唐古特红景天干预高原性肺动脉高压药效物质及多靶点作用机制研究

Sustained pulmonary vasoconstriction and proliferation of pulmonary arterial smooth muscle cells (PASMCs) as experienced at high altitude can lead to high-altitude pulmonary hypertension (HAPH). In preliminary studies, Bioactive fraction of Rhodiola algida could induce rat pulmonary artery vasorelaxation by NO-cGMP- PKG-BKCa pathway and the opening of K+ channels (BKCa and KIR) in vitro. Additionally, Administration of bioactive fraction prevented pulmonary arterial hypertension by alleviating mPAP and right ventricular hypertrophy index based on decreasing cyclin D1, CDK4 expression level and inhibiting p27Kip1 degradation. And constituents were identified using a UHPLC-Q-TOF-MS/MS method..Therefore, the strategy of “bioactive equivalent combinational components” will applied to screening active compounds based on chemical family classification on models of pulmonary arterial rings and pulmonary arterial smooth muscle cells proliferation induced by hypoxia. And uncovering pharmacologically active compounds based on individual compounds and their interaction. Then, dominant active compounds combination will designated based on activity contribution assay. And multi-target mechanism of dominant active compounds combination would be illuminated based on NO-cGMP- PKG-BKCa pathway and K+ channels and protein (cyclin D1, CDK4 and p27Kip1) expression level. It would be a unique natural template in discovery, development and design of multi-compounds and multi-targets innovative drug against high-altitude pulmonary hypertension (HAPH).

高原低氧环境下，持续的肺血管收缩和肺动脉平滑肌细胞增殖能够诱导高原性肺动脉高压的发生和发展。本课题组在前期研究中发现，唐古特红景天有效部位群在体外可以通过NO-cGMP- PKG-BKCa 通路和开放钾离子通道，使肺小动脉血管发生舒张，在体内，能明显抑制肺动脉压的升高及肺动脉血管的重构作用，发现有效部位群能抑制低氧诱导的Cyclin-D1和CDK4蛋白的表达及p27kip1蛋白的降解，从而使细胞周期阻滞于G0/G1期，抑制肺动脉平滑肌细胞的增殖，从而减轻血管重构，并明确了其中的化学组成。.本项目基于有效成分等效群策略及体外肺小动脉血管换模型及低氧诱导的肺动脉平滑肌细胞模型，通过阐明各组分间的交互作用及对整体活性的贡献度，表征出发挥药效学的主要药效物质，并确定主要药效物质对NO-cGMP- PKG-BKCa 通路和开放钾离子通道的活性及对Cyclin-D1、CDK4和p27kip1蛋白的调节

高原低氧性肺动脉高压(HAPH)是人体对高原低压性缺氧失代偿所引起的特发性慢性高原病之一，严重者可发展为右心衰竭而死亡。.本项目利用低氧诱导的肺动脉平滑肌细胞(PASMCs)增殖模型及低氧诱导的肺血管内皮细胞(PAECs)损伤模型，明确了唐古红景天活性部位主要活性成分。并发现香草酸、对香豆酸、红景天素、红景天苷、圣草酚、没食子酸乙酯对精氨酸酶的非竞争性抑制活性。以量丰活性成分为研究对象，获得由没食子酸、香草酸、对香豆酸、咖啡酸、阿魏酸、红景天苷、圣草酚、山奈酚、槲皮素、异槲皮苷、表儿茶素、木犀草素等12个化合物组成的等效成分群，有效改善HAPH大鼠的平均肺动脉压力的升高、肺血管及右心室的重构。.明确了主要活性成分木犀草素通过抑制HIF-2α-Arg轴、调节PI3K-AKT-eNOS来改善HAPH大鼠肺血管内皮功能，并通过上调钾离子通道蛋白KV1.5的表达，平衡肺动脉平滑肌细胞的凋亡和增殖，进而改善肺血管的重构作用。发现主要活性成分山奈酚通过抑制Akt/GSK3β信号通路，改善HAPH大鼠肺血管的重构。确定了精氨酸酶抑制剂香草酸能够通过调节Arg2及eNOS平衡，促进NO的合成，保护肺血管内皮功能，进而抑制肺血管的重构及纤维化作用。并通过ROCK信号通路改善肺动脉高压大鼠的右心结构与功能。.通过本项目实施，进一步将其药效物质明确到12个主要化学成分中，并进一步明确了主要活性成分的关键靶蛋白，为阐明其多组分-多靶点作用机制，提供了科学依据，并为进一步设计、优化更加有效、低毒的“多组分-多靶点”复方组合药物提供了研究模板。为保障青藏高原地区的人居健康及国防安全提供高效、低毒的药物干预具有深远意义。