were both set at 10 nm for the luminescence measurements . a prescan was taken first to obtain the wavelengths of the maximum excitation and emission intensities ; then the emission spectrum was recorded by exciting the sample at the appropriate excitation wavelength . in order to study the anion quenching or enhancement of the luminescence of each osmium complex solution , 2.00 ml of the stock solution was placed in the cuvette , and the emission spectrum was measured ; then a small amount ( 10 l or 20 l ) of the anion solution ( 0.010~1.0 mg / ml for heparin and 0.10 to 0.0010 m for small anions ) was added to the cuvette and the emission spectrum was recorded after the solution was well mixed . the anion solution was continuously added to the cuvette , and the emission spectrum was recorded after each addition . the values of the luminescence intensity were taken at a fixed wavelength at or near the emission maximum . first , 10 or 20 l of injectable heparin solution was added to 2 ml of the aqueous [ os(phen)2co(4-phpy)](pf6)2 solution . next three portions of 5 l of 1 mg / ml standard heparin solution were added , and the emission spectrum of the solution was measured before and after each addition . the concentrations of the heparin in the injectable solution were calculated using the standard addition curve . the excitation and emission spectra of each complex in their ethanol solution ( except for [ os(bpy)2co(py)](pf6)2 which could not dissolve in ethanol ) and their aqueous solution were first recorded at room temperature under ambient air . all the spectra exhibited broad emission and excitation bands up to 200 nm in width . figure 2 shows the emission spectra in ethanol solution of the two complexes : [ os(phen)2co(pph3)](pf6)2 , and [ os(phen)2co(py)](pf6)2 , which exibit the largest difference in luminescence intensity . as shown in table 1 and figure 2 , the two os - pph3 complexes have much higher luminescence intensities and shorter emission wavelengths than the two os-(4-phpy ) complexes and the two os - py complexes ( 520 nm versus 560 nm ) . the shorter wavelength ( higher energy ) of the emission is caused by the -backbonding ability of the phosphine which stabilizes the homo of the molecule relative to pyridine . the higher luminescence intensities are attributed to the larger steric effect of the pph3 ligand which shields the complex from colliding with the solvent . this slows down the vibrational relaxation to the ground vibrational level of the excited state that must occur prior to emission of a photon of light that occurs when the electron returns to the ground electronic state . all these six complexes exhibit higher luminescence intensities in aqueous solution than in ethanol solution due to the hydrophobicity of the ligands surrounding the osmium center . the spectral changes that occur before and after the addition of small inorganic anion solutions suggest that those anions clo4
, so4
, ch3coo , cl , br and no3
exhibit very little quenching of the luminescence of the osmium complex with anion concentrations up to 0.01 m , while no2
, scn and i anions can dramatically quench the luminescence of the osmium complexes with concentrations at 0.01 mm . however , oh exhibits a small enhancement of the luminescence of the osmium complex ; luminescence intensity slightly increased as the solution ph increased from 6 to 9 in aqueous solution . it was expected that heparin would quench the luminescence of the osmium complexes due to its high negative charges . surprisingly , however , it was found that heparin actually enhanced the luminescence intensity of the osmium complexes in ethanol solutions . the luminescence of each complex in the ethanol and aqueous solution changed quite differently after heparin was added . in ethanol solution , each complex exhibited an increase in luminescence intensity after heparin was added in concentrations greater than 0.5 g / ml . however , the amount of increase varied significantly for each complex . the solutions of [ os(bpy)2co(4-phpy)](pf6)2 , [ os(phen)2co(4-phpy)](pf6)2 , and [ os(phen)2co(py)](pf6)2 exhibited a very large increase in luminescence intensity while the solutions of [ os(phen)2co(pph3)](pf6)2 and [ os(bpy)2co(pph3)](pf6)2 exhibited a small luminescence increase . figure 3 shows the luminescence spectra of the ethanol solution of [ os(bpy)2co(4-phpy)](pf6)2 in the absence and presence of heparin at different concentrations . the luminescence intensity was increased from a relative emission intensity of 136 in the absence of heparin to 462 by addition of heparin in a concentration range of 5.038.5 g / ml . further , there is an 8 nm blue shift in wavelength of the emission maxima with high heparin concentrations . a plot showing the wider range of heparin concentrations on the enhancement ratio ( f / f
0 ) is shown in figure 4 . here , f and f
0 are the luminescence ( fluorescence ) intensity of the osmium complexes solutions at 560 nm with and without heparin , respectively . the luminescence intensity increased steadily up to a heparin concentration of 56 g / ml . in the lower concentration range ( up to 30 g / ml ) , there is a linear relationship between the f / f
0 ratio and the concentration with an r
of 0.99 . the heparin enhancement of the [ os(phen)2co(4-phpy)](pf6)2 ( 2.4 mg/100 ml etoh ) luminescence has a similar pattern to that seen with [ os(bpy)2co(4-phpy)](pf6)2 . in this case , the luminescence was enhanced by three and half times when the added heparin concentration was increased to 50 g / ml . once again , there is an 8 nm blue shift of the emission maxima at higher heparin concentrations . in the concentration range of 1 to 40 g / ml , there is a linear relationship between the f / f
0 ratio and the heparin concentration with an r
of 0.99 . the heparin enhancement of the luminescence of [ os(phen)2co(py)](pf6)2 ( 2.4 mg/100 ml etoh ) is smaller than that seen for the 4-phpy complexes . the luminescence intensity was enhanced by two and half times when the added heparin concentration was increased to 2030 g / ml , and then the luminescence started to decrease at heparin concentrations in excess of 30 m / ml . in this case , there is also an 8 nm blue shift of the emission maxima at higher heparin concentrations . the heparin enhancement of the luminescence of the [ os(phen)2co(pph3)](pf6)2 ( 2.4 mg/100 ml etoh ) and the [ os(bpy)2co(pph3)](pf6)2 ( 2.1 mg/100 ml etoh ) solutions is very small compared to the os-(4-phpy ) and os - py complexes . here , the shift in the emission maxima is insignificant ( less than 2 nm ) . figure 5 shows the plot of f / f
0 versus heparin concentration for the two phenanthroline complexes with pyridine and pph3 as the sixth ligand . from figures 4 and 5 , it can be seen that the 4-phpy complexes , which have the lowest solution luminescence in the absence of polyanions , showed the highest enhancement in the concentration range of 540 g / ml added heparin concentrations . by contrast , the pph3 complexes showed the smallest enhancement in the same concentration range . the osmium complexes luminescence enhancement by heparin is likely contributed by the close contact of heparin and the complexes , which forms a rigid structure that reduced the solvent collision and quenching . heparin has a long linear structure with anionic sites on its branches when it is dissolved in aqueous solution . when its aqueous solution is added to the ethanol solution of the osmium complex , the anion sites on its branches attract the double positively charged osmium complex cations . as a result , the osmium complex could electrostatically bind to heparin and become surrounded by the heparin molecules ( figure 6 ) . in this case , solvent molecules can no longer get close enough to the osmium complex to help relax the excited state , resulting in the emission maximum moving to higher energy . this effect would also explain the increased emission intensity since the rate of nonradiative decay would be greatly reduced in the absence of favorable interaction with solvent molecules . the sixth ligand in these complexes plays an important role in the interaction of these charged sites . the complexes [ os(bpy)2co(4-phpy ) ] , [ os(phen)2co(4-phpy ) ] , or [ os(phen)2co(py ) ] have small narrow 4-phenylpyridine or pyridine ligand . when heparin is added to their ethanol solution , these osmium complexes could readily approach the heparin polyanion and bind up to two heparin anion sites . these two sites may well form two heparin polyanions branches , thus forming ion pairs between the polyanion and osmium complex . therefore , heparin can cause significant enhancement to the luminescence of the os-(4-phpy ) and os - py complexes . further , the luminescence only stops increasing at very high heparin concentrations when all the osmium complex cations are bound to heparin anion sites . by contrast , the [ os(phen)2co(pph3 ) ] and [ os(bpy)2co(pph3 ) ] complexes have a sterically bulky pph3 group , which prevents them from approaching the heparin polyanion closely . therefore , the luminescence intensity of the os - pph3 complexes could not be significantly enhanced by heparin . the effect of heparin on the emission of these osmium complexes in their aqueous solution is quite different from that of their ethanol solution . the aqueous solution of osmium complexes with a pph3 or pyridine group exhibits a decrease in luminescence intensity at low added heparin concentrations ; then the luminescence increases at heparin concentration above 15 to 19 g / ml . however , complexes with the 4-phpy group exhibit a decrease of luminescence at heparin concentrations below 0.5 g / ml then an increase in luminescence intensity is observed with the addition of heparin in the range of 1 to 15 g / ml . this shows that as a polyanion , when not binding to the osmium complexes , heparin is an efficient luminescence quencher in aqueous solutions . this is especially true with the pph3 complexes , where binding only occurs at very high ( > 15 g / ml ) heparin concentrations . by contrast , the 4-phpy complexes bind more strongly to heparin , and so less of a quenching effect is observed . figure 7 shows the plots of the luminescence enhancement ratio ( f / f
0 ) as a function of heparin concentration for the two os-(4-phpy ) complexes in aqueous solution . the high luminescence enhancement ratio of the 1,10-phenanthroline complexes by heparin shown in figure 7 suggests that the [ os(phen)2co(4-phpy ) ] complex binds more strongly to the heparin polyanion than the analogous 2,2-bipyridine complex complexes . comparing figures 4 , 5 , and 7 , the enhancement of heparin to the [ os - co(bpy)2(4-phpy ) ] complex is much smaller in aqueous solutions , which indicate a weaker binding to heparin , possibly due to the higher solubility of heparin and the complexes in aqueous solution . in the aqueous solutions of [ os(phen)2co(pph3)](pf6)2 or [ os(bpy)2co(pph3)](pf6)2 , it is even harder for the osmium complex to bind to heparin because of the high solubility of heparin in aqueous solution and the hindrance of the bulky pph3 group . in fact , at low heparin concentration , heparin quenches the luminescence of these two complexes heavily as polyanion . the decrease of the luminescence intensity in the aqueous solution of these two complexes fits the stern - volmer equation
( 1)f0f=1+ksv[q ] .
here f
0 and f are the luminescence signals in the absence and in the presence of a quencher , respectively ; k
sv is the stern - volmer quenching constant ; and [ q ] is the concentration of the quencher . stern - volmer plot is shown in figure 8 for the [ os(phen)2co(pph3 ) ] complex . the high linearity of the stern - volmer curve , shown at the concentration range of 1 to 12 g / ml in the insert , confirms that heparin exhibits dynamic quenching to the os - pph3 complexes . the nonlinearity at higher concentrations may indicate that osmium complex - heparin ion - pairs have formed . further addition of heparin above 12 g / ml caused the luminescence of these complexes to increase slightly . table 2 summarizes the heparin concentration ranges that induced significant luminescence changes to the complexes studied in ethanol or aqueous solutions . in ethanol solution , [ os(bpy)2co(4-phpy)](pf6)2 exhibits the greatest enhancement in the presence of heparin as measured by the highest slope of the f / f
0 versus heparin concentration linear curve . this complex exhibits the longest response range and the highest sensitivity to heparin concentration . in aqueous solutions , the [ os(phen)2co(4-phpy)](pf6)2 and both phosphine complexes showed significant luminescence changes at low heparin concentrations . the detection of heparin was then performed with commercial medicinal solutions to evaluate the feasibility . two commercial heparin sodium injection solutions ( 40 and 100 units / ml in 5% dextrose , in 100 ml plastic bags ) from b. braun medical inc . were tested . with the [ os(phen)2co(4-phpy ) ] complex in aqueous solution , the emission intensities of the osmium solution before and after adding 20 l of these two heparin solutions were measured . the ratios ( f / f
0 ) obtained were 1.30 and 1.97 ( n = 5 ) , respectively , showing that the osmium complex indeed had a greater response to higher heparin concentration preparations . however , these medical preparation responses are much higher than the pure heparin solution response . when response curve of heparin in the presence of 5% dextrose was performed , an enhanced osmium complex response to heparin concentration was observed . this enhanced response is likely due to the hydrophobic environment that the polysaccharide , dextrose , created around the osmium complex in aqueous solution , thus reducing the extent of nonradiative decay of the excited states and increasing the luminescence intensity of the osmium complex . to eliminate the background effect , first , 10 or 20 l of the commercial heparin sample solutions was added to the aqueous osmium complex solution ; then three portions of 5 l of the 1.0 mg / ml heparin standard solution were added . the luminescence values of the complex solution after each addition of the sample and standard heparin solutions were measured at 560 nm . with the standard addition curves and using the conversion of 180 units / mg of sodium heparin , the average heparin concentration in the two sample bags was calculated to be 46 3 and 111 the relative standard deviation , rsd for the two corresponding samples are 5.6 ( n = 3 ) and 2.3 ( n = 6 ) respectively , indicating that the precision of the method is acceptable considering the volume added is very small , although they are 11% higher than the labeled value . further tests , with other commercial sources having different backgrounds , are currently under study . the luminescence of osmium carbonyl complexes is stable at room temperature in ethanol and aqueous solutions , though the intensity and wavelength of this emission are heavily dependent on the other five ligands . the effect of heparin on the luminescence of the osmium complexes depends heavily on both the ligand and solvent used . in ethanol solutions , heparin could enhance the luminescence of five osmium complexes at low heparin concentrations , with the greatest enhancement occurring with the os-(4-phpy ) complexes . this increased emission intensity may be due to binding of heparin to the double charged complexes which changed the solvent environment of the complexes . in aqueous solutions , heparin quenching the osmium complex emission was more significant as heparin is more soluble in water and behaves as an anion quencher . these osmium complexes are shown to be useful for the fast and sensitive detection of heparin in commercially injectable samples . | the luminescence characteristics of six osmium carbonyl complexes with phenanthroline ( phen ) or bipyridine ( bpy ) and pyridine ( py ) , 4-phenylpyridine ( 4-phpy ) , or triphenylphosphine ( pph3 ) complexes in the presence of polyanion heparin were studied in both ethanol and aqueous solutions .
the influence of heparin on the luminescence of the complexes is heavily dependent on the type of ligands in the complexes and the solvent used . in the ethanol solutions , the heparin solution enhanced the luminescence of the five osmium complexes , with the strongest enhancement to the 4-phenylpyridine complexes ;
linear curves were obtained in the luminescence enhancement ratio ( f / f
0 ) versus the heparin concentration range of 140 g / ml . in aqueous solutions , heparin quenching of the complexes was more significant ; a linear quenching curve was obtained with [ os(phen)2co(pph3)](pf6)2 in the lower concentration range of 112 g / ml . the interaction of these complexes with heparin in the solutions is discussed .
the complexes are shown to be successful in the fast and sensitive detection of heparin in commercial injectable samples . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
success rates of dental implants in the edentulous mandible exceed 95% in most long - term clinical studies.1,2 however , implant failure is still considerable in areas of poor quality bone3,4 such as the maxillary region . the long - term success of implant therapy is achieved by the primary stability of the implant for mechanical support from the surrounding bone in the early stage and osseointegration between the surrounding bone and implant through bone regeneration and remodeling in the late stage.5 primary stability is especially necessary in poor quality bone . the instability of dental implants results in fibrous encapsulation and failure to achieve osseointegration.6 primary stability is fundamental for successful osseointegration . clinicians and implant manufacturers have made a major effort to increase the success rates of dental implants by improving primary stability . one of the methods of increasing primary stability is by modifying the surgical technique for implant placement . studies have reported that the undersized surgical technique , which uses a final drill diameter smaller than the diameter of the implant , results in higher primary stability than dose the press - fit technique.5,7 other studies have reported higher implant stability with the bone - condensing technique compared with bone - drilling technique8,9 and the conventional technique compared with the osteotome technique.10,11 surface roughness is also important for achieving good primary stability . in mechanical evaluation , dental implants having higher average roughness showed better primary stability than machined implants did5,12,13 and good osseointegration between the implant and surrounding bone in animal studies.14,15,16 another method for increasing primary stability is to change the implant design , such as the shape of the implant body and thread , the length , or the diameter . various thread designs in tapered implants and various designs of dental implants have been reported to have an effect on primary stability . it has been reported that tapered implants showed higher primary stability than cylindrical implants.17,18,19 in addition , dental implants with a long length or wide diameter showed a significant increase in insertion torque.17,19 one study has reported that using dental implants without self - tapping blades increases the primary stability compared with that with self - tapping blades.20 several studies have described the stress distribution in ti implants with various thread depths by using finite element analysis ( fea ) and reported the most effective thread depth for stress distribution.21,22,23 thread depth has a greater contribution than thread width to stress distribution to the bone.23 ti implants with a deeper thread depth provide a larger surface area and have an advantage in areas of poor quality bone by increasing stability.24 ti implants with deeper thread depths may increase loads on and mechanical interlocking with poor quality bone . although several studies have measured the mechanical stability through stress distribution in ti implants with various thread depths by fea , to our knowledge , no mechanical studies investigating the effects of the thread depth of dental implants on enhancing primary stability have been published . we used tapered implants with deeper thread depths than commercially available dental implants to increase the primary stability . we investigated the primary stability in tapered implants with various thread depths by mechanical testing . to measure the primary stability of implants , periotest , resonance frequency analysis ( rfa ) , insertion torque , and removal torque are possible methods . however , the periotest is not able to identify minor differences25,26 and with rfa an implant stability quotient ( isq ) cutoff value for sufficient primary stability has not been defined , so isq values for different implant systems can not be compared.27 the insertion torque and removal torque is commonly used to measure the primary stability of dental implants . increasing the insertion torque can increase the primary stability through reduction of the micromotion in soft bone,28 but excessive insertion torque causes a high incidence of failure.29 therefore , the control of insertion torque for implant placement is important . we evaluated the primary stability of dental implants with various thread depths by using the insertion torque test , and the stability of deeper threads from breakage after mechanical strength testing was also analyzed . commercial ti implants ( anyridge internal implants ) roughened by grit - blasting were used in this study ( megagen co. ltd . , kyungsan , korea ) . detailed information about the length , diameter , and thread depth of the ti implants is specified in table 1 . we selected ti implants possessing the same length and inner diameter but with different outer diameters . we compared the test values of group a and group b and those of group c and group d. ez post abutments were prepared for static compressive strength . the ez post had a profile diameter of 5.0 mm , a cuff height of 3.0 mm , and a post height of 5.5 mm for groups a and b and a profile diameter of 6.0 mm , a cuff height of 3.0 mm , and a post height of 7.0 mm for groups c and d. solid rigid polyurethane blocks ( sawbones pacific research laboratories inc . , vashon island , wa , usa ) with uniformity were used as an alternative to human cancellous bone . there is little variation in the qualities of the material among samples , and thus the blocks can be used as materials for comparative tests . insertion tests using solid rigid polyurethane blocks provide more objective data than those using animal or human cadaver bones . the bone mineral density of the solid rigid polyurethane blocks was 0.16 g / cm , 0.24 g / cm , and 0.32 g / cm . all of the bone holes for implant placement were prepared according to the recommended surgical protocol supplied by the manufacturer . the ti implants were installed using a surgical engine ( elcomed sa200c , w&h , brmoos , austria ) with a rotation speed of 30 rpm and a torque value of 70 ncm . the recorded torque value was read by impdat software ( kea software gmbh , poecking , germany ) . the ti implant and the ez post abutment were tightened with the recommended torque ( 30 ncm ) by using a digital torque meter ( mark-10 , new york , ny , usa ) . the ti implants tightened with the ez post containing the hemispherical loading member were fixed with a specimen holder that was made from brass and clamped in the jig of a universal test machine ( instron , 3366 , instron , corp . , norwood , ma , usa ) . the implant / abutment assembly was placed at 30 against the axis of the loading direction and set at a distance of about l = 11 mm from the center of the hemispherical loading member to the clamping plane of a jig , as shown fig . 2 . the loading device was positioned in contact with the top of the hemispherical loading member and loaded at a rate of 1.0 mm / min in a unidirectional vertical direction until the failure load that induced buckling was observed . the value of the load and displacement was recorded by series ix software ( instron , 3366 , instron , corp . , the hemispherical loading member was removed from the implant / abutment assembly after the static compressive strength tests . the implant / abutment was mounted with an acrylic resin ( r&b , daejeon , korea ) using an automatic mounting press ( r&b , daejeon , korea ) and was polished by using a polisher ( r&b , daejeon , korea ) changing the grit of the sand paper ( 400 , 800 , 1500 ) . the paired student 's t - test was performed to compare the significance of the differences . the results of the mean insertion torque found for a bone density of 0.16 g / cm are shown in table 2 . the mean insertion torques of group a and group b were 12.37 0.40 and 20.53 1.07 , and those of group c and the group d were 28.93 1.07 and 36.17 0.40 , respectively . the results of the mean insertion torque found for a bone density of 0.24 g / cm are shown in table 3 . the mean insertion torques of groups a and b were 20.77 1.07 and 32.67 2.02 , and those of groups c and d were 26.83 1.46 and 50.87 2.83 , respectively . the results of the mean insertion torque found for a bone density of 0.32 g / cm are shown in table 4 . the mean insertion torques of groups a and b were 9.10 1.21 and 35.47 0.40 and those of groups c and d were 35.70 4.20 and 68.83 2.65 , respectively . the ti implants with deeper threads had significantly higher insertion torque for all bone densities tested ( p<.001 ) . the 10 load - displacement curves in the same group showed a similar pattern and the distance of displacement in the implants with the same inner diameter ( group a and b ; group c and d ) was similar . the maximum compressive values , that is , the maximum compressive load , are shown in fig . the ti implants with the same length and inner diameter showed a similar maximum compressive load regardless of the thread depth ( p>.05 ) . the failure mode was observed to be deformation in the abutment and being torn horizontally at the upper side of the ti implant ( fig . the threads in the ti implants with deeper threads did not show breakage ( fig . we enlarged the surface area by creating a deeper thread depth to improve the primary stability . our study investigated the primary stability in tapered implants with various thread depths using mechanical tests . tapered implants with deeper thread depths resulted in higher insertion torque and thus showed better primary stability . the periotest has a low correlation with torque and does not explain the variation between the parameters.25 it is more effective at gathering information on osseointegration than primary stability.26 rfa is not comparable to the isq values obtained from different implant systems and has a low correlation with insertion torque.27 thus rfa is not suitable as a single method for the measurement of primary stability . the insertion torque and removal torque is commonly used to measure the primary stability of dental implants in mechanical testing . insertion torque is a more effective indicator of primary stability than the rfa and the periotest . our studies investigated the effect of thread depth on the insertion torque values in various bone densities . dental implants with deeper thread depth showed higher insertion torques than those with shallower thread depth when having the same inner diameter in the same bone density ( table 2 , table 3 , table 4 ) . this means that the increase in the thread depth in dental implants with the same inner diameter provides better primary stability at lower bone densities . thread depth affects bone stresses and implant / abutment complex stability , and the effect of thread depth differs according to the bone density.21 therefore , it was thought that confirming the optimal thread depth at specific bone densities is necessary . the insertion torque is increased with increasing bone density.5 in our data , the insertion torque did not show a direct association with bone density . we used different drill diameters for different bone densities of the solid rigid polyurethane block . specifically , we used a drill with a larger diameter in higher bone density and a smaller diameter in lower bone density . we have demonstrated that ti implants with the same length and inner diameter have a similar maximum compressive strength . the mechanical strength is more related to the length and diameter than the thread depth . ti implants with deeper threads did not induce the breakage of threads applying the maximum compressive strength . dental implants may fracture at load levels below the maximum compressive strength of the implant / abutment complex . thus , the maximum compressive strength may suggest a standard point of acute overload . mechanical failures of dental implants appear through a repeated loading process at low loads.30 the fatigue test is a general method used in the laboratory to mimic actual intraoral use . the fatigue limits of the dental implants with a diameter of 4.0 mm and thread depth of 0.6 mm ( 636 n ) and those with a diameter of 4.0 mm and thread depth of 0.35 mm ( 619 n ) in the fatigue test on the basis of the international organization for standardization ( iso14801 ) were both more than 600 n ( data is not shown ) . the fatigue limit of the ti implants with deeper threads is similar to that of ti implants with shallow thread depth . our study indicated that the ti implants with the deeper threads have similar mechanical stability . this is the first study that attempted mechanical test by varying the thread depth in ti implants with the same implant body ( tapered implants ) . recently , we published our finding that the bone is formed until it is inside the deep threads ( root portion ) and that ti implants with deeper thread depth are osseointegrated in animal studies.31 we also have ongoing research on clinical application of ti implants with deeper thread depth in areas of poor quality bone in order to confirm their primary stability and survival rate . this study suggests that ti implants with deeper threads have greater insertion torque without a concomitant decrease in mechanical strength . implants with deeper thread depth may increase the primary stability in areas of poor quality bone without decreasing the mechanical strength . dental implants with primary stability increase the percentage of direct bone - to - implant contact ( bic),6 while dental implants without primary instability result in a lack of osseointegration , and ultimately implant failure.32,33 an increase in primary stability may increase biological stability through bone regeneration and remodeling between the surrounding bone and implant . dental implants with deeper thread depth may lead to successful osseointegration and decrease implant failure in areas of poor quality bone . | purposethis study aimed to evaluate the effect of implant thread depth on primary stability in low density bone.materials and methodsthe insertion torque was measured by inserting ti implants with different thread depths into solid rigid polyurethane blocks ( sawbones ) with three different bone densities ( 0.16 g / cm3 , 0.24 g / cm3 , and 0.32 g / cm3 ) .
the insertion torque value was evaluated with a surgical engine .
the static compressive strength was measured with a universal testing machine ( utm ) and the ti implants were aligned at 30 against the loading direction of the utm . after the static compressive strength test , the ti implants were analyzed with a measurescope.resultsthe ti implants with deeper thread depth showed statistically higher mean insertion torque values ( p<.001 ) .
groups a and group b had similar maximum static compressive strengths , as did groups c and d ( p>.05 ) .
after the static compressive strength , the thread shape of the ti implants with deeper thread depth did not show any breakage but did show deformation of the implant body and abutment.conclusionthe implants with deeper thread depth had higher mean insertion torque values but not lower compressive strength .
the deep threads had a mechanical stability .
implants with deeper thread depth may increase the primary stability in areas of poor quality bone without decreasing mechanical strength . |
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umbilical hernia occurs most frequently in middle - aged women , and is caused by a weakening of fibrous connective tissue in the umbilical ring secondary to obesity , frequent pregnancies or deliverance , ascites from liver cirrhosis or renal failure . the standard repair is an open umbilical hernioplasty by primary closure of the fascial defects ; however , the high recurrence rate associated with this procedure is somewhat problematic . recently , many cases of tension - free mesh repair for umbilical hernia have been reported [ 1 , 2 ] . here , we present a case of an incarcerated umbilical hernia treated by laparoscopic mesh repair . furthermore three years later , she was admitted to the emergency room at our hospital with incarceration of the umbilical hernia . the hernia had a size comparable to that of a tennis ball . her body mass index ( bmi ) was 38.3 kg / m , which is categorized as obese . computed tomography imaging showed an umbilical hernia with an incarcerated portion of small intestine and a hernial orifice of 2 cm ( fig . 1 ) . a gentle attempt at reducing the hernia was unsuccessful ; therefore , we decided to perform an emergency operation . after the induction of general anesthesia using a muscle relaxant agent , the hernia was reduced . a 12-mm trocar was inserted just below the epigastric region , and pneumoperitoneum was established by insufflation with carbon dioxide to a 10-mmhg abdominal pressure . two 5-mm trocars were placed in the right and left lateral abdominal region , respectively . on laparoscopic examination of the abdominal cavity , we identified the portion of incarcerated small intestine . it was reddish and congested ; however , there was no evidence of necrosis or perforation ( fig . the size of the umbilical hernial orifice was 2 2 cm ( fig . we selected a 12 12 cm composite mesh ( composix , davol , inc . warwick , ri , usa ) to cover the hernial defect by at least 5 cm in all directions . the composite mesh was inserted through the 12-mm trocar , and it was fixed to the abdominal wall circumferentially by a permanent fixation system ( permafix , davol , inc . , subsidiary of c. r. bard , inc . ; fig . figure 1:a computed tomographic scan showing the umbilical hernia and an incarcerated portion of the small intestine . figure 2:(a ) the strangulated portion of the small intestine was reddish and congested ; however , there was no evidence of necrosis or perforation ( white arrows ) . ( b ) the size of the umbilical hernial orifice was 2 2 cm . ( c ) composite mesh covering the hernial defect by at least 5 cm in all directions . the mesh was fixed to the abdominal wall circumferentially using a tacking instrument . a computed tomographic scan showing the umbilical hernia and an incarcerated portion of the small intestine . ( a ) the strangulated portion of the small intestine was reddish and congested ; however , there was no evidence of necrosis or perforation ( white arrows ) . ( b ) the size of the umbilical hernial orifice was 2 2 cm . ( c ) composite mesh covering the hernial defect by at least 5 cm in all directions . ohira et al . reported that the average age of patients with umbilical hernia was 59.2 years , and the percentage of female patients was 63.6% ( 21/33 ) , with an average bmi of 30.6 kg / m , which was much higher than the japanese standard bmi of 22 kg / m . they also reported that 12 ( 36.4% ) patients were excessively obese ( bmi > 30 kg / m ) and 10 ( 30.3% ) had liver cirrhosis accompanied with ascites . there is no argument that an operative procedure is the primary treatment for umbilical hernia ; the reduction of the incarcerated contents and closure of the hernial orifice are the basic principles of the surgery . open umbilical herniorrhaphy with the simple fascia suture technique has been widely used by many surgeons and is a long - standing procedure , because it is very simple and may sometimes be performed with local anesthesia . however , the recurrence rate is as high as 1020% [ 1 , 2 ] . tension - free repair with a mesh was introduced for umbilical or ventral hernia since the 1990s and evidence suggests that open mesh repair has significantly lowered the recurrence rates [ 1 , 2 ] . demonstrated a reduced recurrence rate of 1% for open mesh repairs of umbilical hernias , compared with 11% for primary suture repairs . recently , an increasing number of cases of umbilical hernia treated by a laparoscopic approach have been reported . gonzalez et al . reported that the laparoscopic repair group presented lower complication and recurrence rates and faster rehabilitation into society compared with the open mesh repair group . . demonstrated that laparoscopic mesh repair was associated with a significantly lower rate of postoperative wound infection compared with open mesh repair in obese patients . advantages of the laparoscopic procedure include measurement of the size of the hernial orifice , a more definite placement of the mesh and verification of the status of the strangulated intestine by a laparoscopic view . it is very important to determine the hernial orifice size , because the ideal coverage provides a 5-cm overlap in every direction from the defect [ 6 , 7 ] . because the size of the hernial orifice was 2 cm in the present case , we chose a 12 12 cm composite mesh to cover the defect and provide an 5-cm circumferential coverage . by open mesh repair , it may be difficult to ensure an adequate overlap and fixation of the mesh through the small incision ; however , it is not difficult to perform such a procedure using the tacking instrument by laparoscopy . it is also important to verify the conditions of the strangulated intestine because the gangrenous intestine must be resected . this can be easily confirmed by a laparoscopic view once the strangulated intestine is reduced into the abdominal cavity . many surgeons hesitate to use a mesh in cases of gangrenous strangulated intestines , because it may pose a potential risk for mesh infection . however , abd ellatif et al . divided patients with incarcerated hernia into two groups : those who underwent mesh hernioplasty with resection and anastomosis , and those who did not ; there was no significant difference between the two groups in terms of wound infection and recurrence rates , and no patients had to undergo reoperation to remove the mesh . considering these advantages , we suggest that laparoscopic mesh repair should be introduced more aggressively in cases of intestinal gangrene secondary to umbilical strangulated hernia . however , late - onset mesh infection should be taken account when using composite expanded polytetrafluoroethylene mesh . | a 42-year - old , obese woman was admitted to our hospital 3 h after the sudden development of abdominal pain .
her umbilical region was swollen and she was diagnosed with incarceration of an umbilical hernia by computed tomography .
although we tried , we were unable to reduce the hernia with a manipulative procedure .
we decided to perform an emergency laparoscopy .
once general anesthesia was induced , we achieved hernia reduction . from a laparoscopic view , the portion of strangulated small intestine was neither necrotic nor perforated .
the size of the hernial orifice was 2 2 cm , and thus , we selected a 12 12 cm composite mesh to cover the hernia defect by at least 5 cm in all directions .
the surgical procedure was uneventful and the total operation time was 112 min .
the patient recovered uneventfully and was discharged on postoperative day 9 .
she remains free of recurrence 20 months after surgery . |
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teaching is a complex process that involves the interweaving of content knowledge , pedagogy skills and knowledge and appreciation of the multi - faceted nature of students too , in the end , be able to point to evidence that learning has occurred . since the primary purpose of institutions of higher education is teaching , it becomes necessary to have a measure of teaching effectiveness . this can be carried out through a variety of ways such as student ratings , peer ratings , self - evaluation , videos , student interviews , alumni ratings , employer ratings , administrator ratings , learning outcome measures , and teaching portfolios . of the various procedures in place , student evaluation of teacher ( set ) effectiveness is a significant method for assessing the teaching effectiveness of college teachers . despite this , student evaluations of teaching performance is still a controversial tool in the improvement of teaching quality . fundamentally , student evaluations of teaching serve two purposes . from a formative ( or developmental ) perspective , the evaluations can be used to provide feedback to faculty in order to help them to improve their teaching or alter lesson content . evaluations may also serve summative ( or administrative ) purposes for organizational decisions regarding the faculty . the set process is typically implemented by asking students at the end of the academic semester or year to rate the teaching effectiveness of their instructor by anonymously completing a questionnaire . the items included in such evaluation forms refer to various dimensions of perceived teaching effectiveness ( e.g. , the ability of the instructor to communicate clearly ) , as well as characteristics of the taught educational content . the completed questionnaires are analyzed by designated entities within the university , and the results are returned to the department administration and the instructor for review . since student evaluation of faculty is an essential part of the academic process , the study was designed to compare with two formats of sets with a view to determine the method with minimum bias . the study aimed to compare the student ratings of teacher effectiveness obtained from two different set formats , determine different factors contributing to student bias , and established the preferred set format with minimal perceived bias . this research study was conducted at a medical college in the state of uttarakhand , after obtaining due permission and clearance from the institutional research and ethics committee . an ongoing method for student evaluation of teaching effectiveness using a marking questionnaire ( set - i format ) , was already in place . first - year medical students of the class of 2013 , who had recently cleared the first professional examinations ( n = 71 ) were assigned numbers 171 randomly . these students were administered the set - i format as per routine protocol of the institute . using a semi - structured questionnaire the perceptions and acceptability of the faculty regarding the set - i format were recorded . the types of bias , as identified by the responses of the faculty , were also determined from this feedback . taking these into consideration , as well as other suggestions given by the faculty , a new questionnaire based on a likert scale the set - ii format was also administered to the same mbbs students as before , and the rating obtained by the faculty from both formats was compared . the faculty was apprised of their obtained scores , and their acceptability of the preferred type of set format was determined with the help of another semi - structured questionnaire . on comparison of the faculty ratings between the set - i and set - ii formats , no significant difference was observed [ table 1 and figure 1 ] . mean faculty ratings by two formats comparison of student ratings from student evaluation of teacher-1 and student evaluation of teacher-2 the percentage satisfaction of the students with the individual faculty as an effective educator is depicted in figure 2 a comparison of the level of student satisfaction with teacher effectiveness with the total student ratings [ figure 1 ] , showed that with a decrease in total score of faculty below 50% , the level of student satisfaction reduced considerably . over 78% of the faculty 64% faculty felt that students would show positive bias toward senior faculty and those who had a number of classes . the opinion was equally divided for gender as a possible cause for bias [ figure 3 ] . furthermore , the analysis of the responses obtained from the second faculty questionnaire showed that although they were satisfied that student bias would be less if the set - ii format were used , they still felt that some further modification of the format was required . although student ratings should serve as feedback to the faculty for self - improvement , many of them view the process as a matter of concern . the study made an attempt to address these issues by changing the format of the set . . the limitations of the study were the small sample size and the fact that it was conducted only for first professional faculty . the research was conducted in order to get some preliminary results that could be used as a direction for future researches in this field . results from this study will be used for practical improvement of the existing questionnaires , as well as for working with students on their evaluation skills as participants in the appraisal of the educational process . | context : student evaluation of faculty is an essential part of the academic process . the study was designed to compare two formats of student evaluation of teachers ( sets ) with a view to determine the method with minimum bias.aims:to compare student ratings of teacher effectiveness obtained from two different set formats and determine factors contributing to the student bias.materials and methods : faculty members of first professional were evaluated by mbbs students using a set - i questionnaire already in use .
faculty perceived types of bias were documented using a separate semi - structured questionnaire . based on this ,
a second set - ii questionnaire with likert scale was designed and filled out by the same students as before .
the faculty was apprised of the scores granted to them , and their acceptance of the preferred set format was determined with the help of another questionnaire.results:ratings obtained from 71 students using both the set - i and set - ii formats showed no difference .
however , the level of students satisfaction with teacher effectiveness , compared with the total teacher score , indicated that when a score of the faculty was below 50% , the level of students satisfaction reduced considerably .
the major causes of perceived negative bias identified were strictness , seniority , gender , classes taken , less interest in the subject , and lower student grades .
set - ii was preferred by faculty but didnt eliminate all bias factors.conclusions:although it was not possible to remove all causes of bias from the modified student questionnaire , the faculty perception of bias affecting the students rating seems to be largely ungrounded as there was no difference between the scores obtained . |
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they have traditionally been considered benign , but recent evidence indicates that frequent apbs are a strong predictor of atrial fibrillation development and may be associated with increased risk of cardiovascular death in general population . in addition , frequent apbs may cause cardiomyopathy . her heart was structurally normal heart and there was no evidence of any sustained tachyarrhythmias . to our knowledge , this is the first report on successful ablation of frequent apbs in the non - coronary aortic cusp . a 59-year - old female with a long history of highly symptomatic frequent apbs presented to our hospital for catheter ablation . p wave was negative in inferior leads ii , iii , avf , positive in avr and v1v3 , indifferent in lead i , avl and v4v6 ( fig . 1 ) . apb burden in repeated holter recordings was 2040% ( 29000 apbs in the last holter recording ) . neither atrial fibrillation nor any other sustained supraventricular tachyarrhythmia was detected . clinical examination and thyroid function were normal . diagnostic catheters were placed in the coronary sinus and right ventricular apex vie right femoral vein . mapping and ablation was performed retrogradely via right femoral artery with a 3.5 mm tip open irrigated ablation catheter ( biosense webster navistar rmt , diamond bar , ca , usa ) using remote magnetic navigation ( epoch , stereotaxis inc . , st louis , mo , usa ) and 3d electroanatomical mapping system ( carto3 , biosense webster ) . mapping was started from the right atrium because of the p wave morphology and coronary sinus activation sequence . fast anatomical activation mapping demonstrated that the earliest activation in the right atrium was in the vicinity of the his bundle ( fig . 2 ) . no ablation attempt was made although local activation at this site preceded the p wave because of high risk of damage to the atrioventricular conduction system . meticulous mapping at the aortic root demonstrated earliest local activation of the clinical apb in the non - coronary sinus of valsalva anterior and superior to the his bundle . at this site no his potential was detected , and the local electrogram was earlier than in the para - hisian area in right atrium and preceded the p wave by about 50 ms ( fig . 3 ) . radiofrequency energy delivery at the non - coronary aortic cusp resulted in immediate cessation of the apbs . no junctional beats or pr prolongation were observed during ablation ( 120 s with maximum power of 35 w ) . total duration of the procedure was 110 min and fluoroscopy exposure 2 min 35 s ( 3.0 gy / cm2 ) , respectively . at discharge a routine holter recording was scheduled at 3 months and the patient was advised to contact her physician for ecg monitoring if any arrhythmia symptoms recurred . during the follow - up of 12 months the patient reported no recurrence of symptoms and there were no clinically relevant apbs in the holter recording . atrial premature beats are present in 1020% of the general population . in most cases they are benign , but frequent apbs have been associated with development of atrial fibrillation and cardiomyopathy as well as with increased risk of cardiovascular mortality and stroke , , . in our case no sustained atrial tachyarrhythmias were documented , and left ventricular function was normal despite long history of frequent apbs . nevertheless , it is possible that the high apb burden could have caused atrial fibrillation and/or apb - induced cardiomyopathy if not treated . the majority of apbs arise from within and around the pulmonary veins ( pv ) . other sites of origin include left atrial posterior wall , ligament of marshall , coronary sinus , superior and inferior vena cava , crista terminals and tricuspid annulus . catheter ablation has become the treatment of choice in patients with various paroxysmal supraventricular tachycardias and ventricular premature beats , but it is used quite rarely to eliminate isolated apbs . the frequently multiple sites of origin and capricious manifestation have made it difficult to ablate extrapulmonary abps . however , in patients with frequent monomorphic apbs the focus can usually be identified by careful activation mapping and treated by ablation . to the best of our knowledge this is the first report on successful ablation of frequent apbs in the non - coronary aortic cusp . based on our experience the main benefit of using magnetic navigation in cases like this is that mapping with a softer and more flexible magnetic catheter is less likely to cause perforation and to provoke catheter - induced extrasystoles than mapping with a manually steered ablation catheter . in addition , the magnetic technology has proven to reduce personnel and patient radiation exposure . it offers important safety benefit by reducing the risk of perforation but also makes creation of transmural lesions difficult in some areas ( e.g. , cavo - tricuspid isthmus ) . the p wave morphology and right atrial mapping demonstrated that the apbs originated from a site close to the his bundle . we and others have previously shown that sustained atrial tachycardia in this area can often be effectively and safely ablated within the non - coronary aortic cusp which is anatomically in close proximity to the interatrial septum and his bundle , . in the current case , the local atrial activation at the non - coronary aortic cusp was about 20 ms earlier than that recorded at the his region in the right atrium . at the ablation site no his potential was visible , and the catheter was stable and far from the coronary artery ostia . this case demonstrated that in addition to focal atrial tachyarrhythmias and ventricular premature beats aortic root can be a source for frequent apbs . due to the close anatomic relationship between the atrioventricular node and the non - coronary aortic cusp mapping of the aortic root and non - coronary cusp | a 59-year - old female with structurally normal heart was admitted to our hospital for treatment of highly symptomatic , drug refractory atrial premature beats ( apb ) .
ecg revealed atrial parasystolic trigeminy .
the arrhythmogenic focus was mapped and ablated using magnetic remote navigation and 3d electroanatomical mapping system . to our knowledge , this is the first report on successful ablation of frequent apbs in the non - coronary aortic cusp . |
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current approaches in pain management involve traditional non - steroidal anti - inflammatory drugs and opioid narcotics but neither is devoid of side - effects , emphasizing the need for new analgesic agents that would be efficacious but without the risk of tolerance , addiction or intolerable side - effects . piper betle is a shade - loving , perennial , dioecious , semi - woody , ever - green climber of tropical origin and is known as its pharmacological properties include anti - oxidant , anti - inflammatory and hepatoprotective effect , but to date its analgesic activity has not been validated . accordingly , this study aimed to establish the analgesic activity of piper betle leaf extract ( pbe ) and elucidate its putative mechanism of action . healthy adult male swiss albino mice ( 25 5 g ) were housed under standard environmental conditions ( 12 h light / dark cycle , temperature 20 - 25c , humidity 65 - 70% ) , with free access to drinking water and normal laboratory chow diet . all animal experiments were carried out as per protocol approved by the institutional animal ethics committee . the resultant pbe was dissolved in propylene glycol ( pg ) and stored at 4c ( 10 mg / ml ) ; for all experiments , pg served as the vehicle . healthy adult male swiss albino mice ( 25 5 g ) were housed under standard environmental conditions ( 12 h light / dark cycle , temperature 20 - 25c , humidity 65 - 70% ) , with free access to drinking water and normal laboratory chow diet . all animal experiments were carried out as per protocol approved by the institutional animal ethics committee . the resultant pbe was dissolved in propylene glycol ( pg ) and stored at 4c ( 10 mg / ml ) ; for all experiments , pg served as the vehicle . all other chemicals were obtained from sigma aldrich ( st . louis , missouri , usa ) except acetic acid , pg ( merck , mumbai , india ) and diclofenac sodium injection ( novartis , mumbai , india ) . for screening purposes , acetic acid ( 10 ml / kg b.w . , 0.8% v / v in h2o , i.p . ) was administered to mice wherein , the writhing response ( abdominal constriction , trunk twisting and extension of hind limbs ) was counted for 15 min and expressed as the pain response ; in addition , the onset of writhing was noted . animals showing a substantial pain response 60 writhing in 15 min were included to obtain a homogenous study population . the selected animals were divided into five groups of six animals each and administered pg ( 10 ml / kg b.w . , p.o . , group 1 ) , diclofenac ( 20 mg / kg b.w . , i.p . , group 2 ) , pbe ( 25 , 50 and 100 mg / kg b.w . the peritoneal exudate was obtained as previously described ; briefly , 10 ml of ice - cold , sterile phosphate buffered saline ( pbs , 0.02 m , ph 7.2 ) was injected i.p . , after which the peritoneal exudate was aspirated , centrifuged ( 1500 rpm 10 min ) ; the resultant cell pellet was washed twice and finally resuspended in pbs . intracellular ros was measured using the 2,7,-dichlorodihydrofluorescein diacetate ( h2dcfda ) , a non- fluorescent dye that fluoresces on reaction with free radicals . briefly , cells present in the murine peritoneal exudate ( 1 10/ml in pbs ) were initially preloaded with h2dcfda ( 20 m , 30 min , 37c ) . thereafter , cells were incubated with pbe ( 0 - 25 g / ml ) or pg ( at the concentration present in 25 g of pbe ) at 37c for 1 h followed by the addition of aa 1 m in dimethyl sulfoxide ( dmso ) , 5 min , 37c . cells were washed , resuspended in pbs and acquired in a flow cytometer ( facs calibur , becton dickinson , san jose , ca , usa ) , using fluorescence1 ( fl1 ) histogram to quantify fl of viable cells . the fl of dichlorofluorescein were collected in the fl1 channel , equipped with a 530/30 nm band pass flter . fl was measured in the log mode using cellquest pro software ( bd biosciences , san jose , ca , usa ) and expressed as geometrical mean fluorescence channel ( gmfc ) . cells were gated on the basis of their characteristic morphology using forward versus side scatter dot plot . subsequent analyses were done using bd cellquest pro software ( bd biosciences , ca , usa ) . dagostino and pearson omnibus normality test was used to assess the distribution of the data and multiple comparisons were carried out among groups by kruskal - wallis followed by the post - hoc dunn multiple comparison test or mann - whitney test . analysis was performed using graph pad prism software , version 5 ( graphpad software inc . , san diego , ca , usa ) ; p < 0.05 was considered statistically significant . . , 0.8% v / v in h2o , i.p . ) was administered to mice wherein , the writhing response ( abdominal constriction , trunk twisting and extension of hind limbs ) was counted for 15 min and expressed as the pain response ; in addition , the onset of writhing was noted . animals showing a substantial pain response 60 writhing in 15 min were included to obtain a homogenous study population . the selected animals were divided into five groups of six animals each and administered pg ( 10 ml / kg b.w . , p.o . , group 1 ) , diclofenac ( 20 mg / kg b.w . , i.p . , group 2 ) , pbe ( 25 , 50 and 100 mg / kg b.w . the peritoneal exudate was obtained as previously described ; briefly , 10 ml of ice - cold , sterile phosphate buffered saline ( pbs , 0.02 m , ph 7.2 ) was injected i.p . , after which the peritoneal exudate was aspirated , centrifuged ( 1500 rpm 10 min ) ; the resultant cell pellet was washed twice and finally resuspended in pbs . intracellular ros was measured using the 2,7,-dichlorodihydrofluorescein diacetate ( h2dcfda ) , a non- fluorescent dye that fluoresces on reaction with free radicals . briefly , cells present in the murine peritoneal exudate ( 1 10/ml in pbs ) were initially preloaded with h2dcfda ( 20 m , 30 min , 37c ) . thereafter , cells were incubated with pbe ( 0 - 25 g / ml ) or pg ( at the concentration present in 25 g of pbe ) at 37c for 1 h followed by the addition of aa 1 m in dimethyl sulfoxide ( dmso ) , 5 min , 37c . cells were washed , resuspended in pbs and acquired in a flow cytometer ( facs calibur , becton dickinson , san jose , ca , usa ) , using fluorescence1 ( fl1 ) histogram to quantify fl of viable cells . the fl of dichlorofluorescein were collected in the fl1 channel , equipped with a 530/30 nm band pass flter . fl was measured in the log mode using cellquest pro software ( bd biosciences , san jose , ca , usa ) and expressed as geometrical mean fluorescence channel ( gmfc ) . cells were gated on the basis of their characteristic morphology using forward versus side scatter dot plot . subsequent analyses were done using bd cellquest pro software ( bd biosciences , ca , usa ) . dagostino and pearson omnibus normality test was used to assess the distribution of the data and multiple comparisons were carried out among groups by kruskal - wallis followed by the post - hoc dunn multiple comparison test or mann - whitney test . analysis was performed using graph pad prism software , version 5 ( graphpad software inc . , san diego , ca , usa ) ; p < 0.05 was considered statistically significant . in this study , the peripheral analgesic effect was determined as the decrease in pain response induced by i.p . diclofenac served as the standard analgesic and significantly reduced the number of acetic acid induced writhing as compared with the vehicle control ( p < 0.001 ) . pbe ( 25 , 50 and 100 mg / kg ) also reduced the number of acetic acid induced writhing in a dose dependent manner being ( p < 0.01 ) and ( p < 0.001 ) as compared with vehicle control respectively [ figure 1a ] . moreover , the reduction in pain response with pbe ( 100 mg / kg ) was comparable with standard drug diclofenac , indicating its promising anti - nociceptive activity . diclofenac and pbe had no effect on the onset of the pain response [ table 1 ] , indicating that they did not delay nociceptive transmission , but reduced the intensity of pain . effect of piper betle leaf extract ( pbe ) on peripheral algesia ( a ) mice were pre - treated with vehicle 10 ml / kg b.w . ( 1 ) , diclofenac 20 mg / kg b.w . ( 2 ) , pbe at 25 mg / kg b.w . ( 3 ) , 50 mg / kg b.w . ( 4 ) or 100 mg / kg b.w . ( 5 ) followed by acetic acid ( 10 ml / kg b.w . , 0.8 % in h2o , i.p . ) and number of writhings per animal counted during a 15 min period as described in materials and methods . values are expressed in mean standard error of mean ; * p < 0.01 , * * p < 0.001 as compared to vehicle control effect of pbe on the onset of acetic acid induced writhing the effect of pbe on aa induced generation of ros was measured in murine peritoneal resident cells wherein , a respiratory burst was induced by aa as evidenced by a significant 1.90 fold increase in gmfc from baseline ( p < 0.01 ) . this was effectively attenuated when cells were pre - incubated with pbe ( 2.5 , 10 and 25 g / ml , 1 h ) thereby translating into a decrease in gmfc , being ( p < 0.01 ) [ figure 1b ] . pg did not influence generation of ros and similarly , dmso representative of the concentration present in aa ( 1 m ) had no effect on generation of ros , confirming their biological inertness ( data not shown ) . effect of piper betle leaf extract ( pbe ) on generation of reactive oxygen species ( ros ) : murine peritoneal exudate ( 1 106 cells / ml pbs ) were incubated with or without arachidonic acid ( aa ) ( 1 m ) in the presence or absence of pbe ( 2.5 , 10 and 25 g / ml ) for 1 h and assayed for generation of ros as described in materials and methods . values are expressed as geometrical mean fluorescence channel , mean standard error of mean ; * p < 0.01 when compared with control , * * p < 0.01 compared with aa treated group . inset : a representative histogram profile of murine peritoneal ( 5 105 cells , 1 ) incubated with dichlorodihydrofluorescein diacetate in the presence of aa ( 1 m , 2 ) along with pbe ( 25 g / ml , 3 ) and assayed for generation of ros in this study , the peripheral analgesic effect was determined as the decrease in pain response induced by i.p . diclofenac served as the standard analgesic and significantly reduced the number of acetic acid induced writhing as compared with the vehicle control ( p < 0.001 ) . pbe ( 25 , 50 and 100 mg / kg ) also reduced the number of acetic acid induced writhing in a dose dependent manner being ( p < 0.01 ) and ( p < 0.001 ) as compared with vehicle control respectively [ figure 1a ] . moreover , the reduction in pain response with pbe ( 100 mg / kg ) was comparable with standard drug diclofenac , indicating its promising anti - nociceptive activity . diclofenac and pbe had no effect on the onset of the pain response [ table 1 ] , indicating that they did not delay nociceptive transmission , but reduced the intensity of pain . effect of piper betle leaf extract ( pbe ) on peripheral algesia ( a ) mice were pre - treated with vehicle 10 ml / kg b.w . ( 1 ) , diclofenac 20 mg / kg b.w . ( 2 ) , pbe at 25 mg / kg b.w . ( 3 ) , 50 mg / kg b.w . ( 4 ) or 100 mg / kg b.w . ( 5 ) followed by acetic acid ( 10 ml / kg b.w . , 0.8 % in h2o , i.p . ) and number of writhings per animal counted during a 15 min period as described in materials and methods . values are expressed in mean standard error of mean ; * p < 0.01 , * * p < 0.001 as compared to vehicle control effect of pbe on the onset of acetic acid induced writhing the effect of pbe on aa induced generation of ros was measured in murine peritoneal resident cells wherein , a respiratory burst was induced by aa as evidenced by a significant 1.90 fold increase in gmfc from baseline ( p < 0.01 ) . this was effectively attenuated when cells were pre - incubated with pbe ( 2.5 , 10 and 25 g / ml , 1 h ) thereby translating into a decrease in gmfc , being ( p < 0.01 ) [ figure 1b ] . pg did not influence generation of ros and similarly , dmso representative of the concentration present in aa ( 1 m ) had no effect on generation of ros , confirming their biological inertness ( data not shown ) . effect of piper betle leaf extract ( pbe ) on generation of reactive oxygen species ( ros ) : murine peritoneal exudate ( 1 106 cells / ml pbs ) were incubated with or without arachidonic acid ( aa ) ( 1 m ) in the presence or absence of pbe ( 2.5 , 10 and 25 g / ml ) for 1 h and assayed for generation of ros as described in materials and methods . values are expressed as geometrical mean fluorescence channel , mean standard error of mean ; * p < 0.01 when compared with control , * * p < 0.01 compared with aa treated group . inset : a representative histogram profile of murine peritoneal ( 5 105 cells , 1 ) incubated with dichlorodihydrofluorescein diacetate in the presence of aa ( 1 m , 2 ) along with pbe ( 25 g / ml , 3 ) and assayed for generation of ros the anti - inflammatory and anti - oxidant activity of pbe or its phytoconstituent allylpyrocatechol has been established in previous studies . the central anti - nociceptive activity of pbe has been shown to be mediated through opioid receptors . the experiments described in this communication are to validate its peripheral analgesic activity along with elucidating the underlying mechanism of action ; thus , paving the way for its use as a promising herbal extract . acetic acid induced writhing was chosen for evaluating peripheral analgesic activity as it established model of peripheral algesia . pbe , by decreasing number of acetic acid induced writhes in mice clearly demonstrated its peripheral analgesic activity [ figure 1a ] . moreover , the reduction in pain response with pbe ( 100 mg / kg ) was comparable with standard drug diclofenac , indicating its promising anti - nociceptive activity . however , neither diclofenac nor pbe had any effect on the onset of pain , indicating that they did not delay nociceptive transmission , but reduced the intensity of pain [ table 1 ] . acetic acid induced stretching is proposed to occur following the release of endogenous mediators of pain , such as prostaglandins e2(pge2 ) and pge2 from peritoneal resident macrophages and mast cells . this release of pge2 is directly dependent on generation of ros , a byproduct of metabolism of aa . as allylpyrocatechol , a phytoconstituent of pbe has been reported to be an effective anti - oxidant through its ability to scavenge ros , it can be proposed that pbe could influence the aa cascade which might account for its analgesic activity . therefore , this study evaluated the effect of pbe on aa induced ros generation in the cellular components of the peritoneal exudate [ figure 1b ] . further studies are needed to delineate the mechanisms of anti - nociceptive action of pbe in vivo by estimating levels of pge2 , expression of cox , generation of peroxynitrite and other pro - inflammatory markers in the peritoneal exudate . | objectives : the objective of this study was to evaluate the peripheral analgesic effect of piper betle leaf extract ( pbe ) along with establishing its putative mechanism of action.materials and methods : male swiss albino mice after pre - treatment ( 1 h ) with different doses of pbe were injected 0.8% ( v / v ) acetic acid i.p . ; the onset and number of writhes were noted up to 15 min . to evaluate the mechanism of action , the murine peritoneal exudate was incubated with pbe for
1 h , followed by exposure to arachidonic acid ( aa ) and generation of reactive oxygen species ( ros ) was measured by flow cytometry using 2,7-dichlorodihydrofluorescein diacetate.results:pbe in a dose dependent manner significantly reduced acetic acid induced writhing response in mice ( p < 0.001 ) . in peritoneal exudates ,
pbe significantly inhibited aa induced generation of ros , p < 0.01.conclusions:the present study indicates that pbe has promising analgesic activity , worthy of future pharmacological consideration . |
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vitus dance and chorea minor , is a rare but impressive clinical manifestation following streptococcal infection . this movement disorder characterised by chorea , emotional lability and muscle weakness is one of the major criteria of acute rheumatic fever ( arf ) . chorea is observed in 1836% of all patients with arf [ 3 , 25 , 29 ] . the incidence of arf and sc reflects the adequacy of preventive medical care in a community and has declined dramatically in the western world [ 9 , 20 ] . however , chorea is still a common manifestation of arf in developing countries , and a number of recent outbreaks in the usa indicate the need for awareness of this disease [ 3 , 25 ] . complaints can last for months to years , and 2042% of the patients have a relapse [ 8 , 16 ] . curative treatment is still in the experimental stage [ 4 , 7 , 30 ] . in this paper , we describe two patients with sc and severe disabilities in daily life . based upon the experience with them , we advocate the use of intravenous immunoglobulins ( ivig ) as a treatment option for sc in view of the latest insights in the pathophysiology of sc . patient number 1 , an 11-year - old girl , presented at the adult outpatient neurology department with involuntary movements since several days , which she described as my arms hit and my legs kick while i do not want them to do so . , we saw a girl with a figure fitting her age showing continual irregular , uncontrolled movements of the upper and lower extremities and to some extend also of the head and trunk . her speech was incomprehensive and , at rest , she made sucking and swallowing movements . coordination was intact , illustrated by the ability to catch a ball despite her chaotic movements . laboratory findings showed a normal complete blood cell count , erythrocyte sedimentation rate ( esr ) of 6 mm / h ( 125 mm / h ) and an antistreptolysin o titre ( asot ) of 8001,600 iu / ml ( < 250 iu / ml ) . rheumatoid factor and anticardiolipin antibody were negative . in the throat culture , performed several weeks after presentation , no pathogens were isolated . computed tomography of the brain did not reveal any anatomical anomaly . concerning the sc , a policy of wait and see was followed because of the well - known high number of patients who recover spontaneously . however , after 2 months , her symptoms had worsened , and she became wheelchair bound . subsequently , she was referred to us . based upon analyses of recent literature ( see below ) , we treated her with ivig : 400 mg / kg per day for 5 days . treatment was tolerated well and had a pronounced positive effect on clinical symptoms . within several days , her mobility increased , and she was no longer wheelchair bound . within 1 week , the sc disappeared fully . a prophylactic regimen of penicillin was administered , and in a follow - up period of 2 years , no relapse of the disease occurred . patient number 2 , a 13-year - old girl , was referred to us by her physiotherapist because of dysarthria , ataxia and balance problems . she had severe balance difficulties , made involuntary movements with her mouth and tongue and suffered from general muscle weakness . a month before , she had been diagnosed with pneumonia for which her general practitioner had treated her with amoxicillin / clavulanate . on physical examination , the patient made involuntary oral movements smacking her lips and moving her tongue in and out of her mouth . her reflexes were normal , but her coordination was slightly abnormal with a deviant tandem walk . internal examination revealed a holosystolic cardiac murmur grade 2/6 with punctum maximum at the apex . additional laboratory research showed a normal blood cell count , esr of 29 mm / h and an asot of > 1,600 iu / ml . in a throat culture , a group a haemolytic streptococcus was found . the patient was given ivig similarly to patient a. after administering the drug , the complaints improved substantially . the same prophylactic penicillin regimen was used , and for the corvitium , carbasalate calcium was prescribed . fatigue persisted for some weeks . until now , 2 years later , this girl also did not have any relapse . the relation between sc and a previous group a streptococcal ( gas ) infection has been established in 1956 . in 1976 , husby et al . discovered in serum of sc patients that antibodies directed against part of the gas bacterium cross - react with neurons of the caudate nucleus and basal ganglia , a mechanism called molecular mimicry . such a pathophysiologic mechanism is also present in other diseases , for example , in guillain - barr syndrome , where antibodies cross - react with the anti - ganglioside ab on motor neurones and in idiopathic thrombocytopenic purpura ( itp ) with glycoprotein iibiiia on platelets [ 21 , 26 ] . n - acetyl - beta - d - glucosamine ( glcnac ) is a structural component of the streptococcal cell wall and is the epitope that actually provokes the antibodies that play a role in some manifestations of arf [ 15 , 17 ] . several studies show that glcnac cross - reacts with glycoconjugates of different molecular composition , for example , glycoproteins on cardiac valve surface in rheumatic heart disease . a second molecule called m protein , the major virulence factor of gas , turned out to be cross - reactive with various host -helical proteins [ 6 , 15 ] . bronze and dale continued to elucidate the pathogenesis of sc by discovering anti - basal ganglia antibodies ( abga ) . they demonstrated that rabbits immunised with the group a m protein produced antiserum that was cross - reactive with central nerve system antigens in western blots . reactivity to those specific neuronal antigens was inhibited by pre - incubation with gas carbohydrate . the specific neuronal target antigens in sc were only recently discovered by kirvan et al . . they studied human antibodies derived from serum of an sc patient and found cross - reactivity with intracellular brain protein tubulin and extracellular lysoganglioside . by binding to those proteins , a signalling cascade was triggered that may cause the neurological manifestations of sc by releasing dopamine to the synapse . textbooks suggest symptomatic treatment for sc with benzodiazepine , anti - epileptics , phenothiazines or neuroleptics . these drugs are central nerve system depressants and control the main neurological signs of sc . the pathophysiology of cross - reactive antibodies in sc implies new treatment options supported by experiences with immune and inflammatory disorders described above . indeed , immune modulating therapy , like corticosteroids or immunoglobulin infusion , is found to improve sc . there is only one randomised double blind study in which 22 children suffering from sc were treated with 2 mg / kg / day prednisone during 4 weeks followed by a gradual discontinuation . on evaluation in weeks 8 and 12 , these patients showed a significant reduction in chorea intensity compared with a 15 patient placebo group . several other studies ( case reports or retrospective cohorts ) demonstrate that prednisone improves the course of the disease [ 1 , 5 , 28 ] . however , many of these reports note a rapid relapse of symptoms or the development of important side effects like cushing 's syndrome and hypertension . in the past decade , ivig have been increasingly used in the treatment of diseases caused by auto - antibodies [ 13 , 31 ] . the effect is based on the hypothesis that pooled ivig may result in the saturation of fc receptors on phagocytes . through competition , they prevent pathogenic auto - antibodies to bind to these cells and prevent immune activation . the beneficial effect of ivig has been shown in several diseases with molecular mimicry as underlying pathophysiology . marked improvement with this regimen is made in guillain - barr syndrome , chronic inflammatory demyelinating polyneuropathy and itp [ 13 , 31 ] . doses vary from 0.4 g / kg / day for 37 days in guillain - barr to 1 g / kg in single dose in itp . when scrutinising the literature , one randomised controlled clinical trial was found . eighteen subjects were entered into this trial , which was designed to determine if ivig or plasma exchange would be superior to prednisone in decreasing severity of chorea . four patients were randomised to receive ivig , eight received plasma exchange and six , prednisone . mean chorea severity was significantly lower for the entire group at the 1-month follow - up evaluation , with an overall improvement in half of them . nevertheless , clinical signs improved more rapidly and robustly in the ivig and plasma exchange group than in the prednisone group . therapeutic treatment options are still in experimental stage . based on pathophysiology and experiences in comparable diseases this double case report on two sc patients indicates that ivig is an effective treatment , although larger studies are needed to confirm this conclusion . | this double case report indicates that treatment with intravenous immunoglobulins ( ivig ) is effective in patients with sydenham chorea ( sc ) .
sc is a rare but impressive clinical manifestation following streptococcal infection .
this movement disorder characterised by chorea , emotional lability and muscle weakness , is one of the major criteria of acute rheumatic fever .
treatment of sc is typically limited to supportive care and palliative medications .
curative treatment is still in the experimental stage .
recent research on patients with sc proved that antibodies against the group a streptococcus cross - react with epitopes of neurons in the basal ganglia , namely , intracellular tubulin and extracellular lysoganglioside .
therefore , immune modulating therapy by means of prednisone , plasma exchange and ivig are mentioned in the literature as possible effective treatment
. beneficial effect of ivig has been shown in several diseases with molecular mimicry as the underlying pathophysiology . in this paper , we describe two girls aged 11 and 13 years , respectively , who presented with sc having severe disabilities in their daily live .
we treated both patients with ivig 400 mg / kg / day for 5 days .
treatment was tolerated well and had a pronounced positive effect . shortly after the drug was administered , all signs and symptoms disappeared in both patients .
based upon these patients , we highlight ivig as a serious treatment option for sc . |
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glaucoma is one of the major leading causes of blindness worldwide.1 in japan , glaucoma has become the primary cause of blindness since 2004.2 normal - tension glaucoma ( ntg ) is the most common type of glaucoma in japan,3 with a prevalence rate of 3.6% in people over 40 years of age . the ophthalmic solutions 0.005% latanoprost ( lat)4,5 and 0.0015% tafluprost ( taf)6,7 are topical prostaglandin analogs . both solutions greatly reduce intraocular pressure ( iop ) by increasing the uveoscleral outflow as a prostaglandin f receptor agonist.8 lat was first approved for clinical use in 1996 and has been used globally since then as a first - line drug for the treatment of glaucoma . on the other hand , the iop reduction efficacy and safety of taf , first launched in 2008 , is reportedly equivalent to that of lat9 for patients with primary open - angle glaucoma or ocular hypertension . although there have been reports on the efficacy of taf for ntg patients,10,11 to the best of our knowledge , no comparison crossover studies have been conducted to investigate the efficacy and safety between lat and taf for ntg patients . in this prospective crossover study involving japanese ntg patients , all study procedures were approved by the institutional review board of kyoto prefectural university of medicine , kyoto , japan , and were performed in accordance with the tenets set forth in the declaration of helsinki . we selected and enrolled 30 japanese ntg patients who had used lat monotherapy at least 4 weeks at the glaucoma special clinic of kyoto prefectural university of medicine . written informed consent was obtained from all patients after receiving a detailed explanation of the nature and possible consequences of their participation in the study . each of the 30 enrolled patients were randomly divided into one of the following two groups : 1 ) lat - to - taf group ( lt group : 15 patients , 7 male eyes and 8 female eyes ; mean age : 63.511.5 years ) and 2 ) taf - to - lat group ( tl group : 15 patients , 8 male eyes and 7 female eyes ; mean age : 69.18.8 years ) . at the beginning of the study , both groups were switched from initial lat to lat or taf for 12 weeks , and then switched over to the other drug ( crossover ) to use for 12 additional weeks ( figure 1 ) . at 0 , 4 , 12 , 16 , and 24 weeks , we evaluated their iop , conjunctival injection score ( grades 03),12 and corneal epitheliopathy score ( area density [ ad ] score).13 then , at 0 , 12 , and 24 weeks , we evaluated adverse events of eyelash changes and pigmentation of the eyelids and irises via a slit - lamp photograph taken at each visit . the incidence of eyelash and pigmentation was judged via slit - lamp photographs , with a comparison between the current and baseline photographs . in all patients , iop was measured by using an applanation tonometer ( h03 r 90030515 ; haag - streit , bern , switzerland ) by the same glaucoma specialist throughout the protocol periods , and those iop measurements were obtained at approximately the same time of day for each patient . if data were available from both eyes , then the right - eye data were used . we evaluated the mean deviation of the humphrey 30 - 2 threshold of static visual field program at baseline or within 3 months . statistical analysis was performed using the student s t - test and fisher s exact test . for each patient , lat and taf eye drops were prescribed at the glaucoma special clinic . as this was not a masked study , a calculated sample size of 15 enrolled subjects per arm providing the power of 80% was based on a noninferiority limit of 1.5 mmhg , a standard deviation of 1.5 mmhg for change in iop . there were no dropout cases , and all patients completed the study . of the total 30 subjects , 28 were newly prescribed lat , yet 1 subject in both the lt and tl groups used lat for more than 4 weeks . there were no significant differences in age , sex ratio , or mean deviation obtained from humphrey perimetry sita standard 30 - 2 between the lt and tl groups . as for the iop time course of all subjects , at 0 , 12 , and 24 weeks , respectively , the mean iop of the lt group ( 15 eyes ) was 10.5 , 10.6 , and 11.1 mmhg , whereas that of the tl group ( 15 eyes ) was 11.7 , 11.1 , and 10.5 mmhg , and no significant differences were found between the two groups ( student s t - test ) ( figure 2 ) . at 0 , 4 , and 12 weeks , respectively , the mean iop of the lat intragroup ( 30 eyes ) was 10.8 , 10.6 , and 10.5 mmhg , whereas that of the taf intragroup ( 30 eyes ) was 10.8 , 10.7 , and 11.1 mmhg , and no significant differences were found between the two intra - groups ( student s t - test ) ( figure 3 ) . the mean conjunctival hyperemia score at 0 , 4 , 12 , 16 , and 24 weeks , respectively , was 1.00.4 , 1.00.5 , 0.90.3 , 0.90.4 , and 0.90.4 in the lt group and 1.00.4 , 1.10.5 , 1.10.6 , 1.00.5 , and 1.10.5 in the tl group , and no significant differences were found between two groups ( student s t - test ) ( figure 4 ) . the respective mean ad score at 0 , 4 , 12 , 16 , and 24 weeks was 0.50.9 , 1.01.4 , 0.71.0 , 0.91.4 , and 0.71.0 in the lt group and 1.01.3 , 0.71.0 , 0.71.0 , 0.51.0 , and 0.81.3 in the tl group , and no significant differences were found between the two groups ( student s t - test ) ( figure 5 ) . the rates of the adverse events of eyelash change and eyelid and iris pigmentation induced by lat or taf are shown in table 2 . the incidence of eyes showing increased eyelash length was 30.0% by lat and 26.7% by taf , and the incidence of eyes showing increased eyelash amount was 16.7% by lat and 30.0% by taf . the incidence of eyes showing increased eyelid pigmentation was 3.3% by lat and 20.0% by taf , and the incidence of eyes showing increased iris pigmentation was 6.7% by lat and 3.3% by taf . there was no significant difference in the incidence of eyelash change ( length and amount ) as well as eyelid and iris pigmentation between the two groups ( fisher s exact test ) . although there have been reports on the efficacy of taf for ntg patients , to the best of our knowledge , this present study is the first crossover prospective report to investigate and compare iop reduction efficacy and safety between lat and taf in ntg patients . in this study , no significant differences were found between lat and taf in regard to iop reduction effects , conjunctival hyperemia score , and corneal epitheliopathy score , as well as eyelash change and eyelid and iris pigmentation . it has been reported that conjunctival hyperemia becomes stronger when switching from lat to travoprost or bimatoprost,14,15 and the same tendency was found to be true in this present study when switching from lat to taf . although there were no statistically significant differences , elongation of eyelash length was more frequently found in lat , whereas increased eyelash amount was more frequently found in taf . since the patients involved in this study were already users of lat for over 1 month , it was difficult to make a simple comparison between the two drugs in regard to the adverse events of newly appearing eyelid and iris pigmentation or eyelash elongation . in this study , there was a tendency of more eyelid pigmentation with taf than with lat . according to past reports for eyelash length , eyelash amount , eyelid pigmentation , and iris pigmentation , the appearance rate was 025.8/0.38% , 26/46% , 1.56.0/1.08%4% , and 012.3/4.0% with lat / taf,7,13,14,1621 respectively . our results were nearly the same as those of the previous reports , except for iris and eyelid pigmentation . our findings showed that eyelid pigmentation was greater with taf than with lat , yet with no significant difference . the first limitation was that this was not a masked study , as both the subjects and the attending doctor knew which eye drop was being used . the second limitation was that since the baseline iop of all subjects using lat was low , it was difficult to find the switching efficacy of the eye drop . the findings of this study show that lat and taf have equivalent safety and efficacy in japanese ntg patients with low iop . | purposeto evaluate and compare the intraocular pressure ( iop ) reduction efficacy and safety between the ophthalmic solutions 0.005% latanoprost ( lat ) and 0.0015% tafluprost ( taf ) in japanese patients with normal - tension glaucoma ( ntg).methodsin this randomized nonmasked study , we prospectively enrolled 30 japanese ntg patients who had used lat monotherapy for more than 4 weeks , and randomly divided them into the following two groups : 1 ) lat - to - taf group ( lt group ) and 2 ) taf - to - lat group ( tl group ) . at the beginning of the study , both groups were switched from initial lat to lat or taf for 12 weeks , and then switched over to the other drug ( crossover ) for 12 additional weeks . at 0 , 4 , 12 , 16 , and 24 weeks
, we evaluated each patient s iop , conjunctival injection , and corneal epitheliopathy score , and at 0 , 12 , and 24 weeks , we evaluated their eyelash changes and pigmentation of the eyelids and irises.resultsthe mean iop of the lt group ( 15 eyes ) was 10.5 , 10.6 , and 11.1 mmhg , at 0 , 12 , and 24 weeks , respectively , whereas that of the tl group ( 15 eyes ) was 11.7 , 11.1 , and 10.5 mmhg at 0 , 12 , and 24 weeks , respectively .
no significant differences were found between the two groups and in the intragroup comparisons .
moreover , no significant differences were found between lat and taf in regard to the conjunctival injection score and corneal epitheliopathy score .
eyelash changes and eyelid and iris pigmentation were similar in both groups.conclusionthe findings of this study show that lat and taf have equivalent efficacy and safety in japanese patients with ntg . |
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the classical interleukin ( il)-1 family cytokines , il-1 and il-1 , as well as il-18 , are related by mechanism of origin , receptor structure , and signal transduction pathways utilized . il-33 ( also named il-1f11 ) , which belongs to il-1 family , is most closely related in structure to il-18 and il-1 . il-33 is also described as a nuclear factor expressed in high endothelial venules ( nf - hev ) . the human il-33 gene is located on chromosome 9p24.1 , while its mouse counterpart can be found on the syntenic chromosome 19qc1 region . the il-33 cdna sequences encode 270 and 266 amino acid polypeptides for human and mouse , respectively , corresponding to full - length proteins with calculated masses of 30 and 29.9 kda . the proteins do not contain a signal sequence for secretion , similar to the il-1 , il-1 , and il-18 genes [ 4 , 5 ] . the precursor protein ( pre - il-33 ) was cleaved by caspase-1 into a mature 18-kda protein ( il-33 ) in in vitro experiments using a recombinant protein . il-33 signals via the il-1 receptor - related protein st2 [ 2 , 6 ] and il-1 receptor accessory protein activate nf-b and mitogen - activated protein kinases to induce t helper type 2 ( th2)-associated cytokines [ 2 , 8 , 9 ] . administration of il-33 to mice induces production of th2 cytokines and severe pathological changes in mucosal organs . moreover , the ability of il-33 to induce th2 responses has functional relevance in the context of intestinal helminth infection . il-33 can promote survival , adhesion , and cytokine production in human mast cells , but the mechanism behind its involvement in inflammatory conditions remains unclear . it is also a key mediator of immune inflammatory hypernociception normally associated with a th1 type of response , revealing a hitherto unrecognized function of il-33 in a key immune pharmacological pathway that may be amenable to therapeutic intervention . in this study , the cloning and expression of mouse il-33 gene in escherichia coli ( e. coli ) , purification of recombinant protein , and generation of polyclonal antibody against il-33 are described . the prepared antibody can be useful for the study of expression and distribution of il-33 in various tissues at protein level and for the elucidation of its biofunctions and regulation mechanism in inflammatory diseases . e. coli strain bl21 ( de3 ) and ni - nta his - bind resins were obtained from novagen ( madison , wi ) . hotstar taq dna polymerase and qiaquick gel extraction kit were purchased from qiagen ( valencia , ca ) . primescript first strand cdna synthesis kit , restriction endonucleases xhoi , ndei , t4 dna ligase , isopropyl--d - thiogalactoside ( iptg ) , dna marker d2000 , and protein marker were from takara ( dalian , china ) . lysozyme , complete freund 's adjuvant ( cfa ) , and incomplete freund 's adjuvant ( ifa ) were obtained from sigma ( st . louis , mo ) . total cellular rna was extracted from spinal cord of balb / c mouse according to trizol reagent instructions . the coding region of mouse il-33 mature protein was amplified by reverse transcription polymerase chain reaction ( rt - pcr ) . the first strand of cdna was synthesized using the primescript first strand cdna synthesis kit , and the pcr reactions were carried out with a ptc-200 peltier thermal cycler ( mj research , waltham , ma ) . primer 1 ( 5-ggaattccatatg acattgagcatccaaggaac-3 ) and primer 2 ( 5-ccgctcgaggattttcgagagcttaaaca-3 ) were synthesized by invitrogen ( shanghai , china ) . primer 1 was synthesized with an ndei site ( shown in bold ) and an additional atg for initiation of translation in italicized style . the amplification profile included one initial hot - start denaturation step at 94c for 15 minutes , followed by 30 cycles of the following conditions : 94c for 30 seconds , 57c for 30 seconds , and 72c for 1 minute , and a final extension at 72c for 10 minutes . plasmid pet-44 ( novagen , darmstadt , germany ) was digested with ndei and xhoi restriction enzymes . the large fragment was cut and purified from agarose gel using the qiaquick gel extraction kit . a recombinant plasmid was constructed by inserting the amplified fragments ( also digested with ndei and xhoi ) into the ndei and xhoi sites of pet-44 and transformed into e. coli strain bl21 ( de3 ) . the transformants ( pet-44-mil-33 ) were confirmed by colony pcr , restriction enzyme digestion , and dna sequencing . expression in bacteria results in the producing of recombinant il-33 protein containing a hexahistidine tag in the c - terminus . a single colony of successfully transformed e. coli was picked up and cultured overnight at 37c in luria bertani ( lb ) medium , supplemented with 100 g / ml ampicillin . the culture mixture was then inoculated to fresh lb medium ( 1 : 100 dilution ) containing ampicillin and grown at 37c under continuous shaking , until the absorbance at 600 nm reached 0.60.8 . to optimize the expression conditions , il-33 expression was induced by adding 1 mm iptg to the transformed e. coli and the bacteria were incubated at 25c for a period of 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , and 9 hours , respectively . the degree of expression was evaluated by sodium dodecyl sulfate - polyacrylamide gel electrophoresis ( sds - page ) . to compare the protein dissolubility , the bacteria with pet-44-mil-33 after 7 hours of induction at 25c , the cells were harvested by centrifugation at 8,000 g for 20 minutes at 4c . the supernatant was discarded and the pellet was resuspended in lysis buffer consisting of 300 mm nacl , 50 mm nah2po4 , 10 mm imidazole , ph 8.0 , and lysed by using lysozyme , and incubated for 30 minutes at room temperature ( rt ) with gentle shaking to assure thorough cell lysis . the cell lysate was further disrupted by sonication on ice with six 10-second pulses at high intensity with a 10 seconds cooling period between each burst . the suspension was centrifuged at 12,000 g for 30 minutes to remove insoluble debris . the supernatant was added to an ni - nta spin column pre - equilibrated with the lysis buffer , and then they were gently shaked on ice for 60 minutes . the column was then washed with native wash buffer ( 300 mm nacl , 50 mm nah2po4 , 20 mm imidazole , ph 8.0 ) . finally , the bound protein was eluted with native elution buffer ( 300 mm nacl , 50 mm nah2po4 , 250 mm imidazole , ph 8.0 ) and analyzed with 12% sds - page . fractions were desalted and concentrated by an ultrafiltration device ( millipore corporation , bedford , ma ) . the new zealand white rabbits were given intradermal injections of il-33 protein ( 500 g / rabbit ) mixed with cfa in 1 : 1 ratio . after 2 week , the rabbits were boosted subsequently 4 times with protein ( 200 g / rabbit ) mixed with ifa in 1 : 1 ratio at 2-weeks interval . before every immunization , blood samples were taken from the marginal vein of the rabbit ear , centrifuged , and the sera were obtained to determine the antibody titer by enzyme - linked immunosorbent assay ( elisa ) . one week after the last injection , antisera of the rabbits were collected and purified by saturated ammonium sulfate ( sas ) and then protein a sepharose affinity chromatography ( amersham biosciences , piscataway , nj ) following manufacturer 's instructions . the titer of antiserum was determined by an indirect elisa . for each well of the 96-well elisa plate , 200 ng of il-33 protein was diluted in 100 l of sodium bicarbonate solution and incubated overnight at 4c . after three washes with phosphate - buffered saline-(pbs- ) tween buffer ( 0.05% tween 20 in pbs ) , the wells were blocked using 100 l of 3% bovine serum albumin for 1 hour at 37c and then incubated with 100 l polyclonal antibodies against il-33 with different dilution ( from 1 : 100 to 1 : 64,000 ) for 1.5 hours . the wells were incubated with 50 l / well horseradish peroxidase-(hrp- ) conjugated goat antirabbit immunoglobulin ( igg , 1 : 250 dilution ) for 1 hour at 37c after thoroughly washed . after final washes , 100 l / well tetramethyl benzidine ( tmb ) solution was added for color development . after 10 minutes , the reaction was stopped by 100 l / well 2 m h2so4 and the absorbance was measured at 450 nm using a plate reader ( molecular devices ) . ratio of antiserum versus control serum absorbency greater than 2.1 was recognized as positive . in order to confirm that the polyclonal antibodies are able to recognize il-33 protein from tissues , we also examined the il-33 protein expression of plasmid pcdna3.1-mil-33 after delivery through hydrodynamics - based gene injection in balb / c mice [ 14 , 15 ] . the plasmid pcdna3.1-mil-33 was constructed as forenamed , but whole mouse il-33 cdna was amplified by pcr and then inserted into mammalian expression plasmid pcdna3.1(+ ) ( invitrogen ) . briefly , slides were dried in an oven ( 60c ) before removing paraffin in several changes of xylene . slides were hydrated through a series of graded alcohols to water , followed by incubation with 0.5% hydrogen peroxide for 5 minutes . sections were incubated with 5% bovine serum albumin ( bsa ) in pbs for 10 minutes at rt to block nonspecific binding . after rinsing with pbs , slides were incubated overnight at 4c with rabbit anti - il-33 polyclonal antibody ( 1 : 1,000 dilution in pbs containing 5% bsa ) . after rinsing with pbst ( pbs containing 0.01% triton x-100 ) , tissue sections were incubated for 40 minutes at rt with hrp - conjugated goat antirabbit igg ( 1 : 100 ) and rinsed with pbst . subsequently , slides were stained with diaminobenzidine ( dab ) and counterstained with hematoxylin , dehydrated , and mounted . in order to express recombinant il-33 in e. coli , the mature protein coding region of il-33 was generated by pcr and was cloned between ndei and xhoi sites of pet-44 vector containing the coding sequence for a hexahistidine - coding sequence in the c - terminal part . amplification of the il-33 gene by pcr produced a single amplified 480 bp dna fragment , coding for a mature il-33 protein of 160 amino acids . the final pet-44-mil-33 construct is able to express the recombinant il-33 protein in fusion with a hexahistidine tag and allow for metal affinity purification . after induction with iptg , e. coli bl21 ( de3 ) transformed with pet-44-mil-33 produced a protein of approximately 18 kda as shown in figure 1 . the size of the protein matched well with its theoretical molecular weight . to determine the optimal induction period , the bacteria were incubated with iptg at 25c for 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , and 9 hours , respectively . the results showed that while the incubation period was below 7 hours , the yield of the ~18 kda protein was increased as the induction time prolonged ; but when the incubation time exceeded 7 hours , the yield of the product was not significantly raised ( figure 1 ) . mmol / l , and the bacteria were cultivated at 25c for 7 hours . induced bl21 ( de3 ) cells at 25c can express the il-33 protein ( figure 2 , lane 5 ) . in contrast , noninduced bl21 ( de3 ) cells did not express this fusion protein ( figure 2 , lane 2 ) . the soluble il-33 was expressed at high levels at 25c ( figure 2 , lane 3 ) , but at low levels at 37c ( data not shown ) . this was most likely due to growth at 37c , which causes some protein to accumulate as inclusion bodies , whereas incubation at 25c decreases the rate of protein synthesis and leads to soluble protein . the target il-33 protein appeared as a single band on sds - page ( figure 2 , lane 1 ) , which is in agreement with the molecular weight reported . polyhistidine tags form high - affinity complexes with immobilized divalent metal ions ( such as ni or co ) , even in the presence of high concentrations of chaotropic agents ( e.g. , urea or guanidine hydrochloride ) , thereby allowing isolation of tagged protein from a crude cellular extract . his - tagged il-33 exists in the bacterial cells in two forms : either soluble protein or inclusion bodies . the soluble protein was incubated with the ni - nta agarose slurry to allow binding of his - tagged il-33 . n - terminal sequencing of the purified il-33 revealed that the first 20 residues were identical to the mouse il-33 sequence deduced from the dna ( tlsiqgtslltqspaslsty ) . after immunizing rabbits with recombinant il-33 according to standard protocol , anti - il-33 serum was purified by sas precipitation and protein a affinity chromatography . the igg fractions showed two bands , a heavy chain ( 50 kda ) and a light chain ( 25 kda ) on sds - page gel after passing through the protein a - sepharose column ( figure 3 ) . the antibody at different dilutions ( 100- to 64,000-fold ) was reacted with an equal amount of the il-33 protein ( 0.2 g ) . the antibody titer is defined as the highest dilution of serum at which the a450 ratio ( a450 of postimmunization sera / a450 of preimmunization sera ) is greater than 2.1 . the antibody titer was found to be approximately 1 : 32,000 ( figure 4 ) . il-33 is a dual - function cytokine that may function as both an intracellular heterochromatin - associated nuclear factor and a potent proinflammatory cytokine . after immunohistochemistry was performed using the anti - il-33 polyclonal antibody in mouse lung and liver tissues , there were brown staining in nuclei or cytoplasm ( figures 5(b ) , 5(d ) , 5(f ) , 5(h ) ) . furthermore , there was a significant increase of il-33 expression in hydrodynamics - injected mice with il-33 cdna plasmid compared with normal mice ( figure 5 ) . bronchial epithelial cells in lung and liver cells revealed the strongest staining with anti - il-33 antibody in hydrodynamics injected mice ( figures 5(h ) , 5(f ) ) . no staining was detected in tissues where no antibody or control igg was used in the primary layer ( igg control , figures 5(a ) , 5(c ) , 5(e ) , 5(g ) ) . our results demonstrated that the endogenous il-33 protein in tissues could be specifically recognized by our prepared anti - il-33 polyclonal antibodies . we have developed a procedure for the efficient expression and purification of il-33 using pet-44 vector and ni - nta affinity chromatography . furthermore , we report the production of specific anti - il-33 antibody and the localization of endogenous il-33 . it has been observed that il-33 mrna is broadly expressed in many tissues but is more restricted at the level of cell type , which includes pulmonary smooth muscle cells , epithelial cells , dermal fibroblasts , keratinocytes , dendritic cells , and activated macrophages . il-33 is also expressed in endothelial cells from chronically inflamed rheumatoid arthritis synovium and crohn 's disease intestine [ 3 , 18 ] and atherosclerotic tissues . however , very little information is available on the expression of il-33 protein in liver and lung tissues . to our knowledge this is the first report so far of production of polyclonal antibody against full - length mouse il-33 and il-33 protein expression in mouse liver and lung determined with anti - il-33 polyclonal antibodies . as il-33 induces gene expression of th2-associated cytokines and pathological changes in the lung , blockade of il-33 may be a new therapeutic strategy for bronchial asthma . we are investigating the effect of exogenously administered anti - il-33 antibody in a murine model of allergen - induced asthma and collagen - induced arthritis . this research may contribute to elucidation of the biological mechanisms of il-33 in human inflammatory diseases in the future . | interleukin ( il)-33 is the latest member of il-1 cytokine family . in this study , the cloning , expression , purification , and polyclonal antibody preparation of mouse il-33 were described .
the coding region of il-33 mature protein was cloned into the prokaryotic expression vector pet-44 . the recombinant protein , il-33 containing a hexahistidine tag in the c - terminal ,
was expressed in escherichia coli .
the expressed soluble protein was purified by immobilized metal - ion affinity chromatography using ni2 + -nitrilotriacetic acid agarose .
the rabbits were immunized with the purified recombinant protein .
the obtained antiserum was precipitated by saturated ammonium sulfate and then purified by protein a affinity chromatography .
the sensitivity and specificity of the antibodies were evaluated by enzyme - linked immunosorbent assay and immunohistochemistry .
the high titer ( 1 : 32000 ) polyclonal antibodies with high specificity were obtained by immunizing rabbits with the purified recombinant protein .
significant expression of il-33 was seen in mouse liver and lung tissues determined with the anti - il-33 .
the production of the polyclonal antibody against il-33 provides a good tool for studying the biofunctions of il-33 . |
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blood pressure ( bp ) levels are the product of both environmental stimuli and internal regulatory mechanisms of the cardiovascular and renal systems . as a result of the complex interaction between extrinsic and intrinsic factors , bp undergoes continuous fluctuations over a 24-hour period . with the development of noninvasive ambulatory bp monitoring , blood pressure variability ( bpv ) , a surrogate marker for the complex interaction between external and internal factors , cross - sectional studies have reported that daytime and 24-hour bpv are both directly associated with the prevalence and severity of target - organ damage ( tod ) , independent of mean bp [ 14 ] . these cross - sectional data have been confirmed by longitudinal studies , as bpv has been identified as an independent predictor of cardiovascular mortality [ 5 , 6 ] . the association between bpv and cardiovascular risk may be attributed to the augmented mechanical stress placed on the vasculature that occurs as a result of increased variability of blood flow . the augmented mechanical stress may in turn facilitate vascular remodeling and contribute to the advancement in atherosclerosis , thereby conferring greater cardiovascular risk . in the renal vasculature , the augmented mechanical stress characterized by greater bpv may lead to injury , activation , and dysfunction of the glomerular endothelium . therefore , bpv may be a contributing factor in the progression of renal disease by exerting a detrimental effect on the glomerular endothelium . cross - sectional studies support a role for bpv in renal dysfunction , as a direct association between bpv and renal damage , as indicated by microalbuminaria , has been found [ 4 , 7 , 8 ] . recently , it has been reported that the rate of bpv , which accounts for both the magnitude of bpv and the time elapsed between two successive bp readings in quantifying how fast or slow bp values change , was a significant predictor of impaired renal function in a large cohort of adults with essential hypertension , suggestive that the rate of bpv may also have clinical importance for renal function . taken together , these results suggest that both the magnitude and rate of bpv may contribute to the accelerated progression of renal disease . however , the association between bpv and renal function has not been previously investigated in the early stages of renal disease when bp - buffering mechanisms , including baroreceptor and endothelial function , are still preserved . to better elucidate this relationship and implicate bpv in the acceleration of renal disease , the purpose of this study was to investigate the relationship between bpv and renal function in a group of normotensive and pre - hypertensive african americans with normal or mildly impaired renal function . because the release of nitric oxide ( no ) from the endothelium may serve as a bp - buffering mechanism in response to fluctuations in blood flow and shear stress , a secondary purpose of this study was to assess the potential association between no and bpv in this cohort of african americans . participants were recruited via mailed brochures and local newspaper advertisements . upon response to either , the participants were contacted by telephone to assess their eligibility . each participant gave written informed consent following the explanation of study protocols during their first laboratory visit . african american men and women between the ages of 40 to 75 years of age who were sedentary ( regular aerobic exercise < 2 day per week ) , nonsmoking , nonmorbidly obese ( bmi < 40 kg / m ) , not on lipid - lowering medication , and had no history of cardiovascular disease , diabetes , hypercholestermia , liver disease , renal disease , or lung disease were initially enrolled in the study . both pre - menopausal and post - menopausal women were included in the study ; all post - menopausal women were not on hormone replacement therapy . participants on more than one anti - hypertensive medication were excluded from the study . to ensure the eligibility of all qualified participants , three screening visits were completed prior to inclusion in the study . screening visit one consisted of blood sampling and urinalysis following a 12-hour overnight fast to assess blood chemistries and renal function . any individual who had total cholesterol > 240 mg / dl or who had fasting blood glucose > 126 mg / dl were excluded from the study . glomerular filtration rate ( gfr ) was calculated using the four - variable modification of diet in renal disease ( mdrd ) study equation specific to african americans : gfr ( ml / min per 1.73 m ) = 186 ( scr ) ( age ) ( 0.742 if female ) ( 1.21 if black ) , where scr represents serum creatinine . any individual who exhibited evidence of renal disease ( gfr < 60 ml / min per 1.73 m ) was excluded from the study . participants were categorized as having normal ( gfr > 90 ml / min per 1.73 m ) or mildly impaired gfr ( gfr 6089 ml / min per 1.73 m ) according to national kidney foundation criteria . screening visits two and three required all qualified participants to undergo a physician administered physical examination and echocardiogram bicycle stress test to confirm that participants displayed no evidence of cardiovascular , pulmonary , or other chronic diseases . participants who met all inclusion criteria after screening underwent dietary stabilization for 6 weeks prior to testing . any participant receiving anti - hypertensive monotherapy ( n = 14 ) was tapered off of their medication during this dietary stabilization period . participants were instructed by a registered dietician on the american heart association ( aha ) dietary guidelines for healthy american adults , a diet formerly known as the aha step 1 diet . this diet consisted of ~55% of total daily calories from carbohydrates , 15% from protein , and < 30% from fat , with saturated fat 10% of total calories , sodium 3 - 4 g / day , and cholesterol intake < 300 mg / day . participants met with the dietician one time per week at which time body weight and bp were recorded for each visit . participants were required to remain within 5% of their study entry body weight for the duration of the study . in addition , participants who exhibited bp consistently greater than 159/99 mmhg during the stabilization period were excluded from the study . compliance to the prescribed diet was monitored by completion of a 3-day food record at the conclusion of dietary stabilization . all participants who were in compliance with the diet underwent testing 1 - 2 weeks after dietary stabilization . office bp measurements were made in accordance with jnc 7 guidelines on three separate occasions . bp was measured using a mercury sphygmomanometer after 5 minutes of quite rest in a chair , with feet on the floor and arm supported at heart level . an appropriate - size cuff was applied around the participant 's nondominant arm and bp values were identified from the first and fifth phase of korotokoff sounds . bp measurements were performed in triplicate , 5 min apart , and the average of the three values was used as the bp for the visit . participants underwent 24-hour ambulatory bp monitoring ( abpm ) using a non - invasive monitor ( spacelabs medical inc . , model 90219 , redmond , wa ) beginning on the morning of each participant 's typical day , with the exclusion of friday through sunday . the bp cuff was fitted to the participant 's nondominant arm with cuff size determined by upper arm circumference . bp measurements were obtained at 30 minute intervals during the day ( 6:00 am10:00 pm ) and 60 minute intervals at night ( 10:00 pm6:00 am ) . participants were instructed not to exercise prior to or during the 24-hour bp monitoring period and to pause momentarily and maintain their body position during each bp measurement . throughout the duration of the 24-hour recording period , participants were required to maintain a diary in which they recorded their activity and emotional status at the time of each bp measurement . only recordings of good technical quality ( > 80% of valid bp measurements ) were included in final analyses . total volume of urine was measured and recorded ; thereafter , urine samples were sent to quest diagnostics for measurement of urinary creatinine , urinary sodium ( na ) , and urinary albumin levels . awake and sleep periods were defined according to self - reported sleep times recorded in participants ' diaries . 24-hour , awake , and sleep mean values were calculated for systolic bp ( sbp ) , diastolic bp ( dbp ) , mean arterial pressure ( map ) , and heart rate ( hr ) . participants were categorized as pre - hypertensive ( 120/80139/89 ) or normotensive ( < 120/80 mmhg ) according to their mean 24-hour bp . abpm measurements were used instead of office bp measurements under the assumption that bp measurements obtained during abpm reflect a participant 's bp in their natural environment , outside the influence of experimental settings . because current criteria for abpm do not have a clearly defined pre - hypertensive category , we defined prehypertension according to jnc 7 guidelines ( 120/80139/89 mmhg ) . the % dip in bp during sleep was calculated as : ( awake bp sleep bp ) ( 100/awake bp ) . bpv was calculated using the average real variability ( arv ) index
( 1)arv=1n1k=1n1|bpk+1bpk| ,
where n is the number of valid bp measurements and bpk+1 and bpk represent two successive bp measurements . the rationale for selecting the arv index for bpv calculations is based on previous studies that have reported the arv index to be a more reliable representation of time series variability than standard deviation or coefficient of variation [ 13 , 14 ] . the rate of bp variation was calculated according to the formula described by zakopoulos et al . ri is defined as follows :
( 3)ri = si+1siti+1ti ,
at time index
( 4)tri = ti+ti+12 ,
si and si+1 represent two successive bp measurements at time indices ti and ti+1 . we selected the rate of bp variation as an additional measure of bpv because it accounts for the order in which the bp measurements are obtained , as well as the time between successive readings . moreover , this parameter permits the evaluation of how fast or slow and in which direction bp values change . the rationale for using both the arv index and the rate of bp variation was to allow us to quantify the absolute magnitude of bpv ( arv index ) , while also considering the magnitude of bpv in relation to the time that elapsed between successive bp readings ( rate of bp variation ) . body composition was assessed by whole - body bia using the single - frequency impedance instrument impedimed df50 ( san diego , ca ) . bia was measured at 50 khz on the right side of the body , with two electrodes placed on each dorsal right hand and dorsal right foot while participants were lying in a supine position with their legs slightly apart and hands resting next to the body palms down . measures were taken after at least 10 min of lying in the supine position to reduce possible errors from acute changes in body fluid distribution . three measurements were taken , and the mean output values of impedance , phase , resistance , and reactance were used for calculations of total fat mass and total lean body mass according to the manufacturer 's standard operating procedures . blood was drawn into k2 edta tubes , centrifuged at 2000 g for 20 minutes at 4c , and then the plasma was frozen at 80c until assay . on the day of assay , plasma samples were ultrafiltrated through a 10,000 mwco amicon ultra filter ( millipore ) by microcentrifuge at 14,000 g for 30 minutes at 4c . urine was aliquoted from the total volume 24-hour urine collection and frozen at 80c until assay . levels of no end - products were measured using a modified griess assay as previously described . inter - assay and intra - assay cvs were 7.6% and 10.6% respectively . data are expressed as means the standard error of the mean ( sem ) . the distribution of all variables was examined using the shapiro - wilk test of normality , and homogeneity of variances was determined using levene 's test . variables that were found to fail the normality test were log adjusted for any statistical analysis , but true physiological values of any variable are reported throughout the paper . statistical comparisons were performed between normal and mildly impaired renal function groups using the independent t - test . diastolic bpv and the rate of variation in dbp were also compared between the two groups by ancova after adjusting for age , gender , bmi , plasma glucose , cholesterol values , and mean bp levels obtained from office and abpm measurements . the relationship of each clinical , renal , and bp variable with gfr and urinary albumin were tested using univariate and multivariate regression analyses . all statistical analyses were performed using spss version 17.0 ( spss inc . , chicago , il ) . the final study group consisted of 38 african american men ( n = 5 ) and women ( n = 33 ) who were classified as having normal ( gfr > 90 ml / min per 1.73 m ; n = 22 ) or mildly impaired renal function ( gfr 6089 ml / min per 1.73 m ; n = 16 ) . the clinical and renal characteristics of normal or mildly impaired renal function groups are presented in table 1 . all clinical and renal characteristics were similar between the two groups with the exception of serum creatinine and gfr . the normal renal function group exhibited significantly higher office dbp than the mildly impaired renal function group ( p < .05 ) . awake , sleep , and 24-hr sbp , dbp , map , and hr were similar between the two groups . the mildly impaired renal function group exhibited significantly lower diastolic bpv during the awake period even after adjusting for age , gender , bmi , prior anti - hypertensive medication usage , and mean bp levels from office and abpm measurements . similarly , diastolic bpv was also significantly lower in the mildly impaired renal function group during the 24-hour period . this difference was significant after adjusting for gender and bmi , but failed to reach significance after adjusting for age and mean bp levels . there were no statistically significant differences between the two groups for systolic bpv during the awake ( normal : 8.2 0.4 versus mildly impaired : 8.4 0.4 mmhg ) , sleep ( normal : 8.7 0.8 versus mildly impaired : 9.2 0.8 mmhg ) , and 24-hour periods ( normal : 8.4 0.4 versus mildly impaired : 8.5 0.4 mmhg ) . likewise , map variability ( mapv ) was similar between the two groups during the awake ( normal : 7.5 0.4 versus mildly impaired : 7.0 0.3 mmhg ) , sleep ( normal : 7.7 0.8 versus mildly impaired : 8.6 1.4 mmhg ) , and 24-hour periods ( normal : 7.6 0.4 versus mildly impaired : 7.1 0.2 mmhg ) . the rate of variation in dbp during the awake , sleep , and 24-hour periods are presented in figure 2 . the rate of dbp variation was significantly lower in the mildly impaired renal function group during the awake and 24-hour periods . these differences remained significant after adjusting for all clinical characteristics , renal characteristics , and mean bp levels from office and abpm measurements . there were no statistically significant differences between the two groups for the rate of variation in sbp during the awake ( normal : 0.258 0.011 versus mildly impaired : 0.269 0.012 mmhg / min ) , sleep ( normal : 0.146 0.014 versus mildly impaired : 0.158 0.013 mmhg / min ) , and 24-hour periods ( normal : 0.237 0.011 versus mildly impaired : 0.246 0.009 similarly , the rate of variation in map was not statistically different between the two groups during the awake ( normal : 0.237 0.013 versus mildly impaired : 0.227 0.011 mmhg / min ) , sleep ( normal : 0.128 0.016 versus mildly impaired : 0.149 0.024 mmhg / min ) , and 24-hour periods ( normal : 0.218 0.011 versus mildly impaired : 0.208 0.008 univariate regression analyses of selected clinical and abpm variables predicting gfr in the entire study group are presented in table 3 . diastolic bpv and mapv during the awake and 24-hour periods , and the rate of variation for dbp and map during the awake and 24-hour periods were identified as significant predictors of gfr . multivariate regression analyses predicting gfr in the entire study group are presented in table 4 . a core regression model was created using variables known to impact renal function : age , bmi , 24-hour sbp , and 24-hour dbp . each variable identified as a significant predictor of gfr with univariate analyses was then inserted separately into the model . awake diastolic bpv , 24-hour diastolic bpv , awake rate of dbp variation , and 24-hour rate of dbp variation all remained significant predictors of gfr after adjusting for age , bmi , and mean 24-hour bp . to determine which abpm parameters were the strongest predictors of gfr , all variables were entered into a forward step - wise linear regression model . in the final model , only the 24-hour rate of dbp variation was included as a predictor of gfr ( = .639 , p < .001 ) , with 40.8% of the variation in gfr accounted for by the final predictor model ( r = .408 ) . no associations were found for any bpv or bp variation parameter and urinary albumin in the entire study group with univariate regression analyses . when the renal function groups were analyzed separately , urinary albumin was positively associated with the rate of dbp variation in the mildly impaired renal function group during the awake ( = .568 , p < .05 ) and 24-hour ( = .570 , p < .05 ) periods . no associations were found for urinary albumin and bpv or bp variation parameters in the normal renal function group . plasma nox ( pnox ) , urinary nox ( unox ) , and the unox / pnox ratio were analyzed to investigate the potential role of no as an underlying mechanism linking renal function to bp fluctuations . pnox levels were significantly higher in the mildly impaired renal function group ( normal : 19.2 2.8 versus mildly impaired : 27.2 3.6 mol / l ; p < .05 ) . there were no statistically significant differences between the two groups for unox ( normal : 620.3 79.7 versus mildly impaired : 753.5 153.4 mol / gcr ) and the unox / pnox ratio ( normal : 39.5 5.6 versus mildly impaired : 28.4 5.8 ) . in univariate regression analyses predicting gfr , both pnox ( = .495 , p < .05 ) and the unox / pnox ratio ( = .457 , p < .01 ) were significantly associated with gfr ( figure 3).when inserted into a multivariate regression model that included age , bmi , 24-hour sbp , and 24-hour dbp , pnox remained a significant of predictor of gfr ( = .372 ; p < .05 ) . the unox / pnox ratio also remained a significant predictor of gfr ( = .393 ; p < .05 ) when inserted into a multivariate regression model that included age and bmi , but did not remain a significant predictor when 24-hour sbp and 24-hour dbp were added to the model . for univariate regression analyses predicting urinary albumin , there was a significant negative association between unox and urinary albumin ( = .497 ; p < .01 ) that remained significant when inserted into a multivariate regression that included age , bmi , 24-hour sbp , and 24-hour sbp ( = .625 ; p < .01 ) . in order to determine the relationship between a potential bp - buffering mechanism ( no ) and fluctuations in bp , univariate regression analyses predicting bpv and bp variation parameters were conducted . a significant negative association existed between pnox and 24-hour diastolic bpv ( = .356 , p < .05 ) . all remaining renal and abpm variables showed no significant associations with either pnox , unox , or the unox / pnox ratio in the entire study group or in renal function subgroups . in the present study , we used two indices of bpv to investigate the relationship between renal function and bp variation : the arv index , which accounts for the order in which bp measurements are obtained , and the time rate of variation , which accounts for the order in which bp measurements are obtained , as well as the time between successive readings . our results show that normotensive and pre - hypertensive african americans with mildly impaired renal function exhibit lower diastolic bpv and a lower rate of variation in dbp when compared to african americans with normal renal function . moreover , diastolic bpv and the rate of variation in dbp were identified as independent determinants of renal function , as both indices of dbp variation positively correlated with gfr in this cohort of african americans . our findings conflict with previous studies in diseased populations , as it has been more often shown that greater bp variation is associated with poorer health outcomes in patients with essential hypertension or chronic kidney disease ( ckd ) . most recently , manios et al . found that systolic and diastolic bpv and the 24-hour rate of sbp variation were significantly greater in hypertensive patients with a gfr less than 60 ml / min per 1.73 m when compared to hypertensive patients with a gfr greater than 60 ml / min per 1.73 m . however , few studies have investigated bp variations in populations without overt clinical disease as the majority of studies investigating bpv have been conducted in hypertensive populations . given that hypertensive bp loads are directly associated with tod , and considering that hypertension may be a cause or consequence of baroreceptor and/or endothelial dysfunction ; the observed associations between bpv and tod in these populations could be a result of the effects of a hypertensive bp load on both organs and bp - buffering mechanisms . thus , our disparate findings may simply be representative of subclinical populations in which some of the bp - buffering mechanisms are still intact , and could suggest that changes in bp regulation mechanisms , resulting in wider bp fluctuations , may occur during the progression of both ckd and hypertension from early , subclincal disease states . this hypothesis is corroborated by our finding that urinary albumin excretion was positively associated with the rate of dbp variation in the mildly impaired renal function group , while no such association was found in the normal renal function group . however , it should be acknowledged that the cross - sectional design of the present study can not determine whether changes in bp variation promoted ckd or vice versa . it has been proposed that no may serve as a bp - buffering mechanism that regulates fluctuations in bp level . several animal studies have shown that bpv increases with blockade or knockout of endothelial nitric oxide synthase [ 17 , 18 ] , however , to the best of our knowledge , this is the first study to demonstrate a relationship between no and bpv obtained from abpm in humans , as a negative association between diastolic bpv and pnox levels was found . interestingly , the mildly impaired renal function group exhibited greater pnox levels than the normal renal function group . moreover , when gfr was analyzed as a continuous variable , pnox negatively correlated with gfr and was identified as an independent determinant of renal function . no plays a major role in renal perfusion and glomerular filtration ; therefore , the elevation in pnox with declining gfr observed in the present study may serve as a compensatory mechanism to maintain blood flow to the kidneys in an effort to preserve gfr . this is the first study to compare nox levels in african americans with normal or mildly impaired renal function ; therefore , our findings need to be confirmed ; however , in support of no serving in a compensatory capacity are several animal studies which have also observed a paradoxical increase in no with declining renal function [ 2022 ] . in keeping with the hypothesis of no as a bp - buffering mechanism , it therefore seems plausible that the lower bp variation observed in the mildly impaired renal function group may be the result of higher levels of no production . it is interesting to note that higher pnox levels were associated with lower gfr , while no such association was observed between unox levels and gfr . because 60%73% of pnox is excreted renally [ 23 , 24 ] , we also calculated the unox / pnox ratio in order assess unox derived from the kidneys after accounting for basal pnox levels . our findings showed that a lower unox / pnox ratio was associated with a lower gfr , which could suggest that no production in the kidney is attenuated with declining renal function . in accordance with these findings , we observed that lower unox was associated with increased urinary albumin excretion . it has been previously shown that no production in localized areas of the kidney , including the cortex and outer and inner medulla , is important in the regulation of renal hemodynamics . moreover , it has been well reported in both clinical and animal studies that deficiencies in no are associated with kidney damage and/or chronic kidney disease progression . as such , diminished no production / bioavailability in the kidney could be underlying the finding of reduced bpv in subjects with mildly impaired renal function . to the best of our knowledge , this is the first study to investigate bp variations in relationship to renal function in a cohort of african americans . therefore , our finding that diastolic bpv and mapv , but not systolic bpv , were strongly associated with renal function may be the result of underlying race specific contributions to bp regulation . increased peripheral vascular resistance caused by arterial vasoconstriction is the key determinant of dbp , particularly before 5060 years of age . in previous studies , when the hemodynamic mechanisms that underlie the bp response were assessed to determine their influence on bp variations from abpm measurements in different racial groups , systemic vascular resistance was found to play a more prominent role in the regulation of bp in african africans when compared to whites . in contrast , cardiac output was a prominent contributor to abpm measurements only in whites and not in african americans . moreover , when the hemodynamic responses were compared among racial group after three separate laboratory stressors , the pressor responses in african americans were found to be mediated to a greater extent by vascular tone when compared to whites . in keeping with the role of peripheral vascular resistance as the key determinant of dbp , it therefore may be reasonable to hypothesize that the strong associations found for the measures of dbp and map variations , and not for the sbp variations in the present study , may be attributed to the greater contribution of systemic vascular resistance to bp regulation in african americans as they are exposed to daily stressors in their natural environment . however , because of our extensive exclusion criteria , many confounding variables that may influence renal function were well controlled for . second , the low frequency of bp measurements with abpm limits the capacity to optimally assess bp variation . conversely , the use of abpm permits the assessment of the variation of bp in a participant 's natural environment where they are exposed to the various psychological and environmental stressors which influence their bp on a daily basis . third , the reproducibility of bp variation may be poor due to the large influence of daily activity on bp variations . finally , our cross - sectional study design allowed us to assess the relationship between renal function and bpv , but not to assess cause - effect relationships . prospective studies are needed to determine whether the progression from the early stages of ckd to end - stage renal disease confers changes in the physiological mechanisms that regulate bpv . in conclusion , our study provides novel information on the relationship between bpv and renal function by assessing their relationship for the first time in a group of normotensive and pre - hypertensive african americans with normal or mildly impaired renal function . our findings suggest that both the magnitude and rate of diastolic bpv are reduced in african americans with mildly impaired renal function , which may in part be the result of a compensatory mechanism in the early stages of hypertension and kidney disease , whereby no levels in the blood are elevated . these results conflict with previous studies in diseased populations and different races as the relationship between bpv and renal function is reversed in patients with more advanced renal dysfunction and higher bp levels . future studies should investigate whether changes in bp regulation mechanisms , resulting in wider bp fluctuations , may be involved in the progression of both renal disease and hypertension , and whether these changes could explain the accelerated progression of ckd found in african americans . | we investigated the relationship between renal function , blood pressure variability ( bpv ) , and nitric oxide ( no ) in a group of african americans with normal or mildly impaired renal function .
24-hour ambulatory blood pressure monitoring was performed , no measured , and glomerular filtration rate ( gfr ) calculated in 38 african americans .
participants were categorized as having normal ( gfr > 90 ml / min per 1.73 m2 ) or mildly impaired ( gfr 6089 ml / min per 1.73 m2 ) renal function .
diastolic bpv was significantly lower in the mildly impaired renal function group .
regression analyses revealed a significant positive association between gfr and diastolic bpv for the entire study group .
plasma no levels were significantly higher in the mildly impaired renal function group and negatively correlated with diastolic bpv . in conclusion ,
diastolic bpv was reduced in african americans with mildly impaired renal function , which may be the result of increased no production .
these results conflict with previous findings in diseased and nonblack populations and could provide rationale for studying bpv early in the disease state when bp - buffering mechanisms are still preserved . |
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we report the cases of 3 patients with whim syndrome who are affected by tof ( figure ) . taking into consideration the relatively poor prevalence of the two disorders , in the present report the first patient is a 19-year - old man with a history of whim syndrome identified at age 2.5 years , manifesting as severe neutropenia and recurrent pneumonias , resulting in bronchiectases ( table ) . these hematologic and clinical findings led to the suspicion of whim syndrome and thereby to cxcr4 gene sequencing that revealed s338x mutation . the congenital heart disease was suspected at birth and was characterized by the anatomic variant of tof associated with pulmonary atresia and with anomalies in branch pulmonary arteries . of note , the patient presented with agenesis of the left - hand fingers with homolateral hypoplasia of the radius . the patient has been maintained on daily subcutaneously administered granulocyte - colony stimulating factor therapy since 2 years of age . in the second case , the patient had tof associated with the presence of patent ductus arteriosus documented at birth ; the heart disease was surgically corrected at the age of 2 years ( table ) . this patient is a 15-year - old girl who has had recurrent respiratory infections since early childhood . severe neutropenia was discovered at age of 2 years , and on that occasion , an analysis of the bone marrow revealed myelokathexis with mature neutrophils presenting morphologic abnormalities consistent with apoptosis . granulocyte - colony stimulating factor therapy was started at 5 years of age to maintain circulating neutrophil count in the normal range . because of the occurrence of repeated pneumonia episodes , she has been maintained on antibiotic prophylaxis . in the following years , the observation of hypogammaglobulinemia suggested whim syndrome and genetic confirmation of the diagnosis was obtained at age 5 by detection of the r334x mutation in the cxcr4 gene . the third patient is 7 years old and presented shortly after birth with a heart murmur that led to the discovery of a tof characterized by ventricular septal defect , overriding aorta , and pulmonary infundibular stenosis . neutropenia , lymphopenia , and hypogammaglobulinemia were present since infancy , and the analysis of bone marrow showed myelokathexis . three other family members have had neutropenia and recurrent infections , but none of them had a congenital heart defect . the genetic analysis of cxcr4 revealed the same mutation ( s338x ) in the 4 subjects . the child underwent treatment with intravenous immunoglobulins , obtaining a satisfactory control of infectious episodes . although tof is the most common form of cyanotic congenital heart disease , its occurrence in 3 unrelated patients with whim syndrome is much higher than expected ; the normal occurrence in the general population is 3 of every 10 000 live births . because of the low number of patients whom we have observed , we can not draw a correlation between the development of the heart defect with a specific mutation . the detection of the same mutation ( s338x ) in a family with 4 members affected by whim syndrome with only 1 showing tof rules out the hypothesis that the heterozygous gain - of - function mutation of cxcr4 directly leads to the development of the cardiac defect . instead , our observations suggest that the whim syndrome associated cxcr4 truncating mutation might increase the risk that this combination of cardiac defects may develop during the formation of the fetal heart . beyond a role for cxcl12 and cxcr4 in heart , nervous system , and blood vessel development , studies show that cxcr7 , the recently described second receptor for cxcl12 , has also a role in fetal endothelial biology and heart development in particular , ventricular septum and heart valve development . its germline deletion results in perinatal lethality , and its mutation affects semilunar valve development , contributing to aortic and pulmonary valve stenosis and , in some cases , septal defects . recent findings support the view that cxcr7 modulates cxcr4 function by acting as a scavenger for cxcl12 and forming heterodimers with cxcr4 . in vitro studies on endothelial progenitor cell function show a role for both cxcr4 and cxcr7 in regulating the response of the cells to cxcl12 and , thus , angiogenesis . this suggests that interactions between the 2 chemokine receptors may be required for proper valve morphogenesis . our observations demonstrate that tof can be a presenting manifestation of whim syndrome and that this rare inherited disease should be suspected in children with a congenital heart defect and neutropenia . the recognition of this manifestation of whim syndrome might help to prevent the diagnostic delay in the identification of this rare genetic disease . a prompt diagnosis should facilitate the management of leucopenia , which might include in the future cxcr4-targeted therapy as supported by 2 recent studies indicating that the specific cxcr4 antagonist plerixafor may be effective in restoring the cellular blood counts to normal . | warts , hypogammaglobulinemia , infections , and myelokathexis ( whim ) syndrome is a rare immunodeficiency disorder . we report three patients with whim syndrome who are affected by tetralogy of fallot ( tof ) .
this observation suggests a possible increased risk of tof in whim syndrome and that birth presentation of tof and neutropenia should lead to suspect whim syndrome . |
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in this prospective , descriptive - analytic study , 523 pregnant women , who received prenatal care in alzahra and shahid behesti hospitals in isfahan , were selected by convenient sampling method from june 2009 until june 2010 . selected pregnant women with renal disease , infection , previous transplantation , collagen vascular disease and chronic hypertension were excluded from the study . the study protocol was approved by the ethics committee of isfahan university of medical sciences and written informed consents were obtained from all studied patients . maternal and perinatal outcomes were defined in accordance with the american college of obstetrics and gynecology guidelines.18 women with and without preeclampsia were determined and the level of cecs and e - selectin were compared in the two studied groups ( figure 1 ) . flow diagram of patient participation through the study
positive rollover test : the rollover test was considered positive if diastolic blood pressure increased 20 mmhg when the patient moved from the left lateral recumbent to supine position.19preeclampsia : preeclampsia was defined as the development of a systolic blood pressure of 140 mmhg or greater , and/or a diastolic blood pressure of 90 mmhg or greater and proteinuria of 300 mg or greater per 24 hours after 20 weeks of pregnancy , or urine dipstick 1 + or greater on 2 occasions for at least 4 hours . blood pressures were measured 2 times for each patient at least 4 hours apart.20 the blood pressure was taken with the patient 's arm supported on the level of the heart . diastolic pressure was reported as phase v korotkoff.21
positive rollover test : the rollover test was considered positive if diastolic blood pressure increased 20 mmhg when the patient moved from the left lateral recumbent to supine position.19 preeclampsia : preeclampsia was defined as the development of a systolic blood pressure of 140 mmhg or greater , and/or a diastolic blood pressure of 90 mmhg or greater and proteinuria of 300 mg or greater per 24 hours after 20 weeks of pregnancy , or urine dipstick 1 + or greater on 2 occasions for at least 4 hours . blood pressures were measured 2 times for each patient at least 4 hours apart.20 the blood pressure was taken with the patient 's arm supported on the level of the heart . diastolic pressure was reported as phase v korotkoff.21 immunomagnetic bead method : we measured cecs according to a previously validated technique.2223 therefore , one milliliter of thoroughly mixed venous blood was collected in an ethylenediaminetetraacetic acid ( edta ) tube while the first 7 ml of blood drawn were discarded to avoid contamination by endothelial cells from the punctured vessel wall . the blood sample was mixed with 1 ml lysis buffer and incubated at room temperature for 10 minutes . the suspension was centrifuged for 10 minutes and the cell pellet was resuspended in 1 ml of phosphate buffered saline ( pbs ) solution . then , 20 l of fc - receptor blocking agent ( miltenyi , germany ) were added and incubated for 20 minutes . followed by further incubation , one hundred microliters of anti - cd146 coated microbeads per 10 cells were then added to this mixture and incubated for 15 minutes at 4c . bead - bound cells were then separated from non - bound cells using a combination of washing with buffer solution and magnetic separation ( miltenyi , germany ) . one hundred microliters of prepared ulex europaeus lectin solution ( 2 mg / ml ) ( sigma , germany ) was then added to the clean cell / bead suspension and gently mixed for 30 minutes at room temperature ( 22 - 24c ) in darkness . the solution was then washed three times in a magnet . the resulting rosetted cells and beads were finally resuspended in 125 l of pbs for counting under epifluorescence microscopy ( zeiss , welwyn garden city , uk ) in a nageotte counting chamber . cells were defined as cecs if they demonstrated rosetting with 4 cd146-immunobeads , were approximately 20 - 50 m in diameter with distinctive cell morphology , and stained positively with endothelial specific ulex europaeus lectin . serum levels of e - selectin were assessed by human se - selectin platinum enzyme - linked immuno - sorbent assay ( elisa ) kits produced by ebioscience according to the manufacturer 's instructions . we hypothesized that increased numbers of cecs early in the third trimester are associated with an increased risk of developing preeclampsia . therefore , the numbers of cecs in preeclamptic and normal patients were compared using student 's t - test . p values less than 0.05 were considered statistically significant . in order to determine the appropriate cutoff point of cecs number for identifying the risk of preeclampsia , the sensitivity , specificity , and likelihood ratio were calculated at different levels of cecs using receiver operating characteristics ( roc ) curve . positive rollover test : the rollover test was considered positive if diastolic blood pressure increased 20 mmhg when the patient moved from the left lateral recumbent to supine position.19preeclampsia : preeclampsia was defined as the development of a systolic blood pressure of 140 mmhg or greater , and/or a diastolic blood pressure of 90 mmhg or greater and proteinuria of 300 mg or greater per 24 hours after 20 weeks of pregnancy , or urine dipstick 1 + or greater on 2 occasions for at least 4 hours . blood pressures were measured 2 times for each patient at least 4 hours apart.20 the blood pressure was taken with the patient 's arm supported on the level of the heart . diastolic pressure was reported as phase v korotkoff.21
positive rollover test : the rollover test was considered positive if diastolic blood pressure increased 20 mmhg when the patient moved from the left lateral recumbent to supine position.19 preeclampsia : preeclampsia was defined as the development of a systolic blood pressure of 140 mmhg or greater , and/or a diastolic blood pressure of 90 mmhg or greater and proteinuria of 300 mg or greater per 24 hours after 20 weeks of pregnancy , or urine dipstick 1 + or greater on 2 occasions for at least 4 hours . blood pressures were measured 2 times for each patient at least 4 hours apart.20 the blood pressure was taken with the patient 's arm supported on the level of the heart . diastolic pressure was reported as phase v korotkoff.21 immunomagnetic bead method : we measured cecs according to a previously validated technique.2223 therefore , one milliliter of thoroughly mixed venous blood was collected in an ethylenediaminetetraacetic acid ( edta ) tube while the first 7 ml of blood drawn were discarded to avoid contamination by endothelial cells from the punctured vessel wall . the blood sample was mixed with 1 ml lysis buffer and incubated at room temperature for 10 minutes . the suspension was centrifuged for 10 minutes and the cell pellet was resuspended in 1 ml of phosphate buffered saline ( pbs ) solution . then , 20 l of fc - receptor blocking agent ( miltenyi , germany ) were added and incubated for 20 minutes . followed by further incubation , one hundred microliters of anti - cd146 coated microbeads per 10 cells were then added to this mixture and incubated for 15 minutes at 4c . bead - bound cells were then separated from non - bound cells using a combination of washing with buffer solution and magnetic separation ( miltenyi , germany ) . one hundred microliters of prepared ulex europaeus lectin solution ( 2 mg / ml ) ( sigma , germany ) was then added to the clean cell / bead suspension and gently mixed for 30 minutes at room temperature ( 22 - 24c ) in darkness . the solution was then washed three times in a magnet . the resulting rosetted cells and beads were finally resuspended in 125 l of pbs for counting under epifluorescence microscopy ( zeiss , welwyn garden city , uk ) in a nageotte counting chamber . cells were defined as cecs if they demonstrated rosetting with 4 cd146-immunobeads , were approximately 20 - 50 m in diameter with distinctive cell morphology , and stained positively with endothelial specific ulex europaeus lectin . serum levels of e - selectin were assessed by human se - selectin platinum enzyme - linked immuno - sorbent assay ( elisa ) kits produced by ebioscience according to the manufacturer 's instructions . we hypothesized that increased numbers of cecs early in the third trimester are associated with an increased risk of developing preeclampsia . therefore , the numbers of cecs in preeclamptic and normal patients were compared using student 's t - test . p values less than 0.05 were considered statistically significant . in order to determine the appropriate cutoff point of cecs number for identifying the risk of preeclampsia , the sensitivity , specificity , and likelihood ratio were calculated at different levels of cecs using receiver operating characteristics ( roc ) curve . positive rollover test : the rollover test was considered positive if diastolic blood pressure increased 20 mmhg when the patient moved from the left lateral recumbent to supine position.19preeclampsia : preeclampsia was defined as the development of a systolic blood pressure of 140 mmhg or greater , and/or a diastolic blood pressure of 90 mmhg or greater and proteinuria of 300 mg or greater per 24 hours after 20 weeks of pregnancy , or urine dipstick 1 + or greater on 2 occasions for at least 4 hours . blood pressures were measured 2 times for each patient at least 4 hours apart.20 the blood pressure was taken with the patient 's arm supported on the level of the heart . diastolic pressure was reported as phase v korotkoff.21
positive rollover test : the rollover test was considered positive if diastolic blood pressure increased 20 mmhg when the patient moved from the left lateral recumbent to supine position.19 preeclampsia : preeclampsia was defined as the development of a systolic blood pressure of 140 mmhg or greater , and/or a diastolic blood pressure of 90 mmhg or greater and proteinuria of 300 mg or greater per 24 hours after 20 weeks of pregnancy , or urine dipstick 1 + or greater on 2 occasions for at least 4 hours . blood pressures were measured 2 times for each patient at least 4 hours apart.20 the blood pressure was taken with the patient 's arm supported on the level of the heart . immunomagnetic bead method : we measured cecs according to a previously validated technique.2223 therefore , one milliliter of thoroughly mixed venous blood was collected in an ethylenediaminetetraacetic acid ( edta ) tube while the first 7 ml of blood drawn were discarded to avoid contamination by endothelial cells from the punctured vessel wall . the blood sample was mixed with 1 ml lysis buffer and incubated at room temperature for 10 minutes . the suspension was centrifuged for 10 minutes and the cell pellet was resuspended in 1 ml of phosphate buffered saline ( pbs ) solution . then , 20 l of fc - receptor blocking agent ( miltenyi , germany ) were added and incubated for 20 minutes . followed by further incubation , one hundred microliters of anti - cd146 coated microbeads per 10 cells were then added to this mixture and incubated for 15 minutes at 4c . bead - bound cells were then separated from non - bound cells using a combination of washing with buffer solution and magnetic separation ( miltenyi , germany ) . one hundred microliters of prepared ulex europaeus lectin solution ( 2 mg / ml ) ( sigma , germany ) was then added to the clean cell / bead suspension and gently mixed for 30 minutes at room temperature ( 22 - 24c ) in darkness . the solution was then washed three times in a magnet . the resulting rosetted cells and beads were finally resuspended in 125 l of pbs for counting under epifluorescence microscopy ( zeiss , welwyn garden city , uk ) in a nageotte counting chamber . cells were defined as cecs if they demonstrated rosetting with 4 cd146-immunobeads , were approximately 20 - 50 m in diameter with distinctive cell morphology , and stained positively with endothelial specific ulex europaeus lectin . serum levels of e - selectin were assessed by human se - selectin platinum enzyme - linked immuno - sorbent assay ( elisa ) kits produced by ebioscience according to the manufacturer 's instructions . we hypothesized that increased numbers of cecs early in the third trimester are associated with an increased risk of developing preeclampsia . therefore , the numbers of cecs in preeclamptic and normal patients were compared using student 's t - test . p values less than 0.05 were considered statistically significant . in order to determine the appropriate cutoff point of cecs number for identifying the risk of preeclampsia , the sensitivity , specificity , and likelihood ratio were calculated at different levels of cecs using receiver operating characteristics ( roc ) curve . in this study , a total of 47 women with positive rollover test were selected out of 523 subjects . the individuals were followed up and 22 pregnant women were diagnosed with preeclampsia while the remainder were normotensive . the characteristics of women with and without preeclampsia are presented in table 1 . as this table shows , spearman 's correlation revealed a positive correlation between age of mothers and preeclampsia ( r = 0.258 ; p = 0.5 ) . the characteristics of women with and without preeclampsia the sensitivity , specificity and predictive characteristics of different levels of cecs and e - selectin based on the roc curve are presented in table 2 . the roc curve analysis , applied to markers values , showed the best diagnostic profile for cecs and e - selectin with an area under the curve ( auc ) of 0.81 and 0.89 , respectively . the best cutoff points were 6.5 cells / ml for cecs and 14.6 ng / ml for e - selectin . the sensitivity , specificity and accuracy of both cecs and e - selectin were 88.9% , 83.3% and 86.1% , respectively . the sensitivity , specificity , positive predictive value ( ppv ) and negative predictive value ( npv ) of different levels of cecs and e - selectin using receiver operating characteristics ( roc ) curve in this study , mean level and predictive values of cecs and e - selectin in preeclamptic and non - preeclamptic pregnant women were evaluated . the results of the current study indicated that the number of cecs and level of serum e - selectin in preeclamptic women were significantly higher than normotensive pregnant women . as mentioned , preeclampsia is related to an imbalance of circulating angiogenic factors which results in endothelial malfunction and vasospasm.24 many studies supported the endothelial origin of preeclampsia . according to these studies , impaired placentation results in releasing different soluble factors in maternal circulation which consequently increases endothelial permeability possibly through protein kinase c - dependent pathways . these soluble factors can also change the expression of adhesion molecules and calcium content of endothelial cells.21 several markers of endothelial damage such as thrombomodulin , soluble von willebrand factor ( svwf ) and tumor necrosis factor- , and their efficacy in determining endothelial dysfunction have been studied previously . though measuring soluble endothelial markers , such as thrombomodulin , is easy , it could be influenced by different factors including renal function . in addition , measuring svwf and thrombomodulin is not reliable enough due to the fact that these factors could not discriminate between activation and damage.2526 in the current study , we evaluated cecs numbers as an indicator of endothelial malfunction among preeclamptic women . evidence supports the efficacy of cecs numbers in this regard and it is considered as a reliable marker for endothelial damage in many diseases . recently , use of this marker has become more widespread due to new methods which facilitate the enumeration of cecs.2728 the findings of this study indicated that median of cecs numbers was significantly higher in preeclamptic pregnant women than in normotensive ones . that evaluated the relation between cecs and preeclampsia using enumeration of cecs with anti - cd146-driven immunomagnetic isolation . they enumerated cecs in pregnant women before delivery and 1 and 3 - 5 days after delivery and reported that the mean value of cecs numbers was significantly higher in preeclampsic women ( 88 cells / ml ) than normal pregnancies ( 16 cells / m ) and healthy non - pregnant women ( 12 cells / ml ) . they also reported a significant correlation between cecs and systolic blood pressure in preeclampsic women and concluded that cecs can be considered as a novel marker of endothelial damage in preeclampsia.27 in another study in turkey , canbakan et al . evaluated the number of cecs in patients with preeclampsia using anti - cd146 antibody similar to our study . they showed that the numbers of cecs were significantly higher in preeclamptic women ( 13.2 5.2 cells / ml ) than hypertensive patients ( 6.9 0.8 cells / ml ) , healthy pregnant women ( 5.2 1.4 cells / ml ) , and non - pregnant controls ( 4.0 they indicated a significant correlation between cecs and systolic and diastolic blood pressure and concluded that cecs number was a valuable marker in evaluating the endothelial dysfunction in preeclampsia.13 in contrast , strijbos et al . investigated the levels of cecs numbers in normotensive and severe preeclamptic pregnancies and indicated that the number of cecs was not significantly different between preeclamptic women and the control group and all the numbers were in the normal range . since other markers including thrombomodulin , e - selectin and endoglin were significantly higher in preeclamptic women , they concluded that the role of endothelial dysfunction and activation in preeclampsia was more significant than the actual endothelial damage characterized by the increased numbers of cecs.29 although the mentioned studies confirmed the correlation between cecs numbers and preeclampsia , they recommended that the prognostic value of this marker in preeclampsia should also be investigated . in the current study , we determined the predictive value of cecs numbers in the diagnosis of preeclampsia early in the third trimester . the results indicated that cecs numbers during the early period of third trimester have a predictive value for later diagnosis preeclampsia . our findings showed that the serum concentrations of e - selectin were significantly higher in preeclamptic pregnant women . this marker was found to have appropriate sensitivity and specificity . in china , xing et al . indicated that e - selectin had an important role in shallow trophoblast invasion , the major pathological change of preeclampsia.30 in a recent study in greece , the level of e - selection was compared in preeclamptic and control subjects . e - selectin was found to increase significantly in preeclamptic women in antepartum and postpartum period . they concluded that endothelial dysfunction may be one of the key processes in the pathogenesis of preeclampsia in pregnancy and the links between preeclampsia and future cardiovascular diseases.31 in line with the findings of this study , kim et al . indicated that serum levels of e - selectin were significantly higher in both mild and severe preeclampsia than normal pregnancy . however , there were no statistical differences in the levels of e - selectin between mild and severe preeclampsia.32 while bretelle et al . reported similar results,16 bersinger et al . suggested that serum levels of e - selectin were within normal pregnancy ranges among preeclamptic women.33 the limitation of this study was the small sample size however , this is the first report of measuring maternal serum cecs and e - selectin levels as a predictor of preeclampsia . we demonstrated that maternal serum cecs and e - selectin levels increase prior to preeclampsia onset in women with positive rollover test . further studies with larger sample sizes are required to validate these results and estimate the sensitivity and specificity of these markers for preeclampsia screening . fm designed the study , coordinated and carried out the clinical investigation and participated in manuscript preparation . shj carried out the experiments , performed the statistical analysis and participated in manuscript preparation . | background : circulating endothelial cells ( cecs ) and e - selectin are known as sensitive and specific markers of endothelial dysfunction .
this study investigated whether cecs and e - selectin are surrogate biomarkers of preeclampsia and if measurement of cecs and e - selectin , early in the third trimester , could be a means of predicting preeclampsia.methods:in this prospective , descriptive - analytic study , rollover test was performed on 523 pregnant women during 28 - 30 weeks of gestation .
cecs were measured by anti - cd 146-driven immunomagnetic isolation in women with positive rollover test .
they were followed up prospectively until delivery without any active intervention .
women with and without preeclampsia were determined .
the number of cecs and level of e - selectin were compared in the two studied groups.results:from the 47 pregnant women with positive rollover test who were selected and followed up , 22 individuals were diagnosed with preeclampsia while the remainder were normotensive .
mean cec numbers was significantly higher in preeclamptic women than normal pregnancies ( 24.7 cells / ml vs. 13 cells / ml ) .
the best cut - off point for cec numbers was 6.5 with a sensitivity of 78.9% and a specificity of 69.1% .
the level of e - selectin was significantly higher in mothers with preeclampsia ( p < 0.05).conclusions : higher levels of cecs and e - selectin in women with positive rollover test who developed preeclampsia prior to onset of the complication were predictive of preeclampsia
. however , larger studies are needed to confirm these findings . |
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although applying the pesticides in rice has increased grain production , their use has raised serious concern . beside the high cost of the pesticides , the use of pesticides may contaminate the products , as well as posing a serious danger to human health ( 1 ) . carbamates , or urethanes , are a group of organic compounds sharing a common functional group with the general structure r1-o-(co)-nr2-r3 . carbamates are esters of carbamic acid , which are unstable compounds because of the liability of the h - groups , leading upon exposure to water to rearrangements that lead to the decomposition to ammonium bicarbonate . a group of insecticides also contains the carbamate functional group , including aldicarb , carbofuran , pirimicarb , propoxur , carbaryl ( sevin ) , ethienocarb , and fenobucarb . these insecticides can cause cholinesterase inhibition poisoning by reversibly inactivating the enzyme acetyl cholinesterase and cause a cholinergic poisoning ( 2 ) . the presence of harmful pesticide residues in food has caused a great concern among the consumers . therefore , it is important to develop some pragmatic procedures to increase the food safety along with the reduction in consumption of pesticide residue ( 3 ) . food processing treatments include baking , bread making , dairy product manufacture , drying , thermal processing , fermentation , freezing , infusion , juicing , malting , milling , parboiling , peeling , peeling and cooking , storage , storage and milling , washing , washing and cooking , washing and drying , washing and peeling , washing peeling and juicing , and wine making ( 4 ) . literature demonstrates that in most cases processing leads to large reductions in residue levels of pesticides in the prepared food , particularly through washing , peeling and cooking operations ( 5- 9 ) . the effect of food processing on pesticide residues in rice has been the focus of a few recently published articles . a report by lee et al . , it showed that washing the rice grains with water , removes approximately 60% of the chlorpyrifos residues from fortified rice samples ( 10 ) . in another study , fukuhara et al . showed that an initial level of 19 ppm permethrin in rice was removed completely by washing with water ( 11 ) . the elimination of pesticide residues from the boiled extract could be due to decomposition by the effect of heat , the stronger adsorption of pesticide onto plant tissues and/or the poor solubility of pesticides in water ( 12 ) . the persistence of trifluralin , chlorpyrifos , decamethrin , cypermethrin and dichlorvos in rice and beans after cooking in a commercial microwave oven was evaluated by castro et al . the results showed that 92% to 99% of the pesticides were eliminated ( 13 ) . the effect of boiling and cooking on reducting some organophosphorus pesticides in rice was evaluated by holand p t , at et al . the fate of malathion and chlorpyrifos after boiling in rice has been the focus by cogburn et al . in 1990 ( 15 ) . investigating about the effect of cooking process on the pesticides residue has also been the subject of studies by satoh and saka ( 16 , 17 ) . the effect of processing on pesticide residues in food is an area where the available information should be consolidated and the missing information needs to be obtained through further research . more than 90% of the world s rice is cultured and consumed in asia ( 1 ) . rice is a staple food for more than 60% of the world s population . in iranian food basket , the consumption of rice in tehran is 110 g per capita / day ( personal communication ) . in this study , we evaluated the effect of cooking on reducing the amount of three carbamates residue i.e. carbaryl , pirimicarb and propoxur pesticides residues in white rice ( figure 1 ) . the iranian traditional method for rice cooking was used , in which a mixture of 15 g of white rice and 45 ml of boiling water was placed in a 100 ml pyrex container . in order to evaluate the percentage of pesticide s reduction , this processing was applied on spiked rice samples . chemical sctructures of carbaryl , pirimicarb and propoxur the percentage of reduction in pesticides concentration after cooking were determined by gas chromatography mass - spectrometry ( gc / ms ) using interpolation of the relative peak areas for each pesticide to internal standard peak area in the sample on the spiked calibration curve . the list of selected pesticides along with some of their physico - chemical properties are presented in table 1 . all organic solvents , intended for extraction , were at least lc grade and purchased from merck ( darmstadt , germany ) . bulk quantities of anhydrous na2so4 and nacl , were obtained from merck ( darmstadt , germany ) . anhydrous mgso4 , was obtained from sigma - aldich co. ( sigma - aldich , japan ) , the mgso4 was baked for 5 h at 500c in a furnace to remove phthalates and residual water . primary secondary amine ( psa ) was purchased from supelco ( supelco , usa ) . an agilent technologies 6890n network gc system chromatographs ( wilmington , usa ) with a sq detector and equipped with an agilent 7683b autosampler ( agilent technologies , usa ) was used . a hp-5 capillary column ( film thickness : 30 m 0.25 mm i.d . calibration standards
individual stock standard solutions ( 1 mg / ml ) were prepared in ethyl acetate and stored in the dark at -20c . a mixed stock standard solution of pesticides was prepared in 15 g / ml ethyl acetate . spiked calibration standards at 6 levels of 25 , 50 , 100 , 250 , 500 and 1000 ng / ml were prepared by adding 25 l , 50 l , 100 l , 250 l , 500 l and 1000 l of mixed standard stock solution respectively to 15 g of cooked blank rice samples in each case . a stock solution of triphenylmethane ( tpm ) in ethyl acetate at concentration of 1 mg / ml was used as internal standard . an aliquot of 10 l of tpm solution in ethyl acetate ( 1000 mg / l ) was added to the spiked rice sample as internal standard . the iranian traditional method for rice cooking ( kateh ) was used , in which a mixture of 15 g of white rice and 45 ml of distilled water was placed in a 100 ml pyrex container . the mixture was heated for 20 min with hotplate and kept warm for another 45 min in order to complete the cooking process . the whole process was divided into three stages : 1 ) five min from the start of heating to the start of boiling of the mixture ; 2 ) fifteen min from the start of boiling to the start of steam coming out of the rice cooker ; 3 ) forty five min from the start of steam coming out of the rice cooker to the end of cooking . a thermo - meter was applied for measuring the variation in temperature ( 100 - 105c ) at the sampling locations during the rice cooking process . two sets of samples were considered to evaluate the effect of cooking on pesticides residues . for the first set , both pesticides and internal standard were added to the rice after cooking . the second set was spiked with the pesticides prior to the cooking process and the internal standard was added after cooking . the second set was used to evaluate the effect of cooking process on pesticides residues . in all cases , pesticides and/or internal standard solutions were added after cooling the cooked rice at room temperature ( 25c ) in order to avoid any possible decomposition of pesticides and internal standard . the spiked samples were left at ambient temperature in dark for 1.5 h. a volume of 120 ml acetonitrile was added to the mixture and then the mixture was blended . six grams of nacl was added to the mixture , and blending was continued for another 60 sec . the supernatant was filtered through 25 g of na2so4 in a 250 ml container and the filtrate was evaporated by the rotary evaporator to 10 ml . the extract was transferred to a 50 ml falcon tube including 6 g mgso4 and 300 mg psa , and the residue in container was rinsed with 15 ml acetonitrile and added to the centrifuge tube . after shaking for 1 min and centrifugation for 7 min at 3000 rpm , the filtrate evaporated to 5 ml . the extract was transferred to a 15 ml falcon tube containing 2 g mgso4 and 200 mg psa ( primary secondary amine ( psa ) whichis one of the most powerful solid- phase extraction adsorbent for clean - up of food matrices and multi pesticide residue analysis . after shaking for 1 min and centrifugation for 7 min at 503 g , the filtrate was evaporated to 0.3 ml . the residue was reconstituted by toluene to obtain 1 ml solution and 2 l of the solution was injected into gas chromatograph . for recovery determination , spiked cooked blank rice samples at concentration levels of 25 , 50 and 500 ng / g were prepared in triplicates and then treated according to the procedure described in sample preparation . evaluation of pesticide s reduction
the percentage of reduction after cooking was determined based on determination of recovery from triplicate spiked rice raw samples at levels 25 , 100 and 500 ng / g of each pesticides before cooking and then after cooking according to rice cooking and treatment according to the procedure described in sample preparation . the gc - sq - ms was employed with helium as the carrier gas at a constant flow rate of 1 ml / min . the oven temperature started at 75c and remained at this temperature for 3 min increasing to 120c in a ramp rate of 25c / min and then increased to 300c at 5 c / min ramp , holding at 300c for 11 min . the injection port was adjusted at 250c and the splitless mode was used . after acquisition of the total ion chromatogram for the mixed stock standard solutions in scan mode , the identification was confirmed by comparing the relative abundances for three ions ( one quantifier and two qualifiers ) of the experimental standards to the known relative abundances of the pest library reference spectra . the most abundant ion that showed no evidence of chromatographic interference and had the highest signal - to - noise ratio was taken for quantification purposes . the concentrations of pesticides were determined by interpolation of the relative peak areas for each pesticide to internal standard peak area in the sample on the spiked calibration curve . in order to compensate for losses during sample processing and instrumental analysis , internal standard ( tpm ) was used . gas chromatographic determination
analysis was performed in the sim mode based on the use of one target and two qualifier ions . the quantitation was based on the peak area ratio of the targets to that of internal standard . table 2 summarizes pesticides studied with their target and qualifier ions used in the sim mode in this study . the retention times , diagnostic ions and the selected quantification ion for the target pesticides and the internal standard calibration curves were constructed for each compound using six different concentration levels . for identification of pesticides , the retention time and three ions ( one for quantitation and two for identification ) were used . a gc sq ms chromatogram of 3 pesticides and internal standard ( tpm ) analyzed in cooked spiked rice is shown in figure 2 . a representative chromatogram obtained for the three pesticides ( propoxur , rt = 10.91 , 21.14 ; carbaryl , rt = 18.72 ; pirimicab , rt = 26.45 ) and internal standard ( tpm , rt = 29.85 ) . linearity of the standards in spiked calibration curves
the three pesticides showed linearity in the sim mode . linear spiked calibration curves for all the carbamate pesticides were obtained with correlation factors > 0.999 . the spiked calibration curve for pirimicarb in rice sample is shown in figure 3 as a representative . spiked calibration curve for pirimicarb in cooked rice
limitation of quantification
limits of quantification ( loqs ) of the proposed method were calculated by considering a value 10 times more than that of background noise . the loqs for all the three pesticides in this method were calculated as 25 ppb . the recovery of pesticides at these three levels was in the range of 89.47 - 100.24% . in terms of repeatability , the vast majority of pesticides gave rsd < 10% with n = 3 at each spiking level . the recoveries and repeatabilities are in accordance with the criteria set by sanco guideline ( commission of the european communities , 2006 ) . average recoveries ( % ) and range of relative standard deviations ( % ) obtained by gc - ms analysis of rice at 3 spiking levels ( n = 3 ) in rice samples
the reduction of pesticides after cooking
the results of percentage of reduction for each carbamate after the boiling process is presented in table 4 . average reduction ( % ) and range of relative standard deviations ( % ) obtained by gc - ms analysis of spiked rice at 3 spiking levels ( n = 3 ) after cooking
figure 4 shows an overlaid chromatogram of carbaryl in spiked rice samples before ( a ) and after ( b ) cooking . the overlaid chromatogram ; ( a ) blank rice which is spiked with carbaryl ( 100 ppb ) after the cooking ; ( b ) blank rice which is spiked with carbaryl ( 100ppb ) before the cooking the carbamates are a family of compounds whose general structures are shown in figure 5 . the great success of carbamates in agricultural applications has led to a continued increase in the use of these pesticides , but their acute toxicity is of great concern and calls for their accurate determination in food stuff ( 18 ) . food is the basic necessity of life and food contaminated with toxic pesticides is associated with severe effects on the human health . it is therefore of significance to develop simple , cost effective methods to enhance food safety and prevent the possible harms caused by pesticides . the disappearance of pesticide residues from boiling extract could be due to the decomposition by the effect of heat , the stronger adsorption of pesticide onto plant tissues and/or the poor solubility of pesticides in water ( 12 ) . processes involving heating , can increase volatilization , hydrolysis or the other chemical degradation and thus reduce the residue levels ( 14 ) . in this study , we evaluated the effect of boiling on the reduction of three carbamate pesticide residues in rice . the selected pesticides , including those for which mrl is issued by the institute of standards of iran ( 19 ) , include carbaryl and pirimicarb . the results of this study show that the cooking process decreases the amount of the three mentioned carbamates . carbaryl showed the highest reduction after cooking ( 78.47% in average ) while pirimicarb showed the lowest reduction ( 34.5% in average ) . no strong correlation was found between a single physico - chemical property of each of the three pesticides and the percentage of their reduction after the cooking . since several parameters including rate of hydrolysis , volatility ( vapor pressure ) , water solubility and molecular weight could affect the amount of reduction after cooking , no decisive conclusion could be drawn about the extent that each parameter contributes to the total loss of pesticides residue after cooking . investigating the effect of cooking process on the amount of reduction for a larger group of carbamates could help to find out more about mechanisms involved in this process . an accurate method was developed to determine residues of the three carbamate insecticide in cooked rice , a main food in the iranian food basket . the method which consists of a simple sample preparation and gc sq - ms - sim analysis showed a high sensitivity and confirmatory power necessary for the determination of pesticide residues at mrl levels in rice set by the institute of standards and industrial research of iran ( isiri ) . the results of present study proves that the traditional iranian rice cooking which involves long boiling time , significantly reduces the amount of carbamate residues . from the safety and toxicological point of view , this could be considered as an advantage for this type of food processing . | a gas chromatography mass spectrometry with spike calibration curve method was used to quantify three carbamate pesticides residue in cooked white rice and to estimate the reduction percentage of the cooking process duration.the selected pesticides are three carbamate pesticides including carbaryl and pirimicarb that their mrl is issued by the institute of standards of iran and propoxur which is used as a widely consumed pesticide in rice.the analytical method entailed the following steps : 1- blending 15 g cooked sample with 120 ml acetonitrile for 1 min in solvent proof blender , 2- adding 6 g nacl and blending for 1 min , 3- filtering upper layer through 25 g anhydrous na2so4 , 4- cleaning up with psa and mgso4 , 5- centrifuging for 7 min , 6- evaporating about 0.3 ml and reconstituting in toluene till 1 ml , 7- injecting 2 l extract into gc / ms and analyzing by single quadruple selected ion monitoring gc / ms - sq - sim .
the concentration of pesticides and the percentage of pesticides amounts after the cooking were determined by gas chromatography mass - spectrometry ( gc / ms ) using with interpolation of the relative peak areas for each pesticide to internal standard peak area in the sample on the spiked calibration curve .
calibration curve was linear over the range of 25 to 1000 ng / g , and loq was 25 ng / g for all three pesticides .
the percent of loss for the three pesticides were 78% , 55% and 35% for carbaryl , propoxur and pirimicarb respectively .
different parameters such as vapor pressure , boiling point , and suspect ability of the compound to hydrolysis , could be responsible for the losing of pesticides during the cooking process . |
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paliperidone palmitate , a long acting injectable ( lai ) antipsychotic , is the palmitate ester of paliperidone and is widely used in patients with schizophrenia.1 ) as with other antipsychotics , the mechanism of action of paliperidone is unclear.2,3 ) the drug primarily acts as an antagonist of the dopamine d2 and serotonin-2a ( 5ht2a ) receptors . paliperidone also acts as an antagonist of -1 and -2 adrenergic receptors and histamine h1 receptors.1,4 ) paliperidone palmitate lai provides continuous medication with a convenient dosing interval , aimed at enhancing treatment adherence for non - compliant patients with psychotic disorders.5 ) peripheral edema has been reported with atypical anti - psychotics , such as olanzapine , risperidone , ziprasidone , quetiapine , amisulpride , ziprasidone and risperidone lai.611 ) although there is only one published case report of peripheral edema related to paliperidone , to our knowledge , there is no case report in the published literature on peripheral edema associated with paliperidone palmitate lai.12 ) herein , we present the case of a patient who developed pretibial edema after paliperidone palmitate lai treatment . a 36-year - old female unmarried unemployed female patient who lived with her family was diagnosed with paranoid schizophrenia approximately 8 years prior . she had previously received multiple antipsychotics such as haloperidol , olanzapine , risperidone , amisulprid , and quetiapine . at the time of this study , the patient was brought to our outpatient clinic by her family because of worsening delusions , hallucinations and poor treatment adherence . the patient had a history of frequent relapse on oral medication due to non - compliance . because of this poor history of compliance , paliperidone oral tablet was discontinued and the patient was initiated with paliperidone palmitate lai 150 mg on day 1 and 100 mg on day 8 . following a month s usage of paliperidone palmitate lai 100 mg , the patient presented with bilateral pretibial 2 + pitting edema . pretibial edema first occurred at the second week of treatment and then continued to increase gradually . there was no ulceration , erythema , discoloration of the skin , or pain accompanying the edema . complete blood count , thyroid stimulating hormone , vitamin b12 , albumin , liver function tests ( aspartate aminotransferase , alanine transaminase , gamma - glutamyl transpeptidase , albumin ) , renal function tests ( urea , creatinine ) , serum electrolytes and rheumatological tests were performed . the results were found to be within normal limits and the patient had no physical problems . the pretibial edema in this case scored 6 on the naranjo causality scale , suggesting that the adverse effect probably occurred as a result of paliperidone palmitate lai.13 ) treatment with paliperidone palmitate lai was replaced with aripiprazole . the pretibial edema was seen to decrease on the 20th day follow - up . by the end of the second month to the best of our knowledge , this is the first report of pretibial edema accompanying pharmacotherapy with paliperidone palmitate lai . in the literature , there are reported peripheral edema cases associated with antipsychotics.6,8,10 ) paliperidone is the major active metabolite of risperidone and is available as both extended release oral tablet and long - acting injectable suspension for intramuscular use.4 ) there is only one published case report of peripheral edema related to paliperidone . chen et al.12 ) described the development of edema in the dorsal side of the patient s feet 2 weeks after the initiation of treatment with 9 mg paliperidone , which disappeared completely after the discontinuation of paliperidone . again , only one report exists in the literature showing the occurrence of periorbital edema within 14 days of treatment of schizophrenia with 50 mg risperidone lai.7 ) there are several reports on risperidone induced edema either with risperidone alone or in combination with other drugs such as citalopram , sodium valproate and/or benzodiazepines.6,14,15 ) the mechanism of action of antipsychotics that lead to the development of peripheral edema is currently unknown . like risperidone , paliperidone primarily acts by blocking 5ht2 , d2 , -1 , -2 and h1 receptors.1 ) these antipsychotics may cause edema by blocking -1 receptors , resulting in vasodilation , decreased vascular resistance and an increase in hydrostatic pressure in the capillaries.16 ) secondly , 5ht2 receptor blockade can increase cyclic adenosine monophosphate concentration , which relaxes vascular smooth muscle and results in edema.11 ) thirdly , m1 , h1 and 5ht2 receptors blockade can prevent the physiological increase of ip3 , reducing smooth muscle contractility and resulting in vasodilation and edema.17 ) chen et al.12 ) reported that paliperidone and risperidone share common pathogenic pathways that may induce edema . the authors hypothesized that the excipients of paliperidone such as titanium dioxide , propylene glycol and ionic compounds containing sodium can cause the edematous events . pelizza7 ) reported that the peripheral edema associated with risperidone lai may have resulted from the nontherapeutic components of risperidone lai , such as a combination of the polylactide - co - glycolide extended release microspheres and the diluent for parenteral use . interestingly , in our patient , pretibial edema was observed during treatment with paliperidone lai but not during treatment with oral paliperidone . this may be explained by the cumulative period of exposure of the patient to paliperidone with different pharmacokinetic characteristics such as absorption , distribution and metabolism of the medication , depending on the route of administration of the drug . additionally pretibial edema may be associated with the excipients of paliperidone lai such as polysorbate 20 , polyethylene glycol 4000 , citric acid monohydrate , disodium hydrogen phosphate anhydrous , sodium dihydrogen phosphate monohydrate and sodium hydroxide . furthermore , as shown in the current case , the development of edema may take up to two months . in conclusion , the present case suggests that even if peripheral edema does not develop with oral paliperidone , paliperidone lai may cause this side effect . more extensive and comprehensive studies are required to better assess the incidence and mechanism of this adverse effect of long - acting injection antipsychotics . | peripheral edema is observed as an adverse effect of the usage of antipsychotics in the literature .
this case report describes a 36-year - old female patient with the diagnosis of paranoid schizophrenia who presented with pretibial edema following initiation of long - acting injectable paliperidone palmitate .
pretibial edema developed within the second week of treatment and completely disappeared after its discontinuation . |
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sickle cell disease ( scd ) is a genetic hemoglobinopathy , in which hemoglobin , is slightly abnormal . cerebral damage is a serious complication in scd patients , with cerebral infarction ( overt stroke ) secondary to occlusive vasculopathy developing in 5.5 - 17% of patients with scd . conversely the incidence of silent cerebral infarcts ( scis ) lies between 3 - 38% in patients with beta - thalassemia and between 5 - 31% in patients with hbsc being far more common in this patient population . the pathophysiology of cerebrovascular disease in sickle cell anemia is probably related to two processes : i ) intimal hyperplasia and ii ) thrombosis . stroke is attributed to both microvascular and macrovascular lesions . according to the microvascular scenario the small vessels are more likely to be involved in cases of peripheral vaso - occlusive events . in such cases the arteriole and postcapillary venule are the major sites of sicklecell adhesion and subsequent occlusion . the deep white matter , which is perfused by arterioles with sparse anastomoses , is more susceptible to inadequate perfusion and infarction . the deoxygenated , less flexible sickled red blood cells , adhere to the endothelium of capillaries and venules , leading to intravascular stasis and microvascular occlusion . in the macrovascular scenario , scd patients develop chronic anemia resulting in hyperdynamic blood circulation and increased velocity of the blood flow , which has severe consequences on the vessels . repeated red blood cell ( rbc ) adhesion , followed by forcible removal under high - shear forces causes endothelial injury , leading to intimal hyperplasia and endoluminal narrowing . subsequently rbc adherence to damaged vessel walls results in secondary thrombosis , occlusion of the vessel and distal branch emboli . the effects of these lesions depend on the vessel involved , as well as the rapidity of the process . approximately 75% of strokes in scd patients are the result of occlusion of large arteries . however , as stated earlier , the most common form of cns damage in scd patients is silent cerebral infarct ( sci ) . the definition of sci comprises both an abnormal mri with absence of physical findings of overt stroke . surprisingly scd patients will have an incidence of sci up to 27% by age 6 and 37% by age 14 . recent evidence suggests that adult scd patients are prone to develop new and ongoing scis . the latter may on the one hand be the cause for poor intellectual performance in these patients and on the other hand either become progressive sci or overt stroke disease . the first study addressing the issue of sci was undertaken with ct and patients were referred to be suffering a covert stroke . subsequently and after the cooperative study of sickle cell disease ( csscd ) the term sci or silent stroke is preferred . the aim of the study is to find out if there is any correlation between the extent of silent cerebral infartcs ( scis ) depicted on mri with the disease severity in scd adult patients . the study group consisted of 24 consecutive patients ( 11 men , 13 women ) with known scd and a mean age of 38.4 ( range 20 - 59 ) years . the study was approved by the local ethical committee and informed consent was obtained prior to each examination from all patients . all patients included in the study had no evidence of focal neurologic deficit from physical examination . patients with any clinical evidence of neurologic deficit were excluded from the study , as well as patients with contraindications to mr imaging , such as the presence of cardiac pacemakers or metallic implants . the disease was classified according to the number of vaso - occlusive episodes which needed hospitalization per year , the number of transfused rbc units per year , the degree of hemolysis , and the incidence of acute chest syndrome ( acs ) , pulmonary hypertension ( ph ) and acute splenic sequestration ( ass ) . a patient with more than 6 hospitalizations per year and/or for more than 30 days in total , or transfusion - dependency with more than 2 units rbc per month , or with a continuously high degree of intravascular hemolysis , or finally with a history of acs , ph , ass , was considered as severely ill ( grade 2 ) . the disease of a patient with none of the above criteria was characterized as mild ( grade 0 ) , whereas the presence of 3 - 5 hospitalizations per year for less than 30 days or sporadic transfusions as moderately severe ( grade 1 ) . treatment with hydroxyurea was applied to patients with severe and moderately severe form of the disease , and it clearly changed disease severity , dramatically . however , for the needs of this study , the definition of disease severity was based on the patient s history since birth , and it did not take into consideration the last period on hydroxyurea therapy . from these patients ( 50% ) 66.67%(16/24 ) of the patients were on treatment with hydroxyurea , due to history of more frequent or pronounced vaso - occlusive episodes . all patients were submitted to brain mri on a 1 tesla , gyroscan intera , philips mr scanner with a dedicated head coil . mri protocol consisted of tse t2-weighted ( tr 5300 ms , te 120 ms ) and flair ( tr 6000 ms , te 110 ms , ti 2000 ms ) images in the axial and coronal planes . the examinations were reviewed by two radiologists , which were blinded to the clinical results , for evidence of scis ( i.e. lacunar infarcts and leukoencephalopathy ) . lacunar infarcts were defined as focal ( less than 1 cm ) high intensity lesions on the t2-weighted or flair images . leukoencephalopathy was defined as multiple high intensity lesions over 1 cm on the t2-weighted or flair images . score 1 was attributed to small unifocal lesions < 1 cm in diameter or mild extent leukoencephalopathy with lesions measuring <3 cm . score 2 was attributed to more severe / diffuse leukoencephalopathy with lesions > 3 cm . irrespective of their sci score , patients were also evaluated for evidence of brain atrophy , with binary criteria either existing or non - existing . atrophy was defined as lesser volume of brain tissue than the one expected in a healthy person of similar age . the study group consisted of 24 consecutive patients ( 11 men , 13 women ) with known scd and a mean age of 38.4 ( range 20 - 59 ) years . the study was approved by the local ethical committee and informed consent was obtained prior to each examination from all patients . all patients included in the study had no evidence of focal neurologic deficit from physical examination . patients with any clinical evidence of neurologic deficit were excluded from the study , as well as patients with contraindications to mr imaging , such as the presence of cardiac pacemakers or metallic implants . the disease was classified according to the number of vaso - occlusive episodes which needed hospitalization per year , the number of transfused rbc units per year , the degree of hemolysis , and the incidence of acute chest syndrome ( acs ) , pulmonary hypertension ( ph ) and acute splenic sequestration ( ass ) . a patient with more than 6 hospitalizations per year and/or for more than 30 days in total , or transfusion - dependency with more than 2 units rbc per month , or with a continuously high degree of intravascular hemolysis , or finally with a history of acs , ph , ass , was considered as severely ill ( grade 2 ) . the disease of a patient with none of the above criteria was characterized as mild ( grade 0 ) , whereas the presence of 3 - 5 hospitalizations per year for less than 30 days or sporadic transfusions as moderately severe ( grade 1 ) . treatment with hydroxyurea was applied to patients with severe and moderately severe form of the disease , and it clearly changed disease severity , dramatically . however , for the needs of this study , the definition of disease severity was based on the patient s history since birth , and it did not take into consideration the last period on hydroxyurea therapy . from these patients ( 50% ) 66.67%(16/24 ) of the patients were on treatment with hydroxyurea , due to history of more frequent or pronounced vaso - occlusive episodes . all patients were submitted to brain mri on a 1 tesla , gyroscan intera , philips mr scanner with a dedicated head coil . mri protocol consisted of tse t2-weighted ( tr 5300 ms , te 120 ms ) and flair ( tr 6000 ms , te 110 ms , ti 2000 ms ) images in the axial and coronal planes . the examinations were reviewed by two radiologists , which were blinded to the clinical results , for evidence of scis ( i.e. lacunar infarcts and leukoencephalopathy ) . lacunar infarcts were defined as focal ( less than 1 cm ) high intensity lesions on the t2-weighted or flair images . leukoencephalopathy was defined as multiple high intensity lesions over 1 cm on the t2-weighted or flair images . score 1 was attributed to small unifocal lesions < 1 cm in diameter or mild extent leukoencephalopathy with lesions measuring <3 cm . score 2 was attributed to more severe / diffuse leukoencephalopathy with lesions > 3 cm . irrespective of their sci score , patients were also evaluated for evidence of brain atrophy , with binary criteria either existing or non - existing . atrophy was defined as lesser volume of brain tissue than the one expected in a healthy person of similar age . based on clinical criteria 33.33% ( 8/24 ) of the patients had mild , 37.50% ( 9/24 ) moderate and 29.17% ( 7/24 ) severe clinical disease . more specifically 8.33% ( 2/24 ) of them were classified as score 1 and 58.33% ( 14/24 ) were classified as score 2 ( figure 1a - c ) . the rest 33.3% ( 8/24 ) of the patients had no evidence of cerebral lesions on mri and were classified as score 0 . from these 8 patients with no abnormalities on the mri , 25% ( 2/8 ) had mild , 37.50% ( 3/8 ) had moderate and the remaining 37.50% ( 3/8 ) had severe clinical disease . from the 2 patients with score 1 on mri , 50% ( 1/2 ) had moderate clinical disease and 50% ( 1/2 ) had severe clinical disease . from the 14 patients with score 2 on the mri , 42.86% ( 6/14 ) of them had mild , 35.71% ( 5/14 ) moderate and 21.43% ( 3/14 ) had severe clinical disease . atrophy was found in 54.16% ( 13/24 ) of the patients ( figure 2a , b ) . clinical severity and mri score were investigated both through the spearman s correlation coefficient ( rho=-0.227 , p=0.286 ) and with tests for independence ( p=0.72 , chisquare test ) . considering the 38.4 years as the mean age , the patients were subcategorized in two groups : the younger group whose age was under 38.4 years and the older group aged over 38.4 . in the younger age group 35.71% ( 5/14 ) did not show any abnormalities on the mri and were classified as score 0 , no patients were classified as score 1 and 64.29% ( 9/14 ) of them were score 2 . alternatively from the older age group 30% ( 3/10 ) did not exhibit any abnormalities and were classified as score 0 , 20.00% ( 2/10 ) patients were classified as score 1 , whereas 50% ( 5/10 ) were classified as score 2 , on mri . the observed difference in mri scores between the two age groups was not statistically significant ( p=0.216 , chi - square test ) . moreover the mri score was not correlated either with patient gender ( p=0.950 ) or with hydroxyurea treatment ( p=0.230 ) . sickle cell disease ( scd ) is characterized by hemolytic anemia , an increased susceptibility to infections , growth retardation , painful crises , acute chest syndrome and vaso - occlusion . the latter occurs in almost all vascular beds , resulting in ischemia , including the central nervous system , with high morbidity and mortality . outcome is difficult to predict and few effective treatments are available . stroke is a serious complication in patients with sickle cell disease occurring in 5 - 17% , most often between 9 to 15 years of age . there are two main theories for brain infarcts in these patients including both a microvascular scenario and a macrovascular scenario . recent research indicates that scd is also associated with silent cerebral infarcts ( scis ) . the latter is defined as the presence of abnormalities on neuroimaging studies , in the absence of overt clinical symptoms . scis are thought to rather represent small vessel disease but direct evidence is still lacking . furthermore scis seem to affect more the borderline zones and this may suggest hemodynamic reasons implicated in its pathogenesis . mri has proved to be highly sensitive in the identification of scis . due to its sensitivity to detect increased amounts of water within brain tissue at sites of acute and chronic infarctions , especially with fluid - attenuated inversion recovery ( flair ) images , mri has revolutionized the ability to see not only acute symptomatic infarctions , but also scis , especially those within the deep white matter of the centrum semiovale . in our study in contrast in the csscd cohort the prevalence of scis at baseline was 21.2% , while in the sit trial the presence of scis reached a total of 30.8% . both of these studies however have included children and teenagers . the overall percentage of reported sci incidence in the literature is between 3 - 38% in patients with beta - thalassemia and between 5 - 31% in patients with hbsc . scis are known to be more common in homozygous ss patients but they are also known to happen to heterozygous patients like sickle beta - thalassemia and hbsc disease . while one would probably expect a lower overall incidence of scis in our heterozygous scd patients this was surprisingly not the case as the incidence was far greater than that reported in the literature for heterozygous scd adult patients . more specifically a study by kugler et al . found that 50% of the homozygous sdc adult patients ( mean age 20 years ) had scis and of these lesions 16% were at least 1.5 cm . reported an sci incidence of 13% in his study with adult ( 19 - 55 years ) homozygous scd patients . , comprising adult patients with a mean age of 26.99.3 years with both homozygous ss and heterozygous s thal was 20% . studied adult patients younger than 50 years with thalassemia intermedia and homozygous sickle - cell thalassemia . they found scis in 37.5% in patients with thalassemia intermedia and 52% in patients with homozygous sickle cell - thalassemia disease . differences in results between studies may reflect study population differences and differences concerning the study protocol and definitions . there seems to be different ways to define scis and there is no wider consensus . for example in the silent cerebral infarct trial ( sit trial ) it included lesions that were at least 3 mm in their greatest dimension and visible in at least two planes on the t2-weighted images . evaluating adult patients the definition of sci included lesions of at least 5 mm , hyperintense on the t2-weighted image but had to be hypointense on the t1-weighted image . according to zhu et al . discrepancies in the literature are mainly due to : i ) differences in mr parameters , like slice thickness , ii ) different criteria for sci definition and iii ) different criteria for defining dilated virchow - robin spaces ( dvrs ) . while a high percentage 58.33% ( 14/24 ) of our patients had a high mri score of 2 , only a small percentage 29.17% ( 7/24 ) of these patients had severe disease according to clinical criteria . found that brain damage was more severe in patients with sickle cell - thalassemia disease who had more vaso - occlusive episodes per year . this was not the case in our study , where the extent of scis was not well - correlated with clinical severity . however according to other previous studies , various imaging techniques reveal a more extensive and diffuse pattern of vascular involvement than the one expected from the patient s clinical presentation . this may imply the silent , insidious nature and complex pathophysiology of microvascular disease in scd patients . in the study of vichinski et al . lacunae ( lesions 5 mm ) were seen in 13% and white matter lesions in 15% and in our study lacunar lesions ( 5 - 9 mm ) were seen in 8.33% and larger lesions ( > 10 mm ) in 58.33% . although the study design is different it seems that while moderate mri disease scores are rather similar in both studies , severe mri disease scores are much higher in our study . in our study younger and older patient groups did not show statistically significant differences in the extent of scis . more specifically a high percentage 64.26 % ( 9/14 ) from the patients of the younger age group ( < 38.4 years ) showed scis and interestingly all of them were graded score 2 , which would be unusual for the general population of similar age . in the older ( > 38.4 years ) age group patients exhibited scis in a total percentage of 70% ( 7/10 ) , whereas 20% ( 2/10 ) were score 1 and 50% ( 5/10 ) were scored 2 on mri . in contrast , manrfe et al . reported that in thalassemia intermedia patients , the frequency of brain damage increased with age . previous estimates have shown that the incidence of sci with mri in the general population is between 5.8% and 17.7% . more specifically the incidence of these hyperintensities has been 15.6% and 10.7% in two large studies . the framingham offspring study found an incidence or 10.7% , using 1 tesla mri and 4 mm section thickness in a patient population with a mean age of 629 years . on the other hand the incidence was 15.6% in the helsinki aging brain study , using patients between 55 and 85 years and a 0.2-t mri and 10 mm section thickness . in our case this may , however be counteracted by the fact that the younger patient s group < 38.4 years still showed a significant extent of scis . the fact that there was no statistically significant difference between the two age groups , in our patients , implies that scis in adult heterozygous scd patients may not be age - related and may start and accumulate from younger ages . this may be of clinical significance , as presence of scis may result in further brain damage as they are known to progress to symptomatic disease . in fact results from the csscd study indicate that sci poses further risk for extra neurological damage and a higher risk for both progressive sci or overt clinical stroke . furthermore , brain atrophy was commonly observed in our patients , as it occurred in 54.16% ( 13/24 ) of the whole population studied . our results show a higher rate of brain atrophy than those reported in the literature . silva et al reported a lower incidence of brain atrophy ( 28% ) in sdc patients , aged 16 years and above . in the manfre study atrophy was detected in 31% in the thalassemia intermedia group , and reached 51% in the homozygous scd group . in the study by vichinski et al . atrophy in scd patients is considered to suggest a chronic underlying process . in the literature , the risk factors for sci are reported in the csscd study as being history of seizures , pain event rate , elevated wbc and the sen beta globin gene haplotype . in the sit trial risk factors for sci included : hemoglobin < 7.6 g / dl , systolic blood pressure > 113 mmhg and male gender . in our study hydroxyurea and hematopoietic stem cell transplantation are considered for primary and secondary prevention of sci but there is still no high level evidence to support this . in our study there were no statistically significant differences in the extent of scis between patients receiving or not receiving hydroxyurea . some other study limitations in comparison to other studies are : the rather small number of patients studied and the use of 1 t mri scanner instead of the 1.5 t used in other studies . according to the field strength and study parameters ( slice thickness of 5 mm ) the smallest lesions that could be picked - up , would be around 5 mm . we did not use lesion subcategorization for lesions < 10 mm but instead of it all lesions > 5 mm and < 10 mm were given score 1 . this may have made it more difficult to compare our results with those of larger studies . the clinical criteria for disease severity were rather qualitative , but all patients were on follow - up for many years , as the thalassemia and hemoglobinopathies unit of our hospital is a reference center for scd patients . concerning the incidence of brain atrophy it should be noted that the definition is rather subjective and this may also create bias when comparing with previous studies . this is reflected in previous studies with the reported incidence being between 23 - 51% finally it should be noted that although it is known that sci may occur in infants just 1 year of age and continue during childhood , reaching maximum incidence rate in children around 6 years of age , little is known about the natural history of scis in the adult , which is not thoroughly understood . our study may further contribute to the understanding of the disease in the adult as it is one of a few focusing on heterozygous sickle - cell/-thal adult patients . in contrast to larger cohorts that focus on homozygous scd patients of school - age or teenager patients . in conclusion , the extent of silent cerebral infarcts on mr imaging in compound heterozygous ( hbs/-thal ) adult patients is not correlated with the severity of clinical disease . moreover the extent of scis on mr imaging in these patients is not age - related and may be quite severe even in the younger ones . | the aim of this paper is to correlate the extent of silent cerebral infarcts ( scis ) on magnetic resonance imaging ( mri ) with the clinical severity of sickle cell disease ( scd ) in adult patients .
twenty - four consecutive adult asymptomatic scd patients ( 11 male and 13 female ) with a mean age of 38.4 years ( range 20 - 59 ) were submitted to brain mri on a 1 tesla gyroscan intera , philips mr scanner with a dedicated head coil . the protocol consisted of tse t2-weighted and flair images on the axial and coronal planes .
mri readings were undertaken by two radiologists and consensus readings .
patients were compound heterozygotes ( hbs/-thal ) .
the extent of scis was classified from 0 - 2 with 0 designating no lesions .
clinical severity was graded as 0 - 2 by the hematologist , according to the frequency and severity of vaso - occlusive crises .
there was no statistically significant correlation between the severity of clinical disease and the extent of scis on mr imaging .
the extent of sci lesions did not differ statistically between younger and older patients .
patients receiving hydroxyurea had no statistically significant difference in the extent of sci lesions .
the extent of scis in heterozygous ( hbs/-thal ) scd patients is not age related and may be quite severe even in younger ( < 38.4 years ) patients
. however the extent of scis is not correlated with the severity of clinical disease . |
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some diversity in the surgical management of gctds still exists among neurosurgeons . here , we present 18 cases of gctds , which represents the second largest cohort in the literature , and we describe the trench vertebrectomy via a thoracotomy as a safe and effective surgical technique . twenty - nine patients who underwent surgical treatment in our unit for herniated thoracic disks between the years 2000 and 2013 were reviewed . following radiologic review , 18 patients were found to have gctds as defined by hott et al.6 retrospective data was collected on patient demographics , presentation , operative details , and imaging findings including the fusion rates . the modified japanese orthopaedic association ( mjoa ) score was used to assess the outcomes . there was a female preponderance in our series with a male - to - female ratio of 4:14 . the median age at diagnosis was 51.2 years with a range of 37 to 70 years . the mean duration of presenting symptom(s ) was 9.9 months , with a range of 2 weeks to 3 years . seventeen ( 94.4% ) patients in this series presented with signs and/or symptoms of myelopathy , with the remaining 1 ( 5.6% ) patient presenting with symptoms more in keeping with a radiculopathy . the commonest presenting symptom was lower limb weakness in 66.7% of the series , and the commonest clinical sign was hyperreflexia in 83.3% ( table 1 ) . all patients in our center had both a preoperative computed tomography ( ct ) scan and mri ( figs . 1 and 2 ) . a ct scan defines the degree of calcification , disk morphology , and aids in preoperative planning ( fig . the commonest affected level in our cohort was at t8t9 in 6 patients ( 33.3% ) , followed by t7t8 in 22.2% ( table 1 ) . ( a ) preoperative axial computed tomography ( ct ) showing the giant calcified thoracic disk ( gctd ) within the spinal canal . ( b ) ( a ) preoperative axial magnetic resonance imaging ( mri ) of the same disk space as fig . 1 . ( b ) the access was through a thoracotomy in all but one patient , in whom a costotransverse approach was used . the ultrasonic bone cutter was used to facilitate the vertebrectomy in the last three patients in our series . the patient was positioned in the lateral position with the side of the thoracotomy uppermost . for localization of the affected level the thoracotomy was usually from the left side , because the chest cavity is larger on this side and it is also easier to mobilize the aorta than the great veins . in the upper thoracic levels ( above t5 ) , a right - sided thoracotomy may be used to avoid the arch of the aorta . to gain access to the affected level the lung was collapsed at this stage and the pleura opened . once the affected level was identified and confirmed with an x - ray , the parietal pleura was opened over the vertebral bodies below and above and reflected away from the spine . to allow better visualization of the lateral disk , the costovertebral and costotransverse ligaments were divided , and up to 2 cm of rib head were removed with bone nibblers or a drill if necessary . a trench vertebrectomy was performed with half of vertebra on either side excised ; it was not necessary to extend this procedure to remove the anterior part of the vertebral body ( figs . 3 and 4 ) . the ultrasonic bone cutter was useful here as an alternative to piecemeal resection or using a pneumatic drill . the groove was deepened until enough bone was removed to access the disk and normal dura on either side ( fig . ( a ) intraoperative image using the misonix bone cutter ( msb ; misonix , farmingdale , new york , united states ) to drill the giant calcified thoracic disk . ( b ) intraoperative image of the bone graft being placed . ( a ) postoperative axial computed tomography ( ct ) outlining decompression using the trench technique . the base of the disk was eased away from the posterior longitudinal ligament , a layer of which may be left if adherent to the dura . after complete excision , the dura will move forward . in 8/18 cases ( 44.4% ) , the disk material was noted to be adherent to the cord . in such instances , a remnant layer of disk 6a , b. it shows fragments of calcified material within the degenerate disk associated with peripheral ossification . fifteen ( 83.3% ) patients experienced a postoperative improvement of at least 1 point in the mjoa score on last follow - up . six ( 33.3% ) patients required a blood transfusion during their hospital stay , which ranged from 2 to 5 u. one ( 5.6% ) patient was transferred to the intensive care unit for ventilatory support due to development of adult respiratory distress syndrome . the average length of stay was 10.7 days , with a range of 4 to 35 days . the mean duration of follow - up was 19.8 months ( range 7 months to 2 years ) . postoperative imaging was performed on all patients but was heterogeneous in modality including plain films , ct scans , or mris at 6 to 12 months . fusion was reported in all patients on the follow - up radiology reports ; the criteria used were the documentation of bony trabeculation and absence of bony lucency at the graft / vertebral end plate interface ( fig . the postoperative imaging was not standardized compared with the preoperative imaging , and formal sagittal alignment measures were not calculated ; however , no patient developed a clinically or radiologically evident kyphotic deformity . all the bluish areas in right side of the slide represent calcium ; this tissue has been decalcified to be able to cut it , hence the rest of the right side is pink . there was a female preponderance in our series with a male - to - female ratio of 4:14 . the median age at diagnosis was 51.2 years with a range of 37 to 70 years . the mean duration of presenting symptom(s ) was 9.9 months , with a range of 2 weeks to 3 years . seventeen ( 94.4% ) patients in this series presented with signs and/or symptoms of myelopathy , with the remaining 1 ( 5.6% ) patient presenting with symptoms more in keeping with a radiculopathy . the commonest presenting symptom was lower limb weakness in 66.7% of the series , and the commonest clinical sign was hyperreflexia in 83.3% ( table 1 ) . all patients in our center had both a preoperative computed tomography ( ct ) scan and mri ( figs . 1 and 2 ) . a ct scan defines the degree of calcification , disk morphology , and aids in preoperative planning ( fig . the commonest affected level in our cohort was at t8t9 in 6 patients ( 33.3% ) , followed by t7t8 in 22.2% ( table 1 ) . ( a ) preoperative axial computed tomography ( ct ) showing the giant calcified thoracic disk ( gctd ) within the spinal canal . ( a ) preoperative axial magnetic resonance imaging ( mri ) of the same disk space as fig . 1 . the access was through a thoracotomy in all but one patient , in whom a costotransverse approach was used . the ultrasonic bone cutter was used to facilitate the vertebrectomy in the last three patients in our series . the patient was positioned in the lateral position with the side of the thoracotomy uppermost . for localization of the affected level the thoracotomy was usually from the left side , because the chest cavity is larger on this side and it is also easier to mobilize the aorta than the great veins . in the upper thoracic levels ( above t5 ) , a right - sided thoracotomy may be used to avoid the arch of the aorta . to gain access to the affected level once the affected level was identified and confirmed with an x - ray , the parietal pleura was opened over the vertebral bodies below and above and reflected away from the spine . to allow better visualization of the lateral disk , the costovertebral and costotransverse ligaments were divided , and up to 2 cm of rib head were removed with bone nibblers or a drill if necessary . a trench vertebrectomy was performed with half of vertebra on either side excised ; it was not necessary to extend this procedure to remove the anterior part of the vertebral body ( figs . 3 and 4 ) . the ultrasonic bone cutter was useful here as an alternative to piecemeal resection or using a pneumatic drill . the groove was deepened until enough bone was removed to access the disk and normal dura on either side ( fig . 3 ) .
( a ) intraoperative image using the misonix bone cutter ( msb ; misonix , farmingdale , new york , united states ) to drill the giant calcified thoracic disk . ( b ) ( a ) postoperative axial computed tomography ( ct ) outlining decompression using the trench technique . ( b ) postoperative sagittal ct clearly delineating the trench vertebrectomy . at this stage , the base of the disk was eased away from the posterior longitudinal ligament , a layer of which may be left if adherent to the dura . after complete excision , the dura will move forward . in 8/18 cases ( 44.4% ) , the disk material was noted to be adherent to the cord . in such instances , a remnant layer of disk 6a , b. it shows fragments of calcified material within the degenerate disk associated with peripheral ossification . fifteen ( 83.3% ) patients experienced a postoperative improvement of at least 1 point in the mjoa score on last follow - up . six ( 33.3% ) patients required a blood transfusion during their hospital stay , which ranged from 2 to 5 u. one ( 5.6% ) patient was transferred to the intensive care unit for ventilatory support due to development of adult respiratory distress syndrome . the average length of stay was 10.7 days , with a range of 4 to 35 days . the mean duration of follow - up was 19.8 months ( range 7 months to 2 years ) . postoperative imaging was performed on all patients but was heterogeneous in modality including plain films , ct scans , or mris at 6 to 12 months . fusion was reported in all patients on the follow - up radiology reports ; the criteria used were the documentation of bony trabeculation and absence of bony lucency at the graft / vertebral end plate interface ( fig . the postoperative imaging was not standardized compared with the preoperative imaging , and formal sagittal alignment measures were not calculated ; however , no patient developed a clinically or radiologically evident kyphotic deformity .
delayed postoperative sagittal computed tomography showing fusion at t8t9 . all the bluish areas in right side of the slide represent calcium ; this tissue has been decalcified to be able to cut it , hence the rest of the right side is pink . gctds are rare and present a technical neurosurgical challenge . because of their size and degree of calcification , a significant proportion of gctds display some degree of thecal involvement , with penetration into the subarachnoid space via tears of the posterior longitudinal ligament and the dura mater.7
8 presentation may be with clinical signs and symptoms of a compressive myelopathy , radiculopathy , and/or back pain.1
6
8
9
10
in our series , the median age and female preponderance is a finding echoed in other studies,11
12
13
14 which is speculated to be secondary to altered calcium - phosphate metabolism that occurs postmenopausally.8 the pathophysiology of the calcification process is also poorly understood , although it has been suggested that calcium deposition originates from the posterior longitudinal ligament with extension into the disk space.15 all of the lesions in our cohort occurred at the lower thoracic levels , which is consistent with the findings by other authors.1
6
8
10 it is estimated that 75% of herniated thoracic disks occur below the level of t8 , which is a probable combined consequence of the greater weight load in the lower thoracic spine and its greater range of movement.6
12
14
16
myelopathy was present in 17 ( 94.4% ) of our patients . the other large case series in the literature describing gctds reported similar high rates of myelopathy,1
6
8
10 with hott et al comparing these high rates to 47% in smaller , nongiant thds.6
various surgical techniques and approaches to manage gctds are advocated in the literature . the choice of surgical approach has evolved over the last century and has included laminectomy , transpedicular , costotransversectomy , and transthoracic methods . posterior approaches are more or less obsolete owing to extremely poor results , particularly from laminectomy.7
17
18
19
20 despite the refinement of several approaches , the optimal operative treatment for thoracic disk herniations remains open to debate and may in part be dictated by the skills of the operating surgeons , whether the disk is central , and the presence of calcification.4
14
17
21 in their review , mulier and debois compared 7 of their own patients between 1986 and 1993 with 384 others to compare transthoracic , lateral , or posterolateral approaches to thds ; their results illustrated that transthoracic surgeries were associated with better neurologic outcomes.21
most authors advocate an open thoracotomy approach , in particular for centrally located calcified disk herniations.1
4
6
21
22 technical variations in the anterior transthoracic approach exist , with moran et al recommending a mini - open retropleural transthoracic approach with comparable results,10 although some authors advocate a two - level vertebrectomy followed by instrumented fusion.6
22 barbanera et al suggest using a mini - vertebrectomy , although instrumentation was still required in 2 ( 28.5% ) cases of their series.1
in our view , a trench vertebrectomy has the advantage of visualizing the dura on either side of the herniated disk , allowing a safe excision , especially in the cases of a broad , central , heavily calcified disk prolapse with severe cord compression . in addition , this technique enables minimal bone removal compared with a complete two - level vertebrectomy . a costotransverse approach may be an alternative and can be utilized if the anatomy of the prolapsed disk is suitable ( e.g. , a more lateral cord compression ) and good visualization of the anterolateral dura above and below the compressed segment can be safely attained . it enables direct visualization of the ventral spinal column , which is essential in anterior spinal pathologies such as gctds . the approach enables the greatest access and avoids manipulation of the spinal cord , which is already compromised in gctds.23
24 a left - sided thoracotomy evades the vena cava and liver on the right , though a right - sided approach may be undertaken if the pathology is predominately on that side or to evade poor visualization from structures such as the aortic arch around the level of t3 and t4.24 disadvantages of this approach are that it may require a longer hospitalization and longer intensive care stay than with other procedures , with a greater risk of pulmonary complications ; also , postoperative pain may be greater.4
17
21
thoracic excision of gctds is known to be associated with possible significant blood loss,10 particularly where vertebrectomy is conducted,25 which can mandate the ligation of two radicular arteries above and below the disk space . the use of the bone cutter in our experience allows all radicular arteries to be preserved , even those at the disk space level , thereby reducing the risk of spinal cord stroke secondary to ligation of the radicular artery of adamkiewicz . spinal cord injury and dural tears are also less likely due to the selective tissue - cutting properties of the ultrasonic bone cutter . bone is cut in preference to soft tissue , which responds elastically in contact with the blade and therefore moves and vibrates , causing dissemination of the energy.26
27
28
the complications associated with anterior approaches to the spine vary depending upon the specific approach , surgical indication , and patient factors . a large meta - analysis by mcdonnell et al reviewed 447 patients undergoing anterior surgery to the thoracic , thoracolumbar , or lumbar spine.25 indications included correction of deformity , tumor , vertebral osteomyelitis , or diskitis . the authors ' aim was to determine the perioperative complications , which were divided by the authors into major and minor as per table 3 . others have also used this system to report their complication rates.22 in a large meta - analysis of thoracic spine surgery , fessler and sturgill reported a complication rate of 11% in transthoracic approaches.17 ayhan et al reported a complication rate of 21.4% , with two of these complications arising secondary to instrumentation.22
in utilizing the criteria for perioperative complications from mcdonnell et al,25 we report a major complication rate of 5.6% with one of our patients being intubated and ventilated for adult respiratory distress syndrome . minor complications occurred in 2 ( 11.1% ) patients ; both had an intraoperative cerebrospinal fluid leak , which was repaired at the time with no sequelae . intraoperative blood transfusion occurred in 6 ( 33.3% ) patients , which is an expected complication of transthoracic surgery with vertebrectomy as previously mentioned . in our series , 15 ( 83.3% ) patients had a functional improvement of at least one grade on the mjoa scoring system , with the remaining patients ' mjoa scores remaining unchanged . these results compare favorably with those from the other gctd series.1
6
10
22
none of the major series on gctds reported on fusion rates or development of kyphosis.1
6
10
22 in our case cohort , all patients had evidence of fusion on postoperative imaging as defined by documentation of bony trabeculation and absence of bony lucency at the graft / vertebral end plate interface on radiology reports . no patient developed a clinically or radiologically evident kyphotic deformity on follow - up ; we acknowledge the limitations of our cohort study in this regard as cobb angles were not recorded and compared pre- and postoperatively . however , others have also noted that when the amount of bone removed is minimal in an anterior approach , it is not likely to produce frank spinal instability , and instrumentation is not required in these cases.4
10 better long - term data from cohort studies is required to make firm conclusion . thoracic vertebrectomy for a variety of indications have reported worsening of the kyphotic angle postoperatively , albeit the change was minimal.29
30
31
32 a trench vertebrectomy via a thoracotomy allows safe identification of normal dura on either side of the compressed segment prior to performing a diskectomy . a rib autograft has achieved 100% fusion rates with no kyphotic deformities reported in our series . overall , 83% of patients showed postoperative improvement in their mjoa score with none of our cohort experiencing neurologic decline . | study design
case series and review of the literature.objective
to review the management of giant calcified disks in our large cohort and compare with the existing literature .
we discuss our surgical technique.methods
twenty - nine cases of herniated thoracic disk between 2000 and 2013 were reviewed .
eighteen patients were identified as having giant calcified thoracic disks , defined as diffusely calcified disks occupying at least 40% of the spinal canal .
demographic data was collected in addition to presentation , imaging findings , operative details , and outcomes using the modified japanese orthopaedic association ( mjoa ) scale.results
giant calcified thoracic disks ( gctds ) are unique clinical entities that require special neurosurgical consideration owing to significant ( 40% ) involvement of the spinal canal and compression of the spinal cord , often leading to myelopathy .
the median age at diagnosis was 51.2 years ( range 37 to 70 ) with the mean duration of presenting symptoms being 9.9 months ( range 2 weeks to 3 years ) .
seventeen ( 94.4% ) patients presented with at least one sign of myelopathy ( hyperreflexia , hypertonia , bladder or bowel dysfunction ) with the remaining 1 ( 5.6% ) patient presenting with symptoms in keeping with radiculopathy .
thoracotomy was performed on 17 ( 94.4% ) patients , and 1 ( 5.6% ) patient had a costotransverse approach .
mean follow - up was 19.8 months ( range 7 months to 2 years ) .
mjoa score improved in 15 ( 83.3% ) patients .
mjoa scores in the other patients remained stable.conclusions
gctds are difficult neurosurgical challenges owing to their size , degree of spinal cord compression , and consistency . we recommend a trench vertebrectomy via a thoracotomy in their surgical management .
this procedure safely allows the identification of normal dura on either side of the compressed segment prior to performing a diskectomy .
excellent fusion rates were achieved with insertion of rib head autograft in the trench . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
a nucleosome contains 146 bp of dna wrapped twice around an octamer composed of two copies of each histone protein : h2a , h2b , h3 , and h4 ( kornberg , 1974 ; kornberg and lorch , 1999 ) . nucleosomes are observed as a series of beads on a string via electron microscopy with the beads being the individual nucleosomes connected by the linker dna , the string . structural studies have demonstrated that histone n - terminal tails protrude outward from the nucleosome and can be posttranslationally modified by different enzymes ( luger et al . , 1997 ; such posttranslational modifications of histone tails include phosphorylation , acetylation , sumoylation , adp ribosylation , ubiquitination , and methylation . the first h3k4 ( histone h3 lysine 4 ) methylase , set1/compass , was isolated from saccharomyces cerevisiae and was demonstrated to be capable of mono- , di- , and trimethylating h3k4 ( miller et al . , 2001 this posttranslational modification of h3k4 by compass requires prior h2bk123 ( histone h2b lysine 123 ) monoubiquitination in yeast and h2bk120 in vertebrates , which is a process known as histone cross talk ( dover et al . , 2002 ; sun and allis , 2002 ; wood et al . , 2005 it has also been demonstrated that histone h3k79 methylation by dot1 also requires h2bk123 monoubiquitination ( briggs et al . , 2002 ; wood et al . , 2003a ) . monoubiquitination of histone h2bk123 is mediated by the macromolecular complex containing the e2-conjugating enzyme rad6 and the e3 ligase bre1 in s. cerevisiae ( robzyk et al . , 2000 ; hwang et al . , 2003 ; wood et al . , 2003a ) . this modification has been linked to transcriptional activation and elongation ( henry et al . , 2003 ; kao et al . , 2004 ; xiao et al . , 2005 ; pavri et al . , 2006 ; shilatifard , 2006 ; tanny et al . , 2007 ; kim et al . , 2009 ) studies in other eukaryotic organisms also have confirmed that this mode of regulation is well conserved from yeast to human ( pavri et al . however , a recent study performed by foster and downs ( 2009 ) argued that monoubiquitination of h2bk123 in yeast is not the sole determinant for the methylation of h3k4 and h3k79 . in their study , foster and downs ( 2009 ) observed the presence of both h3k4 and h3k79 trimethylation in the h2bk123r mutant strain derived from an fy406 parental strain , which is contradictory to the previous findings , whereas both modifications were lost in an h2bk123r mutant derived in the y131 background . surprisingly , a triple kss mutant ( h2bk123r - s125a - s126a ) in the fy406 background resulted in the loss of trimethylation of both h3k4 and h3k79 . additionally , foster and downs ( 2009 ) demonstrated that flag - tagged h2b containing the k123r mutation ( flag - h2bk123r ) abolished the trimethylation of h3k4 and h3k79 , whereas untagged h2bk123r still possessed normal levels of trimethylation of both of the lysine residues in strain fy406 . these observations led them to conclude that monoubiquitination of histone h2b alone is not required for the trimethylation of either h3k4 or h3k79 , but that an additional unknown alteration to h2b or the mutation in the genome in combination with the k123 mutation caused the loss of histone h3k4 and h3k79 trimethylation . in this study , we have addressed a possible role for the flag tag on histone h2b in the regulation of h3k4 and h3k79 methylation . our collective experiments have demonstrated that histone h3k4 and h3k79 methylation is solely dependent on h2b monoubiquitination and is independent of any other unknown factors or genetic backgrounds . to further investigate the role of h2bk123 monoubiquitination in h3k4 and h3k79 trimethylation and a possible role for the flag epitope of h2b in this process , we set out to analyze the role of flag - less h2b mutated in k123 for h3k4 and h3k79 methylation in several different strain backgrounds . we first wanted to be able to detect the presence of monoubiquitinated h2b in a flag - less h2b background because the observation made by foster and downs ( 2009 ) that h3k4 trimethylation could be detected in a flag - less h2bk123r strain could be because of the inadvertent presence of a wild - type copy of h2b . to address this 1 a ) and then shuffled a plasmid expressing h2b lacking an n - terminal flag tag ( fig . 1 b ) into several different backgrounds , including y131 , fy406 , dy20d , and jhy205 . we analyzed the trimethylation levels of both h3k4 and h3k79 as well as the monoubiquitination of h2b in these backgrounds ( fig . our antibody generated in this study is specific for monoubiquitinated h2b in yeast and is capable of detecting this modification in the presence or absence of a flag tag ( figs . 1 a and 2 a ; and see figs . 3 b and 5 b ) . when this antibody was used , flag - less wild - type h2b showed a band migrating slightly faster than that of flag - tagged wild - type h2b as a result of the deletion of the flag tag ( fig . however , no monoubiquitinated form of h2b was detected from k123a mutants in both y131 and fy406 backgrounds and k123r mutants in dy20d and jhy205 , regardless of the attachment of a flag tag to h2b ( fig . 2 , a and b ) . furthermore , we tested these strains for the methylation of histone h3k4 and h3k79 ( fig . 2 ) . regardless of the presence or absence of a flag tag on h2b , h3k4 methylation was detected in strains bearing wild - type h2b but not in strains in which h2b carries a point mutation at the k123 residue . similar strains were generated independently in all of our laboratories , and we have each confirmed that there is no difference between flag - tagged h2b and untagged h2b and that all of the histone modifications are consistent among the different strain backgrounds , including the fy406 background used in the study performed by foster and downs ( 2009 ; fig . , we conclude that h3k4 and h3k79 trimethylation is indeed solely dependent on h2b monoubiquitination regardless of an additional alteration to the h2b sequence or strain backgrounds . ubiquitinated h2b specific antibodies generated in rabbit were affinity purified and characterized by western blot analysis of whole cell extracts prepared from wild type ( wt ) , h2bk123a , ubp8 , and rad6. both wild - type and ubp8 strains show the presence of monoubiquitinated h2b ( 24 kd and 23 kd , respectively ) ; however , an increased amount of the ubiquitinated form of h2b was observed in the ubp8 strain as expected . no bands were detected in either h2bk123a or the rad6 strains , showing that our h2bub antibody is capable of specifically recognizing ubiquitinated h2b in yeast . psn888 was generated from pzs145 by deletion of the n - terminal flag tag ( fg ) from the htb1 gene . similarly , the flag on h2b was removed in prs315-hht2-hhf2-hta1-flaghtb1 to generate h2b flag - less strains . di- and trimethylation of histone h3k4 and trimethylation of h3k79 are dependent solely on monoubiquitination of h2bk123 . ( a ) western blotting of whole cell extracts from strains transformed with a plasmid carrying wild - type ( wt ) h2b or mutant h2bk123 ( k123a ; k123r ) either with or without an n - terminal flag tag . cell extracts were prepared from wild - type h2b or h2bk123 mutants in three different strain backgrounds and were subjected to sds - page and analyzed by western blot analysis with antibodies to dimethyl h3k4 , trimethyl h3k4 , trimethyl h3k79 ( h3k4me2 , h3k4me3 , and h3k79me3 ) , monoubiquitinated h2bk123 ( h2bub ) , or flag ( flag ) . the calculated molecular masses of h2b , ubiquitinated h2b , and h3 are 14 kd , 23 kd , and 15 kd , respectively . the calculated molecular masses of flag - tagged h2b and ubiquitinated , flag - tagged h2b are 15 kd and 24 kd , respectively . ( b ) western blotting of whole cell extracts from strains transformed with plasmid carrying wild - type or mutant h2bk123r in fy406 background . the y131 strain was originally generated for plasmid shuffling of h2a - h2b genes , as a simultaneous deletion of the hta1-htb1 and hta2-htb2 loci is lethal ( robzyk et al . , 2000 ) . the genotype of y131 describes that hta1-htb1 was replaced with leu2 , whereas hta2-htb2 was replaced with ura3 in the presence of a his3 hta2-htb2 plasmid . after selection for tight 5-foa resistance , a ura3 plasmid carrying hta1-htb1 was substituted for the his3-hta2-htb2 plasmid , thus creating the shuffle strain that was used to introduce a his3-marked plasmid containing hta1 and flag - htb1 . the y131 strain expressing flag - tagged wild - type h2b under either glucose or galactose media has normal levels of h2b monoubiquitination and methylation of h3k4 and k79 ( fig . when flag - h2bk123r in the y131 background was grown continuously in a galactose - containing media , the methylation of both of the h3 residues was present at levels close to those in the wild - type cells ( fig . 3 a , lanes 710 ) . because the n terminus of h2bk123r was tagged with flag , we tested for the presence of monoubiquitination using a flag antibody . there was no slower - migrating band in the histone h2bk123r bearing strain , indicating that there is no monoubiquitinated form of flag - h2b ( fig . the h2a / h2b shuffle strain y131 contains a galactose - regulated copy of hta2-htb2 genes on chromosome ii . ( a ) western blot analysis of whole cell extracts prepared from flag - tagged wild - type h2b and flag - h2bk123r grown under either glucose or galactose media . cell extracts were subjected to sds - page and western analysis with antibodies specific to h3k4me2 , h3k4me3 , h3k79me3 , and flag . ( b ) western analysis of whole cell extracts prepared from wild - type ( wt ) by4742 ( fm392 ) and its derivative rad6 , flag - h2b ( y131 background ) , and flag - h2bk123r ( y131 background ) . cell extracts were subjected to sds - page and western analysis with antibodies to trimethyl h3k4 ( h3k4me3 ) , monoubiquitinated h2bk123 ( h2bub ) , or flag ( flag ) . the h2b monoubiquitination specific antibody detected a faster migrating band , which is indicated by red arrows . this band represents an untagged version of h2b , which is only seen in y131 when cells are grown in galactose - containing media . blue arrows indicate the slower migrating flag - tagged , monoubiquitinated h2b seen under both dextrose and galactose growth conditions only in wild - type cells and not h2bk123r . black lines indicate that intervening lanes have been spliced out . because h2b monoubiquitination is required for the methylation of these lysine residues , we suspected that one of the two genomic htb genes was not deleted in y131 and that the genomic wild - type h2b was expressed in the k123r mutant strain in a galactose - dependent manner . we tested this possibility by using our antibody specific to monoubiquitinated histone h2b ( fig . in wild - type y131 under glucose media , the flag - tagged wild - type h2b produced only an upper band ( flag - tagged , monoubiquitinated h2b ; fig . however , in wild - type y131 under galactose media there are two bands ( fig . 3 b , lane 6 ) . the upper band is flag - tagged , monoubiquitinated histone h2b , and the lower band is flag - less , monoubiquitinated histone h2b ( fig similarly , the flag - h2bk123r strain shows no h2b monoubiquitination under the glucose media ( fig however , the flag - tagged h2bk123r strain grown under galactose media shows the band corresponding to flag - less , monoubiquitinated histone h2b ( fig we have also detected the expression of wild - type h2b under the galactose condition when using h2b - specific antibodies ( unpublished data ) . this suggests that y131 contains a galactose - inducible version of wild - type h2b somewhere in the genome . to understand the basis for these results between the two growth conditions , we sequenced the genomic regions around the deleted hta1-htb1 ( chromosome iv ) and hta2-htb2 ( chromosome ii ) genes in strain y131 . we found that intact hta2 and htb2 genes were still present on chromosome ii in the y131 strains , and surprisingly , the bidirectional gal1/10 promoter was inserted between the two genes to exactly replace the bidirectional hta2-htb2 promoter ( fig . 4 ) . the gal10 promoter drives the expression of hta2 , and gal1 drives the expression of htb2 . the presence of the gal1/10 promoter explains why wild - type h2b was expressed only under the galactose media and not under the glucose media . it remains somewhat of a mystery how gal1/10 was inserted exactly between the two genes in the first place , although the his3 plasmid in the original y131 strain carried a gal1/10-regulated hta2-htb2 locus that may have been incorporated into the genome by a rare recombination event . because y131 is a widely used strain , mutants that affect gal1/10 transcription may not appropriately repress wild - type hta2-htb2 in glucose in this strain , and data obtained from its usage under prolonged growth in galactose conditions must be interpreted carefully . studies using this strain to analyze effects of the h2b - k123r mutation on gal1 transcription were performed for short periods of galactose induction ( 2 h ) when the expression of the genomic hta2-htb2 genes was not detected , and the transcription results have been recapitulated in other strain backgrounds ( xiao et al . , 2005 ; unpublished data ) . in the y131 strain , the gal1/10 promoter is inserted between hta2 and htb2 on chromosome ii . although this region was deleted and replaced with ura3 in the y131 strain , a wild - type copy of hta2-htb2 controlled by the gal1/10 promoter is present in this region . finally , we have now generated a histone h2a and h2b alanine mutant library in the fy406 background ( fig . 5 a ) in addition to the library we reported earlier in y131 ( nakanishi et al . , 2008 ) , as one may wish to use this collection under galactose conditions for different genetic and biochemical screens . from our new library in fy406 , we found three h2a ( y58 , e62 , and d91 ) residues and one h2b ( l109 ) residue that are essential for viability . these data are in agreement with our previous published results in y131 ( nakanishi et al . , 2008 ) . furthermore , we tested our new library for h3k4 methylation and h2b monoubiquitination and identified the same residues in y131 , specifically those that regulate normal levels of h3k4 methylation in fy406 , as well ( fig . these residues include four residues within h2a ( glu65 , leu66 , asn69 , and asp73 ) and three residues within h2b ( his 112 , arg119 , and lys123 ) . thus , strain differences did not cause any discordant results when grown under glucose media . generation of the entire h2a / h2b alanine - scanning collection in an fy406 background . ( a ) the h2a / h2b alanine - scanning library was generated as described in our previous study ( nakanishi et al . the complete removal of wild - type ( wt ) h2b was ensured by multiple rounds of 5-foa selection and verified by sequencing . each colony represents a strain expressing histones containing a single alanine substitution mutation of each of the residues of h2a and h2b . red squares indicate the location of strains that are inviable in sd media containing 5-foa ( lethal mutants ) . for the key to the corresponding mutant strains within each plate , see tables s2 and s3 . ( b ) western blot analysis of mutant strains in the fy406 background identified as defective for proper methylation of h3k4 . cell extracts prepared from each mutant strain were subjected to sds - page and analyzed by western analysis with antibodies to dimethyl h3k4 , trimethyl h3k4 ( h3k4me2 and h3k4me3 , respectively ) , monoubiquitinated h2bk123 ( h2bub ) , or flag ( flag ) . our data clearly demonstrate that monoubiquitination of histone h2b on lysine 123 and the machinery required for its implementation are the sole requirements for the regulation of the trimethylation of h3k4 and h3k79 in yeast s. cerevisiae . unknown mutations as proposed by foster and downs ( 2009 ) that function with h2bk123r in the regulation of h3k4 and h3k79 trimethylation . furthermore , we have discovered that the widely used y131 strain background expresses a previously undetected copy of the hta2-htb2 genes when this strain is grown for many generations in the presence of galactose . therefore , we generated the entire h2a / h2b mutant collection in a background that can be readily used under galactose conditions and have now made this collection available to the entire scientific community . we used a plasmid containing hta1 and htb1 genes and a plasmid containing all four histones , hta1 , htb1 , hht2 , and hhf2 genes , as shown in fig . 1 b. plasmids bearing an alanine or arginine mutation in the flag - tagged htb1 gene and flag - less htb1 gene were generated by site - directed mutagenesis ( quickchange ii kit ; agilent technologies ) . mutated targets were confirmed by sequencing using the primers ( htbseqf ) 5-ggcaaatactaccttggttgg-3 and ( htbseqr ) 5-tttcgagaacacaattttacaaccga-3. each plasmid was transformed manually into yeast shuffle strains y131 , fy406 , dy20d , and jhy205 using a standard yeast transformation protocol , and strains were grown on a synthetic dropout ( sd ) medium lacking histidine , sd - his ( for y131 and fy406 strains ) , or a medium lacking leucine , sd - leu ( for dy20d and jhy205 mutants ) . after 2 d of incubation , transformants were replica plated onto plates containing either sd - his plus 5-foa or sd - leu plus 5-foa to select single - colony cells that had lost the plasmid containing the wild - type histones . each colony was inoculated into ypd medium ( 1% yeast extract , 2% peptone , and 2% dextrose ) plus 5-foa . cells are grown in ypd or ypgal ( 1% yeast extract , 2% peptone , and 2% galactose ) to mid - log phase . whole cell extracts were prepared from the wild - type and histone mutant strains as previously described ( wood et al . , 2003b ) with some minor modifications . in brief , cell pellets were washed and resuspended in 400 l nib ( 0.25 m sucrose , 60 mm kcl , 14 mm nacl , 5 mm mgcl2 , 1 mm cacl2 , and 0.8% triton x-100 ) . after the addition of 250 l of 0.5-mm glass beads to the tubes containing the suspension , the tubes were vortexed for 20 min at 4c . cell lysates were recovered by puncturing the bottom of the tube and centrifuging the contents at 3,000 rpm . after removal of the supernatant , the pellet was suspended in 150 l of sterile water and 75 l of 4 loading buffer and heated at 95c for 5 min . extracts were subjected to 18% sds - page electrophoresis , transferred to nitrocellulose membrane , and probed with either anti - flag or h2b ubiquitin specific antibodies , as well as antibodies specific for h3k4 di- and trimethylation and h3k79 trimethylation , followed by detection of the bound antibody with horseradish peroxidase conjugated secondary antibodies . an antibody to histone h3 was used as a loading control . the h2a / h2b histone mutant library , plasmids bearing alanine mutations in the hta1 and htb1 genes were systematically generated by site - directed mutagenesis in 96-well format . after transformation into e. cloni 10 g elite electrocompetent cells using a 96-well electroporator , plasmids were prepared with a biomekfx ( beckman coulter ) using the cosmcprep kit ( agencourt ) . each mutation was confirmed by sequencing using the aforementioned primers , and each plasmid was transformed into fy406 using a standard yeast transformation protocol . transformants were selected on an sd - his medium followed by the second selection on a plate containing sd - his plus 5-foa . to ensure the complete removal of the plasmid containing the wild - type histones , each single colony was inoculated into ypd medium plus 5-foa in 96-well plates . finally , histone mutant strains were confirmed by sequencing , and the glycerol stocks of the library were generated . tables s2 and s3 show the key for the histone h2a / h2b library in fy406 background that corresponds to the mutant strains within each plate shown in fig . we used a plasmid containing hta1 and htb1 genes and a plasmid containing all four histones , hta1 , htb1 , hht2 , and hhf2 genes , as shown in fig . 1 b. plasmids bearing an alanine or arginine mutation in the flag - tagged htb1 gene and flag - less htb1 gene were generated by site - directed mutagenesis ( quickchange ii kit ; agilent technologies ) . mutated targets were confirmed by sequencing using the primers ( htbseqf ) 5-ggcaaatactaccttggttgg-3 and ( htbseqr ) 5-tttcgagaacacaattttacaaccga-3. each plasmid was transformed manually into yeast shuffle strains y131 , fy406 , dy20d , and jhy205 using a standard yeast transformation protocol , and strains were grown on a synthetic dropout ( sd ) medium lacking histidine , sd - his ( for y131 and fy406 strains ) , or a medium lacking leucine , sd - leu ( for dy20d and jhy205 mutants ) . after 2 d of incubation , transformants were replica plated onto plates containing either sd - his plus 5-foa or sd - leu plus 5-foa to select single - colony cells that had lost the plasmid containing the wild - type histones . each colony was inoculated into ypd medium ( 1% yeast extract , 2% peptone , and 2% dextrose ) plus 5-foa . cells are grown in ypd or ypgal ( 1% yeast extract , 2% peptone , and 2% galactose ) to mid - log phase . whole cell extracts were prepared from the wild - type and histone mutant strains as previously described ( wood et al . , 2003b ) with some minor modifications . in brief , cell pellets were washed and resuspended in 400 l nib ( 0.25 m sucrose , 60 mm kcl , 14 mm nacl , 5 mm mgcl2 , 1 mm cacl2 , and 0.8% triton x-100 ) . after the addition of 250 l of 0.5-mm glass beads to the tubes containing the suspension , the tubes were vortexed for 20 min at 4c . cell lysates were recovered by puncturing the bottom of the tube and centrifuging the contents at 3,000 rpm . after removal of the supernatant , the pellet was suspended in 150 l of sterile water and 75 l of 4 loading buffer and heated at 95c for 5 min . extracts were subjected to 18% sds - page electrophoresis , transferred to nitrocellulose membrane , and probed with either anti - flag or h2b ubiquitin specific antibodies , as well as antibodies specific for h3k4 di- and trimethylation and h3k79 trimethylation , followed by detection of the bound antibody with horseradish peroxidase conjugated secondary antibodies . an antibody to histone h3 was used as a loading control . the h2a / h2b histone mutant library was generated in a fy406 background as described previously ( nakanishi et al . , 2008 ) . in brief , plasmids bearing alanine mutations in the hta1 and htb1 genes were systematically generated by site - directed mutagenesis in 96-well format . after transformation into e. cloni 10 g elite electrocompetent cells using a 96-well electroporator , plasmids were prepared with a biomekfx ( beckman coulter ) using the cosmcprep kit ( agencourt ) . each mutation was confirmed by sequencing using the aforementioned primers , and each plasmid was transformed into fy406 using a standard yeast transformation protocol . transformants were selected on an sd - his medium followed by the second selection on a plate containing sd - his plus 5-foa . to ensure the complete removal of the plasmid containing the wild - type histones , each single colony was inoculated into ypd medium plus 5-foa in 96-well plates . finally , histone mutant strains were confirmed by sequencing , and the glycerol stocks of the library were generated . tables s2 and s3 show the key for the histone h2a / h2b library in fy406 background that corresponds to the mutant strains within each plate shown in fig . 5 a. online supplemental material is available at http://www.jcb.org/cgi/content/full/jcb.200906005/dc1 . | histone h2b monoubiquitination by rad6/bre1 is required for the trimethylation of both histone h3k4 and h3k79 by compass and dot1 methyltransferases , respectively .
the dependency of methylation at h3k4 and h3k79 on the monoubiquitination of h2bk123 was recently challenged , and extragenic mutations in the strain background used for previous studies or epitope - tagged proteins were suggested to be the sources of this discrepancy . in this study , we show that h3k4 and h3k79 methylation is solely dependent on h2b monoubiquitination regardless of any additional alteration to the h2b sequence or genome . furthermore , we report that y131 , one of the yeast histone h2a / h2b shuffle strains widely used for the last decade in the field of chromatin and transcription biology , carries a wild - type copy of each of the hta2 and htb2 genes under the gal1/10 promoter on chromosome ii .
therefore , we generated the entire histone h2a and h2b alanine - scanning mutant strains in another background , which does not express wild - type histones . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
although bone marrow transplantation ( bmt ) can be a successful treatment modality for a variety of hematological disorders , malignancies and metabolic defects , a number of obstacles to be overcome still exist for a better outcome with the transplantation technique . patients with engraftment failure or poor graft function following allogeneic bmt are at high risk of morbidity and mortality . therapy of engraftment failure or poor graft function includes the administration of human recombinant growth factors and reinfusion of donor marrow . recently , infusion of allogeneic peripheral blood stem cells ( pbsc ) mobilized by granulocyte - colony stimulating factor ( g - csf ) has been used in patients with graft failure after allogeneic bmt . allogeneic pbsc transplantation has some advantages over allogeneic bmt : general anesthesia is not required to collect hematopoietic stem cells and faster hematologic recovery has been reported . there may , however , be an increased risk of graft - versus - host - disease ( gvhd ) as leukapheresis products usually contain 1 log higher t lymphocytes than bone marrow . here we report a case of a long - term follow - up who developed chronic gvhd following allogeneic pbsc rescue for poor graft function after allogeneic bmt . an 18-year - old man with severe aplastic anemia , who was diagnosed in june 1991 , underwent an allogeneic bmt from his hla - identical 21-year - old sister . the conditioning regimen consisted of procarbizine orally at a dose of 6.25 mg / kg / day for 3 days and cyclophosphamide at 50 mg / kg / day for 4 days and antithymocyte globulin at 1.25 mg / kg / day for 3 days . a conventional regimen of short course methotrexate and cyclosporin a ( csa ) was used for the prophylaxis of gvhd . after initial excellent graft take for 8 months , his blood counts gradually decreased to 2.8 10/l of white cells and 28 10/l of platelets . there was no infectious evidence including cytomegaloviral infection or acute exacerbation of hepatitis b responsible for marrow suppression . dna finger printing , using variable - number - of - tandem - repeats ( vntr ) , revealed a donor engraftment . recombinant human granulocyte macrophage - colony stimulating factor ( gm - csf ) at 5 g / kg / day was tried for three weeks to no avail . after obtaining informed consents from both donor and recipient the donor received filgrastim at a dose of 5 g / kg / day subcutaneously for 7 consecutive days . on day 8 , the donor underwent a 7 l leukapheresis with fenwal cs-3000 plus cell separator . the donor had no side effects during pbsc mobilization and collection . on day 429 following bmt , a total of 1.6 10/kg mononuclear cells and 4.3 10/kg cd34 + cells was infused without further processing . antilymphocyte globulin ( 10 mg / kg / day for 5 days ) was used as a sole preparative therapy . as a gvhd prophylaxis full engraftment was achieved and he was monitored as an outpatient after discharge from hospital on day 31 after the boost . around 1 year after the boost ( january 1995 ) , while he was on oral csa at 50 mg , he developed abdominal pain , diarrhea and jaundice . blood counts were normal ( hgb 12.5 g / dl , wbc 7.2 10/l , platelet 125 10/l ) , but liver function tests were markedly abnormal ( maximum total bilirubin 19.4 mg / dl ; direct bilirubin 12.4 mg / dl ; alkaline phosphatase 169 iu / l ; ast 54 u / l ; alt 70u / l ) . on the clinical ground , cytomegaloviral ( cmv ) enteritis and hepatitis , acute exacerbation of hepatitis b , or chronic gvhd , were suspected . cmv was excluded by negative early antigenemia assay , polymerase chain reaction and shell vial culture . a percutaneous liver biopsy and gastrofiberoscopic stomach biopsy showed the evidence of chronic gvhd ( fig . he developed extensive chronic gvhd involving skin , eye , liver , oral and gastrointestinal mucosa . the pulse therapy of prednisolone 60 mg / day and csa 600 mg / day resulted in gradual improvement of gvhd and he is now well on low - dose steroid at day + 1055 after pbsc rescue . poor graft function , as indicated by a decrease in the peripheral blood counts to less than 40% of the maximal preceding values post - transplant , in association with reduced marrow cellularity , is an infrequent but serious complication of bmt . therapy of poor graft function is not always successful with a second marrow infusion or , more recently , the administration of human recombinant growth factors ( csfs ) . in our case , recombinant human gm - csf at 5 g / kg / day was tried for three weeks to no avail . as the addition of csf - mobilized peripheral blood progenitor cells ( pbpc ) to autologous bone marrow grafts markedly improves hematopoietic recovery , infusion of donor - derived pbsc they reported rapid tri - lineage engraftment using pbsc collected during steady - state haematopoiesis ; however , a total of nine leukaphereses were required to obtain sufficient cells . both patients were reported to have sustained tri - lineage engraftment with good hematopoietic function . the clinical advantages of pbsc transplants over bmt for allogeneic transplantation have been described in a number of studies . for the donor , the harvest can be performed without the need for a general anesthesia and avoids the usually minor but not negligible morbidity associated with bm harvesting . modern automated continuous flow cell separators can process 9 l of blood in 3 h without the need for central vascular catheters , providing there is good venous access . g - csf has a high safety profile with only minor side - effects . for the recipient , potential advantages of the use of pbsc include accelerated neutrophil and platelet engraftment as has been documented following autologous pbsct . the high number of progenitor cells that can be harvested from g - csf - mobilzed peripheral blood may be particularly advantageous in transplantation for aplastic anemia where an increased risk of rejection has been associated with the infusion of less than 3 10 bm cells / kg . potential disadvantages of allogeneic pbsct include the theoretical increased risk of gvhd due to the presence of approximately 1 log larger numbers of t lymphocytes in the leukapheresis products than in bm . however , tendency toward an increased incidence of chronic gvhd has been reported in some studies , correlating with t cell doses . the chronic form of gvhd seemed to be less sensitive to csa given either prophylactically or therapeutically . improvement in the prophylaxis and control of chronic gvhd , such as the addition of steroids or a partial t cell depletion , should be considered in case the chronic gvhd in allogeneic pbsct might affect the survival and performance adversely . this case suggests that mobilized pbscs are capable of providing a complete , sustained hematopoietic engraftment , and the use of primed pbsc for allogeneic transplantation is a feasible alternative to bm in treating poor graft function after bmt . nevertheless , further studies are required to fully assess its pros and cons , particulary in terms of gvhd and survival . | to overcome poor graft function after allogeneic bone marrow transplantation ( bmt ) , the use of peripheral blood stem cells ( pbsc ) instead of bone marrow is gaining more popularity because of its advantages .
there may , however , be an increased risk of graft - versus - host - disease ( gvhd ) because of the large number of lymphocytes present in a leukapheresis product.an 18-year - old man with severe aplastic anemia underwent an allogeneic bmt using his hla - identical sister .
after initial excellent graft take for 8 months , his blood counts gradually decreased to 2.8 109/l of white cells and 28 109/l of platelets with marrow cellularity of < 10% .
after allogeneic granulocyte - colony stimulating factor mobilized pbsc rescue , the patient s blood counts recovered satisfactorily . around 1 year after the boost , he developed chronic gvhd that responded to prednisolone and cyclosporin a. he is now well on low - dose steroids at day + 1055 after pbsc rescue .
the present case is the first experience of a long - term follow - up who underwent allogeneic pbsc rescue in korea . |
You are an expert at summarizing long articles. Proceed to summarize the following text:
compared with the synovial joints of the limbs , the temporomandibular joint ( tmj ) develops relatively late . by the eighth week of development in humans ( orahilly s stage 23 ) , the joint cavity of the elbows , hips , and knees are already visible , while the tmj has only mesenchymal condensation of the mandibular condyle , articular disc , and squamous part of the temporal bone , and no evidence of its cavity . , by approximately week nine , the condylar chondrification begins in the center of the condylar blastemal . the inferior joint cavity starts to develop at the end of the ninth week , as the density of the mesenchymal cells in the central region decreases because of their differentiation into fibroblasts , constituting the primordium of the articular disc . as the density of the ectomesenchyme decreases , small adjacent spaces between the cells coalesce , forming the inferior joint cavity between the future articular disc and the mandibular condyle . in the 10 week , these newly differentiated fibroblasts become more compact , forming collagen fibers and the articular disc , but there is still no sign of the superior joint cavity . in the eleventh week , the organization of the superior joint cavity begins between the squamous part of the temporal bone and the articular disc . more detailed studies of the tmj development , as well as of their condylar cartilage , articular disc and the matrix proteins on the fetus development were researched through immunohistochemistry and in situ hybridization . morphologically , the synovial membrane consists of two layers : an inner cell layer called the intima and a support layer called the vascular subintima , which mixes with the fibrous capsule . the intima consists of cells embedded in an amorphous , fiber - free matrix with an approximate thickness of one to four cells . the subintima consists of loose connective tissue with blood vessels , spread - out fibroblasts , macrophages , mastocytes , adipose cells , and some elastic fibers that prevent membrane folding . the intima contains macrophage - like type a cells with phagocytic ability , and fibroblast - like type b cells that synthesize proteins , glycoproteins , and proteoglycans . immunohistochemical techniques allowed to detect macrophage - like type a cells and fibroblast - like type b cells by using anti - macrophage and anti - fibroblast antibodies , respectively . grabowski et al . , nozawa - inoue et al . and suzuki et al . successfully marked macrophage - like type a cells with the antibodies ox6 , edi , cd68 , and cd31 . on the other hand , the laminin antibodies mab67 , vcam-1 , lumican , fibromodulin , udpgd , and hsp25 mark fibroblast - like type b cells , and have been used in studies of the synovial intima . although a number of groups have studied about the synovial membrane of the tmj of children , adults , and animals models , there is little in the literature regarding the development of this structure in human fetuses . understanding the morphology of fetal development is important not only to better understand the embryological steps that culminate with the individual s anatomic constitution but also to elucidate the intrinsic mechanisms that may be involved in congenital anomalies and postnatal pathologies . this study aims to improve the anatomic and histologic knowledge of the synovial membrane by determining when the synovial meml.o . carvalho de moraesbrane begins to form during morphogenesis and the chronological occurrence and dynamics of cell types present in the synovial membrane . it also aims to analyze the morphological differences that appear during the development of the superior and inferior articular cavities . we studied twenty one human fetuses from the institute of embryology of the university complutense of madrid and associao de fundo incentivo pesquisa . the specimens ranged from 40 mm to 100 mm greatest length ( gl ) with the following ages : three nine - week fetuses ; four ten - week fetuses ; five eleven - week fetuses ; four twelve - week fetuses ; five thirteen - week fetuses ; post - conception age was determined by measuring gl and external and internal criteria . all specimens were from ectopic pregnancies or spontaneous abortions , and there were no signs of malformation . all fetuses were fixed in 10% formalin and separated into groups studied by light microscopy and immunohistochemistry . all samples were decalcified in edta for 21 days and rinsed with tap water for 10 min . they were then fixed in 10% formaldehyde for 24 h. next , they were dehydrated as follows : one 24-h immersion in 50% ethanol ; two 6-h immersions in 70% ethanol ; and two 6-h immersions in absolute ethanol . finally , the samples were cleared by a 2-h treatment with xylol and fixed in paraffin . semi - serial , frontal , and sagittal 4 m cuts of the tmj were made by the microtome leica , model rm2035 . the sections , which included three spatial planes , were stained with hematoxylin - eosin ( he ) , azocarmine , and masson s trichrome stain . the following antibodies were used : cd68 ( santa cruz biotechnology , santa cruz , ca , usa ) ; monoclonal anti - human igg1 produced in mouse and diluted to 1:100 and hsp27 ( santa cruz biotechnology ) ; monoclonal anti - human igg1 produced in mouse and diluted to 1:150 using the indirect immunoenzyme method in three stages and the streptavidin - biotin - peroxidase complex . the antibodies cd68 and hsp27 react with the cells macrophage - like type a and fibroblast - like type b , respectively . the positive controls for the antibodies cd68 and hsp27 were palatine tonsil and breast adenocarcinoma , respectively . the negative controls were the same cases used as positive controls , but a buffer instead of the primary antibody was used for immunohistochemical incubation . the study was approved by the ethics committee of the faculty of medicine of the university complutense of madrid and the federal university of so paulo research ethics committee . all specimens studied during the ninth week showed small spaces or clefts between the primordium of the articular disc and the mandibular condyle that defined the initial formation of the inferior joint cavity . some vascular branches were located in the periphery of the articular disc primordium ( figure 1a ) . all specimens studied during the tenth week showed the organization of the inferior joint cavity complete although crossed by a few tracts of connective tissue . blood vessels in the inferior part of the articular disc primordium were seen during the eleventh week . fusiform - like cells separate the vessels of the inferior articular cavity ( figure 1 c , d ) . the organization of the superior joint cavity began between the squamous part of the temporal bone and the primordium of the articular disc . the superior joint cavity was crossed by few tracts of the connective tissue ( figure 1c ) . numerous vessels were seen in the lateral and medial parts of the tmj ( figure 1d ) . all specimens studied this week showed the joint cavities clearly defined and decrease in the number of septa of connective tissue that crossed them was noted . the vessels in the inferior part of the articular disc did not reach the middle area of the disc ( figure 1 e , f ) . some macrophage - like cells that expressed cd 68 and fibroblast - like type b cells that express hsp27 ( figure 2 a , c , e ) appeared bottoms of the inferior articular cavity . all specimens studied during the thirteenth week showed the vessels located bottoms of the inferior articular cavity . some vessels surrounded by fusiform cells protruded from the inferior sac bottoms , which corresponds to the primordium of the synovial villi ( figure 1 g ) . panels b and d of figure 2 show the positive control of the antibodies cd68 and hsp27 in palatine tonsil and breast adenocarcinoma samples , respectively . all specimens studied during the ninth week showed small spaces or clefts between the primordium of the articular disc and the mandibular condyle that defined the initial formation of the inferior joint cavity . some vascular branches were located in the periphery of the articular disc primordium ( figure 1a ) . all specimens studied during the tenth week showed the organization of the inferior joint cavity complete although crossed by a few tracts of connective tissue . blood vessels in the inferior part of the articular disc primordium were seen during the eleventh week . fusiform - like cells separate the vessels of the inferior articular cavity ( figure 1 c , d ) . the organization of the superior joint cavity began between the squamous part of the temporal bone and the primordium of the articular disc . the superior joint cavity was crossed by few tracts of the connective tissue ( figure 1c ) . numerous vessels were seen in the lateral and medial parts of the tmj ( figure 1d ) . all specimens studied this week showed the joint cavities clearly defined and decrease in the number of septa of connective tissue that crossed them was noted . the vessels in the inferior part of the articular disc did not reach the middle area of the disc ( figure 1 e , f ) . some macrophage - like cells that expressed cd 68 and fibroblast - like type b cells that express hsp27 ( figure 2 a , c , e ) appeared bottoms of the inferior articular cavity . all specimens studied during the thirteenth week showed the vessels located bottoms of the inferior articular cavity . some vessels surrounded by fusiform cells protruded from the inferior sac bottoms , which corresponds to the primordium of the synovial villi ( figure 1 g ) . panels b and d of figure 2 show the positive control of the antibodies cd68 and hsp27 in palatine tonsil and breast adenocarcinoma samples , respectively . according to mrida - vela et al . , the tmj joint cavities are not formed synchronously . first , the inferior joint cavity begins to develop in the ninth week as small spaces or fissures appear between the articular disc and the mandibular condyle . subsequently , the superior joint cavity begins to form approximately in the eleventh week of gestation , between the articular disc and the squamous part of the temporal bone . sperber , burdi , and gutcen - toller , who reported that the inferior joint cavity begins to form during the tenth week of gestation , while the superior joint cavity begins to form from the eleventh week of gestation . according to ohnuki , the inferior joint cavity above the mandibular condyle is still being formed during the twelfth week of development , while the superior joint cavity is only visible in the posterior region of the joint . in humans , buccal movements begin during weeks 7 and 8 of development at the level of the incudomalleolar joint . tmj movements help to form the joint cavity and their absence could results in congenital craniofacial anomalies . the presence of blood vessels in the articular disc of the tmj of human fetuses has been described by many authors . van der linden et al . noted that capillaries were distributed in the periphery of the anterior and posterior portions of the articular disc in human fetuses . however , they did not report any vessels in the central region of the articular disc . wong et al . observed that in fetuses , 13 to 17.5 weeks of development vessels are mainly located anteriorly and posteriorly , but no evidence of vascularization was found at the center of articular disc . studied 61 specimens of articular disc aged 2 days to 26 years and concluded that vascularization only exists in the first year of life , after which it disappears . interestingly , sab et al . observed vessels containing red blood cell in the central portion of the tmj articular disc of human fetuses . according to our findings , from the ninth week of development the number of vessels increases around the temporomandibular joint region , sending some vascular branches to the periphery of the articular disc primordium . in specimens at eleven weeks of development , the vessels reach the central portion of the articular disc and fusiform - like cells separate the disc from the inferior articular cavity . during the twelfth week of development , the articular cavities are well defined and the vessels do not reach the central region of the articular disc . these findings can suggest that the vessels are necessary for the vascularization of the articular disc until the synovial membrane is well developed . the synovial membrane of the tmj is areolar and only this type of synovial membrane is characterized by the presence of two cell types in its intima layer : macrophage - like type a cells and fibroblast - like type b cells . their basic function is to engulf , by phagocytosis , the proteins and carbohydrates present in the synovial . fibroblast - like type b cells are ultrastructurally characterized by the presence of a well - developed rough endoplasmic reticulum with numerous secretory granules . macrophage - like type a and fibroblast - like type b cells in the synovial membrane can be identified by immunohistochemistry and transmission electron microscopy in fetuses , adults , and animals before and after birth . ikeda et al . reported that macrophage - like type a cells in the synovial membrane of the tmj of rats were only visible after birth . found macrophage - like type a and fibroblast - like type b cells only during the gestational period . our findings confirm those of other authors , including who found both macrophage - like type a and fibroblast - like type b cells in the synovial membrane before birth . during the 12 week of development of the peripheral parts of the inferior articular cavity , macrophage - like type a and fibroblast - like type b cells were marked by immunohistochemical reaction with the antibodies cd68 and hsp27 , respectively . fibroblast - like type b cells were also marked in the peripheral portions of the superior articular cavity . these findings could suggest that from the twelfth week of development , with the articular cavities well formed , the synovial membrane is functional and the vessels are only seen in the peripheral areas of the articular disc . there is also a third type of cell not yet fully studied called an intermediate lining cell . this study demonstrates that development of the synovial membrane depends on the fibroblast - like type b cells located on the surface of the synovial membrane and macrophage - like type a cells that supply the vessels . our results also suggest that in human specimens at twelve weeks of development , the articular cavities are well formed and the synovial membrane begins to be functional . | the development of the synovial membrane was analyzed in serial sections of 21 temporomandibular joints of human fetuses at 9 to 13 weeks of gestation .
sections of two fetuses at 12 weeks of development were used to perform immunohistochemical expression of the markers cd68 and hsp27 on the synovial lining .
macrophage - like type a and fibroblast - like type b cells , which express cd68 and hsp27 , respectively , were observed at the twelfth week of development .
our results suggest that the development of the synovial membrane is related to the vascularization of the joint and the formation of the articular cavities . |
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prior to the study , an administrative authorization was obtained from the divisional officer of the biyem - assi health district . respondents were informed of the study and the confidentiality of the data to be collected . the study was carried out from october 2013 to march 2014 in the biyem - assi health district ( 0384n , 1149e ) which is located in the south west of yaound , the capital of cameroon ( fig . 1 ) . its population is estimated at 310,000 ( 29 ) and is characterized by increasing numbers of stray dogs in the streets and the large number of wild garbage dumps positioned closely to households . the head of each household or its substitute ( > 18 years old ) was interviewed in french for 1015 min using a structured questionnaire . the questionnaire designed for this study consisted of closed and a few open - ended questions . it had four parts : anthropometric and socio - demographic information , risk factors of rabies transmission ( stray dogs and ogds ) , knowledge / attitude / perception of rabies , and its measures of control . the questionnaire was pretested in a neighborhood quarter with the similar socio - demographic characteristic prior to the survey and was modified to improve its clarity and analysis . an observational method was considered more appropriate to characterize household wastes in ogds . through a systematic check , ogds were identified and solid wastes were characterized as fermentable , hazardous , or non - hazardous wastes . vaccinated dogs were defined as those that had been immunized ( oral or parenteral ) 1 year before the survey . the chi - square test was used to set the difference in proportions or frequencies . prior to the study , an administrative authorization was obtained from the divisional officer of the biyem - assi health district . respondents were informed of the study and the confidentiality of the data to be collected . the study was carried out from october 2013 to march 2014 in the biyem - assi health district ( 0384n , 1149e ) which is located in the south west of yaound , the capital of cameroon ( fig . its population is estimated at 310,000 ( 29 ) and is characterized by increasing numbers of stray dogs in the streets and the large number of wild garbage dumps positioned closely to households . the head of each household or its substitute ( > 18 years old ) was interviewed in french for 1015 min using a structured questionnaire . the questionnaire designed for this study consisted of closed and a few open - ended questions . it had four parts : anthropometric and socio - demographic information , risk factors of rabies transmission ( stray dogs and ogds ) , knowledge / attitude / perception of rabies , and its measures of control . the questionnaire was pretested in a neighborhood quarter with the similar socio - demographic characteristic prior to the survey and was modified to improve its clarity and analysis . an observational method was considered more appropriate to characterize household wastes in ogds . through a systematic check , ogds were identified and solid wastes were characterized as fermentable , hazardous , or non - hazardous wastes . vaccinated dogs were defined as those that had been immunized ( oral or parenteral ) 1 year before the survey . the chi - square test was used to set the difference in proportions or frequencies . the main education levels of participants were primary ( 39.8% ) and secondary ( 34.3% ) schools . slightly under a quarter ( 22.1% ) had attended higher education institutions ( table 1 ) . approximately 32.14% of respondents had at least one dog and 67% of them had vaccinated dogs . demographic and socio - demographic characteristics a total of 134 ogds were numbered in the biyem - assi health district and the average distance between these ogds and the nearest households ranged from 5 to 15 m with a median distance of 10 m. some solid wastes including plastic , textile , paper , paperboard , and food wastes were steadily found in every ogd ( table 2 ) . solid wastes characterization questioned about risks of closed dumps to households , 35 and 28.1% of respondents estimated that this predominantly leads to an increase of stray dog density and mosquitoes , respectively . ogds were considered as the source of unhealthy environment and disease at 21.9 and 15% , respectively . regarding the definition of stray dog , 66.9% of heads of households knew what a stray dog is while for 14.76% stray dogs are domestic dogs , pet dogs for 8.81% , or guard dogs for 6.90% . feeding places for stray dogs were wild garbage dumps ( 68.1% ) , markets ( 18.3% ) , and houses ( 13.6% ) . the chi - square test analysis of feeding behavior of stray dogs as well as risk they incur revealed that 42% of respondents consider the feeding behavior of stray dogs as a potential risk for rabies transmission ( table 3 ) . there was a direct association between feeding behavior of stray dogs and rabies transmission at a significant level (
= 154.12 , df=4 , p<0.05 ) . stray dogs and risk linked to feeding behavior all participants involved in the study have full knowledge concerning rabies transmission by dog bites . in contrast , only 32% of respondents attribute rabies transmission to scratching and licking of a rabid dog . concerning transmission of rabies by bites of animals other than dogs , 21 and 15% of respondents attributed rabies to cats and cows , respectively . concerning the perception of rabies symptoms in humans , rabies was characterized by fever ( 10% ) , wound infection ( 51.43% ) , and diarrhea ( 6.67% ) . a total of 31.90% of respondents were unable to state the symptoms of rabies in humans ( fig . rabies was characterized by irritability ( 49% ) , skin lesions ( 26.9% ) , and alopecia ( 11% ) . a total of 13.1% of respondents had no idea about the symptoms of rabies in dogs ( fig . 3 ) . as measures to be taken in the case of dog bites , 56% of participants accepted that they would disinfect the wound after washing with water , 35% reported that they would go to hospital , and 9% did not know what to do in this situation ( table 4 ) . concerning control measures , participants recommended the destruction of ogds to reduce stray dog population ( 36% ) , the regulation of stray dog density in the district ( 48% ) , and the vaccination of dogs ( 16% ) . the main education levels of participants were primary ( 39.8% ) and secondary ( 34.3% ) schools . slightly under a quarter ( 22.1% ) had attended higher education institutions ( table 1 ) . approximately 32.14% of respondents had at least one dog and 67% of them had vaccinated dogs . a total of 134 ogds were numbered in the biyem - assi health district and the average distance between these ogds and the nearest households ranged from 5 to 15 m with a median distance of 10 m. some solid wastes including plastic , textile , paper , paperboard , and food wastes were steadily found in every ogd ( table 2 ) . questioned about risks of closed dumps to households , 35 and 28.1% of respondents estimated that this predominantly leads to an increase of stray dog density and mosquitoes , respectively . ogds were considered as the source of unhealthy environment and disease at 21.9 and 15% , respectively . regarding the definition of stray dog , 66.9% of heads of households knew what a stray dog is while for 14.76% stray dogs are domestic dogs , pet dogs for 8.81% , or guard dogs for 6.90% . feeding places for stray dogs were wild garbage dumps ( 68.1% ) , markets ( 18.3% ) , and houses ( 13.6% ) . the chi - square test analysis of feeding behavior of stray dogs as well as risk they incur revealed that 42% of respondents consider the feeding behavior of stray dogs as a potential risk for rabies transmission ( table 3 ) . there was a direct association between feeding behavior of stray dogs and rabies transmission at a significant level (
= 154.12 , df=4 , p<0.05 ) . stray dogs and risk linked to feeding behavior all participants involved in the study have full knowledge concerning rabies transmission by dog bites . in contrast , only 32% of respondents attribute rabies transmission to scratching and licking of a rabid dog . concerning transmission of rabies by bites of animals other than dogs , 21 and 15% of respondents attributed rabies to cats and cows , respectively . concerning the perception of rabies symptoms in humans , rabies was characterized by fever ( 10% ) , wound infection ( 51.43% ) , and diarrhea ( 6.67% ) . a total of 31.90% of respondents were unable to state the symptoms of rabies in humans ( fig . rabies was characterized by irritability ( 49% ) , skin lesions ( 26.9% ) , and alopecia ( 11% ) . a total of 13.1% of respondents had no idea about the symptoms of rabies in dogs ( fig . 3 ) . as measures to be taken in the case of dog bites , 56% of participants accepted that they would disinfect the wound after washing with water , 35% reported that they would go to hospital , and 9% did not know what to do in this situation ( table 4 ) . concerning control measures , participants recommended the destruction of ogds to reduce stray dog population ( 36% ) , the regulation of stray dog density in the district ( 48% ) , and the vaccination of dogs ( 16% ) . questioned about risks of closed dumps to households , 35 and 28.1% of respondents estimated that this predominantly leads to an increase of stray dog density and mosquitoes , respectively . ogds were considered as the source of unhealthy environment and disease at 21.9 and 15% , respectively . regarding the definition of stray dog , 66.9% of heads of households knew what a stray dog is while for 14.76% stray dogs are domestic dogs , pet dogs for 8.81% , or guard dogs for 6.90% . feeding places for stray dogs were wild garbage dumps ( 68.1% ) , markets ( 18.3% ) , and houses ( 13.6% ) . the chi - square test analysis of feeding behavior of stray dogs as well as risk they incur revealed that 42% of respondents consider the feeding behavior of stray dogs as a potential risk for rabies transmission ( table 3 ) . there was a direct association between feeding behavior of stray dogs and rabies transmission at a significant level (
= 154.12 , df=4 , p<0.05 ) . all participants involved in the study have full knowledge concerning rabies transmission by dog bites . in contrast , only 32% of respondents attribute rabies transmission to scratching and licking of a rabid dog . concerning transmission of rabies by bites of animals other than dogs , 21 and 15% of respondents attributed rabies to cats and cows , respectively . concerning the perception of rabies symptoms in humans , rabies was characterized by fever ( 10% ) , wound infection ( 51.43% ) , and diarrhea ( 6.67% ) . a total of 31.90% of respondents were unable to state the symptoms of rabies in humans ( fig . rabies was characterized by irritability ( 49% ) , skin lesions ( 26.9% ) , and alopecia ( 11% ) . a total of 13.1% of respondents had no idea about the symptoms of rabies in dogs ( fig . 3 ) . as measures to be taken in the case of dog bites , 56% of participants accepted that they would disinfect the wound after washing with water , 35% reported that they would go to hospital , and 9% did not know what to do in this situation ( table 4 ) . concerning control measures , participants recommended the destruction of ogds to reduce stray dog population ( 36% ) , the regulation of stray dog density in the district ( 48% ) , and the vaccination of dogs ( 16% ) . instead of its preventive possibility , rabies remains a public health problem of major concern . the situation is alarming in rural africa and asia where canine rabies has been severely neglected ( 30 ) . moreover , the periodicity of epidemics leads to only intermittently perceiving rabies as a problem ( 31 ) . in these rural areas , the disease persistence is due to poverty and the increase of wild carnivore populations ( 32 ) . regarding canine rabies in africa , most studies focus on dog vaccinations while little is known about the origin and factors that favor the free roaming of dogs and human rabies transmission . thus , transmission and identification of risk factors are the most important processes underlying infectious disease dynamics ( 33 ) , but it is also the least understood . therefore , knowledge and attitudes of the population , perception of rabies , and the impact of ogds on the transmission of rabies were investigated in biyem - assi health district of yaounde through a cross - sectional study . to the best of our knowledge , this study stands as the first conducted to highlight the public health threat of rabies in cameroon . findings of this study indicate that rabies is an important public health problem in the biyem - assi health district and that community awareness , knowledge , and perception of rabies were high among the respondents . these data corroborate with those obtained in some endemic countries of south asia that revealed a high level of knowledge on rabies transmission . in fact , about 98.6% of the population knew that rabies is transmitted by rabid dogs ( 3436 ) . this high level of awareness among the respondents may be attributed to their level of education . in fact , 96% of the respondents had attended , at least , primary school ( table 1 ) . about 31% of respondents did not know what stray dogs were , 32% of dogs were not vaccinated and 31.90% of respondents did not know the symptoms of rabies in humans . educating the population on rabies and the risk factors associated with its transmission could fill this knowledge gap . the study also showed that within dogs owners , there is a good coverage of vaccination ( > 67% ) and the rabies pep behaviors in the population are considerable . this was confirmed by the fact that 56% of participants would disinfect their wounds and 35% would go to hospital for treatment after a dog bite . unfortunately , poor households are facing increasing financial difficulties , resulting in substantial delays in pep delivery ( 7 ) . moreover , the current results indicate that respondents believe that ogds are a public health problem in the community and lead to stray dogs proliferation . most wastes had a household origin and stray dogs were mostly attracted by fermentable wastes from cooking or lapsed foods . these measures should be implemented by the cameroonian government , which took the initiative to increase the number of veterinary vaccination centers all over the country . this decision is facing some discrepancies as many dogs in this country , like elsewhere in africa , had no owner . an integrated rabies control measure coupling dog vaccination , elimination of factors that contribute to dog free - roaming and proliferation must be implemented . these integrated measures can lead to rabies elimination for the reason that in cameroon like elsewhere in central and west africa , domestic dogs are the only rabies vectors ( 39 ) . studies in other countries have shown that children are more frequently bitten ( 6 ) . therefore , awareness education should be planned , targeting children who are usually responsible for the transport of garbage to dumps ; that is , they are more likely to meet stray dogs scavenging for food around dumps . this study reveals that ogds near residential houses and stray dogs increase the risk factor for canine rabies transmission in biyem - assi health district . therefore , if a sustainable effort to maintain sufficient vaccination coverage in the domestic dog populations is a crucial step towards global elimination of canine rabies ( 32 ) , an integrated control method including environmental cleaning up through the improvement of solid waste management to avoid the proliferation of ogds is therefore necessary . the authors have not received any funding or benefits from industry or elsewhere to conduct this study . | backgroundrabies is a neglected enzootic disease which represents a serious public health problem . in cameroon ,
efforts to prevent human deaths caused by rabies are often thwarted by the lack of community awareness .
the community knowledge , as well as attitudes and perception on rabies , is therefore important for both prevention of human deaths and control in animals.methodsa cross - sectional study was carried out to evaluate the level of community knowledge as well as the role of open garbage dumps ( ogds ) in the epidemiology of human rabies .
overall 420 heads of household were interviewed in the biyem - assi health district of yaound .
ogds were identified through a systematic check , and household wastes they contained were characterized.resultsalthough 66.9% of respondents have knowledge on stray dogs , only 35% of respondents knew the role of ogds in the increase of stray dog population .
overall ogds consisted of fermentable wastes .
nutrition places for stray dogs were wild garbage dumps ( 68.1% ) , markets ( 18.3% ) , and houses ( 13.6% ) .
the feeding behavior of stray dogs correlated significantly with the human rabies transmission (
2=154.12 , df=4 , p<0.05).conclusionmost participants knew that rabies could be transmitted by a dog bite as well as the measures to be taken in this type of situation .
increased knowledge of respondents on rabies showed ogds and stray dogs as significant risk factors for canine rabies in biyem - assi health district . |
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the best evidence for the role of vitamin d in health is related to its importance to bone health [ 1 , 2 ] . for example , early life supplementation in children has been associated with decreased risks of type 1 diabetes and influenza [ 3 , 4 ] . despite its positive role in health , given the important role of vitamin d in health , there has been much discussion about the best ways to prevent and treat vitamin d deficiency [ 3 , 6 ] . prevention in children is a priority as it may relate to skeletal and immune development . previous research has demonstrated that the vitamin d levels of women and their neonates are highly related [ 711 ] . the goal of this work was to examine whether there are any particular subgroups in which the mother 's prenatal 25-hydroxyvitamin d [ 25(oh)d ] level and her child 's cord blood level of 25(oh)d are not strongly related . these analyses could identify pregnant women who need a prenatal vitamin d intervention tailored to them based on the various characteristics , as well as identifying children who may have a lower level of 25(oh)d at birth and may be prioritized for vitamin d screening . the birth cohort studied here is part of an nih and institutionally funded cohort study that enrolled pregnant women receiving care at obstetrics clinics in a health system in the detroit , michigan , usa , area for longitudinal study of their children through early childhood with the goal of examining early life exposures related to childhood allergies and asthma . briefly , women were enrolled in their 3rd trimester at which time they provided a blood sample and completed an interview about their health . the child 's cord blood was collected at delivery . only maternal - child pairs in which the child was black / african american or white / non - hispanic / non - middle eastern had their 25(oh)d levels determined as there were insufficient maternal - child pairs for analyses in other race and ethnicity groups . 25(oh)d3 ( cholecalciferol which is sun related ) , was measured in frozen plasma samples ( 80c ) in the laboratory of dr . an hplc method was used and has been used in previously published research [ 1522 ] . 25(oh)d is expressed in ng / ml . 25(oh)d was measured in the stored samples from pregnancy ( 3rd trimester ) and delivery ( cord blood ) . for those with 25(oh)d levels below the lowest detectable limit of 5 ng / ml , a value of 2.5 ng / ml was assigned . this assignment is a common practice with lab values in research as it allows results to be retained in analyses of continuous measures rather than being removed from analyses due to lack of an actual value . for the analyses , we examined whether maternal - child correlations in subgroups would vary between those in which maternal 25(oh)d was and was not low . we chose the a priori cutpoints of 20 ng / ml and 15 ng / ml . while 20 ng / ml defines deficiency , we knew that many of the mothers in the analyses had even lower levels of 25(oh)d . thus , we also chose to examine the cutpoint of 15 ng / ml . although limited by sample size , we also examined the associations when maternal levels were less than 40 ng / ml versus levels at least 40 ng / ml ( n = 218 and n = 23 , resp . ) as it has previously been shown that 40 ng / ml is the level at which 25(oh)d conversion to 1,25-dihydroxyvitamin d3 [ 1,25(oh)2d3 ] is maximized . the maternal prenatal chart was reviewed for key information used in the analyses including maternal height , parity , delivery type , gestational age at delivery , and the child 's birthweight . the maternal weight taken at the first prenatal appointment was recorded and this measurement occurred an average of 7.5 ( sd = 5.6 ) months prior to the child 's birth ; maternal weight was defined as obese if body mass index ( bmi ) 30 kg / m or as classes ii - iii obese if bmi 35 kg / m . allergen - specific ige ( sige ) levels in mothers ' prenatal blood samples were assessed for a set of allergens ( dermatophagoides farinae , dog , cat , timothy grass , ragweed , alternaria alternata , egg , and german cockroach ) . maternal atopy was defined as having at least one sige 0.35 iu / ml . descriptive statistics were calculated to provide an overview of the factors and the 25(oh)d levels . we then took a multistep approach to examine the association between the maternal prenatal and cord blood 25(oh)d levels . subgroups were defined by child race ( black or white ) , delivery type ( c - section or vaginal ) , winter birth ( december , january , or february ) , firstborn status ( yes / no ) , maternal bmi ( kg / m at first prenatal appointment , < 30 kg / m/30 kg / m , and < 35 kg / m/35 kg / m ) , preterm birth ( < 37 weeks/37 weeks ) , birth weight ( 2500 g/>2500 g ) , maternal prenatal 25(oh)d , and maternal atopic status ( yes / no ) . these variables were chosen as they are among the most commonly studied characteristics of pregnancy and delivery . maternal atopic status was included because the primary goal of the cohort was to study allergic outcomes in the children and prenatal and early life vitamin d levels have been investigated for their role in allergic disease development [ 22 , 2427 ] . we then calculated cord blood as a percentage of the prenatal level and compared characteristics between those who did and did not have a percentage at least 25% and 50% of the maternal level . finally , we compared characteristics of those who were and were not above the lowest limit of detection . factors compared between the groups include child race , winter birth , delivery type , firstborn status , maternal atopic status , birthweight , gestational age , and maternal prenatal level of 25(oh)d . maternal average age was 30.3 ( sd = 5.4 ) years and most of the children were black ( n = 175 , 72.6% ) and were not born in the winter ( 81.7% ) . almost half the women were obese ( 46.4% , bmi > 30 kg / m ) , about a third of the pregnancies were by c - section ( 36.5% ) and most of the children were not firstborn ( 81.3% ) . descriptive information about the overall prenatal and cord blood levels of 25(oh)d is provided in table 1 . the mean cord blood 25(oh)d level ( 10.9 ng / ml ) is nearly half of the mean maternal prenatal level ( 23.6 ng / ml ) . white children and their mothers tended to have higher 25(oh)d levels compared to black children and their mothers ( tables 1 and 2 ) . lower prenatal and cord blood levels were found in those who were not firstborn , those mothers with low 25(oh)d levels ( < 40 ng / ml , < 20 ng / ml , and < 15 ng / ml ) , and those who were obese ( 30 kg / m and 35 kg / m ) . lower prenatal levels were found among mothers who had a low birthweight infant . no statistically significant differences were found between those who were and were not born in the winter and were of preterm or low birthweight , or by delivery mode or by maternal atopic status . the prenatal and cord blood levels were highly correlated , overall ( r = 0.75 ) and for most subgroups including those defined by winter birth , firstborn status , maternal bmi , preterm birth , low birth weight status , delivery type , and maternal atopic status . the correlation among black children was less than that of the white children , although both correlations were quite strong ( r = 0.65 and 0.87 , resp . ) . when maternal levels of 25(oh)d were low , the correlation with the cord blood was weak . for example , when the maternal level was less than 15 ng / ml , the correlation was only 0.16 . the correlation was stronger , albeit not as strong as the overall , when the maternal prenatal level was less than 20 ng / ml ( r = 0.29 ) . most of the children had a 25(oh)d cord blood level that was 50% or less of their mother 's prenatal level ( n = 149 , 61.8% ) and 37 children ( 15.4% ) had a level that was less than 25% of their mother 's level . no factors were associated with the level being less than 50% ( data not shown ) ; however , there were two factors associated with the level being less than 25% : winter birth and maternal prenatal level . those children with the percentage less than 25% were more likely to be born in the winter months ( 29.7% versus 16.2% , p = 0.05 ) and have a lower prenatal maternal level ( mean maternal prenatal level = 18.9 ng / ml versus 24.5 ng / ml , p = 0.003 ) . we also examined the characteristics of those children whose 25(oh)d levels were above and below the lowest detectable limit ( table 4 ) . children who had levels below the detectable limit were more likely to be black and less likely to be firstborn or have an atopic mother . the mean prenatal level tended to be higher for those who had a detectable 25(oh)d level . in these analyses of this birth cohort , the correlation between maternal prenatal and cord blood 25(oh)d is quite strong overall . however , the maternal - child correlation is much weaker when the maternal level is low . white maternal - child pairs had higher correlations than the black maternal - child pairs and black children had lower percentages of their maternal prenatal 25(oh)d levels . these results are likely due to the generally lower level of 25(oh)d among black women and suggest that mothers may insufficiently contribute to the child 's 25(oh)d supply once their own level falls too low ( lower threshold ) . the data also demonstrate that it is inappropriate to use a prenatal 25(oh)d level to represent the 25(oh)d in a child 's early life . furthermore , these data suggest that children who are not firstborn will have a lower percentage of their maternal level . this could be attributed to the fact that maternal prenatal 25(oh)d levels were higher in women carrying their firstborn child . this could also reflect that ( 1 ) maternal stores may be depleted from prior births and have not recovered and ( 2 ) prenatal supplementation in parous women may not be sufficient to eliminate low 25(oh)d levels . in a study of 92 pregnant women in saudi arabia , women with two or more previous births were significantly more likely to have lower 25(oh)d3 levels compared with those with one previous birth ( p < 0.05 ) . the authors also reported a significant correlation between maternal serum and neonatal 25(oh)d3 ( r = 0.89 , p = 0.01 ) . our results complement and , by adding extensive subgroup analyses , add an additional important dimension to previously published analyses of examinations of prenatal - cord correlations and factors predicting cord blood 25(oh)d levels [ 7 , 29 ] . reported that neonatal vitamin d had a positive correlation with maternal vitamin d levels ( r = 0.69 , p < 0.001 ) in those mothers with normal vitamin d levels but not in those with hypovitaminosis ( n = 123 maternal - child pairs ) . godang found a strong positive association between maternal 25(oh)d and cord 25(oh)d ( = 0.42 , p < 0.001 ) in a subset of 202 scandinavian women but did not examine the association in subgroups . examined whether prepregnancy obesity predicted poor vitamin d status in neonates in their study of 400 women in pittsburgh , pennsylvania . prenatal ( 422 weeks ) levels of 25(oh)d were lower for women who were obese before pregnancy compared to women who were lean even after adjusting for season and other factors . this difference likely led to the result in which women who were obese before pregnancy were more likely to have delivered a child with vitamin d deficiency compared to women who had a normal bmi ( odds ratio = or = 2.1 , 95% confidence interval = ci 1.2 , 3.6 ) . while we did not see differences by antenatal bmi , the work of bodnar et al . a limitation of our study is that maternal prenatal 25(oh)d , while measured in the 3rd trimester , was not measured at the same time in the 3rd trimester for all women . we did not have maternal prepregnancy weights and relied on the weight measured at the time of the first prenatal care visit ; however , holland et al . suggest similar bmi categorization based on first measured pregnancy weight and self - reported prepregnancy weight . furthermore , there may be maternal characteristics that were not collected for these analyses that may identify other subgroups with variable prenatal - cord correlations . our goal was to examine whether the maternal - child 25(oh)d correlation varied within subgroups rather than to examine predictors of a child 's cord blood 25(oh)d level as previous studies have already highlighted the importance of the maternal level [ 8 , 9 , 11 , 30 ] . the results from this birth cohort suggest that a child 's 25(oh)d level is only a fraction of their mother 's prenatal level . furthermore , the degree to which the newborn 's level is associated with their mother 's prenatal level varies by several interrelated factors including maternal race , birth order , and the actual maternal level . not only do these studies indicate that use of the prenatal level of 25(oh)d to represent the child 's early life vitamin d level is not specific , but also the data suggest that a maternal threshold exists below which the mother limits her contribution of 25(oh)d to the fetus . the designs and analytical plans of future studies of prenatal dietary interventions should consider the possibility of a threshold effect when considering the maternal contribution to the fetus . furthermore , children who are black and not firstborn and those born to women with very low 25(oh)d may identify a priority group for vitamin d deficiency screening . the degree to which a newborn 's vitamin d [ 25(oh)d ] level is associated with their mother 's prenatal level varies by several interrelated factors including maternal race , birth order , and the actual maternal level . a maternal threshold may exist below which the mother limits her contribution of 25(oh)d to the fetus ; low thresholds should be assessed for other prenatal nutrients . the designs and analytical plans of future studies of prenatal dietary interventions should consider the possibility of a threshold effect when considering the maternal contribution to the fetus . children who are not firstborn , those who are black , and those born to women with very low 25(oh)d may identify a priority group for vitamin d deficiency screening . | vitamin d levels of pregnant women and their neonates tend to be related ; however , it is unknown whether there are any subgroups in which they are not related .
25-hydroxyvitamin d [ 25(oh)d ] was measured in prenatal maternal and child cord blood samples of participants ( n = 241 pairs ) in a birth cohort .
spearman correlations were examined within subgroups defined by prenatal and delivery factors .
cord blood as a percentage of prenatal 25(oh)d level was calculated and characteristics compared between those who did and did not have 25% and 50% of the maternal level and those who did and did not have a detectable 25(oh)d level .
the correlation among black children was lower than in white children .
when the maternal 25(oh)d level was < 15 ng / ml , the overall correlation was r = 0.16 .
most children had a 25(oh)d cord blood level less than half of their mother 's ; 15.4% had a level that was < 25% of their mother 's .
winter birth and maternal level were associated with the level being less than 25% .
children with undetectable levels were more likely to be black and less likely to be firstborn .
these data suggest mothers may reduce their contribution to the fetus 's 25(oh)d supply once their own level becomes low . |
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coffee intake was assessed using a semiquantitative frequency questionnaire sent to the participants in 1986 , 1990 , 1994 , 1998 , and 2002 . the validity and reliability of the frequency questionnaire has previously been described ( 6 ) . the end points were incident chd ( defined as nonfatal myocardial infarction or fatal chd ) , stroke , and mortality . the diagnosis of outcomes has previously been described ( 4 ) . briefly , myocardial infarction was confirmed if it met the criteria of the world health organization of symptoms and the patient 's records showed diagnostic electrocardiographic changes or elevated cardiac enzyme levels . stroke was confirmed by medical records according to the criteria of the nationalsurvey of stroke , which define it as a constellation of neurological deficits , sudden or rapid in onset , lasting at least 24 h or until death . deaths were reported by next of kin or the postal system or ascertained through the national death index . cox proportional hazards regression was used to investigate the association between coffee consumption and incidence of cardiovascular events and all - cause mortality . multivariable models were adjusted for age , smoking status , bmi , physical activity , alcohol intake , parental history of myocardial infarction , hypertension , hypercholesterolemia , duration of diabetes , hypoglycemic therapy , and dietary factors ( total energy intake ; use of multivitamin and vitamin e supplements ; polyunsaturated , saturated , and trans fat intake ; glycemic load ; and cereal fiber and folate intake ) using categorical variables . the median value of each category of coffee consumption was modeled as a continuous variable to test for linear trends . all analyses were performed with sas software ( version 8.2 ; sas institute , cary , nc ) . coffee intake was assessed using a semiquantitative frequency questionnaire sent to the participants in 1986 , 1990 , 1994 , 1998 , and 2002 . the validity and reliability of the frequency questionnaire has previously been described ( 6 ) . the end points were incident chd ( defined as nonfatal myocardial infarction or fatal chd ) , stroke , and mortality . briefly , myocardial infarction was confirmed if it met the criteria of the world health organization of symptoms and the patient 's records showed diagnostic electrocardiographic changes or elevated cardiac enzyme levels . stroke was confirmed by medical records according to the criteria of the nationalsurvey of stroke , which define it as a constellation of neurological deficits , sudden or rapid in onset , lasting at least 24 h or until death . deaths were reported by next of kin or the postal system or ascertained through the national death index . cox proportional hazards regression was used to investigate the association between coffee consumption and incidence of cardiovascular events and all - cause mortality . multivariable models were adjusted for age , smoking status , bmi , physical activity , alcohol intake , parental history of myocardial infarction , hypertension , hypercholesterolemia , duration of diabetes , hypoglycemic therapy , and dietary factors ( total energy intake ; use of multivitamin and vitamin e supplements ; polyunsaturated , saturated , and trans fat intake ; glycemic load ; and cereal fiber and folate intake ) using categorical variables . the median value of each category of coffee consumption was modeled as a continuous variable to test for linear trends . all analyses were performed with sas software ( version 8.2 ; sas institute , cary , nc ) . between 1986 and 2004 ( 24,121 person - years of follow - up ) , we documented 435 cases of incident cvd ( 324 chd and 111 stroke ) and 538 deaths from all causes ( 215 from chd or stroke , 145 from cancer , and 178 from other causes ) . in both age- and smoking - adjusted analyses and multivariable analyses adjusting for lifestyle and other cardiovascular risk factors , we observed no association between caffeinated coffee consumption and a higher risk of chd , stroke , or all - cause mortality ( table 1 ) . additional adjustment for dietary factors did not substantially change the results . similarly , caffeinated coffee consumption was not associated with risk of cardiovascular death ( relative risks [ rrs ] 0.64 [ 95% ci 0.351.17 ] for once per month to four times per week , 0.84 [ 0.491.44 ] for five to seven times per week , and 0.58 [ 0.311.10 ] for two or more cups per day compared with the risks for those who did not consume caffeinated coffee ; p for trend = 0.26 ) . we also examined decaffeinated coffee consumption in relation to risk for cvd and mortality and did not observe significant associations ( data not shown ) . rrs ( 95% ci ) for cvds and total mortality by caffeinated coffee consumption and total caffeine intake among men with type 2 diabetes ( 19862004 ) adjusted for age ( 5-year categories ) , smoking status ( never , past , or current at 114 or 15 cigarettes / day ) , bmi ( < 23.0 , 23.024.9 , 25.029.9 , or 30.0 kg / m ) , alcohol intake ( 0 , 0.14.9 , 5.014.9 , or 15 g / day ) , parental history of myocardial infarction , history of hypertension , hypercholesterolemia , physical activities ( quintiles of mets / week ) , duration of diabetes ( < 5 , 510 , or 10 years ) , and hypoglycemic medication ( yes or no ) . adjusted for the variables cited for model i and dietary factors , including total energy intake ; multivitamin use and vitamin e supplement use ; intake of polyunsaturated , saturated , andtrans fat ; long - chain n-3 fatty acids ; cereal fiber ; folate ; glycemic load ( all in quintiles ) ; and decaffeinated coffee and tea consumption . stratified analyses showed no direct association between coffee consumption and cvd risk in any subgroups by risk factor status , including overweight , smoking status , duration of diabetes , hypertension , parental history of myocardial infarction , and aspirin use ( supplemental table 1 , available in the online appendix [ http://care.diabetesjournals.org/cgi/content/full/dc08-2251/dc1 ] ) . in this prospective study in diabetic men , higher habitual coffee consumption was not associated with a higher risk of cvd or all - cause mortality . several studies showed that caffeine acutely impaired postprandial glucose metabolism in diabetic patients ( 8,9 ) . in addition , concerns have been raised in short - term trials that caffeine increases blood pressure ( 10 ) and homocysteine levels ( 11 ) . however , findings from short - term caffeine intervention studies can not be extrapolated to the effects of chronic coffee consumption on risk of cvd . it has been shown that caffeine results in a larger increase in epinephrine concentrations than intake of the same amount of caffeine in coffee ( 12 ) . moreover , coffee contains various substances such as antioxidants ( i.e. , chlorogenic acid ) that may improve glucose metabolism and insulin sensitivity ( 13 ) . second , the acute effects of caffeine could be transient because partial tolerance to the humoral and hemodynamic effects of caffeine among habitual drinkers might develop after several days of use ( 14 ) . in this study , the availability of updated measures of coffee and covariates during the follow - up enabled us to incorporate changes in coffee consumption into the analysis . because coffee drinking is often thought to be an unhealthy habit , people may quit or reduce the consumption of coffee to improve their health after developing hypertension or hypercholesterolemia . these changes would dilute a possible positive association between coffee and chd or stroke . to reduce this bias , we excluded subjects with hypertension or hypercholesterolemia at baseline . we also conducted a sensitivity analysis where we used short - term caffeinated coffee consumption in relation to cvd and mortality , which yielded very similar results . as illustrated by the upper limits of 95% cis of our rr estimates , we can not exclude the possibility that we missed an association between coffee consumption and a modestly higher risk of cvd due to chance . however , results from a previous study in finnish individuals with diabetes support the lack of a direct association and even suggest an inverse association between coffee and cvd mortality ( 5 ) . in conclusion , in this large prospective study of u.s . men , our findings do not support the hypothesis that habitual caffeinated coffee consumption increases risk of cardiovascular events or mortality among individuals with type 2 diabetes . | objectivecoffee consumption has been linked to detrimental acute metabolic and hemodynamic effects .
we investigated coffee consumption in relation to risk of cvds and mortality in diabetic men.research design and methodswe conducted a prospective cohort study including 3,497 diabetic men without cvd at baseline.resultsafter adjustment for age , smoking , and other cardiovascular risk factors , relative risks ( rrs ) were 0.88 ( 95% ci 0.501.57 ) for cvds ( p for trend = 0.29 ) and 0.80 ( 0.411.54 ) for all - cause mortality ( p for trend = 0.45 ) for the consumption of 4 cups / day of caffeinated coffee compared with those for non coffee drinkers .
stratification by smoking and duration of diabetes yielded similar results .
rrs for caffeine intake for the highest compared with the lowest quintile were 1.02 ( 0.701.47 ; p for trend = 0.96 ) for cvds and 0.96 ( 0.641.44 ; p for trend = 0.69 ) for mortality.conclusionsthese data indicate that regular coffee consumption is not associated with increased risk for cvds or mortality in diabetic men . |
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robust evidence demonstrates that sexual minority populations have poorer access to care , poorer care , and higher morbidity than heterosexual populations in areas such as mental health , sexual health , and cancer [ 13 ] . over the last decade , public health authorities and international health organisations have developed policies to eliminate these health disparities [ 48 ] . effectively developing such policies and monitoring their outcomes require official data on sexual orientation that is timely , accurate , reliable , comparable , and of overall high quality [ 9 , 10 ] . producers of official health statistics throughout the world have started collecting sexual orientation data by adding sexual orientation questions in official health surveys . the united states department of health and human services has obtained data on sexual orientation in several surveys since the early 1990s . the united kingdom office for national statistics conducted the sexual identity project from 2006 to 2009 to develop and trial sexual identity questions and recently began the standard implementation of these questions in several surveys . new zealand 's official statistics system ( oss ) , which comprises around 70 government departments that produce official statistics , has also gained significant experience in this area . more specifically , the ministry of health has conducted various health surveys which included sexual orientation questions , and statistics new zealand has led the initiative of considering sexual orientation as an official social ( including health domain ) statistic . despite these advancements in collecting official sexual orientation data , little progress has been made in developing standard definitions and concepts for these data . the united kingdom office for national statistics came closest to developing such standards when it explored the concept of sexual identity ( one dimension of sexual orientation ) in focus groups . however , to our knowledge no producer of official statistics has developed explicit standard definitions and concepts to guide the collection of survey data on sexual orientation . this is despite experts calling for the development , testing , and selection of unambiguous standard definitions and concepts as the foundation for the measurement of sexual orientation in official surveys [ 9 , 10 ] . conceptual frameworks are well - recognised tools that producers of official statistics commonly use to support measurement , data analysis , and analytical commentary . the australian bureau of statistics noted that a primary function of a framework is to map the conceptual terrain surrounding an area of interest [ ] . frameworks can define the scope of inquiry , delineate the important concepts associated with a topic , and organize these into a logical structure ( page 15 ) . according to the bureau , the nature and scope of the topic , the purpose of the framework , and the perspectives of its designers ultimately determine the final content and form a framework takes . successful frameworks are logical in structure , comprehensive but concise , dynamic and flexible to allow for change , and cognisant of other frameworks , classifications , and standards . these frameworks should also be widely accepted to promote standards , consistency and comparability across data collections and between jurisdictions ( e.g. , states and countries ) and sectors ( e.g. , public and private ) ( page 15 ) . we conducted the sexual orientation data collection study in new zealand on behalf of the ministry of social development and its partners , the ministry of health and statistics new zealand . through this study we sought to develop a conceptual framework for the collection of sexual orientation data in official probability surveys in new zealand that would be coherent and theoretically robust and that would adhere to oss requirements . it then discusses the implications of the framework in the context of the emerging global movement of collecting sexual orientation data in official surveys . we used a multimethod , multisource research approach to develop the conceptual framework of sexual orientation . a systematic search of academic and grey literature was carried out across various scientific databases ( academic search complete ; international bibliography of the social sciences ; ovid medline(r ) 1946 to present with daily update ; psycinfo ; sociological abstracts ) , using a range of keywords to identify literature on the definition , conceptualisation , and measurement of sexual orientation and on the collection , analysis , and reporting of sexual orientation data in probability surveys . we reviewed this literature and conducted a secondary analysis that identified key definitions and concepts of sexual orientation . we drafted a first version of the sexual orientation conceptual framework . to ensure that the framework 's development was informed by community expertise , we sought feedback on our draft from sexual minority individuals . we held 90-minute focus groups in new zealand 's two largest cities auckland and wellington ( four participants and five participants , resp . ) and one 60-minute key - informant interview with a total of ten sexual minority individuals , including takatpui
, fa'afafine
, lesbians , gays , and bisexuals . purposive sampling of the participants ensured a diversity of knowledge and opinions , with an emphasis on significant representation of indigenous ( mori ) perspectives . to ensure the appropriate consideration of oss requirements , we also conducted 60-minute key - informant interviews with four producers and consumers of official statistics . key informants included senior policy staff from the ministry of health , the ministry of social development , and statistics new zealand . we conducted a thematic analysis of the focus - group and interview data to identify the key sexual orientation concepts identified by the sexual minority key informants . the advisory board of the sexual orientation data collection study comprised 14 national and international experts on official statistics and on the collection of sexual orientation data . this board provided us with ongoing peer review throughout the development of the sexual orientation conceptual framework . several staff from the ministry of social development and statistics new zealand , as well as mori experts , including from te puni kkiri / the ministry of mori development , commented on draft versions of the framework . the frameworkproposes working definitions for the sexual orientation statistical topic and its measurement concepts , describes key dimensions of the sexual orientation measurement concepts , discusses important variables framing the sexual orientation measurement concepts , outlines conceptual grey areas . proposes working definitions for the sexual orientation statistical topic and its measurement concepts , describes key dimensions of the sexual orientation measurement concepts , discusses important variables framing the sexual orientation measurement concepts , outlines conceptual grey areas . the conceptual framework is about sexual orientation and hence does not discuss gender identity concepts per se . gender identity is a conceptually distinct variable that must not be confused with sexual orientation . however , gender identity does inform and frame sexual orientation in some sexual minority groups and is therefore discussed as a variable that frames sexual orientation in the framework . the conceptual framework is currently limited in scope to adult sexual orientation though it includes some literature on adolescent sexual orientation , where this literature is relevant to adult populations . to fully cover adolescent sexual orientation concepts , the framework may need to be extended . through our study we came to understand sexual orientation in the context of official statistics as a statistical topic with three key measurement concepts : sexual attraction , sexual behaviour , and sexual identity . we reviewed existing definitions of sexual orientation but did not consider these fit for adoption by the oss . consequently we developed a working definition for the sexual orientation statistical topic ( table 1 ) , which defines the key measurement concepts , their relationships , and conceptual features . we reviewed existing definitions of the three sexual orientation measurement concepts and adopted or adapted those that promoted standards suitable for the oss ( table 2 ) . if applied consistently , the proposed definitions will contribute to the conceptual standardisation of health data on sexual orientation and thus improve the conceptual quality of these data . these working definitions should be developed further as scientific knowledge and experience in this area advance . the working definitions differentiate three key dimensions of sexual orientation : sexual attraction , sexual behaviour , and sexual identity . the academic literature generally suggests that sexual orientation comprises these concepts [ 19 , 2123 ] . some definitions of sexual orientation include other concepts , including sexual fantasy and sexual desire . however , such concepts overlap the three measurement concepts proposed in the framework or are peripheral to the conceptualisation of sexual orientation for official data collection purposes . the three concepts overlap considerably , but substantial research demonstrates that individuals often do not report consistently across them . for example , many women who report sexual attraction to women engage in same - sex behaviour , but others do not . sexual attraction can , but does not necessarily , result in various sexual behaviour and the adoption of sexual identities . multiple survey questions need to be asked to differentiate the populations encompassed by the three key measurement concepts . ( 3 ) the continuous nature of sexual attraction and the categorical nature of sexual behaviour and identity . sexual attraction to men and women can be conceptualised along two separate continuous scales marked by the extreme poles of not at all sexually attracted and extremely sexually attracted . asexuality ( defined as the absence of sexual attraction ) is indicated by low scores on both scales . official questions that measure sexual attraction along a continuum acknowledge that sexual attraction is a continuous variable [ 19 , 28 , 29 ] . data from such questions is likely to be more accurate than data from questions using distinct categories . sexual behaviour and identity are categorical variables . for official statistical purposes of measuring sexual orientation , sexual behaviour is best understood as having four categories ( i.e. , opposite - sex behaved , both - sex behaved , same - sex behaved , and not sexually active ) . ( 4 ) the fluidity of sexual attraction , behaviour , and identity . the sexual attraction , behaviour , and/or identity of some individuals change over time and across social contexts . for example , one longitudinal study found that sexual attraction was unstable among young adults , who experienced some same - sex sexual attraction , especially amongst women . those with major same - sex attraction experienced less change in sexual attraction over time . . two crucial factors that can lead to changes in sexual attraction , behaviour , and identity are an individual 's level of certainty about their sexual attraction and identity and differences in the extent of their exploration of sexual attraction , behaviour , and identity [ 19 , 31 , 32 ] . others adopt alternative sociopolitical sexual identities , such as queer , which challenge more traditional , rigid ones . the emergence of sexual identity categories such as those that incorporate interrelated gender , gender - identity , and gender - role dimensions also indicates that sexual identity terminology changes and has become increasingly complex over time [ 31 , 33 , 35 ] . an important issue for the measurement of sexual identity is the adoption of multiple current sexual identities ( i.e. , more than one identity at the same time ) . mori and pacific sexual minority people commonly adopt and manage multiple sexual identities . to address this issue , the oss can draw on its vast experience in measuring other multiple current social identities , such as multiple ethnic identities . the framework considers sexual orientation in the context of cultural and gender - related variables . new zealand mori are familiar with sexual orientation as a concept and with its three measurement concepts . they tend to report mori - specific minority sexual identities used historically and in contemporary times ( i.e. , takatpui whine and takatpui tne ) , as well as common western sexual identities ( e.g. , straight / heterosexual , gay , lesbian , and bisexual ) , sometimes alongside each other [ 36 , 3945 ] . new zealand europeans / pkeh are also familiar with sexual orientation as a concept and its three measurement concepts . people from pacific island backgrounds traditionally do not appear to be familiar with the concepts of sexual orientation and sexual identity . however , same - sex sexual attraction and behaviour are relatively common among young , unmarried pacific men . moreover , some pacific people who engage in male same - sex sexual relationships adopt specific sexual or social identities ( e.g. , samoan fa'afafine ) [ 37 , 46 ] . some people from asian backgrounds do not have traditional cultural concepts equivalent to western sexual orientation and sexual identity . however , same - sex attraction and behaviour are not uncommon amongst these groups in new zealand . new migrants are inclined to adopt the sexual orientation concepts of the host culture . whether they do or not depends on several factors , including country of origin and level of acculturation to the host culture [ 33 , 48 , 49 ] . the understandings of sizeable populations need to be considered when official standard measures of sexual orientation are developed . the framework could potentially be extended to include cultural factors other than ethnicity such as national customs , religion , and globalisation . as more details will be added to the framework over time , a discussion of which specific measurement concept is shaped by which specific cultural variable might be valuable . in any case , the oss requires that official statistics span across cultures . therefore , the conceptual framework would benefit from additional research that could determine whether sexual orientation is a concept well understood by all sizeable new zealand populations . differences in how men and women experience their sexual orientation have been described by golden ( see [ 22 , 50 ] ) and others ( e.g. , ) . whether pacific males adopt a gender - typical or gender - atypical role can influence how they conceptualise their sexual attraction , behaviour , and identity . similarly , whether an individual takes an insertive ( active ) or receptive ( passive ) role during ( same - sex ) sexual behaviour can affect how they conceptualise their sexual identity [ 37 , 49 , 50 ] . for example , taking a receptive role during sexual activity is associated with the adoption of a sexual minority identity ( such as fa'afafine among samoan males ) , whereas taking an insertive role is not . some transgender people conceptualise their sexual attraction , behaviour , and identity differently depending on whether they predicate these on their ( biological ) sex or gender identity [ 15 , 37 , 52 , 53 ] . if a sexual orientation question does not specify whether it expects respondents to base their response on biological sex or gender identity , then transgender people are likely to respond to it in relation to their gender identity . however , in one official health survey conducted in the united states , transgender participants reported their sexual orientation in relation to their biological sex because they assumed that biological sex was more important than gender identity in a health context ( sell , unpublished ) . this highlights that some transgender people may consider the survey context when deciding which variable to base their sexual orientation on , while others explicitly reject the notion that they must necessarily base their responses on an assessment of the potential relevance of their assigned sex . also , some do not identify completely with traditional gender identities of male and female . for these people and for those who are discovering or coming to terms with their gender identity , traditional sexual orientation identities that assume a stable male or female gender identity ( such as heterosexual / straight , gay , and lesbian ) are likely to be inappropriate . the framework highlights the following culture - specific conceptualisations of sexual orientation that require further investigation : mori preferences relating to sexual identity categories.pacific people 's understandings of the sexual orientation topic and of their sexual identity concepts , including especially the samoan fa'afafine concept
.asian people 's understandings of the sexual orientation topic and of their sexual identity concepts . pacific people 's understandings of the sexual orientation topic and of their sexual identity concepts , including especially the samoan fa'afafine concept
. asian people 's understandings of the sexual orientation topic and of their sexual identity concepts . topics of special interest include comparing : how transgender and non - transgender people understand their sexual attraction , behaviour , and identityhow survey respondents conceptualise sexual attraction to and behaviour with transgender and non - transgender people . how transgender and non - transgender people understand their sexual attraction , behaviour , and identity how survey respondents conceptualise sexual attraction to and behaviour with transgender and non - transgender people . while this research would be conceptually relevant , we acknowledge the presumed very small estimated size of the transgender population in new zealand . nationally , producers of official health statistics currently engage in or are in the process of setting up the routine collection of sexual orientation data as the basis for robust health policy making and outcomes monitoring . broadly applicable , nationally ( and potentially internationally ) agreed - upon standard definitions and concepts for sexual orientation need to be established to ensure the quality and comparability of the data obtained . this paper has described the development and content of a conceptual framework of sexual orientation tailored to the new zealand context and oss requirements . the united kingdom office for national statistics has gone some way towards establishing a standard definition and measurement concept for sexual identity . however , to our knowledge the conceptual framework described in this paper is the first statistical framework that establishes standard definitions and concepts for the broader sexual orientation statistical topic , which can be applied in an official statistics system . the framework synthesises a large body of contemporary research on the definition , conceptualisation , and measurement of sexual orientation and on the collection , analysis , and reporting of sexual orientation data in probability surveys . the framework has been developed by a small team of researchers but has been peer - reviewed rigorously throughout its development . it has not undergone formal public consultation to date , but its development has been guided by feedback from a small number of demographically diverse individuals with sexual minority orientations , mori reviewers , and a considerable number of producers and consumers of official statistics . the advisory board of the sexual orientation data collection study also gave feedback , which has been integrated . the validity of the framework and the standards it sets will only be established over time , as the framework is applied in the oss . one initial milestone in applying the framework will be the development of standard questions on sexual attraction , behaviour , and identity for use in official surveys . the framework could also form the foundation for developing standard statistical classifications for the three sexual orientation measurement concepts . additionally , it could be used to guide data commentary , including the evaluation of the quality of sexual orientation data . the scope of the current study was to define , conceptualise , and measure a specific statistical concept ( i.e. , sexual orientation ) , for new zealand 's oss . this study identified how cultural and gender - related variables frame sexual orientation and considered how sexual identity fluidity and multiple identities might influence responding to oss sexual orientation questions . there is a growing understanding of a complex interaction between gender identity and sexual orientation . given this , there is a need for future research to investigate collecting gender identity data for official statistics alongside sexual orientation data . such research could build on the findings of the present research and others ( e.g. , ) . globally , as more producers of official statistics collect data on sexual orientation , the cross - country comparability of these data will need to be assured through international coordination and standard setting . we believe that the standard definitions and concepts proposed in our framework are potentially transferable to some countries , especially those that are historically and demographically similar to new zealand such as australia and the united states . these countries also have sizeable indigenous and pacific island populations that require consideration of concepts and categories other than the common western ones . other countries will need to develop equivalent frameworks of sexual orientation to suit their specific contexts and the requirements of their producers and consumers of official statistics . the producers of official statistics will need to establish a systematic process that allows them to share information related to the definition and conceptualisation of sexual orientation . this will enable them to compare across countries the sexual orientation data that they collect . we have developed a coherent and theoretically robust conceptual framework that sets standard definitions and concepts for collecting official sexual orientation data in new zealand 's oss . the validity of the framework and the standards it sets will be established over time , as the framework is applied in the oss . the framework is potentially transferable to official statistics systems in countries historically and demographically similar to new zealand . the framework offers a first model that could help improve the quality of sexual orientation data collected in official surveys . | objective . effectively addressing health disparities experienced by sexual minority populations requires high - quality official data on sexual orientation .
we developed a conceptual framework of sexual orientation to improve the quality of sexual orientation data in new zealand 's official statistics system .
methods .
we reviewed conceptual and methodological literature , culminating in a draft framework . to improve the framework , we held focus groups and key - informant interviews with sexual minority stakeholders and producers and consumers of official statistics .
an advisory board of experts provided additional guidance .
results .
the framework proposes working definitions of the sexual orientation topic and measurement concepts , describes dimensions of the measurement concepts , discusses variables framing the measurement concepts , and outlines conceptual grey areas .
conclusion .
the framework proposes standard definitions and concepts for the collection of official sexual orientation data in new zealand .
it presents a model for producers of official statistics in other countries , who wish to improve the quality of health data on their citizens . |
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this complication occurs in up to 3% of the newborn cases with central venous access . we report a case of cardiac tamponade following central venous cannulation that was diagnosed using a transthoracic echocardiography , managed by urgent surgical pericardial drainage . complications with central line insertion in neonates occur most likely with guide wire aided insertion , with blind needling of a chest vein and with dilators . the risk appears to be greatest when the end of the catheter creates an acute angle to the vessel or cardiac wall . a 10-day - old 2.2 kg female child was operated for obstructed total anomalous pulmonary venous connections ( tapvc ) [ figure 1 ] . intra - operatively right upper lobe vein was not traceable , left upper and lower pulmonary veins were well seen . rerouting of the three pulmonary veins to the left atrium , ligation of vertical vein and patch closure of atrial septal defect was done [ figure 1 ] . child was extubated on the 3 post - operative day . while positioning after extubation , the central line slipped out . nitroglycerin , dobutamine , milrinone and noradrenaline infusions were on flow , when the line slipped out . schematic representation of pre - operative diagnosis , procedure done and the catheter piercing the innominate vein attempted cannulations of right femoral vein were unsuccessful . left internal jugular vein was cannulated with 4f triple lumen catheter . in two lumens , the blood was aspirated freely and the aspiration of blood in the other lumen was negative . at the end of the procedure central vein catheter tip was seen in the thoracic cavity after puncturing the innominate vein [ figure 2 ] . haemodynamics gradually picked up and later right internal jugular vein was cannulated with 4f triple lumen catheter . most serious complications of central lines in neonates are typically mechanical and occur early and a delay in diagnosis can be catastrophic . careful insertion techniques , as well as continued vigilance in the correct position and function of central venous catheters are imperative to help prevent serious complications . when a suspicion of a serious mechanical complication arises , the infusion should be immediately discontinued and adequate interventions such as pericardial or pleural drainage should be performed immediately . in our patient , the acute cardiorespiratory distress necessitated a bedside sternotomy and the patient was found to have a relatively complex injury of catheter coming through the innominate vein and tip lying in the thoracic cavity . the cardiac arrest was immediate as the rent in innominate vein was big and the amount of saline used to flush all three lumen was enough in 2.4 kg baby to cause immediate tamponade ; roughly 5 ml of saline plus and the blood loss caused by innominate vein perforation could have triggered the arrest . furthermore , as it was a state of post - operative obstructive tapvc , any acute insult would be poorly tolerated . complications of perforation from central venous catheters in neonates include cardiac tamponade with catheter tips in the right atrium , right ventricle , superior vena cava and myocardial infiltration in the right atrium . pleural effusion and ascites can occur due to the tip in the inferior vena cava and pericarditis can occur due to the tip in the superior vena cava . rare complications include diaphragmatic paralysis , paraplegia and myoclonus due to tip in ascending lumbar vein and venous sinus thrombosis due to tip in the jugular vein . another rare complication is the erosion of the catheter tip into pulmonary vessels . as a general rule , direct vessel perforation from malposition occurs immediately as happened in our case , whereas osmotic erosion is more likely to occur days to weeks after catheter positioning . in our patient , there was no significant difference between survivors and non - survivors relative to gestational age at birth , birth weight and days to pericardial effusion diagnosis . however , mortality was significantly less in patients who underwent immediate pericardiocentesis . therefore , emergent drainage of a pericardial effusion must be considered in any patient with a central vein catheter with sudden cardiorespiratory instability . the coexistence of acute pericardial and bilateral pleural effusions from central line perforation remains a very uncommon finding , which was present in our case . after subsequent pleural fluid drainage and central line removal , there was a rapid and complete recovery of the cardiorespiratory status . the incidence for central line associated symptomatic perforation is 3% and fatal perforations are 1% , despite a policy of careful placement . the data also indicate that perforation complications occur regardless of the size , material , location of the catheter tip and the access route . with regard to size of the catheter certainly , it is logical to expect that larger catheters may have a higher propensity to cause trauma to vascular or cardiac tissue , and thus , the smallest catheter required should always be used . however , catheter size is generally standardised and somewhat difficult to alter , and therefore , no definitive relationship is shown between size and development of cardiac tamponade . the recent technical advancement like development of thin silastic and polyurethane catheters has not eliminated the perforation of the great veins of the heart and cardiac tamponade . this is most likely due to two practical problems ; first , because of the difficulty in visualising the course of central line and second , because of the migration of the catheter after insertion . the speed and the amount of volume infused during cardiopulmonary resuscitation as it happened in our case may also be a contributing factor for perforation . smaller children , haemodialysis central line catheter and the team 's learning curve can also be risk factors for perforation . studies in vitro and in adults suggest that an increased angle of incidence between the catheter tip and the cardiac / vessel wall increases the likelihood of perforation . the tip of the catheter is not fixed and lateral neck flexion or arm movement may cause perforation , with arm movement causing greater catheter tip movement than neck movement . in response to reports of death from cardiac tamponade , some catheters now bear a manufacturer 's recommendation that they should not be placed with the tip in the right atrium . however , catastrophic cardiac tamponade was reported even when the catheter tip was outside the right atrium as happened in our case . although isolated cases have been reported in adults , a search of our entire paediatric database showed nil occurrence of catheter coming through the innominate vein into the thoracic cavity . it remains uncertain whether catheter insertion site , location of the catheter tip , material or size play any associated role in the development of this complication . most important to us are restricting operators to experienced personnel , to ensure that blood can be readily and repeatedly withdrawn from all lumens of the catheter , where it is fixed at the completion of insertion , and also to ensure that it is in a large vessel and is not lodged against a vessel or cardiac chamber wall . when a suspicion of a serious mechanical complication arises , the infusions should be immediately discontinued and adequate interventions such as pericardial or pleural drainage should be performed immediately . | cardiac tamponade following central line in a neonate is rare and an uncommon situation ; however , it is potentially reversible when it is diagnosed in time .
we report a case of cardiac tamponade following central line insertion . a 10-day - old 2.2 kg girl operated for obstructed total anomalous pulmonary venous connections had neckline slipped out during extubation . attempted cannulations of right femoral vein were unsuccessful . at the end of the left internal jugular vein cannulaton
, there was a sudden cardiorespiratory arrest .
immediate transthoracic echocardiogram showed left pleural and pericardial collection .
chest was opened and the catheter tip was seen in the thoracic cavity after puncturing the innominate vein .
the catheter was removed and the vent was repaired . |
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epithelioid hemangioendothelioma is a rare vascular tumor with low - grade malignant potential commonly seen in the soft tissues of the extremities , cases have been reported in lung , liver , bone , and skin and few cases in head and neck region . this lesion is usually asymptomatic , can occur in any age group and has female predilection with a female - to - male ratio of 2.5:1 . a 25-year - old female patient presented with a complaint of progressive difficulty in swallowing since 1-year . there was no history of throat pain , cough , fever , stridor , hoarseness of voice , hemoptysis , loss of appetite , weight loss , associated comorbidities , or substance abuse . on examination , hopkins 's laryngoscopy revealed a globular mass projecting into the supraglottis from the lateral wall of the right pyriform fossa [ figure 1a ] . the epiglottis , pharyngoepiglottic folds , vallecula , and base of tongue were also normal . routine blood , urine , chest x - ray , and electrocardiogram investigations were within normal limits . the mass was grayish - white and measured 2.5 cm 2 cm [ figure 1b ] . follow - up laryngoscopy on the 10 postoperative day revealed some slough in the right pyriform fossa . hematoxylin and eosin ( h and e ) stained sections showed round to polygonal tumor cells arranged in cords and small nests having prominent cytoplasmic vacuolization . intraluminal erythrocytes were noted in many of the vacuoles , reminiscent of primitive vascular channels that were confirmed by cd34 immunostain . based on the microscopic findings of h and e and immunostaining , the lesion was diagnosed as epithelioid hemangioendothelioma of pyriform fossa [ figure 2a and b ] . patient has been on follow - up since 1-year without any evidence of loco - regional recurrence or dysphagia [ figure 1c ] . ( a ) hopkins telescopic image preoperative ; ( b ) excised specimen ; ( c ) hopkins telescopic image 1-year postoperative histopathological image ( a ) h and e staining showing numerous proliferating capillaries lined by plump endothelium arranged in lobules and horn - like pattern ; ( h and e , 400 ) . epithelioid hemangioendothelioma is a rare neoplasm of vascular origin initially described by weiss and enzinger in 1982 . vascular tumors composed of histocytoid or epithelioid endothelial cells are divided into epithelioid hemangioma , epithelioid hemangioendothelioma and epithelioid angiosarcoma . however , owing to the overlapping histologic features they are also considered as a continuous spectrum of lesions , where epithelioid hemangioendothelioma represents a borderline or low - grade malignant variant . it is commonly seen in liver , spleen , bone , skin , heart , soft tissues , and vascular system . cases have been reported in the oral cavity , thyroid gland , submandibular area , neck , scalp , larynx , parapharyngeal space , parotid gland , and mandible . the most common site in the head neck region is at or below the level of the mandible and submandibular region . histopathological findings of epithelioid hemangioendothelioma suggest round or spindle- shaped epithelioid cells with pale cytoplasm . on immunohistochemistry , they are positive for endothelial cell markers ( cd31 , cd34 and factor vii - related antigen ) . the present case also shows a typical histopathological picture with cd34 positivity on immunohistochemistry . due to noticeable malignant potential , wide local excision and regular follow - up are the preferred management option . | epithelioid hemangioendothelioma is a rare vascular neoplasm of borderline or low - grade malignant potential uncommonly seen in head and neck region and has not been reported in the hypopharynx .
we present here a case of epithelioid hemangioendothelioma arising from the hypopharynx in a young female patient who presented with progressive dysphagia for 1-year and difficulty in breathing for 1-month . this is first reported case in published english literature to the best of our knowledge .
recognition of this borderline entity is necessary because of its potential for malignant transformation and recurrence
. a wide excision and regular clinical follow - up would be an appropriate treatment protocol .
the role of other therapeutic modalities such as chemotherapy and/or radiotherapy is not yet well established . |
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there are several studies dealing with sexual behavior , people 's attitude towards hiv / aids and hiv testing[14 ] . in zimbabwe , one study found that the prevalence of hiv among monogamous women was 21.8% . these factors are the husbands having children with other women , more than five years age difference within couples , the woman 's age , and the likelihood of the woman discussing monogamy with her husband in the next three months as well as economic disparities between the sexes . in zambia there is a sizable group of hiv positive neurology patients who suffer from myelopathy , neuropathy / radiculopathy , cerebrovascular diseases and infectious diseases . a multi - country study including thailand , south africa , tanzania and zimbabwe , which used a sample of 14,818 in 48 communities , found that frequent discussions about hiv significantly influenced prior hiv testing . a study in vietnam , dealing with hiv risk behavior and the determinants among people living with hiv / aids , found that 82% of the participants were sexually active . some 20% of the participants reported having sex with multiple partners , and only one third of the participants consistently used a condom . jamaica manifests some of these problems relating to hiv / aids , hiv testing , and determinants of risky behaviors seen in other developing countries . sexual risk taking behavior among working class women were influenced by being in a relationship with a physically violent intimate partner , attitudes towards children as resources , having multiple sexual partners , being economically vulnerable , the perception about men being providers and the vetoing power of males on women 's reproductive health matters . choices and the lack of economic dependency are accounting for increasing hiv / aids / stds in developing countries . the risk of contracting hiv also affects the incarcerated where the rate of hiv infection among prisoners is higher than the incidence in the general population . a majority of university students stated that they had no avoidance intentions towards their relatives and friends living with hiv / aids . however , less than one - half of the students were sympathetic to gay and lesbians living with hiv / aids who were not their relatives or friends . it is important to understand the cultural factors at the societal , family and individual levels that influence hiv risk among jamaican youth . in 2004 , from a survey of jamaican youth , males reported greater condom use ( 67.9% ) during their last sexual intercourse with their most recent and regular partner compared to 57% in 1996 . there was no change in the use of condoms among adolescent females for the same period . the youth perceived that they were at a greater risk for hiv infection in 2004 compared 1996 . despite the aforementioned fact , there are still many youths who are inconsistently using a condom , indicating the divide between knowledge and practice . public health effort to improve the case management of sexually transmitted diseases now includes the private sector . private medical practitioners are now receiving continuing medical education to address the hiv / aids epidemic and youths reproductive health matters . despite the increased risk of contracting hiv owing to inconsistent and improper condom usage[811 ] , many jamaicans do not want to know their hiv status . a person knowing his or her hiv status allows the person to get the appropriate medical treatment if they are positive and prevent the spread of the virus through safe sexual practices . douglas opined that the major cause of mortality among women of 15 - 44 years in the caribbean is aids , and that 1 in every 50 caribbean national was infected with hiv / aids , suggesting that there are some premature mortality arising owing to not being cognizant of one 's hiv status . sexual relation is primarily the medium through which most people contract hiv / aids . in 2007/2008 , wilks et al . conducted a study of some 2,848 jamaicans between the ages of 15 - 74 years ; they found that 3.4% of females reported having been infected with a sti in the studied period compared to 1.3% of males . although the prevalence of males having had sti is greater than that of females , which is concurred by wilks et al study ( males , 18.1% ; females , 11.0% ) , with the context that more males ( having multiple partners in last 12 months , 41% ) are promiscuous than females ( 8.4% ) , there is a pending public health problem that is underlying the females allowing males to have vetoing power over their reproductive health issues and having male partners inconsistently using condoms . an extensive research of the literature found no study which has examined the attitude of non - hiv testers towards future hiv testing as well as their current attitude and sexual behavior . within this context , the objectives of this paper are to ( 1 ) investigate the sexual behavior of non - hiv testers in jamaicans , and ( 2 ) their attitudes towards future hiv testing . the current study extracted a sample of 1,192 participants , from a nationally representative survey , who indicated that they had never had an hiv test done up to 2004 . the hiv / aids / std national kabp survey comprised 1,800 participants 15 - 49 years of age who resided in jamaica at the time of the survey ( may - august , 2004 ) . the data was collected by hope enterprises limited on the behalf of the ministry of health . each of the 14 parishes in the country is stratified into electoral constituencies , with each constituency stratified into three areas - rural areas , parish capitals ( urban areas ) and main towns ( semi - urban areas ) . the areas which comprised a constituency were then stratified into primary sampling units ( psus ) or electoral enumeration districts ( eds ) . a random sample of each psu 23 eds in the urban areas , 25 eds in the semi - urban areas , and 24 eds in the rural areas . twenty - five households were systematically chosen from each ed , and cluster sampling was carried out with all the people living in the household of the designated ages interviewed for the survey . the interviewers were trained for a 5-day period , of which 2 days were devoted to field practices . interviewers were assigned to a team comprising two females , two males and a supervisor . the participants were informed of their right to confidentiality and their right to stop the interview at any time . no names , addresses or other personal information was collected from the participants to ensure anonymity . the instrument used in the survey utilized indicator measures and definitions consistent with unaids and the usaid priority prevention indicator . data were entered , stored and retrieved using spss for windows , version 16.0 spss inc ; chicago , il , usa ) . multivariate logistic regressions were fitted using one outcome measure : self - reported confirmed positive hiv test results . where collinearity existed ( r > 0.7 ) , variables were entered independently into the model to determine those that should be retained during the final model construction . a p - value < 0.05 ( two - tailed ) was used to establish statistical significance . the current study extracted a sample of 1,192 participants , from a nationally representative survey , who indicated that they had never had an hiv test done up to 2004 . the hiv / aids / std national kabp survey comprised 1,800 participants 15 - 49 years of age who resided in jamaica at the time of the survey ( may - august , 2004 ) . the data was collected by hope enterprises limited on the behalf of the ministry of health . each of the 14 parishes in the country is stratified into electoral constituencies , with each constituency stratified into three areas - rural areas , parish capitals ( urban areas ) and main towns ( semi - urban areas ) . the areas which comprised a constituency were then stratified into primary sampling units ( psus ) or electoral enumeration districts ( eds ) . a random sample of each psu 23 eds in the urban areas , 25 eds in the semi - urban areas , and 24 eds in the rural areas . twenty - five households were systematically chosen from each ed , and cluster sampling was carried out with all the people living in the household of the designated ages interviewed for the survey . the interviewers were trained for a 5-day period , of which 2 days were devoted to field practices . interviewers were assigned to a team comprising two females , two males and a supervisor . the participants were informed of their right to confidentiality and their right to stop the interview at any time . no names , addresses or other personal information was collected from the participants to ensure anonymity . the instrument used in the survey utilized indicator measures and definitions consistent with unaids and the usaid priority prevention indicator . data were entered , stored and retrieved using spss for windows , version 16.0 spss inc ; chicago , il , usa ) . multivariate logistic regressions were fitted using one outcome measure : self - reported confirmed positive hiv test results . where collinearity existed ( r > 0.7 ) , variables were entered independently into the model to determine those that should be retained during the final model construction . a p - value < 0.05 ( two - tailed ) was used to establish statistical significance . the current study extracted a sample of 1,192 participants , from a nationally representative survey , who indicated that they had never had an hiv test done up to 2004 . the hiv / aids / std national kabp survey comprised 1,800 participants 15 - 49 years of age who resided in jamaica at the time of the survey ( may - august , 2004 ) . the data was collected by hope enterprises limited on the behalf of the ministry of health . each of the 14 parishes in the country is stratified into electoral constituencies , with each constituency stratified into three areas - rural areas , parish capitals ( urban areas ) and main towns ( semi - urban areas ) . the areas which comprised a constituency were then stratified into primary sampling units ( psus ) or electoral enumeration districts ( eds ) . a random sample of each psu 23 eds in the urban areas , 25 eds in the semi - urban areas , and 24 eds in the rural areas . twenty - five households were systematically chosen from each ed , and cluster sampling was carried out with all the people living in the household of the designated ages interviewed for the survey . the interviewers were trained for a 5-day period , of which 2 days were devoted to field practices . interviewers were assigned to a team comprising two females , two males and a supervisor . the participants were informed of their right to confidentiality and their right to stop the interview at any time . no names , addresses or other personal information was collected from the participants to ensure anonymity . the instrument used in the survey utilized indicator measures and definitions consistent with unaids and the usaid priority prevention indicator . data were entered , stored and retrieved using spss for windows , version 16.0 spss inc ; chicago , il , usa ) . multivariate logistic regressions were fitted using one outcome measure : self - reported confirmed positive hiv test results . where collinearity existed ( r > 0.7 ) , variables were entered independently into the model to determine those that should be retained during the final model construction . a p - value < 0.05 ( two - tailed ) was used to establish statistical significance . table 1 presents the demographic characteristics of sample . of the sample ( n=1192 participants ) , 54.4% was males , 42.9% were employed , and the majority had secondary level education . sociodemographic characteristics of sample , n= 1192 of those who had never done a hiv test ( n=1192 ) , 78.0% indicated that they would be willing to do this in the future ; 16.3% have sti ; 55.2% are currently and actively practicing a religion ; 14.9% are laborers ; 38.8% are household helpers and office attendants ; 31.1% are security guards , hairdressers , taxi operators , machine operators and cosmetologist , 3.6% teachers , police officers , nurses and technicians ; 6.8% clerks , 2.2% manager , assistant managers and heads of small businesses , 0.2% business executives , and 0.4% professionals ( doctors , lawyers , architects , etc . ) . when the participants were asked what is the chance of [ you ] catching hiv ? 57.7% indicated none , 30.2% said little , 6.3% reported moderate and 5.8% remarked a good chance . however , 20.5% had sex with at least two partners in the last 4 weeks . only 53.4% used a condom the first time with their current partners , and 36.1% indicated that their current partner has other partner(s ) . fewer than 70% of the sample indicated that their next most recent partner had other sexual partner(s ) . a statistical difference was found between the mean number of sexual partners males had in the last 12 months ( 2.6 , sd = 4.0 ) and females ( existed between the mean age of sexual debut for males ( 14.2 years , sd = 3.2 years ) and that of females ( 16.5 years , sd = 2.8 years ) t - test = -11.443 , p < 0.0001 . table 2 shows information on the reasons the participants gave for not using a condom by current , next recent and next most recent partner . the findings reveal that between 10% and 15% of the participants do not like using condoms with current , next recent or next most recent sexual partner(s ) , and between 3 to 5% of the participants stated that their partner objected to using a condom . reason for not using a condom by current , next recent and next most recent partner table 3 provides condom usage ( ever ) and precautions taken to avoid pregnancy by current , next recent and next most recent partner . the percentage of participants who are using a condom with their current partner(s ) is less ( 68% ) compared to the next recent partner ( 89% ) and the next most recent partner ( 91% ) . condom usage ( ever ) and precautions taken to avoid pregnancy by current , next recent and next most recent partner different reasons were given for the moderate - to - good chance of contracting the hiv virus . the reasons were ; have different partners , 13.6% ; spouse has many partners , 4.1% ; do not use a condom , 7.6% ; had blood transfusion , 1.3% ; never too careful , 14.5% ; and condom can burst , 8.0% . when the participants were asked why they had not done a hiv test , 59.7% indicated that they do not want to know their status , 15.4% said they are not sexually active , 10.0% reported that they know that they do not have hiv , 5.0% reported that they are not interested , 0.5% remarked that they always practice safe sexual relations . eight percent of the participants stated that they were forced to have sexual relations , 7.0% indicated that they had forced someone to have sexual intercourse and 1.7% remarked that physical force was used in the process . significantly more males ( 12.5% ) had forced someone compared to 2.5% of females ( = 32.946 , p < 0.0001 ) . however , 35.9% of females indicated that physical force was used in the process of forced sexual relations compared to 6.7% of males ( = 17.485 , p < 0.0001 ) . furthermore , no statistical association existed between were you forced to have sexual intercourse? based on the gender of the participants males , 9.3% and females , 10.1% ( = 0.190 , p = 0.669 ) . in addition , significantly more people with sti were forced to have sexual relations ( 15.5% ) compared to those who were not forced ( 7.0% ) = 12.894 , p < 0.0001 . table 4 examines particular demographic characteristics of the participants based on gender . some demographic characteristic by gender of participants the results of the multivariate logistic regression model , shown in table 5 , were significant ( model = 32.526 , p = 0.027 ) . table 5 indicates that 20.3% of the variances accounted for by the independent variables used in the regression analyses : age of first sexual debut , age at last birthday , having had sti , sex with commercial worker , union status , and protecting self from the possibility of contracting the hiv virus . the model had statistically significant predictive power ( model = 32.526 , p = 0.027 ; hosmer and lemeshow goodness of fit ( = 3.202 , p = 0.921 ) , and correctly classified 82.5% of the sample ( table 5 ) . logistic regression analyses : variables of willing to do hiv test in the future , n = 950 no statistical existed between would you be willing to do a hiv test in the future? and whether the individual was forced to have sexual intercourse = 0.444 , p = 0.801 : yes to be willing , 9.3% ; no to be willing , 11.0% ; and do nt know , 9.4% . this study provides invaluable insights into those who have done a hiv test and their sexual behavior and attitude towards future hiv testing . from the findings , clearly , people are linking a low risk of contracting the hiv virus with having one sexual partner . based on the perception of respondents in this sample , they are omitting the close association between inconsistent condom usage , not knowing the hiv status of current and past partners , and having unprotected sexual intercourse in a heterosexual way with high risk of contracting the hiv virus . other issues which emerged from the current work are ( 1 ) the low probability of those who have had sex with commercial sex workers willingness to consider doing a hiv test in the future , ( 2 ) the inverse association between having had a sti and the likeliness of wanting to do a hiv test in the future , and ( 3 ) negative association between age of respondents and wanting to do a hiv test in the future . within the context that 16.3% of the sample ; having had stis , 58% indicated a low risk of contracting the hiv virus , 6.2% having had sexual relations with a commercial sex worker , 50.3% consistently using a condom , and 60% not wanting to know their hiv status , and notion that hiv / aids being a homosexual disease , there is a need to execute immediate public health interventions to address these findings . more programs promoting safe sexual practices are definitely needed because between 10 % and 15 % of the participants did not use a condom during sexual intercourse with their current , next recent and next most recent partner(s ) . what is worse is that ; 3 - 5% of the participants stated that their partner objected to them using a condom . moreover , 32% of the participants do not wear a condom with their current partner compared to the 11% who do not wear a condom with their next recent partner . some 16.3% of the participants have stis . despite these risks , only 49.1% of the participants gave several reasons ( participant or spouse having multiple partners , condom failure , negligence , blood transfusion , not wearing a condom ) for a moderate to good chance of contracting hiv . more than one- half of the participants gave no reasons for a good to moderate chance of contracting hiv . it is not surprising then , that 59.7% of the participants did not want to know their hiv status , which creates a problem in the reduction of hiv infection since more than 10% reported that they did not use a condom and 36.1% reported that their current partner has other partners . another explanation which accounts for high risk of contracting stis and hiv / aids , despite the current respondents perception of their seemingly low risk , is women 's acquiescence to the vetoing power of men in reproductive health decisions . a research conducted by who showed that some women opined that reproductive health decisions are made by their partner because he looks after me , suggesting a rationale for inconsistent condom use . significantly more men compared to women declared that they had coerced someone to have sex . similarly , a greater proportion of women ( 35.9% ) stated that physical force was used in coerced sexual intercourse compared to men ( 6.7% ) . participants who experienced coerced sexual relations are less likely to use a condom because of the powerful influence of the situational context in which they find themselves . the data supports this assumption because a significant majority of the participants who were forced to have sex had stis compared to those who were not forced . within the context that hiv is more easily passed from infected males to females than in the reverse and it is predicted that proportionately more cases will occur in females than in males in jamaica in the future , and females being coerced to have sexual intercourse . if this abusive practice is occurring without the use of condoms , it will be accounting for greater risk of hiv infection in the future . an important issue which must be addressed in public health intervention is financial inadequacies of females which account for them relishing their reproductive health choices , often carte blanche to males . jamaica is a middle - income developing country with the prevalence of poverty rate being 9.9% in 2007 ( urban poverty rate , 6.2% ; semi - urban poverty rate , 4.0% ; rural poverty rate , 15.3% ; prevalence of poverty among males , 7.2% ; prevalence of poverty among females , 8.0% ) . economic deprivation of females more than males can also be explained by the unemployment rates , and earnings . the unemployment rate for females was almost 2.3 times more than that of males ( 6.2 ) , indicating that if this economic inequity continues , females will relinquish their reproductive health choices to males as they are providing the economic livelihood . in 2004 , the economic and social survey of jamaica , publication showed that on an average the earnings of males ( mean wage = $ 2.4 million ) was 2 times more than that of females ( $ 1.7 million ) ; and that 76 per cent of senior positions were held by males although 54 per cent of executive and managerial positions were held by females . poverty and unemployment , therefore , are explanations of why people become engaged in risky activities as well as take short - term health risks ; in order to earning a living as they seek a change for the better . it is this belief in socio - economic development that spurs women to relinquish their reproductive health choices to men as they perceive men , as having the economic resources to make an improvement in their current status , even if it is temporary . who stated that , on a yearly basis , there are approximately 340 million new cases of stis and 5 million new cases of hiv infection . clearly , inconsistent condom usage can account for an increase in sti / hiv , indicating that the attitude of non - hiv testers in this sample is reinforcing the pending public health problems that need to be addressed with some degree of urgency . a study by henry - lee found that reasons given by jamaican women for inconsistent contraceptive use or even discontinuation were ( 1 ) cost of contraception , and ( 2 ) long waiting queues at clinics ( 3 ) . she contended that scheduling of appointments could reduce the long waiting time at the clinics . while henry - lee is correct about using appointments to reduce waiting at clinics , but the vetoing supremacy of males in the reproductive health decisions of females could account for their non - return to the clinics . due to the economic disparity , poverty , low education , low income , and inconsistent condom usage , many people do not want to know their hiv status yet they believe that they are at a low risk of contracting the virus . furthermore , significantly more females than males who have never had a hiv test indicated that they are not at risk of contracting the hiv virus , yet statistically more males than females consistently use a condom . clearly , there is a denial among jamaicans of their probability of contracting the hiv virus as people still perceive that the virus is a homosexual disease . a study by dunkle et al . showed that more people that have had heterosexual intercourse had contracted the hiv virus in zambia and rwanda than those who are engaged homosexual relationships . 's work provide some understanding of the rationale behind jamaicans reluctance to have a hiv test done even though they may be in heterosexual relations , as the predominant cultural belief is that it is a homosexual disease . one researcher noted that among jamaican males , there is a substantial peer group pressure to engage in sex in order to avert the stigma of homosexuality , and sexual initiation may occur as early as the age of 8 years indicating the dominance of the culture in causing or reverting to particular and even potentially deleterious actions . the irony here is that caribbean men are socialized to be promiscuous and indulge in premarital sexual relations[2528 ] , which is evident in the male - to - female hiv prevalence ratio ( 1.3:1)[2932 ] , but if they are infected with hiv as a direct consequence of their sexual behavior , they will be scoffed at for having being homosexual . the public health dilemma of hiv infection is that we have been estimating a probability of prevalence among jamaicans[2931 ] . the jamaican ministry of health estimated that 65% of those persons who are infected with hiv are unaware of their status , and this reiterates the rationale for public health intervention programs . due to the sobering reality that people do not want to know their hiv status owing to the stigmatization associated with the virus . currently the nation is not seeing the economic impact of hiv / aids , which may justify why non - hiv testers were not researched on their sexual behavior and attitude towards hiv testing . research findings are in on non - hiv testers , and these should be integrated with public health intervention so as to effectively address some of the present challenges of non - hiv testing in the jamaica and in the wider developing nations . world bank revealed that over 95% of persons living with hiv / aids were in low and middle income countries . of this figure over 20 million of these persons have died from aids in 2002 . this hiv / aids epidemic has reduced life expectancy by more than ten years in many countries . gebre postulated that the caribbean countries have the highest incidence rates of hiv / aids in the americas and the second highest prevalence rates in the world among adults aged 15 44 years . statistics have showed that between 270,000 to 780,000 adults are living with hiv in the region , which is about 2.3% . almost 9453 cases of aids were reported in jamaica and of this amount 1100 of these cases were identified in 2004 . this is an estimated 1.6% of the adult population that has been infected with hiv . according to who , 60% of global mortality is caused by chronic illness , suggesting that hiv / aids will substantially influence the economic and social development of developing countries in the future and that we can no longer allow risky health practices to continue unabated . in 2006 , the reasons persons have given for non - hiv testing were ( 1 ) they do not want to hear their status ( 11% ) , ( 2 ) 1.3% perceived seronegative status , ( 3 ) 1.0% stated that they were not sexually active and ( 4 ) had no partner , 0.7% said that they were not interested in doing an hiv test . based on the findings of the current study the prevalence of jamaicans not wanting to know their hiv status has increased exponentially , which speaks to the rise of inconsistent condom usage , increase in stis , increase in premarital sexual relations and promiscuity . those are the rationales which should vehemently dictate the urgency of hiv / aids registry in seeking to thoroughly understand this threat and how to effectively tailor intervention that the spread of stis and hiv / aids can be reduced . the time has come for the government to require hiv and aids to be reportable by name , and a national hiv / aids registry be kept in order to monitor the epidemic in jamaica . while the researchers recognize the stigmatization surrounding the disease , the issue of privacy , discrimination , and understand that this may result in the continuation of the low willingness of people to become tested for hiv ; the findings are far reaching and can not be allowed to continue into the indefinite future . | background : hiv / aids is a problem in developing countries including jamaica
. there are several studies dealing with hiv / aids in jamaica but given the increasing rate of the infection , ongoing studies are necessary.aims:this study examines the sexual behavior and attitude of non - hiv testers in jamaica in order to provide research evidence that will direct public health policies and interventions.materials and methods : this study extracts a sample of 1,192 participants who indicated not having done a hiv test from 1,800 respondents from a 2004 hiv / aids / std national kabp survey . a detailed questionnaire was developed and used to collect data from people ages 15 - 49 years old.results:some 20.3 % of the variances , which is self reported positive hiv test results , are a function of relationship status , using protection against hiv , having sex with a commercial sex worker , having stis , the age at which the participant first had sex and age at last birthday .
these findings hold across gender , occupational status and education . the majority ( 87.9% ) of the participants
said they had little or no chance of getting hiv , and 59.7% did not want to know their hiv status .
however , 46.6% did not wear a condom the first time they had sex with current partner , 32% do not wear a condom with their current partner and 16.3% reported having had stis.conclusion:the existing hiv prevention programs need to be expanded and fortified to target young jamaicans , particularly those who do not engage in safe sexual practices . |
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