Galacto-oligosaccharides regulate intestinal mucosal sialylation to counteract antibiotic-induced mucin dysbiosis.

Abstract

Intestinal mucin offers a physical barrier to maintain host-commensal homeostasis. Glycosylation is essential for the appropriate functioning of mucin. Galacto-oligosaccharides (GOS) have been used as a prebiotic with proven intestinal benefits, while their regulatory mechanism on mucin remains unclear. This study employed an antibiotic-treated rat model to mimic gut dysbiosis and attempted to restore gut dysbiosis using GOS. The gut microbiome and intestinal mucus O-glycosylations (O-glycans) in the small intestine were profiled by high-throughput sequencing and glycomics. The sialic acid phenotype at the end of O-glycans was further validated with lectin staining. Expressions of key enzymes in sialic acid metabolism and epithelial morphology were determined as well. Antibiotics significantly increased the relative abundance of Escherichia/Shigella and decreased the relative abundance of Lactobacillus. This was accompanied by decreased microbial sialidase activity and increased sialic acid in the digesta, as well as an increase in epithelial sialidase activity. Analysis of key sialylation enzymes showed the upregulation of α 2,6 sialylation (e.g. ST6GALNACs) and downregulation of α 2,3 sialylation (e.g. ST3GALs) after antibiotic treatment. The glycomics results revealed that antibiotics increased core 4 and α 2,6 sialylated O-glycans and decreased core 1, core 3 and α 2,3 sialylated O-glycans in the intestinal mucus of rats, which was further confirmed by lectin staining. Intestinal histology results demonstrated that antibiotic treatment led to the dysbiosis of intestinal mucus homeostasis. To further test the role of microbiota in regulating intestinal mucus sialylation, we supplemented GOS with antibiotics. The results showed that GOS reversed the effects of antibiotics on the gut microbiota and intestinal mucus O-glycans (especially sialylated O-glycans), characterized by an increase of Lactobacillus and α 2,3 sialylated O-glycans and a decrease of Escherichia/Shigella and α 2,6 sialylated O-glycans. What's more, GOS reduced the stimulation of the intestinal mucosa by lipopolysaccharide (LPS) by increasing α 2,3 sialylated intestinal alkaline phosphatase (IAP) to enhance IAP activity, thereby restoring intestinal mucus homeostasis. Overall, GOS counteracts antibiotic-induced mucin deficiency by remedying the gut ecology and changing the mucin sialylation pattern, as reflected by the increase of α 2,3 sialylated O-glycans and the decrease of α 2,6 sialylated O-glycans.