Gegen Qinlian decoction alleviates ulcerative colitis by remodeling the gut microbiota-bile acid axis and inhibiting NLRP3-mediated pyroptosis.

GQD ameliorated gut dysbiosis in HFD+DSS-induced UC mice by altering key bacterial populations.

• GQD decreased the Firmicutes/Bacteroidetes ratio in the UC mouse model.
• GQD increased the abundance of Akkermansia muciniphila, a beneficial mucosal bacterium.
• GQD reduced Clostridium perfringens, a pathogenic bacterium associated with intestinal inflammation.
• Gut microbiota composition was assessed using 16S rRNA gene sequencing.

GQD remodeled bile acid metabolism by shifting the balance from primary to secondary bile acids.

• GQD decreased primary bile acids including CA, TCA, and CDCA.
• GQD increased secondary bile acids including DCA, 6,7-DKLCA, and 3β-DCA.
• Bile acid profiling was conducted by targeted metabolomics.
• The shift in bile acid profile is consistent with microbiota-mediated biotransformation changes.

Transcriptomic analysis indicated that GQD inhibited NOD-like receptor signaling in the colon.

• Transcriptomics was used to investigate the mechanism of GQD in UC mice.
• NOD-like receptor signaling pathway was identified as a key pathway inhibited by GQD.
• This finding directed further mechanistic investigation toward the NLRP3 inflammasome.

GQD suppressed NLRP3 inflammasome activation and downstream pyroptosis-related protein expression in UC mice.

• GQD reduced expression of ASC, pro-caspase-1, cleaved caspase-1, GSDMD-N, and IL-18.
• Protein expression was assessed by Western blotting and ELISA.
• GSDMD-N is the pore-forming fragment of gasdermin D, a key executioner of pyroptosis.
• Reduction of these markers indicates inhibition of the canonical NLRP3-caspase-1 pyroptosis pathway.

Molecular docking prioritized deoxycholic acid (DCA) as a candidate NLRP3-interacting metabolite.

• Molecular docking was performed to assess bile acid interactions with the NLRP3 protein.
• DCA was identified as the top candidate among the altered bile acids for binding to NLRP3.
• This finding linked the GQD-induced increase in secondary bile acids to direct NLRP3 inhibition.

DCA dose-dependently reduced LPS-induced NLRP3 inflammasome gene expression and partially restored tight junction proteins in Caco-2 cells.

• LPS-stimulated Caco-2 cells were treated with 20 or 50 µM DCA.
• DCA reduced mRNA expression of NLRP3, ASC, GSDMD, IL1B, and IL18 in a dose-dependent manner.
• DCA partially restored ZO-1 and Occludin immunofluorescence signals, indicating improved barrier integrity.
• DCA treatment did not cause appreciable cytotoxicity at the tested concentrations.
• Cell viability was assessed alongside gene expression and immunofluorescence.

Fecal microbiota transplantation (FMT) supported the involvement of gut microbiota in GQD-mediated anti-inflammatory effects.

• FMT experiments were conducted to causally link gut microbiota changes to GQD's therapeutic effects.
• FMT supported that the gut microbiota remodeling induced by GQD contributes to its anti-inflammatory properties.
• This finding provided in vivo evidence that the microbiota-bile acid axis is a functional mediator of GQD's effects.

GQD improved colonic pathology and barrier integrity in HFD+DSS-induced UC mice.

• A UC mouse model was induced by combined high-fat diet (HFD) and dextran sulfate sodium (DSS) exposure.
• Colonic pathology was assessed by H&E staining.
• Barrier integrity was assessed by immunofluorescence.
• GQD constituents were identified by UPLC-MS/MS prior to the experiments.