慢性低氧激活ADP/P2Y12信号通路调控平滑肌细胞增殖促进肺动脉高压机制研究

Hypoxia induced smooth muscle cells (SMC) proliferation contributes to pulmonary arterial hypertension (PAH) development. Platelet activation promotes SMC proliferation, however, the mechanism of how platelet is activated and contributes to SMC proliferation during PAH development is unknown. Our previous data demonstrated that the infiltration of platelet in mice lung of chronic hypoxia induced PAH (CH-PAH) was significantly increased. P2Y12, an important receptor for platelet activation, knockout suppressed vascular remodeling. P2Y12 receptor also expresses on SMC. Our previous study also found that hypoxia increased adenosine diphosphate (ADP) release from SMC in vitro, and ADP induced SMC proliferation. We hypothesize that hypoxia induced activated platelet releases many cytokines promoting to SMC proliferation; hypoxia also induces ADP release from SMC, which stimulates SMC proliferation via P2Y12 receptor. In the present study, we will use bone marrow transplantation to confirm the role of P2Y12 on platelet and SMC. Then we are going to explore the important role of ADP/P2Y12 signal pathway in PAH by using P2Y12 knockout mice in CH-PAH with pathologic analysis, flow cytometry and western blotting.

低氧诱导血管平滑肌细胞（smooth muscle cell, SMC）异常增殖促进肺动脉高压（pulmonary arterial hypertension, PAH）。血小板活化调控SMC增殖，但在PAH中血小板活化及其调控SMC增殖的分子机制不清楚。前期在低氧诱导小鼠PAH模型中发现，肺血管周围血小板浸润增多，敲除血小板活化重要受体-P2Y12抑制血管重塑；同时该受体也表达在SMC，我们体外低氧诱导SMC释放ADP，通过与P2Y12结合刺激SMC增殖。据此提出假设：低氧可诱导P2Y12受体介导血小板活化，释放多种介质，诱导SMC增殖；低氧还可诱导SMC分泌ADP，作用于表面P2Y12调控SMC增殖，促进血管重构，参与PAH发生。本研究拟应用P2Y12敲除鼠构建小鼠PAH模型，通过骨髓移植实验明确P2Y12受体发挥作用的细胞定位，利用病理、流式、分子生物学等实验技术证明该科学假设。

肺动脉高压（pulmonary arterial hypertension, PAH）是以肺血管阻力进行性升高及右心衰竭为主要表现的临床病理综合征或疾病，其发病率、致残率及死亡率高。血管SMC异常增殖在肺动脉高压血管重塑中至关重要，因此明确致病因素如何激活、如何引起血管SMC异常增殖，导致血管重塑，将为防治肺动脉高压提供重要理论依据。在本项目支持下，我们发现：1）通过持续4周低氧可稳定建立小鼠肺动脉高压模型，并且构建了经肋间肌穿刺测定小鼠右心室压力曲线的方法,保持动物胸腔密闭无需呼吸机辅助通气,简便快捷,能够快速评估小鼠右心室的血流动力学状态,本研究以共同作者发表文章；2）在低氧诱导的小鼠肺动脉高压模型中，外周血血小板活化水平及肺血管周围血小板浸润增多，利用血小板活化的受体P2Y12敲除鼠及骨髓抑制实验证实，骨髓来源的P2Y12主要参与低氧诱导小鼠肺动脉高压的发生及发展，其机制包括血小板活化介导炎症反应，同时促进肺动脉平滑肌细胞增殖，进一步利用P2Y12受体抑制剂氯吡格雷处理小鼠，可明显降低低氧诱导肺动脉高压的发生及发展，文章正在撰写中；3）在肺动脉高压发生过程中，miR-15a-5p表达上调，利用miR-15a-5p 类似物以及 anti-miR-15a-5p过表达与抑制miR-15a-5p，发现miR-15a-5p可通过抑制VEGF、促进 p-p38及 MMP-2表达，进而促进SMC凋亡、抑制SMC增殖，在PAH发生过程中起保护作用，本研究成果发表International Journal of Molecular Medicine。综上所诉，在本青年课题的资助下，研究了肺动脉高压过程中肺动脉平滑肌细胞增殖的调控机制，为疾病的预防和治疗提供新靶点和新思路。