Tolerance to proinsulin-2 is due to radioresistant thymic cells.

Proinsulin is a key Ag in type 1 diabetes, but the mechanisms regulating proinsulin immune tolerance are unknown. We have shown that preproinsulin-2 gene-deficient mice (proins-2(-/-)) are intolerant to proinsulin-2. In this study, we analyzed the mechanisms underlying T cell-mediated tolerance to proinsulin-2 in 129/Sv nonautoimmune mice. The expression of one proinsulin-2 allele, whatever its parental origin, was sufficient to maintain tolerance. The site of proinsulin-2 expression relevant to tolerance was evaluated in thymus and bone marrow chimeras. CD4+ T cell reactivity to proinsulin-2 was independent of proinsulin-2 expression in radiation-sensitive bone marrow-derived cells. A wt thymus restored tolerance in proins-2(-/-) mice. Conversely, the absence of the preproinsulin-2 gene in radioresistant thymic cells was sufficient to break tolerance. Although chimeric animals had proinsulin-2-reactive CD4+ T cells in their peripheral repertoire, they displayed no insulitis or insulin Abs, suggesting additional protective mechanisms. In a model involving transfer to immunodeficient (CD3epsilon(-/-)) mice, naive and proinsulin-2-primed CD4+ T cells were not activated, but could be activated by immunization regardless of whether the recipient mice expressed proinsulin-2. Furthermore, we could not identify a role for putative specific T cells regulating proinsulin-2-reactive CD4+ T in transfer experiments. Thus, proinsulin-2 gene expression by radioresistant thymic epithelial cells is involved in the induction of self-tolerance, and additional factors are required to induce islet abnormalities.

Role of beta-cells in type 1 diabetes pathogenesis.

Whether autoimmunity results primarily from a defect of the immune system, target organ dysfunction, or both remains an open issue in most human autoimmune diseases. The highly multigenic background on which diabetes develops in the NOD mouse and in the human suggests that numerous gene variants associate in contributing to activation of autoimmunity to beta-cells. Both immune genes and islet-related genes are involved. The presence of beta-cells is required for initiation of diabetes autoimmunity to proceed. Available experiments in the NOD mouse and epidemiological evidence in the human point to proinsulin as a key autoantigen in diabetes. The functional importance of insulin, the high number of autoantigens characterized at different stages of diabetes, and their clustering within beta-cell subparticles point to the islet as a starting point in the initiation phase of the disease. Genes that direct the autoimmune reaction toward the beta-cell target, autoantigens that are recognized by autoreactive B- and T-cells along the autoimmune process, the importance of beta-cells in the activation of autoreactive lymphocytes, and the expression level of key beta-cell molecules along diabetes development are successively considered in this review.

Expression of preproinsulin-2 gene shapes the immune response to preproinsulin in normal mice.

Deciphering mechanisms involved in failure of self tolerance to preproinsulin-2 is a key issue in type 1 diabetes. We used nonautoimmune 129SV/Pas mice lacking preproinsulin-2 to study the immune response to preproinsulin-2. In these mice, a T cell response was detected after immunization with several preproinsulin-2 peptides and confirmed by generating hybridomas. Activation of some of these hybridomas by wild-type (wt) islet cells or recombinant murine proinsulin-2 demonstrated that two epitopes can be generated from the naturally expressed protein. Although T cells from wt mice responded to preproinsulin-2 peptides, we could not detect a response to the naturally processed epitopes in these mice. Moreover, after immunization with recombinant whole proinsulin-2, a T cell response was detected in preproinsulin-2-deficient but not in wt mice. This suggests that islet preproinsulin-2-autoreactive T cells are functionally eliminated in wt mice. We used a transplantation model to evaluate the relevance of reactivity to preproinsulin-2 in vivo. Wild-type preproinsulin-2-expressing islets transplanted in preproinsulin-2-deficient mice elicited a mononuclear cell infiltration and insulin Abs. Graft infiltration was further increased by immunization with preproinsulin-2 peptides. Preproinsulin-2 expression thus shapes the immune response and prevents self reactivity to the islet. Moreover, islet preproinsulin-2 primes an immune response to preproinsulin-2 in deficient mice.