Immunostimulatory Effects of Polysaccharides from In Vivo and Vitro and Their Activation Mechanism on RAW246.7 Macrophages.

In this study, (S.p.) polysaccharide (PSP) was obtained by ultrasonic-microwave-assisted extraction (UMAE) and purified by an aqueous two-phase system (ATPS). Two different methods were applied to purified (S.p.) polysaccharide (PSP), respectively, due to PSP as a complex multi-component system. Three polysaccharide fractions (PSP-1, PSP-2, and PSP-3) with different acidic groups were obtained after PSP was fractionated by the diethyl aminoethyl (DEAE)-52 cellulose chromatography, and two polysaccharide fractions (PSP-L and PSP-H) with different molecular weight were obtained by ultrafiltration centrifugation. The chemoprotective effects of PSP in cyclophosphamide (Cy) treated mice were investigated. The results showed that PSP could significantly increase spleen and thymus index, peripheral white blood cells (PWBC), and peripheral blood lymphocytes (PBL). The in vivo immunostimulatory assays demonstrated that PSP could in dose-dependent increase of TNF-α, IL-10, and IFN-γ production in sera. The in vitro immunostimulatory assays showed that PSP and its fractions (PSPs) could evidently enhance the proliferation of splenocytes and RAW 264.7 cells and increase the productions of nitric oxide (NO), tumor necrosis factor-α (TNF-α), and interleukin 6 (IL-6). PSPs could also enhance phagocytic activity of RAW 264.7 cells. The acidic polysaccharide fractions of PSP-2, PSP-3, and PSP-L with small molecular weight had the higher immunostimulatory activity. Signaling pathway research results indicated that PSP-L activated RAW264.7 cells through MAPKs, NF-κB signaling pathways via TLR4 receptor.