甘油三酯代谢相关基因与冠心病的关联分析及功能研究

Disease caused by atherosclerosis (AS), especially coronary heart disease (CHD), is a major factor of mortality in China. AS is a condition in which an artery wall thickens as a result of the accumulation of cholesterol. The relationship of plasma triglyceride (TG) concentration to CHD is uncertain. In recent years, hypertriglyceridemia is re-emerging as an important predictor of CHD. Because AS is a chronic disease that remains asymptomatic for decades, carotid intima-media thickness (IMT) and pulse wave velocity (PWV) are applied to evaluate the degree of AS. The overall objective of the proposed study is localize and identify new variants associated with IMT, PWV, TG levels or the risk of CHD and determine biological mechanism on the development of AS. .In present project, 5000 CHD cases and controls are enrolled. A total of 2000 volunteers who have no history of cardiovascular disease from community in Beijing are also included. Phenotypes including IMT, PWV and lipid levels were examined from each volunteer at baseline and will be collected again during follow-up in 2013-2015 year. Using the database from NCBI, HapMap, miRBase and miRanda, we select related genes implicated in TG metabolism and genotype tagSNPs and SNPs in 3′untranslated regions with the potential to either disturb or create microRNA-target on ABI 7900 fast real-time PCR system. .Toward achieving our overall objective, we will pursue the following specific aims: .Specific Aim 1: We select the related 22 genes encoding enzymes regulating TG synthesis or lipolysis, including glycerol-phosphate acyltransferase, acylglycerolphosphate acyltransferase, diacylglycerol acyltransferase, monoacylglycerol acyltransferase, lipases, PAT family, lipins. We will analyze the relationships of each tagSNP with IMT, PWV and the risk of CHD, and find novel loci associated with AS and CHD..Specific Aim 2: Using programs of CART and MARS, we will detect the interactive effects of gene-gene and gene-environmental factors on the different phenotypes of AS, and evaluate the effects of these genetic variations on the development of AS..Specific Aim 3: Genetics mechanisms of these SNPs will be explored using molecular biology experiment. Luciferase reporter constructs containing different alleles or haplotypes of SNPs significantly associated with AS/CHD were evaluated for their effects on luciferase activity in cell lines. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecitation (ChIP) were performed to determine the nuclear factors that might bind to oligonucleotides corresponding to the genomic sequence of different alleles of loci associated with AS/CHD. We will select about 200 subjects with three different genotypes of important loci and compare levels of concerned mRNA from peripheral blood mononuclear cells to determine the influence of SNPs on the expression of related genes. Finally, we synthetically evaluate the effects and mechanisms of genetic variations and AS/CHD.

以冠心病(CHD)为主的动脉粥样硬化(AS)性疾病已成为我国人群的重要死因之一。血浆甘油三酯(TG)水平升高是CHD 的重要预测因子。利用5000例CHD病例对照个体和2000例前瞻性社区人群，选择22个TG 代谢相关基因(GPAT、AGPAT、DGAT、MOGAT、脂肪酶、PAT、lipin等家族)的标签SNP及潜在影响miRNA结合的SNP，用TaqMan 技术进行基因型鉴定。分析多态与CHD 及颈动脉内膜中膜厚度、脉搏波速度及血脂水平的关系，发现新的易感位点。采用CART和MARS分析基因-基因之间、基因-环境危险因素之间的交互作用，评价基因多态与CHD及AS亚临床表型和中间表型的关系。选择重要位点不同基因型个体共200例，比较目标mRNA的表达水平，分析多态对基因表达水平的影响。结合报告基因研究、蛋白质结合因子研究，探讨易感位点的分子机制。综合评价TG代谢相关基因变异与AS的关系。

以冠心病为主的动脉粥样硬化性疾病已成为我国人群的重要死因之一。大量研究证实，脂代谢紊乱是动脉粥样硬化的主要危险因素，血浆甘油三酯水平升高是冠心病的重要预测因子。甘油三酯代谢相关基因是脂质代谢紊乱的重要候选基因，可能在冠心病发生发展中起重要作用。.队列人群随访：于2013-2014年对北京首钢和石景山社区队列人群随访调查，除问卷调查、体格检查、实验室生化指标检测外，还测定了脉搏波速度（PWV），并获取了颈动脉内膜中膜厚度（IMT）及颈动脉斑块情况、血管内皮功能等数据。并收集了研究对象的外周血单核细胞。.候选基因与冠心病关联：在冠心病病例对照个体中完成了甘油三酯代谢相关基因标签单核苷酸多态（SNPs） 的基因型鉴定。并发现ADM基因标签SNPs与血脂水平呈线性关联关系。肾上腺髓质素（ADM）基因常见多态与中国人群血脂水平关联，提示ADM基因可能参与调节脂质代谢，从而影响我国人群的血脂水平。.SIRT1基因功能研究：首次探索了SIRT1（沉默交配型信息调节因子2同源体1）基因表达与冠心病的关系，发现在冠心病患者中SIRT1基因mRNA表达下降，提示SIRT1基因在人群具有心血管保护作用。首次发现SIRT1基因rs3758391可能通过改变转录因子结合位点调节SIRT1基因的表达，进而影响冠心病的发生发展。.GALNT家族基因功能研究：探讨GALNT（多肽:N-乙酰氨基半乳糖转移酶4）家族中GALNT2和GALNT3基因与冠心病的关系，发现GALNT3基因多态与冠心病关联。GALNT3基因通过调控p38/MAPK信号通路及基质金属蛋白酶2（MMP-2）和14（MMP-14）的表达导致内皮细胞损伤，参与动脉粥样硬化的发生。研究首次揭示了GALNT3基因在冠心病中的作用及可能机制。.TUG1基因功能研究：与对照组相比，长链非编码RNA（LncRNA）TUG1(taurine up-regulated 1)基因在冠心病患者中出现高表达。体外实验结果显示，敲低TUG1能抑制内皮细胞凋亡，降低IL-8的表达水平。提示TUG1可能影响血管内皮细胞功能，参与冠心病的发生发展过程。