目的 探讨人蛋白原转化酶枯草杆菌蛋白酶9(PCSK9)基因I474V位点在冠心病(CAD)患者和健康人群中的分布特点以及与血脂水平、CAD发生、病变程度的相关性,为阐明CAD的遗传发病机理、患者危险性分级评估等提供实验依据.方法 采集经冠状动脉造影确诊的CAD患者血液标本288例,健康对照者血液标本300例,提取基因组DNA模板,应用双向等位基因特异性引物聚合酶链反应(Bi-PASA PCR)技术检测PCSK9基因I474V多态性位点,并用基因测序法验证结果.血清总胆固醇(TC)、甘油三酯(TG)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)水平用酶法测定.结果 PCSK9基因I474V多态性位点主要以Ⅱ和Ⅳ基因型存在.CAD组和对照组在I474V位点的等位基因频率、基因型频率分布差异无统计学意义(P＞0.05).CAD组和对照组之间TC、LDL-C、HDL-C水平差异具有统计学意义(P＜0.05),TC水平差异无统计学意义(P＞0.05).CAD组中Ⅱ和Ⅳ基因型之间TC、TG水平的差异具有统计学意义(P＜0.01),HDL-C、LDL-C水平差异无统计学意义(P＞0.05).与非I474V携带者比较,I474V携带者血TC、TG水平增高.Ⅳ基因型的冠状动脉病变支数明显高于Ⅱ基因型(P＜0.05).结论 PCSK9基因I474V多态性位点主要以Ⅱ和Ⅳ基因型存在,CAD患者PCSK9基因I474V位点的基因型与TC、TG水平升高相关联,同时与CAD病变严重程度存在一定关联.%Objective To investigate the distribution characteristics of the 1474V loci of proprolein convertase subtilisin/kexin type 9 (PCSK9) gene in coronary artery disease (CAD) patients and healthy subjects, study the correlation of blood lipid levels, the development of CAD and the severity of coronary lesions, and provide the reference to elucidate the pathogenesis and genetic risk assessment of CAD. Methods According to the results of coronary angiography, blood samples from 288 CAD patients and 300 healthy controls were collected. 1474 V polymorphism loci in PCSK9 gene was detected by bi-directional polymerase chain reaction (PCR) amplification of specific alkies (Bi-PASA PCR). The results were further confirmed by cycle sequencing. Serum levels of total cholesterol (TC) , triglyceride (TG), high-density lipoprotein cholesterol ( HDL-C) and low-density lipoprotein cholesterol (LDL-C) were measured by enzymatic methods. Results Ⅱ and Ⅳ genotypes were dominant in the I474V polymorphism loci of PCSK9 gene. No significant difference was found in allele frequencies of 1474V loci between CAD patients and healthy controls (P > 0.05). There was statistical significance in TC, LDL-C and HDL-C levels between CAD patients and healthy controls (P<0.05), and the TG level had no statistical significance (P>0.05). Serum levels of TC and TG (P<0.01), but not HDL-C and LDL-C (/0.05) were statistically different between Ⅱ and Ⅳ genotypes in CAD patients. Subjects with I474V polymorphism of PCSK9 gene had significantly higher serum levels of TC and TG than subjects without I474V polymorphism. The number of coronary lesions was more in patients with Ⅳ genotype than Ⅱ genotype with statistical difference (P<0. 05). Conclusions Ⅱ and Ⅳ genotypes are dominant in the I474V polymorphism loci of PCSK9 gene. The genotype of PCSK9 gene I474V polymorphism is correlated with serum increasing levels of TC and TG and the severity of coronary lesions in CAD patients.
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