The effects of hepatitis C virus (HCV) E2 protein on host immunomodulation

描述(由申请人提供)： 丙型肝炎病毒是慢性肝病、肝硬变和肝细胞癌的重要的世界性病因。在退伍军人中，丙型肝炎病毒的比例过高，退伍军人管理局的医疗系统为美国最大的丙型肝炎病毒感染者提供护理。宿主对丙型肝炎病毒的免疫识别在感染后发展缓慢，而CD4+和CD8+T细胞反应在感染后数周至数月内发展。在病毒持续存在后，在大多数感染者中很容易检测到丙型肝炎病毒特异性的CD4+和CD8+T细胞，但这些T细胞在控制病毒复制方面无效。尽管已有多种病毒蛋白被证明可干扰宿主天然免疫和获得性免疫反应的各个方面，但目前尚缺乏一个明确的机制来阐明丙型肝炎病毒通过何种途径降低CD4+和CD8+T细胞的免疫反应。除了肝脏疾病，丙型肝炎病毒还导致了各种非肝脏并发症。非肝性丙型肝炎相关疾病的发病机制可能反映了宿主免疫反应的异常调节(如冷球蛋白血症)、慢性炎症和/或免疫监视功能减弱(如非霍奇金淋巴瘤[NHL])。幸运的是，在过去的几年里，丙型肝炎病毒感染的抗病毒治疗有了显著的进步，联合使用直接抗病毒药物在病毒学治疗方面具有巨大的潜力。然而，由于丙型肝炎病毒治疗可能有严重的副作用，费用昂贵，而且不是完全有效的，因此更好地了解宿主对这种病原体的免疫反应对于确定替代治疗靶点和促进疫苗开发非常重要。我们提供的初步数据表明，丙型肝炎病毒包膜糖蛋白通过T细胞受体竞争性抑制T细胞信号转导。基于对最接近的人类黄病毒(GB病毒C；GBV-C)的研究，我们假设E2蛋白通过作为磷酸化的替代底物与关键的近端淋巴细胞特异性蛋白酪氨酸激酶(LCK)竞争。这种相互作用的结果是，T细胞受体参与后信号转导减少，导致持续的丙型肝炎病毒感染，宿主免疫监视机制减少，从而增加改变的宿主免疫反应的发展。我们进一步假设，丙型肝炎病毒E2蛋白和丙型肝炎病毒病毒粒子调节包括淋巴细胞在内的未感染旁观者细胞的免疫反应。已发表的工作与丙型肝炎病毒感染在体内改变TCR介导的信号转导一致，并且丙型肝炎病毒蛋白在体外刺激人和小鼠的TCR后抑制了增殖。我们的初步工作表明，这可能至少部分是由丙型肝炎病毒E2蛋白介导的，并且对旁观者细胞的影响涉及到外切体中的E2向周围细胞的转移。在这项建议中，我们将表征丙型肝炎病毒包膜糖蛋白E2对TCR信号的影响，并确定其对旁观者T细胞的影响机制。此外，我们还将描述含有外切体的丙型肝炎病毒包膜糖蛋白E2的成分，以及存在于丙型肝炎病毒感染者血浆中的外切体。这些研究将描述一种新的丙型肝炎病毒介导的免疫调节机制，这种机制可能有助于持续性，并可能影响非肝性丙型肝炎病毒相关的疾病过程。这种丙型肝炎病毒E2-淋巴细胞相互作用的鉴定和表征可能有助于开发新的治疗和/或疫苗方法来治疗丙型肝炎病毒，并潜在地治疗黄病毒科内的相关病毒。

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) is an important and worldwide cause of chronic liver disease, cirrhosis, and hepatocellular carcinoma. HCV is over-represented among veterans, and the VA medical system provides care for the largest population of HCV-infected people in the U.S. Host immune recognition of HCV develops slowly following infection, and CD4+ and CD8+ T cell responses develop weeks to months after infection if they develop at all. After viral persistence is established, HCV-specific CD4+ and CD8+ T cells are readily detected in most infected individuals, yet these T cells are ineffective in controlling viral replication. Although several vral proteins have been shown to interfere with various aspects of the host innate and adaptive immune response, a clear mechanism by which HCV diminishes CD4+ and CD8+ T cell immune responses is lacking. In addition to liver disease, HCV also is responsible for a variety of non- hepatic complications. The pathogenesis of non-hepatic HCV-related diseases may reflect abnormal regulation of host immune responses (e.g. cryoglobulinemia), chronic inflammation and/or decreased immune surveillance (e.g. non-Hodgkins lymphoma [NHL]). Fortunately, antiviral therapy for HCV infection has dramatically improved in the past few years, and the use of combination direct antiviral agents has a great potential for virological cure. However, because HCV treatment may have serious side effects, is expensive, and is not completely effective, it is important to better understand host immune responses to this pathogen for identifying alternative therapeutic targets and to facilitate vaccine development. We provide preliminary data demonstrating that the HCV E2 envelope glycoprotein competitively inhibits T cell signaling through the T cell receptor. Based on work in the most closely related human flavivirus (GB virus C; GBV-C), we hypothesize that E2 protein competes with the key proximal lymphocyte-specific protein tyrosine kinase (Lck) by serving as an alternative substrate for phosphorylation. As a consequence of this interaction, signaling is reduced following T cell receptor engagement, contributing to persistent HCV infection, a decrease in host immune surveillance mechanisms, and resultant increase in the development of altered host immune responses. We further hypothesize that HCV E2 protein and HCV virions modulate immune responses in uninfected bystander cells including lymphocytes. Published work is consistent with HCV infection altering TCR-mediated signaling in vivo, and that HCV proteins blunt proliferation following TCR stimulation in vitro in humans in and mice. Our preliminary work suggests that this may be mediated at least in part by HCV E2 protein, and that the effect on bystander cells involves the transfer of E2 in exosomes to surrounding cells. In this proposal we will characterize the HCV envelope glycoprotein E2 effects on TCR signaling, and confirm the mechanism of the effects on bystander T cells. In addition, we will characterize the constituents of HCV envelope glycoprotein E2 containing exosomes, and in exosomes present in the plasma of HCV-infected subjects. These studies will characterize a novel mechanism of HCV-mediated immunomodulation that is likely to contribute to persistence and may influence non-hepatic HCV- related disease processes. Identification and characterization of this HCV E2 - lymphocyte interaction may contribute to the development of novel therapeutic and/or vaccine approaches to HCV, and potentially to related viruses within the Flaviviridae.