Differential protection in two transgenic lines of NOD/Lt mice hyperexpressing the autoantigen GAD65 in pancreatic beta-cells.

Abstract

Although expressed at very low levels in islets of NOD mice, GAD65 is a candidate islet autoantigen. Two transgenic lines of NOD/Lt mice expressing high levels of human GAD65 from a rat insulin promoter were generated. Transgenes were integrated on proximal chromosome 15 of the A line and on the Y chromosome of the Y line. Transgenic A-line mice were obligate hemizygotes, since homozygous expression resulted in developmental lethality. A twofold higher level of hGAD65 transcripts in A-line islets from young donors was associated with higher GAD protein and enzyme activity levels. Y-line males developed diabetes at a similar rate and incidence as standard NOD/Lt males. In contrast, A-line mice of both sexes exhibited a markedly lowered incidence of diabetes. Insulitis, present in both transgenic lines, developed more slowly in A-line mice and correlated with a reduction in the ratio of gamma-interferon to interleukin-10 transcripts. Splenic leukocytes from young A-line donors transferred diabetes into NOD-scid recipients at a retarded rate compared with those from nontransgenic donors. Further, nontransgenic NOD T-cells transferred diabetes more slowly in NOD-scid recipients that were congenic for A-line transgenes as compared with standard NOD-scid recipients. Primed T-cell responses and spontaneous humoral reactivity to GAD65 failed to distinguish transgenic from nontransgenic mice. Quantitative differences in expression level or insertional mutagenesis are possible mechanisms of protection in the A line.

N‐terminal flanking residues of a diabetes‐associated GAD65 determinant are necessary for activation of antigen‐specific T cells in diabetes‐resistant mice

“…Finally, although the role of GAD65 in NOD diabetes remains ambiguous [35][36][37], there is consensus that GAD65 is an important target antigen that is also expressed in islet beta cells (Dai et al, unpublished data). Many studies involving GAD65 in NOD mice have centered on the immune response to the whole molecule.…”

“…A plethora of proteolytic enzymes could become engaged in this truncation process, and the activity of a protein such as Nramp1 (natural resistance-associated macrophage protein 1), which regulates intravesicular pH, could thereby be responsible for favoring the enzymatic activity of certain enzymes and disfavoring others. It is of great interest that the susceptible NOD and resistant NOR strains differ only in five major Idd regions, one of them, Idd5.2, coding for the Nramp1 molecule.Finally, although the role of GAD65 in NOD diabetes remains ambiguous [35][36][37], there is consensus that GAD65 is an important target antigen that is also expressed in islet beta cells (Dai et al, unpublished data). Many studies involving GAD65 in NOD mice have centered on the immune response to the whole molecule.…”

Prevention of type I diabetes transfer by glutamic acid decarboxylase 65 peptide 206-220-specific T cells

“…Peripheral blood leukocytes of all F 1 progeny were screened by flow cytometry for G206 or G286 TCR expression. The presence of RIP-huGAD transgene in genomic DNA was determined by PCR as described (23). NOD.RIP-huGAD.Yline males were crossed with G206 and G286 females.…”

Streptozotocin Upregulates GAD67 Expression in MIN6N8a Mouse Beta Cells

“…GAD is one of the putative autoantigens in both human and experimental autoimmune IDDM [4][5][6][7]. Transfer of GAD-specific T cells into non-diabetic recipients induced insulitis and IDDM [8][9][10][11] and the incidence of diabetes could be modulated according to GAD expression on beta cell [12][13][14]. More recent data suggested that GAD could also inhibit glucosestimulated insulin secretion by reducing glutamate levels in beta cell [15].…”