Immune Mechanisms of HCV Persistence and Pathogenesis

描述（由申请人提供）：目前近400万美国人感染丙型肝炎病毒（HCV）。HCV感染每年造成大约8000 - 10000人死亡，是美国肝移植的主要原因。目前还没有控制丙型肝炎的疫苗，少数人感染。丙型肝炎病毒在最初感染时克服病毒并将其从血液中清除。其余75%-85%发展为慢性感染。这种疾病的严重程度因人而异。病毒持续存在、疾病进展和新疗法的潜在病毒/宿主靶点的分子基础尚不清楚。我们假设实质HCV抗原呈递导致不适当的T细胞功能有助于免疫介导的肝脏炎症，并且病毒蛋白的直接免疫抑制是HCV生存综合策略的一个组成部分。三种互补的方法将用于验证这些假设：1)为了测试免疫共刺激分子（如CD40）在HCV相关肝损伤中的功能，我们将在肝脏中建立有条件表达HCV结构蛋白和CD40分子的小鼠谱系。通过FACS、ELISPOT和免疫组织学分析检测小鼠肝脏病理变化、肝T细胞启动和效应功能。2)为了研究HCV非结构蛋白（NS3）是否会干扰宿主抗病毒反应，从而导致病毒的持续存在，我们将培养有条件表达HCV-NS基因的小鼠，该基因由一种新型的他莫昔芬诱导系统控制。我们将检查这些动物的干扰素(IFN)- α / β和适应性免疫反应，以应对肝脏中的病毒攻击。这些动物的肝损伤和病毒持久性将通过生化、组织病理学和免疫化学以及实时PCR分析来确定。3)最后，我们将在有条件表达HCV全基因的动物肝脏中研究HCV- s和ns基因对疾病发病和病毒持久性的协同作用。总补体HCV蛋白的免疫抑制作用将在病毒攻击后进行测试。这些研究将提供有关病毒在体内持续存在和疾病进展的分子机制的重要信息，并可能有助于设计未来的靶向阻断特定候选药物的策略。

DESCRIPTION (provided by applicant): Nearly 4 million Americans are currently infected with hepatitis C virus (HCV). HCV infection is responsible for approximately 8,000-10,000 deaths annually and is the leading reason for liver transplantation in the United States. There is no vaccine for the control of hepatitis C. A small number of people infected with. HCV overcome the virus during the initial infection and clear it from the bloodstream. The remaining 75%-85% develop a chronic infection. The severity of the disease varies greatly from person to person. The molecular basis for viral persistence, disease progression and potential virus/host targets for novel therapeutics is not well understood. We hypothesize that parenchymal HCV antigen presentation leading to inappropriate T cell function contributes to immune-mediated liver inflammation and that direct immunosuppression by viral proteins is a component of an integrated strategy of HCV survival. Three complementary approaches will be used to test these hypotheses: 1) To test the function of immune costimulatory molecules (e.g. CD40) in HCV-related liver injuries, we will establish a lineage of mice that conditionally express HCV structural proteins and CD40 molecules in the liver. Pathological changes in the liver, hepatic T cell priming and effector functions in these mice will be measured by FACS, ELISPOT and immunohistological analyses. 2) To examine whether HCV nonstructural (NS) proteins, especially NS3, interfere with the host antiviral responses leading to viral persistence, we will generate mice conditionally expressing HCV-NS genes, which are controlled by a novel tamoxifen-inducible system. We will examine these animals' interferon (IFN)-alpha/beta and adaptive immune responses in response to a viral challenge in the liver. Hepatic injury and viral persistence in these animals will be determined by biochemical, histopathological and immunochemical, and real-time PCR analyses. 3) Finally, we will examine the synergistic effects of HCV-S and-NS genes on disease pathogenesis and viral persistence in animals conditionally expressing the entire HCV genes in the liver. The immunosuppressive effect of the total complement HCV proteins will be tested following a viral challenge. These studies would provide important information concerning molecular mechanisms responsible for viral persistence and disease progression in vivo, and may aid in devising future strategies targeted at blocking specific candidates.