This project proposes to utilize a transgenic animal model of virus induced IDDM (RIP-LCMV) to study the pathogenesis and test novel therapeutic approaches for autoimmune diabetes. In previous studies, I has developed and characterized an interesting model of virus-induced diabetes in which an LCMV-derived viral protein (NP or GP) is placed under the regulation of an insulin promoter and this construct is incorporated into the mouse germline as a transgene. This transgenic animal will not develop IDDM; however, if it is infected with LCMV, then diabetes is induced in a rapid fashion (14 to 21 days). Further, I have demonstrated that if this same LCMV protein is simultaneously expressed in the thymus, then disease onset is significantly delayed (up to as long as 200 days, depending on the strain of mouse and H-2 haplotype). In addition, the delayed onset form of IDDM is mediated by CD4+ T helpers together with low affinity CD8+ T cells, whereas the acute version is mediated by high affinity CD8+ T cells. I propose to utilize this model system to study several interesting features of IDDM pathogenesis. There are four specific aims: 1. To determine the regulatory role of cytokines in autoimmune diabetes by assessing cytokine profiles during pathogenesis and using targeted short-term application of anti-cytokine antibodies to modulate IDDM. Th1 and Th2 cytokine profiles of lymphocytes infiltrating islets will be assessed at various times in both the fast and slow IDDM model. This information will be used to design targeted short-time application of anti-cytokine antibodies during the pathogenesis of IDDM to attempt to modify the outcome of the autoimmune process and minimize side effects. 2. To assess the efficacy of therapy in the slow-onset IDDM by oral tolerance induction to insulin and insulin analogs. These experiments will test the hypothesis that oral tolerance to insulin prevents IDDM via a """"""""bystander"""""""" effect of Th2 induced cytokines. 3. To determine the mechanism of beta-cell destruction by anti-self CTL in fast-onset and slow-onset IDDM by assessing the role of perforin and affinity of the CTL. These experiments will utilize double transgenic RIP-LCMV/perforin knockout mice to assess the effect of perforin and transfer experiments to assess the importance of affinity. 4. To assess the prevention of slow-onset and fast-onset IDDM by specific tolerization of anti-self CTL by the use of DNA vaccination. This will be assessed using plasmids expressing LCMV proteins and various injection routes.
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