Upregulating CD4+CD25+FOXP3+ Regulatory T Cells in Pancreatic Lymph Nodes in Diabetic NOD Mice by Adjuvant Immunotherapy

Background. Immunotherapy with Complete Freund's adjuvant (CFA) is effective in ameliorating autoimmunity in diabetic nonobese diabetic (NOD) mice. We investigated whether CFA treatment up-regulates CD4(+)CD25(+)Foxp3(+) regulatory T cells and increases transforming growth factor (TGF)-beta 1 production in diabetic NOD mice. Methods. New-onset diabetic NOD mice were treated with CFA and exendin-4, a potent analog of glucagon-like peptide-1. Reversal of diabetes was determined by monitoring blood glucose level. Ameliorating autoimmunity through immunoregulation was assessed by adoptive transfer. Regulatory T cells in the peripheral blood, spleen, thymus, and pancreatic nodes were measured. TGF-beta 1 in plasma and the insulin content in the pancreas were also measured. Immunostainings for insulin and BrdU were performed. Results. New-onset diabetes could be reversed in 38% of NOD mice treated with CFA alone and in 86% of NOD mice treated with both CFA and exendin-4. Diabetes adoptive transfer by splenocytes from CFA-treated NOD mice was delayed. The percentage of CD4(+)CD25(+)Foxp3(+) regulatory T cells in the pancreatic lymph nodes of CFA-treated NOD mice was significantly increased at 1, 5, and 15 to 17 weeks after treatment. TGF-beta 1 in the plasma of CFA-treated NOD mice was also significantly increased. Combining CFA with exendin-4 treatment significantly increased the insulin content and the numbers of insulin and BrdU double-labeled beta cells in the islets. Conclusions. Our results demonstrated that CFA treatment ameliorates autoimmunity in diabetic NOD mice by up-regulating CD4(+)CD25(+)Foxp3(+) regulatory T cells and increasing TGF-beta 1 production. Exendin-4 enhanced the effect of CFA on reversing diabetes in NOD mice by stimulating P-cell replication.