PATHOGENIC AUTOREACTIVE T CELLS IN MURINE AUTOIMMUNITY

描述（改编自应用程序）系统性红斑狼疮 (SLE)是一种系统性自身免疫性疾病， 导致大约1%的成人病例和5%的儿童病例， 肾衰竭的早期症状MRL小鼠品系，当 与Fas（MRL-1 pr/1 pr）或Fas配体（MRL-1 pr/1 pr）缺陷结合， gld/gld），发展出类似于人类SLE的疾病，并且在更广泛的意义上， 感觉，作为由于外周功能故障引起的自身免疫模型 宽容这个应用程序的长期目标是了解 病原性抗原特异性自身反应性CD 4 + T细胞的功能与调控 在该模型中，通过以下方式分离淋巴细胞（ART）：1）分离ART克隆，理想地 特异性的核小体决定簇，并提供其致病性 在过继转移实验中，这些ART将用作 T细胞受体转基因小鼠将从其获得T细胞受体DNA 被建造。ART将源自双突变体1 pr/gld MRL 小鼠（具有Fas和Fas配体的组合缺陷），以促进 这些过继转移实验;除了常规的抗原 呈递细胞（APC），表达类风湿因子的转基因B细胞 用免疫复合物脉冲将用作APC的来源; 2） 开发T细胞α和β链转基因小鼠 使用适当的分子标记， 获得重排的V-α J-α和VDJ-β序列的技术， 将其插入到适当的载体中， 胚泡注射构建体; 3）表征T细胞 TCR转基因小鼠的教育、功能和再循环 开发;这将通过分析胸腺选择，TCR-转基因 表达、体外反应性、体内疾病诱导特性 转基因T细胞和转基因T细胞的体内相互作用位点 T细胞与产生自身抗体的B细胞。该项目将提供 了解自身免疫的基本机制，不仅与SLE相关， 还涉及其它免疫介导的肾病， 自身反应性T细胞起致病作用。它也将作为一个 宝贵的培训工具，申请人将扩大他的专业知识 在细胞免疫学和获得新的理解和技能， 分子生物学、转基因技术和 免疫组化

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.