PATHOGENIC AUTOREACTIVE T CELLS IN MURINE AUTOIMMUNITY

小鼠自身免疫中的致病性自身反应性 T 细胞

• 批准号: 2681329
• 负责人: IAN R RIFKIN
• 金额: $ 10.69万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2681329
• 关键词: B lymphocyte; T cell receptor; antigen presenting cell; autoantibody; autoimmunity; disease /disorder model; gene expression; gene rearrangement; genetic manipulation; genetically modified animals; helper T lymphocyte; immunocytochemistry; laboratory mouse; lymphocyte proliferation; model design /development; mutant; passive immunization; receptor expression; southern blotting; systemic lupus erythematosus; transfection /expression vector

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.

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，作为由于外周功能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