Prebiotic β (1-4) Galacto-oligosaccharides Strengthen Intestinal Barrier Integrity via Mucus Production and...

Preserving a functional intestinal barrier is crucial for overall host health, as increased permeability can lead to systemic pathologies, including a dysregulated immune system and increased susceptibility to infections. Pure β (1-4) galacto-oligosaccharides (GOS) and Type 2 LacNAc-enriched β (1-4) GOS (humanized GOS, hGOS) modulate the gut microbiome, increasing the abundance of beneficial microorganisms, including Bifidobacterium, Akkermansia, and Lactobacillus. In this study, we report that direct exposure of monolayers of human primary colonic cells to β(1-4) GOS and β(1-4) hGOS enhances barrier integrity by significantly upregulating MUC2 (2-5 fold) and tight junction genes (1.5-8 fold) (p<0.05). RNA sequencing revealed that β(1-4) GOS and β(1-4) hGOS activated the Aryl Hydrocarbon Receptor (AHR) pathway, with GOS specifically inducing CYP1A1 (log2FC = 3.57) and TIPARP, while hGOS enriched pathways related to apical junction integrity. This suggests that AHR activation may enhance gut barrier function, in part, by repressing IL-1β-mediated inflammation. These findings were validated in vivo in young and old C57BL/6 mice, where GOS and hGOS enhanced intestinal permeability by increasing the expression of Muc2 and promoting mucus production, resulting in a thicker mucus layer (GOS: 95% CI 0.054-0.146; hGOS: 95% CI 0.080-0.176). Tight junction integrity genes were also upregulated by the prebiotics in our study (two-way ANOVA, p < 0.05). Our findings suggest that, beyond their modulation of the gut microbiome, GOS and hGOS enhance intestinal barrier function by directly inducing the expression of mucin and tight junction genes, likely through the AHR pathway. These results highlight the potential of prebiotic supplementation in improving gut barrier function and maintaining intestinal health.