The pathogenesis of toxoplasmic encephalitis (TE) in the immunocompromised patients who develop this disease is still poorly defined. To understand the mechanisms underlying the development of TE it is necessary to understand how protective immunity to Toxoplasma gondii is mediated in the brain. Data from this laboratory, and from other workers, indicate that T cells can mediate immunity to T. gondii in the brain, likely through production of IFN-gamma and that resident glial cell populations are involved in the immunopathogenesis of TE. The long term objective of this proposal is to understand how protective immunity against Toxoplasma gondii is mediated in the brain. This proposal has four specific aims designed to focus on the events that result in T cell mediated immunity to T. gondii in the brain; 1. Determine if the protective activity of IFN-gamma is through enhanced macrophage activation. These studies will be performed using a murine model of TE in which mice are treated with monoclonal antibodies to deplete IFN-gamma and exacerbate the TE or treated with recombinant IFN-gamma to ameliorate TE. The brains of treated mice will be analyzed using RT-PCR and immunocytochemistry to determine if these treatments affect macrophage activation. These studies are based on the use of immune-intervention to elucidate the mechanism whereby IFN-g mediates resistance to TE in the brain. 2. Determine the functional characteristics of the intracerebral T-cells involved in resistance or susceptibility to TE. Studies will be performed to characterize the cytokines produced by the intracerebral CD4+ and CD8+ T cells isolated from the brains of infected mice which differ in ability to resolve TE; if these T cells are cytotoxic for T gondii infected cells; and characterize the T cell expression of markers of activation such as CD45 and LFA-1. To test different T cell populations for ability to mediate resistance to T-gondii in the brain a model of reactivates of TE in SCID mice will be used. 3. Investigate the role of adhesion molecules in trafficking of T cell populations into the brain during TE. The expression of adhesion molecules during TE will be characterized using RT-PCR and immunocytochemistry. To determine which adhesion molecules are involved in trafficking of T cells into the brain during TE specific antibodies will be used to blockade adhesion molecules and the binding of parasite specific cells to brain endothelium measured in an in vitro assay and the ability of T cell to traffic into the brain assayed in vivo. Further experiments will be performed in which genetically altered strains of mice deficient in select adhesion molecules are used to characterize the mechanism whereby T cells traffic into the brain of mice with TE. 4. Characterize the role of astrocytes and microglia in the regulation of T cell mediated immunity and the immunopathogenesis of TE. Cultures of astrocytes or microglia will be stimulated with T.gondii to determine if this parasite will stimulate these cells to produce cytokines which affect T cell function. Further experiments will be performed to assess the role of these glia as accessory cells for parasite specific T cell responses. To assess the role of astrocyte production of cytokines on the immune response during TE, transgenic mice in which astrocytes over express either IL-3, IL-6, TNF-alpha or IFN-alpha will be infected with T-gondii and the development of TE assessed.
Showing the most recent 10 out of 49 publications
