| This study of liver injury in mice induced by Diosbulbin B(LIDB)based on nuclear magnetic resonance(l^-NMR)and gas chromatography-time of flight mass spectrometry(GC-TOFMS)of metabonomics approach and 16S rRNA high-throughput sequencing approach,which provide toxicity of Dioscorea bulbifera L a new experimental evidence from the perspective of metabolic networks and intestinal flora.The main contents of the study are as follows:Firstly,the experiment was divided into Diosbulbin B group and Normal group.The corresponding drugs were administered to the rats for 14 consecutive days by intragastric administration.The levels of alanine aminotransferase(ALT)and aspartate aminotransferase(AST)in serum,malondialdehyde(MDA)and superoxide dismutase(SOD)in liver tissue were detected 24h after last administration in both group.Hepatic histopathological changes were observed by hematoxylin-eosin staining.The results showed that:(1)serum ALT,AST and MDA levels in liver tissues of mice in the both group were increased,while SOD levels in liver tissues were decreased.(2)the liver cells in normal groups had clear boundaries,neat arrangement and normal morphology,and no histopathological abnormalities were observed.Fatty degeneration and edema were observed in liver tissue cells of mice in the Diosbulbin B group.In addition,Diosbulbin B group steatosis,edema and pathological changes were observed in some cells of liver tissue.Secondly,the metabolic profiles of urine and liver in mice were measured by ~1H-NMR approach,and the metabolic phenotypic changes induced by Diosbulbin B in mice were studied by Orthogonal Partial Least-Squares Discriminant Analysis(OPLS-DA).Variable Importance in the Projection(VIP)and t-test were used to screen potential biomarkers Results:(1)Diosbulbin B can cause metabolism phenotype of urine and faeces of mice in disorder,characterized by 8 biomarkers of urine(acetone,pyruvate,2-oxoglutarate,taurine,glycine,phenylacetylglycine,hippurate,indoxylsulfate)and 9 biomarkers of liver tissue(bile acids,propionate,glutamine,lactate,threonine,?-Glucose,?-glucose,tyrosine,phenylalanine)related to LIDB have a significant change.(2)the metabolic pathway of LIDB mainly involves the metabolism of tricarboxylic acid,amino acid and intestinal flora.Thirdly,the metabolic profiles of serum in mice were measured by GC-TOFMS approach,and the metabolic phenotypic changes induced by Diosbulbin B in mice were studied by Orthogonal Partial Least-Squares Discriminant Analysis(OPL S-DA).Variable Importance in the Projection(VIP)and t-test were used to screen potential biomarkers.Results:(1)9 markers related to LIDB were selected(alpha ketone isovaleric acid,allantoin,taurine,caproic acid,D-Sue sugar alcohol,sugar,glutaric acid).(2)LIDB metabolic pathways may be related to abnormal energy metabolism,oxidative stress,lipid peroxidation and intestinal flora metabolism and related amino acid metabolism.Fourthly,the changes of the structure and composition of intestinal microflora in mice were studied by using 16S rRNA high-throughput sequencing approach.The construction of a small fragment library was carried out by using the Paired-End method,and sequenced by the Illumina HiSeq sequencing platform,after the splicing and filtration of the Reads,the OTU(Additional Taxonomy Units)cluster was carried out,and the OTU was analyzed by the taxonomy database.The potential biomarkers were identified by the PCoA,analysis of variance(ANOVA)and LEfSe analysis.The results show that:(1)Diosbulbin B can cause structure changes in the intestinal microflora of mice and reduce the diversity of the microflora.(2)eleven potential biomarkers were found including:Bacteroidetes,Firmicutes,Saccharibacteria,Succinivibrionaceae,acidaminococcaceae,Rikenellaceae,Candidatus Saccharimonas,Selenomonas 1,Succiniclasticum,Escherichia-Shigella,Rikenellaceae_RC9_gut_group,suggesting that the mechanism of liver injury induced by biosbulbin B was related to short-chain fatty acid metabolism,bile acid metabolism and bile acid reabsorption. |
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