RNA-seq based elucidation of mechanism underlying the protective effect of Huangshui polysaccharide on intestinal barrier injury in Caco-2 cells

[Display omitted] •A heteropolysaccharide named NLS-2 with a MW of 51.9 kDa was extracted from Huangshui.•NLS-2 has beneficial effects on LPS-damaged intestinal barrier.•NLS-2 reduced the content of inflammatory cytokines and increased the expression of TJ proteins.•RNA-seq and WB results showed that the main signaling pathways involved TRL4/MyD88/NF-κB and MAPK. Polysaccharides from Huangshui (HS) have the function of antioxidant and immunoregulation, but its intestinal barrier protection activity and the underlying mechanism remains unclear. The present work mainly studied the intestinal barrier protection function and its potential molecular mechanism of a heteropolysaccharide named NLS-2 with a molecular weight of 51.9 kDa. NLS-2 reduced intestinal permeability by decreasing the content of inflammatory cytokines and increasing the expression of tight junction (TJ) protein in LPS-damaged Caco-2 cells, thus protecting the intestinal barrier function. RNA-seq results showed that the differentially expressed genes (DEGs) were mainly enriched in the signaling pathways of MAPK, Toll-like receptor, and NF-κB. Subsequent western blot validation experiments proved that NLS-2 could indeed inhibit the two pathways of MAPK and NF-κB by reducing the expression of TRL4, thereby down-regulating the release of downstream pro-inflammatory cytokines and playing the role of intestinal barrier protection. Collectively, NLS-2 has beneficial effects on LPS-damaged intestinal barrier by inhibiting the TRL4/MyD88/NF-κB and MAPK signaling pathways.