Vascular Mechanisms in Homocysteinemia and Atherosclerosis

描述（由申请方提供）：高同型半胱氨酸血症和高胆固醇血症是常见的心血管风险因素，通常在血管血栓形成事件患者中同时发生。需要更好地了解高同型半胱氨酸血症和高胆固醇血症（单独或联合）易导致血管功能障碍的病理生理学途径，以开发更有选择性的血管事件预防和治疗靶点。在上一个资助期间，该项目研究了高同型半胱氨酸血症小鼠模型中血管功能障碍的机制。我们现在建议关注高同型半胱氨酸血症和高胆固醇血症之间的相互作用，导致血管功能障碍，新生内膜形成和血栓形成。我们的初步数据表明，高同型半胱氨酸血症和高胆固醇血症诱导细胞应激反应（氧化应激和内质网（ER）应激）通过共同的机制，包括激活NADPH氧化酶和T细胞死亡相关基因51（TDAG 51）。本项目的总体目标是确定高同型半胱氨酸血症和高胆固醇血症单独或联合导致血管功能障碍和血栓形成的分子机制。我们的具体目标是确定NADPH氧化酶的机制作用，我们假设这是一个主要的氧化应激源，和TDAG 51，我们假设这是一个主要的调解ER压力。我们将使用基因改变的小鼠来探测导致和防止血管功能障碍、新生内膜形成和由高同型半胱氨酸血症和/或高胆固醇血症诱导的血栓形成的机制。在目的1中，我们将检验高同型半胱氨酸血症和高胆固醇血症通过血管NADPH氧化酶机制诱导细胞氧化应激、血管功能障碍、新生内膜形成增加和血栓形成易感性增强的假设。我们建议通过确定NADPH催化亚基Nox 1和Nox 4以及过氧化物酶体增殖物激活受体γ（PPAR 3）在血管平滑肌细胞（SMC）和小鼠中表达改变的血管表型效应来验证这一假设。在目标2中，我们将检验高同型半胱氨酸血症和高胆固醇血症诱导ER应激，导致TDAG 51介导的血管功能障碍，增加新生内膜形成，并增强血栓形成的易感性的假设。我们建议通过确定TDAG 51和葡萄糖调节蛋白78（GRP 78;一种ER伴侣蛋白，可保护免受ER应激）在内皮细胞（EC）、SMC和小鼠中的表达改变对血管表型的影响来验证这一假设。我们还建议确定的作用，PPAR 3下调介导的血管效应的ER应激和TDAG 51的血管内皮或血管肌肉中的显性负性PPAR 3的组织特异性过表达的血管表型效应。因此，我们建议使用这些模型和方法来定义两个基本的细胞应激途径的作用，其中每一个都有可能被靶向治疗。公共卫生相关性：心血管疾病及其并发症是死亡、发病和卫生保健支出的主要原因。高同型半胱氨酸血症和高胆固醇血症是常见的心血管危险因素，经常在临床血管事件患者中同时发生。该项目的目标是定义高同型半胱氨酸血症和高胆固醇血症单独或联合导致不良血管事件的分子机制，并确定新的治疗靶点。

DESCRIPTION (provided by applicant): Hyperhomocysteinemia and hypercholesterolemia are common cardiovascular risk factors that often occur together in patients with vascular thrombotic events. A better understanding of the pathophysiological pathways by which hyperhomocysteinemia and hypercholesterolemia, alone or in combination, predispose to vascular dysfunction is needed to develop more selective targets for the prevention and treatment of vascular events. During the previous funding period, this project investigated mechanisms of vascular dysfunction in murine models of hyperhomocysteinemia. We now propose to focus on interactions between hyperhomocysteinemia and hypercholesterolemia that lead to vascular dysfunction, neointima formation, and thrombosis. Our preliminary data suggest that hyperhomocysteinemia and hypercholesterolemia induce cellular stress responses (oxidative stress and endoplasmic reticulum (ER) stress) through common mechanisms, including the activation of NADPH oxidase and T cell death-associated gene 51 (TDAG51). The overall goal of this project is to define the molecular mechanisms by which hyperhomocysteinemia and hypercholesterolemia, alone or in combination, lead to vascular dysfunction and thrombosis. Our specific objectives are to determine the mechanistic roles of NADPH oxidase, which we hypothesize to be a major source of oxidative stress, and TDAG51, which we hypothesize to be a major mediator of ER stress. We will use genetically altered mice to probe mechanisms that lead to, and protect against vasomotor dysfunction, neointima formation, and thrombosis induced by hyperhomocysteinemia and/or hypercholesterolemia. In Aim 1, we will test the hypothesis that hyperhomocysteinemia and hypercholesterolemia induce cellular oxidative stress, vascular dysfunction, increased neointima formation, and enhanced susceptibility to thrombosis through a mechanism involving vascular NADPH oxidase. We propose to test this hypothesis by determining the vascular phenotypic effects of altered expression of the NADPH catalytic subunits Nox1 and Nox4, as well as peroxisome proliferator-activated receptor gamma (PPAR3), in vascular smooth muscle cells (SMC) and mice. In Aim 2, we will test the hypothesis that hyperhomocysteinemia and hypercholesterolemia induce ER stress, leading to TDAG51-mediated vasomotor dysfunction, increased neointimal formation, and enhanced susceptibility to thrombosis. We propose to test this hypothesis by determining the vascular phenotypic effects of altered expression of TDAG51 and glucose-regulated protein 78 (GRP78; an ER chaperone that protects from ER stress) in endothelial cells (EC), SMC, and mice. We also propose to ascertain the role of PPAR3 downregulation in mediating the vascular effects of ER stress and TDAG51 by determining the vascular phenotypic effects of tissue-specific overexpression of dominant-negative PPAR3 in endothelium or vascular muscle. Thus, we propose to use these models and approaches to define the roles of two fundamental cellular stress pathways, each of which has the potential to be targeted therapeutically. PUBLIC HEALTH RELEVANCE: Cardiovascular disease and its complications are major causes of death, morbidity, and health care expenditures. Hyperhomocysteinemia and hypercholesterolemia are common cardiovascular risk factors that often occur together in patients with clinical vascular events. The goals of this project are to define the molecular mechanisms by which hyperhomocysteinemia and hypercholesterolemia, alone or in combination, lead to adverse vascular events, and to identify new targets for therapy.