Since PPARγ2 is expressed mainly in adipose tissue, a moderate reduction of its activity influences the sensitivity to insulin, diabetes, and other metabolic parameters. The present study aims to contribute to the elucidation of the impact of the Pro12Ala polymorphism associated with T2D and obesity through a meta-analysis study of the literature that included approximately 11500 individuals, from which 3870 were obese and 7625 were diabetic. Statistical evidence supports protective effect in T2D of polymorphism Pro12Ala of PPARγ2 . Conversely the same polymorphism Pro12Ala of PPARγ2 seems to favor obesity since 1.196 more chance than nonobese was found . Our results suggest that Pro12Ala polymorphism enhances both adipogenic and antidiabetogenic physiological role of PPARγ. Does Pro12Ala polymorphism represent an evolutionary step towards the stabilization of the molecular function of PPARγ transcription factor signaling pathway?Obesity and type 2 diabetes mellitus (T2D) are two major public health problems that have motivated the scientific community to investigate the high contribution of genetic factors to these disorders. The peroxisome proliferator activated by gamma 2 (PPAR Peroxisome proliferator-activated receptors (PPARs) are transcription factors from nuclear receptor's protein family that regulate target genes' expression by connecting to response elements of peroxisomes proliferators (PPERs) in regulating sites of each gene. The signal transduction mechanism of these receptors involves retinol X receptor (RXR) and PPARs form heterodimers that regulate the transcription of several genes  1–3].–3.1–3].α, PPARβ, and PPARγ, the latter is the most studied  Pro12Ala, (2) Pro12Ala [AND] obesity, and (3) Pro12Ala [AND] diabetes mellitus 2. The study was conducted according to PRISMA directives. In order to identify the studies, an electronic search, with no linguistic restriction, was conducted, researching two databases, the ScienceDirect and Medline (PubMed). The Boolean search was conducted in both databases according to the following keywords: (1) PPARTwo investigators evaluated, independently, the titles and abstracts of the papers identified as of potential relevance in the inclusion criteria. Where a title or abstract could not be included for certainty, a third corrector would help decide whether to reject or not. 12Ala polymorphism as a comparison between control groups and obese or diabetic groups, were then used for systematic revision. The papers that satisfied the initial inclusion criteria and that were identified as of randomized or quasi-randomized clinical trial conducted in human, and used the ProAfter the papers' analysis, the data of interest was used based on the place where the study was conducted, year of publication, outcome, and number of individuals. P value, and odd ratios (OR), with and with a confidence interval of 95%.The statistical analysis included false positives, false negatives, true positives, and true negatives to establish sensitivity and specificity and correlate both in a Roc curve, For the data analysis was used the program RevMan version 5.1 and the SPSS Statistics version 17.0.12Ala polymorphism in obesity and 8 to the presence of Pro12Ala polymorphism in T2D.The research identified 11 studies, from which 6 were related to the presence of ProThe studies related to obesity investigated a total of 3870 individuals, with 2260 being obese and 1610 nonobese . 12Ala polymorphism and T2D investigated a total of 7625, with 4464 being diabetic and 3161 nondiabetic , despite possessing a lower abdominal diameter [González Sánchez and collaborators were the firsts to conduct a population-based nationwide multicenter study in Spain, suggesting that Prodiameter .P value of 0.04 and OR = 1.37  [Ghoussaini and collaborators studied the French population with obesity and T2D, in 2005. Regarding the T2D study, they affirmed that the polymorphism has a risk role, since a significant association between the polymorphism and T2D was found, with a 2; 1.85) . 12Ala polymorphism and T2D incidence, in a high-risk population with diminished glucose tolerance, in a longitudinal study. No significant association of the Ala12 allele and T2D incidence in the studied group [Lindi and collaborators studied a diabetic Finnish population. They were the first to try an association between Proed group  was founChistiakov and collaborators analyzed Russian individuals with T2D. Their results suggest that the polymorphism reduces T2D risk, performing a protective role in this pathology. In the studied population, the Pro12 allele contributes to higher risk in developing T2D, according to OR = 1.69  . 12Ala polymorphism and T2D, in diabetic Koreans. They compared individuals with normal and diminished glucose tolerance and with diabetes. No significant differences were found among Ala12 allele frequency in the study groups, concluding an inexistent significant association between Pro12Ala polymorphism and T2D. However, despite these results, they affirm that there is the possibility of an effect, at minor scale, of the polymorphism on the pathology, when associated with other mutations on other genes, as well with environmental factors [Oh and collaborators analyzed the association between Pro12Ala polymorphism and Tunisian T2D carriers. They concluded that there was no significant differences between diabetic and control groups and that, therefore, the polymorphism does not display a role in the pathology [Bouassida and collaborators studied the association of Proathology . 12Ala polymorphism with T2D incidence in a polish population. They compared the polymorphism incidence in a diabetic and a nondiabetic (control) group and notice that the Pro12 and Ala12 alleles' frequency were similar , as well as the genotypic distribution. They could not conclude that the polymorphism confers higher risk to T2D susceptibility, as in numerous European studies in Caucasians [Malecki and collaborators studied the Proucasians . 12Ala polymorphism and T2D. In order to do so, they studied the Czech population, comparing a study group of 133 diabetic individuals to a nondiabetic control group of 97 individuals. In the study group, 3 individuals (2.26%) were identified as homozygotes for the Ala/Ala genotype, 99 individuals (74.44%) were identified as homozygotes for the Pro/Pro genotype, and 31 individuals (23.31%) were identified as heterozygotic. In the control group, 6 individuals (6.19%) were identified as homozygotes for the Ala/Ala genotype, 61 individuals (62.89%) were identified as homozygotes for the Pro/Pro genotype, and 30 individuals (30.93%) were identified as heterozygotic. The allelic frequency for the Ala allele was lower in the diabetic group . There was no difference (P = 0.05) among phenotypic characteristics  in the studied group regarding genotype Pro12Ala. Therefore this study supports the hypothesis that the polymorphism plays an important role in T2D for the Czech population. The results showed that the allelic frequency of for Ala12 is higher in control group than in study group. Overall the polymorphism is associated with reduced risk of T2D, assuming a protective role in the pathology [Pintérová and collaborators, in 2004, wanted to understand the association between the Proathology . Mori and collaborators analyzed the polymorphism and T2D in a Japanese population. They included in their study 2201 individuals with T2D (study group) and 1212 individuals without the pathology (control group). The allelic frequency of Ala12 was higher in in control group (4.13% versus 2.39% for the study group). They concluded that the polymorphism is associated with reduced risk of T2D development, acting out protectively factor in the pathology .12Ala polymorphism in obesity is showed on The combination of the studies that analyzed the presence of Pro12Ala polymorphism in T2D, the combination of all included studies is presented in Regarding the presence of Pro12Ala polymorphism were included in the present meta-analysis. From these, 6 studies investigated the hypothetical association of the polymorphism and obesity and 8 investigated the hypothetical association of the polymorphism, Tables Eleven studies that investigated the ProIn In 12Ala polymorphism is not a good test (regarding sensitivity and specificity) to predict the presence of obesity, since the test line is very close to the random line. The same conclusion can be taken from the analysis of the Pro12Ala polymorphism which is not a good test to predict the presence of T2D. The results from these tables allowed constructing the respective ROC curve (not shown) and we conclude that the Proodds ratio (OR) and P values, by the Chi-squared test. Tables P value for obesity and T2D, respectively, and Tables We also proceed to the determination of the P < 0.004).Regarding the association between the polymorphism and obesity, obese individuals possess a 1.196 more chance of carrying the polymorphism than non-obese .Concerning the association between the polymorphism and T2D, diabetic individuals possess a 0.702 less chance of carrying the polymorphism than nondiabetic  and as a protective factor for T2D , by the evident statistical significance. Therefore, the Pro12Ala polymorphism is a risk factor in obesity or T2D, while others conclude the opposite for both pathologies. There are also studies that do not have statistically significant data to support either conclusion.Comparing results obtained from this meta-analysis and each of the included studies, we can verify some differences. There are studies that conclude that ProBen Ali et al. , GhoussaResults regarding the role of the polymorphism in T2D, Boussida et al. , Lindi e12Ala polymorphism with body mass in different human populations. Numerous studies have also clearly showed the heterogeneous effects of Pro12Ala polymorphism in decreased risk of T2D several populations. Considering that the two pathologies are multifactorial it is difficult to clarify the real contribution that the polymorphism makes in these metabolic disorders.Association studies clearly show this gene's regulation in adipose tissue is a complex process. Several studies suggest that genetics or the environment (such as diet) participates in the formation of association patterns of Pro12Ala polymorphism from PPARγ2 gene has a protective role in T2D but is a risk factor for obesity accordingly with what we should expected substitute to for about to the physiological role of PPARγ itself. It seems that the Pro12Ala polymorphism may be a transitory state towards the structural and functional stabilization of the gene role in human kind, resembling a genetic drift at the molecular level.The present study includes approximately 11500 individuals, from which 3870 are obese and 7625 are diabetic. From our results we concluded that the Proγ cascades.Therefore, this study supports the need of investment for further research, mainly by population genetics, in order to fully understand the role of this polymorphism in the PPAR


Previous studies have examined the associations between polymorphisms of adiponectin gene (ADIPOQ) and cardiovascular disease (CVD), but those studies have been inconclusive. The aim of this study was to access the relationship between three single nucleotide polymorphisms (SNPs), +45 T > G (rs2241766), +276 G > T (rs1501299) and -11377 C > G (rs266729) in ADIPOQ and CVD.2. Publication bias was estimated using modified Egger’s linear regression test.A comprehensive search was conducted to identify all studies on the association of ADIPOQ gene polymorphisms with CVD risk. The fixed and random effect pooled measures (i.e. odds ratio (OR) and 95% confidence interval (CI)) were calculated in the meta-analysis. Heterogeneity among studies was evaluated using Q test and the IP = 0.004), 0.90  and 1.09 for rs2241766, rs1501299 and rs266729, respectively. Rs2241766 and rs1501299 were significantly associated with coronary heart disease (CHD), yielding pooled ORs of 1.29  and 0.89 , respectively. The pooled OR for rs266729 and CHD was 1.09 . Significant between-study heterogeneity was found in our meta-analysis. Evidence of publication bias was observed in the meta-analysis.Thirty-seven studies concerning the associations between the three polymorphisms of ADIPOQ gene and CVD risk were enrolled in this meta-analysis, including 6,398 cases and 10,829 controls for rs2241766, 8,392 cases and 18,730 controls for rs1501299 and 7,835 cases and 14,023 controls for rs266729. The three SNPs were significantly associated with CVD, yielding pooled ORs of 1.22 (95%CI: 1.07, 1.39; The present meta-analysis showed that the associations between rs2241766, rs1501299 and rs266729 in the ADIPOQ and CVD were significant but weak. High quality studies are still needed to confirm the associations, especially for rs2241766. Adiponectin is a 30-kDa protein that consists of an N-terminal collagenous domain and a C-terminal globular domain, with ciAdiponectin plays a role in preventing atherosclerosis. Epidemiology studies have found that adiponectin levels were associated with risk of cardiovascular disease (CVD). Some cross-sectional studies have demonstrated that hypoadiponectinemia was associated with the prevalence of CVD-7. ProspP-combined = 9.2 × 10−19, n = 14,733) [There have been a number of studies reported that certain polymorphisms in the adiponectin coding gene ADIPOQ were strongly associated with adiponectin levels -13. A re 14,733) . The thrLacquemant et al. reported that rs2241766 was associated with an increased risk of coronary artery disease among patients with type 2 diabetes. Bacci eMeta-analysis is a powerful tool for summarizing results from different studies by producing a single estimate of the major effect with enhanced precision. In this study, we conducted a meta-analysis to examine the associations between the three SNPs in the ADIPOQ gene and CVD.Two independent reviewers (Zhang and Mo) conducted a systematic computerized literature search for papers published before February 12, 2012. PubMed, Embase and Wanfang databases were searched, using various combinations of keywords, such as ‘cardiovascular disease’ or ‘coronary heart disease’ or ‘coronary artery disease’ or ‘myocardial infarction’ or ‘stroke’ combined with ‘ADIPOQ’ or ‘APM1’ or ‘ACDC’ or ‘adiponectin’ and ‘polymorphism’ and ‘genetic association’, without language restriction. The full texts of the retrieved articles were read to decide whether information on the topic of interest should be included. The reference lists of the retrieved articles as well as those of review articles and previous meta-analyses on this topic were searched to identify other studies that were not identified initially. Articles were included in the meta-analysis if they examined the hypothesis that ADIPOQ polymorphisms were associated with CVD using case–control or cohort design, and had sufficient published data on the genotypes or allele frequencies for determining an estimate of relative risk (i.e. odds ratio (OR)) and confidence interval (CI). Our meta-analysis was conducted according to the Meta-analysis of Observational Studies in Epidemiology (MOOSE) guidelines .Two reviewers (Zhang and Mo) independently examined the retrieved articles using a data collection form, in order to extract the information needed. From each study, the following data were abstracted: first author, year of publication, country of the population studied, study subjects and main outcomes, the mean age and body mass index (BMI) and the percentage of men in case and control groups, polymorphisms tested, genotyping methods, the number of persons with different genotypes in cases and controls, and main results. Following data extraction, the reviewers checked for any discordance until a consensus was reached.Quality of studies was also independently assessed by the same two reviewers, using guidelines proposed by the NCI-NHGRI Working Group on Replication in Association Studies . These g2 (ranging from 0 to 100%) was also calculated, where higher values of the index (I2 > 50%) indicate the existence of heterogeneity[2). On the other hand, the random effects hypothesis is appropriate for clinical trials but results in relatively reduced power for genetic association/GWAS detection of SNPs which show association in at least one study [P value < 0.05, except for tests of publication bias where 0.1 was used as significant level.The OR was used to compare alleles between cases and controls. We computed the genetic contrast of the mutant alleles  versus the wildtype alleles. In secondary analyses, we calculated specific ORs according to the racial descent of subjects , study subjects , sample size (<1000 or ≥1000), genotyping methods (PCR-RFLP or Taqman genotyping assay or other). The association for coronary heart disease (CHD) was also examined. We assessed the presence of between-study heterogeneity using the chi-square based Cochran’s Q statistic. The inconsistency index Irogeneity. The poone study . The ranne study . To examAfter the literature searching and the subsequent screening, we came up with 34 research papers consisting of 37 case–control or cohort studies concerning the association of rs2241766 or rs1501299 or rs266729 polymorphisms with CVD  and 1.22 . There was a significant between-study heterogeneity as indicated by the P value of the corresponding test (P < 0.001) and the value of the I2 index (I2 =74.2%). The sensitivity analysis revealed that there was not a single study influencing the result significantly. Cumulative analysis found the influence of the first published study on the subsequent publications and the evolution of the combined estimate over time. Lacquemant et al. reported the significant association in 2003 [P = 0.007). The association for CHD alone was also significant, with an OR of 1.29  and 0.90 . Significant heterogeneity was observed . The sensitivity analysis revealed that there was not a single study influencing the result significantly. Cumulative analysis did not find the influence of the first published study on the subsequent publications and the evolution of the combined estimate over time. There was publication bias in the studies . The combined OR for rs1501299 and CHD was 0.89  and 1.09, and significant heterogeneity  , with significant between-study heterogeneity   of 1.07  Figure . By perfThe present meta-analysis, involving 6 to 8 thousand cases and more than 10 thousand controls for each polymorphism, provides a clear indication of significant associations between the three SNPs, rs2241766, rs1501299 and rs266729, in ADIPOQ and CVD. The findings of this study suggest that the rs2241766G allele and rs266729G allele increase odds of CVD, while the rs1501299T allele decreases. Rs2241766 and rs1501299 are significantly associated with CHD. However, the results were nearly “crude” ones, other important risk factors for CVD or CHD might diminish the significance of the results, so the associations were significant but weak. Nevertheless, the magnitude of these associations is in the range of all positive associations found with SNPs in multifactorial polygenic disorders, even with the “top ten” SNPs from GWAS.Qi et al. have conducted a meta-analysis to summarize the association between rs1501299 and CVD risk among diabetic patients . 827 CVDThe findings of our study may shed light on the underlying disease mechanism in CVD. Previous studies and meta-analysis demonstrated that rs1501299T carriers had higher adiponectin levels ,59, whicSeveral potential limitations of our study should be noted. First, significant between-study heterogeneities were observed in our study, we tried to explored the source of heterogeneities with factors such as ethnicity, study subjects, sample size, genotyping methods in the subgroup analyses. However, for rs2241766 and rs1501299 polymorphisms, significant heterogeneities still remained, only the heterogeneity for rs266729 disappeared by performing subgroup analyses according to ethnicity. Second, as significant publication bias was observed in the analysis of rs2241766, the association for this SNP still needs to be confirmed in future studies. Third, the eligible studies in our research were mainly from Asia and Europe, data of other populations, like African, was limited. Fourth, because we did not have access to individual data, we could not control for population stratification, nor could we adjust for variables in possible intermediate pathways. Finally, we should realize that the results might be distorted by potential weakness and biases of genetic association studies, such as genotyping error, phenotype misclassification, population stratification, gene-gene or gene-environment interactive effect, and selective reporting biases .Further studies should be conducted to confirm the functional variants. More studies are needed to elucidate the complete range of the signal transduction pathways that the variant is implicated in, and thus, throw light in the underlying molecular mechanisms that confer susceptibility to CVD. The particular polymorphism associated with CVD itself may not play a functional role, but rather it may be located physically close to the actual disease-predisposing gene.In addition to the genetic markers described in this meta-analysis, many other polymorphisms have been studied. Those polymorphisms may be candidates for a multivariate analysis. Future haplotypic approaches and further haplotype-based meta-analyses will provide more powerful and informative results than current single genotype-based data.The present meta-analysis showed that the associations between rs2241766, rs1501299 and rs266729 in the ADIPOQ and CVD were significant but weak. The rs2241766G allele and rs266729G allele increase risk of CVD, while the rs1501299T allele decreases. High quality studies are still needed to confirm the associations, especially for rs2241766.ACI: atherothrombotic cerebral infarction; ADIPOQ: adiponectin gene; CAD: coronary artery disease; CHD: coronary heart disease; CVD: cardiovascular disease; CI: confidence interval; HWE: hardy-Weinberg equilibrium; MAF: Minor Allele Frequency; MetS: Metabolic syndrome; MI: myocardial infarction; NA: Not available; OR: odds ratio; RA: Rheumatoid arthritis; RD: Renal disease; T2D: Type 2 diabetes.The authors declare that they have no conflict of interest.HZ and DFG conceived of the study, and participated in its design and coordination. HZ, XBM and YCH carried out the literature searching and data extraction, independently. XBM and HZ performed the statistic analysis and draft the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2350/13/40/prepubTable S1. Characteristics of the eligible studies included in the meta-analysis.Click here for fileTable S2. Main results of each study included in the meta-analysis. Click here for fileTable S3. Quality assessment for each study included in this meta-analysis. Click here for file


An insertion/deletion (I/D) variant in the angiotensin-converting enzyme (ACE) gene was associated with ACE inhibitor (ACEI)–related cough in previous studies. However, the results were inconsistent. Our objective was to assess the relationship between the ACE I/D polymorphism and ACEI-related cough by meta-analysis and to summarize all studies that are related to ACE I/D polymorphism and ACEI-cough and make a summary conclusion to provide reference for the researchers who attempt to conduct such a study.Databases including PubMed, EMbase, Cochrane Library, and China National Knowledge Infrastructure, were searched for genetic association studies. Data were extracted by two independent authors and pooled odds ratio (OR) with 95% confidence interval (CI) was calculated. Metaregression and subgroup analyses were performed to identify the source of heterogeneity.p = 0.46) in the dominant model and 1.61  in the recessive model. Heterogeneity was found among and within studies. Metaregression indicated that the effect size was positively associated with age and negatively associated with follow-up duration of ACEI treatment. Subgroup analysis revealed a significant association between ACE I/D polymorphism and ACEI-related cough in studies with mean age >60 y, but not in studies with mean age ≤60 y. No heterogeneity was found within each mean age subgroup. We also found no association between ACE I/D polymorphism and ACEI-related cough in studies with follow-up>2 mo or in studies in Caucasians. No heterogeneity was detected in these two subgroups.Eleven trials, including 906 cases (ACEI-related cough) and 1,175 controls, were reviewed in the present meta-analysis. The random effects pooled OR was 1.16 (95%CI: 0.78–1.74, Synthesis of the available evidence supports ACE I/D polymorphism as an age-dependent predictor for risk of ACEI-related cough. Angiotensin-converting enzyme inhibitors (ACEI) have been used for 30 years to treat cardiovascular diseases. In addition to their antihypertensive effects, ACEI are now widely used for congestive heart failure, acute myocardial infarction, and diabetic nephropathy The pathogenesis of ACEI-related cough remains unclear. It has been speculated that a local accumulation of bradykinin may play a major role in ACEI-related cough. This accumulation may lead to the activation of proinflammatory peptides  and local release of histamine in the airways An insertion/deletion (ID) polymorphism of the human ACE gene on chromosome 17q23 comprising the presence or absence of a 287 base pair (bp) insert in intron 16 was described by Rigat et al. in 1990 http://www.ncbi.nlm.nih.gov/pubmed; 1950–March 2011), EMbase , China National Knowledge Infrastructure (CNKI) , the Cochrane Library (http://www.cochrane.org), and reviews and reference lists of relevant articles. The last search was performed in March 2011.Using the term “ACE” or “ACEI” paired with “cough” and “polymorphism” without language restriction, we systematically searched PubMed (We selected studies as follows: (1) unrelated case-control studies and cohort studies; (2) complete data with genotype and allele frequencies; (3) genotype frequency of cases and controls is within Hardy–Weinberg equilibrium; and (4) studies in which the ID polymorphism has been determined. All of the literature searches were independently reviewed by two of the authors (Ya-Feng Li and Yun-Fei Bian).We extracted the following information from each study: first author, journal, year of publication, study population, demographics, number of cases and controls for ID genotype, type and dosage of ACEI, and follow-up time. The frequencies of the alleles and the genotypic distributions were extracted or calculated for both the cases and the controls.Data from the meta-analysis were analyzed using Stata software  and REVMAN software . The significance of the association for the dominant model (II/ID versus DD genotypes), the recessive model (II versus ID/DD genotypes), and the allele contrast I versus D were evaluated for 11 studies separately. All associations were indicated as odds ratios (ORs) with 95% confidence interval (CI). The pooled ORs were estimated using random effects (RE) and fixed effects (FE) models based on the individual Ors 2I statistic, which yields values between 0% and 100%, with higher values denoting greater degree of heterogeneity  Statistical heterogeneity between studies was formally tested with Cochran’s test. We also determined the Publication bias was assessed with the Egger regression test and represented graphically by use of Begg’s funnel plots of logOR compared with the standard error (SE) of logOR 2) test was applied to determine if genotype distributions of the control population reported conformed to Hardy-Weinberg equilibrium , and the study that the genotype distributions in the controls were significantly deviated from HWE was excluded from our sensitive analysis p values were two-tailed and p<0.05 was considered statistically significant.A chi-square . No difference was found in the distribution of genotype ID between Asians and Caucasians .p = 0.003) and 1.37 , respectively . To determine the sources of heterogeneity, metaregression and subgroup analyses were performed. Metaregression analysis of data showed that the mean age of hypertensives with ACEI-related cough and the follow-up duration of ACEI treatment contributed to the heterogeneity. The mean age was positively associated with the OR , and the duration of treatment was negatively associated with the OR . The type of ACEI, the proportion of males, and the study population were not effect modifiers.The RE and FE pooled ORs for the recessive model were 1.61 . The FE pooled ORs  were significantly different between the two mean age subgroups (p<0.05). In addition, when subgroups were categorized by the median of follow-up duration of ACEI treatment, heterogeneity was also reduced in both the shorter duration median subgroup (≤2 mo) and the longer duration median subgroup (>2 mo) . The FE pooled ORs of the two ACEI treatment duration median subgroups different significantly  (p = 0.0016).To further investigate the heterogeneity, we performed subgroup analyses . Other t2I = 0%; p = 0.485) and an FE pooled OR of 1.22  for the Caucasian studies. However, the seven Asian population studies were highly heterogeneous  , and the FE pooled ORs were 1.06  and 2.39   . Therefop<0.001) .p = 0.46) and 1.15 , respectively, and a moderate heterogeneity was found  , but the heterogeneity in the other subgroup was still large  (p = 0.787) in studies with a >2 mo follow-up duration of ACEI treatment, and the FE OR was 0.85  in the Caucasian study population. Further, we divided the 11 studies into three subgroups: Asians with mean age of cases ≤60 y, Asians with mean age of cases >60 y, and Caucasians. In each subgroup, heterogeneity was reduced . The FE pooled ORs were 0.82 , 2.95 , and 0.85 , respectively  . We did 2 = 65%) . For theectively .2I = 64%, p = 0.002), and the RE and FE pooled ORs were 1.33  and 1.22  (p = 0.572) in studies with a follow-up duration of >2 mo and 0.99  in the Caucasian studies. We further divided the studies by population and mean ages. Among the three subgroups , no heterogeneity was detected , and the FE pooled ORs were 0.99 , 2.08 , and 0.99 , respectively  . No hete= 0.004) . The FE ectively .p = 0.102).A statistical analysis of the Egger test and funnel plots was performed for all 11 studies . No publOur meta-analysis of 11 studies revealed that a greater I allele frequency gave rise to an increased susceptibility to ACEI-related cough. However, heterogeneity was detected in the 11 studies. The meta-regression analysis indicated that the effect size was positively related to the mean age of the subjects and negatively related to the follow-up duration.In the subgroup analysis, the association depended on three variables: (1) Age: a strong association between the ACE I/D polymorphism and ACEI-related cough was only identified in studies where the mean age was >60 y, not in studies where the mean age was ≤60 y. (2) Follow-up duration: no significant association existed between the ACE I/D polymorphism and ACEI-related cough when the follow-up duration was >2 months. (3) Ethnicity: we did not find any association between the ACE I/D polymorphism and ACEI-related cough in studies on Caucasian populations. This may have resulted from differences in age among studies, because the mean age of patients in two of the Caucasian groups was <57 y The mechanism for ACEI-related cough was not entirely clear. We believe that the lower level of tissue ACE (which correlates with a lower level of blood ACE in the patient) leads to higher concentrations of ACE substrates (both bradykinin and substance P), which may contribute to cough development. Bradykinin, a peptide degraded by ACE, has been suggested to play a major role in ACEI-related cough et al. speculated that ACEI induced a slow, marked reduction in ACE activity, but the loss of activity might be compensated by other peptide-degrading pathways. For example, neutral endopeptidase (NEP), which has been shown to regulate substance P and cough, was one of the candidates for these compensating pathways Some studies indicated that the ACE I/D polymorphism was associated with cough sensitivity, rather than ACEI-related cough (cough sensitivity was defined as a cough reflex response to stimulation with distilled water) Based on dominant and recessive gene models, we estimated the effects of an ACE gene polymorphism on ACEI-related cough. We found that the I allele of the ACE gene increased the susceptibility to ACEI-related cough. However, due to heterogeneity among the 11 studies, we performed a meta-regression analysis. This indicated that the effect size was positively related to the mean age of the subjects and negatively associated with the follow-up duration after ACEI treatment. The meta-analysis results also supported the notion that an ACEI-induced reduction in ACE activity may lead to bradykinin accumulation, which may contribute to ACEI-related cough. However, when the ACEI treatment is long in duration, bradykinin could be degraded by alternative metabolic pathways, and the ACEI-related cough might eventually cease or become tolerable. In some long-term studies, subjects that had recovered from an ACEI-related cough due to delayed bradykinin degradation may have subsequently been used as controls. This could have confounded the association between the ACE I/D polymorphism and ACEI-related cough.It is not clear why the association between the ACE I/D polymorphism and ACEI-related cough was more significant in subjects >60 y compared to those ≤60 y. It is known that older subjects have an age-related attenuation of the cough reflex; this may be related to an attenuation of bradykinin metabolism Our meta-analysis may also provide some guidance in the clinical use of ACEIs. Young patients with ACEI-related cough may be advised to adhere to ACEI treatment for 2 or 3 months to observe whether the cough ceases spontaneously. In cases where the cough ceases or becomes tolerable, the patient can continue ACEI therapy. In older patients with ACEI-related cough, it would also be advisable to adhere to ACEI treatment for a set period of time. This is because ACE activity directly affects the cough reflex in older patients, and this effect was negatively associated with pneumonia risk During our manuscript revision, we noted that Dr. Nishio’s group performed a similar meta-analysis Some limitations of the present study should be mentioned. First, although we excluded six studies that were not within Hardy–Weinberg equilibrium, the studies that we included had inconsistent designs. The categories and dosages of ACEIs varied among studies, and the follow-up duration ranged from 0.5 to 36 months. Second, the sample sizes were relatively small, and only five studies were age and sex matched. Thus, the results of the present meta-analysis should be interpreted with some caution. Third, some misclassification in case-control studies may have occurred. Some subjects that could tolerate a slight ACEI-related cough may have been classified as controls rather than cases.The present study indicated that the ACE I/D polymorphism may be a genetic, age-dependent risk factor for ACEI-related cough. Large case-control studies should be performed to verify the conclusions of our meta-analysis. Future investigations aimed at elucidating the age-dependence of the association between the ACE I/D polymorphism and ACEI-related cough may help to clarify the mechanism of ACEI-related cough.Figure S1PRISMA 2009 Flow Diagram.(TIF)Click here for additional data file.Table S1PRISMA Checklist.(DOC)Click here for additional data file.


We describe a novel two-stage method for the meta-analysis of percentile ratios that avoids distributional assumptions at the study level. Copyright © 2012 John Wiley & Sons, Ltd.Methods for individual participant data meta-analysis of survival outcomes commonly focus on the hazard ratio as a measure of treatment effect. Recently, Siannis  Fiocco et al.et al.In the analysis of a single survival study, the hazard ratio is a commonly used measure of treatment effect and is therefore a natural quantity to consider when undertaking a meta-analysis. However, an assumption of proportional hazards is even more restrictive in meta-analysis because it is imposed on multiple studies. If the proportional hazards assumption does not hold for some studies, then the estimated hazard ratio depends on the length of follow-up in those studies, and meta-analysis may not be appropriate. Methods that relax the proportional hazards assumption, the majority of which focus on a measure or measures of treatment effect that are based on the hazard ratio, have been proposed. For example, Moodie et al.qk at percentile level k comparing the survival distributions of two groups is defined asIn a recent paper, Siannis k = 0.5 represents the expected ratio of times at which half of the population will fail in the treatment group compared with the control group. Similarly, for other percentile levels k, the percentile ratio is the expected ratio for the time at which 100k% of the population will fail in the treatment group compared with the control group. Because the interpretation of percentile ratios can be so easily explained, their use could lead to clearer, more practical understanding of survival differences between treatment groups. In general, percentile ratios may vary across percentile levels. If the survival distributions in question are accelerated failure time models, however, the percentile ratio q is constant across all values of kThe median ratio at et al. estimated percentile ratios using a one-stage parametric model with data at the individual study level being modelled using either accelerated failure time or proportional hazards distributions. In the simplest version of the model, accelerated failure time distributions were used to model the data at the study level. In this case, the combined percentile ratio q is constant across percentile levels and can be modelled using either fixed or random effects. The proposed framework is very general, however, and could be used to model any choice of distribution at the study level. This was illustrated using a combination of accelerated failure time and proportional hazards models.Siannis Motivated by the popularity of simple two-stage analyses, we propose an alternative, two-stage method for meta-analysis of percentile ratios, which in addition avoids all distributional assumptions in the first stage. In stage 1, we use Kaplan–Meier estimates of the survivor functions for the treatment and control groups to estimate percentile ratios and their variance–covariance matrix. In stage 2, percentile ratios are combined using either univariate or multivariate, random-effects meta-analysis  and its variance–covariance matrix is estimated for each study. In Section 2.2, we describe how estimated logPRs from several studies can be combined using multivariate meta-analysis.qi = T for each study i, where qik denotes the percentile ratio in study i at percentile level k, and its variance–covariance matrix Si. Methods for combining these estimates will be discussed in Section 2.2. To estimate qik, we first estimate the kth percentile in the treatment and control groups for study i, which we denote We focus here on the analysis of the data from a single study. The aim is to estimate a vector of logPRs i for the remainder of this section. We will assume throughout that censoring is non-informative, that is, it occurs independently of survival.To simplify the notation, we will ignore study index tk at percentile level k of a distribution of survival times is defined by the equation S(tk) = k, where S is the survivor function. We estimate tk from a Kaplan–Meier estimate of the survivor function The percentile t, this corresponds to the value of t for which a horizontal line at S(t) = k crosses the estimated survival curve. If there is an interval t1≤ t < t2 for which S(t) = k exactly, then the percentile is estimated to be the midpoint of that interval:where the minimisation in  is neceskth percentiles in the treatment and control groups, respectively. Then, an estimate for the logPR is given by substituting the estimates Suppose Equation .S for quantile estimates is not straightforward. We describe two methods, the first of which is based on asymptotic approximations and is relatively fast to compute. The second method uses bootstrapping but is computationally more intensive.Sander A simple asymptotic expression for the variance of an estimated log-percentile f(t) is the probability density function of the survival distribution (see Appendix A.1 for a derivation). To estimate f(t) replaced by suitable estimates. f(t) is less straightforward but can be carried out using the presmooth package in R where Equation  with  anHaving estimated the variances of both treatment and control group log percentiles, an estimate of the variance of the estimated logPR is given byAn asymptotic expression to estimate the covariance between two estimated log percentiles k1 = k2, Equation (S(t)}and f(t) can again be estimated using Greenwood's formula and the presmooth package, respectively. The estimated covariance matrix S between two logPRs is then given by(see Appendix A.2 for a derivation). When Equation  reduces Equation . Var{S b b, alongi, log qik, is then assumed to be normally distributed about the combined logPR qk with between-studies variance The true logPR in study If a vector of logPRs from each study are to be combined, this can be performed either by using a separate univariate meta-analysis at each percentile level or by combining logPRs at all levels simultaneously using multivariate meta-analysis. It has been argued that multivariate meta-analysis is preferable when combining multiple correlated outcomes because estimates of combined effect sizes borrow strength across outcomes through the correlations between them qi is a vector of true, underlying logPRs in study i. Si is the within-study variance–covariance matrix from study i, which is again assumed fixed and known. The within-study correlation for a given study arises from the use of data from the same set of participants in the estimation of percentile ratios for that study. For random-effects meta-analysis, the vectors of true logPRs are then assumed to be distributed multivariate normally about a vector of combined logPRs log q,where logΣ is the between-studies variance–covariance matrix, to be estimated from the data. Σ comprises the between-studies variances and the between-studies correlation. The between-studies variances measure the variation in the true effect sizes across studies, equivalent to where stata, which uses restricted maximum likelihood (REML) to estimate log q and ΣK variance parameters and the K(K− 1) / 2 correlation parameters contained in the matrix Σ. The number of percentile levels for which estimation is possible is therefore limited by the number of studies in the dataset. The number of parameters to be estimated in Σ could be reduced by imposing constraints, but options to do this within the mvmeta package are currently limited.The model given by  and 13)13) can bIn this section, we present the results of a simulation study designed to assess the performance of stage 1 of the two-stage method. Bivariate meta-analysis has already been investigated in simulation studies μ = 4, scale parameter σ = 0.3 and log hazard ratio θ = − 0.4. The baseline parameter values were chosen to be similar to parameter estimates for the example dataset of Section 4. The hazard ratio was chosen to represent a moderately beneficial effect of treatment. For simplicity, no censoring was assumed to take place, but results should be comparable to censored datasets with a similar numbers of events.Data were simulated from a Cox proportional hazards model with a log-logistic baseline distribution with location parameter Simulated data were generated for studies with 20, 100 and 500 participants, with equal numbers in each treatment arm, and percentile ratios were calculated for percentile levels 0.9, 0.7, 0.5, 0.3 and 0.1 . Because of the absence of censoring in these simulated datasets, estimated survival probabilities were always equal to the chosen percentile levels for some time interval. Percentile estimates were therefore always calculated using Equation . To inveFor asymptotic variance estimation, the presmoothing method for estimation of the survival density function requires two bandwidths to be specified, as explained in Appendix B. The ‘presmooth’ package offers two built-in options for bandwidth selection, ‘plug-in’ and ‘bootstrap’. However, we found these to be unreliable for some simulated datasets, giving undersmoothed density function estimates that were inappropriately close to zero. For the simulation study, we instead fixed the bandwidths for all datasets to be the median of the bandwidths selected by the ‘plug-in’ method for 50 simulated datasets for each treatment arm in each study size.n = 18 and 20. For the smaller sample sizes, the estimated bias is greatest for the lower percentile levels, rising to 0.086 for n = 18 and 0.046 for n = 20 at k = 0.1. The true logPR at k = 0.1 is 0.362.From the simulations study results (not presented here), the bias in the estimated logPRs is estimated to be less than 0.01 for all percentile ratios for all sample sizes except the smallest, Results for the estimated coverage probabilities are given in We illustrate our method using a dataset consisting of nine studies examining the post-surgery treatment of patients with high-grade glioma. Patients in the treatment groups underwent a post-surgery course of radiotherapy and chemotherapy, whereas in the control groups, patients were treated with radiotherapy alone. The original meta-analysis contained 12 studies. Hazard ratios were estimated by log-rank statistics and combined using fixed-effects meta-analysis, giving a pooled hazard ratio of 1.18  et al.The data were investigated for departures from the proportional hazards assumption of Siannis et al.These data were also analysed using the accelerated failure time models of Siannis k = 0.9, …, 0.2 using the two-stage method of Section 2. There were insufficient data available to estimate logPRs at k = 0.1. Variance–covariance matrices were calculated for each study using both asymptotic and bootstrap methods.We estimated logPRs for each study in the glioma dataset at For the asymptotic variance estimation method, bandwidths were selected using the plug-in option as a default. To ensure that appropriate bandwidths had been selected, estimated density functions were plotted for each treatment arm in each study. For the control arms of studies 9 and 19, the estimated density functions appeared to be undersmoothed. We then used the bootstrap bandwidth selection option as an alternative. If the estimated curves still appeared to be undersmoothed, we then multiplied the bootstrap bandwidth by successive integers until the estimated curves appeared smooth.k = 0.2 could not be estimated for 140 out of 1000 bootstrap samples and for four out of 1000 at k = 0.3. We therefore calculated logPRs only at k = 0.9, …, 0.3, discarding the small number of samples at k = 0.3 for which logPRs could not be calculated.When applying the bootstrap variance estimation method, we were unable to calculate the logPR at lower percentile levels for some bootstrap samples when Kaplan–Meier estimates of survival curves did not decrease below a survival probability of 0.2. Removing all bootstrap samples where this is the case may bias the variance estimate because those samples would tend to have higher survival rates in the treatment group and therefore larger differences between treatment groups. For study 9, the logPR at stata with unconstrained between-studies covariance matrix Σ. Parameter values for the multivariate meta-analysis model described in Equations stata witet al. using accelerated failure time models, which gave a combined percentile ratio of 1.18 . The widths of the estimated CIs are similar for all analyses, suggesting that the new method has similar power to detect non-null treatment effects compared with the standard method. The value of I2, which measures the proportion of the variation in the data due to heterogeneity We first present in k-values for logPRs combined using univariate meta-analysis in the left-hand side of k, whereas for the bootstrapped results, combined logPRs appear to be more constant over time. Estimated standard errors are also less variable when the bootstrap method is used. Results from the simulation study of Section 3 suggest that the bootstrapped results are more reliable. In general, CIs are wider at early k, when fewer events have taken place, and at later k, when fewer participants remain in the studies. Also presented in k = 3 (using the bootstrap method).We present results for the full range of To compare our results with a method based on time-varying hazard ratios, we re-analysed the data using Cox relative risk regression models with different treatment effects in years 1, 2, 3 and beyond of the studies. The combined hazard ratio was 0.84 for year 1 , 0.85 for year 2  and 0.86 for year 3 and beyond . For this example, both methods are in agreement, suggesting a small, beneficial effect of treatment, which is maintained over time.k = 0.8, …, 0.3 have been combined to avoid convergence difficulties when too many correlation parameters are estimated. The asymptotic and bootstrapped results both show similar trends to those observed in the univariate analyses. The multivariate standard error estimates are slightly higher than the corresponding univariate estimates, in contrast with expectations that the borrowing of strength among outcomes would lead to smaller standard errors. The larger standard errors in the multivariate case appear to be caused by higher estimates of the between-studies standard deviation . For the multivariate meta-analyses, logPRs at levels k = 0.7,0.6,0.5 and 0.4 were combined simultaneously. Results for combined median ratios are presented in k = 0.7 and 0.6 for those studies. Estimated standard errors in Our analysis of the glioma dataset suggests that there is limited benefit in the use of multivariate meta-analysis in this context. However, it may still be advantageous to use multivariate meta-analysis for some datasets. For example, if some of the studies had only sufficient follow-up to estimate a subset of the logPRs of interest, multivariate meta-analysis would enable the combination of logPRs that were missing in those studies to borrow strength from the logPRs that had been observed Meta-analyses of survival data usually focus on the estimation of constant hazard ratios. If an assumption of proportional hazards seems inappropriate, then models that incorporate time-varying hazards could be used. However, it may be preferable to consider alternative measures of treatment effect, which are more readily interpretable. We have proposed a novel, two-stage method for meta-analysis of survival IPD using percentile ratios. The advantage of our method lies in the avoidance of distributional assumptions at the study level, which makes it suitable to use when proportional hazards or accelerated failure time assumptions are inappropriate.Stage 1 of the proposed meta-analysis involves estimation of logPRs and their variance–covariance matrix from Kaplan–Meier estimates of the survivor function. We have presented two methods for estimation of the variance–covariance matrix, an asymptotic method and a bootstrap method. Results from the simulation study of Section 3 suggest that the bootstrap method has the superior coverage properties, and we therefore recommend its use for variance estimation. If bootstrapping is too computationally intensive, the asymptotic method could be used to estimate mid-range logPRs for sufficiently large studies.Σ to have, for example, an auto-correlated form; we leave this for future work.In stage 2, logPRs can be combined using either univariate or multivariate meta-analysis. When logPR estimates are available for all percentile levels of interest, we recommend the use of univariate meta-analysis because multivariate meta-analysis can lead to inflated estimates of between-studies variances. However, when some studies lack sufficient data to estimate all logPRs, then a multivariate meta-analysis may be more appropriate to allow combined logPR estimates to borrow strength across percentile levels. We have not yet explored the possibility of constraining the between-studies variance–covariance matrix Our method could be adapted to allow either parametric or non-parametric methods to be used in stage 1 of a two-stage meta-analysis. Where appropriate, study-level data could be modelled using parametric distributions, with the non-parametric approach being used when distributional assumptions appear unjustified. Alternatively, the non-parametric method could be extended to estimate other quantities of interest from survival curves, such as percentile differences, ratios of survival probabilities at a given time, or differences in survival probabilities. Further work would be required to investigate the asymptotic properties of these alternative measures of treatment effect.et al.et al.et al.et al.Our focus has been on methods for meta-analysis of IPD. Our approach could also be used in the meta-analysis of aggregate data if methods were available for the extraction of IPD from published survival curves. To reconstruct the individual patient data, a model would be required for the censoring mechanism. Parmar


To examine the relationship between nonarteritic anterior ischemic optic neuropathy (NAION) and genetic polymorphisms of enzymes influencing endothelial function.The subjects were 34 patients with NAION  and 102 controls . Genetic polymorphisms were investigated in three candidate genes associated with endothelial function: endothelin-1 (ET-1), angiotensin-converting enzyme (ACE), and methylenetetrahydrofolate reductase (MTHFR). The genotype distributions in the patients with NAION were compared with those in the controls.There were no significant differences in the genotype distributions of the ACE I/D and MTHFR C677T polymorphisms between the NAION and control groups , whereas the genotype distribution of the G/T (Lys198Asn) polymorphism of the ET-1 gene varied significantly between the groups (p=0.009). After adjusting for covariates, individuals with the TT genotype of the Lys198Asn polymorphism were more likely to develop NAION compared with those with the GG genotype .We found an increased prevalence of a G/T polymorphism of the ET-1 gene in patients with NAION. Our data suggest that this polymorphism may be an important risk factor in developing NAION in the Japanese population. Nonarteritic anterior ischemic optic neuropathy (NAION) is a disease caused by vascular insufficiency leading to optic nerve head ischemia, and characterized by acute, monocular, and painless visual loss . AlthougEndothelin (ET)-1 is a 21 amino acid peptide that is a potent vasoconstrictor with an important role in the pathophysiology of the vascular system. Previous studies have shown that increased plasma ET-1 levels are associated with acute myocardial infarction , essentiElevated serum homocysteine is another possible aggravating factor for vascular endothelial cell damage in NAION . A C/T pThe renin-angiotensin system has important roles in atherosclerotic and cardiovascular disease, with effects ranging from endothelial dysfunction to vascular occlusion. Angiotensin-converting enzyme (ACE) governs conversion of angiotensin (Ang) I to Ang II and degradation of bradykinin. An insertion/deletion (I/D) polymorphism of the ACE gene (presence or absence of a 287 base pair insert in intron 16) significantly influences circulating and tissue ACE levels and results in greater production of Ang II . This maThe relationship between NAION and genetic polymorphisms in enzymes influencing endothelial function is incompletely understood. In this study, candidate genes were selected based on published evidence of involvement in risk factors of NAION or suspected biochemical contribution to a disease pathway. We examined genetic polymorphisms of ET-1, ACE, and MTHFR in patients with NAION. These polymorphisms have been reported to be associated with ocular vascular diseases.Written informed consent was obtained from all subjects. The protocol was approved by the ethics committee at our institution, and the study was performed in accordance with the ethical standards laid down in the Declaration of Helsinki. The subjects were 34 patients with NAION . NAION was diagnosed by neuroophthalmologists following the criteria of the Ischemic Optic Neuropathy Decompression Trial Research Group : 1) acutSubjects were defined as having diabetes mellitus if they were receiving treatment for insulin- or non-insulin-dependent diabetes mellitus. Hypertension was defined based on a history of hypertension with intake of antihypertensive drugs. Seven patients took these drugs at bedtime. Subjects were classified with hyperlipidemia if they were currently being treated for hyperlipidemia. The definition of smoking status included past and current smokers.Venous blood was collected from all subjects. The whole blood samples (5 ml) from all the patients were collected in EDTA tube and stored at -20°C for fewer than one month storage. Genomic DNA was isolated from peripheral leukocytes according to a kit protocol . Polymorphisms of the ET-1, ACE, and MTHFR genes were examined. Genotypes were determined with a fluorescence- or colorimetry-based allele-specific DNA primer assay system  . PolymorTo confirm the accuracy of genotyping with this method, DNA samples were subjected to PCR, followed by restriction-fragment-length polymorphism analysis or direct DNA sequencing of the PCR products. In each instance, the genotype determined with the allele-specific DNA-primer-probe assay was identical to that determined using the confirmatory methods.Statistical analysis was performed using STATA 8.0 . Logistic regression analysis was used to evaluate the independent effect of ET-1 mutations on the occurrence of NAION by adjusting for covariates such as diabetes mellitus, hypertension, hyperlipidemia, and smoking status. The ET-1 mutations and other covariates were coded with indicator variables. The criterion for significance was p<0.05.The distribution of the three polymorphisms of ET-1, ACE I/D, and MTHFR in the case and control groups was under Hardy–Weinberg equilibrium. The genotype and allele frequencies of the ET-1 gene polymorphisms are shown in Multivariable logistic regression analysis showed that the TT genotype of the ET-1 G/T polymorphism emerged as a significant risk factor for NAION after adjustments for age, diabetes mellitus, hypertension, hyperlipidemia, and tobacco smoking . The oddThe genotype and allele frequencies of the ACE and MTHFR gene polymorphisms are shown in In this study, we found a significant association between the G/T polymorphism of the ET-1 gene and the occurrence of NAION in Japanese subjects. To the best of our knowledge, this is the first report of a positive association between an ET-1 gene polymorphism and NAION. In a previous study, we showed that patients with NAION have higher circulating levels of ET-1 than controls . This reAnalysis of a multifactorial disease requires that several coexisting and risk-modifying factors be taken into account. Hypertension, diabetes mellitus, and tobacco smoking have been related to NAION ,10, and In contrast to the ET-1 polymorphism, we found no positive association of development of NAION with the I/D polymorphism of the ACE gene or the C677T polymorphism of the MTHFR gene. Many reports have shown that the ACE I/D polymorphism is significantly associated with subclinical structural arteriolar changes , diabetiAn important limitation of our study is the small number of cases. However, the distribution of the G/T genotype in the Japanese patients in our study is similar to the distribution found in previous studies ,32 in a


Studies on the risk of chromosomal abnormalities in early spontaneous abortion after assisted reproductive technology (ART) are relatively controversial and insufficient. Thus, to obtain a more precise evaluation of the risk of embryonic chromosomal abnormalities in first-trimester miscarriage after ART, we performed a meta-analysis of all available case–control studies relating to the cytogenetic analysis of chromosomal abnormalities in first-trimester miscarriage after ART.Literature search in the electronic databases MEDLINE, EMBASE, and Cochrane Central Register of Controlled Trials (CENTRAL) based on the established strategy. Meta-regression, subgroup analysis, and Galbraith plots were conducted to explore the sources of heterogeneity.A total of 15 studies with 1,896 cases and 1,186 controls relevant to the risk of chromosomal abnormalities in first- trimester miscarriage after ART, and 8 studies with 601 cases and 602 controls evaluating frequency of chromosome anomaly for maternal age≥35 versus <35 were eligible for the meta-analysis. No statistical difference was found in risk of chromosomally abnormal miscarriage compared to natural conception and the different types of ART utilized, whereas the risk of fetal aneuploidy significantly increased with maternal age≥35 .ART treatment does not present an increased risk for chromosomal abnormalities occurring in a first trimester miscarriage, but incidence of fetal aneuploidy could increase significantly with advancing maternal age. Assisted reproductive technology (ART) has been an important therapy method and a basic technique in many infertile couples to have children. Although the ART pregnancy rate of multiple Infertility treatment centers is stable at around 40%, the take home baby rate is still 20–30%, one of the important reasons is the high rate of early spontaneous abortion Cytogenetic analysis of products of conception (POC) is essential to examine the cause of the spontaneous abortion. Multiple cytogenetic analysis have indicated aneuploidy rates of first trimester miscarriages ranging from 50%–80% in various populations. The abnormal karyotypes from of cytogenetic studies include autosomal trisomies, sex chromosome monosomy, triploidy, double trisomies, polyploidies, as well as structural rearrangements. Among these autosomal trisomies are the most common chromosomal abnormalities There are adverse factors of advanced maternal age, altered karyotype, multiple assisted reproductive technologies (ART) failure, repeated miscarriages, spermatozoa obtained by Mesa-Tese reported, which lead to an increased risk of embryonic chromosomal abnormalities A number of studies have been conducted to investigate the pregnancy outcomes after ART, and the relationship between risk of chromosomal abnormalities occurring in first-trimester miscarriage and ART treatment, but their results are somewhat controversial and underpowered. With regards to whether an elevated risk of chromosomal abnormalities resulting in a first trimester miscarriage after ART, to the best of our knowledge, no meta-analyses on this issue have ever appeared. To obtain a more precise evaluation of the association between embryonic chromosomal abnormalities and risk of first-trimester miscarriage after ART, we performed a meta-analysis of all available case–control studies relating the cytogenetic analysis of chromosomal abnormalities to risk of first-trimester miscarriage after ART.To select eligible studies, a search was performed in the electronic databases MEDLINE, EMBASE, and Cochrane Central Register of Controlled Trials (CENTRAL), using the search strategy depended on various combinations of the keywords “assisted reproductive technology, ART or intracytoplasmic sperm injection, ICSI or in vitro fertilization-embryo transfer, IVF-ET” and “chromosomal abnormalities or cytogenetic analysis” and “first trimester miscarriages or spontaneous abortion or pregnancy loss ” only permitted the articles published in English. The last search was updated on April 01, 2013. And then reference lists of the relevant studies were also examined and the literature retrieval was completed independently by two reviewers (Lihong Pang and Junzhen Qin). Discordance was settled by consultation of a third reviewer (Mujun Li). When a study reported the outcomes on different subtypes of ART, we considered it as separate studies in the meta-analysis.The main searching strategy identified 229publications, 64 publications were excluded because of duplication . Manual Data extraction was conducted independently by two reviewers (Li-Hong Pang and Jun-Zhen Qin) from all included studies. Information was summarized as follow: the first author's name, publication date, study period, matching criteria, pregnancy loss definition, source of controls, the way of POC received, cytogenetic analysis obtained, method of conception, cytogenetic analysis and ratio of cases to control. The two reviewers examined the data extraction results and reached an agreement on all of the data extracted. If inconsistent opinion appeared on the data extraction results, a third reviewer (Xing Zhou) was invited to take part in discussion till the problem solved. Try best to communicate with the authors when data incomplete. Major characteristics are summed up in http://www.Iri.ca/programs/ceu/oxford.htm). The total scores ranged from 0 (lowest) to 9 (highest), and a study with scores≥6 was considered a high-quality study, whereas studies with scores<6 were classified as low-quality studies.Quality of the studies was determined with the use of the partially validated Newcastle-Ottawa Quality Assessment Scale: Case-Control Studies  or I2>50%, indicating the existence of statistically significant heterogeneity, a random-effects model (the DerSimonian and Laird method) was used to combine the data 2 tests >0.1 and I2<50%, showing the absence of heterogeneity, a fixed-effects (the Mantel–Haenszel method) was used Odds ratios (OR) with 95% confidence intervals (CI) for dichotomous data were used to assess the risk of chromosomal abnormalities in different compared models: ART versus natural pregnancy (NP); ICSI versus NP; IVF versus NP; IVF versus ICSI. The statistical heterogeneity among studies was detected by χAccording to the inclusion and exclusion criteria, 15 studies relevant to the risk of chromosomal abnormalities resulting in a first trimester miscarriage after ART were identified  [17]–[30A total of 3278 patients with first trimester abortions were included in our analysis cytogenetic analyses. Cytogenetic analysis of products of conception (POC) indicated that 1603(48.9%) were karyotypic abnormalities, and among them 1143 (71.3%) were autosomal trisomy, 118 (7.3%) were monosomy X, 62(3.8%) were structural anomalies, 54 (3.37%) were polyploidy, 39 (2.4%) were mosaic, 33 (2.1%) were double trisomies, and14 (0.87%) were tetraploidy. Therefore, autosomal trisomies represented the most common chromosomal abnormalities in spontaneous abortions from our data. 2: P = 0.092, I2 = 39.9%; 2: P = 0.67, I2 = 0.0%; 2: P = 0.125, I2 = 42.1%; 2: P = 0.318, I2 = 13.6%; The meta-analysis of the compared model for ART vs. natural pregnancy indicated that risk of chromosomal abnormalities resulting in a first trimester miscarriage after ART was not significantly increased , pregnancy loss diagnosis (Yes versus no), or quality score  were the major sources which contributed to heterogeneity. The three covariates were failed to relate with the ORs in the compared model of ART vs. NC , maternal age model≥35 versus<35 . Subsequently, subgroup analyses by quality scores, cytogenetic analysis of chorionic villi and pregnancy loss diagnosis indicated that heterogeneity still existed in ART vs. NP and maternal age≥35 vs. <35 comparison models.For the compared models of ART versus natural pregnancy and maternal age≥35 versus<35, the I2 values decreased lower than 50% and P values were greater than 0.10 . The summary ORs for compared models of ART vs. NC changed into significance after omitting the study of Kim et al. (fixed effects OR 0.73 (95% CI: 0.59–0.90), but the significance of summary OR for maternal age model was not influenced by excluding the study of Bingol et al.For further exploration of the heterogeneity we performed Galbraith plots analysis to identify the outliers which might contribute to the heterogeneity. Our results indicated that Kim et al. To prove reliability of the available evidence, sensitivity analyses were performed to examine the influence of the individual data-set to the pooled ORs by sequentially omitting a single study each time. But in our results, the corresponding pooled ORs of chromosomal abnormalities rate in a first trimester miscarriage were not materially altered (data not shown), demonstrating that our results were statistically robust. Begg's funnel plot and Egger's test were performed to assess the publication bias of literatures in all comparison models. The shapes of the funnel plots did not suggest any obvious asymmetry . Then, tFirst trimester miscarriage is the most common complication of pregnancies conceived through assisted reproductive treatment (ART), with embryonic chromosome anomalies accounting for approximately 50% of these losses Our meta-analysis results indicated that infertile couples with ART treatment did not have an increased risk of chromosomal abnormalities resulting in a first trimester miscarriage compared to those with natural conception, especially among the different types of ART using . However, when we excluded the study of Kim et al. With respect to compared model of maternal age≥35 vs. <35, 8 studies were found in our meta-analysis. Several studies have shown that increasing miscarriage and aneuploidy rates with increasing maternal age, and the main cause of this risk is considered to be errors arising at meisois I in the oocyte 2 values decreased lower than 50% and P values were greater than 0.10 after excluding the studies of Kim et al. Heterogeneity analysis of different compared models suggested significant heterogeneity in ART vs. NP and maternal age≥35 vs. <35. To explore the sources of heterogeneity, we performed meta-regression and subgroup analyses. Meta-regression analysis of maternal age data showed that the quality scores but not cytogenetic analysis of chorionic villi (Yes versus no), pregnancy loss diagnosis (Yes versus no) might substantially influence the initial heterogeneity, whereas this three covariates were failed to explain the significant heterogeneity in the compared model of ART vs. NC. Subgroup analyses by quality scores, cytogenetic analysis of chorionic villi and pregnancy loss diagnosis indicated that heterogeneity still existed in ART vs. NP and maternal age≥35 vs. <35 comparison models. To further investigate the heterogeneity, Galbraith plots analysis was performed to identify the outliers which might contribute most to the heterogeneity. Our results indicated that Kim et al. It should be noted that the present meta-analysis existed some limitations, caution is always required when interpreting the results. First, all of the data included in our meta-analysis were not based on individual adjusted ORs, individuals of some studies were not matched by maternal age and reproductive history within the case and control groups. To guarantee synthesis of the best available evidence, a more accurate evaluation should be adjusted by potentially suspicious factors, including maternal age, reproductive history, the number of embryos transferred and spermatozoa obtained. Second, the methods of ovarian stimulation were not uniformly defined in our analysis. Therefore, whether this confounding factor of ovarian stimulation may play a role on the risk of chromosomally abnormal miscarriages wan unclear. Third, the number of studies included in this study for compared models of ICSI vs. NP and IVF vs. NP were small. Forth, published bias may result from the unpublished data, and languages of the articles included in our meta-analysis only permitted published in English.In conclusion, the present meta-analyses t suggested infertile couples with ART treatment did not have an increased risk of chromosomal abnormalities occurring in a first trimester miscarriage compared to natural conception, and no statistical difference in frequency of chromosome anomaly among the different types of ART utilized, whereas the risk of fetal aneuploidy significantly increased with maternal age≥35. Although spontaneous abortion of ART may be a natural selection way of optimization to reduce the occurrence of birth defects, it not only decreases the successful rate of ART but also adds emotional pressure as well as economic burden to the infertile couples. Therefore, large sample studies with standardized unbiased ovarian stimulation methods, the number of embryos transferred, spermatozoa obtained and well matched controls for further investigation should be conducted. Such knowledge may help better preventing the occurrence of early spontaneous abortion, taking intervention measures to birth defects on time, and greatly increasing the take home baby rate.Checklist S1PRISMA checklist.(DOC)Click here for additional data file.


Plasma levels of adiponectin which is secreted from adipose tissue are associated with various parameters of metabolic syndrome. This effect seems to be a result of interactions between genetic and environmental factors including central obesity. The present study was carried out to investigate the possibility of relation between single nucleotide polymorphisms of adiponectin gene (+45 T/G and −11391 G/A) and waist circumferences (WC) in patients with type 2 diabetes.This cross-sectional study was conducted on n = 238 diabetic patients selected as cases and n = 159 as healthy control who were recruited from Rafsanjan city in south – east of Iran. The possible association of +45 T/G and –11391 G/A adiponectin gene polymorphisms with WC according to age and sex was evaluated.There was no significant difference in distribution of frequencies of +45 T/G and –11391 G/A adiponectin gene polymorphisms in each group. We only found a significant association between –11391 G/A adiponectin gene polymorphism with WC in diabetic group (p = 0.021). This association was remained significant after adjustment in multivariate regression model  and also this effect was independent of sex and age.We found higher abdominal obesity in GA or AA carriers of adiponectin – 11391 G/A genotype in type 2 diabetes patients independent of age and sex. Obesity and diet-dependent development of metabolic syndrome are considered as a global health problem ,2. AdipoThis was a case–control study. The case group included 238 patients with type 2 diabetes who were selected randomly among patients referred to Rafsanjan diabetes clinic. Control group were 159 healthy people who have been chosen from the same population. (The sample of the population is the same as previous studies from our group) . DiabeteAfter obtaining written consent from patients, a questionnaire comprising personal and demographic information for each participant was completed. Waist measurement was performed by a constant co-worker by measuring the widest area which is located between lower edge of lower rib and iliac crest and the unit was centimeter. Abdominal obesity was considered as a wais circumference over 90 cm. Waist circumference has been considered as an index for abdominal obesity. In this study, according to results of other studies which were performed on Iranian population, measures more than 90 cm were considered as abdominal obesity ,21,30-33DNA extraction and molecular analysis of adiponectin gene polymorphism: DNA was extracted from blood samples which were collected in tubes containing EDTA through salting out method. Molecular analysis of the +45 T/G polymorphism was conducted by using the report of Schaffer and his colleagues . For mol5Á-CATCAGAATGTGTGGCTTGC-3Á.5Á-AGAAGCAGCCTGGAGAACTG-3Á.The PCR product was cut by MspI restriction enzyme .Enzyme digestion lead to production of 137 and 26 base pairs . Digestion products were visualized on Agarose gel (%3.5) which was stained with ethidium bromide Statistical analysis.T-test, multivariate regression model, POST-Hoc, chi- square and Software version 15 called SPSS were used for data analysis ≤ 0.05 is considered as a data with statistically significance.In this study, We recorded characteristics of 397 participants of whom 238 subjects suffered from diabetes. Majority of the participants in diabetic group [170 (42.8%)] were females. But in non-diabetic group 36 (9.1%) of participants were female. Mean age of participants was 53 ± 10 and 50 ± 13 years in diabetic and non-diabetic group, respectively. The mean ± SD for WC in diabetics was 91 ± 9 cm, and in non-diabetics was 97 ± 11 cm. All of above analyses had significant p values.Genotype frequencies of +45 T/G or −11391 G/A for our groups was shown in Table We found significantly increased abdominal obesity in GA or AA carriers of adiponectin – 11391 G/A genotype in type 2 diabetes patients. This relation was independent of age and sex. In a study on French population the effects of adiponectin gene polymorphisms at positions  were examined with obesity and a strong relationship between adiponectin polymorphisms and severe obesity was observed. Furthermore, they found that haplotypes associated with obesity were related to elevated levels of plasma adiponectin, which indicates a relationship between hyper-adiponectinemia and weight gain . It has Previous studies suggest that the relation between incidence of type 2 diabetes and high blood pressure with adiponectin genes polymorphism is depends on the presence of obesity. It means that these effects are less observed in lean individuals who posses this polymorphism . HoweverFinally it seems that more studies in a larger population will be helpful to further confirm our Results. Also study of other common polymorphisms in adiponectin gene in Iranian population is recommended.The authors have no competing interests.SH-R was the principle investigator, conceived of the study, and wrote draft the manuscript. MMA was the principle investigator, participated in its design and coordination and wrote the draft of the manuscript. OT-M conceived of the study, helped to draft the manuscript and performed the statistical analysis. YR collected the data. JT-B conceived the study. HS collected the data. EA collected the data. All authors read and approved the final manuscript.


Helicobacter pylori (HP) infection status, pepsinogen I/II ratio, smoking, and alcohol. We further performed meta-analysis using the random effects model to redefine the relationship between coffee intake and peptic ulcer disease. The eligible 8,013 study subjects comprised of 5,451 coffee drinkers and 2,562 non-coffee drinkers. By univariate analysis, age, BMI, pepsinogen I/II ratio, smoking, and alcohol showed significant associations with coffee consumption. By multiple logistic regression analysis, positively correlated factors with significance were HP infection, current smoking, BMI, and pepsinogen I/II ratio for GU; HP infection, pepsinogen I/II ratio, and current smoking for DU; HP non-infection, male, BMI, pepsinogen I/II ratio, smoking, age, and alcohol for RE; younger age, smoking, and female for NERD. The meta-analyses could detect any association of coffee consumption with neither GU nor DU. In conclusion, there are no significant relationship between coffee consumption and the four major acid-related upper gastrointestinal disorders.Probably due to caffeine-induced gastric acid secretion, negative effects of coffee upon various upper-gastrointestinal diseases have been precariously accepted, despite the inadequate epidemiological evidence. Our aim is to evaluate the effect of coffee consumption on four major acid-related diseases: gastric ulcer (GU), duodenal ulcer (DU), reflux esophagitis (RE), and non-erosive reflux disease (NERD) based on the large-scale multivariate analysis. Of the 9,517 healthy adults, GU, DU, and RE were diagnosed by endoscopy, and NERD was diagnosed by the symptoms of heartburn and regurgitation without esophageal erosion. Associations between coffee consumption and the four disorders were evaluated, together with age, gender, body mass index (BMI),  Coffee is one of the most widely consumed beverages in the world; in particular, Japan is one of the biggest coffee markets in Asia Helicobacter pylori (HP) infection, obesity, smoking, alcohol drinking, etc. should be taken into consideration HP infection, which is an evident risk factor for peptic ulcer diseases HP infection, obesity, smoking, or alcohol drinking HP infection To evaluate the effect of coffee consumption on four upper gastrointestinal disorders precisely, effects of many causative factors such as Study participants were 9,517 adults who received a medical checkup at Kameda Medical Center Makuhari from October 2010 to September 2011. In this study, all the participants were asked to respond to the Frequency Scale for the Symptoms of GERD (FSSG) Gastric ulcer (GU) and duodenal ulcer (DU) were diagnosed by endoscopy. In the present study, only active ulcers were considered as GU or DU respectively. Peptic ulcer (PU) was defined as the presence of GU and/or DU. Reflux esophagitis (RE) was also diagnosed by endoscopy, according to the modified Los Angeles (LA) classification Helicobacter pylori antibody was measured using a commercial EIA kit . According to the manufacture’s instruction, the antibody titer above 10 U/ml was considered as HP-positive. Serum pepsinogen I and II were measured using a commercial LAR kit .Serum anti-Helicobacter pylori?”; iv) “Do you have a history of gastric surgery?”; v) “Are you taking proton pump inhibitors (PPIs) or histamine H2-receptor antagonists (H2RAs)?”; and vi) “How much coffee do you drink?”. The answers for question i) were selected from five classifications , which were further categorized into two groups as nominal variables: rarely drinking group (never or seldom) and usually drinking group . The answers for question ii) were categorized into two groups as nominal variables: current or past habitual smoking (smoker group), and lifelong nonsmoking (nonsmoker group). The answer for iii), iv), and v) were “yes” or “no”. The answers for question vi) were categorized into three groups as ordinal variables: drinking less than a cup of coffee per day, 1–2 cups of coffee per day, and 3 or more cups of coffee per day.The Frequency Scale for the Symptoms of GERD (FSSG) is a widely used questionnaire for diagnosis of GERD and also for evaluating the effectiveness of digestive drug treatment 2 statistics.The meta-analysis was conducted according to the PRISMA guidelines . Previou2RAs, and having past history of HP eradication were further excluded from the study population, since such background factors might adversely affect accurate analysis. In the present study, we used eight factors as explanatory variables: age, body mass index (BMI), gender, drinking habit, smoking habit, Helicobacter pylori infection status, ratio of pepsinogen I/pepsinogen II (PG I/II ratio), and coffee consumption.The association of candidate background factors with the four major upper-gastrointestinal acid-related diseases was evaluated by univariate and multivariate analyses using the JMP® 9 program . After subjects with missing values were omitted, subjects with prior gastric surgery, taking PPIs and/or HHP infection status were divided into two groups: smoker and nonsmoker; drinking and rarely drinking; HP-positive and HP-negative.We categorized age into five groups to apply a univariate analysis: <40, 40–49, 50–59, 60–69, and ≥70. BMI and PG I/II ratio were respectively categorized into three groups: <18.5 (underweight), 18.5–24.9 , and ≥25.0 (overweight) for BMI; <2.0, 2.0–2.9, and ≥3.0 for PG I/II ratio. Based on the above-mentioned criteria, smoking, alcohol drinking, and HP infection status, and coffee consumption were analyzed as ordinal or nominal variables. A p-value of less than 0.05 was considered significant.Univariate analyses were done using Pearson's chi-square test, Student's t-test, and Welch's t-test to evaluate association between coffee consumption and other background factors. In addition, multiple logistic regression analysis was applied for evaluating the relationship between the above four esophago-gastro-duodenal diseases and eight background factors respectively. Specifically, we applied firth's penalized-likelihood method to deal with issues of separability, small event sizes, and bias of the parameter estimates for GU and DU. Age, BMI, and PG I/II ratio were evaluated as continuous variables, whereas smoking, alcohol drinking, 2RAs (493), and a history of H. pylori eradication (900). Among the eligible 8,013 study subjects , 5,849 had neither PU nor GERD , intake of PPI and/or Hnor GERD . Among tnor GERD . Distribnor GERD , in whicHP infection status did not , positively correlated factors of gastric ulcer in order of significance are HP infection , current smoking , higher BMI , and higher PG I/II ratio . Coffee consumption as well as age, sex, and alcohol drinking did not show significant association with GU.For gastric ulcer, we compared GU patients (n = 43) with GU-free subjects . By multiple logistic regression analysis , BMI, PGHP infection status showed a significant association with DU. Judging from the value of standardized coefficients (β), positively correlated factors of duodenal ulcer in order of significance are HP infection , higher PG I/II ratio , and current smoking . Coffee consumption as well as age, sex, BMI, and alcohol drinking did not show significant association with DU.For duodenal ulcer, we compared DU subjects (n = 32) with DU-free subjects . By multiple logistic regression analysis , PG I/IIHP infection showed a significant association with RE. Judging from the value of standardized coefficients (β), positively correlated factors of reflux esophagitis in order of significance are HP non-infection , male gender , higher BMI , higher PG I/II ratio , current smoking , alcohol drinking , and former smoking . Among the examined variables, only coffee consumption did not show significant association with RE.For reflux esophagitis, the subjects with RE (n = 994) were compared with GERD-free subjects . By multiple logistic regression analysis , age, geHP infection status did not show significant association with NERD.For non-erosive reflux disease (NERD), the subjects with NERD  were compared with GERD-free subjects . By multiple logistic regression analysis , age, geTo get an overview of the accumulated reports concerning association between coffee consumption and the four upper gastrointestinal disorders, we performed meta-analyses using the random effects model. For peptic ulcer diseases, we found six case-control and four cohort studies fulfilling the inclusion criteria for meta-analysis . We havep for heterogeneity, 0.0008; I2, 78.9%). The meta-analysis of five case-control and three cohort studies also detected no significant association between coffee consumption and DU . For PU, we analyzed three case-control and five cohort studies . To sum up, the meta-analyses could not detect a significant association of coffee consumption with the upper gastrointestinal ulcer diseases.Meta-analysis was executed using a random effects model, because a test of heterogeneity was statistically significant. As shown in  studies , which ap-values less than 0.1 were considered statistically significant. In all meta-analyses we performed, significance tests of the asymmetry were not significant . We thence concluded that publication bias can be considered as nonsignificant.Publication bias of each meta-analysis was further assessed by a funnel plot and funnel plot regression, in which All the four upper gastrointestinal disorders examined in the present study have been considered as acid-related diseases For GERD, we also could not detect a significant association between coffee intake and the incidence of GERD (both RE and NERD), although some past study have reported that coffee intake may predispose to GERD syndrome One of limitations of our present study was of course a cross-sectional design, which should be precisely validated in the future prospective study. We are following the present large-scale cohort to validate our present conclusion in the upcoming trial. Another limitation of our study was lacking more detailed information of coffee, such as kinds of coffee beans, use of milk or sugar, regular coffee or not, the time of coffee drinking, etc. These minute data concerning coffee intake will be added to the future study, which will make our next research more accurate and polished to verify our present conclusion.Figure S1Flow diagram of the meta-analysis literature search results.(DOC)Click here for additional data file.Table S1Summary characteristics of cohort or case-control studies were included from the meta-analysis that compare the relationship of coffee and peptic ulcer.(XLS)Click here for additional data file.Table S2Summary characteristics of cohort or case-control studies were excluded from the meta-analysis that compare the association of coffee and peptic ulcer.(XLS)Click here for additional data file.Document S1References used in (DOC)Click here for additional data file.