REGULATION OF FAS-MEDIATED APOPTOSIS IN B CELLS

FAS 介导的 B 细胞凋亡的调节

• 批准号: 2457884
• 负责人: THOMAS L ROTHSTEIN
• 金额: $ 25.26万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-08-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2457884
• 关键词: B lymphocyte; CD antigens; T lymphocyte; anergy; antiantibody; apoptosis; autoantibody; autoimmunity; biological signal transduction; cell mediated lymphocytolysis test; ceramides; endopeptidases; enzyme activity; gene expression; genetically modified animals; interleukin 4; laboratory mouse; lymphokines; phosphorylation; protein biosynthesis; protein kinase C; tyrosine; western blottings

DESCRIPTION (Adapted from the Applicant's Abstract): B-cell anergy is one mechanism utilized to expunge autoreactive immunoglobulin specificities. Anergic B-cells survive in a state of diminished antigen receptor responsiveness, and are deleted by Fas-mediated apoptosis induced by T effector cells. In normal B-cells sensitivity to Fas-directed cell death can be regulated by IL-4R and sIg signals that produce a state of Fas-resistance. The hypothesis guiding this study is that anergic B-cells may, as a result of specific immune signals, become resistant to Fas-mediated apoptosis, thereby avoiding elimination and presenting the potential for subsequent activation and autoantibody production. Elements of this hypothesis have ramifications for B-cell immunocompetence and the regulation of apoptosis in general. For this reason, and because so little is known about the means by which Fas-resistance is triggered and established, it is proposed to explore protection against Fas-directed cell death in 3 stages. 1) Evaluate the capacity of IL-4 to oppose Fas-mediated apoptosis by anti-Ig, and determine the role of other T-cell-derived lymphokines, using standard cytotoxicity assays. 2) Determine the metabolic level at which resistance to Fas-mediated apoptosis is produced by IL-4 and by anti-Ig, by examining Fas-triggered events of protein tyrosine phosphorylation, ceramide production, and protease activation, using Western blotting, identification of radiolabeled lipids, and enzymatic assays of protein digestion, and evaluate the role of Bcl-x, through tests of gene and protein expression, and evaluation of overexpressing transgenic animals. 3) Assess induction of Fas-resistance in anergic B-cells derived from double transgenic mice, and monitor autoantibodies rescued through somatic cell hybridization of B-cells from normal animals in which resistance is generated in vitro. This work should lead to a fuller understanding of the signals and mechanisms that produce B-cell resistance to Fas-mediated apoptosis, and validate or refute the hypothesis described above, which has important implications for the pathogenesis of serological autoimmunity.

描述（改编自申请人的摘要）：B 细胞无反应性是一种 用于消除自身反应性免疫球蛋白特异性的机制。 无能 B 细胞在抗原受体减少的状态下存活 响应性，并被 T 诱导的 Fas 介导的细胞凋亡删除 效应细胞。 正常 B 细胞对 Fas 引导的细胞死亡敏感 可以通过 IL-4R 和 sIg 信号调节，从而产生以下状态： Fas 抗性。 指导这项研究的假设是无能 B 细胞 由于特定的免疫信号，可能会产生耐药性 Fas 介导的细胞凋亡，从而避免消除并呈现 随后激活和自身抗体产生的潜力。 元素 该假设对 B 细胞免疫能力和 细胞凋亡的一般调节。 正因为如此，也因为如此之少 了解触发 Fas 抗性的方式，并且 建立后，建议探索针对 Fas 定向细胞的保护 死亡分3个阶段。 1）评估IL-4对抗Fas介导的能力 抗 Ig 导致细胞凋亡，并确定其他 T 细胞来源的作用 淋巴因子，使用标准细胞毒性测定。 2) 确定代谢 IL-4 和 IL-4 对 Fas 介导的细胞凋亡产生抗性的水平 通过抗 Ig，通过检查 Fas 触发的蛋白质酪氨酸事件 使用 Western 进行磷酸化、神经酰胺生成和蛋白酶激活 印迹、放射性标记脂质的鉴定以及酶测定 蛋白质消化，并通过基因测试和评估Bcl-x的作用 蛋白质表达，以及过度表达转基因动物的评估。 3） 评估源自双核细胞的无能 B 细胞对 Fas 抗性的诱导 转基因小鼠，并监测通过体细胞拯救的自身抗体 正常动物的 B 细胞杂交，其中抗性 体外产生。 这项工作应该使人们对 产生 B 细胞对 Fas 介导的抗性的信号和机制 细胞凋亡，并验证或反驳上述假设，该假设 对血清学自身免疫的发病机制具有重要意义。