Xianglian Pill attenuates ulcerative colitis through TLR4/MyD88/NF-κB signaling pathway

Ethnopharmacological relevance: Xianglian Pill (XLP) is a classical Chinese medicine prescription applied for controlling ulcerative colitis (UC). Whereas, the underlying mechanism remains unclear. Aim of the study: The present work was aimed to investigate the mechanism of XLP in dextran sulfate sodium (DSS)-induced UC via the Toll Like Receptor 4 (TLR4)/Myeloid Differentiation factor 88 (MyD88)/Nuclear Factor kappa-B (NF-kappa B) signaling in mice. Materials and methods: The major components of XLP were detected by high-performance liquid chromatographydiode array detection (HPLC-DAD). The ulcerative colitis model was induced by DSS in mice. 5-Amino Salicylic Acid (5-ASA) group and XLP group were intragastrically treated. Disease activity index (DAI) and colon length were monitored and hematoxylin-eosin (HE) staining was conducted. Gasdermin D (GSDMD)-N and TLR4 expressions in colon tissues were visualized by immunofluorescence. TLR4 mRNA was measured by Real Time Quantitative PCR (RT-qPCR). The expressions of NOD-like receptor thermal protein domain associated protein 3 (NLRP3), active-caspase-1, GSDMD-N, TLR4, MYD88, NF-kappa B, p-NF-kappa B, and the ubiquitination of TLR4 in colon tissues were detected by Western blot. Myeloperoxidase (MPO) enzyme activity was examined and serum inflammatory factors Interleukin (IL)-1 beta, IL-6, Tumor Necrosis Factor-alpha (TNF-alpha), and IL-18 were determined by Enzyme-linked Immunosorbent Assay (ELISA). TLR4(-/-) mice were applied for verifying the mechanism of XLP attenuated DSS symptoms. Results: The XLP treatment extended colon length, reduced DAI, and attenuated histopathological alteration in DSS-induced mice. XLP administration suppressed MPO activity and reduced the content of IL-1 beta, IL-6, TNF-alpha and IL-18 in serum. XLP also inhibited the expression levels of GSDMD-N, TLR4, NLRP3, active-caspase-1, MyD88, p-NF-kappa B/NF-kappa B in colon tissues of DSS-induced mice. TLR4(-/-) mice proved that TLR4 was involved in XLP-mediated beneficial effect on DSS-induced ulcerative colitis. Conclusions: XLP might treat ulcerative colitis by regulating the TLR4/MyD88/NF-kappa B signaling pathway.