Investigation on the molecular mechanisms of antigen presentation and recognition.

抗原呈递和识别的分子机制研究。

• 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
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14370115/
• 关键词: T cell; MHC; autoimmune diseases; epitope; intracellular signal transduction; embryonic stem cell; dendritic cell; tumor immunology; ES細胞; TCRアンタゴニスト; T細胞シグナル伝達; ZAP-70; PKC; ヘルパーT細胞; 抗原ペプチド; 抗原認識; 交叉反応; 分子擬態; 自己免疫; インバリアント鎖; 発現クローニング

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以及随后刺激 CD4^+ T 细胞对于免疫反应的调节至关重要。在这个研究项目中，我们研究了T细胞受体（TCR）对抗原肽的识别以及用改变的肽配体（APL）刺激的CD4^+ T细胞中的信号转导途径。此外，我们还建立了一种基于ES细胞的树突状细胞（DC）基因工程方法。获得以下结果：1)我们开发了一个实验系统，使用基于CLIP取代的不变链的表位呈递载体从T细胞表位表达库中鉴定不同的T细胞表位。使用随机氨基酸残基缩小为三个连续残基的文库，我们表征了来自 IDDM 患者的 GAD65 反应性 Th 细胞克隆表位的简并性，并鉴定了 Th 细胞克隆交叉反应的几种微生物衍生抗原。2) 我们发现……更多 d，人类 CD4^+ T 细胞克隆在响应 过度表达部分激动肽/HLA-DR4 复合物。然而，蛋白酪氨酸激酶 ZAP-70（被认为对于 TCR 介导的 T 细胞激活至关重要）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，这表明存在新的信号通路，导致 T 细胞完全增殖，且与 ZAP-70 激活无关。3）我们建立了一种产生小鼠 ES 细胞来源的 DC (ES-DC) 的方法。 ES-DC的遗传修饰可以通过ES细胞转染并随后分化为DC来完成。通过向小鼠注射引入 OVA 表达载体的 ES-DC，在体内有效启动 OVA 抗原特异性细胞毒性 T 淋巴细胞。表达趋化因子和 OVA 的双转染子 ES-DC 提供了针对表达 OVA 的肿瘤细胞的有效保护。此外，我们还证明了通过转基因 ES-DC 治疗可以预防实验性自身免疫性脑脊髓炎。较少的

项目成果

Irie, A. et al.: "Unique T cell proliferation associated with PKCm activation and impaired Zap-70 phosphorylation in recognition of overexpressed HLA/partially agonistic peptide complexes"Eur.J.Immunol.. 33. 1497-1507 (2003)

Irie, A. 等人：“识别过表达的 HLA/部分激动肽复合物时与 PKCm 激活和 Zap-70 磷酸化受损相关的独特 T 细胞增殖”Eur.J.Immunol.. 33. 1497-1507 (2003)

Senju, S., Hirata, S., Matsuyoshi, H., Masuda, M., Uemura, Y., Araki, K., Yamamura, K-I., Nishimura, Y.: "Generation and genetic modification of dendritic cells derived from mouse embryonic stem cells."Blood. 101. 3501-3508 (2003)

Senju, S.、Hirata, S.、Matsuyoshi, H.、Masuda, M.、Uemura, Y.、Araki, K.、Yamamura, K-I.、Nishimura, Y.：“衍生自树突状细胞的生成和遗传修饰

Nakatsura, T., Yoshitake, Y., Senju, S., Monji, M., Komori, H., Motomura, Y., Hosaka, S., Beppu, T., Ishiko, T., Kamohara, H., Ashihara, H., Katagiri, T., Furukawa, Y., Fujiyama, S., Ogawa, M., Nakamura, Y., Nishimura, Y.: "Glypican-3, overexpressed speci

Nakatsura, T.、Yoshitake, Y.、Senju, S.、Monji, M.、Komori, H.、Motomura, Y.、Hosaka, S.、Beppu, T.、Ishiko, T.、Kamohara, H.、

Soejima, H.: "The preference to a Th1-type response in patients with coronary spastic angina"Circulation. (in press).

Soejima, H.：“冠状动脉痉挛性心绞痛患者对 Th1 型反应的偏好”循环。

Nakatsura T et al.: "Cellular and Humoral Immune Responses to A Human Pancreatic Cancer Antigen, CLP, Originally Defined by the SEREX Method"Eur.J.Immunol. 32. 826-836 (2002)

Nakatsura T 等人：“对人类胰腺癌抗原 (CLP) 的细胞和体液免疫反应，最初由 SEREX 方法定义”Eur.J.Immunol。