Effect Of Dihydromyricetin On Abnormal Glucose And Lipid Metabolism In Rats

Vine tea,as traditional medicinal and edible resource rich in flavonoids,is commonly used as traditional medicine or functional tea by Tujia and Zhuang nationality in Hunan,Hubei,Guizhou and other provinces.Because of its functions of reducing fat and glucose,antiinflammatory and antibacterial,and regulating microbiotas.Dihydromyricetin is the main flavonoid in Vine tea and its main active ingredient.Recent studies have shown that Dihydromyricetin has a significant effect in improving metabolic syndrome and protecting liver,but the specific molecular mechanism remains unclear.Obesity is an important inducement of type 2 diabetes,non-alcoholic fatty liver disease and cardiovascular and cerebrovascular diseases,and is closely related to the occurrence and development of metabolic syndrome.Liver——one of the most important metabolic organs of human body,it undertakes important life activities such as digestion,detoxification and biosynthesis,and its metabolic disorder will directly endanger human health.In this study,Dihydromyricetin was used as experimental material and mixed with feed to feed obese rats,and microbiome,untargeted metabolomics and transcriptomics were used to conduct correlation analysis,and the key microorganisms,target substances and target genes that were regulated by Dihydromyricetin in glucolipid metabolism were initially screened out.Furthermore,Western Blot,q RT-PCR,ELISA and other modern molecular biological techniques were used to reveal the effect and mechanism of DHM on improving metabolic syndrome and protecting liver in high-fat rats,combined with ABTS,DPPH,ORAC and other antioxidant activities in vitro.The results are as follows:1.Using different concentrations of Dihydromyricetin(low dose,150 mg/kg;high dose,450 mg/kg)to feed obese rats induced by high-fat diet,it was found that dihydromyricetin can reduce triglyceride(TC),total cholesterol(TG)and low density lipoprotein(LDL-C),and restore the level of high density lipoprotein(HDL-C)(p < 0.05),and reduce body weight and blood glucose level,decreased lipid drop content in liver cells,decreased alanine aminotransferase(ALT),aspartate aminotransferase(AST)activities and malondialdehyde(MDA)content,and increase the activities of catalase(CAT),glutathione peroxidase(GSHPX)and superoxide dismutase(SOD)(p < 0.05),reduce the accumulation of reactive oxygen species(ROS)in tissues.2.Using 16 S r DNA sequencing technology to detect fecal samples of obese rats,it was found that Dihydromyricetin could significantly change the richness and diversity of microbiotas.For example,the ratio of Firmicutes /Bacteroidetes and Proteobacteria was reduced,and the relative abundance of Verrucomicrobia was restored at the phylum level(p< 0.05).At the genus level,reduce the relative abundance of Ruminococcaceae＿UCG-005、Clostridiales and Bacteroides,and increase the levels of probiotics such as Parabacteroides and Akkermansia(p < 0.05).By analyzing the correlation between physical and chemical indexes and intestinal flora,it was found that probiotics such as Akkermansia and Parabacteroides were positively correlated with SOD,CAT,GSH-Px and HDL-C indexes.Ruminococcus,Clostridiales and Lachnospiraceae＿UCG-010 were positively correlated with TG,TC,MDA,LDL-C,ALT,AST,blood glucose and body weight.3.Using non-target metabolomics analysis technology to detect fecal samples of obese rats,it was found that Dihydromyricetin could p-regulated the levels of hydroxyl cinnamic acid family,Paraxanthine and 13-Oxo ODE,and down-regulated the levels of fatty acids such as phthalic acid and 5-hydroxyindole-3-acetic acid.Through the correlation analysis of physical and chemical indexes,intestinal microflora and differential metabolites,Trans-3-hydroxycinnamic acid,13-Oxo ODE and paraxanthine were positively correlated with physical and chemical indexes(SOD,CAT,etc)and probiotics(Akkermansia,Parabacteroides,etc).Metabolites such as phthalic acid and traumatic acid were positively correlated with physical and chemical indexes(AST,ALT,etc).4.RNA sequencing was used to determine the gene expression in the liver tissue of highfat obese rats,and using PCR technology to verify the transcriptome datas.Dihydromyricetin could reduce the expression of lipid synthesis genes(FABPs,CD36,etc.)(p < 0.05),increased the expression of PPARγ gene,and regulated the expression of glucose metabolism genes(ENO2,Pfkl,etc.)and oxidative stress genes(ATP1A1,Xiap)(p < 0.05).Multiple omics correlation analysis showed that PPARα,SLC25A4,Pgam2 were negatively correlated with MDA,ROS,LDL-C,and positively correlated with GSH-Px.It was positively correlated with microorganism Clostridiales,Ruminococcaceae＿UCG-005 and metabolites such as phthalic acid and traumatic acid.After safe toxicological analysis of transcriptome sequencing data,it was found that Dihydromyricetin could restore the expression of CYP7A1 gene,and gene NTRK2,ADCY3,TCF7L2 and other pathogenic genes were not induced by Dihydromyricetin.5.The protein expression of rat liver was detected by in vitro antioxidant capacity test and Western blot.It was found that Dihydromyricetin can up-regulate the expression of antioxidant proteins Nrf2 and HO-1 in tissues.Dihydromyricetin can regulate the expression of p-Akt and p-GSK-3β protein expression.Although Dihydromyricetin did not regulate NF-κB and i NOS,it down-regulated the expression of inflammatory factor COX-2.DHM can increase the expression of lipid metabolism protein PPARγ.In conclusion,through multi-omics association analysis and molecular validation research,we found that Dihydromyricetin can affect the intestinal absorption and digestion of short chain fatty acids and small molecule metabolites by interfering with the relative abundance of microbiotas like Parabacteroides and Akkermansia,and regulate the expression of genes related to glucose and lipid metabolism(ATP1A1,Xiap,PPARγ),as well as the expression of related proteins in the redox pathway(Nrf2 / HO-1,NF-κB / i NOS / COX-2)and glucose and lipid metabolism pathway(Akt / GSK,PPARγ),so as to intervene the disorder of glucose and lipid metabolism and protect the liver in the obesity model of rats induced by high-fat diets.The results of this study will provide theoretical support for the development of functional foods using Dihydromyricetin as raw materials and the research and development of related hypoglycemic and lipid lowering products.