Demonstration of the effective intestinal immunity activity of a high branched rhamnogalacturonan-I type pectin polysaccharide from wolfberry via exploration its interaction with mechanical barrier.

Rhamnogalacturonan-I type pectin polysaccharide from wolfberry has immunity activity, but its intestinal immunity activity and structure-activity relationship in the small intestine was still unclear. This study comparatively investigated the intestinal immune activity of wolfberry-derived high and low branched Rhamnogalacturonan-I type pectin polysaccharides (H-LBP and L-LBP) and explored the interaction mechanism with mechanical barrier. In the normal mechanical barrier model, both H-LBP and L-LBP could cross mechanical barrier with transport rates of 23.2 % and 25 %, thereby directly enhancing macrophage viability and phagocytic ability after crossing the mechanical barrier. The transport mechanism of H-LBP in mechanical barrier included the clathrin- and caveolin-mediated pathways. In the damaged mechanical barrier model, H-LBP could significantly enhance mechanical barrier integrity, reduce the production of neurotransmitter (NO) and inflammatory cytokine (TNF-α), thereby exerting indirectly intestinal immune activity. Transcriptome analysis showed that the interaction mechanism between H-LBP and damaged mechanical barrier mainly involved signaling pathway regulating cell growth and survival (PI3K-AKT). Western blot experiment and molecular docking simulation confirmed that H-LBP could reduce the expression of p-PI3K, p-AKT and cleaved Caspase3. H-LBP had stronger directly and indirectly intestinal immune activity than L-LBP. These findings were useful for the application of H-LBP in improving intestinal immunity oral formulations.