Structural properties of glucan from Russula griseocarnosa and its immunomodulatory activities mediated via T cell differentiation

The polysaccharide, RGP2, was isolated from Russula griseocarnosa and its immunostimulatory effects were confirmed in cyclophosphamide (CTX)-induced immunosuppressed mice. Following purification via chromatography, structural analysis revealed that RGP2 had a molecular weight of 11.82 kDa and consisted of glucose (Glc), galactose (Gal), mannose, glucuronic acid and glucosamine. Bond structure analysis and nuclear magnetic resonance characterization confirmed that the main chain of RGP2 was formed by →6)-β-D-Glcp-(1→, →3)-β-D-Glcp-(1→ and →6)-α-D-Galp-(1→, which was substituted at O-3 of →6)-β-D-Glcp-(1→ by β-D-Glcp-(1→. RGP2 was found to ameliorate pathological damage in the spleen and enhance immune cell activity in immunosuppressed mice. Based on combined multiomics analysis, RGP2 altered the abundance of immune-related microbiota (such as Lactobacillus, Faecalibacterium, and Bacteroides) in the gut and metabolites (uridine, leucine, and tryptophan) in the serum. Compared with immunosuppressed mice, RGP2 also restored the function of antigen-presenting cells, promoted the polarization of macrophages into the M1 phenotype, positively affected the differentiation of helper T cells, and inhibited regulatory T cell differentiation through the protein kinase B (AKT)/mechanistic target of rapamycin (mTOR) pathway, ultimately exerting an immune boosting function. Overall, our findings highlight therapeutic strategies to alleviate CTX-induced immunosuppression in a clinical setting.