| BackgroundsIschemic stroke (IS) is a chronic disease influenced by environmental and genetic factors. Gene-environmental interaction are closely related with the susceptibility of IS. With the completement of Human Hapmap, it has become the focus in medical fields to study the pathogenesis of IS from the view of candidate Gene and whole-genome. In recent years, more and more studies have suggested that Inflammation play important roles in IS and coronary heart disease (CHD). However, studies on the association of the inflammatory cytokines and the interaction between the gene polymorphisms and environment exposures with IS are rare presently in the Han population.Objectives1. To explore the association of the G-174C, G-572C polymorphisms of IL-6 with the susceptibility of IS.2. To explore the association of the G-238A, G-308A polymorphisms of TNF-αwith the susceptibility of IS.3. To explore the association of the VNTR polymorphisms of IL-1RN and the VNTR with the susceptibility of IS.4. To explore two key SNPs on chromosome 12p13 with the susceptibility of IS.5. Classification tree model and Multifactor dimensionality reduction were applied to explore the possible interaction between gene and environment and build the risk model for IS. MethodsCase collection was made according to the diagnostic criteria of TOAST, two 1:1 matched hospital- based case-control studies were performed. We enrolled inpatients attending the stroke units of five large general hospitals in Shenzhen, Heinongjiang and Xinjiang from September 2003 to February2009. A total of 748 subjects presenting within 24h of symptom onset were enrolled in two case–control studies. Additionally, 748 age-, gender- and ethnicity-matched normal healthy controls were randomly selected from healthy volunteers from four local community-based populations. A structured questionnaire was used to record general information, clinical history of IS and associated clinical parameters, and epidemiological data. Genotypes were examined by using Taqman Primers and a novel tailed primers protocol. Logistic regression models were used to study the association between the polymorphisms and ischemic stroke. Classification tree model was applied to build up the risk model for IS, multifactor dimensionality reduction and Haplotype analyses were used to assess the possible interaction between gene and environment.Results1. Univariate logistic regression showed that traditional risk factors of IS including education, income, BMI, smoking, WHR,hypertension, diabetes, negative events and TG. In two multivariate logistic models, WHR, smoking, Diabetes, and Hypertension were positively associated with IS, with OR=2.158 (95%CI: 1.554~2.997), OR=2.777 (95%CI: 2.084~3.699), OR=1.519 (95%CI: 1.107~2.086), OR=3.143 (95%CI: 2.445~4.042) respectively, while tea-drinking was inversely associated with IS, with an OR of 0.347 (95%CI: 0.269~0.448) in the Hans. However, we only find WHR and Hypertension were independent risk factors of IS with OR=2.158 (95%CI: 1.554~2.997) and OR=2.777 (95%CI: 2.084~3.699) respectively. 2. For IL-6 G-572C polymorphisms, significant difference was found in the distributions of genotypes between two populations. The G allele of the promoter single nucleotide polymorphism (SNP) G-572C was more common in IS subjects than controls (P = 0.004, corrected for multiple testing) in the Han population. GC carriage therefore increased the risk of IS in the Han ethnic group (OR 1.45, 95% CI 1.13–1.86). However, GG carriage decreased the risk of IS in the Uyghur ethnic group (OR= 0.36, 95% CI:1.19-1.88;P<0.05). The G allele was also less common in IS subjects than controls (OR= 0.67, 95% CI:0.45-0.99;P<0.05) in the Uyghur population.3. No significant difference was found in the association between TNF-α?238G/A and IS in both ethnic populations. However, the result showed that carriage of the TNF-α?308GA was a decreased risk of IS in both Han and Uyghur populations (OR:0.453, 0.213). GA heterozygous may be an independent protective factor for IS in the Chinese Han and Uyghur populations.4. The distributions of both IL-4 and IL-1RN VNTR polymorphisms were not significantly associated with IS subjects after adjustment for risk factors. We, however, found that subjects with the 2R-2R haplotype had about two fold lower risk of IS than those with the 4R-2R haplotype (OR=0.504, 95%CI: 0.322-0.789, P=0.002).5. Significant allelic association was identified between rs11833579 and IS in the Han population (OR 1.27, 95% CI 1.08–1.49). The association remained more significant after adjusting for IS covariates in the Han population in a recessive model but not in the Uyghur population. The significant difference in genotypes frequency of Rs11833579 was found between the two ethnic controls (P=0.000), which indicated that there may be genetic variations between Chinese Han and Uyghur populations that may affect the risk of IS. One risk haplotype (A-G; OR 1.52, 95% CI 1.21-1.92) was identified in the Han population. However, the associations between rs12425791 and IS were insignificant in both ethnic populations.6. Classification regression smodel were applied to build the risk model for IS in two ethnic groups. The model had five stratums including seven explanatory variations in Han population. There were three stratums and three explanatory variations in Uyghur population model. Additionally, the indexes of the screening test were applied to assess the fitness of the model. The results showed that sensitivity and specificity were reasonable in both ethnic populations. There were interactions between gene and environment by multi-factor dimensionality reduction analysis.Conclusions:1. The traditional risk factors were the main risk for IS in the Han population and Uyghur population. 2. IL-6 G allele may be an independent risk factor for IS in the Chinese Han population. However, G allele may be protective factor for IS in the Uyghur population.3. TNF-α?308GA heterozygous may be an independent protective factor for IS in the Chinese Han andUyghur populations.4. Significant associations were not found between Single VNTR polymorphisms and IS after adjustment for risk factors. 2R-2R haplotype of IL-4 and IL-1RN is significantly associated with lower IS risk in Chinese Han population.5. The A allele of SNP rs11833579 on chromosome 12p13 may play a role in mediating susceptibility to IS in the Han Chinese population in a recessive model. The A-G haplotype is also significantly associated with higher IS risk in the Han Chinese population.6. There were interactions between gene and environment by multi-factor dimensionality reduction analysis; Classification tree model can reasonably predict the occurrences of IS.Innovations:The study was the first research based on signal transduction pathways associated with Inflammation and related functional polymorphisms in Chinese two populations. Meanwhile, the study was the first to explore the association between two key SNPs on chromosome 12p13 and ischemic stroke in Chinese two populations based on the American genome-wide association study results.The study was the first to apply a novel tailed primers protocol to identify the association of IL-4 and IL-1RN (receptor antagonist) gene variable number of tandem repeats with ischemic stroke in Chinese Han population.In addition, the study was the first to apply Classification tree and Multifactor dimensionality reduction model to build up the risk model for IS in Chinese two populations.
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