Bifidobacterium breve and lacto-N-neotetraose mediate gut microbiota-derived acetate to regulate defecation performance and intestinal barrier function in constipated mice.

Constipation has emerged as an important public health concern, and novel therapeutic approaches, such as those, are attracting increasing attention. However, the effects and mechanisms of Bifidobacterium breve and lacto-N-neotetraose (LNnT) in relieving constipation remain incompletely understood. Moreover, the potential synergistic effects of B. breve and LNnT in alleviating constipation are still unclear. In this study, we used 4-wk-old female BALB/c mice (n = 60), which were randomly assigned to normal control (NC) group, model control (MC) group, LNnT group, B. breve group, and B. breve+LNnT group. We evaluated the effects of B. breve+LNnT on defecation performance and intestinal mucus secretion. We also analyzed gut microbiota composition and metabolic functions. Additionally, an independent cohort of 45 mice was used to assess the effect of gut microbiota and microbiota-derived metabolites on constipation. The results demonstrated that B. breve+LNnT increased the gastrointestinal transit rate and fecal water content while reducing whole-gut transit time. It also elevated serum concentrations of gastrin (GAS) and motilin (MLT) while decreasing those of vasoactive intestinal peptide (VIP) and nitric oxide (NO). Mice receiving B. breve+LNnT showed increased intestinal mucus content, a more organized mucus layer structure, and higher expression of mucus-related genes (Muc2, Agr2, Muc1, Muc4, Muc13). Additionally, B. breve+LNnT increased gut microbiota diversity and enriched potentially beneficial microbiota, including Ruminococcus, Bifidobacterium, Tyzzerella, and Roseburia. Consistently, levels of the acetate and propionate were elevated in the B. breve+LNnT group. Correlation analysis indicated positive associations between these microbiota and acetate. Finally, we explored the role of gut microbiota, acetate, and propionate in constipation. We found that the alleviation of constipation by B. breve+LNnT depended on the presence of gut microbiota and was associated with microbiota-derived acetate.