B7/CD28/CTLA-4 COSTIMULATION IN PATHOGENESIS OF R-EAE

描述(改编自申请者摘要)：激活幼稚 T细胞需要TCR占位加上CD28介导的共刺激信号 通过将B7-1和B7-2连接到抗原提呈细胞(APC)来传递。在……里面 相比之下，B7连接活化T细胞上表达的CTLA-4可提供 强烈的负调控信号可能参与耐受诱导。 在之前的申请期内的这些研究清楚地表明， 针对这些互动的策略有潜在的积极和消极 对急性和慢性肝炎的发生和发展的调节作用 复发-缓解型实验性自身免疫性脑脊髓炎 CD4+T细胞介导的自身免疫性疾病可作为多发性硬化的模型 硬化症。在此续订申请中测试的假设是 B7-CD28共刺激相互作用在积极中发挥关键作用 调节自身反应性T细胞的激活和效应功能，而 B7-CTLA-4共刺激相互作用负性调节自身免疫性疾病 是诱导/维持免疫耐受所必需的。在基础上建设 他们的初步数据描述了一种新的NOD小鼠EAE模型 用蛋白质脂蛋白多肽PLP56-70，AIM 1将使用 多方面的方法，利用靶向缺失B7-1，B7-2， B7-1/B7-2，或CD28，与抗体阻断研究相比较，以进一步 描述单个共刺激受体和配体在 EAE的诱导期、效应期和髓鞘激活期 多肽特异性T细胞反应。这一目标也将检查个人 B7-1和B7-2在诱导细胞表面活化和归巢中的作用 抗原、增殖、细胞因子/趋化因子mRNA和蛋白的模式 表达，以及激活野生型T细胞和Th1克隆的能力 R-EAE的过继转移。此外，他们还将探索 CD28基因敲除小鼠对EAE诱导的抵抗力。目标2将 扩展他们已发表的研究，检查修改的影响 既往存在自身免疫性疾病的动物的共刺激信号 PLP139-151诱导SJL和(SJL×B10.PL)F1小鼠R-EAE模型的建立 其中复发是由于脑源性T细胞的激活 内源性的特异性；由表位扩散诱导的髓鞘表位。 抗B7-1完整单抗治疗小鼠的初步临床研究 发作导致复发率和恶化的显著增加 疾病的严重性。然而，对F(Ab)片段的类似处理 抗B7-1单抗阻断表位扩散，显著改善中枢神经系统 组织病理学检查，防止临床复发。细胞靶标和 完整抗B7-1介导的R-EAE加重及分子机制的研究 抗B7-1 F(Ab)片段诱导的疾病复发保护作用 通过检测Th1/Th2表型和功能反应进行评估 外周和中枢神经系统驻留的APC和T细胞 启动和复发相关的髓鞘表位。《目标3》将探讨 CTLA-4对R-EAE发病和表位扩散的负性调控作用 并确定B7-1、B7-2和CTLA-4在诱导和 静脉注射诱导的外周耐受性维持。注射法 多肽脉冲，ECDI固定的APC。这些研究应该增进我们的理解 共刺激分子在疾病发生和调控中的作用 表位在慢性自身免疫中的扩散并提供重要信息 与共刺激分子治疗的潜在靶向性有关 已经存在的免疫介导性疾病。

DESCRIPTION (Adapted from the Applicant's Abstract): Activation of naive T-cells requires TCR occupancy plus CD28-mediated co-stimulatory signals delivered by ligation of B7-1 and B7-2 on antigen presenting cells (APCs). In contrast, B7 ligation of CTLA-4 expressed on activated T-cells delivers a potent negative regulatory signal and may be involved in tolerance induction. These studies during the previous application period have clearly shown that strategies targeting these interactions have potent positive and negative regulatory activity on the initiation and progression of both acute and relapsing-remitting forms of experimental autoimmune encephalomyelitis (EAE), a CD4+ T-cell-mediated autoimmune disease which serves as a model for multiple sclerosis. The hypothesis under test in this renewal application is that the B7-CD28 co-stimulatory interactions play a critical role in positively regulating the activation and effector functions of autoreactive T-cells, while B7-CTLA-4 co-stimulatory interactions negatively regulate autoimmune disease and are required for the induction/maintenance of immune tolerance. Building on their preliminary data describing a new EAE model in NOD mice induced by priming with proteolipid protein peptide, PLP56-70, Aim 1 will employ a multifaceted approach, utilizing NOD mice with targeted deletion of B7-1, B7-2, B7-1/B7-2, or CD28 in comparison with antibody blocking studies, to further delineate the roles of the individual co-stimulatory receptors and ligands in the induction and effector phases of EAE and in activating myelin peptide-specific T-cell responses. This aim will also examine the individual roles of B7-1 and B7-2 in induction of cell surface activation and homing antigens, proliferation, patterns of cytokine/chemokine mRNA and protein expression, and the ability to activate wildtype T-cells and Th1 clones for adoptive transfer of R-EAE. In addition, they will explore the mechanistic basis behind the resistance of CD28 knockout mice to EAE induction. Aim 2 will expand their published studies examining the effects of modifying co-stimulatory signals in animals with a pre-existing autoimmune disease employing the PLP139-151-induced R-EAE model in SJL and (SJL x B10.PL)F1 mice in which relapses are due to the activation of encephalitogenic T-cells specific for endogenous; myelin epitopes induced by epitope spreading. Treatment of mice with intact anti-B7-1 mAb following the initial clinical episode results in a significant increase in relapse incidence and exacerbation of disease severity. However, similar treatment with the F(ab) fragments of anti-B7-1 mAb blocked epitope spreading, and significantly ameliorated CNS histopathology and prevented clinical relapses. The cellular targets and molecular mechanisms of intact anti-B7-1-mediated R-EAE exacerbation and anti-B7-1 F(ab) fragment-induced protection from disease relapses will be assessed by examining the Th1/Th2 phenotype and functional responses of peripheral and CNS-resident APCs and of T-cells specific for both the initiating and relapse-associated myelin epitopes. Aim 3 will explore the role of CTLA-4 in negatively regulating R-EAE pathogenesis and epitope spreading, and determine the role of B7-1, B7-2 and CTLA-4 in the induction and maintenance of peripheral tolerance induced by the i.v. injection peptide-pulsed, ECDI-fixed APCs. These studies should enhance our understanding of the role of co-stimulatory molecules in disease initiation and regulation of epitope spreading in chronic autoimmunity and provide vital information relative to the potential targeting of co-stimulatory molecules for treatment of pre-existing immune-mediated disorders.