obstructive sleep apnea ( osa ) is a chronic disorder characterized by repetitive apneas , oxygen desaturation , and disruption during sleep [ 13 ] . osa affects 3%24% of the general population and an even higher percentage ( 3545% ) of individuals who are obese or are suffering from diabetes mellitus ( dm ) [ 47 ] . recent studies have found that osa increases the risk of cardiovascular disease ( cvd ) independent of age , body mass index ( bmi ) , systolic blood pressure ( sbp ) , diastolic blood pressure ( dbp ) , or smoking habit [ 810 ] . importantly , atherosclerosis is considered to be the main cause of cardiovascular complications in osa patients . endothelial dysfunction , as an early marker of atherosclerosis , correlates significantly with osa . in general , the multiple methods measuring markers of endothelial function were invasive procedures . recently , flow - mediated dilatation ( fmd ) , a non - invasive method that evaluates nitric oxide ( no)-dependent vasodilatation , has been used to detect atherosclerosis in its subclinical phase . as a safe and convenient procedure , therefore , we performed this meta - analysis to assess whether atherosclerosis could be detected based on brachial artery fmd in patients with osa . the meta - analysis was performed in accordance with the recommendations of the cochrane collaboration and the quality of reporting of meta - analyses ( quorum ) guidelines . studies reported in english concerning osa and endothelial function were identified by searching electronic databases , including pubmed , embase , the cochrane library , and google scholar . the databases were searched from the earliest available dates until may 16 , 2016 . only those papers published with full - length text were considered . unpublished data from scientific meetings were also searched but not included since their abstracts did not provide detailed data . the search terms used were  obstructive sleep apnea  or  sleep apnea  or  osa  or  sleep - breathing disorders  and  flow - mediated  or references for all relevant articles , review articles , and relevant non - electronic literature were searched manually to identify additional relevant studies . manuscripts were scored in duplicate , and if their scores differed , a third author ( dr . fmd was selected as a marker of endothelial function based on a review of the literature . studies were included if they met the following criteria : ( 1 ) they were published in english and performed on adult humans ; ( 2 ) full - text manuscripts were available ; ( 3 ) they included at least two separate groups , one diagnosed with osa and the other made up of control subjects without osa ; ( 4 ) osa was diagnosed by polysomnography ( psg ) ; ( 5 ) initial fmd values recorded by ultrasound were available ; and ( 6 ) the reported values were presented as means and standard deviation or standard error or interquartile range . studies were excluded from the analysis for the following reasons : ( 1 ) fmd was not used to measure endothelial function ; ( 2 ) the results of comparison were not reported , or the data could not be extracted from the published results ; ( 3 ) they were non - human studies , letters , reviews , or case reports ; ( 4 ) the patients had other medical conditions that may have interfered with sleep , such as chronic respiratory disorders , heart failure , or uncontrolled allergies , or they were being treated with continuous positive airway pressure ( cpap ) ; ( 5 ) the outcomes of the same patient group were reported in another publication ( in that case , the higher - quality article was included ) ; ( 6 ) the trials were not published in english . the definition of osa varied in the different publications , because apnea - hypopnea index ( ahi ) cutoffs in epidemiological investigations conducted to date have been variable . thus , our meta - analysis also accepted osa as defined by the authors and not by the ahi criteria . wang and xu ) independently assessed the content of all studies to be included , and selected those that met the inclusion criteria while annotating the reasons for study exclusion . included studies were carefully scanned and the following information was extracted : first author , year of publication , number of participants , subject demographics ( age , sex , and bmi ) , ahi , fmd values , and confounding factors ( sbp , dbp , high - density lipoprotein [ hdl ] cholesterol , low - density lipoprotein [ ldl ] cholesterol , and glucose ) . for studies in which osa groups were divided based on severity , all sets of data were combined into one single group . for example , chami and colleagues   divided osa patients into mild , moderate , and severe groups , and the control group into two groups ( ahi 1.54.9 , ahi < 1.5 ) . therefore , we combined the three osa groups as well as the two control groups into one osa and one control group according to the methodology of the cochrane handbook ( 9.2.4 effect measures for ordinal outcomes and measurement scales ) . if the study enrolled participants with other diseases , then the methodology was processed in the same way . continuous values such as fmd were analyzed using the weighted mean difference ( wmd ) and were reported with 95% confidence intervals ( cis ) . the wmd summarizes the differences between two groups with respect to continuous variables , while accounting for sample size . for studies that presented continuous data as means and ses , the sds were calculated using the formula sd = sen . for studies that presented continuous data as means and quartiles , the sds were calculated using statistical algorithms according to the cochrane handbook : 
 sd = v73-v231.35 . an i of 2549% was considered to represent a low level of heterogeneity , 5074% a moderate level , and 75100% a high level . a fixed - effects or random - effects model was used according to the heterogeneity between studies . the covariates in the regression analyses included age , sex , bmi , blood pressure , glucose , and hdl and ldl cholesterol . all meta - analyses were conducted using the cochrane collaboration s review manager software version 5.0 and stata version 12.0 ( statacorp . , college station , texas , usa ) software packages . studies reported in english concerning osa and endothelial function were identified by searching electronic databases , including pubmed , embase , the cochrane library , and google scholar . the databases were searched from the earliest available dates until may 16 , 2016 . only those papers published with full - length text were considered . unpublished data from scientific meetings were also searched but not included since their abstracts did not provide detailed data . sleep apnea  or  osa  or  sleep - breathing disorders  and  flow - mediated  or references for all relevant articles , review articles , and relevant non - electronic literature were searched manually to identify additional relevant studies . manuscripts were scored in duplicate , and if their scores differed , a third author ( dr . fmd was selected as a marker of endothelial function based on a review of the literature . studies were included if they met the following criteria : ( 1 ) they were published in english and performed on adult humans ; ( 2 ) full - text manuscripts were available ; ( 3 ) they included at least two separate groups , one diagnosed with osa and the other made up of control subjects without osa ; ( 4 ) osa was diagnosed by polysomnography ( psg ) ; ( 5 ) initial fmd values recorded by ultrasound were available ; and ( 6 ) the reported values were presented as means and standard deviation or standard error or interquartile range . studies were excluded from the analysis for the following reasons : ( 1 ) fmd was not used to measure endothelial function ; ( 2 ) the results of comparison were not reported , or the data could not be extracted from the published results ; ( 3 ) they were non - human studies , letters , reviews , or case reports ; ( 4 ) the patients had other medical conditions that may have interfered with sleep , such as chronic respiratory disorders , heart failure , or uncontrolled allergies , or they were being treated with continuous positive airway pressure ( cpap ) ; ( 5 ) the outcomes of the same patient group were reported in another publication ( in that case , the higher - quality article was included ) ; ( 6 ) the trials were not published in english . the definition of osa varied in the different publications , because apnea - hypopnea index ( ahi ) cutoffs in epidemiological investigations conducted to date have been variable . thus , our meta - analysis also accepted osa as defined by the authors and not by the ahi criteria . two reviewers ( dr . wang and xu ) independently assessed the content of all studies to be included , and selected those that met the inclusion criteria while annotating the reasons for study exclusion . included studies were carefully scanned and the following information was extracted : first author , year of publication , number of participants , subject demographics ( age , sex , and bmi ) , ahi , fmd values , and confounding factors ( sbp , dbp , high - density lipoprotein [ hdl ] cholesterol , low - density lipoprotein [ ldl ] cholesterol , and glucose ) . for studies in which osa groups were divided based on severity , all sets of data were combined into one single group . for example , chami and colleagues   divided osa patients into mild , moderate , and severe groups , and the control group into two groups ( ahi 1.54.9 , ahi < 1.5 ) . therefore , we combined the three osa groups as well as the two control groups into one osa and one control group according to the methodology of the cochrane handbook ( 9.2.4 effect measures for ordinal outcomes and measurement scales ) . if the study enrolled participants with other diseases , then the methodology was processed in the same way . continuous values such as fmd were analyzed using the weighted mean difference ( wmd ) and were reported with 95% confidence intervals ( cis ) . the wmd summarizes the differences between two groups with respect to continuous variables , while accounting for sample size . for studies that presented continuous data as means and ses , the sds were calculated using the formula sd = sen . for studies that presented continuous data as means and quartiles , the sds were calculated using statistical algorithms according to the cochrane handbook : 
 sd = v73-v231.35 . an i of 2549% was considered to represent a low level of heterogeneity , 5074% a moderate level , and 75100% a high level . a fixed - effects or random - effects model was used according to the heterogeneity between studies . the covariates in the regression analyses included age , sex , bmi , blood pressure , glucose , and hdl and ldl cholesterol . all meta - analyses were conducted using the cochrane collaboration s review manager software version 5.0 and stata version 12.0 ( statacorp . , college station , texas , usa ) software packages . we identified 713 titles of potentially relevant articles from the literature search . from these , 622 articles were excluded based on preliminary title and abstract screening ( irrelevant=361 , case report or review=191 , non - adult population=44 , non - english=26 ) . the remaining 91 studies were scanned for full - text evaluation , and a further 63 articles were excluded for the following reasons : 31 articles did not use fmd to measure endothelial function , 11 did not contain original data , 16 did not have a control group , 3 included osa patients under cpap treatment , and 2 used medians with a range ( minimum , maximum ) to measure fmd ( figure 1 ) . thus , 28 articles covering 1496 osa patients and 1135 controls were finally included in the meta - analysis [ 11,12,1439 ] . a total of 28 articles providing 28 data sets were pooled for this meta - analysis . in these case - control studies , 11 were from asian researchers , 10 from american researchers , and 7 recruited patients from european countries . a total of 24 were clinically based , recruiting patients from hospitals or research centers , while 4 were population - based . most studies used in - hospital complete psg . there were no significant differences between the methods to measure fmd . the outcomes of the studies are discussed in table 1 . the pooled data from the eligible studies suggested that fmd was significantly reduced in osa patients . for fmd , the wmd was 3.07 and the 95% ci was 3.71 to 2.43 ( p<0.00001 ) . we re - performed another meta - analysis of studies using only full psg to exclude the impact of portable monitoring devices . the relationship between osa and fmd remained the same ( wmd : 3.30 , 95% ci : 3.83 to 2.78 , p<0.00001 ) . significant heterogeneity was observed between the studies in both analyses ( i=90% , i=78% ) . thus , a random - effects model was applied ( figures 2 , 3 ) . in addition , when studies with an inadequate number of osa subjects ( < 20 in each group ) [ 11,12,16,19,2530,3234,39 ] were excluded , according to the guidelines for the measurement of fmd , the pooled data also provided a robust result ( wmd : 2.42 , 95% ci : 3.32 . to 1.52 , p<0.001 ) ( supplementary figure 1 ) . a power calculation showed that seven studies [ 14,16,21,23 , 24,38,39 ] lacked sufficient power ( < 80% ) ( table 2 ) . however , their exclusion did not change the pooled result ( wmd : 3.50 , 95% ci : 4.04 . to 2.96 , p<0.001 ) ( supplementary figure 2 ) . multiple meta - regression analyses were performed to evaluate the effect of the covariant variables on fmd when reported . age ( slope=0.149 , p=0.136 ) , sex ( slope=6.970 , p=0.095 ) , bmi ( slope=0.014 , p=0.817 ) , sbp ( slope=0.193 , p=0.147 ) , dbp ( slope=0.204 , p=0.080 ) , glucose ( slope=0.058 , p=0.486 ) , triglycerides ( slope=0.017 , p=0.592 ) , total cholesterol ( slope=0.028 , p=0.607 ) , hdl cholesterol ( slope=0.091 , p=0.280 ) and ldl cholesterol ( slope=0.167 , p=0.066 ) were shown to not have a significant effect as confounding factors ( table 3 ) . to study the effect of complications , such as hypertension , diabetes , dyslipidemia , and treatment , we conducted a subgroup analysis according to whether the recruited subjects were free of the above disorders . the result showed that there was a significant association between osa and decreased fmd in subjects with ( wmd : 2.83 , 95% ci : 3.67 to 1.98 , p<0.001 ) versus those without ( wmd : 3.51 , 95% ci : 4.41 to 2.60 , p<0.001 ) these disorders ( supplementary figure 3 ) . thus , 11 studies were from asia , 10 were from the usa , and 7 were from europe . in all of the subgroups , decreased fmd was related to osa : the wmd ( 95% ci ) values from each subgroup were 2.68 ( 2.96 to 2.40 ) , 1.46 ( 1.76 to 1.16 ) , and 2.54 ( 3.00 to 2.09 ) , respectively . the i values from the three subgroups were 88% , 92% , and 75% , respectively ( asia , america , and europe ) . in the european subgroup , the heterogeneity was no longer apparent ( i=0% ) ( wmd : 3.03 , 95% ci : 3.55 . to 2.51 ) after the two studies that recruited subjects from italy were excluded ( figures 4 , 5 ) . test ( p=0.249 ) and funnel plot ( figure 6 ) showed no evidence of publication bias . we identified 713 titles of potentially relevant articles from the literature search . from these , 622 articles were excluded based on preliminary title and abstract screening ( irrelevant=361 , case report or review=191 , non - adult population=44 , non - english=26 ) . the remaining 91 studies were scanned for full - text evaluation , and a further 63 articles were excluded for the following reasons : 31 articles did not use fmd to measure endothelial function , 11 did not contain original data , 16 did not have a control group , 3 included osa patients under cpap treatment , and 2 used medians with a range ( minimum , maximum ) to measure fmd ( figure 1 ) . thus , 28 articles covering 1496 osa patients and 1135 controls were finally included in the meta - analysis [ 11,12,1439 ] . a total of 28 articles providing 28 data sets were pooled for this meta - analysis . in these case - control studies , 11 were from asian researchers , 10 from american researchers , and 7 recruited patients from european countries . a total of 24 were clinically based , recruiting patients from hospitals or research centers , while 4 were population - based . most studies used in - hospital complete psg . there were no significant differences between the methods to measure fmd . the pooled data from the eligible studies suggested that fmd was significantly reduced in osa patients . for fmd , the wmd was 3.07 and the 95% ci was 3.71 to 2.43 ( p<0.00001 ) . we re - performed another meta - analysis of studies using only full psg to exclude the impact of portable monitoring devices . the relationship between osa and fmd remained the same ( wmd : 3.30 , 95% ci : 3.83 to 2.78 , p<0.00001 ) . significant heterogeneity was observed between the studies in both analyses ( i=90% , i=78% ) . thus , a random - effects model was applied ( figures 2 , 3 ) . in addition , when studies with an inadequate number of osa subjects ( < 20 in each group ) [ 11,12,16,19,2530,3234,39 ] were excluded , according to the guidelines for the measurement of fmd , the pooled data also provided a robust result ( wmd : 2.42 , 95% ci : 3.32 . to 1.52 , p<0.001 ) ( supplementary figure 1 ) . a power calculation showed that seven studies [ 14,16,21,23 , 24,38,39 ] lacked sufficient power ( < 80% ) ( table 2 ) . however , their exclusion did not change the pooled result ( wmd : 3.50 , 95% ci : 4.04 . to 2.96 , p<0.001 ) ( supplementary figure 2 ) . multiple meta - regression analyses were performed to evaluate the effect of the covariant variables on fmd when reported . age ( slope=0.149 , p=0.136 ) , sex ( slope=6.970 , p=0.095 ) , bmi ( slope=0.014 , p=0.817 ) , sbp ( slope=0.193 , p=0.147 ) , dbp ( slope=0.204 , p=0.080 ) , glucose ( slope=0.058 , p=0.486 ) , triglycerides ( slope=0.017 , p=0.592 ) , total cholesterol ( slope=0.028 , p=0.607 ) , hdl cholesterol ( slope=0.091 , p=0.280 ) and ldl cholesterol ( slope=0.167 , p=0.066 ) were shown to not have a significant effect as confounding factors ( table 3 ) . to study the effect of complications , such as hypertension , diabetes , dyslipidemia , and treatment , we conducted a subgroup analysis according to whether the recruited subjects were free of the above disorders . the result showed that there was a significant association between osa and decreased fmd in subjects with ( wmd : 2.83 , 95% ci : 3.67 to 1.98 , p<0.001 ) versus those without ( wmd : 3.51 , 95% ci : 4.41 to 2.60 , p<0.001 ) these disorders ( supplementary figure 3 ) . thus , 11 studies were from asia , 10 were from the usa , and 7 were from europe . in all of the subgroups , decreased fmd was related to osa : the wmd ( 95% ci ) values from each subgroup were 2.68 ( 2.96 to 2.40 ) , 1.46 ( 1.76 to 1.16 ) , and 2.54 ( 3.00 to 2.09 ) , respectively . the i values from the three subgroups were 88% , 92% , and 75% , respectively ( asia , america , and europe ) . in the european subgroup , the heterogeneity was no longer apparent ( i=0% ) ( wmd : 3.03 , 95% ci : 3.55 . to 2.51 ) after the two studies that recruited subjects from italy were excluded ( figures 4 , 5 ) . test ( p=0.249 ) and funnel plot ( figure 6 ) showed no evidence of publication bias . to the best of our knowledge , this is the first meta - analysis to pool the available data and provide a summary of the relationship between decreased fmd and osa patients . twenty - eight studies , pooling 1496 osa patients and 1135 controls , were included . though many previous studies have assessed the relationship between osa and endothelial dysfunction , the results are conflicting . the majority of studies reviewed for this analysis reported a decrease in fmd in osa patients compared with controls . after summarizing all of the data , our results showed a statistically significantly lower fmd in osa patients than in controls . in addition , the results of several studies support a correlation between osa and decreased fmd , while also demonstrating that fmd values become even smaller as the severity of osa increases . in those studies , the subjects were divided into control , mild , moderate , and severe osa groups [ 2124,35 ] , and the results showed that moderate - severe osa patients suffered more from decreased fmd compared with mild osa patients . however , it is regrettable that the validity of this relationship remains to be established due to the small number of relevant studies ; thus , large - scale studies are required to obtain high - level evidence . the mechanism underlying osa impairment of endothelial function is unclear but is likely to involve several pathways . the three acute consequences of osa , intermittent hypoxia , intra - pleural pressure swings , and recurrent arousals , are thought to be the main causes of impaired endothelial function [ 4143 ] . of these , intermittent hypoxia is considered the most important factor promoting the production of reactive oxygen species ( ros ) , thereby increasing oxidative stress and decreasing nitric oxide ( no ) synthetase activity . some studies   have reported that age , bmi , and sbp were correlated with fmd ; in contrast , others   found no significant relationship between fmd and age , bmi , sbp , dbp , lipids , or fasting glucose levels . females were also more vulnerable than males to early osa - related cardiovascular diseases . after adjustment for variables significantly associated with fmd , namtvedt et al .   and jelic et al .   reported an independent association between increasing ahi and a reduction in fmd . according to chami et al . , however , there was no apparent association between osa and fmd after adjustment for age , sex , race , and all covariates . since the data in the majority of included studies were not adjusted for covariates , significant findings about the impact of confounding factors heterogeneity was observed in our meta - analysis ; meta - regression and subgroup analysis were conducted to determine the potential sources of the heterogeneity . meta - regression analysis excluded age , sex , bmi , blood pressure , glucose , hdl cholesterol , and ldl cholesterol as sources of heterogeneity . the european subgroup exhibited an i of 75% ; however , the exclusion of two studies that recruited subjects from italy lessened the heterogeneity ( i=0% ) . we assumed that this was due to ethnic diversity among the subgroups , leading to different physiological responses . the majority of studies included were clinically rather than community based , which could have introduced referral bias , resulting in heterogeneity . also , fmd might be affected by environmental effects such as noise , temperature , alcohol , caffeine , or fasting . although these factors were well controlled in the majority of studies according to the methods described by celermajer et al . first , the included studies were limited to publications in english , which may have increased the possibility of publication bias . in addition , it is known that positive results are more likely to be published ; since we included data only from published studies , publication bias was likely . third , this meta - analysis was not an overview of all methods of evaluating endothelial dysfunction . other indicators of endothelial function , such as no levels , endothelin-1 ( et-1 ) levels , measurements of circulating endothelial cells ( cecs ) , and peripheral artery tonometry ( pat ) , were not searched for and evaluated . also , most included articles did not clearly describe their osa patients , such as with respect to compliance and complications , both of which may affect the results . nonetheless , the subgroup analysis showed that decreased fmd was related to osa in all subgroups . recently , several articles   declared that fmd is concomitantly dependent on initial artery diameter , which may itself be higher in osa patients . other limitations pertaining to the methods of the individual studies included in this meta - analysis should also be addressed . finally , some of the included studies lacked enough power to detect an association or were not based on the use of standard psg to diagnose osa , but our results were also robust when these inadequate studies were excluded . future larger randomized studies of longer duration should focus on the effect of treatment of osa on endothelial dysfunction . forest plot excluding studies with an inadequate number of osa subjects ( < 20 in each group ) . meta - analysis of the relationship between osa and fmd according to whether other disorders were excluded or not .
backgroundendothelial dysfunction , which can be measured by flow - mediated dilatation ( fmd ) , is an early clinical marker of atherosclerosis , which is considered to be the main cause of the observed cardiovascular complications in obstructive sleep apnea ( osa ) patients . 
 the association between osa and endothelial dysfunction has been reported in a number of studies ; however , the findings are not entirely consistent . 
 our aim was to meta - analytically synthesize the existing evidence to explore the association between osa and endothelial dysfunction.material/methodsdata from pubmed , embase , the cochrane library , and google scholar for all trials that investigated the relationship between endothelial dysfunction and osa were systematically reviewed . 
 the minimum inclusion criteria for the studies were reporting of the apnea - hypopnea index ( ahi ) and fmd measurements ( as an indicator of endothelial dysfunction ) for both osa and control groups . 
 data from case - control studies that met the inclusion criteria were extracted.resultstwenty-eight studies comprising a total of 1496 osa patients and 1135 controls were included in the meta - analysis . a random - effects model was used . 
 the weighted mean difference in the fmd measurements was 3.07 and the 95% confidence interval was 3.71 to 2.43 ( p<0.01 ) . 
 meta - regression analysis showed that age , sex , body mass index ( bmi ) , blood pressure , glucose , high - density lipoprotein ( hdl ) cholesterol , and low - density lipoprotein ( ldl ) cholesterol did not explain the heterogeneity.conclusionsthis meta - analysis showed that patients with osa have decreased fmd , which may contribute to the development of atherosclerosis .