Induction of Autoantigen-Specific Th2 and Tr1 Regulatory T Cells and Modulation of Autoimmune Diabetes

Abstract

Autoantigen-based immunotherapy can modulate autoimmune diabetes, perhaps due to the activation of Ag-specific regulatory T cells. Studies of these regulatory T cells should help us understand their roles in diabetes and aid in designing a more effective immunotherapy. We have used class II MHC tetramers to isolate Ag-specific T cells from nonobese diabetic (NOD) mice and BALB/c mice treated with glutamic acid decarboxylase 65 peptides (p206 and p221). Based on their cytokine secretion profiles, immunization of NOD mice with the same peptide induced different T cell subsets than in BALB/c mice. Treatment of NOD mice induced not only Th2 cells but also IFN-γ/IL-10-secreting T regulatory type 1 (Tr1) cells. Adoptive transfer experiments showed that isolated tetramer+ T cells specific for p206 or p221 could inhibit diabetes development. These cells were able to suppress the in vitro proliferation of other NOD mouse T cells without cell-cell contact. They performed their regulatory functions probably by secreting cytokines, and Abs against these cytokines could block their suppressive effect. Interestingly, the presence of both anti-IL-10 and anti-IFN-γ could enhance the target cell proliferation, suggesting that Tr1 cells play an important role. Further in vivo experiments showed that the tetramer+ T cells could block diabetogenic T cell migration into lymph nodes. Therefore, treatment of NOD mice with autoantigen could induce Th2 and Tr1 regulatory cells that can suppress the function and/or block the migration of other T cells, including diabetogenic T cells, and inhibit diabetes development.

Presence of Diabetes-Inhibiting, Glutamic Acid Decarboxylase-Specific, IL-10-Dependent, Regulatory T Cells in Naive Nonobese Diabetic Mice

“…8B). Consistent with our previous studies using T cells isolated from p221-immunized NOD mice, these results suggest that the migration of diabetogenic T cells into pancreatic islets was blocked by N221 ϩ cells (20). Therefore, these N221 ϩ T cells exhibit regulatory functions in vivo and efficiently prevent both insulitis and diabetes development.…”

“…The p221 peptide has been identified as one of the most dominant epitopes derived from the GAD protein and may play an important role in diabetes development (19,26). We have previously shown that immunization of NOD mice with p221 could activate p221-specific regulatory T cells, leading to the suppression and delay of diabetes development (20). In this report we provide evidences demonstrating that T cells specific for p221 are present in unimmunized naive NOD mice.…”

“…To date, we have focused our studies on the GAD 221-235 peptide (p221), one of the most immunogenic GAD determinants. Our previous studies have shown that immunization of NOD mice with p221 led to the generation of p221-specific Tr cells that could prevent diabetes development (20). However, those studies left unclear whether p221-specific Tr cells arise after immunization with p221 or whether such cells develop spontaneously in NOD mice.…”

“…2A). We then performed experiments to compare simultaneous secretion of 16 different cytokines/chemokines by N221 ϩ cells with the previously described p221-specific T cells isolated from p221-immunized NOD mice (20). In addition to the secretion of IFN-␥ and IL-10, both p221-specific T cell lines secreted MIP-1␣ and GM-CSF (Ͼ10,000 pg/ml) and a lower level of RANTES (ϳ500 -1,000 pg/ml), TNF-␣ (ϳ1,500 -2,500 pg/ml), and IL-6 (ϳ400 pg/ml; data not shown).…”

“…However, those studies left unclear whether p221-specific Tr cells arise after immunization with p221 or whether such cells develop spontaneously in NOD mice. Attempts to address this question have been hampered by the fact that autoantigen-specific T cells are present at a very low frequency in unimmunized naive NOD mice (20). To avoid this problem, we used a novel approach involving class II MHC I-Ag7 tetramers specific for p221 to identify and isolate a large number of p221-specific T cells from unimmunized naive NOD mice.…”