3, 3'-diindolylmethane, a natural aryl hydrocarbon receptor agonist, alleviates ulcerative colitis by enhancing "glycolysis-lactate-STAT3" and TIP60 signals-mediated Treg differentiation

Background: Aryl hydrocarbon receptor (AhR) plays an important role in the occurrence and development of ulcerative colitis (UC). In this study, the effect and mechanism of 3, 3'-diindolylmethane (DIM), the classical AhR agonist, on UC was investigated from the angle of recovering the balance of Th17/Treg.Methods: The in vivo colitis model was established in mice by using dextran sulfate sodium, and CD4+ T cells were used to simulate the in vitro differentiation of Treg and Th17 cells. The proportions and related factors of Th17 and Treg cells were measured using flow cytometry, Q-PCR and western blotting. The glycolysis was evaluated by examining the glucose uptake, glucose consumption and lactate production using kits or immunofluorescence. The activation of AhR was detected by western blotting and the XRE-luciferase reporter gene. The coimmunoprecipitation, transfection or other methods were selected to investigate and identify the signaling molecular pathway.Results: DIM significantly attenuated symptoms of colitis mice by rebuilding the balance of Th17/Treg in anoxic colons. In hypoxia, a more potent promotion of Treg differentiation was showed by DIM relative to normoxia, and siFoxp3 prevented DIM-suppressed Th17 differentiation. DIM repressed the excessive glycolysis in hypoxia evidenced by down-regulated glucose uptake, lactate production, Glut1 and HK2 levels. Interestingly, IL-10, the function-related factor of Treg cells, showed the feedback effect of DIM-suppressed glycolysis. Besides, 2-deoxyD-glucose, HK2 plasmid and IL-10 antibody prevented increase of DIM on the expression of Foxp3 at the transcriptional level and subsequent Treg differentiation through the lactate-STAT3 pathway, and reasons for the direct improvement of DIM on Foxp3 protein was attributed to promoting the formation of HIF-1 alpha/TIP60 complexes as well as subsequent acetylation and protein stability. Finally, AhR dependence and mechanisms for DIM-improved Treg differentiation in vitro and in vivo were well confirmed by using plasmids or inhibitors. Conclusions: DIM enhances activation of AhR and subsequent "glycolysis-lactate-STAT3 '' and TIP60 signalsmediated Treg differentiation.