Recently, it has been reported that the A930G and C242T polymorphisms within p22phox (CYBA) gene are involved in the pathogenesis of hypertension. However, the results remain controversial. Furthermore, no previous meta-analysis has been conducted to evaluate the relationship between the A930G and C242T polymorphisms and hypertension. Therefore, we performed this meta-analysis to clarify these controversies.2 statistic. A cumulative meta-analysis was conducted to estimate the tendency of pooled OR. Funnel plots and Egger’s tests were performed to test for possible publication bias.All of the included articles were retrieved from the PubMed and Embase databases, as well as the CNKI, CBM, Chongqing VIP and Wan Fang databases according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Odds ratios (OR) with corresponding 95% confidence intervals (CI) were used to assess the strength of the association. Accounting for heterogeneity, a fixed or random effects model was respectively adopted. Heterogeneity was checked using the Q test and the IFive articles on A930G with 2003 cases/2434 controls and eight articles on C242T with 2644 cases/1967 controls were identiﬁed. A signiﬁcant association of A930G polymorphisms with the risk of hypertension was found in the dominant model  and allelic model . In the stratified analysis, a signiﬁcant association could be found among the hospital-based and population-based studies. However, no evidence of a significant association of the C242T polymorphism with hypertension was found in the overall analysis and subgroup analysis.This meta-analysis indicates that the A930G polymorphism, but not the C242T variation, might be a protective factor for hypertension. Hypertension is now considered a major public health issue  that affCYBA C852G and C536T polymorphisms have not been reported in association with hypertension [The p22phox (CYBA) gene is located on the long arm of chromosome 16q24 -16. It ertension . CYBA A930G and C242T polymorphisms with hypertension. One study conducted by Sales et al. [Recently, more studies have focused on the association of the s et al.  indicates et al. , Moreno s et al.  and Ha es et al.  support s et al. . Subsequs et al. -40 have In accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines , we searCYBA A930G or C242T polymorphisms and hypertension; b) case-control or cohort studies; c) sufficient data provided with information on the genotypes and allele frequencies. The exclusion criteria were: a) pedigree and family-based studies, b) duplicate publications. All the included original studies were selected according to the following inclusion criteria: a) studies evaluating the association between The data were carefully extracted from all the included publications independently by two investigators (Liang and Peng) according to the selection criteria listed above. If there was any disagreement, it was discussed among the authors or by consulting another reviewer (Su) to reach a consensus. Data extracted from the studies included the name of the ﬁrst author, publication year, country, ethnicity, study design, source of controls, number of cases and controls, diagnostic criteria and selection criteria of cases and controls, genotyping methods, genotype or allele distribution, and the matching method. Incomplete data regarding the genotypes and allele frequencies were calculated using the available information.AA/CC versus CT +TT), codominant model 1 (GG versus GA/CC versus CT), codominant model 2 (GA versus AA/CT versus TT), and the allelic model . The strength of the association between the CYBA A930G or C242T polymorphism and hypertension was estimated by calculating ORs with corresponding 95% CIs. Heterogeneity was analyzed using the Q test and the I2 statistic. If significant heterogeneity existed , the random effects model was used to calculate the pooled OR [CYBA C242T polymorphism and hypertension to assess the trends in the pooled OR over time under the dominant contrast in the random effect model. Studies included in the cumulative meta-analysis were sorted by the year of publication.All statistical analyses were implemented in Stata statistical software version 11.1. The Hardy-Weinberg equilibrium (HWE) for the genotype distributions in controls was assessed by the chi-square test (p>0.05). We used the following genetic models to pool the data: dominant model (GG + GA versus AA/CC + CT versus TT), recessive model , Embase (n=67), CNKI (n=72), CBM (n=7), Chinese Wan Fang (n=62), and Chongqing VIP (n=22). After reviewing these titles and abstracts, we obtained 47 potential eligible studies. By scanning the full texts, 34 articles were excluded according to the selection criteria. Finally, thirteen qualiﬁed articles (five articles for A930G and eight articles for C242T) were included in the meta-analysis .Five included studies investigating A930G polymorphisms in hypertension were examined, including 2003 cases and 2434 controls. All of the included studies were published between 2003 and 2007. The ethnicities of the research populations were as follows: three studies involved Asians and two involved in Caucasians. There were two population-based studies ,29 and t As shown in Significant associations between the A930G polymorphism and the risk of hypertension were identified in the dominant model  and the allelic model  . HoweverIn the further subgroup analysis according to the source of controls, a statistically significant association of the A930G polymorphism with hypertension was found in the dominant model in the hospital-based studies  and the population-based studies . Similar results were found in the allelic model among the hospital-based studies  and the population-based studies  . StatistThere was no statistically significant association between the C242T polymorphism and hypertension in any of the genetic models: dominant model , recessive model , co-dominant model 1 , co-dominant model 2 , and allelic model  . SimilarSensitivity analysis was performed to strengthen the confidence in the results by limiting the included studies according to HWE. After two studies ,37 withoCumulative analysis of the C242T polymorphism with hypertension was performed according to the publication date. A signiﬁcant association was conﬁrmed in the dominant contrast among the studies from 2003 to 2005. However, subsequent publications from 2006 to 2013 failed to repeat the initial results: Moreno et al. , Wang et al. , Kuznetsova et al. , Schreiber et al. , Petrovicet al.  .Funnel plots, Begg’s test and Egger's test were performed to assess the publication bias of the studies. The shapes of the funnel plots were visually symmetrical. No publication bias was found regarding the A930G and C242T polymorphism and its association with hypertension by using Egger’s test under the allelic model , dominant model , recessive model , codominant model 1 , or codominant model 2 .In this meta-analysis, thirteen qualiﬁed articles (five articles with 2003 cases/2434 controls for A930G and eight articles with 2644 cases/1967 controls for C242T) were included. To the best of our knowledge, this is the ﬁrst meta-analysis to assess the relationship of the A930G and C242T polymorphisms with hypertension. Our results suggest that the A930G polymorphism is associated with hypertension under both the dominant and allelic model, which was in accordance with the results of the studies by Kokubo et al. , Moreno In the stratified analysis by ethnicity, no significant association of the A930G polymorphism with hypertension was detected in any of the genetic models. Similarly, no significant association between the C242T polymorphism and hypertension could be found in any of the genetic models in the further subgroup analysis by ethnicity. According to the data from the International HapMap project, the frequency of the minor G allele in the A930G polymorphism (rs9932581) among the CEU population (northern and western European ancestry) is 0.350; the minor allele frequency (MAF) of the A930G polymorphism is similar among Japanese in Tokyo (JPT) (MAF=0.477), whereas it is A with a frequency of 0.360 among Han Chinese in Beijing (CHB). The MAF of the A930G polymorphism was found to be different between Caucasian populations and Asian Chinese populations according to the HapMap data. However, a sample size of 1515 cases and 2125 controls, which was used to evaluate the genetic susceptibility of hypertension in the Japanese population was larger than that in the Chinese population (206 cases/171 controls). The MAF of the A930G polymorphism is similar in Caucasian and Asian Japanese populations, which is in accordance to our findings that the effect of the A930G polymorphism on hypertension is similar across different ethnic populations. Furthermore, the C242T MAF varies between European and Asian populations, as the frequency of the T allele is higher in Europeans than in Asians . Our result is not consistent with the data from the International HapMap project, probably because of the limited eligible studies and sample sizes. Thus, large-scale studies are required to further evaluate our ﬁndings.A significant association between the A930G polymorphism and hypertension was found for the dominant and allelic models in both the hospital-based studies and the population-based studies. However, a signiﬁcant association was found only in the hospital-based studies for the recessive model and co-dominant model 1, but not in the population-based studies, which might be attributed to differences in the sources of controls. The controls in hospital-based studies might suffer from other diseases that could possibly involve in the same genetic pathogenesis of hypertension. They may not be representative of the general population and result in a false positive. Thus, selective bias, which is likely to affect the quality and reliability of our findings, could not be ignored in the hospital-based studies. Therefore, this positive result should be interpreted with caution. As to the subgroup analysis of studies based on population, only two articles were included in the analysis; therefore, studies with larger sample sizes are required to further confirm our findings. Moreover, no significant association between the C242T polymorphism and hypertension could be found in any of the genetic models in the subgroup analysis according to the source of controls.The biological mechanism of the A930G and C242T polymorphisms in the physiopathogenesis of hypertension is unclear. ROS have been suggested to play a major role in oxidative stress, and contribute significantly to the development of hypertension . Higher According to the publication date, a cumulative analysis of the C242T polymorphism in hypertension was performed. A signiﬁcant association between the C242T polymorphism and hypertension was conﬁrmed with an increasingly narrow 95% confidence interval (CI) from 2003 to 2005. Nevertheless, no trend for the association between the C242T polymorphism and hypertension was found in subsequent studies from 2006 to 2013, which is in accordance with the results of the current meta-analysis. However, only eight studies were included in the cumulative analysis. To be sure of these results, further studies re required to conﬁrm our ﬁndings.An exploration in the source of heterogeneity was conducted by subgroup analysis. Strong heterogeneity was still observed in studies assessing the relationship between the A930G polymorphism and hypertension when stratified by ethnicity. Thus, Asian and Caucasian ethnicity was likely to be one of the causes of heterogeneity. In addition, the environment that people lived in and genetic variations should be considered as sources of heterogeneity. Similarly, signiﬁcant heterogeneity still persisted in the subgroup analysis according to the source of controls, which suggested that the source of controls, whether studies were hospital-based or population-based, also had an influence on heterogeneity. Additionally, no obvious changes in heterogeneity could be observed among the studies of Asian populations in the association between the C242T polymorphism and hypertension when stratified by ethnicity. Nevertheless, heterogeneity decreased in studies on Caucasian populations, which indicated that the source of heterogeneity might originate from the Asian ethnicity. Heterogeneity was still signiﬁcant in hospital-based studies, but obviously reduced in the population-based studies, which suggested that heterogeneity might be attributed to the hospital-based studies. There was considerable variation in the sample sizes of the studies included in this meta-analysis; of the qualiﬁed studies on the A930G polymorphism in hypertension, the largest sample size was 1515 cases/2125 controls and the smallest sample size was 83 cases/66 controls. Similarly, of the included studies used to evaluate the association between the C242T polymorphism and genetic susceptibility to hypertension, the largest sample size was 1030 cases/826 controls and the smallest sample size was 57 cases/106 controls. These different sample sizes might also contribute to the source of heterogeneity. Furthermore, other confounding factors, such as matching methods, study design, and individual biological characteristics were identified as a potential source of heterogeneity. Furthermore, there was no publication bias in the present meta-analysis, as a relatively comprehensive search strategy was conducted. Finally, our results should be considered with some limitations in the present meta-analysis. First, the sample size in this meta-analysis was not large, which limited the statistical power. As to the subgroup analysis by ethnicity, only two Caucasian studies and three Asian studies were included for the A930G polymorphism in hypertension. Similarly, in the subgroup analysis according to the source of controls, only two population-based studies were included for the A930G polymorphism in hypertension, in contrast to three hospital-based studies. In order to better decipher our results, more studies with larger sample sizes are needed in the future. Second, this meta-analysis only focused on the articles from English and Chinese databases without language restrictions, which might lead to a potential language bias. Third, information on confounding factors, such as age, sex, smoking, and drinking, could not be obtained from all the original articles; these are considered effective inﬂuencing factors in the pathogenesis of hypertension. In addition, we did not perform an evaluation of potential interactions such as gene-gene or gene-environment, which might be involved in susceptibility to hypertension.In summary, in the overall analysis, the present meta-analysis shows that the A930G polymorphism contributes to susceptibility to hypertension. Similarly, a statistically signiﬁcant association was found in the subgroup analysis according to ethnicity. However, there is a lack of evidence to support the association of C242T with hypertension. Further studies considering gene-gene and gene-environment interactions, larger sample sizes, and well-matched controls are needed to identify the association between the A930G and C242T polymorphisms and hypertension.Checklist S1PRISMA checklist.(DOC)Click here for additional data file.File S1PRISMA 2009 Flow Diagram.(DOC)Click here for additional data file.


Glutathione S transferase (GST) polymorphisms have been considered risk factors for the development of glaucoma, including primary open angle glaucoma (POAG) and other types of glaucoma. However, the results remain controversial. In this study, we have conducted a meta-analysis to assess the association between polymorphisms of GSTM1, GSTT1 and GSTP1 and glaucoma risk.GSTM1, GSTT1 and GSTP1 polymorphisms and glaucoma risk were included. Pooled odds ratio (OR) and 95% confidence interval (CI) were calculated using random- or fixed-effects model.Published literature from PubMed and other databases were retrieved. All studies evaluating the association between GSTM1 (1109 cases and 844 controls), ten studies on GSTT1 (709 cases and 664 controls) and four studies on GSTP1 (543 cases and 511 controls) were included. By pooling all the studies, either GSTM1 or GSTT1 null polymorphism was not associated with a POAG risk, and this negative association maintained in Caucasian. The GSTP1 Ile 105 Val polymorphism was significantly correlated with increased POAG risk among Caucasian in a recessive model . Interestingly, increased glaucoma risk was associated with the combined GSTM1 and GSTT1 null genotypes , and with the combined GSTM1 null and GSTP1 Val genotypes .Twelve studies on GST polymorphisms are associated with glaucoma risk. Given the limited sample size, the associations between single GST polymorphism and glaucoma risk await further investigation.This meta-analysis suggests that combinations of  MYOC, OPTN and WDR36) for juvenile-onset and late-onset primary open angle glaucoma (POAG) Glaucoma is a heterogeneous group of diseases characterized by the death of the retinal ganglion cells and progressive degeneration of the optic nerve. It is the second most frequent cause of irreversible blindness in the world and affects primarily the older population, estimated to affect about 80 million people worldwide by 2020 Growing evidence supports the involvement of oxidative stress as a common component of glaucomatous neurodegeneration in different subcellular compartments of retinal ganglion cells (RGCs), by acting as a second messenger and/or modulating protein function by redox modifications of downstream effectors through enzymatic oxidation of specific substrates GSTT1 and GSTM1 genes caused by a deletion in each gene, and a single-nucleotide polymorphism (SNP) of GSTP1 resulting in the coding sequence change Ile 105 Val GSTT1 or GSTM1, or GSTP1 Ile 105 Val polymorphism results in an absence of their enzyme activity GST have been associated with altered risk of a variety of pathologies including cancer GST polymorphisms and risk of glaucoma has been studied for more than 10 years. Several studies have found GST polymorphisms to be protective or risk factors in POAG GST polymorphisms and risk of glaucoma GSTM1, GSTT1, and GSTP1 polymorphisms and the risk for glaucoma.Previous studies of allelic variants in these classes have identified two major polymorphisms of the GST polymorphism with glaucoma, we conducted a literature search in the PubMed databases up to August 2012 using the following MeSH terms and keywords: “glutathione S transferase”, “polymorphism” and “glaucoma”. Additional studies were identified by a manual search from other sources , references of original studies or review articles on this topic. Eligible studies included in this meta-analysis had to meet the following criteria: (a) evaluation of the association between GSTM1 or GSTT1 null genotypes, or GSTP1 Ile 105 Val polymorphism and glaucoma, (b) an unrelated case-control study, if studies had partly overlapped subjects, only the one with a larger sample size was selected, (c) available genotype frequency, (d) sufficient published data for estimating an odds ratio (OR) with 95% confidence interval (CI), and (e) papers published in English from 2000.To identify all articles that examined the association of Two investigators independently assessed the articles for inclusion/exclusion and extracted data, and reached a consensus on all of the items. For each study, the following information was extracted: name of the first author; publication year; ethnicity (country); sample size (numbers of cases and controls); gene polymorphisms investigated; types of glaucoma; sources of samples; genotyping methods.GSTM1, GSTT1 or GSTP1 polymorphism and glaucoma was estimated by calculating pooled odd ratios (ORs) and 95% CIs. The significance of the pooled OR was determined by Z test . The risk of GSTM1 or GSTT1 null genotype on glaucoma was evaluated by comparing with their reference wild type homozygote. For the GSTP1 polymorphism, we first estimated the risks of the Ile/Val and Val/Val genotypes on glaucoma, compared with the reference Ile/Ile homozygote, and then evaluated the risks of  and  on glaucoma, assuming dominant and recessive effects of the variant Val/Val allele, respectively. The I2-based Q statistic test was performed to evaluate variations due to heterogeneity rather than chance. A random-effects (DerSimonian-Laird method) or fixed-effects (Mantel-Haenszel method) model was used to calculate pooled effect estimates in the presence (P≤0.10) or absence (P>0.10) of heterogeneity. Publication bias was detected by Egger’s test GSTM1 and GSTT1 null genotypes, and recessive model of GSTP1. Additionally, Begg’s funnel plot was drawn. Asymmetry of the funnel plot means a potential publication bias. Stratified analyses were also performed by types of glaucoma and ethnicities of study populations. For the one-way sensitivity analysis, one single study was excluded each time, and the new pooled results could reflect the influence of that deleted study to the overall summary OR. All analyses were done with Stata software , using two-sided P values.The association between in vitro study to evaluate the sensitivity to oxidative stress of anterior chamber tissues Thirteen abstracts were retrieved through the search “glutathione S transferase”, “polymorphism” and “glaucoma”, and ten studies meeting the inclusion criteria were identified as eligible GSTM1 genotype , ten studies for GSTT1 genotype  and four studies for GSTP1 polymorphism . For the ethnicities, eleven studies of Caucasians and one study of Asians were included on the GSTM1 genotype. As to GSTT1, night studies of Caucasians and one study of Asians were included. The four studies on GSTP1 were all based on the Caucasians. For the glaucoma type, this meta-analysis included ten, eight, and three studies on the relationship between GSTM1, GSTT1, and GSTP1 polymorphism and risk of POAG (the most common form of glaucoma), respectively. In addition, we included four and three studies on the association between the GSTM1 and GSTT1 polymorphism and risk of other types of glaucoma (including exfoliative glaucoma and primary closed angle glaucoma), respectively. In addition to the study by Juronen et al. GSTM1 and GSTT1 phenotypes were determined with monoclonal antibody based enzyme-linked immunosorbent assay (ELISA), the genotyping for GSTM1, GSTT1 or GSTP1 was performed using polymerase chain reaction (PCR) in all other studies. The detailed characteristics of each study included in the meta-analysis are presented in GST polymorphism genotype distributions from each study are presented in Twelve studies were included in the meta-analysis of GSTM1 or GSTT1 null polymorphism and risk of glaucoma. By pooling all the studies, either GSTM1 or GSTT1 null polymorphism was not associated with a glaucoma risk, and this negative association maintained in Caucasian  but not in other three models. Interestingly, these three studies were all based on Caucasian populations, thus, GSTP1 Ile 105 Val polymorphism in recessive model was associated with increased POAG risk in Caucasians . When stratified by the types of glaucoma, combination of GSTM1 and GSTT1 null genotypes was associated with increased risk of POAG  but not other types of glaucoma . We also examined if the risk of glaucoma was associated with combinations of GSTP1 and GSTM1, or GSTT1 genotypes, in which the homozygous Ile/Ile genotype for GSTP1 was used as reference and the individuals heterozygous and homozygous for the Ile 105 Val allele was combined GSTM1 null and GSTP1 Val genotypes , but the combined GSTT1 null and GSTP1 Val genotypes played a protective role in glaucoma risk which, which remained of borderline statistical significance .To investigate if the profiles of GSTM1 and GSTT1 null genotypes, and the recessive model of GSTP1 polymorphism. The Begg’s test showed that the P value for GSTM1, GSTT1 and GSTP1 polymorphism was 1.00, 0.371 and 1.00 respectively, and the corresponding funnel plots showed symmetric distribution . Thus, no evident publication bias was found in present study. Sensitivity analysis was conducted by deleting each study in turn from the pooled analysis to examine the influence of the removed data set to the overall ORs. As shown in GST polymorphism, suggesting that the results of synthetic analysis were robust.Publication bias was firstly detected by Begg’s test for the overall pooled analysis of ribution . The EggGST polymorphisms and susceptibility to glaucoma. Our meta-analysis showed that single GSTM1 or GSTT1 null polymorphism was not associated with glaucoma risk, and GSTP1 Ile 105 Val polymorphism in recessive model was positively correlated with increased glaucoma risk. The combination of GSTM1 null and GSTT1 null, or GSTM1 null and GSTP1 genotype was associated with increased risk of glaucoma. Although different types of glaucoma have their own clinical characteristics and pathogenesis, our meta-analysis indicates that GST polymorphisms may contribute to increased risk of glaucoma.In the present study, we systemically reviewed all available published studies and performed a meta-analysis to examine the association between the GSTM1 positive phenotype may be a genetic risk factor for development of POAG GSTM1 null genotype is a risk factor for development of POAG GSTM1 polymorphism and POAG, suggesting that previous controversial data may be due to small size of population. As to GSTP1, previous studies did not identify differences between POAG patients and control individuals in the frequencies of GSTP1 Ile 105 Val genotypes GSTP1 polymorphism was significantly correlated with increased POAG risk in Caucasian in a recessive model. It should be noted that the statistical significance of the association between the GSTP1 polymorphism and POAG risk was at borderline level. The GSTP1 105-Val allele homozygote was correlated with increased POAG risk but the difference did not reach statistical significance . Since the studies included were very limited, it is necessary to validate the association between GSTP1 Ile 105 Val polymorphism and glaucoma risk in future studies.Previously, the study by Juronen et al. suggests that the GST polymorphism, or combination of GST polymorphisms and glaucoma. Previously, Yildirin et al. reported an increasing glaucoma risk with higher numbers of the combined of GSTM1 null and GSTP1 105-Val allele genotypes but this association was not significant GSTM1 and GSTT1 null genotypes, or GSTM1 null and GSTP1 Val genotypes, and the risk for glaucoma is statistically significant in Caucasians. The study by Yildirin et al. also found a trend of increasing glaucoma risk with higher numbers of the combined GSTM1 null, GSTT1 null and GSTP1 105-Val allele genotypes  GSTM1 null, GSTT1 null and GSTP1 105-Val allele genotypes and glaucoma risk. The current available data support the multifactorial nature of glaucoma, and both genetic and environmental factors are involved in pathogenesis of glaucoma. However, most studies did not provide GST polymorphisms when stratified by environmental factors . The relationship between polymorphic GST with other genetic and environmental glaucoma risk factors may be highly complicated, and extensive research is required to ascertain how exactly the GST genotype affects the individual susceptibility to glaucoma.To the best of our knowledge, this is the first meta-analysis assessing the association between single GSTP1 polymorphism), which needs further investigations. Second, basic methodological differences among the studies might have affected the results. In addition to three studies GSTM1 and GSTT1 genotypes, the association between GSTP1 polymorphism and POAG risk in Caucasian was not significant due to the limited studies (n = 2) (Data not shown). Third, the studies differed in their procedure for sampling. Three studies used trabeculectomy specimens GST polymorphism and glaucoma risk in each subgroup analysis when the samples were stratified as trabeculectomy specimens or blood, and excluding one study using trabeculectomy specimens as samples did not affect the results on the associations between the combinations of GST polymorphisms and glaucoma risk (Data not shown). Fourth, most of the studies included in this meta-analysis did not categorize the POAG patients as high- and normal-tension glaucoma. So, we did not analyze the association between GST polymorphism and risk of high- or normal-tension glaucoma, and future studies should address this point. Last, the Caucasian group might have been genetically heterogeneous, with differences in terms of lifestyle and environment . These factors may explain the heterogeneity in meta-analysis for Caucasian populations.Meta-analysis has advantages compared to individual studies, however, some potential limitations in our study should be considered. First, this meta-analysis was limited by the small sample size, especially in subgroup analysis aforementioned Click here for additional data file.Figure S2Forest plots of the association between GSTT1 null polymorphism and glaucoma risk.(DOC)Click here for additional data file.Figure S3Forest plots of the association between GSTP1 Ile 105 Val polymorphism and glaucoma risk.(DOC)Click here for additional data file.Figure S4Sensitivity analysis for GSTM1 null polymorphism.(DOC)Click here for additional data file.Figure S5Sensitivity analysis for GSTT1 null polymorphism.(DOC)Click here for additional data file.Figure S6Sensitivity analysis for GSTP1 Ile 105 Val polymorphism.(DOC)Click here for additional data file.Table S1GSTM1 and GSTT1 polymorphism genotype distribution of each study included in the meta-analysis.(DOC)Click here for additional data file.Table S2GSTP1 Ile 105 Val polymorphism genotype distribution of each study included in the meta-analysis.(DOC)Click here for additional data file.


Numerous studies in Chinese populations have evaluated the association between the A-6G and A-20C polymorphisms in the promoter region of angiotensinogen gene and hypertension. However, the results remain conflicting. We carried out a meta-analysis for these associations.heterogeneity = 0.96). The A-20C polymorphism was significantly associated with the increased risk for hypertension in the allele comparison  and recessive genetic model . In the subgroup analysis by ethnicity, significant association was also found among Han Chinese for both A-6G and A-20C polymorphisms. A borderline significantly decreased risk of hypertension between A-6G and Chinese Mongolian was seen in the allele comparison .Case–control studies in Chinese and English publications were identified by searching the MEDLINE, EMBASE, CNKI, Wanfang, CBM, and VIP databases. The random-effects model was applied for dichotomous outcomes to combine the results of the individual studies. We finally selected 24 studies containing 5932 hypertensive patients and 5231 normotensive controls. Overall, we found significant association between the A-6G polymorphism and the decreased risk of hypertension in the dominant genetic model (AA+AG vs. GG: P = 0.001, OR = 0.71, 95%CI 0.57–0.87, POur meta-analysis indicated significant association between angiotensinogen promoter polymorphisms and hypertension in the Chinese populations, especially in Han Chinese. Essential hypertension (EH), the major section (over 95%) of hypertension, represents a serious health problem all over the world. In China, hypertension affects more than 18.8% of the adult population, and a total of 170 million people suffer from this disease The renin-angiotensin-aldosterone system (RAAS) plays a crucial role in the maintenance of BP In a 2008 meta-analysis by Pereira et al. To search for all the studies that examined the association of the A-6G and A-20C polymorphisms with hypertension in Chinese, we conducted a computerized literature search of the PubMed, EMBASE, CNKI (China Nation Knowledge Infrastructure Platform), Wanfang, CBM  and VIP databases (up to May 2011), using the following keywords and subject terms: ‘AGT or Angiotensinogen’, ‘polymorphism’, ‘hypertension’ and ‘Chinese or China or Taiwanese or Taiwan’. We only included studies published in Chinese or in English. References of retrieved articles were also screened. When a report overlapped with another publication, to prevent data duplication, only the more detailed one was kept. If an article reported results on different ethnic sub-populations, each sub-population was treated as separate study in our meta-analysis. We used the following inclusion criteria for a study to be included in the meta-analysis: (a) studies investigating the association of the A-6G or A-20C polymorphisms with hypertension in Chinese individuals, (b) use of an unrelated case–control design (family-based study design with linkage considerations was excluded), (c) available genotype frequency, (d) the genotype distribution of the control population must be in Hardy–Weinberg equilibrium (HWE) and (e) hypertension defined as systolic blood pressure (SBP)≥140 mmHg and/or diastolic blood pressure (DBP)≥90 mmHg and/or treatment with anti-hypertensive medication. If the genotype frequency was not reported, we contacted the original authors by e-mail in order to obtain the missing data.Two authors (W. Gu and J. Liu) independently reviewed and extracted the data needed. Disagreements were resolved through discussion among the authors to achieve a consensus. The following information was abstracted from each study: first author, year of publication, racial background and resident region of study population, diagnostic criteria, matching, source of samples, genotype detecting method of each study, number of cases and controls, distribution of genotypes and alleles in both case and control groups.2 was also calculated to evaluate the variation which was caused by heterogeneity rather than by chance, and higher values of the index indicate the existence of heterogeneity Odds ratios (OR) corresponding to 95% confidence interval (CI) was applied to measure the strength of the association of A-6G and A-20C with hypertension as case–control studies were used, and OR was calculated according to the method of Woolf When unexpected heterogeneity was present, sensitivity analysis was performed to examine specific sensitivity of the findings. This analysis was conducted by examining and recalculating the pooled association sizes and joint values of P in homogeneous subgroups, as well as after excluding studies one by one.http://ihg.gsf.de/cgi-bin/hw/hwa1.pl). All statistical analyses were performed using ReviewManager 5.0.25  and the software Stata version 10.0 . All P-values were two-sided.Publication bias was investigated by funnel plot, in which the standard error of the log (OR) of each study was plotted against its OR. An asymmetric plot suggested possible publication bias. Funnel-plot asymmetry was assessed by the method of Egger's linear regression test After literature search and selection applying our inclusion criteria, we identified a total of 27 relevant articles (40 studies) heterogeneity = 0.09, I2 = 35%. AA vs. AG+GG: P = 0.4, OR = 0.93, 95%CI 0.8–1.09, Pheterogeneity = 0.04, I2 = 43%)   . However I2 = 0) .heterogeneity = 0.64, I2 = 0) , two studies involved Chinese Mongolian (350 cases and 366 controls) and two studies involved Chinese Tibetan (259 cases and 325 controls). The -6A allele had a much higher representation in cases and controls of Han Chinese  than that of Tibetan  and of Mongolian . In Han Chinese population, significant association between the A-6G polymorphism and the decreased risk for hypertension was observed in the dominant genetic model . In the , I2 = 0). For the, I2 = 0). For Mon2 = 31%) . There w2 = 31%) .heterogeneity = 0.24, I2 = 29%)  .Considering the fact that the sex discrepancy might bias the overall association, a further subgroup analysis was conducted according to gender. As reflected in 2 = 29%) . HoweverI2 = 0%)  and receheterogeneity = 0.92, I2 = 0)     and rece I2 = 0) . There w I2 = 0) .heterogeneity = 0.67, I2 = 0) and recessive genetic model  , and only one study considered Kazakh, Li, Tibetan and Mongolian populations. In the Han population, we found significantly elevated risk of hypertension with the A-20C polymorphism in the allele comparison  . No posi I2 = 0) . Finallyheterogeneity = 0.15; Kong et al. heterogeneity = 0.16; Jiang et al. heterogeneity = 0.16; Wang (Amis) et al. heterogeneity = 0.37, AA vs. AG+GG, Pheterogeneity = 0.23). In addition, our analysis showed that the corresponding pooled ORs were materially altered with the sequential removal of these four studies (data not shown). The findings revealed that these independent studies might be the main cause of heterogeneity across all subjects.Significant between-study heterogeneity only existed among all studies in the meta-analysis of the A-6G polymorphism. Sensitivity analyses were conducted by sequentially removing a single study each time to find out the origin of heterogeneity. As a result, the heterogeneity no longer existed for the A-6G polymorphism when four studies were excluded (Liu (Yi) et al. The Egger's test and Begg's funnel plot were applied for allele comparison to asses the publication bias of the literatures. As indicated by the Egger's test and funnel plot, there was no publication bias for the A-6G polymorphism  , and a pThe literature examining the relationship between the A-6G and A-20C polymorphisms and hypertension in the Chinese populations abounds in small studies reporting controversial findings. No clear consensus has yet been reached. Therefore, we restricted our research to Chinese populations and did a meta-analysis with 24 studies totaling 5932 hypertensive patients and 5231 normotensive controls to form a more precise estimation of their association. To our knowledge, this was one of the largest meta-analysis to date investigating the association of angiotensinogen promoter polymorphisms with hypertension in Chinese. In the current study, we found that the A allele of the A-6G polymorphism was associated with a significant decrease in the risk of hypertension in all subject, and the C allele of A-20C could increase the risk of hypertension. These positive associations should be treated with caution as the P values obtained were reported without correction for multiple testing.In 1997, Inoue et al. Regarding the A-20C polymorphism, a report by Zhao et al. China is a huge multi-ethnic country with 56 identified ethnic groups. Among these groups, Han Chinese is the largest ethnic group, making up over 93% of the total population The positive findings about A-20C should be interpreted in light of the fact that publication bias was detected. The statistical test and funnel plot inspection in the meta- analysis have indicated the potential for such bias. Publication bias is a relatively common phenomenon in clinical literature Several limitations of our meta-analysis need to be noted. First, because of a small number of available studies, we failed to perform additional subgroup analysis in other minority populations (such as Bai), and by gender for the A-20C polymorphism. Second, publication bias was present, and might distort the final conclusion. Third, due to the lack of original data, an evaluation of potential interactions such as gene-gene or gene-environment was not considered in this meta-analysis, which might confound our results.In conclusion, our meta-analysis suggested significant association between the A-6G and A-20C polymorphisms and hypertension in the Chinese populations, particularly in Han Chinese. More large–scale studies, and especially studies stratified for different minorities and different sexes, should be performed to further elucidate the association between the A-6G and A-20C polymorphisms and hypertension in the Chinese populations.Table S1The PRISMA checklist for this meta-analysis.(DOC)Click here for additional data file.


Purpose: Folate is essential to DNA methylation and synthesis and may have a complex dualistic role in prostate cancer. Alcohol use may increase risk and epigenetic factors may interact with lifestyle exposures. We aimed to characterize the independent and joint effects of folate intake, alcohol consumption, and the MTHFR C677T gene polymorphism on prostate cancer risk, while accounting for intakes of vitamins B2, B6, B12, methionine, total energy, and confounders. Methods: A case-control study was conducted at Kingston General Hospital of 80 incident primary prostate cancer cases and 334 urology clinic controls, all with normal age-specific PSA levels (to exclude latent prostate cancers). Participants completed a questionnaire on folate and alcohol intakes and potential confounders prior to knowledge of diagnosis, eliminating recall bias, and blood was drawn for MTHFR genotyping. Joint effects of exposures were assessed using unconditional logistic regression and significance of multiplicative and additive interactions using general linear models. Results: Folate, vitamins B2, B6, B12, methionine, and the CT and TT genotypes were not associated with prostate cancer risk. The highest tertile of lifetime alcohol consumption was associated with increased risk . Consumption of >5 alcoholic drinks per week was associated with increased prostate cancer risk among men with low folate intake , and higher risk among those with the CC MTHFR genotype . Increased risk was also apparent for average weekly alcohol consumption when accounting for the multiplicative interaction between folate intake and MTHFR C677T genotype . Conclusion: Alcohol consumption is associated with increased prostate cancer risk, and this association is stronger among men with low folate intake, with the CC MTHFR genotype, and when accounting for the joint effect of folate intake and MTHFR C677T genotype. Prostate cancer is the second most common malignancy among men  gene. A meta-analysis in 2009 showed that consistently heavy alcohol drinking confers higher risk for prostate cancer  levels  and a normal digital rectal examination (DRE). Of 1288 men seen in clinic, 746 did not meet these criteria. Of the 542 eligible subjects, 71 (13%) were excluded due to diagnosis of prostate intraepithelial neoplasia, use of a hormonal medication, or missing questionnaire, and 56 declined to participate (10%). Participants were 414 men (76%) enrolled prior to knowledge of diagnosis who completed a questionnaire regarding demographic, lifestyle, medical, diet and occupational histories, and donated 15 mL of blood. After biopsy and pathology exams for 214 eligible biopsy subjects, 80 were diagnosed as incident primary prostate cancer cases, and the other 134 were included as “biopsy controls.” In addition, 200 eligible cancer-free men of the same age distribution who were diagnosed with a non-cancer urologic condition were enrolled as “urology controls,” for a total of 334 controls. Among these groups, 43 cases (54%), 102 biopsy controls (76%), and 68 urology controls (34%) consented to The dietary section of the questionnaire was adapted from Byers and colleagues' food frequency questionnaire , B6, and B12, fat, energy, and methionine levels were calculated due to their possible roles as confounding factors. Specifically, food nutrient levels were determined using the 1991 Canadian Nutrient file , the Canada Food Statistics 1996 , which cleaves the DNA when the variant allele is present. The wildtype CC fragment showed a 198 base pair (bp) fragment when analysed by gel electrophoresis. The cleaved homozygous TT variant showed two fragments of 175 and 23 bp. The cleaved heterozygous CT variant produced three fragments of 198, 175, and 23 bp. To ensure quality control, PCR-RFLP results for a subset of gene fragments were verified against exact nucleotide sequences for these fragments, as sequenced by Cortec DNA Service Laboratories .Blood samples were stored at –80°C and were defrosted to room temperature for analysis. DNA isolation was performed using the QIAamp DNA Blood Mini Kit spin protocol , and PCR-RFLP analysis was based on methods described previously , as tertiles based on intake among controls, and as ever use of a vitamin or mineral supplement containing folate for ≥6 months (yes/no). Intakes of vitamins B2, B6, and B12, and methionine were each analysed in these ways. Folate intake was also examined by Gleason score among cases, stratified into highly (score ≥7) and less aggressive (score 5–6) groups. Alcohol consumption was analysed as age started drinking, total alcohol intake per week (>5 drinks vs. ≤5), total beer, wine, and liquor per week (1–7 drinks and ≥8 drinks vs. <1 drink), and total lifetime alcohol intake . Analyses of alcohol variables were performed excluding never drinkers (n = 68 cases and 292 controls included). MTHFR genotype was analysed with risk of the CT (heterozygous mutant) and TT genotypes (homozygous mutant) compared with the CC genotype (wildtype), both individually and combined as CT + TT in a dominant genetic model , and low alcohol consumption was defined as ≤5 drinks per week (with >5 defined as high). Statistical significance of multiplicative and additive interactions was assessed from the likelihood ratio tests performed using the GENMOD procedure in SAS, which allows testing of these two types of interactions separately in general linear models .2, B6, B12, and methionine was similar (not shown).While most characteristics of cases and controls were similar Table , control2, B6, B12, or methionine (folate only shown in Table p-trend = 0.06).No association with prostate cancer risk was apparent for dietary folate analysed continuously, by tertile, or according supplement use as the source, nor for vitamins BMTHFR C677T genotype, and the multiplicative interaction between the two, the magnitude of risk was even higher .In terms of alcohol consumption, also seen in Table MTHFR C677T genotype among consenting cases, 44% displayed the CT genotype, 5% displayed the TT genotype, and 51% displayed with wildtype CC genotype. Among genotyped controls, these numbers were 51, 7, and 42%, respectively. The CT and TT MTHFR genotypes were not associated with prostate cancer risk  within the CC genotype  reduced the possibility of recall error (Middleton Fillmore et al., Dietary folate consumption among our study population was measured prior to 1998, when folate fortification of white wheat flour became mandatory in Canada and the United States. Dietary folate consumption and folate status of our study population are likely lower than that of the more recent general Canadian and US populations (Baily et al., MTHFR genotype. Taking into account potential combined effects and interactions is important for future studies, and may in part explain inconsistent findings to date. Future work should also capture a greater range of folate intakes to determine the true role of folate in prostate carcinogenesis, and whether this is role is dualistic in nature, to inform public policy regarding folate fortification.In summary, we found that alcohol consumption is associated with increased prostate cancer risk, with higher risk among men with low folate intake and among men with the CC The authors declare that this research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Infection with Epstein-Barr virus (EBV) has been suggested to contribute to the pathogenesis of systemic lupus erythematosus (SLE). We sought to determine whether prior infection with the virus occurs more frequently in patients with SLE compared to matched controls.We performed a systematic review and meta-analyses of studies that reported the prevalence of anti-EBV antibodies in the sera from cases of SLE and controls by searching Medline and Embase databases from 1966 to 2012, with no language restriction. Mantel-Haenszel odds ratios (OR) for the detection of anti-EBV antibodies were calculated, and meta-analyses conducted. Quality assessments were performed using a modified version of the Newcastle-Ottawa scale.Twenty-five case–control studies were included. Quality assessment found most studies reported acceptable selection criteria but poor description of how cases and controls were recruited. There was a statistically significant higher seroprevalence of anti-viral capsid antigen (VCA) IgG  1.15 – 3.76, p = 0.007) but not anti-EBV-nuclear antigen1 (EBNA1)  in cases compared to controls. The meta-analyses for anti-early antigen (EA) /D IgG and anti-VCA IgA also showed significantly high ORs , p = 0.0009 respectively). However, funnel plot examination suggested publication bias.Overall, our findings support the hypothesis that infection with EBV predisposes to the development of SLE. However, publication bias cannot be excluded and the methodological conduct of studies could be improved, with regard to recruitment, matching and reporting of blinded laboratory analyses. The pathogenesis of autoimmune diseases involves a complex interplay between genetic, environmental and stochastic factors. However, the precise nature and relative importance of these remain unclear. Advancement of our understanding of the environmental factors responsible for autoimmunity has, on the whole, lagged that of genetic factors. Much interest has focused on the role of infection in triggering autoimmune disorders, including systemic lupus erythematosus (SLE), by such mechanisms as molecular mimicry, bystander activation and epitope spreading . EpsteinEBV has the structure common to all herpes viruses of a large double-stranded DNA genome enclosed with an icosahedral capsid, including viral capsid antigen (VCA). Infection is usually not associated with symptoms when contracted in the first decade of life. Primary infection during adolescence may result in infectious mononucleosis. The virus infects B cells and establishes a latent cycle, persisting for life within the long-lived memory B-cell population of the host . During The nature of the association between EBV and autoimmunity, in particular the question of causality, remains to be fully elucidated. If the virus is an important factor, it follows that prior infection must be more common in those with the disease than healthy controls. Indeed, an association between MS and the virus has been demonstrated in several meta-analyses and infection predates development of the disease . SeveralThe systematic review was carried out according to a prespecified protocol developed by the authors . The study had to assay serum IgG or IgA antibodies to EBV in each group. We considered studies from any region and in any language. Studies were considered regardless of serological assay used and EBV antigen detected. Nonhuman studies were excluded, as were studies that only measured IgM responses.Two authors reviewed a sample of 200 titles and abstracts to identify those for which full texts should be sought and those likely to be eligible for inclusion. A kappa statistic of agreement  was high for both outcomes . Titles and abstracts of all articles were read by one author, all potentially relevant articles were identified and full texts of these articles were screened for eligibility. Any uncertainties were resolved by discussion between the authors. Reference lists of included studies were hand searched and citation searches of all relevant articles were conducted using the Web of Science: Thomson Reuters, New York, NY 10036 USA citation search tool to identify any additional articles not identified from the database search. Relevant data from each article were extracted independently by two authors using a standardised data extraction form  of seropositivity to EBV were calculated for each anti-EBV antibody  using Review Manager Version 5.1: Review Manager (RevMan) [Computer program]. Version 5.1. Copenhagen: The Nordic Cochrane Centre, The Cochrane Collaboration, 2011. The ORs for each antibody were combined in a meta-analysis. As we anticipated study heterogeneity, we used a conservative random-effects model with a 95% confidence interval (CI).  studies .One study analysed cases and controls in two, separate, race-matched groups . For thepost-hoc subgroup analysis testing of the meta-analysis to compare the OR of VCA seropositivity in the following categories to investigate potential sources of heterogeneity: studies using community controls versus those in which the source of controls was different or unspecified; studies conducted in Far Eastern populations versus black populations versus other populations, including where specific details on ethnicity were not available; and studies using age-matched controls versus those that did not.We conducted We sought to conduct similar analyses for blinding of laboratory analyses; however, only one study in the meta-analysis specified that this was done.Once duplicates had been removed, the search revealed 1,024 records. Screening of all titles and abstracts identified 55 studies potentially eligible for inclusion, for which full texts were obtained. Thirty-one studies were excluded  and 50 for the control groups (range 20 to 392). Most studies included cases of SLE defined by the American College of Rheumatology criteria (1982 or 1997 revision) ,42. The P = 0.02), indicating a significantly higher seroprevalence of anti-VCA in patients with SLE versus controls , indicating that the result should be interpreted cautiously. The distribution in the funnel plot appeared asymmetrical, raising the possibility that the effect may be due to publication bias . Meta-analysis revealed an overall OR of 2.08  . For the, which may reflect a tendency towards viral exposure at a younger age in Taiwan versus the USA [James and colleagues’ 1997 study is one of only two studies that specifically recruited young patient and control populations  . The ser the USA .On the basis of the use of age-matched and sex-matched community controls, with analyses conducted in a blinded manner, the highest quality studies were those by Parks and colleagues  and StraP = 0.32), indicating that the difference in seropositivity rates between SLE and control groups for this antibody was not significant .Eleven studies measured anti-EBNA1 IgG. A total of 705 SLE patients and 928 controls were included. There was a higher proportion of anti-EBNA1-positive lupus cases than controls . Four studies showed a higher anti-EBNA1 seroprevalence in the control group, and one study found no difference. Meta-analysis showed OR = 1.45  .Seven studies reported data on anti-EA seropositivity, with a total of 568 SLE cases and 368 controls. All assays were of antibodies to the EA/D antigen. Rates were significantly lower compared with anti-VCA and anti-EBNA-1 . Meta-analysis showed a significant association between these antibodies and SLE, with OR = 5.76  . In the subgroup analysis carried out by Parks and colleagues the OR was higher in the African American group than the white group (5.6 (95% CI 3.0 to 10.6) and 1.6 (95% CI 0.94 to 2.7), respectively) [Four studies also assayed anti-VCA antibodies of the IgA subclass, which we undertook as a ctively) .post-hoc subgroup analyses are shown in Additional file P = 0.12). No trends were seen in the other subgroups.The results of the exploratory post-hoc subgroup analyses failed to demonstrate a significant difference between any of the subgroups considered, although they did show a trend towards higher ORs in studies lacking a community control group. This observation, coupled with the observation that the highest quality studies according to the quality assessment failed to show any positive association, raises concern that differences in methodology and study population may account for the heterogeneity and perhaps the effect seen.This meta-analysis of case–control studies investigating the association between SLE and serological markers of EBV infection shows a significant association between the disease and anti-VCA IgG  but not between SLE and anti-EBNA1 IgG . We also found stronger associations between SLE and both anti-EA/D IgG and anti-VCA IgA (OR 5.76 (95% CI 3.00 to 11.06) and 5.05 (95% CI 1.95 to 13.13), respectively). This analysis is, to our knowledge, the first attempt to combine such estimates of association with SLE in a meta-analysis and should therefore provide a more robust estimate of the association than individual studies, which have tended to include relatively small numbers of participants. One should note, however, that the studies included varied widely in their quality, and there was considerable heterogeneity in the results of the meta-analyses. Our While a number of observations have been made linking EBV to autoimmune disease generally, and SLE specifically ,44, it rStrengths of this study include its comprehensive search strategy, without language restriction. Conversely, meta-analyses such as this are susceptible to publication bias, and there is a suggestion of this in the asymmetry of the funnel plot of those studies included in the anti-VCA analysis. The majority of these studies do not, however, claim a significant association between the antibody and SLE, as their sample sizes are, on the whole, too small to detect the effect and it is only when they are combined that a significant association is seen. Of further concern is the high level of heterogeneity between the ORs of the individual studies, the lack of consistency in matching of cases and controls, and the lack of reporting of recruitment and laboratory methodology, as seen in the quality assessment. We hoped to find prospective studies that would allow analysis of the relative timing of infection and the development of SLE, but our search revealed none. Nevertheless, infection in the general population occurs biphasically, generally below the age of 20, and would be expected to precede the onset of symptoms of SLE.Our results could perhaps be explained if there were higher false negative rates among controls or a greater proportion of false positive results among SLE sera. The observation that anti-VCA has been detected in nearly 100% of some normal, healthy populations makes the former scenario unlikely. That the assays could be more sensitive when using SLE sera, which have been shown to have elevated overall titres of anti-EBV antibodies as well as hypergammaglobulinaemia in general, is a more plausible explanation for the result being spurious, and this cannot be ruled out.The associations between anti-EA IgG and anti-VCA IgA seen here are more marked than with anti-VCA or anti-EBNA1. Unlike anti-VCA and anti-EBNA1 IgG, which persist following exposure, antibodies to EA/D are held to be detectable for only a transient period following primary infection and to rise again following reinfection or reactivation of EBV, in which the virus switches from latent to lytic cycles of replication within memory B cells. The fact that SLE sera in our analysis had significantly higher rates of anti-EA/D reactivity may implicate EBV reactivation in precipitating an autoimmune reaction; conversely, it may reflect an alteration in the relationship between the immune system and the virus in the setting of SLE. A third possibility is that immunosuppressive therapy used to treat the disease might lead to viral reactivation and an anti-EA/D response. Cross-linking of immunoglobulins on latently infected memory B cells has been shown to precipitate the lytic cycle of viral replication , and thiThe study with highest OR for anti-VCA IgG was by James and colleagues . This waIn summary, these findings support the hypothesis that prior infection with EBV is important in the development of SLE, as indicated by a higher seroprevalence of anti-VCA IgG. The results also suggest dysregulated  immune responses to EBV in the context of SLE. The studies included in this meta-analysis, however, are heterogeneous and, on the whole, have small sample sizes. Many do not match for age and sex, few match for race, and a specific description of recruitment and laboratory practice is lacking in many of the reports. The role of publication bias cannot be excluded. Further, larger studies with more precise matching and descriptions of recruitment and measurement, ideally using prospective serological measurements, addressing the questions of both precedence and the possibility of spurious results from abnormal humoral immune reactivity secondary to the disease, are needed to establish the relationship between infection with EBV and SLE.CI: Confidence interval; EA: Early antigen; EBNA: Epstein–Barr virus nuclear antigen; EBV: Epstein–Barr virus; MS: Multiple sclerosis; OR: Odds ratio; SLE: Systemic lupus erythematosus; VCA: Viral capsid antigen.The authors declare that they have no competing interests.PH, AA and MAV devised the protocol, made decisions on study inclusion, extracted data and wrote the paper. PH performed the systematic review and meta-analysis. LA was the statistical advisor. All authors read and approved the final manuscript.Figure S1 showing a flow diagram of search results and included/excluded studies. Figure S2 showing a funnel plot of studies of VCA seropositivity. Figure S3 showing random effects meta-analysis of seroprevalence of anti-VCA IgG between SLE and control – subgroup analysis of community controls and noncommunity controls. Figure S4 showing random effects meta-analysis of seroprevalence of anti-VCA IgG between SLE and control – subgroup analysis based on ethnicity of study populations. Figure S5 showing random effects meta-analysis of seroprevalence of anti-VCA IgG between SLE and control – subgroup analysis of age-matched and nonage-matched studies. Table S1 presenting the search strategy, detailing search terms and combinations used for each database. Study protocol: data extraction form and quality assessment tool.Click here for file


ESR1) has been suggested as a possible candidate gene for hip fractures; however, published studies of ESR1 gene polymorphisms have been hampered by small sample sizes and inconclusive or ambiguous results. The aim of this meta-analysis is to investigate the associations between two novel common ESR1 polymorphisms (intron 1 polymorphisms PvuII-rs2234693: C>T and XbaI-rs9340799: A>G) and hip fracture.Genetic factors are important in the pathogenesis of fractures. Notably, estrogen receptor α (Crude odds ratios (ORs) with 95% confidence intervals (CIs) were used to evaluate the strength of the association.Five case-control and three cohort studies were assessed, including a total of 1,838 hip fracture cases and 14,972 healthy controls. This meta-analysis revealed that the PvuII T allele is a highly significant risk factor for hip fracture susceptibility, with an effect magnitude similar in male and pre-menopausal and post-menopausal female patients. In stratified analysis based on ethnicity, the PvuII T allele remained significantly correlated with increased risk of hip fracture in Caucasian populations; this correlation, however, was not found in Asian populations. Unlike the PvuII polymorphism, we did not find significant differences in the XbaI (A>G) polymorphism allele or genotype distributions of hip fracture patients and controls. We also found no obvious association between the XbaI polymorphism and hip fracture in any of the racial or gender subgroups.ESR1 PvuII T allele may increase the risk of hip fracture and that the XbaI polymorphism is not associated with hip fracture.Our findings show that the  VDR) gene, insulin-like growth factor I (IGF-I) gene, collagen type I alpha 1(COL1A1) gene and estrogen receptor α gene (ESR1), etc Hip fracture, the most serious complication of osteoporosis, is a common musculoskeletal disorder ESR1, which are caused by alternative splicing that alters the expression of ESR1, are likely to affect hip fracture susceptibility ESR1 gene, including ESR1 XbaI  and PvuII , have been investigated for a possible association with hip fracture risk ESR1 region are associated with bone mineral density of the hip and spine, providing rapid insights into the genetic association between ESR1 gene and hip fracture risk ESR1 PvuII pp genotype and vertebral fracture existed in five case-control studies, but no association between the PvuII polymorphism and hip fracture were observed in one study Estrogen receptor α, a member of the nuclear receptor super-family of ligand-activated transcription factors, is one of the key mediators of hormonal response in estrogen-sensitive tissues ESR1 XbaI polymorphism may be associated with decreased risk of vertebral fracture in women and that the PvuII polymorphism is not associated with vertebral fracture risk ESR1 gene and hip fracture susceptibility. We hope this meta-analysis will help with the early identification and therapeutic treatment of hip fracture.Two previous meta-analyses have concluded that the ESR1’). The reference lists of the prospective articles were also reviewed to identify additional relevant publications and studies.We performed an electronic search for papers published before June 19th, 2013 in PubMed, Embase, Web of Science, Chinese National Knowledge Infrastructure (CNKI), and Chinese Biomedical Literature Database (CBM). Literature searches were performed by an expert using the following key words and MeSH terms: , (‘genetic polymorphism’ or ‘single nucleotide polymorphisms’ or ‘gene mutation’ or ‘genetic variants’) and  hip fracture cases must be diagnosed as pathologic fractures or fractures resulting from trauma other than a fall; (c) inclusion of sufficient data on sample size, odds ratio (OR), and 95% confidence interval (CI); and (d) published in the English or Chinese language.Studies included in our meta-analysis had to meet the following criteria: (a) population-based case-control, cohort or cross-sectional study focusing on associations of Studies were excluded when they represented duplicates of previous publications, or were meta-analyses, letters, reviews or editorial articles. Additionally, when data were included in multiple studies using the same case series, either the study with the largest sample size or the study with the most recently publication date was selected. All disagreements on study inclusion were resolved through discussions. To ensure the rigor of the current meta-analysis, we designed and reported the meta-analysis according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) statement. The relevant checklist is shown in All data from included studies were extracted independently by two investigators using a piloted data standardized form : the first author’s surname, year of publication, country of origin, published language, gender and ethnicity of study subjects, study design, number of subjects, SNP genotyping methods, genotyping method and detected sample, allele and genotype frequencies, and evidence of Hardy-Weinberg equilibrium (HWE) in controls. In addition, we also compared key study characteristics, such as location, publication date and authorship, to determine the existence of multiple publications from the same study.Two authors independently assessed the quality of the published articles according to the modified STROBE quality score systems ESR1 PvuII and/or XbaI polymorphisms and hip fracture risk under five genetic models: allele, dominant, recessive, homozygous, and heterozygous models. The deviation of frequency from those expected under Hardy-Weinberg equilibrium (HWE) was assessed by Chi-squared goodness of fit tests in the controls. We explored inter-study variation by prespecified subgrouping of studies according to ethnicity , gender  and menopausal status among women , where applicable. The statistical significance of the pooled OR was assessed with a Z test. Between-study variation and heterogeneity were estimated using Cochran’s Q-statistic, with P<0.05 as a cutoff for statistically significant heterogeneity Crude ORs together with their corresponding 95% CIs were used to calculate and assess the strength of associations between 2I test (ranges from 0 to 100%), which represents the proportion of inter-study variability that can be attributed to heterogeneity rather than to chance Mantel-Haenszel method) was used, except when a significant Q-test (P<0.05) or 2I>50% indicated the existence of heterogeneity among studies; otherwise, the random effects model (DerSimonian-Laird method) was applied to the meta-analysis. In order to ensure the reliability of results, a sensitivity analysis was performed by omitting individual studies. Begger’s funnel plots were used to detect publication bias. In addition, Egger’s linear regression test, which measures funnel plot asymmetry via a natural logarithm scale of OR, was also used to evaluate publication bias P-values were two-sided. Analyses were conducted with STATA Version 12.0 software . Meta analysis, sensitivity analysis and publication bias were conducted using the following STATA’s user-written functions: metan, metaninf and metabias, respectively.We also quantified the effect of heterogeneity with the ESR1, PvuII (rs2234693) and XbaI (rs9340799) in intron 1. A flow diagram of the studies selection process, as well as the specific reasons for exclusion from the meta-analysis is shown in Literature search yielded 129 reports, of which five population-based case-control ESR1 PvuII (C>T) polymorphism and hip fracture was investigated in eight studies, with a total of 1,838 cases and 14,972 healthy controls. Among these studies, six were conducted in Caucasian populations and two in Asian populations. The findings of this meta-analysis on the correlation between the polymorphism and hip fracture risk are summarized in The association of the P<0.001; dominant model: OR = 1.25, 95%CI: 1.09−1.44, P = 0.001; recessive model: OR = 1.26, 95%CI: 1.12−1.42, P<0.001; homozygous model: OR = 1.40, 95%CI: 1.19−1.64, P<0.001; heterozygous model: OR = 1.21, 95%CI: 1.07−1.36, P = 0.003), and the magnitude of the effect was similar in female  and male  . In postmenopausal women, we also observed that TT genotype is associated with higher risk of hip fracture than either TC or CC subjects  , while such result was not found in Asian populations   . In addi= 0.043) . Interes P>0.05) .ESR1 XbaI (A>G) polymorphism and hip fracture was investigated only in four studies, with a total of 853 cases and 4,522 healthy controls. Among these studies, three were on Caucasians and one on Asians. A summary of the meta-analysis findings on the association of the polymorphism with hip fracture risk is shown in P = 0.125; dominant model: OR = 1.14, 95%CI: 0.89−1.47, P = 0.290; recessive model: OR = 1.12, 95%CI: 0.96−1.30, P = 0.161; homozygous model: OR = 1.20, 95%CI: 0.93−1.57, P = 0.167; heterozygous model: OR = 1.10, 95%CI: 0.94−1.29, P = 0.245). The association between either ethnicity or menopausal status and the polymorphic genotypes were also not statistically significant for any of these analyses  (The association between the t shown) .ESR1 PvuII (C>T) or XbaI (A>G) under the allele model  or XbaI (A>G) .Begger's funnel plot and Egger's linear regression test were performed on the metadata to assess the publication bias of individual studies. The shapes of the funnel plots did not reveal any evidence of obvious asymmetry for either the aI (A>G) . Egger'sESR1 gene ESR1 region showed an association with bone mineral density of both the hip and spine, suggesting the possible role of the ESR1 gene in the pathogenesis of osteoporosis ESR1 gene and osteoporotic fractures ESR1 gene are the PvuII and XbaI polymorphisms, which have been linked to a lower sensitivity to estrogen ESR1 gene polymorphisms and fracture risk is still controversial and ambiguous. To date, there are only two published meta-analyses that evaluate the possibility of a significant association between the ESR1 PvuII polymorphism and risk of fracture. The meta-analysis by Ioannidiset al. showed that the PvuII polymorphism was not associated with fracture risk ESR1 PvuII pp genotype and vertebral fracture ESR1 gene and risk of hip fracture.Over the past decade, many linkage and association studies have been performed to find candidate genes in order to analyze the relationship between genetic factors and complex traits, such as BMD and/or fracture risk ESR1 polymorphisms and hip fracture. The results of this meta-analysis show that the PvuII polymorphism has a statistically significant association with hip fracture, especially in Caucasian populations but not in Asian populations. The discrepancy between ethnicity subgroups may be due to sample size since only 188 cases and 248 controls were studied in Asian populations. Other than family history and ethnicity, the most well-established risk factors for hip fracture are gender and menopausal status in females ESR1 XbaI (A>G) polymorphism. We found no obvious association between the XbaI polymorphism and hip fracture in either racial or gender subgroups. This discrepancy in the associations of the two polymorphisms may be due to sample size since there are only eight studies on PvuII and four on XbaI. Although these variants alone are not clinically useful in the prediction of risk to the individual person, their interactions with candidate genes identified by the GWAS may play important roles in the pathogenesis underlying hip fracture.This meta-analysis evaluated the association between two common ESR1 gene, including polymorphisms of the VDR gene, the COL1A1 gene, the IGF-I gene, and several other candidate genes. Thus, our meta-analysis emphasizes that elucidating the pathogenesis of fracture demands an investigation into the association of many gene variants constituting distinct pathophysiological pathways. Second, we should emphasize that this meta-analysis does not address the question of whether the ESR1 gene shows evidence of genetic linkage to BMD. Third, we identified only two studies from Asian populations and obtained no data from African populations, thus these two racial groups demand further studies. Last, this meta-analysis was based on unadjusted ORs estimates as a result of the lack of available information, preventing a more precise evaluation with adjusted ORs by certain covariates such as age, BMI, and smoking status, etc; thus, our data is only a conservative estimates of the association between ESR1 gene and hip fracture. Ideally, our conclusions will be tested by future studies. Despite the limitations listed above, our meta-analysis has some strength. To the best of our knowledge, this is the first meta-analysis on the relationship between the ESR1 gene polymorphisms and hip fracture. We explored inter-study variations by subgrouping studies according to ethnicity, gender and menopausal status for females. Furthermore, although this meta-analysis does not accommodate all previously published data, that is limited compared to the evidence here generated.In interpreting the results of this meta-analysis, some specific issues should be mentioned. First, as with other complex traits, hip fracture risk may be modulated by several other genetic markers besides ESR1 gene may be a risk factor for hip fracture, with a similar effect magnitude in both pre-menopausal and post-menopausal females, and male. However, unlike with the PvuII polymorphism, no significant difference was found in the XbaI (A>G) polymorphism allele or genotype distribution between hip fracture patients and controls, either in racial or gender subgroups. Thus, our results do support the hypothesis that the PvuII (C>T) polymorphism has potential clinical value as a predictors of hip fracture. Based on the limitations mentioned above, it is critical that large, well-designed studies are performed to re-evaluate the potential associations between ESR1 gene polymorphisms with other candidate gene polymorphisms and hip fracture risk.In summary, this meta-analysis of eight studies indicates that allelic variation in the PvuII (C>T) polymorphism of Supplement S1PRISMA Checklist.(DOC)Click here for additional data file.Supplement S2Modified STROBE quality score systems.(DOC)Click here for additional data file.Supplement S3The genotype distribution and allele frequencies of ESR1 PvuII and XbaI polymorphisms.(XML)Click here for additional data file.


Several studies have investigated whether the polymorphism in the apolipoprotein A5 (APOA5) is associated with type 2 diabetes mellitus (T2DM) risk. However, those studies have produced inconsistent results. The purpose of this study was to investigate whether the APOA5 -1131T/C polymorphism (rs662799) confers significant susceptibility to T2DM using a meta-analysis.PubMed, Embase, Web of Science, Cochrane database, CBMdisc, CNKI and Google Scholar were searched to get the genetic association studies. All statistical analyses were done with Stata 11.0.p<0.00001; for C/C vs. T/T: OR = 1.57, 95% CI = 1.35–1.83, p<0.00001; for C/C vs. T/C+T/T: OR = 1.36, 95% CI = 1.18–1.57, p<0.0001; for C/C+T/C vs. T/T: OR = 1.32, 95% CI = 1.16–1.51, p<0.0001). In the subgroup analysis by ethnicity, significant association was also found among Asians . However, no significant association was found between the APOA5 -1131T/C polymorphism and T2DM risk among Europeans.A total of 19 studies included 4,767 T2DM cases and 10,370 controls (four studies involving 555 T2DM cases and 2958 controls were performed among Europeans and 15 studies involving 4212 T2DM cases and 7412 controls were performed among Asians) were combined showing significant association between the APOA5 -1131T/C polymorphism and T2DM risk (for C allele vs. T allele: OR = 1.28, 95% CI = 1.17–1.40, The present meta-analysis suggests that the APOA5 -1131T/C polymorphism is associated with an increased T2DM risk in Asian population. Type 2 diabetes mellitus (T2DM) is a common, multifactorial disease that causes significant morbidity and mortality worldwide Recently, numerous epidemiological studies have focused on the associations between the apolipoprotein A5 (APOA5) gene polymorphisms and T2DM risk, and indicate that the APOA5 gene polymorphisms exert important role in the development of T2DM We performed this meta-analysis in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) criteria The studies included in the meta-analysis must meet all the following inclusion criteria: (1) evaluation of the association between APOA5 -1131T/C polymorphism and T2DM risk; (2) the design had to be a case-control study or cohort study; (3) sufficient published data for calculating odds ratios (ORs) with their 95% confidence intervals (CIs); and (4) not republished data.For each study, the following information was extracted: name of the first author, year of publication, study population , source of controls , sample size , and genotype frequency in cases and controls. Two authors (Yin YW and Sun QQ) independently assessed the articles for compliance with the inclusion criteria, resolved disagreements and reached a consistent decision.To determine the methodological quality of each study, we used the Newcastle-Ottawa scale (NOS), which uses a “star” rating system to judge the quality of observational studies p<0.0125 was considered to be statistically significant). Hardy-Weinberg equilibrium (HWE) in the control group was assessed using the chi-square test (p<0.05 was considered significant deviation from HWE).The strength of the association between the APOA5 -1131T/C polymorphism and T2DM risk was evaluated by the ORs with 95% CIs under the allelic model , additive model (C/C vs. T/T), recessive model (C/C vs. T/C+T/T), and dominant model (C/C+T/C vs. T/T). To adjust for multiple comparisons, we applied the Bonferroni method, which control for false positive error rate  p<0.05 was considered representative of statistically significant publication bias). Data analyses were performed using Stata software  and Review Manager software 5.1.4 .Cochran's Q statistic and the IThe present study met the PRISMA statement requirements . The inip<0.00001; for C/C vs. T/T: OR = 1.57, 95% CI = 1.35–1.83, p<0.00001; for C/C vs. T/C+T/T: OR = 1.36, 95% CI = 1.18–1.57, p<0.0001; for C/C+T/C vs. T/T: OR = 1.32, 95% CI = 1.16–1.51, p<0.0001). The main results of meta-analysis were shown in Given that significant heterogeneity existed in the allelic model and dominant model, we chose the random-effects model to synthesize the data of these two genetic models. As for the additive model and recessive model, no heterogeneity was found in these two genetic models. Therefore, we used fixed-effects model to analyze the data of additive and recessive models. The overall results showed evidence of significant association between the APOA5 -1131T/C polymorphism and T2DM risk, suggesting that the APOA5 -1131T/C polymorphism was a risk factor for T2DM , but not in Europeans . The main results of subgroup analysis were shown in In the subgroup analysis based on ethnicity, significant association was also found between the APOA5 -1131T/C polymorphism and T2DM risk in Asians , NOS score (studies with score ≥7 were included), and source of controls (studies with population-based controls were included). Overall, the pooled ORs and 95% CIs were not materially altered when any part of the study was omitted, indicating that our results were statistically robust. The results of sensitivity analyses were shown in Q = 0.05, I2 = 38%) and dominant model . In contrast, the additive model  and recessive model  did not present significant heterogeneity. To clarify the sources of heterogeneity, we first conducted the subgroup and sensitivity analyses. We only effectively removed the heterogeneity in the European group. We next created a Galbraith plot to graphically assess the sources of heterogeneity. A total of four studies Significant heterogeneity existed in the allelic model . Therefore, there was no risk of publication bias in the present meta-analysis.Begg's funnel plot and Egger's regression test were performed to assess potential publication bias. The funnel plot did not show obvious asymmetry in any genetic model . In addiThe APOA5 gene was identified by Pennacchio et al. in 2001 To the best of our knowledge, this is the first comprehensive meta-analysis to date investigating the association between the APOA5 -1131T/C polymorphism and T2DM risk. Nineteen studies involving 4,767 T2DM cases and 10,370 controls were included in the present meta-analysis. The overall results showed that there was significant association between the APOA5 -1131T/C polymorphism and the T2DM risk, suggesting the C allele was a independent risk factor for the development of T2DM. The findings showed that the risk of developing T2DM in C allele carriers was 1.32-fold higher than those without. Furthermore, the individuals with C/C genotype had a significantly higher risk for developing T2DM (for OR = 1.57 in additive model and OR = 1.36 in recessive model) compared to those with T/C genotype and T/T genotype.Considering that potential ethnic difference might be associated with the distribution of genotypes, we also performed subgroup analysis by ethnicity of study population. We only obtained the positive result in the Asians, which further consolidated the overall results. For Europeans, we did not find significant association between the APOA5 -1131T/C polymorphism and T2DM risk. Actually, the minor allele frequency (MAF) of APOA5 -1131T/C is very low among utah residents with Northern and Western European ancestry (CEU) (MAF = 0.015 in HapMap CEU) In the present meta-analysis, we found significant association between the APOA5 -1131T/C polymorphism and T2DM risk, particularly in Asian population. However, so far no definite evidence exists to support that the APOA5 -1131T/C polymorphism is directly related to T2DM susceptibility. It is well known that high plasma TG level is an important risk factor in developing T2DM Between-study heterogeneity should not be ignored and should be carefully factored in the interpretation of the finally results For better interpreting the results, some limitations of this meta-analysis should be acknowledged. Firstly, there was significant between-study heterogeneity in the allelic model and dominant model. Heterogeneity is a problem that may affect the precision of overall results. Secondly, in 19 studies included in this meta-analysis, only four of them are European samples, which may develop a partial result. Thirdly, the controls were not uniformly defined. Both healthy individuals and patients without T2DM in hospital were included in the control group. The hospital-based controls may not be representative of the randomized population. Therefore, some inevitable selection bias might be brought into the meta-analyses, which might affect the interpretation of the final results. Moreover, meta-analysis is a type of retrospective study, and recall and selection bias might inevitably exist.In conclusion, the present meta-analysis suggests that the APOA5 -1131T/C polymorphism is associated with an increased T2DM risk in Asian population. However, the result should be interpreted with caution because of its limitations. Larger and well-designed studies based on different ethnic samples are needed to confirm our results in the future.Checklist S1PRISMA 2009 checklist.(DOC)Click here for additional data file.


NKX2-5, 63A>G (rs2277923) and 606G>C (rs3729753), have been investigated in the risk of congenital heart disease (CHD), although with inconsistent results. Thus, a meta-analysis was performed to clarify the associations between the two variants and CHD risk in the Chinese population.NKX2-5 is a transcriptional factor, which plays an important role in heart formation and development. Two genetic variants in the coding region of Pheterogeneity = 0.009, I2 = 65.1%), but not for 606G>C . Regarding 63A>G variant, positive results were also obtained in the subgroups of atrial septal defect and large-sample-size study. Besides, the sensitivity analysis indicated that significant association was still detected after deletion of the individual studies with positive result and striking heterogeneity.Relevant studies were identified by searching PubMed, ISI Web of Science and CNKI databases and by reviewing the reference lists of retrieved articles. Then, the data from eligible studies were combined in an allelic model. A total of 7 and 4 studies were ultimately included for 63A>G and 606G>C, respectively. The results of overall meta-analyses showed that significant association was detected for 63A>G (OR = 1.26, 95% CI = 1.02–1.56, NKX2-5, but not the 606G>C, may contribute to CHD risk for Chinese.Our results revealed that the 63A>G variant in  Congenital heart disease (CHD) is defined as a gross structural anomaly of the heart or intrathoracic great vessels that is actually or potentially of functional significance NKX2-5 gene, located at chromosome 5q34, was firstly identified to be involved in heart development of many organisms, such as zebrafish, frog, chicken, mouse and human being. A series of clinical manifestations for CHD including arrhythmia, cardiac contractility defects, cardiac structural defects and premature death were observed in NKX2-5 knocked out mice NKX2-5 would also lead to CHD NKX2-5 not only individually acts in transcriptional regulation, but also interacts with other transcription factors in the early stage of cardiac development NKX2-5, was also suspected to implicate in CHD risk in the Chinese population. Based on the controversial conclusions and limited statistical power of individual studies, as well as almost all relevant data obtained from the Chinese population, we performed a meta-analysis of published studies to provide more precise estimations for the associations between 63A>G and 606G>C variants and the risk of CHD in Chinese population.Cardiac transcription factors, such as NKX2-5, GATA4, TBX5 and TBX20, strictly regulate the development of heart through a regulatory network NKX/CSX” and “congenital heart disease/defects” without language restriction. To expand the coverage of our searches, we further performed search in China National Knowledge Infrastructure (CNKI) database applying the above key words. References of retrieved articles were also scanned. We conducted the meta-analysis and reported its results according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) statement (Relevant articles published up to February 2013 were identified by searching PubMed and ISI Web of Science databases using the key words “tatement .NKX2-5 63A>G and/or 606G>C variant(s) and CHD risk; (3) non-syndromic CHD confirmed histologically or pathologically; (4) presentation of data necessary for calculating odds ratios (ORs) with 95% confidence intervals (CIs). All clinical types, such as atrial septal defect (ASD), ventricular septal defect (VSD), patent ductus arteriosus and patent formen ovale were included in this meta-analysis. Review, case report, simply commentary, animal study, unpublished report and the study with CHD being considered as a component of well-known genetic syndromes or a multiple congenital anomaly syndrome were excluded.We used following criteria to select the eligible studies: (1) case-control studies in the Chinese population; (2) evaluation of the Data were extracted independently by two reviewers. The following information was extracted from the eligible studies: first author, year of publication, design type of study, types of CHD, source of DNA, whether Hardy-Weinberg equilibrium (HWE) accorded in control population and counts of alleles or genotypes in case and control groups.x2 test was applied to assess HWE in controls if there were no definitive statements about whether the genotype frequencies in control groups were compatible with HWE in the original. The associations between NKX2-5 63A>G, 606G>C variants and risk of CHD were evaluated by allelic ORs and their 95% CIs because of only allele frequencies reported in some studies. The Cochran’s Q statistic test was employed to estimate the between-study heterogeneity P value of Q statistic test was <0.1, the random-effects model was used due to significant heterogeneity P values less than 0.05 were considered statistically significant for all tests except for Q test. Besides, the power analysis was carried out by Power V3.0 software (http://dceg.cancer.gov/tools/design/POWER).Goodness-of-fit x2 test due to lack of genotype frequencies Pheterogeneity = 0.009, I2 = 65.1%). Individuals with the NKX2-5 63G allele showed a significantly increased CHD risk compared with those with the A allele . For 606G>C, no substantial heterogeneity was detected (Pheterogeneity = 0.412). In the fixed-effects model, no significant association between this variant and CHD was found . However, no statistical evidence for the association between 63A>G and VSD was detected . While no significant risk of CHD associated with 63A>G was observed in the small-sample-size subgroup  and Egger’s test, no publication bias was detected for 63A>G (P = 0.249) and 606G>C in NKX2-5 (P = 0.797).As reflected by funnel plots , TN and NK2-specific (NK2-SD) domains. It is estimated that there are more than 33 variants detected in NKX2-5 as yet, in which, the 63A>G and 606G>C polymorphsims studied most extensively were included in our meta-analysis NKX2.5 by approximately 20% To explore the sources of heterogeneity, we further conducted stratified and sensitivity analyses for 63A>G variant. Stratified analyses by the types of CHD and sample size suggested that heterogeneity only presented in the VSD and small-sample-size subgroups, while heterogeneity was effectively removed after deletion of the study by Shi et al. in the sensitivity analysis. Interestingly, the study by Shi et al. was included in the VSD and small-sample-size subgroups, implying that the heterogeneity found in the two subgroups might result from this study. Under review of this report, Shi et al. proposed that 63A>G variant was not significantly associated with ASD, but with VSD. Individuals carrying G allele had a 4.32 - fold increased risk of VSD than those with A allele, which did not conform to other studies. But positive result was still observed after removal of the study conducted by Shi et al., indicating the relatively stability of current meta-analysis. Besides, marginal statistical significance was presented in the sensitivity analysis when several studies were omitted, which was probably caused by limited data and modest effect of this variant.NKX2-5 gene may differ across racial backgrounds, while the current meta-analysis was specifically based on the Chinese population, baffling the generalization of our conclusion to other ethnic populations. Second, the sample size of our meta-analysis was relatively small, especially for NKX2-5 606G>C. Besides, all included studies were retrospectively designed. Thus, additional large-scale and well designed studies will be required to further confirm our results. Third, we only considered the allelic model in current study because of limited data. Other genetic models should be taken into account to validate our results. Fourth, some heterogeneous natures of studies, including different phenotypes of CHD, female/male ratios and match conditions of control group probably affected our results. However, we were unable to perform further analyses due to lack of detail data.Some limitations of this meta-analysis should be acknowledged. First, there is evidence that the roles of variants in NKX2-5 63A>G and 606G>C variants with CHD and revealed that the 63A>G, but not the 606G>C was significantly associated with the risk of CHD in the Chinese population. Although the summary risk for developing CHD with the variants of NKX2-5 gene may be small, CHD occurs in high incidence in China and even a small increase in risk will translate to a large number of potential CHD cases.In summary, our meta-analysis helped for clarifying the discrepancies of genetic studies into associations of Figure S1The funnel plot for NKX2-5 63A>G.(TIF)Click here for additional data file.Figure S2The funnel plot for NKX2-5 606G>C.(TIF)Click here for additional data file.Checklist S1The PRISMA 2009 Checklist.(DOC)Click here for additional data file.