Investigating the Digestive and Absorptive Mechanisms of Polysaccharides from Huaishan Based on Intestinal Organoids and Their Effects on Intestinal Cell Proliferation and Differentiation

To contribute to better understanding of the structural composition, bioactivity, and intestinal health effects of Huaishan polysaccharides (HSP), analyzed the changes in the molecular weight and monosaccharide composition of HSP after digestion with simulated gastric juice, and evaluated its impact on intestinal cell proliferation and differentiation using a three-dimensional (3D) organoid model. The study findings showed that Huaishan polysaccharide molecular weights averaged 139.173 kDa prior to source plant digestion, and 1.676 kDa after digestion. Prior to digestion, D-glucose (89.72%) was the dominant monosaccharide, but its levels decline significantly after digestion, while levels of other monosaccharides (e.g., D-mannose, D-galactose) tend to rise. Immunofluorescence analyses confirmed key intestinal cell markers, including Ki67 (proliferation), Chromogranin A (secretory), Mucin2 (goblet cells), Lysozyme (Paneth cells), Villim (absorptive), and Olfm4 (ISCs), were present in the 3D organoids. Chinese yam polysaccharide-treated organoids exhibited significantly enhanced mRNA levels and immunofluorescence signals for Ki67 and Muc2. Huangshan polysaccharides also upregulated respective Lgr5, Olfm4, Wnt3, and Hes1 mRNA levels 1.32, 1.15, 1.1, and 1.12 fold (P<0.05). These data suggested that Huaishan polysaccharides enhance ISC proliferation via Wnt-mediated Lgr5/Olfm4 activation while promoting goblet cell differentiation via Notch inhibition. These data revealed the structure of Huaishan polysaccharides at stages during digestion, and its intestinal proliferation and differentiation promoting effects, laying a foundation for Huaishan polysaccharide application in functional foods.