PATHOGENIC AUTOREACTIVE T CELLS IN MURINE AUTOIMMUNITY

描述(改编自应用)系统性红斑狼疮 系统性红斑狼疮(SLE)是一种系统性自身免疫性疾病，常累及肾脏。 导致约1%的成人病例和5%的儿科病例 每年在美国都会出现肾功能衰竭。MRL小鼠品系，当 合并Fas缺陷(MRL-1PR/1PR)或Fas配体缺陷(MRL-1PR- GLD/GLD)，会发展成一种类似于人类SLE的疾病，而且在更广泛的意义上 SENSE，作为外周设备故障引起的自身免疫的模型 宽容。这个应用程序的长期目标是理解 致病抗原特异性自体反应性CD4+T细胞的功能与调控 此模型中的淋巴细胞(ART)通过：1)分离ART克隆，理想情况下 核小体决定簇的特异性，并提供它们的致病性 在采用迁移实验中；这些ART将被用作 T细胞受体DNA，T细胞受体转基因小鼠将从中获得 是被建造的。ART将来自双突变体1PR/GLD MRL 小鼠(Fas和Fas配体联合缺陷)，以便于 这些过继转移实验；除了常规抗原 表达转基因B细胞的提呈细胞(APC)、类风湿因子 免疫复合体脉冲将被用作APC的来源；2) T细胞α链和β链转基因小鼠的建立 选择的致病ART克隆的受体使用适当的分子 获得重排的V-AlphaJ-α和VDJ-β序列的技术， 将这些插入到适当的载体中，通过以下方式建立创始人 构建物的囊胚注射；3)T细胞的特性 TCR转基因小鼠的培养、功能和再循环 开发；这将通过分析胸腺选择，TCR转基因来完成 的表达、体外反应性和体内致病特性 转基因T细胞及其体内相互作用部位 T细胞和产生自身抗体的B细胞。该项目应提供 自身免疫的基本机制不仅与系统性红斑狼疮有关 也与其他免疫介导的肾脏疾病有关 自身反应性T细胞起致病作用。它还将作为一个 宝贵的培训工具，申请者将借此扩展其专业知识 在细胞免疫学方面取得了新的认识和技能 分子生物学、转基因技术和 免疫组织化学。

DESCRIPTION (adapted from the application) Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that frequently involves the kidney causing approximately 1% of all cases of adult, and 5% of pediatric, end- stage renal failure in the USA every year. The MRL mouse strain, when combined with either a defect in Fas (MRL-1pr/1pr) or in Fas ligand (MRL- gld/gld), develops a disease resembling human SLE and also, in a broader sense, serves as a model of autoimmunity due to failure of peripheral tolerance. The long-term goal of this application is to understand the function and regulation of pathogenic antigen-specific autoreative CD4+ T lymphocytes (ART) in this model by: 1) isolating ART clones, ideally specific for nucleosomal determinants, and providing their pathogenicity in adoptive transfer experiments; these ARTs will be used as a source of the T cell receptor DNA from which a T cell receptor transgenic mouse will be constructed. The ART will be derived from double mutant 1pr/gld MRL mice (with combined defects of Fas and Fas ligand) in order to facilitate these adoptive transfer experiments; in additional to conventional antigen presenting cells (APC), rheumatoid factor expressing transgenic B cells pulsed with immune complexes will be used as a source of APC; 2) developing a mouse transgenic for the alpha and beta chains of the T cell receptor of the selected pathogenic ART clone using appropriate molecular techniques to obtain rearranged V-alphaJ-alpha and VDJ-beta sequences, inserting these into appropriate vectors and establishing founders by blastocyst injection of the constructs; 3) characterizing T cell education, function and recirculation in the TCR transgenic mouse developed; this will be done by analyzing thymic selection, TCR-transgene expression, in vitro reactivity, in vivo disease-inducing properties of the transgenic T cell and in vivo sites of interaction of the transgenic T cell with autoantibody-producing B cells. This project should provide insights into basic mechanisms of autoimmunity relevant not only to SLE but also to other immunologically mediated renal disease in which autoreactive T cells play a pathogenic role. It will also serve as a valuable training vehicle whereby the applicant will extend his expertise in cellular immunology and acquire new understanding and skills in state of the art molecular biology, transgenic technology and immunohistochemistry.