Mechanisms of huajiao in alleviating Shigella flexneri-induced colitis via NF-κB/TNF-α pathway

Huajiao, a medicinal and edible spice, is widely recognized for its potential in alleviating intestinal inflammation. This study investigated the antibacterial activity of five plant parts from three huajiao cultivars against S. flexneri, identifying the pericarp of the “Dahongpao” variety as the most potent. Untargeted metabolomic profiling combined with chemometric analysis revealed significant differences in metabolite composition among pericarp extracts prepared with solvents of varying polarity (P < 0.05). Partial least squares (PLS) modeling highlighted several compounds—nepetin 7-glucoside, quercetin 3-O-malonylglucoside, quercitrin, guaiaverin, and hyperoside—as likely contributors to the observed antibacterial effects. Network pharmacology and molecular docking further identified TNF, NF-κB1, BCL2, CASP3, and EGFR as potential molecular targets of the active compounds. Gene Ontology (GO) and KEGG enrichment analyses indicated that the NF-κB/TNF-α signaling pathway plays a central role in mediating the anti-inflammatory response. In a murine model of S. flexneri-induced enteritis, treatment with huajiao extracts alleviated colonic inflammatory responses, increased goblet cell numbers, and significantly reduced colonic mRNA and protein levels of TNF-α, IL-1β, and IL-6 (P < 0.01). Western blot analysis confirmed decreased phosphorylation of IκBα and NF-κB p65, indicating inhibition of NF-κB activation and alleviation of intestinal inflammation. Collectively, these results indicate that huajiao is a functional food source with potential for alleviating colitis.