冠心病相关线粒体tRNAThr 15927G>A突变的致病机制研究

Coronary heart disease is a complex disease caused by multi-factors including environment and inherited risk factors. Our previous studies identified the mitochondrial tRNAThr 15927G>A mutation associate with coronary heart disease. To further understand the pathophysiology of maternally transmitted coronary heart disease, we will construct cybrids by the fusion of R6G-treated human umbilical cord vein epithelial cells and enucleated lymphocyte cells carrying mitochondrial tRNAThr 15927G>A mutation, examine the mitochondria and cell dysfunctions caused by this mutation in the same nuclear genetic context. Coronary heart disease patient specific iPSCs will be generated from lymphoblastoid cell lines, and subsequently directed to differentiate into vascular endothelial cell. Then, we will examine the effects of mitochondrial tRNAThr 15927G>A mutation on the process of iPSCs induction, characteristics of iPSCs and the function of mitochondria. Thus, the success of this project will help to understand the pathogenesis of mitochondrial tRNAThr 15927G>A mutation related coronary heart disease, provide new perspectives for the research of coronary heart disease pathology and therapy.

冠心病是一种复杂性疾病，由遗传因素、环境因素或两者相互作用所致。在冠心病发生的诸多危险因素中，遗传因素起着十分重要的作用。我们在前期工作中发现一个呈母系遗传特征的冠心病家系，并发现此家系中线粒体tRNAThr 15927G>A突变是冠心病的致病因素之一。本项目拟在此基础上构建转线粒体人脐带静脉内皮细胞系，检测相同核基因背景下线粒体tRNAThr 15927G>A突变对人脐带静脉内皮细胞线粒体功能和细胞功能的影响。诱导携带线粒体tRNAThr 15927G>A突变的冠心病患者特异性iPSCs，并将其定向分化为血管内皮细胞。评估该突变对iPSCs诱导过程及iPSCs干性的影响，检测该突变对血管内皮细胞线粒体功能和细胞功能的影响，从分子和细胞层面探讨冠心病线粒体tRNAThr 15927G>A突变在冠心病病理过程中的作用，为冠心病的致病机制和治疗方法研究提供新的视角。

心血管疾病是全球范围内造成死亡和经济负担最严重的非传染性疾病，而冠心病是患病率和死亡率最高的心血管疾病。线粒体tRNAThr 15927G>A突变可能是冠心病的致病因素之一。为了进一步研究该突变在冠心病发生过程中的作用，我们构建了携带15927G>A突变的转线粒体HUVEC细胞系；同时诱导携带线粒体tRNAThr 15927G>A突变的永生化淋巴细胞系为iPSCs，并将其定向分化诱导为血管内皮细胞。通过对所构建血管内皮细胞模型线粒体功能和细胞功能的检测发现，线粒体tRNAThr 15927G>A突变破坏了tRNAThr反密码子臂的28C-42G碱基对，影响tRNA的结构和稳定性。导致tRNAThr在非变性条件下电泳迁移速率降低，稳态水平和氨酰化水平分别降低23.82%和31.74%。线粒体tRNAThr结构和功能的异常进一步导致线粒体蛋白质合成水平下降，氧耗速率降低，活性氧水平增加，线粒体膜电位降低。对细胞功能的检测发现，突变细胞凋亡水平增高，体外迁移和血管生成能力下降。本研究进一步探究了线粒体tRNAThr 15927G>A突变在冠心病发生过程中可能起到的作用，并为线粒体基因突变相关或血管内皮细胞相关疾病的研究提供了新的细胞模型和研究思路。