Project Summary/Abstract Type 1 Diabetes (T1D) is an autoimmune disease thought to be mediated by autoreactive CD4 and CD8 T cell that orchestrate the death of insulin-producing beta cells within pancreatic islets. This proposal aims to investigate how antigen-specific immunotherapy (ASI) with a CD4-specific neoepitope can selectively suppress autoreactive T cells by tolerogenic delivery of antigens in the NOD mouse model of autoimmune diabetes. Previously, our lab demonstrated that induction of tolerance to a dominant CD4 neoepitope, the 2.5 Hybrid Insulin Peptide (2.5HIP), can prolong islet graft survival in diabetic NOD mice. In the outlined studies, we will investigate the mechanistic impact of ASI on the differentiation and function of effector CD4 and CD8 T cells in islet grafts. The role of IL-10-producing regulatory T cells, expanded by 2.5HIP tolerance induction, will also be examined. We hypothesize that induction of tolerance to the 2.5HIP expands antigen-specific IL10-producing regulatory T cells that, directly or indirectly, arrest the effector differentiation and function of autoreactive CD4 and CD8 T cells within islet grafts. The specific aims of the study are to (1) determine the differentiation and functional states of antigen-specific CD4 and CD8 T cells following induction of tolerance to the hybrid insulin peptide, and (2) determine the contribution of IL10-producing regulatory T cells to prolonged graft survival and the suppression of autoreactive T cells. The completion of the proposed studies is expected to generate high-impact information on the mechanisms underlying autoreactive T cell suppression and differentiation following induction of antigen- specific tolerance. Findings from this research will also critically inform antigen-specific therapeutic approaches for human T1D and other autoimmune diseases.