Regulatory CD4/CD8 T cells in autoimmune demyelination

描述(由申请人提供)：免疫调节在控制成熟的自我反应性、潜在致病的T细胞方面发挥着关键作用。不同的细胞成分及其在调控机制中的相互作用还不是很清楚。在这里，我们将研究显性负反馈机制如何控制髓鞘碱性蛋白(MBP)反应性Vbeta8.2+T细胞介导实验性自身免疫性脑脊髓炎，H-2U小鼠是多发性硬化症的原型。这种调节集中在调节CD8和CD4T细胞(Treg)分别识别I类和II类MHC背景下的TCR肽/MHC复合体。我们已经提出(见图1)，CD8Treg在CD8Treg的募集/激活中提供帮助，最终诱导活化的Vbeta8.2+Th1细胞凋亡。致病Th1细胞的耗尽和非致病Th2细胞的扩张最终导致抗MBP应答的免疫偏离和对EAE的保护。CD4Treg可与骨架3区多肽BS反应，而CD8Treg可识别TCR Vbeta8.2链上的CDR1/2区肽B3或p41-50。通过CDR3长度分型，或免疫显微镜分析和CFSE标记细胞的Annexin-V染色，我们将确定Vbeta8.2+T细胞在体内经过诱导或生理调节后的命运。我们将分别确定1型和2型细胞因子在调节的初始和后期阶段中的作用。将产生CD8 Treg克隆，并对其MHC限制性、TCR-V基因用途和功能特性进行鉴定，包括其体外和体内的细胞溶解活性，以及过继转移后控制疾病的能力。我们将用B10.PL.lpr、B10.PL.gld和B10.PL.Proublin-/-小鼠来确定CD8 Treg的杀伤机制。了解这里涉及的免疫原理不仅对于理解免疫调节的生物学很重要，而且对于设计基于TCR的治疗人类自身免疫疾病的方法也很重要。

DESCRIPTION (provided by applicant): Immune regulation plays a critical role in the control of mature self-reactive, potentially pathogenic T cells. Different cellular components and their interactions in regulatory mechanisms are not well understood. Here we will investigate how a dominant negative feedback mechanism controls myelin basic protein (MBP)-reactive Vbeta8.2+ T cells mediating experimental autoimmune encephalomyelitis in the H-2u mouse, a prototype for multiple sclerosis. This regulation is focused on the recognition of TCR peptide/MHC complexes in class I and II MHC contexts by regulatory CD8 and CD4 T cells (Treg) respectively. We have proposed (see Figure 1) that CD4 Treg provide help in the recruitment/activation of CD8 Treg, which ultimately induce apoptosis in activated Vbeta8.2+ Th1 cells. The depletion of pathogenic Th1 and expansion of non-pathogenic Th2 cells eventually leads to immune deviation of the anti-MBP response and protection from EAE. CD4 Treg are reactive to a framework 3-region peptide, BS, whereas CD8 Treg recognize the CDR1/2 region peptide, B3 or p41-50, from the TCR Vbeta8.2 chain. Using CDR3 length spectratyping, or immunoscope analysis and annexin-V staining of CFSE-labeled cells we will determine the fate of Vbeta8.2+ T cells following induced or physiological regulation in vivo. We will determine the role of type 1 and type 2 cytokines in the initial and late phases of regulation, respectively. CD8 Treg clones will be generated and characterized for their MHC-restriction, TCR-V gene usage and functional properties, including their cytolytic activity both in vitro and in vivo, and their ability to control disease upon adoptive transfer. We will determine the mechanism of killing by the CD8 Treg using B10.PL.lpr, B10.PL.gld and B10.PL.perforin-/- mice. Knowledge of the immune principles involved here is important not only for understanding the biology of immune regulation but also in the design of TCR-based therapeutic approaches for autoimmune conditions in humans.