Dabupi decoction mitigates fluorouracil-induced intestinal mucositis via farnesoid X receptor activation and dihydropyrimidine dehydrogenase upregulation

Ethnopharmacological relevance: 5-Fluorouracil (5-FU) is a widely utilized antimetabolite in colorectal Cancer chemotherapy, primarily exerting its cytotoxic effects by irreversibly inhibiting Thymidylate Synthase (TS). This inhibition leads to a reduction in deoxythymidine monophosphate (dTMP), which disrupts DNA synthesis and repair. A major challenge in 5-FU treatment is the dose-limiting toxicity of chemotherapeutic-induced intestinal mucositis. Da-Bu-Pi Decoction (DBPD), a well-established formula in traditional Chinese medicine for treating spleen-stomach deficiency, is often used to enhance spleen qi and balance the middle jiao. While growing clinical evidence points to the therapeutic benefits of DBPD in alleviating 5-FU-induced mucositis, the underlying molecular mechanisms remain largely undefined.

Aim of the study: To elucidate the molecular mechanisms underlying chemotherapy-induced intestinal mucositis (CIM) induced by 5-FU, with a focus on the disruption of bile acid metabolism and the downregulation of the key detoxifying enzyme DPYD expression.

Materials and methods: DBPD was administered orally to C57BL/6 mice with 5-FU-induced intestinal mucositis over a seven-day period. The evaluation of intestinal damage included assessments of diarrhea, morphology, intestinal barrier function and inflammatory factors, alongside techniques such as immunofluorescence, immunohistochemistry, transmission electron microscopy and Western blot. Transcriptome analysis of mouse ileal tissue was applied to reveal differentially expressed genes (DEGs) in different treated mice, and bile acid metabolism-related genes (UDP-glucuronosyltransferase 1A1 (UGT1A1), UGT1A9, Farnesoid x receptor (FXR), TGR5) and DPYD, a key detoxification enzyme for 5-FU, were confirmed by qRT-PCR. Additionally, changes of DCA and LCA were measured using ELISA. Bioinformatics helped delineate the association of these genes with pan-cancer versus normal tissues. Meanwhile, 5-FU-induced intestinal epithelial cells (HIEC) were treated with serum containing DBPD. It was further explored how DBPD modulates the UGT1A1/TGR5/FXR signaling pathway and enhances DPYD activity to reduce Apoptosis and intestinal barrier damage in 5-FU induced HIEC. AlphaFold3 and further bioinformatics analysis predicted the binding interactions and expression correlations among UGT1A1, UGT1A9 and FXR.

Results: DBPD is protective by reducing inflammation and intestinal barrier dysfunction in 5-FU-induced intestinal mucositis. Transcriptome analysis and in vivo validation highlighted the crucial function of bile acid metabolism-related pathways and DPYD in 5-FU-induced CIM. 5-FU increased the levels of deoxycholic acid (DCA) and lithocholic acid (LCA) in the mouse ileum. DBPD activated the UGT1A1/TGR5/FXR pathway and up-regulated DPYD to suppress the pathological accumulation of these specific cytotoxic bile acids within the local intestinal microenvironment and ameliorate 5-FU-induced CIM. It was indicated by the analysis of bioinformatics that low levels of UGT1A1, UGT1A9, and FXR exhibited a connection with poor prognosis within colorectal Cancer. In vitro studies confirmed that DBPD significantly improved the UGT1A1/TGR5/FXR pathway and increased the expression of DPYD in 5-FU treated HIEC, thereby improving intestinal barrier integrity and alleviating Apoptosis. Further validation using the FXR inhibitor (Gly-β-MCA) showed that DBPD ameliorated Gly-β-MCA induced HIEC Apoptosis and barrier attenuation. Finally, AlphaFold3 and bioinformatics predictions suggested potential binding interactions between UGT1A1, UGT1A9 and FXR with positively correlated expression.