线粒体融合蛋白1/2（Mfn1/2）在缺氧肺血管稳态失衡、重构中的作用及分子机制

Vascular homeostasis and remodeling is a key scientific problem of the malignant cardiovascular diseases, including pulmonary arterial hypertension. The pathophysiology and the mechanism of the disorders are incompletely understood. We and other have discovered that mitochondria and mitofusin-2 involved in the hypoxia-induced proliferation and remodeling of pulmonary arterial smooth musle cells（PASMCs）. However, the unifying mechanism for the anti-apoptosis and proliferation of Mfn2 in PASMCs are not yet well defined, and the role of Mfn1 has not been reported in pulmonary circulation. In addtion, how hypoxia regulation of Mfn1/2 is not well understood We hypothesize that hypoxia-induced functional and morphylogical changes and signal transduction abnormity are mediated by Mfn1/2, which subsequently regalates cell circle and phenotypes of PASMCs, leading to PASMCs proliferation and pulmonary vessel remodeling. We propose 3 specific aims to test the hypothesis: (1) to examine the influence of hypoxia on the expression of mitofusin-1/2; the role of Mfn1/2 on mitochondrial morphology and function as well as PASMC proliferation and vasculau remodeling under hypoxic condition, (2) to identify the signal transduction pathways, protien-protien interaction network and key protiens involved in mitofusin-1/2 induced PASMC proliferation, and (3) to demonstrate the role of microRNAs in the regulation of mitofusin-1/2 and the underlying molecular mechanisms. This project will significantly increase our understanding of the role of mitofusin-1/2 on hypxia induced imbalance of vascular homeostasis and remodeling, and is likely to shed insight into a novel mechanism of hypoxia induced PASMC proliferation，and develop a poterntial therapeutic strategies for interference of the disorder. Furthermore, the study may provide the basic data to reveal the mechanism of pulmonary vascular homeostasis and remodeling for the on going key research project of National Natural Science Foundation of China.

血管稳态失衡与重构是重大心脑血管疾病，包括肺动脉高压的共性核心科学问题，其发生的病理生理学基础和机制不完全清楚。我们和他人发现线粒体融合蛋白2（Mfn2）与缺氧肺血管平滑肌凋亡、增殖和重构有关，但Mfn2作用的分子机制，Mfn1在肺循环中的作用、缺氧如何调控Mfn1/2未见报告。我们假设缺氧通过microRNA调控Mfn1/2，使线粒体形态、功能和信号转导系统等异常，导致肺血管增殖和重构。通过下述研究证明该假设：1）缺氧对Mfn1/2表达的影响、Mfn1/2对线粒体形态、功能的影响及在缺氧肺血管重构中的作用；2）Mfn1/2调控缺氧肺血管重构的信号转导系统、调控网络和关键分子；3）miRNAs在缺氧调控Mfn1/2中的作用和分子机制。本申请将明确Mfn1/2在缺氧肺血管稳态失衡，重构中的作用和机制;发现治疗肺血管重构相关疾病的靶点；为重大项目总体目标提供肺循环血管稳态和重构调控的关键数据。

《线粒体融合蛋白1/2（Mfn1/2）在缺氧肺血管稳态失衡、重构中的作用及分子机制》课题获得国家自然科学基金资助，课题负责人朱大岭。已完成超预期的研究目标，研究内容超额完成，发表14篇SCI文章，部分结果发表在肺动脉高压领域有国际影响力的杂志上。本课题组前期研究发现缺氧上调线粒体融合蛋白Mfn2表达，通过PI3K-AKT通路介导肺动脉平滑肌细胞凋亡。而作为肺血管重构的重要环节——线粒体功能对血管稳态失衡的调控机制还需进一步探讨。在本项目资助下，继续深入研究线粒体功能对缺氧条件下肺血管稳态的影响，研究发现: 1.缺氧诱导肺血管及肺动脉平滑肌细胞线粒体形态及功能改变，上调Mfn1表达，揭示缺氧对线粒体超微结构及线粒体融合/分裂动态过程的影响。2. 缺氧通过Mfn1调控肺血管稳态失衡和血管重构。3. 缺氧通过miR-125a介导Mfn1加速肺动脉平滑肌细胞周期，调控细胞增殖。4. miR-125a肺靶向治疗能够改善缺氧引起的线粒体功能失衡，减少肺血管稳态失衡及降低肺动脉高压形成。研究结果支持研究假设线粒体融合蛋白参与缺氧引起的肺血管重构。同时，在项目的开展过程中，本课题组还完成超预期的重大拓展研究：1.在肺动脉高压状态下，线粒体分裂蛋白Drp1以及具线粒体分裂活性的p-Drp1表达上调，Drp1发生胞浆向线粒体的过度转位活化；2.平滑肌特异性Drp1表达缺陷，能逆转肺动脉高压的各项临床指标，阻止肺动脉高压进程；3.Drp1通过调控线粒体形态和功能介导肺动脉平滑肌细胞表型转变，引起细胞病理性增殖，该调控作用依赖于ERK信号转导系统；4.缺氧通过下调miR-449a-5p诱导靶基因Myc的表达，介导转录因子Myc与Drp1的相互作用，调控Drp1的表达与活化；5. miR-449a-5p/Myc通路在缺氧诱导Drp1介导线粒体功能损伤诱导肺动脉平滑肌表型转变，导致肺血管稳态失衡及稳态失衡中发挥重要作用。以上研究结果证实线粒体在肺动脉高压血管重构中的作用，发现了参与肺血管重构的线粒体融合分裂蛋白及新的作用机制，并提出了缺氧调控线粒体稳态的具体环节和分子机制，为完善肺动脉高压的发病机制研究及治疗提供理论依据。