Experimental allergic encephalomyelitis (EAE) is an inflammatory demyelinating disease of the central nervous system (CNS) that is an animal model for the human disease multiple sclerosis (MS). EAE is initiated by CD4 TCR alpha beta lymphocytes reactive to myelin antigens that initiate an inflamatory cascade in the central nervous system (CNS) through the release of cytokines. Recent results from this laboratory have shown that gamma delta T cells are present in lesioned areas of the brain at sites that colocalize with altered expression of heat shock proteins (HSP) and contribute to disease pathogenesis. This is the first report that documents a role for these cells in disease expression. At the present time nothing is known about how gamma delta T cells might facilitate the development of EAE. In this continuation application, we plan to define the role of gamma delta T cells in EAE and extend our studies of altered HSP expression in the CNS. The specific hypothesis to be tested is that gamma delta T cells contribute to the pathogenesis of EAE by augmenting the inflammatory cascade, and are activated by altered HSP expression in the inflamed CNS.
Four specific aims are proposed. In the first aim, using a combination of FACS analysis and in vitro assays, the phenotypic and functional characteristics of gamma delta T cells present in the lesion at varying times in the disease process will be defined.
This aim will determine if different subsets of gamma delta T cells, with distinct functional attributes, are correlated with specific phases of EAE. The second specific aim will seek to define the affects of gamm delta T cell depletion on EAE by determining changes in the distribution and nature of the inflammatory infiltrates, the types and relative levels of different cytokines and chemokines in the CNS, as well as changes in potential effector mechanisms. The third specific aim will investigate the role of altered heat shock proteins as immunogenic targets of gamma delta T cells using a combination of in vitro proliferation and cytotoxicity assays to HSP expressing targets, as well as analyses of T cell receptor usage. The fourth specific aim will examine the expression of HSP 70 and HSP 27 at varying times in the disease process using a combination of immunohistochemistry and semi-quantitative western blotting. The experiments planned are designed to provide insights into the pathogenic mechanisms involved in the development of the primary demyelinating diseases, and to provide an experimental paradigm that complements parallel studies in MS.
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