Immunomodulatary Effects Of Three Marine Polysaccharides On RAW264.7 Cells

Both Laminaria japonica and Stichopus japonicus have great edible and medicinal values.Polysaccharide is the main active ingredient of Laminaria japonica and Stichopus japonicus.The present study investigated the immunomodulatory effects of F1 and F2,which were isolated from Laminaria japonica.In addition,HS extracted from Stichopus japonicus was also researched.RAW264.7 was adopted as macrophage.The main results were as followed:1.The influence of F1,F2 and HS on active substances in macrophagesMTT method was adopted to determine the macrophage proliferation.The result showed that the proliferation effect of F1,F2 and HS on macrophages was:HS(31.9%)>F2(23.1%)>F1(20.7%).The fluorescence degree of each macrophage was studied by acridine orange fluorescent staining method.The results showed that the fluorescence degree improvement effect of F1,F2 and HS(at the concentration of 166.7 μg/ml)on macrophage was:HS(1.89)>F1(1.85)>F2(1.60).The lysozyme activity was researched by Mouse LZM ELISA Kit,the result showed that the lysozyme activity enhancement effect of F1,F2 and HS on macrophages was:HS(272.1 U/L)>F1(267.7 U/L)>F2(235 U/L).Neural red method was supplied to determine phagocytic indexes,it showed that the phagocytic indexes enhancement effect of F1,F2 and HS on macrophage was:HS(1.89)>F1(1.85)>F2(1.60).2.The influence of F1 F2 and HS on cytokines production of macrophagesDetermined interleukin-1β(IL-1β)production by the Enzyme-Linked Immunosorbent Method(ELISA),the results showing that the enhancementeffect of IL-1βproduction by F1,F2 and HS on macrophage were:HS(65.5 U/L)>F1(59.1 U/L)>F2(56.4 U/L);the enhancementeffect of Tumor necrosis factor-α(TNF-α)production by F1，F2 and HS on macrophage was:HS(351.4 U/L)>F1(334.5 U/L)>F2(320.5 U/L);the enhancementeffect of interleukin-12(IL-12)production by F1，F2 and HS on macrophage was:HS(21.6 U/L)>F1(20.5U/L)>F2(18.6 U/L);the enhancementeffect of interleukin-6(IL-6)production by F1,F2 and HS on macrophages was:HS(64.7 U/L)>F1(63.6U/L)>F2(60.3 U/L);The enhancementeffect of inducible nitric oxide synthase(iNOS)production by F1 F2 and HS on macrophage was:HS(4.0 U/L)>F1(3.8 U/L)>F2(3.4 U/L).3.The influence of F1,F2,HS on cytokines mRNA expression of macrophagesThe expression of TNF-amRNA was studied by RT-PCR methods.The results showed that the enhancement effects of TNF-a mRNA expression was:HS(1.82)>F1(1.75)>F2(1.31);the enhancement effect of IL-6 mRNA expression was:HS(2.1)>F1(1.8)>F2(1.7),which was consistent with the results of ELISA.Althoug F2 showed the weakest up-regulation immunomodulatory activity to RAW264.7 cells at the concentration of 166.7 μg/ml compared to F1 and HS,it reached far higher proliferation rate of RAW264.7 cells than F1 and HS at the concentration of 500 μg/ml.Although blood concentration could not reach 500 μg/ml,the above results provided the possibility of getting polysaccharides with smaller molecular weight and higher activity to promote the proliferation of RAW264.7 cells.And make it possible that by using some kinds of fucoidans from Laminaria japonica at low cost together,the immunomodulatory activity to RAW264.7 cells could be better than polysaccharides derived from Stichopus japonicus at high cost.