Immunization of SJL (H-2s) mice with the PLP 139-151 peptide (HSLGKWLGHPDKF) induces a very potent autoimmune disease of the CNS called Experimental autoimmune encephalomyelitis (EAE). By replacing principal TcR contact residues in the encephalitogenic peptide we have generated a panel of altered peptide ligands (APLs) that can prevent and/or reverse EAE. These APLs not only inhibit EAE induced with the PLP 139-151 peptide but also disease induced by other encephalitogenic myelin antigens including myelin basic protein (MBP) and myelin oligodendrocyte glycoprotein (MOG). Thus APLs of self peptides provide a powerful tool for the regulation of autoimmune diseases. Preliminary evidence suggests that the APLs that protect mice from EAE induce a shift in the cytokine balance from a pro-inflammatory Th1 (IFN- gamma, TNF) to an anti-inflammatory Th2 (IL-4, IL-10) cytokine phenotype in the responding T cells. However, the cellular and molecular mechanisms involved in this cytokine shift have not yet been determined. To study this mechanism, we propose the following specific aims: I. Determine a) whether APLs interact with the encephalitogenic precursor T cells and differentiate these cells along the Th2 pathway; b) whether APLs induce a protective Th2 T cell repertoire that is distinct from the autopathogenic Th1 repertoire or c) both. II. Determine whether APLs derived from self peptides, that induce production of pro-inflammatory cytokines from Th2 cells, can break self tolerance and promote induction of autoimmunity in the EAE-resistant B10.2 mouse strain that is H-2 congenic with the susceptible SJL strain. III. Define the intracellular signaling events induced in T cells that is associated with deviation of cytokine production. This will be undertaken in collaboration with Dr. Steven Burakoff's laboratory by analyzing membrane proximal signaling events including molecules that positively (CD3-zeta and ZAP-70 phosphorylation) and negatively (Cb1 and SHP protein tyrosine phosphatases) regulate TcR-mediated signals. These molecules will be undertaken in parallel with the human T cell clones (project 3) and will thus provide a more comprehensive analysis on the mechanisms by which APLs achieve cytokine shift. These studies will provide fundamental information on the cellular and molecular mechanisms by which APLs induce protective or pathogenic autoreactive T cell responses and alter the course of the CNS autoimmune disease, EAE.
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