Retroviral delivery of GAD-IgG fusion construct induces tolerance and modulates diabetes: a role for CD4+ regulatory T cells and TGF-beta?
Previous studies have demonstrated that antigen-specific tolerance could be induced by lipopolysaccharide (LPS)-stimulated B cells retrovirally transduced with an immunoglobulin-antigen (or epitope-containing peptide) fusion construct. To investigate the mechanism of this gene therapy system, we now adapted this approach to immunotherapy of spontaneous diabetes in nonobese diabetic (NOD) mice, a T-cell-mediated autoimmune disease triggered, in part, by a pathogenic response to glutamate decarboxylase (GAD) 65. We demonstrate that LPS-stimulated splenocytes, retrovirally transfected with GAD-IgG fusion construct, induce a significant antigen-specific hyporesponsiveness at both cellular and humoral levels and reduce the incidence of diabetes in female NOD mice. Parallel with disease protection, we observed a prolonged increase of the numbers of CD4+CD25+ T cells in the periphery of GAD-IgG-treated mice, compared to those treated with a control IgG vector, both in the prediabetic period and persisting even 8 months after gene therapy. This increase appeared to be induced by the repeated stimulation of the antigen in the periphery instead of a result of differentiation of T-cell precursor in the thymus. Moreover, CD4+CD25+ T cells induced by GAD-IgG fusion construct were capable of suppressing the proliferative response of CD4+CD25- T cells in vitro; and ablation of the activity of CD4+CD25+ T cells by blocking antibody against CD25 could reverse GAD-specific T-cell hyporesponsiveness. These results suggested that CD4+CD25+ T-cell subset induced in GAD-IgG-treated NOD mice represented the regulatory or suppressive CD4+CD25+ T cells (Treg) and might play an important role in the induction and maintenance of tolerance in NOD mice. Furthermore, the numbers of splenic CD4+CD62L+ regulatory T cells in GAD-IgG-treated mice during the prediabetic period and serum TGF-beta levels in 34-38-week-old GAD-IgG-protected mice were also increased, compared to control IgG-treated ones. Therefore, we propose that the induction of tolerance and the prevention of diabetes incidence in NOD female mice induced by the GAD-IgG fusion construct may require CD4+ regulatory T cells, and the possible mediation of TGF-beta.
Song
L
L
Department of Molecular Immunology, Institute of Basic Medical Sciences, Beijing, PR China.
Wang
J
J
Wang
R
R
Yu
M
M
Sun
Y
Y
Han
G
G
Li
Y
Y
Qian
J
J
Scott
D W
DW
Kang
Y
Y
Soukhareva
N
N
Shen
B
B
D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D018920:Coculture Techniques; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology / Q000628:therapy*; D005260:Female; D005434:Flow Cytometry; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000235:genetics; D005968:Glutamate Decarboxylase / Q000235:genetics* / Q000378:metabolism; D007108:Immune Tolerance; D007074:Immunoglobulin G / Q000235:genetics* / Q000378:metabolism; D016753:Interleukin-10 / Q000276:immunology; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D016688:Mice, Inbred NOD; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012190:Retroviridae / Q000235:genetics*; D016212:Transforming Growth Factor beta / Q000276:immunology
D007074:Immunoglobulin G; D008070:Lipopolysaccharides; D011993:Recombinant Fusion Proteins; D016212:Transforming Growth Factor beta; D016753:Interleukin-10; D005968:Glutamate Decarboxylase