MHC class II-Associated Endothelial Dysfunction

描述(申请人提供)：动脉粥样硬化性心血管疾病是长期观察到的类风湿关节炎(RA)患者早期死亡的主要原因。在这个项目中，将探索RA共享表位(SE)直接促进动脉粥样硬化发展的可能性。这项拟议的研究是基于我们最近的发现，即SE作为一种配体，能够激活内皮细胞(EC)内的促氧化信号和促动脉粥样硬化因子的分泌。在以往的成纤维细胞研究中，SE结合分子已被确定为细胞表面钙网蛋白(CRT)，它是一种已建立的天然免疫受体，在其他系统中也参与了EC的功能、免疫调节和自身免疫。值得注意的是，已知的几种动脉粥样硬化调节蛋白，如C1q、甘露糖结合凝集素和脂联素，都被证明通过细胞表面CRT传递信号。因此，我们认为SE可能通过在EC中异常激活CRT介导的信号而具有直接的促动脉粥样硬化作用。作为检验这一假说的第一步，这里提出了一个探索性的研究项目，有两个具体的目的：1.确定EC中SE激活的信号通路中的关键信号元件：这个目的的总体目标是更好地表征EC中SE触发的促氧化促动脉粥样硬化信号通路。基于已发表的相关途径的研究和我们自己的初步数据，我们将进行实验，以回答以下问题：1.1)CRT是EC中的SE结合受体吗？1.2)CD91是否在EC中扮演辅助受体的角色？1.3)2整合素在EC的SE信号中起什么作用？1.4)SE-CRT途径是否导致内皮细胞一氧化氮合酶(ENOS)解偶联？2.为了研究SE致动脉粥样硬化的作用：我们将研究SE-EC相互作用的结果，重点放在表征内皮功能障碍和AS的生物标志物上。为此，我们建议研究EC的SE激活的后果：2.1)体外，在细胞培养水平；2.2)这些分子在体内对SE阳性的HLA-DR转基因小鼠的影响；2.3)体外对从这些转基因小鼠获得的组织样本的影响。这里提出的研究检验了一个与重大健康问题有关的高度新颖的假说。拟议研究的成功完成可能为免疫系统在健康和疾病中的作用提供新的线索，并确定新的治疗方法。公共卫生相关性：动脉粥样硬化是一种常见的疾病，影响许多类风湿关节炎患者，在许多情况下会导致早期死亡。该项目将研究类风湿性关节炎患者发生动脉粥样硬化风险较高的新机制。这项研究将集中在由已知在类风湿性关节炎患者中发现频率较高的基因编码的特定蛋白质片段的影响上。该片段激活血管细胞的能力以及这种激活的生物学效应将被研究。这项研究的结果可能会为这种缩短类风湿性关节炎患者预期寿命的疾病提供亟需的线索，并可能为未来的治疗确定新的靶点。

DESCRIPTION (provided by applicant): Atherosclerotic cardiovascular disease is the main cause of the long observed early mortality in rheumatoid arthritis (RA) patients. In this project, the possibility that the RA shared epitope (SE) is directly contributing to atherosclerosis development will be explored. The proposed research is based on our recent findings that the SE acts as a ligand capable of activate pro-oxidative signaling in - and secretion of pro-atherogenic factors by - endothelial cells (EC). In prior studies in fibroblasts, the SE-binding molecule has been identified as cell surface calreticulin (CRT), an established innate immunity receptor, whose participation in EC function, immune regulation and autoimmunity has been documented in other systems. It is noteworthy that several known atherosclerosis-modulating proteins, such as C1q, mannose binding lectin and adiponectin, have all been previously shown to transduce signaling via cell surface CRT. We therefore propose that the SE may have a direct pro-atherogenic effect due to aberrant activation of CRT-mediated signaling in EC. As a first step toward examining this hypothesis, an exploratory research project is proposed here with 2 specific aims: 1. To identify key signaling elements in the SE-activated pathway in EC: The general goal of this aim is to better characterize the SE-triggered pro-oxidative pro-atherogenic signaling pathway in EC. Based on published studies in related pathways and our own preliminary data, experiments will be carried out in order to answer the following questions: 1.1) Is CRT the SE-binding receptor in EC?; 1.2) Is CD91 acting as a co-receptor in EC?; 1.3) What is the role of 2 integrins in SE signaling in EC?; 1.4) Does the SE-CRT pathway lead to endothelial 1 nitric oxide (NO) synthase (eNOS) uncoupling? 2. To characterize the atherogenic effects of the SE: We will study the outcomes of SE-EC interaction, focusing on biomarkers, which characterize endothelial dysfunction and AS. To that end, we propose to study the consequences of SE activation of EC: 2.1) in vitro, at the cell culture level; 2.2) the effect of these molecules in vivo in SE-positive HLA-DR transgenic mice and; 2.3) ex-vivo on tissue samples obtained from these transgenic mice. The studies propose here examine a highly novel hypothesis pertaining to a significant health issue. Successful completion of the proposed studies could shed new light on the role of the immune system in health and diseases and identify novel therapeutic approaches. PUBLIC HEALTH RELEVANCE: Atherosclerosis is a common condition which affects many patients with rheumatoid arthritis and in many cases causes early death. This project will investigate a novel mechanism why patients with rheumatoid arthritis have a higher risk to develop atherosclerosis. The research will focus on the effect of a particular segment of a protein encoded by a gene that is known to be found in higher frequency in patients with rheumatoid arthritis. The ability of that segment to activate blood vessel cells and the biologic effect of that activation will be studied. The results of this study could shed much needed light on a disease that shortens life expectancy of rheumatoid arthritis patients and could identify novel targets for future therapy.