Targeting smooth muscle cell BMAL1 as a new therapeutic strategy against restenosis

项目摘要/摘要 在美国，冠心病(CHD)是唯一主要的死亡原因。冠状动脉 血运重建，包括冠状动脉搭桥术和经皮冠状动脉介入治疗，是最常见的 冠心病患者的常见方式。然而，它常常与再狭窄的高发生率相关。 尽管使用裸金属支架(BMS)可以显著降低再狭窄率，尤其是使用 药物洗脱支架(DES)、BMS后持续高的再狭窄率和支架内风险增加 血栓形成与DES一直是该领域的一大重大空白。当前应用程序 建议弥合这一差距，并专门研究平滑肌细胞(SMC)BMAL1在 小鼠动脉损伤模型中的新生内膜增生。BMal1是BMAL1的核心分子成分 生物钟。长期以来，人们一直认为BMAL1功能的丧失对人类的健康有害。然而， 这一教条受到了我们最近报告的挑战，即SMC中BMal1的基因缺失(SM-BMal1-KO) 保护小鼠免受醛固酮或血管紧张素II诱导的腹主动脉瘤的影响。与此一致 我们的初步发现表明，SMC特异性的BMal1缺失也能保护小鼠股动脉丝 损伤或颈动脉结扎所致的新生内膜增生。当潜在的机制仍然存在时 难以捉摸的是，我们的初步数据显示，花生四烯酸代谢受到SMC特异性的影响最大 BMal1缺失。有趣的是，在与花生四烯酸代谢有关的酶中，胞浆 磷脂酶A2α(CPLA2)受SMC特异性BMAL1缺失的影响最大。从机械上讲，我们的 初步数据表明，BMAL1与小鼠cPLA2启动子结合，SMC BMAL1是必需的 血管损伤诱导新生内膜cPLA2表达上调。鉴于cplA2在血管平滑肌细胞中的潜在作用 增殖和新生内膜增生，我们假设血管上调SMC中BMAL1的表达 损伤促进cPLA2的表达，从而介导新生内膜增生的发生和发展。 显著促进冠状动脉血运重建后再狭窄的发生。目标1将测试 假设血管损伤导致SMC BMAL1表达上调在血管病变的发生和发展中起关键作用 新生内膜增生。目标2将定义SMC BMAL1调节cPLA2的机制，从而 介导血管损伤诱导的新生内膜增生。为了实现这些目标，SM-BMal1-KO，SM-CPLA2- KO和SM-BMal1/cPLA2-KO小鼠将造成股动脉钢丝损伤或颈动脉结扎 诱导新生内膜增殖及SMC BMAL1和cPLA2调节机制的研究 血管损伤所致的新生内膜增生。拟议研究的结果将提供新的机制 深入了解SMC BMAL1如何通过cPLA2介导血管损伤诱导的新生内膜增生。此外， 拟议研究的结果将提供临床前证据，抑制BMAL1/cPLA2信号转导 在病变部位是针对冠状动脉血运重建后再狭窄的新治疗策略。

Project Summary/Abstract Coronary heart disease (CHD) is the single leading cause of death in the United States. Coronary revascularization, including coronary artery bypass graft and percutaneous coronary intervention, is the most common modality in patients with CHD. However, it is often associated with a high incidence of restenosis. Although the rate of restenosis is reduced significantly by using bare-metal stent (BMS) and particularly with drug-eluting stent (DES), a persistently high rate of restenosis after BMS and an increased risk of in-stent thrombosis with DES has been encountered as a major significant gap in the field. The current application proposes to bridge this gap and specifically investigates the role of the smooth muscle cell (SMC) BMAL1 in neointimal hyperplasia in mouse models of arterial injury. BMAL1 is a core molecular component of the circadian clock. It has long been believed that loss of function of BMAL1 is detrimental to well-being. However, this dogma was challenged by our recent report that genetic deletion of Bmal1 in SMCs (SM-Bmal1-KO) protects mice from aldosterone or angiotensin II-induced abdominal aortic aneurysm. Consistent with this finding, our preliminary demonstrate that SMC-specific Bmal1 deletion also protects mice femoral artery wire injury- or carotid artery ligation-induced neointimal hyperplasia. While the underlying mechanism remains elusive, our preliminary data reveal arachidonic acid metabolism is most significantly affected by SMC-specific Bmal1 deletion. Interestingly, among enzymes implicated in arachidonic acid metabolism, cytosolic phospholipase A2 alpha (cPLA2) is most downregulated by SMC-specific Bmal1 deletion. Mechanistically, our preliminary data illustrate that BMAL1 binds to the mouse cPLA2 promoter, and SMC BMAL1 is required for vascular injury-induced cPLA2 upregulation in neointima. Given the potential role of cPLA2 in VSMC proliferation and neointimal hyperplasia, we hypothesize that upregulation of BMAL1 in SMCs by vascular injury promotes cPLA2 expression, thus mediating the initiation and progression of neointimal hyperplasia and significantly contributing to the development of restenosis after coronary revascularization. Aim 1 will test the hypothesis that upregulation of SMC BMAL1 by vascular injury is critical for the onset and progression of neointimal hyperplasia. Aim 2 will define the mechanism by which SMC BMAL1 regulates cPLA2 and thereby mediates vascular injury-induced neointimal hyperplasia. To achieve the goals, SM-Bmal1-KO, SM-cPLA2- KO, and SM-Bmal1/cPLA2-KO mice will be subjected to femoral artery wire injury or carotid artery ligation to induce neointimal hyperplasia and to determine the mechanism by which SMC BMAL1 and cPLA2 mediate vascular injury-induced neointimal hyperplasia. Results from the proposed studies will provide new mechanistic insight on how SMC BMAL1 mediates vascular injury-induced neointimal hyperplasia via cPLA2. Moreover, results from the proposed studies will provide preclinical evidence that inhibiting the BMAL1/cPLA2 signaling at the lesion site is a new therapeutic strategy against restenosis after coronary revascularization.