Integrated 16S rRNA sequencing technology and metabolomics to explore mechanism of hepatotoxicity of Dictamni Cortex on larvae zebrafish
Objective To investigate the mechanism of hepatotoxicity of Baixianpi (Dictamni Cortex) in larvae zebrafish using 16S rRNA equencing technology combined with non-targeted metabolomics techniques. Methods The zebrafish were placed in 0,100,200,300,400,500,600,700,800,1000 μg/mL concentration gradients for 24 h,the number of deaths and the lethality rate were counted,and the sublethal concentration (LC10) was calculated,and the low-,medium-and high-doses of Dictamni Cortex were set according to this,and the exposure was 24h under LC10. The activities of ALT,AST,ALB,SOD,MDA,LN and GLU in larvae zebrafish were detected. To analyze the effect of Dictamni Cortex on the distribution of larvae zebrafish by 16S rRNA sequencing,to explore the metabolic pathways of the biomarkers based on non-targeted metabolomics technology. Combining Spearman analysis for correlation between differential intestinal flora and differential metabolites. Results The LC10 of Dictamni Cortex on larvae zebrafish is 572.43 μg/mL. Compared with the control group,100,300,500 μg/mLgroups of Dictamni Cortex can increase the content of ALT,AST,MDA,LN and GLU (P<0.05,0.01),and decreased the content of SOD and ALB (P<0.05,0.01). Results of 16S rRNA sequencing revealed that Dictamni Cortex can increase the abundance of Proteobacteria,Chlamydiae,Deinococcota,Verrucomicrobiota (P<0.05,0.001),decreasing the abundance of Bacteroidota (P<0.01). Metabolomic analysis identified 32 key differential metabolites,and pathway analysis showed that Dictamni Cortex could produce toxicity by participating in sphingolipid metabolism,purine metabolism,arachidonic acid metabolism,galactose metabolism,glutathione metabolism,amino sugar and nucleotide sugar metabolism,biosynthesis of unsaturated fatty acids,drug metabolism-cytochrome P450. Conclusion Dictamni Cortex can cause hepatotoxicity in larval zebrafish and change intestinal flora structure and influence sphingolipid metabolism,purine metabolism,arachidonic acid metabolism and then promote inflammatory response,oxidative stress,apoptosis and metabolic activation.
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