心肌衰老中mtDNA甲基化的作用及分子机制

Increased cardiac vulnerability in aged heart is the leading cause of increased incidence of heart diseases and death among elderly population. Loss of mitochondrial homeostasis which includes the function, biogenesis, degradation and dynamics of mitochondria might be a major factor causing the degeneration of cardiac structure and function as well as the increase in cardiac vulnerability. However it remains unknown whether the mtDNA methylation alters during cardiac aging and affects mitochondrial homeostasis. Our preliminary data show that the level of DNMT1 in mitochondria was increased and the mRNA levels of mtDNA-encoded components of respiratory chain were decreased in aged mouse heart, indicating that the mtDNA methylation might regulate cardiac aging. Therefore we propose the hypothesis that mtDNA methylation elevates during cardiac aging, which then disturbs mitochondrial homeostasis and causes the degeneration of cardiac structure and function as well as the increase of cardiac vulnerability. In this study, we will draw the mtDNA methylation map during cardiac aging based on aging models at cellular and animal levels. By gene manipulation techniques, we will illustrate the molecular mechanisms by which mtDNA methylation regulates mitochondrial homeostasis and cardiac function. Moreover, we will verify our hypothesis by targeting mtDNA methylation in animal aging models and observe the changes of mitochondrial homeostasis and cardiac structure and function. This study shall reveal new mechanisms on how disturbance of mitochondrial homeostasis contributes to cardiac aging from a cutting-edge mitochondrial epigenetic view, and provide novel therapeutic targets for the prevention and treatment of aging-associated heart diseases.

衰老心脏心肌细胞易损性增加是老年人群心脏病发病和致死率增高的主要原因，而线粒体稳态(功能、生成、降解和动态变化)失衡可能是导致心肌结构和功能衰退及易损性增加的重要因素，但心肌衰老中mtDNA甲基化是否发生改变并影响线粒体稳态尚无报道。近期我们发现衰老小鼠心肌线粒体中DNA甲基转移酶1显著升高，同时mtDNA编码的呼吸链亚基mRNA水平显著降低，提示mtDNA甲基化可能调控心肌衰老。我们据此假设：心肌衰老中mtDNA甲基化水平增高，影响线粒体稳态，使心肌结构和功能失常及易损性增加。本课题将利用细胞和动物的衰老模型绘制心肌衰老中的mtDNA甲基化图谱；并采用基因操作技术阐明mtDNA甲基化影响线粒体稳态和心肌功能的分子机制；最后通过靶向调控mtDNA甲基化确认其影响线粒体稳态进而调控心肌衰老的研究假设。本课题有望从线粒体表观遗传学角度揭示心肌衰老的新机制，并为衰老相关心脏病的防治提供新靶点。

近年来表观遗传学在衰老及代谢紊乱相关疾病的研究中得到了广泛关注。线粒体作为细胞的能量工厂，其稳态失衡是导致衰老及代谢紊乱相关疾病的重要因素，但mtDNA甲基化是否影响线粒体稳态尚待阐明。在本课题中，我们发现mtDNA甲基化能够通过调控线粒体稳态，参与衰老相关心肌重塑及代谢紊乱疾病。本课题取得的主要研究进展如下：（1）明确mtDNA甲基化能够调控线粒体稳态。在棕榈酸诱导的ROS升高和线粒体稳态失衡模型中，发现DNMT1的线粒体定位增多，mtDNA多个区域的甲基化程度升高，同时mtDNA编码基因转录水平降低；线性相关分析发现mtDNA甲基化频率与mtDNA编码基因的转录水平以及线粒体复合物活性呈负相关；（2）发现mtDNA甲基化参与调控心肌衰老和衰老相关代谢紊乱。在高脂饮食诱导的代谢紊乱小鼠中得到单碱基分辨率的mtDNA甲基化图谱，在衰老小鼠和AngⅡ诱导的心肌肥大模型小鼠的心肌线粒体中发现DNMT1的线粒体定位增多，同时mtDNA编码基因转录水平降低；（3）明确参与mtDNA甲基化调控的主要分子为DNMT1，发现DNMT1过表达后其线粒体转位增多，通过调控mtDNA的甲基化水平进一步调节mtDNA编码基因转录水平及线粒体功能；（4）发现AMPK可能作用于DNMT1上游，通过促进DNMT1的线粒体转位调控mtDNA甲基化；（5）筛选调控mtDNA甲基化的功能分子，发现羟基酪醇可能通过调节mtDNA COX2基因的甲基化水平改善线粒体复合物Ⅳ的活性。.综上所述，本课题从线粒体表观遗传学角度揭示了线粒体稳态失衡参与心肌衰老和衰老相关代谢紊乱的新机制，并为衰老相关代谢综合征和心脏病的防治提供了新靶点，较好地丰富了衰老的自由基和线粒体理论。.