Compound Kushen Injection Alleviates Experimental Colitis in Mice by Regulating Macrophage Polarization through TLR4/NF-κB Signaling.

OBJECTIVE: To explore the therapeutic effect and mechanism of Compound Kushen Injection (CKI) in dextran sulfate sodium (DSS)-induced ulcerative colitis (UC). METHODS: Network pharmacology was used to predict the potential targets and pathways for CKI's therapeutic effects on UC. Thirty-five male C57BL/6 mice were randomized into 5 groups, including control, DSS model, 5-aminosalicylic acid (5-ASA), low- and high-dose CKI groups (n=7 per group). A 7-d protocol of ad libitum administration of 3% (w/v) DSS in drinking water was implemented to induce experimental colitis in mice of all groups except the control. Mice received daily intragastric 5-ASA (50 mg/kg), low-dose (2 mL/kg) or high-dose (4 mL/kg) CKI accordingly. In vivo therapeutic effects of CKI were evaluated via histology, ELISA and immunohistochemistry. In vitro, LPS-stimulated RAW264.7 inflammatory cells were treated with CKI for 24 h to detect cytokine changes. Immunofluorescence, flow cytometry and Western blot were applied to analyze macrophage polarization phenotypes in mouse peritoneal macrophages and RAW264.7 cells. RESULTS: Network pharmacology analysis indicated that CKI's therapeutic effects on UC involved macrophage polarization regulation. CKI substantially alleviated DSS-induced colitis symptoms, improved intestinal barrier function, and reduced both pro-inflammatory cytokine secretion and oxidative stress in colonic tissues (P<0.05 or P<0.01). Upon DSS or LPS stimulation, CKI treatment markedly decreased M1 macrophage marker expression and increased M2 markers (P<0.05 or P<0.01). Mechanistic studies further revealed that CKI modulated macrophage polarization via Toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) signaling pathway (P<0.05 or P<0.01). CONCLUSIONS: CKI alleviates experimental colitis in mice by modulating macrophage polarization, reducing inflammatory responses, and repairing the intestinal barrier; this effect may closely associated with inhibition of TLR4/NF-κB signaling pathway.