Study On Immunomodulatory Activity Of Cyclocarya Paliurus Polysaccharide

Polysaccharides are one of the most important active ingredients in Cyclocarya paliurus.In order to investigate the structure of Cyclocarya paliurus polysaccharide,the immunoregulatory effect of Cyclocarya paliurus polysaccharide on RAW264.7 macrophage and the immune regulation of Cyclocarya paliurus polysaccharide on immunosuppressed mice.DEAE-cellulose column was used to purify the crude polysaccharide of Cyclocarya paliurus,0.3% Na Cl was eluted to obtain CPP-3.Fourier transform infrared spectroscopy(FT-IR),high performance gel permeation chromatography(HPGPC),gas chromatography(GC)and scanning electron microscope(SEM)were used to analyze of the CPP-3 structure.MTT colorimetric assay was used to detect cell proliferation.Neutral red phagocytosis assay was used to detect macrophage phagocytosis.Griess reagent was used to detect NO content in cell supernatant.Enzyme-linked immunosorbent assay(ELISA)was used to detect cytokines(IL-1β,TNF-α)and proinflammatory mediators(PGE2)content in cell supernatant.RT-PCR was used to detect the expression of IL-12,IL-18,IFN-γ,TNF-α and i NOS genes.After a week of adaptive feeding of SPF kunming mice,60 kunming mice were randomly divided into normal control group,model control group,low-dose,medium-dose,high-dose CPP-3 groups and positive control group.They were given saline 0.1m L/10 g,subcutaneous injection of cyclophosphamide 30 mg/kg on 1,3,5,7,9,11,13,15,17,19 days,levomisole aqueous solution 300 mg/(kg/d),CPP-3 aqueous solution 150,300,600 mg/(kg/d),respectively.Feeding for 20 days,at the end of the experiment,total cholesterol(TC),triglyceride(TG),high-density lipoprotein cholesterol(HDL-C)and low-density lipoprotein cholesterol(LDL-C)levels were determined in serum of immunosuppressed mice feeded with CPP-3.The activities of superoxide dismutase(SOD),glutathione peroxidase(GSH-PX),catalase(CAT),malondialdehyde(MDA)and total antioxidant capacity(T-AOC)were determined in the spleen of immunosuppressed mice feeded with CPP-3.The content of red blood cells(RBC),white blood cells(WBC),hemoglobin(Hb)and platelet(PLT)was measured in blood of immunosuppressed mice by using a fully automated hematology analyzer.The main test results are as follows:(1)CPP-3 was comprised of two polysaccharides with average molecular weight(Mw)of 5.6×104 Da and 4.94×103 Da.CPP-3 contains six monosaccharides,of which are rhamnose(Rha),arabinose(Ara),xylose(Xyl),mannose(Man),glucose(Glu),galactose(Gal),the molar ratio of six monosaccharides is 0.060: 0.109: 0.053: 0.128: 0.293: 0.357.(2)CPP-3 increased the amount of NO released from mouse macrophage RAW264.7 and significantly increased the levels of TNF-α,IL-1β and PGE2(P < 0.01).CPP-3 suppressed LPS-stimulated RAW264.7macrophage to release NO,TNF-α,IL-1β and PGE2(P < 0.01).CPP-3 and LPS accounted for synergistic effect on the release of NO and TNF-α,CPP-3 and LPS accounted for antagonistic effect on the release of IL-1β and PGE2.(3)CPP-3 reduced TC,TG and LDL-C levels in serum of immunosuppressed mice induced by cyclophosphamide,and increased HDL-C level and LDH activity(P < 0.01 or P <0.05).(4)CPP-3 increased the activities of superoxide dismutase(SOD),glutathione peroxidase(GSH-PX)and catalase(CAT)in serum of immunosuppressed mice induced by cyclophosphamide,and decreased the content of malondialdehyde(MDA),enhanced total antioxidant capacity of T-AOC(P < 0.01 or P <0.05).(5)CPP-3 increased the content of red blood cells(RBC),white blood cells(WBC),hemoglobin(Hb)and platelets(PLT)in blood of immunosuppressed mice induced by cyclophosphamide.