Polysaccharide from Atractylodes macrocephala Koidz. protects against avian pathogenic Escherichia coli-induced intestinal barrier Dysfunction via Suppressing PI3K/Akt-Mediated Claudin-2 upregulation

Avian pathogenic Escherichia coli (APEC) is a leading cause of intestinal barrier dysfunction and diarrhea in chicks, resulting in substantial economic losses to the poultry industry. This study investigated the protective effects and underlying mechanism of polysaccharide from Atractylodes macrocephala Koidz. (PAMK) against APEC-induced intestinal injury using an integrated in vivo (chick) and in vitro (IPEC-J2 cell) approach. In vivo, PAMK intervention significantly reduced necropsy scores, alleviated growth retardation, and improved villus height-to-crypt depth ratios as well as brush border height. PAMK also attenuated the APEC-induced increase in goblet cell counts and upregulated MUC-2 mRNA expression, indicating enhanced chemical barrier function. Furthermore, PAMK suppressed the mRNA expression of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) and the pore-forming tight junction protein Claudin-2 in the jejunum. In IPEC-J2 cells, PAMK pretreatment inhibited APEC-induced upregulation of TNF-α and Claudin-2 at both mRNA and protein levels, effects that were phenocopied by the PI3K inhibitor LY294002. Mechanistically, PAMK suppressed APEC-induced phosphorylation of Akt and its downstream target GSK-3β, thereby blocking the PI3K/Akt/GSK-3β signaling cascade. Immunohistochemical analysis confirmed that APEC infection increased Claudin-2 protein abundance along the villus apical membrane. Collectively, these findings demonstrate that PAMK alleviates APEC-induced intestinal barrier dysfunction by inhibiting the PI3K/Akt/GSK-3β pathway to downregulate Claudin-2 expression, suggesting its potential as a dietary supplement to improve gut health in poultry production.