Peptide antigens in CD45RBhigh CD4+T cell colitis model

CD45RBhigh CD4 T 细胞结肠炎模型中的肽抗原

• 批准号: 6468191
• 负责人: LAWRENCE J SAUBERMANN
• 金额: $ 8.05万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6468191
• 关键词: CD antigens; MHC class II antigen; SCID mouse; T cell receptor; antigen presentation; antigens; bacterial antigens; chimeric proteins; colitis; cytokine; disease /disorder model; genetic library; helper T lymphocyte; laboratory mouse; leukocyte activation /transformation; lymphocyte proliferation; recombinant proteins; tissue /cell culture

DESCRIPTION (provided by applicant): Chronic human inflammatory bowel diseases (IBD), such as Crohn?s Disease and Ulcerative Colitis, afflicts approximately two million individuals in this country alone. These disorders are primarily characterized by the presence of activated forms of T lymphocytes in the intestinal mucosal compartment. Through a number of investigations in murine models of IBD, as well as human studies, it is becoming increasingly recognized that IBD is associated with luminal or environmental stimuli in a genetically susceptible host. One of the best-characterized murine models of IBD is the CD45RBhigh CD4+ T cell transfer model. Adoptive transfer of this particular subset of T cells into a severe combined immunodeficient (SCID) congeneic recipient, who lacks functional B or T cells, results in colitis. However, adoptive transfer of the CD45RBlow CD25+ CD4+ T cells inhibits colitis. Recent evidence indicates that both T cell subsets are dependent on MHC class II and are activated through a restricted set of T cell receptors, which indicates expansion to a limited set of antigens. The model is also dependent on environmental antigen exposure, as SCID mice raised under germ-free conditions, do not develop colitis following cell transfer. Taken together, this implies that the effector and regulatory CD4plus T cells in this transfer model of colitis are most likely activated by a limited number of peptide antigens presented in a MHC class II dependent fashion, and possibly derived from the luminal environment. Therefore, the primary goal of this project is to elucidate the peptide antigen(s) involved in the CD45RBhigh CD4+ T cell population effector response or the CD45RBlow CD25+ CD4+ inhibitory response. Specifically, we plan to use a methodological approach involving cDNA expression libraries to identify unknown peptide antigens for these subsets of T cells. Preliminary experiments with murine DO11.10 CD4+ T cell hybridoma cells, which recognize a 17 amino-acid peptide epitope of chicken ovalbumin in the context of MHC class II, has indicated the initial parameters of this methodology. The primary goal is to identify expressed peptide(s) from recombinant fusion proteins created from cDNA libraries made from tissue and cecal bacteria extracts. The libraries will be screened for reactive epitopes by proliferation and cytokine production. Next, these important T cells will undergo T cell receptor evaluation. Potentially, by this approach, antigens responsible for CD45RBhigh CD4plus T cell adoptive transfer model of colitis can be elucidated, this may lead to a better understanding of human inflammatory bowel disease.

描述（由申请人提供）： 慢性人类炎症性肠病 (IBD)，例如克罗恩病和 溃疡性结肠炎，困扰着大约 200 万人 国家独自。这些疾病的主要特征是存在 肠粘膜室中 T 淋巴细胞的活化形式。通过 对 IBD 小鼠模型以及人类研究进行的多项研究， 人们越来越认识到 IBD 与管腔或 遗传易感宿主的环境刺激。中的一个 最具特征的 IBD 小鼠模型是 CD45RBhigh CD4+ T 细胞 转移模型。将这一特定 T 细胞亚群过继转移至 严重联合免疫缺陷 (SCID) 同基因受者，缺乏 功能性 B 或 T 细胞，导致结肠炎。然而，收养转移 CD45RBlow CD25+ CD4+ T 细胞抑制结肠炎。最近的证据表明 两个 T 细胞亚群都依赖于 MHC II 类并被激活 通过一组有限的 T 细胞受体，这表明扩展到 有限的抗原组。该模型还依赖于环境抗原 在无菌条件下饲养的 SCID 小鼠在接触暴露后不会发育 细胞移植后结肠炎。综合起来，这意味着效应器 在这种结肠炎转移模型中，调节性 CD4+ T 细胞最多 可能被 MHC 中存在的有限数量的肽抗原激活 II 类依赖时尚，可能源自光环境。 因此，该项目的首要目标是阐明肽 参与 CD45RBhigh CD4+ T 细胞群效应子的抗原 反应或 CD45RBlow CD25+ CD4+ 抑制反应。具体来说， 我们计划使用涉及 cDNA 表达库的方法学方法 鉴定这些 T 细胞亚群的未知肽抗原。初步的 使用小鼠 DO11.10 CD4+ T 细胞杂交瘤细胞进行实验，该细胞识别 MHC 背景下鸡卵清蛋白的 17 个氨基酸肽表位 II 类，表明了该方法的初始参数。初级 目标是从重组融合蛋白中鉴定表达的肽 由组织和盲肠细菌提取物制成的 cDNA 文库创建。这 将通过增殖和细胞因子筛选文库的反应性表位 生产。接下来，这些重要的T细胞将经历T细胞受体 评估。潜在地，通过这种方法，负责 CD45RBhigh 的抗原 结肠炎的CD4+ T细胞过继转移模型得以阐明，这可能 有助于更好地了解人类炎症性肠病。