Experimental autoimmune encephalomyelitis (EAE) is a Th1 and Th17 cell-mediated autoimmune disease of the CNS. IDO and tryptophan metabolites have inhibitory effects on Th1 cells in EAE. For Th17 cells, IDO-mediated tryptophan deprivation and small molecule halofuginone-induced amino acid starvation response were shown to activate general control nonrepressed 2 (GCN2) kinase that directly or indirectly inhibits Th17 cell differentiation. However, it remains unclear whether IDO and tryptophan metabolites impact the Th17 cell response by mechanisms other than the GCN2 kinase pathway. In this article, we show that IDO-deficient mice develop exacerbated EAE with enhanced encephalitogenic Th1 and Th17 cell responses and reduced regulatory T cell (Treg) responses. Administration of the downstream tryptophan metabolite 3-hydroxyanthranillic acid (3-HAA) enhanced the percentage of Tregs, inhibited Th1 and Th17 cells, and ameliorated EAE. We further demonstrate that Th17 cells are less sensitive to direct suppression by 3-HAA than are Th1 cells. 3-HAA treatment in vitro reduced IL-6 production by activated spleen cells and increased expression of TGF-β in dendritic cells (DCs), which correlated with enhanced levels of Tregs, suggesting that 3-HAA–induced Tregs contribute to inhibition of Th17 cells. By using a DC–T cell coculture, we found that 3-HAA–treated DCs expressed higher levels of TGF-β and had properties to induce generation of Tregs from anti-CD3/anti-CD28–stimulated naive CD4⁺ T cells. Thus, our data support the hypothesis that IDO induces the generation of Tregs via tryptophan metabolites, such as 3-HAA, which enhances TGF-β expression from DCs and promotes Treg differentiation.

Experimental autoimmune encephalomyelitis (EAE) serves as a model for multiple sclerosis and is considered a CD4⁺, Th1 cell-mediated autoimmune disease. IL-12 is a heterodimeric cytokine, composed of a p40 and a p35 subunit, which is thought to play an important role in the development of Th1 cells and can exacerbate EAE. We induced EAE with myelin oligodendrocyte glycoprotein (MOG) peptide 35–55 (MOG_35–55) in C57BL/6 mice and found that while IL-12p40-deficient (−/−) mice are resistant to EAE, IL-12p35^−/− mice are susceptible. Typical spinal cord mononuclear cell infiltration and demyelination were observed in wild-type and IL-12p35^−/− mice, whereas IL-12p40^−/− mice had normal spinal cords. A Th1-type response to MOG_35–55 was observed in the draining lymph node and the spleen of wild-type mice. A weaker MOG_35–55-specific Th1 response was observed in IL-12p35^−/− mice, with lower production of IFN-γ. By contrast, a Th2-type response to MOG_35–55 correlated with disease resistance in IL-12p40^−/− mice. Production of TNF-α by microglia, CNS-infiltrating macrophages, and CD4⁺ T cells was detected in wild-type and IL-12p35^−/−, but not in IL-12p40^−/−, mice. In addition, NO production was higher in IL-12p35^−/− and wild-type mice than in IL-12p40^−/− mice. These data demonstrate a redundancy of the IL-12 system in the induction of EAE and suggest that p40-related heterodimers, such as the recently cloned IL-23 (p40p19), may play an important role in disease pathogenesis.