Microbial metabolite butyrate modulates granzyme B in tolerogenic IL-10 producing Th1 cells to regulate intestinal inflammation

CD4⁺ T cells play a critical role in regulating autoimmune diseases, and intestinal microbial metabolites control various immune responses. Granzyme B (GzmB)-producing CD4⁺ T cells have been recently reported to participate in the pathogenesis of autoimmune diseases. Here, we found that GzmbB-deficient CD4⁺ T cells induced more severe colitis in Rag1^−/− mice than wild-type (WT) CD4⁺ T cells. Germ-free (GF) mice exhibited a lower expression of GzmB in intestinal CD4⁺ T cells compared to specific pathogen-free (SPF) mice. Intestinal microbial metabolite butyrate increased GzmB expression in CD4⁺ T cells, especially in IL-10-producing Th1 cells, through HDAC inhibition and GPR43, but not GPR41 and GPR109a. Butyrate-treated GzmB-deficient CD4⁺ T cells demonstrated more severe colitis compared to butyrate-treated WT CD4⁺ T cells in the T cell transfer model. Butyrate altered intestinal microbiota composition, but altered microbiota did not mediate butyrate induction of intestinal CD4⁺ T cell expression of GzmB in mice. Blimp1 was involved in the butyrate induction of GzmB in IL-10-producing Th1 cells. Glucose metabolism, including glycolysis and pyruvate oxidation, mediated butyrate induction of GzmB in Th1 cells. In addition, we found that IKZF3 and NR2F6 regulated GzmB expression induced by butyrate. Together, our studies underscored the critical role of GzmB in mediating gut bacterial metabolite butyrate regulation of T cell tolerance at the mucosal surface.