Structural characterization and immunomodulatory activity of a novel polysaccharide from wild Chinese cordyceps (Ophiocordyceps sinensis).

Wild Chinese Cordyceps polysaccharides (CPs) exert immunomodulatory effects that are closely associated with their structural characteristics. However, research on its structure-function relationship remains limited. This work aims to compare the immunological efficacy of CPs with different molecular weights (MW), together with characterizing their structures and exploring the potential mechanisms of the isolated components. Four CPs with varying MWs were extracted by a stepwise ethanol precipitation technique and subsequently purified through column chromatography. They were named CP-20-1a, CP-40-2a, CP-60-2, and CP-80-2 in descending order of MW. The immunomodulatory effects of these CPs were evaluated using a cyclophosphamide (CTX)-induced immunosuppression mouse model. Among them, CP-20-1a, possessing the largest MW, had the most potent immunomodulatory activity, primarily modulating CD4+ T cells. Structural analysis revealed that CP-20-1a is a branched α-D-glucan with a → 4)-α-D-Glc-(1→/→4,6)-α-D-Glc-(1 → backbone and O-6-linked α-D-Glc-(1 → or α-D-Glc-(1 → 3)-α-D-Glc-(1 → branches. In vitro investigations based on fluorescent labeling demonstrated that CP-20-1a predominantly accumulated on the cell membrane of CD4+ T cells. Transcriptomic analysis and Western blot results demonstrated that CP-20-1a reinstated T lymphocyte proliferation through the Frizzled/AKT/β-catenin pathway along with mTOR signaling, thereby fulfilling its immunomodulatory role. In conclusion, these findings elucidated the chemical foundation underlying the immunomodulatory activity of CPs and provide a foundation for developing targeted immune-regulatory agents in subsequent studies.