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 CD4CD25Foxp3 regulatory T cells and increases transforming growth factor (TGF)-β1 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-β1 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 CD4CD25Foxp3 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-β1 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 β cells in the islets. CONCLUSIONS.: Our results demonstrated that CFA treatment ameliorates autoimmunity in diabetic NOD mice by up-regulating CD4CD25Foxp3 regulatory T cells and increasing TGF-β1 production. Exendin-4 enhanced the effect of CFA on reversing diabetes in NOD mice by stimulating β-cell replication.