PCSK9促动脉粥样硬化新机制：NF-κB/KLF4/MYOCD途径介导的血管平滑肌细胞表型转化

Evidence indicates a non-lipid role for proprotein convertase subtilisin/kexin 9 (PCSK9) in atherogenesis, but the underlying molecular mechanisms remain unclear. The phenotypic switch of vascular smooth muscle cells (VSMCs) plays a major role in the development of atherosclerosis. Our preliminary studies suggest that PCSK9 has a potential new effect of VSMC phenotypic switch. Therefore, we hypothesize that PCSK9 may accelerate atherosclerosis by stimulating VSMC phenotypic switch. To test this hypothesis, we will first confirm and characterize the expression of PCSK9 in VSMCs with contractile and synthetic phenotypes. Then, gene transfection and RNA interference will be used to determine the effects of PCSK9 on the regulation of VSMC phenotypic switch, and the possible role of the NF-κB/KLF4/MYOCD axis will be determined as well. Finally, we will generate VSMC-specific PCSK9 overexpression and knockout mice in an atherosclerosis-prone ApoE−/− background and examine the roles of PCSK9 in VSMC phenotypic switch and atherogenesis in vivo. Our proposed studies will provide new insights into whether PCSK9-regulated VSMC phenotypic switch contributes to the development of atherosclerosis and unveil the molecular mechanisms underlying the PSCK9 regulation of VSMC phenotypic switch, which will shed light on the PCSK9 biological functions. Establishing a direct role for VSMC PCSK9 in atherosclerosis will open a new avenue to treat and prevent atherosclerosis-related diseases.

PCSK9的非脂质调节作用在动脉粥样硬化（As）发生发展中扮演重要角色，但机制未明。血管平滑肌细胞（VSMCs）表型转化是As进程的关键环节。我们前期发现PCSK9具有调控VSMCs表型转化的新潜在效应。据此，提出“PCSK9调控VSMCs表型转化促As”的科学假说。项目将首先观察VSMCs不同表型状态时PCSK9的表达变化；然后，在细胞水平运用基因转染和RNA干扰技术，明确PCSK9对VSMCs表型转化的调控作用，并从NF-κB/KLF4/MYOCD途径阐明其作用的分子机制；最后，构建VSMCs特异性PCSK9过表达和敲除的小鼠As模型，整体水平进一步观察PCSK9对As中VSMCs表型转化的调控以及对As病变形成的影响。本研究将从VSMCs表型转化调控这个新视点揭示PCSK9促As的新作用机制，从而为PCSK9的生物学效应增添新认识，也为As的防治提供新思路。

本课题观察了PCSK9定位于人类和小鼠的主动脉粥样硬化（atherosclerosis，As）斑块的平滑肌细胞中并且高表达，为了更好的明确VSMCs中PCSK9过表达在As斑块形成的作用，课题组成功构建了C57BL/6J背景的VSMCs特性PCSK9过表达小鼠，从整体水平明确了PCSK9对As病变形成，平滑肌细胞表型由收缩型向合成型转化，MYOCD表达促进作用以及对KLF4表达抑制作用；在细胞层面，利用构建的PCSK9过表达腺病毒载体进行感染或siRNA干扰PCSK9，改变小鼠原代主动脉VSMCs的PCSK9内源性表达，从而有助于更好地理解PCSK9对平滑肌细胞表型转换和NF-κB/KLF4/心肌蛋白途径的影响。进一步在改变PCSK9的基础上分级干预NF-κB或KLF4的表达，明确了PCSK9可能通过NF-κB/KLF4/MYOCD作用途径促进VSMCs向合成型表型转化。本课题的研究结果为PCSK9非脂质调节作用的研究开拓新途径，也将为 PCSK9作为As性疾病的治疗靶点提供更充分的科学依据。