circ_stard13竞争性调控RhoA/ROCK通路对动脉平滑肌细胞增殖与迁移的作用及机制研究

Excessive vascular smooth muscle cells (VSMCs) proliferation and migration play pivotal steps in the genesis and process of arteriosclerosis obliterans (ASO) of the lower extremities. CircRNA is recognized as an important governor of VSMCs phenotype switch. Previously, we revealed the circRNA expression profile in ASO samples. We found that circ_stard13 was significantly upregulated in ASO media, and down-regulation of circ_stard13 markedly attenuated VSMCs proliferation and migration. Further study showed that it likely associated with RhoA/ROCK downstream pathway activation which affects the phenotypic switching of VSMCs. In the present study, we intend to use methods such as oligo nucleotide transfection, dual-luciferase assay, lentivirus mediated overexpression and in situ hybridization-immunofluorescence double staining to prove that circ_stard13 regulates VSMCs behavior via RhoA/ROCK pathway, and to explore the role of circ_stard13 in ASO. The present study would shed light on the possible role and mechanism of circ_stard13/miR-125b/RhoA/MRTF-A in modulating VSMCs behavior in ASO, and provide solid evidence for proving circ_stard13/RhoA as a new potential therapeutic approach for preventing and treating ASO.

血管平滑肌细胞（VSMCs）增殖迁移失控是导致下肢动脉硬化闭塞(ASO)的重要原因，circRNA是调控VSMCs表型转换的重要分子。通过对人ASO病变动脉进行circRNA芯片筛选发现，circ_stard13在中膜显著上调，下调其表达后可明显抑制VSMCs增殖及迁移。深入分析发现circ_stard13可能通过调控下游RhoA/ROCK通路的激活，影响VSMCs的表型切换有关。本项目拟采用寡核苷酸转染、双荧光素酶报告基因、慢病毒表达、原位杂交-免疫荧光双染等方法，旨在获得circ_stard13通过RhoA/ROCK调控VSMCs表型转换的可靠证据，评价和证实其在ASO发病过程中的作用。本项目将揭示ASO发病中circ_stard13/miR-125b/RhoA/MRTF-A调控VSMCs生物学行为变化的新机制，为确立其作为干预ASO的新途径提供科学依据。

血管平滑肌细胞（VSMCs）增殖迁移失控是导致下肢动脉硬化闭塞(ASO)的重要原因，circRNA是调控VSMCs表型转换的重要分子。本研究对人ASO病变动脉进行circRNA芯片筛选入手，发现circ_stard13在中膜显著上调，下调其表达后可明显抑制VSMCs增殖及迁移。本项目采用了寡核苷酸转染、双荧光素酶报告基因、RNA pulldown、质谱、原位杂交-免疫荧光双染等方法，发现circ_stard13作为锚定剂促进ROCK2分子与m6A效应蛋白YTHDF2的结合，导致YTHDF2对ROCK2的降解，从而调控RhoA/ROCK通路调控VSMCs表型转换，促进ASO疾病进展。本研究首次探讨了非编码RNA和表观遗传学在ASO中的可能致病机制，并为ASO的治疗提供了新的靶点和理论依据。课题组在完成了项目计划的基础上，还深入挖掘circRNA作为锚定分子促进蛋白与RNA结合行使功能，从而促进了ASO的发生发展。