Zanthoxylum nitidum-derived active alkaloid nitidine chloride mitigates ulcerative colitis by remodeling gut...

Ulcerative colitis (UC) is a recurrent inflammatory bowel disorder with rising prevalence and limited efficacy of current treatments. Nitidine chloride (NC), a characteristic bioactive constituent of Zanthoxylum nitidum (Liang-Mian-Zhen), possesses diverse pharmacological properties, including anti-inflammatory, antitumor, and antioxidant activities. Nevertheless, its role in UC and the associated regulatory mechanisms remain incompletely defined.

This study investigated whether NC protects against experimental colitis and clarified its effects on epithelial barrier integrity, gut microbial composition, and SIRT1-associated regulation of endoplasmic reticulum (ER) stress and TLR4/NF-κB signaling.

Experimental colitis was established using 3% dextran sulfate sodium (DSS)-treated mice, while LPS-stimulated Caco-2 cells were used to evaluate epithelial barrier dysfunction in vitro. Disease severity, barrier integrity, and gut microbial alterations were assessed by histological, permeability-related, and 16S rRNA sequencing analyses. Transepithelial electrical resistance (TEER) and FITC-dextran assays were applied to evaluate epithelial permeability. ER stress, TLR4/NF-κB signaling, inflammatory mediators, and SIRT1-related mechanisms were examined by Western blot, qRT-PCR, ELISA, molecular docking, cellular thermal shift assay (CETSA), pharmacological inhibition with EX-527, and SIRT1 overexpression.

NC markedly alleviated DSS-induced colitis, as evidenced by reduced clinical symptoms, attenuated histological injury, and improved intestinal barrier function. NC restored ZO-1, occludin, and mucin 2 (MUC2) expression, improved Alcian blue-positive goblet cell area, and reduced serum DAO and d-lactate levels. Microbiota analysis showed that NC partly corrected DSS-induced gut microbial dysbiosis and reshaped predicted microbial functions. In LPS-stimulated Caco-2 cells, NC improved barrier integrity by increasing TEER, reducing FITC-dextran permeability, and restoring tight junction protein expression. Mechanistically, NC increased SIRT1 expression and suppressed the PERK-eIF2α-ATF4-CHOP and TLR4/NF-κB pathways both in vivo and in vitro. CETSA further supported target engagement between NC and SIRT1 in Caco-2 cells. Moreover, EX-527 largely abolished the protective effects of NC, whereas SIRT1 overexpression attenuated LPS-induced ER stress and inflammatory signaling.

NC exerted significant protective effects against experimental UC by alleviating inflammation, preserving epithelial and mucus barrier integrity, and partially restoring gut microbial homeostasis. These effects were associated, at least in part, with SIRT1-mediated suppression of ER stress and TLR4/NF-κB signaling. NC may represent a promising natural agent for UC therapy.