Intervention Mechanism Of Auricularia Auricula Polysaccharides On High Fat Diet-Induced Obesity

Obesity has become a disease that affects human health and has a high incidence.Auricularia auricula polysaccharide has anti-obesity biological activity,but its mechanism of intervention on high fat diet-induced obesity is not clear.In this research,based on the extraction,purification,and preliminary structural characterization of polysaccharides from Auricularia auricula,this study established a high fat diet-induced obesity model,and used microbiology and metabolomics techniques to study the possible mechanism of Auricularia auricula polysaccharides to exert obesity intervention effects.The technology of pseudo-sterile mouse and faecal transplantation has further been used and verified the possible mechanism of Auricularia auricula polysaccharides intervention in high fat-induced obesity.The purpose of this study is to elucidate the mechanism by which Auricularia auricula polysaccharides interfere with high-fat-induced obesity,to screen the target intestinal microorganisms for Auricularia auricula polysaccharides,and to provide a theoretical basis for the prevention and treatment of obesity.The research mainly includes the following four aspects:1.Extraction,purification and preliminary structural characterization of polysaccharides from Auricularia auriculaThe polysaccharide was extracted from Auricularia auricula fruit body by hot water extraction and ethanol precipitation,and the protein was removed by repeated freeze-thaw method,and Auricularia auricula polysaccharide was obtained by freeze-drying.On this basis,phenol-sulfuric acid method and Coomassie Brilliant Blue method were used to determine the purity of Auricularia auricula polysaccharides,and it was confirmed that the obtained polysaccharides were relatively pure,with a total sugar content of 97.6% and a protein content of 2.4%.In terms of structure characterization,Fourier transform infrared spectroscopy was used to detect the structure of Auricularia auricula polysaccharide.The results showed that the prepared polysaccharide was an acidic polysaccharide containingβ-D-glucopyranose and glycoprotein.2.Study on the intervention effect of Auricularia auricula polysaccharide on high fat diet-induced obesityC57BL/6 male mice were first treated with a high-fat diet to establish the high fat diet-induced obesity model,which was verified by comparison with chow-fat diet mice.The results show that a high-fat diet could significantly increase the body weight,body weight gain,obesity index,liver weight,heart weight,fat weight,average area of fat cells,blood glucose concentration,T-CHO and LDL-C concentrations,and significantly reduce mouse glucose tolerance.The results showed that the high fat diet-induced obesity model was successfully established.The possible mechanism for the intervention of Auricularia auricula polysaccharides(200 mg/kg BW)on high fat diet-induced obesity was investigated based on metabolomic techniques and 16 S r RNA sequsencing.The results show that Auricularia auricula polysaccharide could inhibit the process of obesity,including significantly reducing body weight,body weight gain,obesity,liver weight,heart weight,average area of fat cells,and TG concentration in high-fat diet mice.A total of 7 differential metabolites were screened.Compared with the high fat diet-induced obesity model,the intake of Auricularia auricula polysaccharide could significantly increase the content of bis2-hydroxypropylamine and mannose in the serum,and significantly reduce methylmalonic acid,ergosterol,4-hydroxyphenylacetic acid,elaidic acid and monoolein.Changes in relative concentrations of metabolites suggested that Auricularia auricula polysaccharides could improve metabolic disorders induced by a high-fat diet,including significantly affected fructose and mannose metabolism(P=0.03),amino sugar and nucleotide sugar metabolism(P=0.03),galactose metabolism(P=0.03),valine,leucine and isoleucine degradation(P=0.04),fatty acid biosynthesis(P=0.04),biosynthesis of unsaturated fatty acids(P=0.04),and tyrosine metabolism(P=0.09).In addition,using microbiome technology,it was found that compared with the high fat diet-induced obesity model,the intake of polysaccharides from Auricularia auricula could significantly change the diversity of intestinal microbiota,with the tendency to increase the Shannon index.It has also indicated Auricularia auricula polysaccharides could significantly improve the relative abundance of Desulfotomaculum guttoideum,Eubacterium saphenum,Muribacum intestinale,Catabacter hongkongensis,Emergencia timonensis,Parabacteroides goldsteinii,Alistipes putredinis,Papillibacter cinnamivorans and Massilimaliae massiliensis;while significantly reduce the relative abundance of Faecalicatena contorta and Clostridium leptum.These intestinal microorganisms were highly correlated with mouse phenotype and serum metabolism and might play an important role in the anti-obesity mechanism of Auricularia auricula polysaccharides.3.Pseudo-sterile mouse model validated Auricularia auricula polysaccharide intervention in obesityMultiple antibiotics were used to deplete intestinal microorganisms in mice,and a pseudo-sterile model was established.Based on this,the relationship between the effect of Auricularia auricula polysaccharides on obesity and specific intestinal microorganisms was explored through metabolomics and microbiology techniques.The results showed that compared with the pseudo-sterile model group,the pseudo-sterilized Auricularia auricula polysaccharide intervention group had no significant difference in body weight,body weight gain,organ indicators,blood glucose tolerance,and blood lipid content.The results showed that intestinal microbiota might play an important role in the anti-obesity effect of Auricularia auricula polysaccharides.Metabolomics studies indicate that compared with the polysaccharide intervention group,antibiotics significantly increased the content of4-hydroxyphenylacetic acid and elaidic acid in the serum of mice,and significantly reduced the content of l-homoserine,capric Acid,ribulose-5-phosphate,2,3-dihydroxypyridine,dihydroxyacetone and p-cresol.Changes in differential metabolites caused changes in 6metabolic pathways,including glycerolipid metabolism(P=0.04),tyrosine metabolism(P=0.04),fatty acid biosynthesis(P=0.04),biosynthesis of unsaturated fatty acids(P=0.05),pentose phosphate pathway(P=0.06),and purine metabolism(P=0.06).The results also suggested that the intestinal microbiota played an important role in the host’s energy metabolism.In addition,it was found that antibiotics significantly reduced the species diversity and abundance of intestinal microbiota,and significantly reduced or eliminated a variety of intestinal bacteria caused by caused by Auricularia auricula polysaccharides,including Stomatobaculum longum,Clostridium asparagiforme,Desulfotomaculum guttoideum,Papillibacter cinnamivorans,Bacteroides thetaiotaomicron,Bacteroides caccae,Bacteroides cellulosilyticus,Parabacteroides distasonis,Christensenella minuta,Clostridium saccharolyticum,Parabacteroides goldsteinii,Roseburia hominis,Muribaculum intestinale,Bacteroides uniformis,Desulfonispora thiosulfatigenes and Intestinimonas butyriciproducens et al.In addition,correlation analysis showed that the above bacteria were significantly related to the phenotype and serum metabolites.The above results indicated that Auricularia auricula polysaccharide was likely to regulate the intestinal microbial composition,thereby improving lipid metabolism and inhibiting the occurrence of high fat diet-induced obesity.4.Fecal microbiota transplantation confirmed that Auricularia auricula polysaccharide exerted high fat-induced obesity intervention effect through intestinal microorganism The effect of intestinal microbiota on the host phenotype was further investigated using fecal microbiota transplantation technology to screen target microorganisms inducd by Auricularia auricula polysaccharide.Three groups of mice were fed a high-fat diet for 6 weeks and received intestinal microbiota from the high fat diet-obesity model group,Auricularia auricula polysaccharide intervention group,and the pseudo-sterile Auricularia auricula polysaccharide intervention group.Compared with the other two groups,the mice that received fecal microbiota from Auricularia auricula polysaccharide intervention group significantly decreased the body weight,body weight gain,obesity index,liver weight,heart weight,fat weight,average area of fat cells,blood glucose concentration,TG,T-CHO,and LDL-C concentrations,while increased the blood glucose tolerance and HDL-C concentrations.In addition,there was no significant difference in the above indicators in the other two groups.The results indicated that the intestinal microbiota was the key target for Auricularia auricula polysaccharide in obesity.In addition,compared with the other two groups,the abundance of microbiota of the mice receiving fecal microbiota from the polysaccharide intervention group significantly increased.The relative abundance of Papillibacter cinnamivorans was significantly higher than the other two groups.The relative abundance of this microorganism was significantly positively correlated with the serum HDL-C content of the mice;it was significantly negatively related to the body weight,liver weight,fat weight,fat cell area,fasting blood glucose,serum TG and LDL-C.Therefore,Papillibacter cinnamivorans was initially identified as a target microorganism for Auricularia auricula polysaccharides to interfere with obesity.The above results indicated that Auricularia auricula polysaccharide had a significant inhibitory activity to high fat diet-induced obesity,which might be related to the the improment of the abundance and diversity of intestinal microbiota,the modulation of metabolic disorders and energy metabolism pathways induced by high fat diet.