CpG Motifs of Bacterial DNA Exert Protective Effects in Mouse Models of IBD by Antigen-Independent Tolerance Induction

Background & AimsProphylactic treatment of mice with CpG motifs of bacterial DNA protects from experimental inflammatory bowel disease, at least partly via induction of inhibitory T-cells. The aim of this study was to elucidate whether these CpG-dependent protective effects require presence of bacterial flora suggesting antigen-specific regulatory activity.MethodsGerm-free BALB/c and IL-10−/− mice were treated with CpG-oligodeoxynucleotides (ODN), control-ODN, or PBS. CD4+CD62L+ cells of these mice were transferred into SCID recipients. CpG-ODN–treated germ-free IL-10−/− mice were transferred into colitogenic environment. Monoclonal antibodies were used to neutralize TGF-β and IFN-α/β during CpG-ODN treatment. CD4+CD62L+ cells of donors were evaluated for cytokine secretion and FOXP3, PD-1, and CD25 expression.ResultsCompared to PBS or control-ODN treatment, CpG-ODN application to germ-free donors led to decreased intestinal inflammation as indicated by histology, decreased proinflammatory cytokines, and increased IL-10 secretion. Protection was also observed after cotransfer of cells from PBS and CpG-ODN treated donors. Anti-TGF-β and anti-INF-α/β partly reversed the protective CpG-ODN effect. CpG-ODN–treated germ-free IL-10−/− mice transferred into colitogenic environment developed significantly less colitis than controls but not recipients of IL-10−/− CD4+CD62L+ cells. CD4+CD62L+ cells of CpG-treated germ-free animals displayed increased expression of regulatory markers.ConclusionsEven without pre-existence of bacterial flora CpG-ODN exposition induces tolerance, indicating that CpG-ODN–induced regulatory T-cells are not bacterial antigen specific. TGF-β and IFN-α/β play major roles in induction of regulatory cells, and although IL10–independent mechanisms play a role in CpG-ODN protection, this cytokine likely is important for the effector mechanism of CpG-ODN–induced regulatory T-cells. Prophylactic treatment of mice with CpG motifs of bacterial DNA protects from experimental inflammatory bowel disease, at least partly via induction of inhibitory T-cells. The aim of this study was to elucidate whether these CpG-dependent protective effects require presence of bacterial flora suggesting antigen-specific regulatory activity. Germ-free BALB/c and IL-10−/− mice were treated with CpG-oligodeoxynucleotides (ODN), control-ODN, or PBS. CD4+CD62L+ cells of these mice were transferred into SCID recipients. CpG-ODN–treated germ-free IL-10−/− mice were transferred into colitogenic environment. Monoclonal antibodies were used to neutralize TGF-β and IFN-α/β during CpG-ODN treatment. CD4+CD62L+ cells of donors were evaluated for cytokine secretion and FOXP3, PD-1, and CD25 expression. Compared to PBS or control-ODN treatment, CpG-ODN application to germ-free donors led to decreased intestinal inflammation as indicated by histology, decreased proinflammatory cytokines, and increased IL-10 secretion. Protection was also observed after cotransfer of cells from PBS and CpG-ODN treated donors. Anti-TGF-β and anti-INF-α/β partly reversed the protective CpG-ODN effect. CpG-ODN–treated germ-free IL-10−/− mice transferred into colitogenic environment developed significantly less colitis than controls but not recipients of IL-10−/− CD4+CD62L+ cells. CD4+CD62L+ cells of CpG-treated germ-free animals displayed increased expression of regulatory markers. Even without pre-existence of bacterial flora CpG-ODN exposition induces tolerance, indicating that CpG-ODN–induced regulatory T-cells are not bacterial antigen specific. TGF-β and IFN-α/β play major roles in induction of regulatory cells, and although IL10–independent mechanisms play a role in CpG-ODN protection, this cytokine likely is important for the effector mechanism of CpG-ODN–induced regulatory T-cells.