Regulatory CD4/CD8 T cells in autoimmune demyelination

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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.