The Association Study On Genetic Polymophisms Of ApoB, ApoC-Ⅰ And LDLR With Coronary Heart Disease

Objective To establish polymerase chain reaction and high resolution melting(PCR-HRM) assays for genotyping the three single nucleotide polymorphisms(SNP) of rs693 in apolipoproteins B(Apo B) gene, rs688 in Low-density lipoprotein receptor(LDLR) gene and rs4420638 in apolipoprotein C-I(Apo C-Ⅰ) gene, and then to investigate the relationship of the three SNPs with the risk of coronary heart disease(CHD) development and the blood lipid levels.Methods The molecular diagnosis methods of PCR-HRM were established by designing the primers of the three SNPs of rs693, rs688 and rs4420638. The accuracy of the developed methods was verified using direct sequencing of the amplicons. The gene polymorphisms of the three SNPs(rs693, rs688, rs4420638) were determined by PCR-HRM. A case-control study was performed in 311 CHD patients and 300 matched healthy controls to analyze the correlation of the three SNPs with the susceptibility of CHD. In addition, It is investigated of the relationship between the tree SNPs and the blood lipid levels in 462 subjects.Result(1) The establishment of the PCR-HRM detection method: The PCR-HRM methods genotyping for the three SNPs of rs693, rs688 and rs4420638 were successfully established and tested the 1073 clinical specimens(CHD group 311 cases, healthy control group 300 cases, blood lipids group 462 cases), so as to achieve the fast genotyping for the three SNPs.(2)CHD susceptibility analysis: 1The three SNPs of rs693, rs688 and rs4420638 were accord with Hardy Weinberg equilibrium in healthy control group(?2:3.055, 2.804, 1.151; P>0.05). 2The allele and genotype frequencies of rs693 and rs4420638 were significantly different between CHD cases and the controls(rs693:?2=8.167, P=0.004; ?2=8.836, P=0.003; rs4420638:?2=8.664, P=0.003; ?2=9.512, P=0.002). 3After adjusting for age, sex and BMI, the multiple logistic regression analyses using an additive model revealed that the gene polymorphism of rs693 and rs4420638 were significantly associated with the susceptibility of CHD development in Chinese Han population. Compared with rs693 CC genotype, the genotype CT of rs693 confers to the low risk of CHD development(OR=0.448, 95%CI: 0.246~0.817, P=0.009); compared with rs4420638 AA genotype, the genotype AG of rs4420638 confers to the high risk of CHD(OR=2.140, 95%CI: 1.241~3.688, P=0.006). 4The haplotype analysis of the two SNPs of rs688(C>T) and rs4420638(A>G) showed that haplotype CG increases the risk of CHD development(OR=1.715, 95%CI: 1.091~2.697, P=0.018).(3) Association analysis of the blood lipid levels: 1The one-way analysis of variance(ANOVA) was used to analyze the relationship between mutations genotype and the blood lipid levels. The results showed that the total cholesterol(TC) levels and low density lipoprotein cholesterol(LDL-C) levels were significantly different in genotypes of rs688(P<0.05); The TC levels and triglyceride(TG) levels were significantly different in genotypes of rs693(P<0.05); the TG levels was significantly different in genotypes of rs4420638(P<0.05). 2Under dominant model, the multiple linear regression analyses was tested by taking the TC, TG, LDL-C and HDL-C as the dependent variable while taking the gender, age, BMI, rs688, rs693 and rs4420638 as independent variables. The results showed that the TC levels and LDL-C levels of rs688 T allele carriers were significantly higher than homozygous type(β=0.301, P<0.001; β=0.335, P<0.001); the TC levels and TG levels of rs693 T allele carriers were significantly lower than homozygous type(β=-0.250, P=0.027; β=-0.162, P=0.029); the TG levels of rs4420638 T allele carriers were significantly higher than homozygous type(β=0.180,P=0.016).Conclusion The PCR-HRM genotyping of SNPs rs688, rs693 and rs4420638 assays was successfully established; the two SNPs of rs693 and rs4420638 and the haplotype CG are significantly associated with the susceptibility of CHD development in Chinese Han population; the present study suggested that SNPs rs688, rs693, rs4420638 were significantly associated with blood lipid levels in Chinese Lan Zhou Han population.