Background CD4+CD25+ regulatory T cell (Treg)-based immunotherapy is considered a promising regimen for controlling the progression of autoimmune diabetes. In this study, we tested the hypothesis that the therapeutic effects of Tregs in response to the antigenic epitope stimulation depend on the structural properties of the epitopes used. Methodology/Principal Findings Splenic lymphocytes from nonobese diabetic (NOD) mice were stimulated with different glutamic acid decarboxylase (GAD)-derived epitopes for 7–10 days and the frequency and function of Tregs was analyzed. We found that, although all expanded Tregs showed suppressive functions in vitro, only p524 (GAD524–538)-expanded CD4+CD25+ T cells inhibited diabetes development in the co-transfer models, while p509 (GAD509–528)- or p530 (GAD530–543)-expanded CD4+CD25+ T cells had no such effects. Using computer-guided molecular modeling and docking methods, the differences in structural characteristics of these epitopes and the interaction mode (including binding energy and identified domains in the epitopes) between the above-mentioned epitopes and MHC class II I-Ag7 were analyzed. The theoretical results showed that the epitope p524, which induced protective Tregs, possessed negative surface-electrostatic potential and bound two chains of MHC class II I-Ag7, while the epitopes p509 and p530 which had no such ability exhibited positive surface-electrostatic potential and bound one chain of I-Ag7. Furthermore, p524 bound to I-Ag7 more stably than p509 and p530. Of importance, we hypothesized and subsequently confirmed experimentally that the epitope (GAD570–585, p570), which displayed similar characteristics to p524, was a protective epitope by showing that p570-expanded CD4+CD25+ T cells suppressed the onset of diabetes in NOD mice. Conclusions/Significance These data suggest that molecular modeling-based structural analysis of epitopes may be an instrumental tool for prediction of protective epitopes to expand functional Tregs.
Geng L, Solimena M, Flavell RA, Sherwin RS, Hayday AC (1998) Widespread expression of an autoantigen-GAD65 transgene does not tolerize non-obese diabetic mice and can exacerbate disease. Proc Natl Acad Sci USA 95: 10055–10060.
Quinn A, McInerney B, Reich EP, Kim O, Jensen KP, et al. (2001) Regulatory and effector CD4 T cells in nonobese diabetic mice recognize overlapping determinants on glutamic acid decarboxylase and use distinct V β genes. J Immunol 166: 2982–2991.
Han G, Li Y, Wang J, Wang R, Chen G, et al. (2005) Active tolerance induction and prevention of autoimmune diabetes by immunogene therapy using recombinant adenoassociated virus expressing glutamic acid decarboxylase 65 peptide GAD500–585. J Immunol 174: 4516–4524.
Song L, Wang J, Wang R, Yu M, Sun Y, et al. (2004) Retroviral delivery of GAD-IgG fusion construct induces tolerance and modulates diabetes: a role for CD4+ regulatory T cells and TGF-β? Gene Ther 11: 1487–1496.
Pop SM, Wong CP, Culton DA, Clarke SH, Tisch R (2005) Single cell analysis shows decreasing FoxP3 and TGF-β1 coexpressing CD4+CD25+ regulatory T cells during autoimmune diabetes. J Exp Med 201: 1333–1346.
Masteller EL, Warner MR, Tang Q, Tarbell KV, McDevitt H, et al. (2005) Expansion of functional endogenous antigen-specific CD4+CD25+ regulatory T cells from nonobese diabetic mice. J Immunol 175: 3053–3059.
Dai YD, Jensen KP, Marrero I, Li N, Quinn A, et al. (2008) N-terminal flanking residues of a diabetes-associated GAD65 determinant are necessary for activation of antigen-specific T cells in diabetes-resistant mice. Eur J Immunol 38: 968–976.
Tarbell KV, Lee M, Ranheim E, Chao CC, Sanna M, et al. (2002) CD4+ T cells from glutamic acid decarboxylase (GAD)65-specific T cells receptor transgenic mice are not diabetogenic and can delay diabetes transfer. J Exp Med 196: 481–492.
Fisson S, Djelti F, Trenado A, Billiard F, Liblau R, et al. (2006) Therapeutic potential of self-antigen-specific CD4+CD25+ regulatory T cells selected in vitro from a polyclonal repertoire. Eur J Immunol 36: 817–827.
Ochoa-Repáraz J, Riccardi C, Rynda A, Jun S, Callis G, et al. (2007) Regulatory T cell vaccination without autoantigen protects against experimental autoimmune encephalomyelitis. J Immunol 178: 1791–1799.