Paeoniae Radix Alba polysaccharides: Structural characterization, macrophage immunomodulation, and potent vaccine...

PRAPs with M_w of 14.391 kDa was purified from Paeoniae Radix Alba. Structural analysis identified it as a homogeneous α-glucan with a → 4)-α-D-Glcp-(1 → backbone, partially O-3 methylated and substituted at C-6 with α-D-Glcp-(1 → side chains. In vitro, PRAPs potently activated macrophages, characterized by the enhanced pinocytic and phagocytic activities, upregulated expressions of surface molecules and increased productions of pro-inflammatory mediators and chemokines. RNA-seq, bioinformatics analysis and mechanistic investigation demonstrated that PRAPs activated macrophages primarily via the TLR2/TLR4-mediated MyD88-dependent NF-κB signaling pathway, and TNF was identified as a hub gene. The potential of PRAPs as a vaccine adjuvant was evaluated in vivo. PRAPs was revealed to significantly enhance OVA-specific IgG, IgG1, IgG2a, and IgG2b levels. Notably, a synergistic adjuvant effect was observed when PRAPs was combined with Alum, resulting in superior antibody level and robust Th1 immune response. Furthermore, PRAPs promoted the generation of central memory T cells and memory B cells, and fostered a potent germinal center reaction, as evidenced by the increased frequencies of T follicular helper cells, germinal center B cells and CD21/CD35-positive cells. These findings collectively demonstrated that PRAPs was a structurally and mechanistically well-defined vaccine adjuvant capable of eliciting potent, balanced Th1/Th2, and durable immune responses.