Investigation on the molecular mechanisms of antigen presentation and recognition.

抗原呈递和识别的分子机制研究。

基本信息

批准号：
14370115
负责人：
NISHIMURA Yasuharu
金额：
$ 8.9万
依托单位：
Kumamoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

项目摘要

Presentation of antigenic peptides by antigen presenting cells and consequent stimulation of CD4^+ T cells is crucial in the regulation of immune response. In this research project, we studied on the recognition of antigenic peptide by T cell receptor (TCR) and signal transduction pathway in CD4^+ T cells stimulated with altered peptide ligands (APL). In addition, we established a method for ES cell-based genetic engineering of dendritic cells (DC). The following results were obtained.1)We developed an experimental system to dentify diverse T-cell epitopes from T-cell epitope-expression library, using an epitope presenting vector based on CLIP-substituted invariant chain. Using the libraries in which randomized amino acid residues were narrowed down into three successive ones, we characterized the degeneracy in the epitopes of the GAD65-reactive Th-cell clones derived from IDDM patients, and identified several microbe-derived antigens to which the Th-cell clones cross-reacted.2)We foun … More d that a human CD4^+ T cell clone showed full proliferation in response to over-expressed partially agonistic peptide/HLA-DR4 complex. However, a protein tyrosine kinase ZAP-70, which is believed to be essential for TCR-mediated T cell activation, was not tyrosine-phosphorylated and activated. Instead, we found that other kinases B-Raf and PKCm were activated in the T cells, suggesting the presence of new signaling pathways leading to full T cell proliferation that is independent of ZAP-70 activation.3)We established a method to generate mouse ES cell-derived DC (ES-DC). Genetic modification of ES-DC can be done by transfection of ES cells and subsequent differentiation to DC. OVA antigen-specific cytotoxic T lymphocytes were efficiently primed in vivo by injecting mice with ES-DCs introduced with an OVA-expression vector. Double-transfectant ES-DC expressing a chemokine along with OVA provided potent protection from OVA-expressing tumor cells. In addition, we demonstrated prevention of experimental autoimmune encephalomyelitis by treatment with genetically modified ES-DC. Less

抗原呈递细胞呈递抗原肽并随后刺激 CD4^+ T 细胞对于免疫反应的调节至关重要。在本研究项目中，我们研究了 T 细胞受体 (TCR) 和信号转导途径对抗原肽的识别。在用改变的肽配体（APL）刺激的CD4+T细胞中，我们建立了一种基于ES细胞的树突状细胞（DC）基因工程方法。1）我们开发了。我们使用基于 CLIP 取代的不变链的表位呈递载体来鉴定来自 T 细胞表位表达文库的不同 T 细胞表位的实验系统，使用其中随机氨基酸残基缩小为三个连续残基的文库。表征了来自 IDDM 患者的 GAD65 反应性 Th 细胞克隆表位的简并性，并鉴定了 Th 细胞克隆所针对的几种微生物衍生抗原2) 我们发现，人类 CD4^+ T 细胞克隆在过度表达的部分激动肽/HLA-DR4 复合物（一种蛋白酪氨酸激酶 ZAP-70）下表现出完全增殖。被认为对于 TCR 介导的 T 细胞激活至关重要，但并未被酪氨酸磷酸化和激活，相反，我们发现其他激酶 B-Raf 和 PKCm 在 T 细胞中被激活，表明其存在。导致独立于 ZAP-70 激活的完全 T 细胞增殖的新信号通路。3)我们建立了一种产生小鼠 ES 细胞衍生 DC (ES-DC) 的方法，可以通过转染完成 ES-DC 的遗传修饰。通过向小鼠注射导入了表达趋化因子的双转染子 ES-DC 的 ES-DC，可以在体内有效地诱导 ES 细胞的分化并随后分化为 DC。与OVA一起提供了针对表达OVA的肿瘤细胞的有效保护。此外，我们证明了通过转基因ES-DC治疗可以预防实验性自身免疫性脑脊髓炎。