Xianglian pill alleviates ulcerative colitis by inhibiting M1 macrophage polarization via modulation of energy metabolite itaconate

Background: Xianglian pill (XLP) is a traditional Chinese medicine (TCM) that is widely used to treat ulcerative colitis (UC). However, its mechanism of action in UC is unclear.

Purpose: This study aimed to investigate the mechanism of action of XLP in treating UC and role of M1 macrophage polarization in this process.

Study design: In vivo experiments were performed using UC mice while in vitro experiments were conducted using RAW264.7 cells.

Methods: Mice were administered 3 % dextran sulfate to induce UC model and then treated with XLP. Changes in histopathology and pro-inflammatory cytokines were evaluated. The levels of M1 macrophages in mesenteric lymph nodes were detected by flow cytometry. Colon metabolite levels were analyzed using an energy metabolomic assay. To assess itaconate's impact, both in vivo (mice) and in vitro (RAW264.7 cells) models were employed. Immunofluorescence staining was used to measure the expression levels of TNF-α, IL-6, and iNOS, while qRT-PCR was utilized to quantify the mRNA levels of TET2, STAT1, and Nfkbiz.

Results: XLP alleviated ulcerative damage and reduced TNF-α and IL-6 levels in colon, and also downregulated the levels of M1 macrophages and modulated the state of energy metabolism. Specifically, XLP significantly increased ITA level in colonic tissue and this increase was significantly associated with decreased levels of M1 macrophages and alleviation of UC following XLP treatment. Moreover, ITA directly suppressed the polarization of macrophage from M0 to M1 phenotype, accompanied by the decrease of TNF-α, IL-6, and iNOS levels. Further, ITA decreased inflammatory responses in M1 macrophage by inhibiting the TET2/STAT1 and TET2/NF-κB signaling pathways.

Conclusion: XLP can treat UC by suppressing M1 macrophage polarization via increasing the level of energy metabolite ITA.