周期性牵张力调节血管平滑肌细胞ACE2表达的分子机制及其介导的生物学效应研究

Cyclic stretch due to the heart beats play key roles for the maintenance of vascular structure and function, the smooth muscle cells located in he medial layer of the vessel wall are the primary target cells. Stretch-induced function disorders of smooth muscle cells could result vascular lesions . ACE2 is a recently discovered RAS system member, has a wide range of cardiovascular and renal protective effects. Research confirmed that other members of RAS system such as ACE, AT1R were involved in stretch mediated biological role, while the relationship between ACE2 and stretch has not been reported. We will adopt in vitro and in vivo experiments to observe the effects of stretch on ACE2 expression in smooth muscle cells and its role in biological effects induced by stretch; comparison to knowledge of ACE2 biological effects, the regulation mechanisms of ACE2 is still poorly understood. This study will focus on elaborating the molecular mechanisms of ACE2 expression mediated by stretch at transcriptional and post-transcriptional levels respectively, through pharmacological inhibitors, gene engineering, luciferase reporter gene, EMSA, ChIP and miRNA relevant technology. It has important theoretical significance for further clarifying the molecular mechanisms of vascular lesion, with providing new ideas and the intervention targets.

由于心脏搏动而产生的周期性牵张力对于血管的结构和功能具有重要的调节作用，病理性牵张力诱导的平滑肌细胞功能变化是血管重构和病变发生、发展的主要机制之一。ACE2是近年来发现的RAS系统新成员，具有广泛的心血管保护作用，已知RAS系统成员如ACE、AT1R是牵张力调节的主要分子靶点，但牵张力对ACE2的作用及机制目前国内外尚未见报道。本课题将从体内外水平系统研究不同的周期性牵张力对ACE2表达的影响，并阐述ACE2在牵张力调节平滑肌细胞生物学效应中的作用；相对于近年来ACE2功能的研究进展，目前对ACE2的表达调控机制还知之甚少，本研究将综合利用药理学抑制剂、基因工程技术、萤光素酶报告基因、EMSA、ChIP以及miRNA相关技术，分别在转录水平和转录后水平重点阐述牵张力调节ACE2表达的分子机制，研究将深入明确血管病变的力学分子机制，为防治血管疾病提供新的思路和新的干预靶点。

明确机械牵张力诱导血管平滑肌细胞功能变化的机制，对于防止血管重构的发生和发展有重要的理论和临床意义。应用Wistar大鼠腹主动脉缩窄技术构建病理性牵张力动物模型，我们发现与生理对照组相比，病理性牵张力可以抑制ACE2的表达，而诱导ACE的高表达。体外人主动脉平滑肌细胞培养发现，病理性牵张力抑制ACE2及Ang（1-7）的表达，促进ACE及Ang II的表达。应用ACE2过表达腺病毒感染人主动脉平滑肌细胞，发现ACE2参与病理性牵张力对平滑肌细胞增殖、迁移的调节，不参与对平滑肌细胞胶原代谢的调节。我们进一步研究发现病理性牵张力通过调节平滑肌细胞内MMP-2及P4Hα1的表达进而影响胶原蛋白的含量，且ACE2并不影响平滑肌细胞内MMP-2及P4Hα1的表达。应用药理学抑制剂、siRNA干扰技术及microRNA技术，发现p38、ATF3及miR-421参与了病理性牵张力对ACE2表达的调节。牵张力通过影响ATF3核转位来进一步影响ACE2的表达。