CTL Effector Mechanisms in Adenoviral Hepatitis

描述（由申请人提供）：本项目的总体目标是阐明负责杀死病毒感染的肝细胞的免疫机制。作为该项目的一部分进行的研究发现，肝细胞对穿孔素和颗粒酶依赖的细胞毒性T淋巴细胞（CTL）杀伤机制具有抗性，并且肝脏中病毒感染的肝细胞的清除主要由其他CTL效应机制介导，这些机制利用Fas/FasL、TNF/TNFR1和/或其他死亡受体介导的途径。这些结果使我们假设肝细胞对穿孔素和颗粒酶介导的CTL效应途径的细胞毒性作用的抵抗在限制肝内免疫应答期间肝损伤的程度方面是重要的。为了解决这一假设，并探讨在肝脏抗病毒免疫过程中CTL效应机制的独特库的机制；我们拟探讨病毒感染的肝细胞对CTL效应的颗粒胞吐途径产生耐药性的机制。初步结果表明，抗病毒免疫应答过程中产生的细胞因子分别诱导颗粒酶B特异性抑制剂蛋白酶抑制剂9 （PI-9）和丝氨酸蛋白酶抑制剂6 （SPI-6）在人和小鼠肝细胞中的表达。在拟议的研究中，我们将确定在正常和病毒感染的小鼠肝细胞中表达的丝氨酸蛋白酶抑制剂（丝氨酸蛋白酶抑制剂），并确定细胞因子在体外和体内调节这种丝氨酸蛋白酶抑制剂表达中的作用。然后，我们将分别探讨蛇肽和/或组织蛋白酶B在介导肝细胞对颗粒酶和穿孔素的抵抗中所起的作用。最后，我们将确定在体内沉默serpin和/或组织蛋白酶B基因是否会使病毒感染的肝细胞对穿孔素和颗粒酶依赖性细胞毒性敏感，从而加速病毒感染的肝细胞的免疫清除，并加剧病毒性肝炎。

DESCRIPTION (provided by applicant): The broad objective of this project is to elucidate the immune mechanisms responsible for killing of virally infected hepatocytes. Studies conducted as part of this project have found that hepatocytes are resistant to perforin and granzyme dependent mechanisms of cytotoxic T lymphocyte (CTL) killing and that clearance of virally infected hepatocytes from the liver is mediated predominately by alternative CTL effector mechanisms that utilize Fas/FasL, TNF/TNFR1 and/or other death receptor mediated pathways. These results lead us to hypothesize that hepatocyte resistance to the cytotoxic effects of the perforin and granzyme mediated CTL effector pathway is important in limiting the degree of liver injury during intrahepatic immune responses. To address this hypothesis and to explore the mechanisms responsible for the unique repertoire of CTL effector mechanisms employed during anti-viral immunity in the liver; we propose to explore mechanisms responsible for resistance of virally infected hepatocytes to the granule exocytosis pathway of CTL effector function. Preliminary findings indicate that cytokines produced during antiviral immune responses induce expression of the granzyme B specific inhibitors proteinase inhibitor 9 (PI-9) and serine proteinase inhibitor 6 (SPI-6) in human and mouse hepatocytes respectively. In proposed studies, we will determine the repertoire of serine proteinase inhibitors (serpins) expressed in normal and virally infected murine hepatocytes and determine the role of cytokines in regulating such serpin expression in vitro and in vivo. We will then explore the role that serpins and/or cathepsin B play in mediating hepatocyte resistance to granzymes and perforin, respectively. Finally, we will determine whether silencing of serpin and/or cathepsin B genes in vivo renders virally infected hepatocytes susceptible to perforin and granzyme dependent cytotoxicity and thereby accelerates immune clearance of virally infected hepatocytes and exacerbates viral hepatitis.