Study On Lipid-lowering Effect And Mechanism Of Agaricus Blazei Murill Polysaccharides

Objective: Investigate the effects of Agaricus Blazei Murill polysaccharides on blood lipid metabolism in hyperlipidemia rats,and the role of oleic acid in the cell model of lipid accumulation in HepG2 cells to clarify the lipid-lowering effect and mechanism of Agaricus Blazei Murill polysaccharides in vivo and in vitro.Methods: Water-soluble alcohol precipitation and chromatography were used to extract and purify Agaricus blazei Murill acid polysaccharides with uniform molecular weight.Establish a hyperlipidemia rat model through a high-fat diet,the normal control group?NG?,the model group?MG?,Positive drug group?PD,4.8mg/kg/d?,Agaricus blazei Murill polysaccharide group?WABM-A,640mg/kg/d?,of which the NG was fed with ordinary diet and the rest three groups were fed with high-fat diet for 8 weeks.During the establishment of the model,rats in the PD and WABM-A groups were treated with lovastatin and Agaricus blazei Murill acid polysaccharides.On the 9th week of the experiment,the rats were anesthetized with urethane and blood was taken from the abdominal aorta to collect spleen,liver,and pancreas tissue.The Visceral indexes were measured;serum lipids and antioxidant-related indicators were measured.HE staining was used to observe the liver pathological manifestations of rats.Westernblot was used to measure PPAR?,LXR?,ABCA1 and ABCG1 of proteins expression.Collect rectal feces to extract intestinal micro-ecological flora DNA,high-throughput sequencing of the 16 S rDNA gene V3+V4 region of intestinal microorganisms using high-throughput sequencing technology to obtain intestinal flora big data,OTU,Alpha and Beta diversity analysis,and biological classification analysis of species differences between groups using academic comparisons.Oleic acid was used to induce fatty degeneration of HepG2 cells,and a lipid droplet accumulation cell model was established,which was divided into 6 groups: the normal control group?NG?,the model group?MG?,Positive drug group?PD,0.4 ug/mL?,Agaricus blazei Murill polysaccharide low dose group?WABM-A-b-L,2.5ug/mL?,Agaricus blazei Murill polysaccharide middle dose group?WABM-A-b-M,5ug/mL?and Agaricus blazei Murill polysaccharide high dose group?WABM-A-b-H,10ug/mL?,observe the effect of WABM-A-b on the HepG2 cell,and observe the HepG2 cell by oil red O staining;measure the lipid content;Westernblot was used to measure PPAR?,LXR?,ABCA1 and ABCG1 of proteins expression in HepG2 cell.Results: Separation and purification of Agaricus blazei Murill acid polysaccharides?WABM-A?to obtain WABM-A-b fractions,and mainly composed of Glc?95.1%?,GlcA?3.7%?,Gal?0.9%?,Xyl?0.2%?and Fuc?0.1%?,its molecular weight is 10 kD,and its structure is dextran with-6-?-D-Glcp-1-as the main chain.In vivo experiments showed that compared with the MG group,the weight of the rats in the WABM-A group was decreased?p < 0.05?,and the organ indexes were significantly reduced?p < 0.05 or p < 0.01?;compared with the MG group,the levels of total cholesterol?TC?,triglyceride?TG?and low density lipoprotein cholesteros?LDL-C?in the WABM-A group were significantly reduced?p < 0.01?,and the content of high density lipoprotein cholesterol?HDL-C?increased significantly?p < 0.01?;compared with the MG group,the total antioxidant capacity?T-AOC?,superoxide dismutase?SOD?,glutathione peroxidase?GSH-Px?,catalase?CAT?and nitric oxide synthase?NOS?were significantly increased?p < 0.05 or p < 0.01?,malondialdehyde?MDA?activity was significantly decreased?p < 0.01?in the WABM-A group;compared with the MG group,hepatic steatosis in the hepatic lobules of rats in the WABM-A group was significantly decreased;the WABM-A group significantly up-regulated the expression of PPAR?,LXR?,ABCA1 and ABCG1 proteins?p <0.05?.Comparison of differences in OTU levels and Alpha diversity analysis showed that the microbial diversity of the intestinal contents of the MG group was significantly decreased than the WABM-A and NG groups after the administration of high-fat diets;Beta diversity analysis showed that the MG group and the WABM-A correlation is strong;comparative analysis of taxonomy showed that at the level of the phylum,the ratio of Firmicutes / Bacteroidetes of the MG group to the WABM-A group decreased from 27.12 to 8.03?p < 0.05?;compared with the MG group,the flora relative abundance of Spirochaetes?p < 0.01?,Actinobacteria?p < 0.05?,Elusimicrobia?p < 0.05?in the WABM-A group was high?p < 0.05?.At the level of the genus,compared to the MG group,the relative richness of Ruminococcaceae_unclassified?p < 0.01?was lower in WABM-A group,and the relative abundance of Clostridium_sensu_stricto?p <0.01?,Allobaculum?p < 0.01?was higher in ABP group.At the level of the family,compared to the MG group,the relative abundance of Ruminococcaceae?p < 0.01?was lower in ABP group,while the relative abundance of the Peptostreptococcaceae,Clostridiaceae₁?p < 0.01?,Erysipelotrichaceae?p < 0.01?was higher in the WABM-A group.In vitro experiments showed that compared with the MG group,WABM-A-b can significantly improve lipid accumulation inhepatocytes;compared with the MG group,the TC and TG contents in the WABM-A-b-H group were significantly reduced?p < 0.01?;compared with the MG group,the expressions of PPAR?,LXR?,ABCA1 and ABCG1 proteins were up-regulated in the WABM-A-b-H group?p < 0.05?.Conclusion: WABM-A has a lipid-lowering effect,and its mechanism is to activate the cholesterol metabolism PPAR? / LXR? / ABCA1 / ABCG1 signaling pathway.It was found that WABM-A reduced the ratio of Firmicutes/Bacteroidetes,reduce the relative abundance of Firmicutes,Ruminococcaceae_unclassified,Ruminococcaceae and increased the relative abundance of Proteobacteria,Clostridium_sensu_stricto,Allobaculum,Clostridiaceae₁ and Erysipelotrichaceae as targets to regulate the blood lipids by regulation of the balance of the intestinal microecological bacteria.WABM-A-b may be the main fraction of WABM-A to play a role in lowering blood lipids,and its lipid-lowering mechanism is achieved by activating the PPAR? / LXR? / ABCA1 / ABCG1 signaling pathway.