虎杖苷在小鼠冠状动脉缺血再灌注损伤中的保护作用及其机制研究

The traditional Chinese medicine monomer, Polydatin, has been reported to be protective against myocardial ischemia-reperfusion injury and the underlying mechanism remains unclear. RhoA/ROCK pathway is proved playing a central pathophysiological role in the coronary artery spasms. Our preliminary studies proved that that Polydatin could inhibit the L-type voltage-gated calcium channel (L-VGCC) but not the contraction of myocardial cells. Additionally, the increase of ROCK activity stimulated by phorbol esters (ROCK agonist) could be prevented by Polydatin in myocardial cells of neonatal rats. Our hypothesis is that Polydatin may dilate the coronary artery and improve the coronary blood supply via the inhibition of RhoA/ROCK pathway which is regulated by the L-VGCC. Techniques of electrophysiological recording and ultrasound usage in small animal combined with the cell culture, measurement of contractility of arterial ring, isolated perfused heart assay and ischemia-reperfusion animal model are used to investigate the protection mechanism. Thus this project is to find out the underlying protective mechanisms from ischemia-reperfusion injury by Polydatin and the theoretical basis and laboratory evidence for its clinical application in myocardial infarction.

研究表明虎杖苷在缺血再灌注损伤中有扩张冠状动脉、提高心功能的保护作用，但其具体机制不明。 RhoA/ROCK通路的激活是缺血再灌注损伤中冠状动脉痉挛收缩最主要的机制之一。本项目组前期研究表明，虎杖苷可抑制L-型Ca2+通道但不减轻心肌收缩力，并且可抑制由佛波酯（ROCK激动剂）刺激引起的ROCK活性的增高。因此，我们认为在缺血再灌注损伤中，虎杖苷可能通过抑制L-型Ca2+通道，调节RhoA/ROCK通路，达到缓解冠状动脉平滑肌痉挛的保护作用。本项目拟采用在细胞培养及血管环模型基础上，利用电生理和分子生物学技术，阐明虎杖苷缓解平滑肌细胞收缩的具体机制；并在离体心脏缺血再灌注模型基础上，利用Powerlab软件分析仪分析虎杖苷对血流动力学的影响；最后利用动物模型，结合小动物超声技术，探明虎杖苷对载体动物心功能的保护作用。该项目的研究为虎杖苷改善冠脉血流、提高心功能提供实验依据和理论基础。

项目背景：新近研究表明肾素-血管紧张素系统（RAS）和RhoA/ Rho kinase (ROCK)通路在心肌细胞缺血再灌注损伤病理生理机制中扮演了举足轻重的作用。目前，中药提取物虎杖苷（Polydatin, PD，其成分类似于白藜芦醇）在心血管疾病治疗中的应用逐渐受到重视。在本研究中，我们主要研究虎杖苷在心肌细胞缺血再灌注损伤中的保护作用。.方法和结果：实验主要包括细胞水平培养大鼠心肌细胞模拟缺血再灌注损伤模型；和动物水平C57成年小鼠左前降支结扎冠状动脉缺血再灌注损伤模型。与对照组相比较，虎杖苷治疗组显著提高细胞在缺血再灌注损伤后的活性并降低细胞凋亡率。同时，虎杖苷可以显著降低在小鼠冠状动脉缺血再灌注损伤，提升心功能。进一步研究表明，虎杖苷显著抑制缺血再灌注损伤时升高的ROCK活性，并且可通过抑制肾素和血管紧张素转换酶来抑制激活的局部和循环RAS系统。.结论：虎杖苷可能通过抑制肾素和血管紧张素转换酶来抑制激活的局部和循环RAS系统，达到保护缺血再灌注下心肌细胞的损伤。并且，虎杖苷可能通过抑制RhoA/ROCK通路抑制RAS系统激活