| Background: The effects of gut microbiota on the response and metabolism of exogenous substance have aroused great interest.The pseudo germ-free animal models established by antibiotic treatment have been widely used as a favorable "tool" to study the relationships between gut microflora and drug metabolism and diseases.However,relatively less is known about the expression profiles of host drug-processing genes of pseudo germ-free rats and the effects of the absence of gut microflora on pharmacokinetic behaviors of chemical drugs.Objective: To characterize the expression profiles of host drugprocessing genes in pseudo germ-free rats and to investigate the effects of the lack of intestinal microbiota on pharmacokinetic behaviors of CYP450 s probe drugs.Methods: Wistar rats were gavaged quadruple antibiotic mixture for continuous 2 weeks to clear intestinal microflora of recipient rats as pseudo germ-free rats model group,while conventional rats(CV)were given the same dose of normal saline.After the modeling was successful,RNA-seq was used to analyze the effects of the loss of intestinal microbiota on the expression profiles of drug-processing genes in host liver and jejunum tissues,and the transcriptome results were verified by RT-q PCR.The mixture of six drugs,phenacetin,tolbutamide,omeprazole,metoprolol,chlorzoxazone and midazolam,was given intragastorally in the manner of Cocktail.Blood samples were collected at each time point by rat tail microcapillary method.Plasma concentration of six probe drugs were measured with a validated UPLC-MS/MS method.Serum bile acid metabolism was analyzed by targeted metabonomics.The protein expression and enzyme activity of major CYP450 s in liver were determined by Western Blot and in vitro liver microsomal incubation.Results: The pseudo germ-free condition altered the expression of116 drug-processing genes in host liver and jejunum tissues.Compared with CV rats,the m RNA expression levels of Cyp2a1,Cyp2c11,Cyp2c13,Cyp2 d,Cyp2e1 and Cyp3a1 of CYP450 family members in pseudo germfree rats were significantly down-regulated.KEGG pathway enrichment analysis showed that the absence of gut microbiota significantly altered biological pathways related to drug metabolism in the body.Pharmacokinetic results showed that,except for tolbutamide,the loss of gut microbiota significantly altered the pharmacokinetic behaviors of phenacetin,omeprazole,metoprolol,chlorzoxazone,and midazolam.The results of in vitro and in vivo experiments preliminarily showed that the lack of gut microflora may down-regulate the protein expression and enzyme activity of Cyp2e1 and Cyp3a1 in liver by affecting the composition of bile acid profiles,thereby altering the pharmacokinetics of their substrate drugs.Conclusion: The absence of gut microflora altered the expression levels of a large number of host drug-processing genes and affected pharmacokinetic behaviors of CYP450 s probe drugs. |
