An examination of the role of GAD65-specific T cells in diabetes pathogenesis and differential allelic regulation of IFN-gamma using transgenic model systems

GAD65 is an important antigen in both human diabetes mellitus and the non-obese diabetic (NOD) mouse, but the role of GAD-specific T cells in diabetes pathogenesis is unclear. We have generated NOD transgenic mice expressing T cell receptors specific for two immunodominant I-Ag7-restricted GAD65 peptide epitopes, 206-220 and 286-300. This is the first description of TCR transgenics for a known β-cell autoantigen. Lymphocytes from p286-300-specific TCR transgenic mice (G286) proliferate and make Th1 cytokines when stimulated in vitro with peptide 286-300, yet G286 mice do not develop diabetes or insulitis. CD4+ T cells from G286 mice delay diabetes transferred to NOD scid mice by diabetic NOD spleen cells, suggesting that T cells from G286 mice have a disease protective capability. I-Ag7/p286-specific T cells have been identified with a tetramer reagent. Activated CD4+tetramer+ cells from G286 mice can also delay diabetes transfer. Furthermore, in vitro activated CD4 T cells from G286 mice express higher levels of CTLA-4 than non-transgenic littermates. Preliminary characterization of transgenic mice expressing a TCR specific for GAD65 peptide 206-220 indicates that these mice do not develop accelerated diabetes, suggesting that this T cell response is also not pathogenic.; Part II of this thesis studies epigenetic regulation of the IFNγ gene. Some cytokine genes, including IL-2 and IL-4, are controlled by monoallelic regulation. We employed transgenic mice expressing the huThy1 gene under the control of the murine IFNγ promoter to determine if IFNγ also displays differential allelic regulation. HuThy1 expression indicates activation from the transgenic promoter, which can be distinguished from activation at the two native alleles. We found both IFNγ and huThy1 were expressed in CD4+ T cells under Th1 but not Th2 growth conditions, but were usually not expressed within the same cells. The low number of cells expressing both IFNγ and huThy1 suggest that separate alleles of the IFNγ promoter can be regulated independently. Increasing histone acetylation in transgenic reporter T cells increases the percentage of cells expressing both IFNγ and huThy1, indicating that allelic differences in histone acetylation may control allelic differences in IFNγ expression.