circRNA与microRNA或功能蛋白互作调控血管平滑肌细胞增殖的分子机制

Vascular smooth muscle cell (VSMC) phenotypic switch and obtaining proliferative capacity, is an important pathological event in vascular remodeling diseases. To discover the regulatory functions of circular RNA (circRNA) in VSMC proliferation, differentially expressed circRNA in proliferative VSMC were detected by gene microarray, further studies suggested that many circRNAs participate in VSMC proliferation regulation, but the molecular mechanisms are unclear. This project is planned to explore the molecular mechanisms of circRNA in regulating VSMC proliferation: (1) Based on circRNA playing function as competing endogenous RNA (ceRNA), to identify circRNA-miRNA-mRNA interaction pathways, and elucidate their regulatory functions in VSMC proliferation; (2) Based on circRNA playing function as circRNA-protein complex, to identify circRNA functional binding proteins, and demonstrate the regulatory functions of circRNA-protein complexes in VSMC proliferation. Finally, to certify that circRNA is a class of important regulator in vascular remodeling from the molecular, cellular and whole levels, and provide novel insights for studying the mechanisms of vascular remodeling diseases，and provide experimental evidences for discovering novel potential therapeutic targets.

血管平滑肌细胞（VSMC）表型发生转化并获得增殖能力，是血管重塑性疾病的重要病理学事件。为了探讨环状RNA（circRNA）在VSMC增殖中的调控功能，我们采用基因芯片筛查获得增殖型VSMC中差异表达的circRNA，进一步研究证明多种circRNA参与调控VSMC增殖，但具体的分子机制还不清楚。本项目拟从两个角度对circRNA调控VSMC增殖的分子机制展开论证：（1）基于circRNA作为竞争性内源RNA角度，鉴定circRNA-microRNA-mRNA互作通路，阐明其参与VSMC增殖的调控功能；（2）基于circRNA-蛋白质互作角度，鉴定与circRNA结合的功能蛋白，论证circRNA-蛋白质复合体调控VSMC增殖的分子机制。最终，从分子、细胞和整体水平证明circRNA是一类重要的血管重塑调控因子，为血管重塑性疾病的机制研究提供新思路，为新型治疗靶点的发掘提供实验依据。

血管平滑肌细胞（vascular smooth muscle cell，VSMC）异常增殖是动脉粥样硬化、高血压和动脉瘤等血管重构性疾病的病理基础。环状RNA（circular RNA，circRNA）是一类具有共价闭合环状结构的非编码RNA，参与多种生物过程的调控，但其在VSMC增殖调控中的作用尚不完全清楚。本研究通过基因芯片技术，分析了增殖型和静止型人主动脉平滑肌细胞（human aortic smooth muscle cell，HASMC）差异表达的circRNA（differentially expressed circRNA，DEcircRNA）。与静止型HASMC相比，增殖型HASMC中有66种circRNA表达水平上调，68种circRNA表达水平下调。通过构建DEcircRNA-miRNA-DEmRNA网络，利用GO和KEGG通路分析对DEcircRNA进行功能预测。通过circRNADb预测分析，筛选得到27种DEcircRNA含有内部核糖体进入位点和开放阅读框，提示它们具有编码蛋白质的潜能。通过表达相关性分析，发现DEcircRNA与其同源的线性mRNA不具有表达相关性。通过敲减和过表达分析，证实hsa_circ_0001402、has_circ_0040705、has_circ_0001304等多种DEcircRNA能够调控VSMC增殖。选择hsa_circ_0001402进行了深入探究，发现hsa_circ_0001402在增殖型HASMC中表达水平异常下调；EdU和CCK-8分析表明，过表达hsa_circ_0001402能显著抑制HASMC增殖和细胞活力；进一步联合报告基因分析、Western blot等技术，揭示hsa_circ_0001402通过miR-183-5p/FKBPL/p21轴抑制HASMC细胞周期。另外，发现hsa_circ_0001402在不稳定型心绞痛患者的血浆中显著下调，是一种有前景的不稳定型心绞痛诊断标志物。我们的研究为探究circRNA在VSMC增殖相关的血管重构疾病中的功能提供了重要的研究线索，hsa_circ_0001402是一种有潜力的血管重构疾病诊断治疗新靶点。