IKK/NF-κB Inactivation by Salidroside via Targeting TNF-α for the Treatment of LPS-Induced Colitis

Tumor necrosis factor-alpha (TNF-α) serves as a central mediator of inflammation and represents key therapeutic target in inflammatory bowel disease (IBD). This study investigates the protective effects of salidroside (Sal) against inflammation and explores its underlying molecular mechanisms.

We employed network pharmacology to identify potential targets of Sal. The anti-inflammatory effects of Sal were evaluated in LPS-Induced cellular models using NCM460 colonic epithelial cells and RAW264.7 macrophages, as well as in a murine model of acute colonic inflammation. Direct target engagement was confirmed through cellular thermal shift assay (CETSA) and co-immunoprecipitation (Co-IP). The mechanism was further elucidated via site-directed mutagenesis and analysis of the IKK/NF-κB signaling pathway.

Network pharmacology predicted TNF-α as a key target. Sal significantly attenuated LPS-Induced inflammation in vitro and ameliorated colitis symptoms in vivo. Notably, CETSA and Co-IP assays confirmed direct interaction between Sal and TNF-α. Mutagenesis studies identified Arg179, Lys188, and Tyr191 as critical residues for this binding. Mechanistically, Sal inhibited TNF-α-mediated activation of the IKK/NF-κB pathway and the subsequent production of pro-inflammatory cytokines.

Our findings demonstrate that Sal alleviates inflammation by directly binding to TNF-α and suppressing the downstream NF-κB signaling cascade, thereby positioning it as a promising therapeutic candidate for TNF-α-driven inflammatory diseases.