Platycodon grandiflorum polysaccharide identified via activity-guided screening: structure, immunostimulatory activity, and glycolysis-mediated mechanism.

Alkaline-extracted Platycodon grandiflorum polysaccharide (APGP) exhibits strong immunostimulatory activity, however, the specific structural features and properties that confer the greatest development potential, as well as the underlying mechanism, remain unclear. In this study, the APGP4 fraction exhibiting significant immunostimulatory activity in mouse monocyte-macrophage cells (RAW264.7 cells) and phorbol myristate acetate (PMA)-induced human acute monocytic leukemia cells (THP-1 cells) was identified. This activity was further confirmed through in vivo experiments in immunosuppressed mice. Subsequently, a new polysaccharide, APGP4-3, was isolated and exhibited considerable immunostimulatory activity. APGP4-3 (molecular weight: 11.7 kDa) comprises 1,4-α-D-GalpA, 1,3,4-α-D-Rhap in the backbone, with T-α-L-Araf, 1,3,5-α-L-Araf, 1,3-α-D-Galp, 1,3,4-α-D-Galp and 1,4,6-α-D-Glcp bound to the 3 position of 1,3,4-α-D-Rhap, as well as a few T-α-D-GalpA and Δ4,5-GalpA. Mechanistic studies revealed that the APGP4-3 treatment enhances the proportion of CD11c+ cells (M1-type macrophages) and reduces the proportion of CD206+ cells (M2-type macrophages). Furthermore, APGP4-3 significantly increases the end product (lactic acid) and key enzyme (pyruvate kinase) involved in glycolytic metabolism, while reducing the key enzyme (acetyl coenzyme A) associated with oxidative phosphorylation in macrophages. These findings suggest that APGP4-3 may induce M1 polarization of macrophages by promoting glycolysis and inhibiting oxidative phosphorylation, thereby exerting an immune-promoting effect. This study advances our understanding of the structural features and potential mechanisms underlying the immunostimulatory activity of Platycodon grandiflorum polysaccharide (PGP), and provides new insights into the development of novel immunopotentiators.