Xiao-Chai-Hu-Tang alleviates irinotecan-induced diarrhea via TGR5-FAK-RhoA/Rock signaling revealed by integrated metabolomics and proteomics

ABSTRACT:

Irinotecan (CPT-11) is widely used for colorectal cancer treatment, with delayed-onset diarrhea as its primary side effect. Xiao-Chai-Hu-Tang (XCHT) has been clinically observed to alleviate diarrhea, but its mechanisms remain unclear. This study aimed to explore the underlying regulatory mechanisms through which XCHT alleviated CPT-11-induced diarrhea. The chemical components of XCHT were detected using UHPLC-Q-Exactive Orbitrap-MS, and 12 active components were quantified via UHPLC-QQQ-MS/MS to ensure quality stability. CPT-11-induced diarrheal mice were established to evaluate the effects of XCHT. Differential metabolites in liver and intestinal tissues among control, CPT-11, and XCHT mice was analyzed by metabolomics, followed by molecular network analysis using IPA. The bile acids were quantified by using the UHPLC-QQQ-MS/MS method. Proteomics of colon tissues identified differentially expressed proteins, with functional enrichment conducted via GO and KEGG. Western blot and flow cytometry were used to validate potential biopathway. XCHT primarily contained flavonoids, carbohydrates and glycosides, and terpenoids. Twelve active components remained stable throughout the experiment. XCHT significantly improved diarrhea symptoms. Metabolomics revealed 133 and 72 differential metabolites in the liver and intestine, respectively, and linked to bile acid metabolism. Further targeted bile acid analysis revealed that CPT-11 reduced hepatic bile acid synthesis and uptake, but increased biliary excretion. Significantly elevated bile acid levels were observed in the gallbladder of CPT-11-treated mice. Furthermore, intestinal bile acid reabsorption was impaired with an 81% decrease in total bile acids and reduced uptake/efflux, resulting in a 56% decline in circulatory bile acids, and 40% increase in colonic accumulation. Proteomics identified 86 differentially expressed proteins in colon tissues, with significant enrichment in ‘Focal adhesion’. Integrated analysis and validation studies suggested the TGR5-FAK-RhoA/ROCK signaling axis as the crucial pathway through which XCHT alleviated diarrhea. These findings provide novel insights into the pharmacological mechanisms of XCHT and suggest potential therapeutic strategies targeting bile acid metabolism for chemotherapy-induced diarrhea.

INSTRUMENT(S): Liquid Chromatography MS - negative - reverse phase, Liquid Chromatography MS - positive - reverse phase

PROVIDER: MTBLS12931 | MetaboLights | 2025-09-03

REPOSITORIES: MetaboLights