Gut microbiota-derived 5-hydroxyindoleacetic acid from Pumpkin Polysaccharides supplementation alleviates colitis through Epac/Rap1 signaling activation

Abstract Aims Polysaccharides from Pumpkin ( Cucurbita moschata Duchesne) (PP) have many pharmacological activities, including anti-oxidant, immune, and intestinal microbiota regulation. These activities have provided some reminders of its potential therapeutic effect on ulcerative colitis (UC), but this has not yet been confirmed. This study explores the potential modulatory roles of gut microbial metabolites (5-hydroxyindole acetic acid, 5-HIAA) after PP treatment in protecting against UC and to elucidate the underlying molecular mechanisms. Results The average molecular weight of PP was 3.10 × 10 5 Da, and PP mainly comprised Mannose, Rhamnose, Galacturonic acid, Galactosamine, Glucose, and Xylose with molar ratios of 1.58:3.51:34.54:1.00:3.25:3.02. PPs (50, 100 mg/kg) could significantly resist dextran sodium sulfate (DSS) induced UC on C57BL/6 mice by improving gut microbiota dysbiosis and intestinal mucosal barrier function. Metabolomic profiling analysis showed that PP supplementation resulted in the enrichment of 5-HIAA, which exhibited individual and synergistic anti-UC activities. The results of the transcriptome and confirmatory experiments suggested that Rap1 pathway activation may participate in the anti-UC effect of PPs. 8-pCPT-2’-O-Me-cAMP (Epac/Rap1 signaling antagonist) showed similar anti-UC activity to PP and 5-HIAA, while the effect of ESI-05 (Epac/Rap1 signaling inhibitor) was opposite. Furthermore, exogenous treatment with 5-HIAA activated the Epac/Rap1 signaling pathway both in vitro and in vivo , which may be related to elevated cAMP levels in the colon. Conclusions This study revealed that PP could be developed to treat UC and other diseases associated with an imbalance in the intestinal microbiota. The gut microbiota-5-HIAA-Epac/Rap1 axis plays an important role in intestinal homeostasis and may provide a novel approach for the prevention of UC by manipulating gut microbial serotonin metabolites.