Study On The Influence Of Gene Polymorphisms And Environmental Factors And Their Interactions On Metabolic Syndrome

Metabolic syndrome（MS）is a cluster of risk factors related to cardiovascular disease（CVD）and diabetes mellitus assemble,and it is becoming an increasingly serious health problem both in developing and developed countries.With the rapid development of economy in China,the prevalence of MS among residents,especially in large cities of developed regions,is going through the stage of rapid growth.The diagnostic criteria of MS haven’t been completely unified,which makes it difficult to compare the prevalence of MS directly in various regions.Currently,the lack of representative MS prevalence data based on the latest diagnostic criteria of MS（JIS standard）is a big issue.Although the pathogenesis of MS is still unclear,studies have shown that MS is determined by genetic and environmental factors,and there may be interaction between them.The relationships between environmental,genetic factors and MS are inconsistent,and above associations in Chinese have been rarely studied so far.In these studies,dietary factors were more concentrated in a single food or nutrient,and the studies of the interaction between genetic and environmental factors are warranted.Based on residents in the developed cities（Nanjing）of eastern China,a multi-stage stratified cluster random sampling method were used to explore the individual health status,physical activity,diet survey,physical examination and laboratory test.According to the diagnosis standard of JIS,the epidemiological characteristics of the MS and its influencing factors were investigated,and the association between single nucleotide polymorphisms（SNPs）and MS were studied by screening and determining SNPs using NCBI-dbSNP database and Haploview4.1 software,and the effects of interaction between environmental factors and gene polymorphism on MS were also discussed.The present thesis was divided into three parts:Chapter 1 Study on Epidemiological Characteristics and Environmental Risk Factors including Dietary Pattern of MSObjective:To investigate the epidemiological characteristics of MS in Nanjing residents and to explore the environmental risk factors of MS.Methods:The 13 districts of Nanjing were divided into three layers according to the economic level by using the method of multi-stage stratified cluster random sampling.2 districts were randomly selected from each layer,followed by 6 community neighborhood committees random selection from each district,and then 75households were randomly selected from each neighborhood committee,and all the family members were selected as the subjects for inquiry,dietary investigation,physical examination and laboratory testing.Multivariate unconditional logistic regression analysis was used to analyze the influencing factors of MS and its components for subjects aged 15 years and old.Based on the food frequency questionnaire,the dietary patterns were identified using factor analysis,the score of each dietary pattern factor was quarters（from low to high Q1,Q2,Q3 and Q4）,and the effects of dietary pattern on MS were analyzed by multivariate unconditional logistic regression.Results:A total of 5029 subjects aged 15 years or old were included in Nanjing.According to the diagnosis standard of JIS,the crude prevalence of MS was 31.0%,and the standardized prevalence rate was 21.9%,of which 24.3%was males,19.2%was females,and the male prevalence rate was higher than that of females（P<0.05）;23.1%was urban,19.6%was rural,and the urban prevalence rate was higher than that in rural areas（P<0.05）.The standardized prevalence rates of MS components were 28.5%for central obesity,35.0%for hypertension,29.3%for high TG,19.0%for HDL-C and 28.5%for hyperglycemia.Multivariate analysis showed that the risk factors for MS included living in cities（OR=1.344,95%CI=1.126-1.604）,older age(OR_{65 group}=10.658,95%CI=5.357-21.204),smoking（OR=1.222,95%CI=1.008-1.480）,long leisure sedentary time（OR=1.225,95%CI=1.044-1.437）,and increased BMI(OR _obesity=18.491,95%CI=14.887-22.969)and family history of hypertension（OR=1.164,95%CI=1.010-1.342）,while high education(OR_{college and above}=0.693,95%CI=0.515-0.932)was protective factor.Risk factors for MS in male population included living in cities（OR=1.598,95%CI=1.245-2.051）,older age(OR_{65 group}=6.366,95%CI=2.684-15.097),smoking（OR=1.307,95%CI=1.046-1.633）,drinking（OR=1.442,95%CI=1.044-1.992）,long leisure time（OR=1.358,95%CI=1.074-1.717）and BMI(OR_obesity=27.245,95%CI=19.365-38.331);Risk factors for MS in female population include living in cities（OR=1.303,95%CI=1.024-1.659）,older age(OR_{65 group}=18.042,95%CI=5.407-60.207),increasing BMI(OR _obesity=13.754,95%CI=0.323-18.324);High educational level(OR _{college and above}=0.514,95%CI=0.321-0.822)and high physical activity level（OR=0.763,95%CI=0.587-0.993）were protected factors.A total of 1746 subjects were included in dietary pattern analysis using the food frequency data,and three dietary patterns were extracted by factor analysis.In the Western meat pattern,subjects in the highest quartile had an increased risk of MS and central obesity compared with those in the lowest quartile(OR=1.595,95%CI=1.095-2.323,P_trend=0.018;OR=1.600,95%CI:1.158-2.211,P_trend=0.015,respectively);but there was no significant correlation between pastry beverage and traditional plant patterns and MS and its components（p>0.05）.Conclusion:The standardized prevalence rate of MS among residents over 15 years old in Nanjing was 21.9%,and the prevalence rate was high.The older age,living in urban areas,low educational level,smoking,long leisure time,increased BMI,and family history of hypertension were risk factors for MS.Among males,older age,living in urban areas,smoking,high alcohol intake,long leisure sitting time and increased BMI were risk factors for MS;Among females,older age,living in urban areas,low educational level,low physical activity level and increased BMI were risk factors for MS.The western meat pattern,which was dominated by livestock,poultry meat and aquatic products was associated with an elevated risk of MS and central obesity.Chapter 2 A Case-Control Study of Gene Polymorphisms and their Interactions with MS Objective:To study the effects of single nucleotide polymorphisms（SNPs）of ADIPOQ,PPARγAPOA5,APOC3,LEPR and CETP genes on MS risk in Nanjing Han population.Methods:Patients with MS met JIS diagnostic criteria were selected as the case group except the non-Han population from the dietary pattern analysis population,and those met one or more diagnostic criteria of MS components were excluded from the non-diagnostic group,while the remaining subjects were taken as the matching control group.A total of 506 chinese han subjects from Nanjing,including 253 MS patients（case group）and 253 control subjects（control group）were randomly matched according to sex and age（within 3 years of difference）through the Empowerstats software.According to the SNPs information from NCBI-dbSNP database,the tagSNPs of each gene was analyzed by Haploview4.1 software.Combined with the results of current epidemiological studies,a total of 19 gene loci which were inconsistent or possibly related to MS but not studied on Chinese population were selected.These loci were rs1801282,rs3856806 and rs292002（located in gene PPARγ）,rs2241766,rs1501299 and rs266799（located in gene ADIPOQ）,rs2854116,rs854117 and rs5128（located in gene APOC3）,rs662799,rs6511821,rs2072560 and rs2266788（located in gene APOA5）,rs1805096,rs1137101 and rs1137100（located in gene LEPR）,rs708272,rs1800775 and rs2897142（located in gene CETP）,respectively.Genomic DNA was extracted from peripheral blood leukocytes.SNPs were detected by the ligase detection reaction（LDR）and SNaPshot SNP typing methods.The genotype frequencies were tested for Hardy-Weinberg equilibrium（HWE）using goodness-of-fitχ² test.Differences in genotype distribution between two groups were compared byχ² test.The association between SNPs and MS risk in different genetic models was analyzed by multivariable Logistic regression after adjusting the environmental factors.Linkage disequilibrium（LD）and haplotype were performed using SHEsis program.The effects of interaction between different SNPs on MS were analyzed by generalized multifactor dimensionality reduction（GMDR）.Results:（1）The genotype distribution of all 19 loci in the control group and the case group conformed to Hardy-Weinberg equilibrium law（P>0.05）,and the sample genotype distribution was representative.Multivariate unconditional logistic regression analysis showed that,in the codominance model,the risk of MS in CC genotype carriers at rs2854116 locus was higher than that in TT genotype carriers（OR=2.219,95%CI:1.163-4.234）,TT genotype carriers at rs2854117 locus was higher than CC genotype carriers（OR=2.619,95%CI）.1.336-5.136),GG genotype carriers at rs5128 locus was higher than CC genotype carriers（OR=4.152,95%CI:1.651-10.442）,AG and GG genotype carriers at rs662799 locus was higher than AA genotype carriers(OR_AG=2.261,95%CI:1.892-3.672;OR_GG=2.502,95%CI:1.062-5.895),CT and CC genotype carriers at rs651821 locus was higher than TT genotype carriers(OR_CT=2.224,95%CI:1.327-3.729;OR_CC=3.411,95%CI:1.342-8.667),CT and TT genotype carriers at rs2072560 locus was higher than CC genotype carriers(OR_CT=1.741,95%CI:1.038-2.919;OR_TT=3.690,95%CI:1.195-11.396),GG genotype carriers at rs2266788 locus was higher than AA genotype carriers（OR=3.692,95%CI:1.107-12.319）,AA genotype carriers at rs708272 locus was lower than GG genotype carriers（OR=0.318,95%CI:0.155-0.652）,AC genotype carriers at rs1800775 was higher than AA genotype carrier（OR=1.848,95%CI:1.069-3.195）.In the dominant model,the association of SNPs at rs2854116,rs2854117,rs5128,rs662799,rs651821,rs2072560,rs2266788,rs1800775 loci with MS was statistically significant（P<0.05）,and the risk of MS was increased in secondary allele carriers.In the recessive model,the association of SNPs at rs2854117,rs5128,rs2072560and rs708272 loci with MS was statistically significant（P<0.05）.and the risk of MS in carriers with secondary alleles at rs708272 locus was reduced,while the risk of MS in carriers with secondary alleles at other loci was increased.（2）The correlations between elevated TG levels and gene polymorphisms of rs2854116,rs2854117,rs5128,rs662799,rs651821,rs2072560,rs2266788,rs708272,rs1800775 were statistically significant（P<0.05）;The correlation between elevated fasting blood glucose and gene polymorphisms of rs662799,rs651821,rs2072560 and rs2266788 was statistically significant（P<0.05）;and the correlation between elevated blood pressure and gene polymorphisms of rs2854117 and rs708272 was also statistically significant（P<0.05）;however the decrease of HDL-C level were significantly correlated with gene polymorphisms of rs2854116、rs1805096、rs708272 and rs1800775（P<0.05）.（3）The linkage disequilibrium among the three SNPs at rs2854117,rs2854116 and rs5129 sites of APOC3 gene were observed.T-C haplotype between rs2854117 and rs2854116and C-G haplotype between rs2854116 and rs5128 increased the risk of MS,and T-C-G haplotypes increased the risk of MS among three SNPs（OR=1.755,95%CI:1.317-2.339）.There were strong linkage disequilibrium among the SNPs at rs2266788,rs2072560,rs651821,rs662799 sites of APOA5 gene.Among them,G-T haplotype between rs2266788 and rs2072560,T-C haplotype between rs2072560 and rs651821,C-G haplotype between rs651821and rs6627991 increased the risk of MS;G-T-C haplotypes among rs2266788,rs2072560 and rs651821,and T-C-G haplotypes among rs2072560,rs651821 and rs662799 increased the risk of MS;G-T-C-G haplotypes significantly increased the risk of MS among four SNPs（OR=1.651,95%CI:1.201-2.270）.There was a linkage disequilibrium between rs1800775 and rs708272 of CETP gene,and the A-A haplotype decreased the risk of MS（OR=0.748,95%CI:0.583-0.960）.（4）The three-order intercation model of APOC3rs2854117,rs2854116and rs5129 was the best model,which may increase the MS risk（P=0.0107）.The four-order intercation model of APOA5rs2266788,rs2072560,rs651821 and rs662799 was the best model,which may increase the MS risk（P=0.0107）.The second-order intercation model（rs2854116-rs662799）between genes of APOC3 and APOA5 is the best model,which may increase the MS risk（P=0.0107）.The four-order intercation model（rs2854117-rs662799-rs708272-rs289714）of the three gene-gene interaction（APOC3,APOA5 and CETP）is the best model,which may increase the MS risk（P=0.001）.Conclusion:（1）In Nanjing Han population,the gene polymorphisms of rs2854116,rs2854117,rs5128,rs662799,rs6511821,rs2072560,rs2266788,rs708272 and rs1800775 were associated with MS.（2）The gene polymorphisms of rs2854116,rs2854117,rs5128,rs662799,rs651821,rs2072560,rs2266788,rs708272 and rs1800775 were associated with elevated TG level.The gene polymorphisms of rs651821,rs2072560 and rs2266788 were associated with elevated fasting blood glucose.The gene polymorphisms of rs2854117 and rs708272 were associated with elevated blood pressure.The gene polymorphisms of rs2854116、rs1805096、rs708272 and rs1800775 were associated with the decrease of HDL-C level.（3）The haplotype T-C-G form rs2854117,rs2854116 and rs5129（located in gene APOC3）may increase MS risk.The haplotype G-T-C-G form rs2266788,rs2072560,rs651821 and rs662799（located in gene APOA5）may increase MS risk.While the haplotype A-A form rs1800775 and rs708272（located in gene CETP）could decrease MS risk.（4）The interactions between some SNPs may increase MS risk,for instance,the interactions of rs2854116-rs2854117-rs51283（gene APOC3）,the interactions of rs662799-rs651821-rs2072560-rs2266788（gene APOA5）,the interactions between APOC3rs2854116-APOA5rs662799 and the interactions of APOC3rs2854117-APOA5rs662799-CETPrs708272-CETPrs289714.Chapter 3 The Influence of Interactions between Environmental Factors and Gene Polymorphisms on MSObjective:To discuss the influence of interactions between environmental factors and gene polymorphisms on MS.Methods:Using multiple logistic regression model,the interaction between environmental factors and gene polymorphisms was investigated.The environmental factors included smoking,drinking,leisure time physical activity,leisure sitting time,dietary patterns,family history of hypertension and diabetes,which were all classified to two levels.The odds ratio（OR）of environmental factors,gene factors and interaction were described as ORe,ORg and OReg,respectively.Results:There were significant interactions between family history of diabetes and SNPs of rs1801282（located in gene PPARγ）,rs662799and rs651821（located in gene APOA5）（P<0.05）,OReg>ORe×ORg.The interactions between leisure time physical activity and SNPs of rs662799,rs651821,rs2072560,rs2266788（located in gene APOA5）were significant（P<0.05）,OReg>ORe×ORg.Also,we have found that there were significant interactions between SNP of rs3856806（located in gene PPARγ）and western meat patterns,leisure sitting time（P<0.05）,OReg>ORe×ORg.The interactions between SNP of rs1800775（located in gene CETP）and smoking or drinking were significant（P<0.05）,OReg<ORe×ORg.Conclusion:The interactions between family history of diabetes and SNPs of rs1801282,rs662799 and rs651821 could influence MS in the form of positive multiplication.Similarly,the interactions between low leisure time physical activity and SNPs of rs662799,rs651821,rs2072560,rs2266788 could influence MS in the form of positive multiplication.Furthermore,the interactions of positive multiplication could also be found between SNP of rs3856806 and western meat patterns,leisure sitting time.In addition,the interactions between SNP of rs1800775 and smoking or drinking could influence MS in the form of negative multiplication.In summary,The prevalence rate of MS among residents over 15 years old in Nanjing was 21.9%.The risk factors of MS were older age,living in urban areas,low educational level,smoking,long leisure time,increased BMI,and family history of hypertension.In Nanjing Han population,APOC3 gene,APOA5 gene and CETP gene SNPs and the interactions of them may be related to MS.The partial loci SNPs of PPARγgene and APOA5 gene may have positive multiplication with family history of diabetes,low leisure time physical activity,western meat patterns,leisure sitting time on MS.