DEFINING HEPATITIS C VIRUS-SPECIFIC CTL ACTIVITY IN MAN

描述：该建议有两个主要的具体目标：1) 证明存在丙型肝炎病毒特异性肝脏浸润性细胞毒T细胞 慢性丙型肝炎病毒感染者的淋巴细胞。申请人已经 确定了许多患者的丙型肝炎病毒基因，并将完成这项工作。 为了检测丙型肝炎病毒特异性CTL的存在，他将使用一个靶细胞 由每个患者的EBV永生化B淋巴母细胞组成 患有慢性丙型肝炎病毒。这些细胞将感染牛痘病毒。 携带丙型肝炎病毒基因的特定候选区域的编码插入 怀疑在CTL识别过程中充当表位。CTL 将通过大量扩增从肝脏收集的淋巴细胞来获得 活组织检查。靶细胞裂解的铬释放试验将用于 检测细胞并定量测定细胞毒活性。2)第二个 具体目的是定义CTL所涉及的实际表位- 中介识别。将克隆在特定AIM 1中鉴定的CTL 用于筛选合成的小分子多肽重叠 在特定目标中确定的区域1.靶细胞的致敏作用 并将使用细胞溶解试验。

DESCRIPTION: This proposal has two major specific aims: 1) To demonstrate the existence HCV-specific liver-infiltrating cytotoxic T lymphocytes in chronic HCV infection. The applicant has already determined the HCV genotype of many patients and will complete this. To detect the presence of HCV-specific CTLs, he will employ a target cell consisting EBV immortalized B-lymphoblastoid cells from each patient with chronic HCV. These cells will be infected with vaccinia virus carrying HCV gene inserts coding for specific candidate regions that are suspected of serving as epitopes in the CTL recognition process. CTLs will be obtained by bulk expansion of lymphocytes harvested from liver biopsies. chromium release assay of target cell lysis will be used to detect cells and quantitate the cytotoxic activity. 2) The second specific aim is to define the actual epitopes involved in the CTL- mediated recognition. CTLs identified in specific aim 1 will be cloned for use in screening small synthetically produced peptides overlapping in regions identified in specific aim 1. Sensitization of target cells and the cytolysis assay will be used.