Regulation of smooth muscle cell function by CD44 in cardiovascular disease

描述(由申请人提供)：血管平滑肌细胞(VSMCs)的增殖是新生内膜形成的基础，这是心血管疾病(CVD)的标志。细胞外基质(ECM)在VSMC的调控中起着重要作用，我们研究了ECM成分透明质酸(HA)及其受体CD44对VSMC增殖和新生内膜形成的影响。我们发现HA和CD44对VSMC的增殖和激活具有双功能作用。在体外，天然的高分子量HA(HMW-HA)拮抗丝裂原诱导的VSMC进入S时相，而聚集在炎症部位的低分子HA(LMW-HA)则与丝裂原协同刺激VSMC进入S时相。这两种作用都是由CD44介导的，并且是由于RAC和ERK依赖的信号对细胞周期蛋白D1的不同调控所致。HMW-HA与CD44结合可抑制RAC的GTP负载以及RAC依赖的信号转导至细胞周期蛋白D1基因，而LMW-HA与CD44结合可刺激ERK依赖的细胞周期蛋白D1基因的表达。在体内也可以检测到类似的双功能，因为CD44的基因缺失增强了C57BL/6小鼠的损伤反应，但抑制了apoE缺陷小鼠的动脉粥样硬化形成。由于LMW-HA/HMW-HA的比例与炎症程度直接相关，并且apoE缺陷小鼠的炎症环境比B6小鼠要大得多，我们假设CD44在体内的生物学反应是由局部炎症程度决定的，炎症环境的变化可以调和CD44在血管损伤和动脉粥样硬化反应中对VSMC激活和新生内膜形成的似乎相互矛盾的影响。在目标1中，我们将比较CD44在调节炎症的同时对血管损伤中VSMC增殖和新生内膜形成的影响，并将利用我们最近开发的CD44小鼠来区分CD44对SMC和炎症细胞的影响。在目标2中，我们将进行类似的实验，监测炎症和细胞类型特异性CD44表达在动脉粥样硬化形成过程中对新生内膜形成的影响。在目标3中，我们将尝试通过对携带SMC特异性缺失的rac1的小鼠进行细丝损伤来消除CD44对VSMC激活的双功能反应。公共卫生相关性：动脉粥样硬化引起的心血管疾病是美国的头号死亡原因，其特征是炎症和血管平滑肌细胞增殖导致的新生内膜形成。炎症和富含平滑肌细胞的纤维帽的形成之间的平衡在一定程度上决定了动脉粥样硬化斑块破裂和心脏病发作等急性事件的风险。微创经皮血管重建术，如血管成形术和支架植入术，目前被认为是许多患者预防和治疗急性冠状动脉事件的首选治疗方法。然而，这样的干预措施与血管再狭窄的显著风险有关，这是由于干预造成的损伤导致血管平滑肌细胞过度增殖。调节平滑肌细胞增殖可能是将动脉粥样硬化患者发生急性事件的风险降至最低的重要方法，还可以防止物理干预后的再狭窄，从而大大降低与标准治疗干预相关的风险。这项拟议的研究将确定调节平滑肌细胞生长的机制，从而为动脉粥样硬化和再狭窄的治疗提供新的靶点。

DESCRIPTION (provided by applicant): Proliferation of vascular smooth muscle cells (VSMCs) is fundamental to neointima formation, a hallmark of cardiovascular disease (CVD). The extracellular matrix (ECM) plays an important role in regulating VSMCs, and we have studied the effects of the ECM component hyaluronan (HA) and the HA receptor, CD44, on VSMC proliferation and neointima formation. We show that HA and CD44 have bifunctional effects on VSMC proliferation and activation. In vitro, the native high molecular weight form of HA (HMW-HA) antagonizes mitogen-induced S phase entry of VSMCs while the lower molecular weight forms of HA (LMW- HA) that accumulate at sites of inflammation synergize with mitogens to stimulate VSMC S phase entry. Both effects are mediated by CD44, and result from differential regulation of Rac- and ERK-dependent signaling to cyclin D1. HMW-HA binding to CD44 inhibits GTP-loading of Rac and Rac-dependent signaling to the cyclin D1 gene while LMW-HA binding to CD44 stimulates ERK-dependent cyclin D1 gene expression. A similar bifunctionality can be detected in vivo as genetic deletion of CD44 enhances the response to injury in C57BL/6 mice but inhibits atherogenesis in apoE-deficient mice. Since the ratio of LMW-HA to HMW-HA is directly related to the extent of inflammation, and the inflammatory environment is much greater in apoE-deficient mice as compared to B6 mice, we hypothesize that the biological response to CD44 in vivo is directed by the degree of local inflammation, and that changes in the inflammatory milieu can reconcile the seemingly contradictory effects of CD44 on VSMC activation and neointima formation during the response to vascular injury and atherogenesis. In Aim 1, we will compare the effects of CD44 on VSMC proliferation and neointima formation in vascular injury as we modulate inflammation, and we will exploit our recently developed floxed CD44 mouse to distinguish the effects of CD44 on SMCs and inflammatory cells. In Aim 2, we will perform similar experiments monitoring the effect of inflammation and cell type-specific CD44 expression on neointima formation during atherogenesis. In Aim 3, we will attempt to eliminate the bifunctional response of CD44 on VSMC activation by performing fine-wire injury on mice carrying an SMC-specific deletion of Rac1. PUBLIC HEALTH RELEVANCE: Cardiovascular disease due to atherosclerosis, the number one cause of death in the United States, is characterized by neointima formation due to inflammation and vascular smooth muscle cell proliferation. The balance between inflammation and formation of smooth muscle cell-rich fibrotic caps in part determines the risk to rupture of atherosclerotic plaques and acute events such as heart attacks. Minimally invasive percutaneous revascularization procedures such as angioplasty and stent implantation are currently considered the treatment of choice for the prevention and treatment of acute coronary events in many patients. However, such interventions are associated with a significant risk of vascular restenosis due to excessive vascular smooth cell proliferation in response to injury caused by the intervention. Modulating smooth muscle cell proliferation may be an important approach to minimize the risk of acute events in patients with atherosclerosis, and may also prevent restenosis following physical interventions, thereby greatly reducing the risks associated with standard therapeutic interventions. The proposed studies will define mechanisms that regulate smooth muscle cell growth and thus provide novel targets for treatment of atherosclerosis and restenosis.