The Study Of Immune Regulation And Its Anti-tumor Mechanisms About The Crude Polysaccharide Extracted From Enteromorpha On Macrophage Cell Line Raw264.7

Objective: In this study, the crude polysaccharide extracted from Enteromorphaimpacted cell activity changes in vitro RAW264.7murine macrophage cell line,such as Proliferation, secretion of cytokines IL-6, TNF-alpha level,to verify itsregulation of macrophage immune function;Under the intervention from thecrude polysaccharide extracted from Enteromorpha, RAW264.7mousemacrophage cell line expressed mTOR mRAN changes to explore that theEnteromorpha polysaccharide activated RAW264.7murine macrophage celllinepossible molecular mechanism, and thus provide a basis for furtherEnteromorpha polysaccharide regulation of the immune anti-tumor mechanism.Methods:A RAW264.7macrophage cell line in vitro test methods: the recovery of cellscultured, passaged, frozen, and experimental pre-treatment.B detected the proliferation of RAW264.7macrophage cell line inferred by thecrude polysaccharide extracted from Enteromorpha: First we observedRAW264.7macrophage cell line changes by a microscope under the differentdose or time;Then we thiazolyl blue (MTT)colorimetric assay the role ofmacrophage RAW264. proliferative effect change;C By the enzyme-linked immunosorbent assay (ELISA) testing,we obvered thechange of the macrophage cell line RAW264.7secretion of cytokines TNF-α,IL-6secretion under different doses or times of the crude extract ofEnteromorpha polysaccharide intervention;D By Real Time PCR we detected the PTEN\mTOR gene expression relativequantitative changes of macrophages RAW264.7RAW264.7intervented thecrude polysaccharide extracted from Enteromorpha under different doses or timesResults:A under microscope, Increased with the intervention dose,the number ofmacrophages increased, the volume of larger cells more, the differentiation is more obvious; but with time longer, reduce the number of macrophages, the cellsbecame larger, more obvious differentiation.The absorbance differences fromeach dose group was significant, a certain role in the concentration of theintervention concentration increased macrophage proliferation upward trend;absorbance of each dose group decreased with time.B the crude polysaccharide extracted from Enteromorpha improved the secretioneffec t of cytokines IL-6, TNF-alpha on macrophage cell line RAW264.7; in15.625-1000μg/mL, with the Intervention material concentration increases,macrophage cell RAW264.7secreted IL-6more, Showing the strongerdose-dependent manner; For the role of TNF-α secretion, its dose-dependentmannerin the low concentration range (500μg/mL below) with the increase of theconcentration to promote TNF-α secretion, under high concentrations(1000μg/mL for),the RAW264.7secretion of TNF-α to promote weakened.C Expression of mTOR gene: the crude polysaccharide extracted fromEnteromorpha on mTOR gene expression of macrophage RAW264.7apparentstimulative effect, and have a certain dose-dependent.Conclusions:A In the concentration range,the crude polysaccharide extracted fromEnteromorpha promote the macrophage cell line RAW264.7proliferation,Itinitially confirmed its activity to promote the proliferation of macrophages,showing the role of cell proliferation in a dose-dependent manner, non-cytotoxic,The result suggested that the crude polysaccharide extracted from Enteromorphamay play a role of the immunomodulatory effects.B the crude polysaccharide extracted from Enteromorpha improved the secretioneffec t of cytokines IL-6, TNF-alpha on macrophage cell line RAW264.7; Theresult suggested that the crude polysaccharide extracted from Enteromorpha hassignificant immune-enhancing activity, and further that help to promotecell-mediated immune responseC Macrophage relative proliferation rate and cytokines of TNF-A and IL-6secretion in cultured macrophages RAW264.7have t increasing trend with theextension of time, but different intervention time increased slightly different.Possibility as follows:1. Culture medium lack of nutrition caused byconsumption, induced apoptosis of Macrophage RAW264.7and Secretion a greatnumber of inflammatory cytokines.2. Inflammatory cytokines in macrophagesRAW264.7caused autophagy.3. The effect of polysaccharides in Enteromorpha. D he crude polysaccharide extracted from Enteromorpha promote mTOR geneexpression on macrophage RAW264.7, threre are consistent effects with that itpromoted macrophage RAW264.7relative-proliferation and promoted immuneactivation, suggesting that P13K/AKT-PTEN-mTOR signal transductionpathway may be associated with immune regulation on macrophage RAW264.7.