Development of model system to analyze the antigen epitope to induce cell-mediated immunity in fish

鱼类具有特异性抗原识别分子以及B淋巴细胞和T淋巴细胞，可引起针对多种抗原的特异性免疫反应。然而，在鱼类中，由于缺乏体内和体外模型系统，与体液免疫相比，关于细胞介导免疫的信息有限。尽管病毒性疾病是水产养殖中的严重问题，但对鱼类的病毒性疾病疫苗有限。研制这类疫苗需要鉴定和鉴定可诱发保护性免疫的病毒抗原。众所周知，细胞介导的杀伤是控制病毒性疾病最重要的防御机制。本研究项目的目的是开发实验模型系统，以分析参与细胞介导免疫的保护性抗原或表位，以开发鱼类疫苗。通过与东京水产大学的Okamoto博士和国立水产养殖研究所的Ototake博士的合作，我们已经建立了几种用于特异性细胞介导免疫的检测系统，特别是使用克隆ginbuna和同基因细胞系的细胞毒性t细胞功能。这些包括移植物抗宿主反应（GVHR）和细胞介导的细胞毒性（CMC）对异体红细胞或细胞系。本课题在此基础上进一步建立了CMC抗病毒感染细胞的模型系统，并利用等基因的银麻和细胞系与鱼类病毒一起分析了CMC的保护机制。我们还广泛分析了虹鳟鱼的MHC多态性，发现虹鳟鱼细胞系（RTG-2）与纯合克隆虹鳟鱼（C25克隆）具有相同的MHC I类等位基因。我们用IHNV- g DNA免疫致敏或IHNV和RTG-2细胞感染致敏的C25克隆鳟鱼淋巴细胞，证明了mhc限制细胞介导的对病毒感染细胞系的细胞毒性。本研究建立的实验模型系统可用于研究参与细胞免疫的保护性抗原或表位，为鱼类疫苗的研制提供依据，也可用于研究特异性细胞免疫的识别和杀伤机制。少

Fish possess specific antigen recognition molecules as well as B and T lymphocytes and can evoke specific immune responses against a variety of antigens. In fish, however, information on cell-mediated immunity is limited when compared to those on humoral immunity due to the lack of in vivo and in vitro model systems. Vaccine against viral disease is limited in fish despite that viral diseases are serious problem in aquaculture. Development of such vaccines requires identification and characterization of viral antigens which induce protective immunity. It is well known that cell-mediated killing is most important defense mechanisms in the control of viral disease. The objective of this research project is to develop experimental model systems to analyze the protective antigens or epitopes involved in cell-mediated immunity for the development of fish vaccines.In collaboration with Dr. Okamoto at Tokyo University of Fisheries and Dr. Ototake at National Research Institute of Aquaculture … More we have established several assay systems for specific cell-mediated immunity, particularly on cytotoxic T-cell function employing clonal ginbuna and a syngeneic cell line. Those include the graft-versus host reaction (GVHR) and cell-mediated cytotoxicity (CMC) against allogeneic erythrocytes or cell lines. In the present research project we further pursued the studies and have established model systems for CMC against virus-infected cells and analyzed the protection mechanism using isogeneic ginbuna and cell lines together with fish viruses. We also extensively analyzed the MHC polymorphism in rainbow trout and found that trout cell line (RTG-2) shares the same MHC class I allele with homozygous clonal rainbow trout (C25 clone). We demonstrated MHC-restricted cell-mediated cytotoxicity against virus-infected cell lines using lymphocytes from C25 clonal trout sensitized by DNA immunization with IHNV-G or by infection with IHNV and RTG-2 cells as target. Experimental model systems established in the present study can be applied to investigate the protective antigens or epitopes involved in cell-mediated immunity for the development of fish vaccines as well as to investigate the recognition and killing g mechanism of specific cell-mediated immunity. Less