T Lymphocyte Apoptosis in Hepatitis C Persistence

描述（由申请人提供）：丙型肝炎病毒（HCV）感染越来越被认为是慢性肝炎、肝硬化和肝细胞癌（HCC）的主要原因。感染时间超过20年的人中，并发症发生率最高的人的比例预计在未来10至20年内将增加。大约85%的感染者未能清除病毒。HCV感染的一个核心未解决的问题是病毒如何在大多数感染个体中建立持续感染。最近的一些研究表明，HCV清除的失败与针对病毒的1-细胞应答的弱点有关。尽管HCV特异性T细胞在大多数慢性感染个体中很容易被证实，但与其他病毒感染相比，它们在肝脏和外周血中的频率异常低。尚不清楚这是由于未能产生病毒特异性T细胞应答还是由于活化的效应1细胞的凋亡增加。越来越多的证据表明，肝脏可能通过捕获和清除活化的T细胞，特别是CD 8 + 1细胞，在T细胞稳态中发挥重要作用。该提议的中心假设是，HCV感染的肝细胞导致肝脏中HCV特异性T细胞的过早凋亡，导致T细胞应答的减弱。为了支持这一假设，我们的初步研究表明，活化的CD 4+和CD 8 + T细胞的凋亡在体外或体内暴露于表达HCV核心、E1和E2转基因的肝细胞后加速。在转基因肝细胞中，Fas配体（FasL或Apo-Ⅰ配体）的表达上调，并可被抗Fas抗体阻断。这里提出的实验是专门设计来扩展我们的观察，并在体内验证我们的假设。具体而言，我们建议：1）确定HCV转基因小鼠中T淋巴细胞凋亡增加的体内意义; 2）确定Fas：FasL相互作用是否对该系统中活化T细胞的凋亡至关重要;和3）使用新型腺相关病毒载体在鼠肝细胞中表达HCV蛋白来表征调节活化T淋巴细胞凋亡的HCV蛋白。这些数据将证实HCV持续存在的一种机制可能是病毒复制主要部位的免疫应答减弱，并可能提出新的治疗策略以增强免疫应答的有效性。

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) infection has been increasingly recognized as a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma (HCC). The proportion of individuals infected more than twenty years, who face the highest rate of complications, is expected to increase over the next ten to twenty years. Approximately 85 percent of infected individuals fail to clear the virus. A central unresolved issue in HCV infection is how the virus establishes persistent infection in most infected individuals. A number of recent studies have suggested that failure of HCV clearance is associated with weakness in 1-cell responses against the virus. Although HCV-specific T cells are readily demonstrable in most chronically infected individuals, their frequency in the liver and peripheral blood is unusually low, when compared with other viral infections. It is not clear whether this is due to failure of generating virus-specific T-cell response or due to increased apoptosis of activated effector 1 cells. Accumulating evidence has shown that the liver may play an important role in T cell homeostasis by trapping and eliminating activated T cells, especially CD8+ 1 cells. The central hypothesis of this proposal is that hepatocytes infected by HCV causes premature apoptosis of HCV-specific T cells in the liver, leading to attenuation of T-cell response. In support of this hypothesis, our preliminary studies have shown that apoptosis of activated CD4+ and CD8+ T cells was accelerated after exposure to hepatocytes expressing HCV core, El and E2 transgenes either in vitro or in vivo. This was accompanied by up-regulation of the expression of Fas ligand (FasL or Apo- I ligand), one of the death-inducing molecules, in transgenic hepatocytes, and could be blocked by antibodies against Fas in vitro and in vivo. The experiments proposed here are specifically designed to extend our observations, and to validate our hypothesis in vivo. Specifically, we propose to: 1) Determine the in vivo significance of increased T lymphocyte apoptosis in HCV transgenic mice; 2) Determine whether Fas: FasL interactions are critical to apoptosis of activated T cells in this system; and 3) Characterize the HCV proteins regulating apoptosis of activated T lymphocytes using novel adeno-associated virus vectors to express HCV proteins in murine hepatocytes. These data would confirm one mechanism of HCV persistence may be attenuation of the immune response in the principal site of viral replication, and might suggest new therapeutic strategies to augment the effectiveness of the immune response.