P0029IL4I1 regulates macrophage senescence and gut microbiota homeostasis in IBD

Abstract Background IL4I1, an L-amino acid oxidase, metabolizes tryptophan into indole-3-pyruvic acid (I3P), activating AHR. While its role in autoimmune diseases and cancer is well documented, its impact on IBD remains unclear. Methods IL4I1 expression in IBD patients was analyzed using single-cell RNA sequencing data and validated by qRT-PCR and immunofluorescence. BMDMs from WT and Il4i1-/- mice were used to assess migration, phagocytosis, polarization, and ROS release. Gut microbiota was analyzed via fecal metagenomics. The role of IL4I1 in vivo was evaluated in DSS-induced colitis and AOM/DSS-induced CAC models. Bone marrow chimeras, clodronate liposome-mediated macrophage depletion, and BMDM transfer experiments were performed to examine IL4I1 function in macrophages. RNA-seq was conducted to analyze transcriptomic changes in Il4i1-/- macrophages. Macrophage senescence was assessed via cell cycle analysis, γ-H2AX levels, and β-galactosidase activity. Ahr and Foxm1 plasmids were used for rescue experiments. The therapeutic potential of recombinant IL4I1 and I3P was evaluated in DSS-induced colitis. Results IL4I1 expression was significantly upregulated in the intestinal mucosa of IBD patients, correlating with disease activity, and was primarily expressed in macrophages. Proinflammatory cytokines and multiple TLR agonists induced IL4I1 expression in macrophages. Il4i1 deficiency impaired phagocytosis, increased migration, and skewed macrophages toward an M1 phenotype. It also disrupted gut microbiota homeostasis, increasing harmful bacteria while reducing beneficial ones. Il4i1 deficiency exacerbated colitis and CAC, leading to heightened macrophage infiltration and impaired maturation. Transcriptomic analysis revealed that Il4i1 deficiency downregulated genes involved in the cell cycle, DNA repair, and DNA damage response pathways, promoting macrophage senescence. Overexpression of Ahr and the key cell cycle transcription factor Foxm1 reduced senescence and restored macrophage function. Treatment with recombinant IL4I1 and I3P activated AHR-mediated cell cycle regulation in macrophages and alleviated colitis. Conclusion IL4I1 plays a crucial role in macrophage senescence and gut microbiota homeostasis, highlighting its potential as a novel therapeutic approach for IBD. Conflict of interest: Li, Ai: No conflict of interest