CYP2E1) polymorphisms and the risk of alcoholic liver diseases (ALD) have yielded conflicting results. Thus, a meta-analysis was performed to clarify the association between CYP2E1 polymorphisms and the risks of ALD.Previous studies investigating the association between cytochrome P4502E1  when ALD patients were compared with alcoholics without ALD. Significant associations between CYP2E1 Pst I/Rsa I polymorphism and ALD risk were also observed in Asians  and in Caucasians  when ALD patients were compared with alcoholics without ALD. However, subgroup analysis stratified by ALD types showed that CYP2E1 Pst I/Rsa I polymorphism was not significantly associated with the risks of alcoholic cirrhosis (ALC). No significant association was observed between CYP2E1 Dra I polymorphism and ALD risks.A total of 27 and 9 studies were finally included for the association between the CYP2E1 Pst I/Rsa I polymorphism might be not significantly associated with advanced form of ALD (ALC), but might be significantly associated with other form of ALD such as steatosis, hepatisis, fibrosis. Furthermore, CYP2E1 Dra I polymorphism might be not significantly associated with the ALD risks. Since potential confounders could not be ruled out completely, further studies were needed to confirm these results.This meta-analysis suggested that  Alcoholic liver disease (ALD) remains to be one of the most common etiologies of liver diseases and is a major cause of morbidity and mortality worldwide CYP2E1 gene might be related with the risk of ALD in individuals.Cytochrome P4502E1 (CYP2E1) is a member of the phage I detoxifying enzymes, which plays important roles in the metabolic activation of many xenobiotics including alcohol. CYP2E1 is physiologically responsible for about 10% of ethanol metabolism, but it could be induced after chronic ethanol administration CYP2E1, located in 10q2403-qter, is a 1104 kb gene consisting of 9 exons and 8 introns. CYP2E1 contains six restriction fragment length polymorphisms (RFLP), among which the Pst I/Rsa I polymorphism in its 5′-flanking region was reported to be associated with higher transcription and increased enzyme activity Dra I in intron 6 is also known to be associated with increased expression and enzyme activity CYP2E1 gene polymorphisms and the risks of ALD have been widely investigated in the past 20 years. Unfortunately, these epidemiological studies have yielded conflicting results, which might be related with the ethnic difference in the distribution of the mutant alleles and the relatively small sample size in some studies underpowered to detect the potential effects. Considering the important roles of CYP2E1 in the etiology of ALD, it would be necessary to summarize all these individual studies and discern whether CYP2E1 gene polymorphisms are associated with the risks of ALD. Thus, we performed a meta-analysis in order to provide more accurate estimate of the association of the above gene polymorphisms and the risks of ALD.A computerized literature search was conducted for the relevant available studies from 3 databases including PubMed, ISI Web of Science, and Embase. The search strategy to identify all possible studies involved use of combinations of the following key words: (“cytochrome P450 2E1” or “CYP2E1”) and “polymorphism” and  and . The reference lists of review articles, clinical trials, and meta-analyses were also hand-searched to identify additional works. There was no restriction on time period, sample size, population, language, or type of reports. If more than one article were published using the same case series, only the study with largest sample size was selected. The literature search was updated to June 2012.CYP2E1 Pst I/Rsa I and/or Dra I polymorphisms and the risk of ALD; (2) providing sufficient data for calculation of odds ratio (OR) with the corresponding 95% confidence interval (95%CI). When genotype frequencies and OR with 95%CI were all not available, authors were contacted to request the relevant information. All identified studies were carefully reviewed independently by two investigators to determine whether an individual study was eligible for inclusion in this meta-analysis.The studies included must meet the following criteria: (1) evaluating the association between Data were extracted independently by two investigators who reached a consensus on all of the items. The following information was extracted from each study: (1) name of the first author; (2) year of publication; (3) country of origin; (4) ethnicity of the study population; (5) numbers of cases and controls; (6) gender and age of enrolled subjects; and (7) numbers of genotypes in cases and controls.CYP2E1 Pst I/Rsa I and/or Dra I polymorphisms and ALD risks were estimated by calculating pooled ORs and 95%CI. The comparisons were made between ALD patients and alcoholics without ALD, and between ALD patients and non-alcoholics without liver diseases , respectively. The significance of the pooled effect size was determined by Z test. χ2 analysis with exact probability was used to test departure from Hardy-Weinberg equilibrium (HWE) for the genotype distribution in control groups . Heterogeneity among studies was assessed by χ2-based Q test as well as the 2I statistic P < 0.10) indicated heterogeneity across studies. Subgroup analyses were performed based on the ethnicity of the enrolled subjects, the type of ALD, and the gender of subjects. Sensitivity analysis was undertaken by removing one individual study each time to check whether any of single study could bias the overall estimate I2 statistic. Data analysis was performed using STATA version 11 .The associations between CYP2E1 Pst I/Rsa I polymorphism and the risks of ALD, and CYP2E1 Dra I polymorphism and the risks of ALD, respectively. All these included studies used peripheral blood samples for DNA extraction and polymerase chain reaction-restriction fragment length polymorphism (PCR-RELP) methods for genotyping. These studies were performed in a wide range of geographical settings leading to a diversity of racial groups. For the Pst I/ Rsa I polymorphisms, 9 studies examined individuals of Asians, 15 studies recruited Caucasians, one study in Brazilian Dra I polymorphism, all the studies were on Caucasians. In these studies, the genotype/allele distribution between ALD patients and the alcoholics without ALD, and/or between ALD patients and non-alcoholics, were compared. The detailed characteristics of the included studies were shown in the  and , respectively. The detailed criteria for the selection of ALD patients, alcoholics without ALD, and non-alcoholics were shown in Table S1.The flowchart of the study selection for this meta-analysis was shown in the CYP2E1 Pst I/Rsa I polymorphism and the risk of ALD were shown in Table S1. Overall, no significant association between c2 allele and the risks of ALD was observed   when ALD patients were compared with alcoholics without ALD, which was not observed in other genotypes contrasts. In regard with the genotypes contrast between ALD patients and non-alcoholics, no significant association was detected in any genetic model .Results of pooled analysis on the associations between 92–2.04) Figure 2vs. c1c1: OR = 4.11, 95%CI 2.32–7.29), while significant associations were also observed in Caucasians  when ALD patient were compared with alcoholics without ALD. However, no significant association was also observed in Asians or Caucasians when ALD patients were compared with no-alcoholics (Table S2).Due to the ethnicity-related distinct discrepancy of allele distribution in non-alcoholics  Figure 3CYP2E1 Pst I/Rsa I polymorphism and the risks of clearly defined alcoholic liver cirrhosis (ALC). As shown in Table S3, no significant association was observed between CYP2E1 Pst I/Rsa I polymorphism and ALC risks in Asians  as well as in Caucasians  when ALC patients were compared with alcoholics without ALD. However, the polled results of the studies, in which cases were composed by several types of ALD patients including steatosis, hepatisis, fibrosis, and cirrhosis, showed significant associations in Asians  and in Caucasians  (Table S3).As ALD contains many types of histological forms, including steatosis, hepatisis, fibrosis, and cirrhosis, we then investigated the associations between CYP2E1 Pst I/Rsa I polymorphisms and ALD risks in men and women, we then combined the results of studies in which only male subjects were enrolled. Meta-analysis for the association between CYP2E1 Pst I/Rsa I polymorphism and the risk of ALD in male subjects were shown in Table S4. As shown in Table S4, no significant association was detected in any comparisons.To investigate whether there was difference in the association between CYP2E1 Dra I polymorphism and the risk of ALD were shown in CYP2E1 Dra I polymorphism and the risk of ALD . We further analyzed the relationship between CYP2E1 Dra I polymorphism and the risks of ALC. Again, the pooled results revealed no significant association (Table S5).Results of pooled analysis on the associations between CYP2E1 Pst I/Rsa I polymorphism and the risk of ALD in worldwide population (2>50% in most comparisonsI). Subgroup meta-analysis has been used as a common method for exploring the heterogeneity CYP2E1 Pst I/Rsa I or Dra I polymorphisms and the risk of ALD in Caucasians, which suggested that ethnicity might be an important contributor to heterogeneity. The heterogeneity test in the subgroup analyses based on the types of ALD showed that lower heterogeneity in the analysis of the association between CYP2E1 Pst I/Rsa I polymorphism and ALC risks, indicating that types of ALD might be another source of the between study heterogeneity. Sensitivity analysis was performed by sequential omission of individual studies in every comparison, and the data showed that no study significantly influenced the pooled effects by omitting any single study. For the comparisons between ALD patients and non-alcoholics, the exclusion of the studies deviated form HWE did not change the results significantly.The between-study heterogeneity was significant in the analyses of association between vs. c1, c1c2 vs. c1c1, and c2c2/c2c1 vs. c1c1  when ALD patients were compared with alcoholics without ALD in Asians. No significant publication bias was detected in any other comparisons .Begg’s funnel plots were generated to assess publication bias. The Egger’s test was performed to statistically evaluate funnel plot symmetry. Potential publication bias was detected by Egger’s test for the comparisons between c2 CYP2E1 functional polymorphisms including Pst I/Rsa I or Dra I polymorphisms might be associated with the ALD risks of individuals. Unfortunately, previous epidemiological studies conducted in the past 20 decades have yielded conflicting results, ranging from strong association to no links. Because of the above-mentioned conflicting results from relatively small sample size which might be underpowered to detect the potential effects, a meta-analysis might be an appropriate approach to obtain a more definitive conclusion.ALD is a multifactorial process involving several mechanisms, in which oxidative stress may play an important role CYP2E1 Pst I/Rsa I polymorphisms and the risk of ALD, and between CYP2E1 Dra I polymorphism and the risk of ALD, respectively. The pooled results showed that c2c2 genotype was significantly associated with increased risk of ALD in Asians (Table S2). These data were well consistent with many previous studies CYP2E1 Pst I/Rsa I polymorphism and the ALD risk was also found in Caucasians , although many pervious studies reported no association vs. 2.7% in Asians and Caucasians, respectively)  In the current study, we made a comprehensive literature search and a total of 27 and 9 studies were finally included for the analysis of the associations between the ctively) Figure 3et al. performed a meta-analysis to evaluate the association between the polymorphisms of ethanol-metabolizing enzymes including CYP2E1 Pst I/Rsa I polymorphism and the risk of ALD vs. c1c1, OR = 4.11, 95%CI 2.32–7.29) as well as in Caucasians  (Table S2). However, the subgroup analysis on the association between CYP2E1 Pst I/Rsa I polymorphism and ALC risks in the current meta-analysis showed similar results to those obtained in the study by Zintzaras et al.  (Table S3). These results suggested that CYP2E1 Pst I/Rsa I polymorphism might be not significantly associated with the ALC risks, but might be significantly associated with other types of ALD. In fact, ALD represents a spectrum of clinical illness and morphological changes that range from alcoholic fatty liver (AFL) to hepatic inflammation and necrosis  to progressive fibrosis (AF) and cirrhosis CYP2E1 Pst I/Rsa I polymorphism might be significantly associated with the earlier liver damages such as AFL and AH, but not with the advanced ALD (ALC), which need to be evaluated in the future studies. This meta-analysis firstly addressed the association between CYP2E1 Dra I polymorphism and the risk of ALD, but no significant association was found based on 9 studies performed in Caucasians.Zintzaras in vivo would be a more reliable method for the evaluation of CYP2E1 status, and may be helpful to evaluate the association of CYP2E1 polymorphisms and the risk of ALD. If subjects with c2c2/c1c2 genotypes have higher CYP2E1 activity, then a solid conclusion about the association between CYP2E1 Pst I/Rsa I polymorphism and ALD risk, would be more rational and reliable, as these persons would be more susceptive to ALD. However, this information is limited in the literature. A preliminary study on the elimination of acetaminophen, which was metabolized mainly by CYP2E1, suggested that individuals with homozygous c2c2 genotypes exhibited higher CYP2E1 activity than individuals with c1c1 or c1c2 genotypes CYP2E1 Pst I/Rsa I polymorphisms and the activity of CYP2E1 in vivo.One of the most characteristic features that distinguish CYP2E1 from other ethanol metabolizing enzymes is its inducibility. Some studies demonstrated that ethanol could prevent the decomposition of CYP2E1 protein without influencing the mRNA levels Despite clear strengths of our study including the larger sample size and comprehensive literature search, it dose have some limitations. Firstly, the present meta-analysis was based on unadjusted effect estimates and CIs, since most studies did not provide the adjusted OR and 95%CI controlling for potential confounding factors. Secondly, moderate to higher heterogeneity existed for the analyses especially for the subgroup of Asians. Thirdly, it has been well known that ALD is a multifactor diseases, however, the effects of gene-gene and gene-environment interactions were not addressed in this meta-analysis, and thus the potential roles of the above gene polymorphism may be masked or magnified by other gene-gene/gene-environment interactions.CYP2E1 Pst I/Rsa I and Dra I polymorphisms and the risk of ALD. The combined results showed that CYP2E1 Pst I/Rsa I polymorphism might be not significantly associated with advanced form of ALD (ALC), but might be significantly associated with other form of ALD such as steatosis, hepatisis, fibrosis. Furthermore, CYP2E1 Dra I polymorphism might be not significantly associated with ALD risks. Since potential confounders could not be ruled out completely, further studies are needed to confirm these results.In summary, this meta-analysis systematically analyzed the association between the Table S1The criteria for selection of cases and controls of included studies.(DOC)Click here for additional data file.Table S2CYP2E1 Pst I/Rsa I polymorphism and the risk of ALD.Meta-analysis for the association between (DOC)Click here for additional data file.Table S3CYP2E1 Pst I/Rsa I polymorphism and the risk of ALDSubgroup analysis on the association between (DOC)Click here for additional data file.Table S4CYP2E1 Pst I/Rsa I polymorphism and the risk of ALD in male subjectsMeta-analysis for the association between (DOC)Click here for additional data file.Table S5CYP2E1 Dra I polymorphism and the risk of ALDMeta-analysis for the association between (DOC)Click here for additional data file.PRISMA Checklist S1PRISMA Checklist.(DOC)Click here for additional data file.


There is no evidence for an association with gout in Caucasian populations. In the present study, we tested rs3733591 for association with gout in New Zealand (NZ) Māori, Pacific Island and Caucasian samples.The C allele of the nonsynonymous Arg265His  and non-gout controls . Further Caucasian sample sets consisting of 67 cases and 4,712 controls as well as 153 cases and 6,969 controls were obtained from the Framingham Heart Study and the Atherosclerosis Risk in Communities study, respectively. The Polynesian samples were analyzed according to Eastern and Western Polynesian ancestry.rs3733591 with gout was found in the sample sets of NZ Māori (odd ratio (OR) = 0.98, P = 0.86), Eastern Polynesians , Western Polynesians  or combined Caucasians . The C allele was significantly overrepresented in Māori tophaceous cases compared to cases without tophi , but not in the other ancestral groupings.No evidence for risk conferred by the C allele of rs3733591 with gout in Eastern Polynesian, Western Polynesian and Caucasian samples. However, consistent with a previous study of Han Chinese and Solomon Island populations, our data suggest that rs3733591 could be a marker of severe gout in some populations. Our results also suggest that the effect of this variant is population-specific, further confirming population heterogeneity regarding the association of SLC2A9 with gout.Noting that our study's power was limited for detecting weak genetic effects, we were unable to replicate associations of  Gout is a common form of inflammatory arthritis predominantly affecting men, with hyperuricemia being an essential predeterminant. As urate concentrations reach saturation in the blood, monosodium urate (MSU) crystals are deposited in the joints and tissues. An acute self-limiting inflammatory reaction to these MSU crystals leads to severe pain and debilitation (gout). Without resolution, the MSU crystals and subsequent inflammation can lead to chronic tophaceous gout, bony erosions and permanent disability. In New Zealand , gout is common in Māori and Pacific Island men, with the prevalence estimated to range from 9.3% to 13.9% and 14.9%, respectively . Renal urs7442295 and surrogate marker rs11942223) within the solute carrier family 2, member 9/facilitated glucose transporter 9 (SLC2A9/GLUT9) gene are associated with high serum urate concentrations and gout [SLC2A9 variant rs11942223 best explains the strong role that SLC2A9 played in the development of gout in NZ Māori, Pacific Island and Caucasian sample sets [SLC2A9 variants have gender-specific effects on serum urate, with the effect being stronger in women [Genomewide association studies in Caucasian cohorts have shown that intronic variants , contributes significantly to the development of elevated urate concentrations and gout in Han Chinese, Solomon Island and Japanese sample sets [rs7442295/rs11942223 variants. The measure of linkage disequilibrium, r2, is < 0.05 between rs3733591 and rs11942223 in HapMap Caucasian, Chinese and Japanese samples. Given the existing evidence for population heterogeneity in association of SLC2A9 variants (intronic and R265H) with gout [rs3733591) in gout in NZ Māori, Pacific Island and Caucasian case-control sample sets in a study adequately powered to detect an effect equivalent to that observed in other populations (odds ratio (OR) > 1.4).An additional ple sets ,13, but ple sets . Han Chiple sets . There aith gout ,11-13, wGenotyping of NZ sample sets of Māori (229 cases and 343 controls), Pacific Island (232 cases and 174 controls) and Caucasian (327 cases and 638 controls) was done  Offspring data set and combined with 17 gout cases from the Generation 3 data. Self-reported gout cases from the Offspring data set were included if the participants reported having gout on two or more survey occasions or reported having gout on one survey occasion and were also taking antigout medication. Self-reported gout cases from the Generation 3 data set were included if the participants had also answered "no" to taking medication for hypertension or high blood pressure. Control participants included those who were Caucasian and were unrelated to the gout patients. Genotypes were available for 67 gout cases and 4,712 control samples. One hundred fifty-three self-reported Caucasian gout patients who were not taking hypertensive medication were obtained from the Atherosclerosis Risk in Communities (ARIC) study and were compared to 6,969 unrelated controls.rs3733591 variant of SLC2A9 was genotyped across the NZ Caucasian, Māori and Pacific Island sample sets using the TaqMan Allelic Discrimination Assay Kit  and a LightCycler 480 Real-Time Polymerase Chain Reaction System .The A priori calculations were performed to test the power of the NZ Polynesian sample sets to detect association of rs3733591 with gout on the basis of previous data [ous data ,13. Statrs3733591 were in Hardy-Weinberg equilibrium (P > 0.01) for all case and control sample sets.Allelic and genotypic frequencies were compared between case and control samples, and ORs and adherence to the Hardy-Weinberg equilibrium were calculated using the SHEsis package . The genrs2075876 (AIRE), rs1816532 (ERBB4), rs13419122 (GFPT1), rs12401573 (SEMA4A), rs6945435 (MGC87315), rs743777 (IL2RB), rs10511216 (Intergenic), rs12745968 (FAM69A), rs1539438 (AP4B1), rs729749 (NCF4), rs3738919 (ITGAV), rs1130214 (AKT1), rs755622 (MIF), rs7901695 (TCF7L2), rs7578597 (THADA), rs2043211 (CARD8), rs10733113 (NLRP3), rs900865 (SOX6), rs2059606 (PGDS), rs4129148 (PseudoY), rs831628 (CD59), rs7725 (GFPT), rs573816 (upstream of ALDOB), rs1929480 (ALDOB) and rs12917707 (UMOD). There was an average allele frequency difference of 0.22 (0.03 to 0.61) between a subset of 469 Māori cases and controls and 505 Caucasian controls and a difference of 0.22  and 0.29 (0.04 to 0.67) between subsets of 417 Eastern Polynesian and 215 Western Polynesian cases and controls and 505 Caucasian controls, respectively. The genotype frequencies for the stratification markers all exhibited Hardy-Weinberg equilibrium P values > 0.003 for all case and control sample sets. STRUCTURE [PSTRAT) of the variant with disease in the presence of admixture. The phenotype of the 505 Caucasian individuals was set as unknown.Twenty-five biallelic markers were used as genomic controls to account for differing levels of non-Māori and non-Eastern Polynesian and non-Western Polynesian ancestry between the case and control samples. The stratification markers used were as follows: http://cran.r-project.org/web/packages/rmeta/index.html) (within Stata) to calculate the combined Mantel-Haenszel OR using a fixed effects model and the Breslow-Day test for heterogeneity between studies. Imputation of rs3733591 genotypes in the FHS and ARIC samples was done with IMPUTE2 using HapMap3 CEU  as reference data and a quality threshold of 0.9.Gender and gender-genotype interaction analysis was performed using Stata software . Meta-analysis was performed to combine data from independent data sets using rmeta software . The r2 values (measure of linkage disequilibrium) between the previously associated SLC2A9 variant, rs11942223 [rs3733591, were 0.04, 0.03, 0.04 and 0.05 for the Māori, Eastern Polynesian, Western Polynesian and Caucasian sample sets, respectively. Similarly, there was no evidence for an association on the basis of meta-analysis of the Eastern and Western Polynesian sample sets (OR = 1.05 (95% CI: 0.86 to 1.29); P = 0.62 and PBreslow-Day = 0.44, respectively). Meta-analysis of data from Polynesian, Han Chinese, Solomon Islands and Japanese sample sets (Table PBreslow-Day = 0.05); however, there was strong evidence for an association of rs3733591 in these combined Asian-Pacific populations . Addition of Caucasian data to the meta-analysis weakened the overall effect; however, a strong association was maintained .The demographic and clinical characteristics of study participants are presented in Table rs3733591 in the group with tophaceous gout in the Māori analysis , but not in the analyses of Eastern Polynesian , Western Polynesian  or NZ Caucasian  population samples  have shown gender-specific effects on serum urate [rs3733591 exerting a gender influence in the gout sample sets comprising NZ Māori , Eastern Polynesians  or NZ Caucasians  ). On the basis of logistic regression models, there was no evidence of an interaction between rs3733591 genotype and gender for the Māori, Eastern Polynesian or Caucasian sample sets . There was also no evidence for an association of rs3733591 with gout in the Māori, Eastern Polynesian or Caucasian sample sets when only males were analyzed .Although the intronic um urate , there wSLC2A9 variant has previously been demonstrated to be associated with gout and tophaceous gout in Han Chinese and Solomon Islanders [PSTRAT = 0.01), but not in the other sample sets .The R265H nonsynonymous slanders , and it slanders . We founslanders , we founts Table . At thisu et al.  and Uran [et al. . Note thrs3733591 with gout in the Māori sample set (OR = 0.98), there was an effect conferred by the C allele when the Māori cases were stratified for the presence or absence of tophi (OR = 2.21). This supports the previous findings of Tu et al. [rs3733591 might be a genetic checkpoint for tophaceous gout. Whether this is true will require further study in larger sample sets drawn from diverse ancestral groups. In the Asia-Pacific region, an effect for rs3733591 in tophaceous gout has been observed in the Han Chinese, Solomon Island and Polynesian NZ Māori populations, but not in Western Polynesian populations . Why this is the case is unclear; however, it is worth pointing out that unexpected heterogeneity in genetic association with gout in the Asia-Pacific region has also been documented at ABCG2 [Despite not finding any evidence for an association of u et al. , who conat ABCG2 . FurtherSLC2A9 is one of these factors. SLC2A9 is expressed in the chondrocytes of human articular cartilage [rs3733591, or one in linkage disequilibrium) may influence the activity of SLC2A9 in articular chondrocytes and increase the risk for deposition of MSU crystals and formation of tophi in joint structures. A further issue is that host factors other than hyperuricemia may contribute to the development of tophi; quantitative analysis of tophus histology indicates that, in contrast to the innate immune responses that are activated in acute gout, both innate and adaptive immunity are implicated in the development of the tophus [It is of interest that tophus formation is not consistently present in all patients with long-standing hyperuricemia and gout, suggesting that additional factors may regulate the development of these lesions. It is possible that genetic variation in artilage . The mine tophus . Thus dirs3733591 is the minor allele in control sample sets of Han Chinese (controls = 0.32) [ABCG2 (rs2231142)  [rs3733591 C allele increases, the risk conferred by rs3733591 for gout decreases, suggesting that population allele frequency is important at R265H to detect an effect on the risk of gout. It is also notable that the situation is reversed for the SLC2A9 intronic variants (rs11942223 and surrogates), with a strong association in Caucasian [Interestingly, the C-allele risk variant of  = 0.32)  Solomon  = 0.32) , Japanes = 0.32)  and Westnesians) . It can CG2 rs22342 , G25R (rs2276961), V253I (rs16890979) and P321L (rs2280205) [rs16890979 is associated with gout in Caucasians [rs6855911 and rs11942223 [Rs16890979 is also associated with gout in Polynesians; however, this is also likely to be secondary to an association with the intronic variants [rs16890979 is rare in Han Chinese and Japanese [In Caucasians, in addition to R265H, there are other nonsynonymous genetic variants within ucasians , possiblJapanese ,13. Of tSLC2A9 and ABCG2 in the Asian and Austronesian populations studied so far, further investigation of these genes in samples derived from Asian and Austronesian people is warranted. These populations have shared geographical history, although it is increasingly evident that considerable genetic heterogeneity exists within Asians and Austronesians, at least in gout and possibly in other complex diseases.Given the heterogeneity evident in risk conferred for gout at rs3733591 variant with gout in NZ Eastern Polynesian, Western Polynesian or Caucasian sample sets. It is possible that rs3733591 has a weak effect in these populations; however, our study's power was limited to detecting an effect less than that previously reported (OR = 1.4). However, consistent with a previous study in Han Chinese and Solomon Islanders, our data suggest that rs3733591 could be a marker of severe gout in some populations.We were unable to replicate association of the P: probability value; SHARe: SNP Health Association Resource; SLC2A9: solute carrier family 2, member 9; SNP: single-nucleotide polymorphism; URAT1: urate transporter 1.ABCG2: ATP-binding cassette-dependent transporter G2; ARIC: Atherosclerosis Risk in Communities; FHS: Framingham Heart Study; MSU: monosodium urate; NHLBI: National Heart, Lung and Blood Institute; NZ: New Zealand; OR: odds ratio; The authors declare that they have no competing interests.JEHM and TRM helped to design the study, oversaw its execution and prepared the manuscript. PJG, AAH, JH, PBBJ, LKS and ND helped to provide clinical recruitment and prepared the manuscript. All authors read and approved the final manuscript.


CTLA4 gene have been associated with autoimmune diseases and the functional CT60 single nucleotide polymorphism  has been proposed to be a casual variant in several of these diseases. The aim of this study was to ascertain whether this polymorphism is associated with inter-individual variation in susceptibility to pemphigus foliaceus. The population sample in this case-control association study comprised 248 patient and 367 controls. We did not found a significant association of pemphigus foliaceus with the CT60 variants. We conclude that the CTLA4CT60 polymorphism is not an important factor for pemphigus foliaceus pathogenesis in the population analyzed.Pemphigus foliaceus is an organ-specific autoimmune disease characterized by autoantibodies against the extracellular region of desmoglein 1, a protein that mediates intercellular adhesion in desmosomes. Cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) is a key negative regulator of the T cell immune response, playing an important role in T cell homeostasis and maintenance of peripheral tolerance. Polymorphisms in the  The CT60*G allele was correlated with higher disease susceptibility and lower mRNA levels of the soluble alternative splice form of CTLA-4 (sCTLA-4). Recent studies also suggested that polymorphisms in the CTLA4 gene may influence the development of autoimmune diseases. CTLA4 gene exert a strong influence on expression levels of both known CTLA-4 isoforms. Moreover, association between CT60 polymorphism and variation in the frequency of regulatory T cells has been reported. Individuals homozygous for allele CT60*A showed an increase from 30-40% in the frequency of regulatory T cells. Although the basic mechanism connecting the CT60*A allele with an increase in regulatory T cells has not yet been established, these differences reveal a relationship between CT60 polymorphism and variation in adaptive immune responses , is an organ-specific autoimmune disease characterized by autoantibodies against desmoglein 1 protein and by loss of adhesion between keratinocytes, leading to intraepithelial blisters of the skin , also known as 40L gene  showed t40L gene . Two SNPCTLA4 polymorphisms, especially the CT60 SNP, the aim of this study was to extend analysis of this candidate gene to evaluate whether the CT60 polymorphism is a factor contributing to differential genetic susceptibility to pemphigus foliaceus in the Brazilian population.Based on the reported associations between susceptibility to autoimmune diseases and We analyzed 248 patients and 367 controls without history of the disease, all unrelated to each other. Diagnosis was according to clinical and histological criteria. Patients and controls were contacted at Hospital Adventista do Pênfigo, Campo Grande, Mato Grosso do Sul State, Brazil. Additional controls were contacted in Curitiba, Paraná State, Brazil. Patients and controls were matched for ancestry. Seventy percent of the individuals were of predominantly European and 30% of predominantly African ancestry. The male:female ratio was close to one, viz. 47% of the patients and 45% of the controls were males. According to age of disease onset, distribution was as follows: from 0 to 9 years, 4.2%; 10-14 years, 8.3%; 15-19, 13.1%; 20-24, 11.3%; 25-29, 9.5%; 30-34, 9.5%; 35-39, 12.9%; 40-44, 8.3%; 45-49, 10.1%; 50-59, 5.4%; 60-69, 4.2%; 70-84, 3.6%. Written informed consent was obtained from all the participants. The study received approval by the Human Research Ethics Committee, in accordance with Brazilian Federal Laws.i.e., the frequency of individuals having the allele in either homozygosity or heterozygosity) were estimated by direct counting. Hardy-Weinberg equilibrium was assessed with the CTLA4 CT60 G/G. Therefore, by definition, the OR for this genotype equals 1 and the OR for A/G and A/A approach the risk of these genotypes relative to the G/G genotype.Genotyping was performed by polymerase chain reaction amplification followed by restriction fragment length polymorphism analysis (PCR-RFLP). The primers and PCR conditions were the same as those described by CT60 variants was detected.Allelic, genotypic and allele carrier frequencies are presented in CT60 polymorphism have been interpreted considering the possible effect of this SNP in the alteration of the ratio of CTLA-4 splicing isoforms, and that elevated levels of sCTLA-4 have already been detected in certain autoimmune disorders (CT60 polymorphism (see above), further studies of the same and other autoimmune diseases in various populations have failed to detect associations, thus concurring with our results (for example, CTLA4 gene is important in some diseases but not in others.Associations previously described between susceptibility to some autoimmune diseases and CTLA4 gene by analyzing CT60 polymorphism. Our results lead to the conclusion that genetic variation in CTLA4 does not play an important role in PF susceptibility and that the effect of variations in this gene differs among autoimmune diseases. We do not exclude the involvement of the CTLA-4 molecule in PF pathogenesis, but show that CT60 genotypes have no significant impact on pemphigus foliaceus disease susceptibility.In this study, we expanded our previous findings regarding variation in the


CYP26B1, NANOS1 and STRA8) have been proposed to play key roles in spermatogenesis.The meiotic program initiation pathway genes  and 363 infertility/normozoospermia subjects (sperm counts >20×106/ml). The control group consisted of 383 subjects with normal semen parameters, all of which had fathered at least one child without assisted reproductive technologies.In this study, all subjects were volunteers from the affiliated hospitals of Nanjing Medical University between March 2004 and July 2009 (NJMU Infertile Study). Total 719 idiopathic infertile cases were recruited and divided into three groups according to WHO semen parameters: 201 azoospermia patients (no sperm in the ejaculate even after centrifugation), 155 oligozoospermia patients  in STRA8 gene showed higher risk of spermatogenic impairment in the groups of abnormospermia (including azoospermia subgroup and oligozoospermia subgroup) and azoospermia than the controls with odds ratios and 95% confidence intervals of 2.52 (1.29–4.94) and 2.92 (1.41–6.06), respectively . Notably, larger sample size studies and in vivo or in vitro functional studies are needed to substantiate the biological roles of these variants.The genetic variant rs10269148 of Our results provided epidemiological evidence supporting the involvement of genetic polymorphisms of the meiotic program initiation genes in modifying the risk of azoospermia and oligozoospermia in a Han-Chinese population. CYP26B1, NANOS1, STRA8 et al) co-modify the process of meiotic program initiation Infertility was reported affecting 10–15% of couples, and roughly half of which are due to the man’s problem STRA8, stimulated by retinoic acid gene 8, was activated by retinoic acid (RA), which can directly enter the nucleus and bind to the corresponding retinoic acid receptor (RAR) to regulate the expression of specific genes. RA plays an important role in the process of cell growth, differentiation and apoptosis, and is catabolized by a family of cytochrome P450 (CYP) enzymes, CYP26A1, CYP26B1 and CYP26C1CYP26B1 is expressed in not the embryonic ovary but the embryonic testis, implying that the CYP26B1 is closely associated with male germ cell development. The expression of CYP26B1 in the testis of wild-type mouse is increased at embryonic day (E) 11.5, prior to mitotic arrest, and then reduced at E13.5 CYP26B1 on RA concentration may inactivate STRA8, thus ensuring the normal development of spermatogonia. Previous study reports that genetic deletion of CYP26B1 leads to increased RA levels and activation of STRA8 in the embryonic testes. As a consequence, male germ cells are prematurely entering into meiosis, arrested at pachytene stage, causing a rapid increase in apoptosis and a lack of spermatogonia of male mice after birth STRA8 inhibitory factors exist, the concentration of RA will be increased, and the STRA8 will be re-activated. To ensure the process of meiosis initiation go normally, other inhibitory factors are required.Of these genes, the NANOS  is one of the evolutionarily conserved proteins implicated in germ cell development and closely related with somite and germ cell development Nanos2 knockout mice’s testis during the embryonic period was similar to that of Cyp26b1 functional defects Stra8 expression in these two gene knockout mice showed certain period difference. In the testis of Cyp26b1 knockout mice, it begins to express highly in embryonic E13.5, but appears at E14.5 and continues until after birth in Nanos2 knockout mice’s testis. It seems that the Nanos2 begins to play an inhibitory effect on Stra8 when the Cyp26b1 expression decreases STRA8, CYP26B1, and NANOS1 were chosen as the typical meiotic program initiation pathway genes in this study.The Considering the essential role of meiotic program initiation pathway genes in spermatogenesis, we speculate that genetic variants of these genes have the potential to affect normal spermatogenesis. To test our hypothesis, we performed genotyping analyses for eight SNPs  in these genes, and investigated the association between these genetic variants and idiopathic infertility in a Chinese population. To the best of our knowledge, this is the first time to investigate the associations between these meiotic program initiation pathway gene polymorphisms and susceptibility to spermatogenic impairment.The study was approved by the Ethics review board of Nanjing medical university. The protocol and consent form were approved by the Institutional Review Board of Nanjing Medical University prior to the study. All participants provided their written informed consent to join in this study. And all the subjects were genetically unrelated ethnic Han-Chinese from East China. Every participant received complete medical history questionnaire, physical examinations and semen analysis. All infertility patients were examined, among which, those with a history of Y chromosome microdeletions, cytogenetic abnormalities, congenital bilateral absence of vas deferens, cryptorchidism and orchitis were excluded CYP26B1, NANOS1 and STRA8. All selected single nucleotide polymorphisms (SNPs) have reported minor allele frequencies (MAF) of >0.05 in Pubmed and are located in exons and UTRs. In the case of multiple SNPs in the same haplotype block (linkage coefficient r2>0.8), only one was selected. Finally, we identified eight potential functional polymorphisms  in three genes involved in the meiotic program initiation . The subjects with idiopathic infertility were first divided into two groups according to semen parameters: the abnormospermia group with sperm count below the WHO reference value and the normozoospermia group with the normal sperm count at or above the WHO reference value. And then the abnormospermia group was further divided into two subgroups: the azoospermia group and the oligozoospermia group. The semen analysis was described previously Statistical analyses were carried out using the Stata 10.0 statistical package . The Hardy-Weinberg equilibrium (HWE) tests were performed on online software, SHEsis  was used to compare the mean age, tea consumption, Body Mass Index (BMI) between the case and control groups. The chi-squared test was used to evaluate the differences in smoking status and drinking status of each SNP in the three genes between the case and control groups. Estimated infertility risks with odds ratios (OR) and 95% confidence intervals (95%CI) were calculated by multivariate logistic regression with adjustment for age, smoking status and BMI, where it was appropriate. Two-sided tests were used and the Bonferroni adjustment for multiple testing was applied. The applied p>0.05). The mean ages were higher in the groups of all infertility, azoospermia and oligozoospermia than the control group . BMI levels in the group of all infertility were significantly higher than those in the control subjects. Smoking prevalence was higher in the azoospermia group than the control group (p<0.05).The final population consisted of 1102 Han Chinese subjects, composed of 383 fertile controls, 201 azoospermia, 155 oligozoospermia and 363 infertility/normozoospermia. The distributions of selected characteristics among the case and control subjects were presented in p value <0.01 for deviation from HWE. The associations between meiosis initiation genes SNPs and the risks of male infertility were shown in The position and minor allele frequency of the 8 potential functional SNPs, were presented in STRA8 were significantly different existed between the infertility/abnormospermia group and the control group. Then the subgroup of infertility/abnormospermia was further stratified into two subgroups: the azoospermia group and the oligozoospermia group. Compared to the control group, statistically significant increased risk of idiopathic spermatogenic impairment was found for carriers of rs10269148CG genotype of STRA8, when compared with homozygous carriers of the C allele in the infertility/abnormospermia group and azoospermia subgroup respectively , while in the oligozoospermia group, the frequency distribution showed no significant difference. These results suggested that the G allele of rs10269148(C>G) may contribute to the severe idiopathic spermatogenic impairment.Initially, we overviewed the frequency distribution between the total case group and the control group, and no significant difference was found. To investigate the possible role of the meiotic program initiation pathway genes in spermatogenesis, we subdivided the case group into two subgroups on the basis of sperm concentration: infertility/abnormospermia and infertility/normozoospermia. The genotype frequencies of rs10269148 in CYP26B1, although the distribution frequencies were lower in the total infertility group [12.52% (CT+CC)] than that in the control group [16.98% (CT+CC)], the distribution difference was not statistically significant after Bonferroni correction . And to the rs707718, logistic regression analysis revealed that compared with the rs707718 AA genotype, subjects in oligozoospermia group carrying the heterozygous rs707718 AC genotype had a significantly 39% decreased risk of spermatogenic impairment . When we combined the rs707718 AC and CC genotype, assuming a co-dominant allele effect, the combined rs707718 AC+CC variant genotypes were associated with a 41%  reduced risk of spermatogenic impairment. The OR values suggested that the variants rs2241057T>C, rs707718A>C may be protective factors against the risk of idiopathic male infertility with abnormal and/or normal semen parameters.To the rs2241057 of As to the other SNPs, no significant differences of distribution frequencies were identified among the case and control groups. In our study, due to the low occurrence frequency in Asians, we didn’t analyse homozygous mutant separately.STRA8 is required for premeiotic DNA replication, while CYP26B1 is decreased. By preventing STRA8’s expression, CYP26B1 and NANOS1 play critical roles in the differentiation of male germ cells CYP26B1and NANOS1 expression in the germ cells CYP26B1, NANOS1) and effectors (STRA8) of the meiosis-inducing action of RA were analyzed in this study. Through information gained from PubMed and Hapmap searches, eight potential functional polymorphisms in meiosis initiation genes were examined.Although meiosis initiation genes have been recognized as key regulators in sperm function and male fertility Vivo studies have demonstrated that male mice lacking STRA8 function produce no sperm p = 0.006). In the following stratified analysis, compared with homozygous type CC, the rs10269148 C>G increased the risk of azoospermia and displayed 2.92 fold increased risk of spermatogenic impairment, which was consistent with the phenotypes in STRA8−/− mice STRA8 is suppressed by a retinoid-degrading enzyme, which is mainly coded by CYP26B1. The CYP26B1, an important member of CYP family, is expressed in sertoli cells, and has a crucial role in the formation of sperm CYP26B1, which causes Leu to Ser substitution at position 264 (rs2241057 T>C) (http://www.ncbi.nlm.nih.gov/projects/SNP/snp_ref.cgi?rs=2241057), may lower the risk of spermatogenic impairment in our study population. As to the rs707718, comparing with homozygous type AA, the A>C mutation significantly reduced the risk of spermatogenic impairment, displaying 0.59 fold decreased risk of spermatogenic impairment . However this difference disappeared after the Bonferroni correction. The statistical power of our study may be limited by the small sample size.CYP26B1 (rs2241057 T>C) enhanced the catabolicing activity of CYP26B1 substantially, both in a cell free system and in cell culture. The actual catabolic rates in vivo depend on the local tissue concentration of atRA and many other factors, which cannot be accurately predicted from the data. But it is plausible that the differences between variants of CYP enzyme alter the equilibrium of etinoid synthesis and catabolism CYP26B1 in biogenesis and postnatal growth, we speculate that the genetic variation may affect the capacity of CYP26B1 enzyme, then the meiotic program of germ cell, which might tightly associates with male infertility.Our results were consistent with that the substitution of CYP26B1 has an important inhibitory effect during the early initiation of meiosis in testicular germ cells, when it comes to E13.5, the expression begins to decrease STRA8. So the other STRA8 inhibitory factors- NANOS1 are required before the spermatogonial cells initiating meiosis. Given that NANOS1 is an RNA-binding protein localized in the cytoplasm, we suspected the regulation of STRA8 transcription may be an indirect effect of NANOS1NANOS1 gene on the STRA8 gene might be moderate and other macromolecules might participate in the initiation of meiosis. The precise mechanism of the variants of NANOS1 in male infertility needs further investigation.Although In conclusion, the results of our study demonstrated for the first time that some representative genetic variants in the meiosis initiation genes might regulate the risk of male infertility. Although the statistical power of our study was limited by the small sample size in the subgroups, our findings might be helpful to understand the mechanism of male infertility. Larger sample size studies and in vivo or in vitro functional studies will be needed to substantiate the biological roles of these variants.


The search to identify disease-susceptible genes requires access to biological material from numerous well-characterized subjects. Archived residual dried blood spot (DBS) samples, also known as Guthrie cards, from national newborn screening programs may provide a DNA source for entire populations. Combined with clinical information from medical registries, DBS samples could provide a rich source for productive research. However, the amounts of DNA which can be extracted from these precious samples are minute and may be prohibitive for numerous genotypings. Previously, we demonstrated that DBS DNA can be whole-genome amplified and used for reliable genetic analysis on different platforms, including genome-wide scanning arrays. However, it remains unclear whether this approach is workable on a large sample scale. We examined the robustness of using DBS samples for whole-genome amplification following genome-wide scanning, using arrays from Illumina and Affymetrix.This study is based on 4,641 DBS samples from the Danish Newborn Screening Biobank, extracted for three separate genome-wide association studies. The amount of amplified DNA was significantly (P < 0.05) affected by the year of storage and storage conditions. Nine (0.2%) DBS samples failed whole-genome amplification. A total of 4,586 (98.8%) samples met our criterion of success of a genetic call-rate above 97%. The three studies used different arrays, with mean genotyping call-rates of 99.385% (Illumina Infinium Human610-Quad), 99.722% (Illumina Infinium HD HumanOmni1-Quad), and 99.206% (Affymetrix Axiom Genome-Wide CEU). We observed a concordance rate of 99.997% in the 38 methodological replications, and 99.999% in the 27 technical replications. Handling variables such as time of storage, storage conditions and type of filter paper were shown too significantly (P < 0.05) affect the genotype call-rates in some of the arrays, although the effect was minimal.Our study indicates that archived DBS samples from the Danish Newborn Screening Biobank represent a reliable resource of DNA for whole-genome amplification and subsequent genome-wide association studies. With call-rates equivalent to high quality DNA samples, our results point to new opportunities for using the neonatal biobanks available worldwide in the hunt for genetic components of disease. Identifying genetic effects in complex disorders usually requires genome studies in large cohorts. Access to DNA from well-characterized patients and healthy controls represents a major bottleneck. This problem may be circumvented by using archived residual blood samples from newborn screening programs, which encompass the entire population under a certain age in several countries. The blood is usually collected by heel-prick and applied to special filter paper; a proven robust and convenient medium for transport and storage . StorageThe Danish Neonatal Screening Biobank (DNSB) contains nearly two million DBS samples collected from almost every Dane born after 1981. It has recently been updated to meet new general guidelines for the establishment and operation of biobanks [In Denmark, all citizens have a unique personal identification number used in all public registration systems, including the DNSB. Denmark also has a well-established public health care system with equal treatment offered to all citizens. These resources allow researchers to study the entire country as a cohort, and make the DNSB an ideal resource for studying common and complex genetic diseases in Caucasians .A major challenge using DBS samples for genetic studies is the small amount of blood available in a spot. The amount of genomic DNA (gDNA) that can be extracted from a 3.2-mm punch of a DBS sample is approximately 60 ng . In geneHere we describe genome-wide association studies (GWAS) using DBS samples from the DNSB. Storage time, storage conditions, and type of filter paper used for DBS collection were evaluated to determine their effects on the amount of amplified wgaDNA material obtained from each sample. The effects of these variables on genotype call rates in three studies, using three different types of array, running on either Illumina or Affymetrix genotyping platforms were also examined.Our 4,641 subjects were obtained from three case-control GWAS studies. The first study, GEMS (Genomic Medicine for Schizophrenia), called "610k" in this manuscript, included 1,808 DBS samples stored from 1981-1996. The purpose was to identify genetic regions associated with schizophrenia . The second study, "Omni1", provided 1,283 DBS samples stored from 1982-2006, and was undertaken to examine the role of genetics in Meningococcal and Pneumococcal infections . The third study, "Axiom", aimed to identify genetic variations associated with asthma, and included DBS samples stored from 1982-2006 . All studies were approved by the DNSB Steering Committee. The current study was conducted as an anonymous register study.Two 3.2-mm disks were punched from each DBS sample, and protein was removed as previously described . GenomicTo evaluate the sample processing we pooled the DBS samples included in the three GWAS studies. Pooling samples was statistically sound as all samples were treated identically up to the step before choosing the SNP genotyping array platform and technology, but possible sample effects were tested statistically. We used a linear regression model to test for interaction between the included variables: years of storage counting from 1981 (years), storage conditions , 1 ), type of filter paper , 1 ) and the wgaDNA concentration.The GCR was used for evaluating the array efficiency and sensitivity to biobank variables. To meet the criteria of being normally distributed, the GCR was transformed using a zero-skewness log (resulting in log(1-GCR), the logarithm of the failure rate). The effect of the years of storage, type of filter paper, and storage conditions on the transformed GCR was analysed in the three studies individually using a linear regression interaction model. A bivariate linear regression model was used to evaluate the effect of the wgaDNA concentration on the GCR. STATA MP11 software  was used for the statistical analyses.A total of 4,641 subjects from the three GWAS disease studies were used to evaluate the use of DBS samples for genetic studies. "610k" included 1,808 samples stored for a mean of 23.9 years : 2.8 years; 45.5% female and 54.5% male). "Omni1" was based on 1,283 samples stored for a mean of 15.7 years . "Axiom" included 1,550 samples stored for a mean of 16.5 years .As the three studies used the same sample preparation process, we were able to pool their results to evaluate the method. As both storage conditions and filter paper are time dependent, bivariate analysis was not appropriate. Results of the multiple regression models summarised in Table In the "610k" and "Omni1" studies, 60 and 18 samples, respectively, failed to meet our predefined GCR criterion of > 97% and were requeued Table . Eleven The three GWAS studies were evaluated separately as their performance was significantly different (data not shown), likely because different array types were used on different genotyping platforms (Affymetrix and Illumina).The mean GCRs in the three studies were: "610k", 99.385385% ; "Omni1", 99.722 ; and "Axiom", 99.206% .Considering that wgaDNA concentration is an intermediate variable between GCR and years of storage and storage conditions Table , we did Dried blood spot samples are being collected and stored in biobanks for diagnostic and research purposes worldwide. In several countries this has been common practice for several decades. We previously showed that DBS samples from the DNSB can be used to generate reliable genetic results using the Illumina genome-wide scanning technology, but this evaluation was restricted to relatively few samples . In the Storage time influenced the wgaDNA amplification negatively, with significantly lower concentrations in the older samples Table Figure 11Figure . The conAccording to our regression model, and as suggested in a previous study , increasThe wgaDNA samples in the three studies performed excellently, with mean GCRs greater than 99.2%, and replication concordance rates greater than 99.9%  in "Omni1" and 200 (~13%) in "Axiom"), so the significant associations could also be artefacts. Future studies will help us to answer this question.The storage conditions significantly affected the GCR in the "Axiom" study, with the GCR increasing when samples were stored at -20°C. Unexpectedly, the older samples performed better than the more recent samples. We speculate that the difference between the Affymetrix (ligation) and Illumina (single base extension) SNP genotyping approaches may contribute to this. Overall, of the three arrays tested, the Illumina "Omni1" array performed best. Compared with the other arrays, "Omni1" had the highest mean GCR, and the highest sample success rate. It is important to point out that none of the arrays performed poorly, and that the effects of the different variables on the GCR were minimal, even when statistically significant. All three arrays should be considered usable for GWS of DBS samples.The robustness of the three GWS studies indicates that filter paper is an excellent way to collect and store whole blood samples for later DNA research purposes. Collecting samples on filter paper has several advantages compared with standard venepuncture, including less discomfort for the patient, especially if several samples need to be collected within a short period of time. The relatively small amount of blood taken limits the number of analyses that can be performed, but techniques such as WGA help mitigate these restrictions with regard to DNA based methods. To date, DBS samples have been used for multiplex protein analysis , VitaminIn summary, we found that DNSB DBS samples constitute a good resource for SNP genotyping and GWS array studies. Samples in neonatal screening biobanks worldwide should be considered an important source of genetic material for future genetic studies. Our results also suggest suggests that new samples for GWS studies can be collected on filter paper with minimal discomfort for patients, potentially higher participation rates, and convenience in collection, shipping, costs, and storage as compared with whole blood obtained by venepuncture. Depending on the array chosen, different variables may marginally affect the GCR, but overall our approach using DBS samples stored for up to 28 years performed as well as good quality DNA from whole-blood samples. Though not significantly affecting the GCR, we emphasize the importance of storing DBS samples at-20°C, to enhance the numbers of biomarkers that can be analysed.MVH drafted the manuscript, managed the DBS samples and genetic material used in the three studies, analysed the genetic GWS data in the "610k" and "Omni" studies, did the statistical analyses, and interpreted the results. JAG revised the statistical analysis, interpreted data, and revised the manuscript critically. JOG performed the GWS of the "610k" and "Omni" studies and was involved in the quality testing of the arrays. EKM performed the initial analysis of the "Axiom" GWS data, and revised the manuscript critically. KB contributed substantially to the conception and design of the "Axiom" study and took part in the initial analysis of the GWS data. MN and TLB conceived the "Omni" study, and participated in its design and coordination, took part in the initial analysis of the "Omni" GWS results, and revised the manuscript critically. PBM, OM, TFO, and BNP conceived the "610k" study, and participated in its design and coordination. HTS contributed substantially to the conception and design of the "Omni" study, and revised the manuscript critically. ZBH contributed substantially to the conception and design of the "Omni" study. ADB conceived the "610k" study, and participated in its design and coordination, and revised the manuscript critically. DD performed the quality test of the "610k" GWS data, and revised the manuscript. HB conceived the "Axiom" study, and participated in its design and coordination. DMH conceived the "610k" study, participated in its design and coordination. DMH also participated in the coordination of the "Omni" and "Axiom" studies, and critically revised the manuscript. All authors read and approved the final manuscript.


Clefts of the lip (CL), the palate (CP), or both (CLP) are the most common orofacial congenital malformations found among live births, accounting for 65% of all head and neck anomalies. The frequency and pattern of orofacial clefts in different parts of the world and among different human groups varies widely. Generally, populations of Asian or Native American origin have the highest prevalence, while Caucasian populations show intermediate prevalence and African populations the lowest. To date, little is known regarding the epidemiology and pattern of orofacial clefts in Tanzania.A retrospective descriptive study was conducted at Bugando Medical Centre to identify all children with orofacial clefts that attended or were treated during a period of five years. Cleft lip and/or palate records were obtained from patient files in the Hospital's Departments of Surgery, Paediatrics and medical records. Age at presentation, sex, region of origin, type and laterality of the cleft were recorded. In addition, presence of associated congenital anomalies or syndromes was recorded.2 = 92.4, p < 0.001), followed by the right (28.8%) and bilateral sides (18.3%). Patients with isolated cleft palate presented at very early age . Associated congenital anomalies were observed in 2.8% of all patients with orofacial clefts, and included neural tube defects, Talipes and persistent ductus arteriosus.A total of 240 orofacial cleft cases were seen during this period. Isolated cleft lip was the most common cleft type followed closely by cleft lip and palate (CLP). This is a departure from the pattern of clefting reported for Caucasian and Asian populations, where CLP or isolated cleft palate is the most common type. The distribution of clefts by side showed a statistically significant preponderance of the left side (43.7%) (χUnilateral orofacial clefts were significantly more common than bilateral clefts; with the left side being the most common affected side. Most of the other findings did not show marked differences with orofacial cleft distributions in other African populations. Clefting of the lip (CL), the palate (CP), or both (CLP) are the most common orofacial congenital malformations found among live births, accounting for 65% of all head and neck anomalies . AetioloOrofacial clefts have long been known to be associated with other congenital defects, though the frequency and type of associated malformations observed varies considerably across studies . ObserveAlthough the epidemiology of cleft lip and palate has been studied extensively in different parts of the world Table  18,19],,19,18,19A retrospective descriptive study was conducted at Bugando Medical Centre, located in the northwest region of Mwanza, Tanzania, to identify all children with orofacial clefts that attended or were treated from October 2004 to July 2009. Bugando Medical Centre (BMC) is one of four referral hospitals in the country, serving a population of approximately 10 million people in five regional districts. Most patients with orofacial clefts in the surrounding regions are usually referred to this hospital as it is the only centre that offers surgical expertise to repair orofacial clefts on the North-Western part of Tanzania. Cleft lip and/or palate records were obtained from patient files in the Hospital's Departments of Surgery, Paediatrics and Medical Records. Patient file notes are usually written by medical officers and surgeons from the time of hospital admission to discharge. Age at presentation, sex, region of origin, type and laterality of the cleft were recorded. Additionally, presence of associated congenital anomalies or syndromes was recorded. Orofacial cleft cases that lacked some of the above information  were excluded. Most of the associated congenital anomalies reported were of a type that can be readily observed by medical officers and/or surgeons, as genetic services are very limited at BMC. The study was reviewed and approved by the institutional review boards of both Bugando University College and the University of Calgary, in accordance with the dual affiliation of the first author.Data were recorded and entered into SPSS version 11.5. Cleft type was classified as follows: 1. CL: Cleft lip and alveolus , 2. CLP: Cleft lip and palate , 3. Cleft palate . Frequencies were obtained for different types of clefts in relation to the total number of orofacial clefts. Deviations from expected frequencies  were analysed using contingency tables and Chi-square analyses.A total of 240 cases with orofacial cleft were seen between October 2004 and July 2009. Among these, 14 cases were admitted in the Paediatrics Department with other medical conditions; orofacial cleft not being the primary reason for hospital admission. About 60 additional cases with orofacial clefts seen at BMC during the study period had incomplete patient files, and thus were not included in the study. The majority of cases came from Mwanza region, followed by Mara and Shinyanga regions . In contrast, the association between cleft type and laterality was statistically significant . The distribution of clefts by side showed a preponderance of the left side (43.7%), followed by right side (28.8%) and bilateral cases (18.3%). Midline clefts accounted for only 10% of all cleft cases. About 2.8% of cleft cases were associated with congenital anomalies, including two cases each of Hydrocephalus, Talipes equinovarus, Spina bifida, and one case of Persistent Ductus arteriosus.The most common cleft type was isolated cleft lip (CL), constituting 49.2% of all cleft deformities Table . OverallA wide range of age at presentation was observed in this study (range 1 month to 18 years), with about 33% of all cleft cases presenting at over one year of age. Patients with isolated cleft palate presented earlier  as compared to those with isolated cleft lip or cleft lip and palate ; however, these differences were not statistically significant .The aim of this retrospective study was to report on key aspects of the epidemiology of orofacial clefts in Tanzania, based on a review of hospital records at Bugando Medical Centre in Mwanza. Retrospective studies are usually small, based on clinic records, subject to underreporting, and may suffer from multiple sources of ascertainment bias (e.g. socioeconomic factors). While prospective studies are ideal, they are not currently feasible in Tanzania. Even so, ongoing studies of the genetics of CLP in East-Africa require analysis and interpretation of the existing clinical records to determine what they can reveal about the epidemiology of CLP in this region.In this study, potential sources of underreporting at BMC include the distance and/or cost of travel to the hospital and referrals to other treatment centres closer to the patient's area of residence. For instance, although patients in this study came from the seven regions served by BMC as a referral hospital, the majority came from the regions closest to the hospital, namely, Mwanza, Mara and Shinyanga. Therefore, as was recently observed in another study in Kenya , distancBugando Medical Centre provides surgical treatment and other supportive care like speech therapy to patients with orofacial clefts. Due to differential reporting among surgeons and the fact that some of the records were missing key post operative information, we did not investigate the fate or outcome  of these patients after surgery.et al [At BMC, isolated cleft lip was the most common cleft type, followed closely by clefts of both the lip and palate. Similar findings have been observed in other African studies Tables  and 3. Fet al  found thet al ,16,23. Aet al . Alternaet al ,25.The sex distribution of isolated cleft lips showed a skew towards females Table , which cThe age at presentation in our study does not support the hypothesis that patients with orofacial clefts in low and middle income countries tend to present at later age due to unavailability of specialized medical facilities . As has The result from this study shows the frequency of patients with orofacial clefts and associated congenital anomalies to be 2.8%. Other studies have reported that this frequency can range from as low as 4.3%  to as hiIn this preliminary study, we found that unilateral clefts were more common than bilateral clefts. Unilateral clefts showed preponderance for the left side. Most of the other findings from our study regarding distribution of orofacial clefts were similar to other African populations. Owing to the nature of this hospital-based retrospective study, it was not possible to estimate the true prevalence of orofacial clefts and their associated congenital anomalies in Tanzania at this time. Further large population- and birth registry-based studies are needed to obtain more representative results regarding both the prevalence and frequency of associated anomalies for orofacial clefts in Tanzania, and more broadly in Africa.The authors declare that they have no competing interests.MM made substantial contributions to the conception and design of this study, participated in data collection, statistical analysis and interpretation of results, drafted and revised the final manuscript, and read and approved the final manuscript. CR contributed substantially to statistical analysis and draft of the manuscript. JG and CM participated in the design and data analysis. KM, EM and EK made substantial contribution to the data collection and analysis. BH made substantial contribution to the statistical analysis and draft of the manuscript. All authors have read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1472-6831/11/5/prepub


Objective: Vascular events are a common complication in patients with polycythemia vera (PV) and essential thrombocythemia (ET). This study aimed to analyze the association between PAI-1 4G/5G and ACE I/D gene polymorphisms, and leukocytosis with thrombosis in patients with PV and ET. Material and Methods: In total, 64 patients with ET and PV were evaluated. Arterial or venous thrombosis, such as cerebral transient ischemic attack, ischemic stroke, myocardial infarction, peripheral arterial thrombosis, deep venous thrombosis, and pulmonary embolism, were defined as a vascular event. DNA samples were screened for mutations via reverse hybridization strip assay. Results: In terms of PAI-1 gene polymorphism, the frequency of the 4G and 5G allele was 48.5% and 51.5%, respectively. The ACE allele frequency was 51.2% and 48.8% for D and I, respectively. There wasn’t an association between occurrence of vascular events and the frequency of any allele. In terms of occurrence of vascular events, there weren’t any significance differences between the patients that were carrying the ACE D/D homozygous allele to ACE I/D and those that carried the I/I allele (P = 0.93). There wasn’t a significant difference in occurrence of vascular events between the PAI-1 5G/5G homozygote allele carriers, and the 4G/5G and 4G/4G allele carriers (P = 0.97). Vascular events were significantly more common in the patients with leukocytosis (leukocyte count >10 × 109 L–1) than in those without leukocytosis (leukocyte count ≤10 × 109 L–1) (P = 0.00). Age >60 years was also a significant risk factor for occurrence of vascular events(P = 0.008). Conclusion: PAI-1 and ACE gene polymorphisms were not considered new risk factors for thrombosis in PV and ET patients. On the other hand, leukocytosis at diagnosis was associated with the occurrence of vascular events in the patients with ET and PV. Thrombotic events, both in the arterial and venous systems, are a common complication in patients with polycythemia vera (PV) and essential thrombocythemia (ET). About 30%-50% of patients with PV and ET suffer from these complications, and vascular mortality accounts for 35%-45% of all deaths ,2,3. StuThe formation of thrombus depends on the balance between fibrinolytic and procoagulant systems. During fibrinolysis plasminogen is converted to plasmin, which in turn dissolves fibrin clots. This conversion is regulated by tissue plasminogen activator and plasminogen activator inhibitor-1 (PAI-1)—a serine protease inhibitor . The PAIRigat et al. 1990) reported a polymorphism involving the presence  or absence  of a 287-bp sequence of DNA in intron 16 of the angiotensin I-converting enzyme (ACE) gene. According to this polymorphism, 3 genotypes  were determined. The D/D genotype has been associated with the highest plasma ACE activity, the I/D genotype with moderate ACE activity, and the I/I genotype with the low ACE activity . It has 90 reportTo date, no study on PAI-1 4G/5G and ACE I/D gene polymorphism in relation to the risk of thrombosis in patients with PV and ET has been published. As such, using our patient database the present study aimed to analyze the association between these polymorphisms, and age and leukocytosis with thrombosis in patents with PV and ET.PatientsThis retrospective cohort study included patients with ET and PV that were diagnosed according to WHO-2008 criteria . Cerebral computed tomography (CT) or magnetic resonance imaging (MRI) was used to diagnose ischemic stroke and neurologic symptoms. In addition to these procedures, electrocardiography and/or increased cardiac enzymes, angiography, ultrasonography of the arms or legs, and pulmonary ventilation-perfusion scan and/ or CT were used to diagnosis TIA, AMI, PAT, DVT, and PE, respectively. Microcirculatory events, such as vascular headaches, dizziness, distal paresthesia, etc., were not considered vascular events and were not analyzed. The study was conducted in accordance with good clinical and laboratory practices, and the principles of the Declaration of Helsinki. The University of Kocaeli Ethics Committee approved the study protocol. GenotypingDNA extractionGenomic DNA was extracted from 3 μL of peripheral blood with EDTA using the MagNA Pure Compact Nucleic Acid Isolation Kit I  and was stored at 4 °C until analyzed. Quantification of the DNA concentration was performed using a NanoDrop ND-1000 spectrophotometer. The ratio of spectrophotometric measurements at 260 nm and 280 nm provided an estimate of the purity of the DNA sample. Multiplex PCR-CVD StripAssay Isolated DNA samples were screened for mutations associated with cardiovascular disease using reverse hybridization strip assay . This assay covers PAI-1, 4G/5G, and ACE I/D mutations. For each reaction 5 μL of DNA was added to 20 μL of PCR reaction mix containing 15 μL of amplification mix and 5 μL of diluted Taq DNA polymerase (1U). PCR was performed in 0.2-mL tubes under the following cycling conditions: initial denaturation at 94 °C for 2 min, 35 cycles at 94 °C for 15 s, 58 °C for 30 s, 72 °C for 30 s, and a final extension step at 72 °C for 3 min. To confirm amplification, each PCR product was electrophoresed on 3% agarose gel. Finally, the amplification products were selectively hybridized to a test strip that contained allelespecific (wild type and mutant) oligonucleotide probes immobilized as an array of parallel lines. Bound biotinylated sequences were detected using streptavidin-alkaline phosphatase and color substrates. Hybridization was performed in an automated incubator . For each polymorphic position, 1 of 3 possible staining patterns may be obtained: normal genotype (wild type probe only), heterozygous genotype (wild type and mutant probe), and homozygous mutant genotype (mutant probe only). Mutation detection using real-time PCR JAK2 gene mutations were detected using a LightCycler 1.5  and JAK2V617F LightMix Kit , and capillary tubes. According to the manufacturer’s protocol, 20 uL of reaction containing 5 uL of genomic DNA and 15 uL of reaction mix  was prepared. Each study was carried out with negative control (5 uL of PCR-grade water) and positive controls that were available in commercial kits. Data obtained were analyzed using LightCycler v.3.5 software. Mutations were evaluated according to the status of the normal allele or the mutant, in consideration of the specific melting points (Tm). Statistical analysisAll analyses were performed using SPSS v.15.0 for Windows XP . Pearson’s chisquare and Fisher’s exact test were used to compare categorical variables. The Mann-Whitney U test was used to measure the statistical significance of the leukocyte count at diagnosis and its association with occurrence of vascular events. The relationship between vascular events and an above normal leukocyte count was evaluated using Pearson’s chi-square test. The frequency of alleles and their significance were determined using the Mann-Whitney U test. P values <0.05 were considered statistically significant. In total, 64 patients were evaluated. Patient characteristics are summarized in The mean leukocyte count at diagnosis in patients with and without vascular events was 13.4x10[u]9[/u] L[u]–1[/u] ± 3.9x10[u]9[/u] L[u]–1[/u] and 9.8 x 10[u]9[/u] L[u]–1[/u] ± 3.8x x 10[u]9[/u] L[u]–1[/u], respectively. Significantly more vascular events were observed in the patients with leukocytosis (leukocyte count >10 x 10[u]9[/u] L[u]–1[/u]) (P = 0.00) than in those without leukocytosis (leukocyte count ≤10 x 10[u]9[/u] L[u]–1[/u]) and the presence of leukocytosis at diagnosis was a significant risk factor for vascular events (OR = 15.625 [95% CI: 4.322-56.492]). In addition to leukocytosis, age >60 years was a significant risk factor for vascular events (P = 0.008) (OR = 4.154 [95% CI: 1.416- 12.184]) .Thrombosis in patients with PV and ET occurs in arterial, venous, or microcirculatory systems. In the present study the majority of vascular complications were arterial, which is in accordance with previous reports . Age aboHereditary thrombophilic states in patients with PV and ET, including factor V Leiden, prothrombin G20210A, and methylenetetrahydrofolate reductase (MTHFR) mutations, have been studied extensively ,12. DataData regarding the association between the presence of the 4G allele and the risk for venous thrombotic episodes are also controversial. A recent meta-analysis reported an unexpected weak association between the 4G allele and the risk of venous thromboembolism, based on analysis of patients with venous thrombosis that were not included in certain risk groups with other genetic or acquired thrombophilic defects . NonetheThe correlation between ACE polymorphism and vascular events, especially arterial events, has been studied. Dilley et al. (1998) reported that patients with ACE D/D polymorphism had moderately higher risk for venous thrombosis risk than those with other genotypes (OR = 1.5 [95% CI: 0.9-2.6]). In addition, females with the ACE D/D genotype did not a have an increased risk (OR = 0.9 ) . CambienSayed-Tabatabaei et al. suggested that the D allele is not clinically important in the general population, but may play an important role in certain groups of patients with coronary heart disease . While 2In recent years new data concerning the pathogenesis of thrombosis in Philadelphia-chromosome negative myeloproliferative disorders have become available, including the role of leukocyte activation, and leukocyte interaction with platelets and endothelial cells. In myeloproliferative disorders neutrophils circulate in an activated state and are able to bind to platelets in a dynamic adhesive process ,42,43,44The research-proven risk factors for thrombosis in patients with ET and PV are age >60 years and a history of thrombosis. The presence of either risk factor places patients in the high-risk category, indicating the need for myelosuppressive therapy . In the Conflict of Interest Statement The authors of this paper have no conflicts of interest, including specific financial interests, relationships, and/ or affiliations relevant to the subject matter or materials included.


Previously, CYP1A1 Ile462Val polymorphism has been indicated to be a risk factor for several malignancies. Increasing reports have focused on the association of CYP1A1 Ile462Val polymorphisms with susceptibility to acute leukemia and have generated controversial results. The goal of the present study was to derive a more precise estimation of the relationship.Relevant literature has been rigorously searched and screened. Eligible studies were identified for the period up to Apr 2012. Meta-analyses evaluating the association of CYP1A1 Ile462Val variation with acute leukemia were carried out. Subgroup analyses on ethnicity, clinical types and source of controls were further performed.A total of thirteen publications including fourteen case-control studies with 2164 cases and 4160 controls were selected for analysis. The overall data indicated a significant association of CYP1A1 Ile462Val polymorphism with acute leukemia risk . In subgroup analysis on ethnicity, increased risk was shown among mixed ethnicities  but not Asians or Caucasians. In subgroup analysis on clinical types, increased risk was observed in the acute lymphocytic leukemia (ALL) subgroup  but not in the acute myeloid leukemia (AML) subgroup.The results of the present study suggest that CYP1A1 Ile462Val polymorphism might be a low-penetrant risk factor for acute leukemia. Subgroup analyses suggest that homozygous Val/Val alleles might modify the susceptibility to ALL. Acute leukemia, a malignant tumor of the hematopoietic system, is characterized by a rapid increase in the numbers of immature blood cells. The disease can be subdivided into two major groups according to the cell affected as acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML), respectively. ALL is the most common type of leukemia in young children while AML occurs more commonly in adults than in children Several genetic variations have been evaluated as possible risk factors for leukemia by meta-analyses. Polymorphisms of GSTM1, GSTT1, MTHFR C677T and XRCC1 Arg399Gln have been indicated to increase leukemia risk Previous evidence indicates that carcinogen-metabolizing genes may play critical roles in determining individual susceptibility to malignancies A number of published studies have been conducted on the relationship between CYP1A1 Ile462Val polymorphism and acute leukemia risk. However, the results are inconclusive. The issue of whether the CYP1A1 Ile462Val polymorphism is a risk factor for acute leukemia has not been clearly addressed. Thus, in this study we aimed to derive a more precise estimation of the relationship by performing a quantitative meta-analysis that increases statistical power to reach more convincible results.Cytochrome P450 1A1, CYP1A1, Ile462Val, exon7, acute leukemia, hematology, malignancy, neoplasm, cancer, variation and polymorphism. All searched studies were retrieved and the bibliographies were checked for other relevant publications. Review articles and bibliographies of other relevant studies identified were electric/hand searched to find additional eligible studies.To obtain eligible literature, we carried out a search in the Medline, EMBASE, OVID, Sciencedirect, Google Scholar and Chinese National Knowledge Infrastructure (CNKI) without a language limitation, covering published publications up to Apr 2012, with a combination of the following keywords: The following criteria were used for the literature selection: first, studies should concern the association of CYP1A1 Ile462Val polymorphism with acute leukemia risk; second, studies must be observational studies (Case–control or cohort); third, papers must offer the sample size, odds ratios (ORs) and their 95% confidence intervals (CIs), the genetic distribution or the information that can help infer the needed results. Accordingly, the following criteria for exclusion were also used: first, the design and the definition of the experiments were obviously different from those of the selected articles; second, the source of cases and controls and other essential information were not offered; third, reviews and duplicated publications. After rigorous searching, we reviewed all papers in accordance with the criteria defined above for further analysis.(Zhuo and Zhang) according to the inclusion criteria mentioned above. For conflicting evaluations, an agreement was reached following a discussion. If a consensus could not be reached, another author was consulted to resolve the dispute and then a final decision was made by the majority of the votes. The extracted information was entered into a database.Data were carefully extracted from all eligible publications independently by two of the authors 2I value was applied to evaluate heterogeneity between the included studies  . In addition, a Chi-square based Q statistic test was conducted to assess the heterogeneity. If the result of the Q-test was P>0.1, ORs were pooled according to the fixed-effect model (Mantel-Haenszel), Otherwise, the random-effect model (DerSimonian and Laird) was used. The significance of the pooled ORs was determined by Z-test. The Hardy-Weinberg equilibrium (HWE) was assessed by Fisher’s exact test. Publication bias was assessed by visual inspection of funnel plots The odds ratio (OR) of CYP1A1 Ile462Val polymorphisms and acute leukemia risk was estimated for each study. The pooled ORs were performed for a homozygote comparison model , a dominant model  and a recessive model , respectively. To detect any possible sample size biases, the OR and its 95% confidence interval (CI) to each study was plotted against the number of participants respectively. Relevant publications were retrieved and screened carefully. A total of ninety-six publications were identified, of which seventy-three irrelevant papers were excluded. As shown in Of the selected publications, one publication was written in Chinese The distributions of CYP1A1 Ile462Val genotypes as well as the genotyping methods of the included studies were presented in 2I = 42.1%), except for the homozygote comparison  and the recessive models . However, when subgroup analyses on ethnicity, clinical types and source of controls were further conducted, we found diminished heterogeneities in some of the subgroups under the dominant model.As shown in  and the recessive  models, indicating that individuals carrying the variant Val allele may have an increased acute leukemia risk compared with those bearing the wild-type Ile allele.The main results of the meta-analysis were listed in . In subgroup analyses regarding clinical types, increased risks for ALL were found under the homozygote comparison  and the recessive models , respectively. When the data regarding ALL were separated by age groups, significant increased risk could be observed in both adult ALL and childhood ALL. No significant associations were shown in subgroup regarding AML under the three genetic models . In subgroup analysis on source of controls, elevated risks were observed in the population-based subgroup  rather than the hospital-based subgroup .Considering the potential impact of the confounding factors on the overall results, we further conducted subgroup analyses. In the primary literature, only the detailed information on ethnicity, clinical types and source of controls were sufficient for analysis. Therefore, subgroup analyses on these issues were performed. In subgroup analysis according to ethnicity, increased leukemia risks were observed in the three genetic models among mixed ethnicities  but not Caucasians or Asians . In subg. In subgWhen the effect-models were changed, the significance of the overall data for the three models was not statistically altered (data not shown). Then, we discarded one study whose genetic distributions in controls exhibited significant deviation from HWE . Additionally, the data of the Egger’s tests also indicate the absence of the publication bias in the three models, respectively  , suggesting that the results of the meta-analyses are relatively stable and the potential publication bias might not have an evident influence on the results.Funnel plots were created to assess the publication bias. Then, Egger’s linear regression tests were used to assess the symmetries of the plots. The funnel plots seemed symmetrical for the overall data of the three genetic models . AdditioFor the overall data, the results showed that CYP1A1 Ile462Val might have a marked correlation with increased acute leukemia risk. Moreover, in subgroup analyses stratified by ethnicity, the data failed to reveal an association among either Asians or Caucasians, but mixed ethnicities. In subgroup analysis according to clinical types, the results indicated that Val/Val alleles might increase susceptibility to ALL.The relations of CYP1A1 Ile462Val variations with cancer risk have been evaluated by several meta-analyses. CYP1A1 Ile462Val polymorphism might have a correlation with increased risks of lung cancer, cervical cancer, colorectal cancer, esophageal cancer and breast cancers In the subgroup analysis according to ethnicity, significant increased leukemia risk was found in the mixed races subgroup but not Asian and Caucasian subgroups, suggesting that CYP1A1 Ile462Val genetic variation may not confer acute leukemia risk among either Asians or Caucasians. Nevertheless, since we could not obtain the data regarding separate ethnicities from the mixed ethnicities subgroup, the possible effects of ethnicity variations on the results could not be precisely evaluated. Evidence suggested that gene polymorphisms could result in ethnic-specific susceptibility to leukemia In the subgroup analysis stratified by clinical types, increased risk for ALL was shown under the homozygote comparison and the recessive models, suggesting that the homozygous Val/Val allele carriers might have an increased ALL risk relative to those of the wild-type Ile carriers. When the data about ALL were further divided by age groups, similar results were obtained in the two subgroups. However, only one study provided the information about adult ALL; thus, the results should be interpreted with care. As for AML, no associations could be observed. The disparity may be due to the different precise mechanisms involved in the genesis of different types of leukemia. ALL is a common childhood leukemia with poor outcomes Smoking and alcohol consumption are also important risk factors for leukemia. Evidence indicates that maternal smoking and drinking prior to and during pregnancy may increase risk of childhood leukemia, particularly ALL In the subgroup analysis according to source of controls, significance increased leukemia risk were observed in the population-based subgroup but not the hospital-based group. Since hospital-based controls might not be always truly representative of the general population, any biases might exist and the results should be interpreted with care. Therefore, use of proper control participants with strict matching criteria and large sample sizes are important for reducing such selection bias in future investigations.In the present meta-analysis, both Q-tests and I-squared values were used to assess the heterogeneities. Evident between-study heterogeneities for overall data were observed in the dominant genetic model, and thus the random-effect models were utilized in this model. However, as shown in Publication bias is an important factor that should be considered in a meta-analysis. We used funnel plots to evaluate the potential publication bias. Then, Egger’s linear regression test was also used as an approach for assessment of the symmetries. The results failed to suggest evident bias in the three genetic models, suggesting little influences of the bias on the results and demonstrating the robustness and credibility of the present meta-analysis.Several limitations should be addressed. First, in this meta-analysis, the primary articles only provided data about Caucasians, Asians and mixed ethnicities. Separate data regarding other ethnicities such as African should be concerned. Second, only studies written in English and Chinese were searched and included in this meta-analysis. Thus, any selection bias should be noted. Third, subgroup analyses regarding age, gender and other factors such as smoking, drinking and radiation exposure have not been conducted in the present study because relevant sufficient data were not available in the primary articles. Furthermore, among the fourteen included studies, only three studies provided the adjusted ORs In summary, the results of the present meta-analysis suggest that variant Val allele of CYP1A1 Ile462Val polymorphism might have an association with excess acute leukemia risk. Moreover, subgroup analyses indicate that homozygous Val/Val might modify the susceptibility to ALL. Further well-designed investigations in view of the confounding factors are needed reach a more convincible conclusion.