Study On Metabolic Toxicology Of Exogenous And Endogenous Small Molecules

Research purposes From the perspective of small molecular substances,this study selected representative exogenous environmental pollutants,drug molecules and endogenous bile acid molecules to research the mechanism of toxic effect based on metabolic toxicology.Research content and methods1.Inhibition of UGT by PAEs and phthalate monoestersIn vitro metabolic enzyme incubation system was established to detect the inhibition of highly exposed PAEs(DAP,DBP,DCHP,DEHP,DEP,Di BP,Di OP,Di PP,DMEP,DMP,DHP,DNP,DPP,BBOP and BBZP)and phthalate monoesters(MBP,MBZP,MCHP,MEHP,MEP,MHP,MMP and MOP)on the main UGT of human body(UGT1A1,UGT1A3,UGT1A6,UGT1A7,UGT1A8,UGT1A9,UGT1A10,UGT2B4,UGT2B7,UGT2B15 and UGT2B17).PAEs and phthalate monoesters which significantly inhibited UGT activity were choosen to determine the median inhibition concentration(IC50),inhibition kinetic parameter(Ki)and type of inhibition kinetics,and determine the binding site,binding energy,hydrogen bonding,and hydrophobic interaction of PAEs and phthalate monoesters with UGT through computer simulation of molecular docking,so as to clarify the dose-effect relationship of the inhibitory effect of PAEs and phthalate monoesters on UGT,and predict its effect on endogenous substances by inhibiting UGT activity and explain its various toxic mechanisms.2.The inhibitory effect of everolimus on UGT and the inhibitory effect of PC,PD and PE on CESBased on the in vitro metabolic enzyme incubation system,the inhibitory effect of everolimus on UGT(UGT1A1,UGT1A3,UGT1A6,UGT1A7,UGT1A8,UGT1A9,UGT1A10,UGT2B7,and UGT2B17)and PC,PD,PE on CES1 and CES2 were tested and the dose-effect relationship between everolimus and PC,PD,PE with the corresponding metabolic enzymes was determined by the IC50,Ki and the type of inhibition kinetics.Through computer simulation of molecular docking,the binding site of the interaction between everolimus and UGT was determined.Binding energy,hydrogen bonding,and hydrophobic interaction were determined to explain the toxic effect of everolimus and PC,PD,PE by affecting the corresponding metabolic enzymes and the mechanism of drug interaction.3.The mechanism of bile acid in the process of gallstone diseaseGallstone disease mice model was established and based on the metabolomics platform the changes of various bile acid levels in mice with gallstones were detected.Molecular biology methods were used to detect changes in receptors and metabolic enzymes related to bile acid metabolism,and clear the key bile acids,receptors and metabolic enzymes regulating gallstone disease.Based on gene knockout technology,the key gene of bile acid metabolism-FXR knockout mouse was established to detect the effect of FXR knockout on bile acid metabolism and gallstone phenotype.Representative bile acid TUDCA was selected to treat gallstone mice to detect the effect of TUDCA on gallstone phenotype and related signaling molecules,so as to explain the mechanism of bile acid in the process of gallstone disease by affecting related metabolic enzymes and signaling molecules.Research results and conclusion1.PAEs and phthalate monoesters affect the metabolism of endogenous substances by inhibiting UGTPAEs have strong inhibitory effect on UGT1A6,UGT1A7,UGT1A9 and UGT2B4,and phthalate monoesters have strong inhibitory effect on UGT1A7 and UGT1A9.PAEs and phthalate monoesters combine with UGT through hydrogen bonding and hydrophobic interaction,and may have inhibitory effect on UGT in vivo.Therefore,PAEs and phthalate monoesters may interfere with the metabolism of endogenous substances by inhibiting UGT and produce toxic effect.2.Everolimus and PC,PD,PE affect the metabolism of endogenous substance and induce drug interactions by inhibiting UGT and CES,respectivelyEverolimus has a strong and significant inhibitory effect on the activitie of UGT1A1,UGT1A3 and UGT2B7.Computer simulation of molecular docking shows that the small molecule everolimus binds to UGT1A3 and UGT2B7 proteins through hydrogen bonding and hydrophobic interaction,thereby inhibiting the catalytic effect of UGT1A3 and UGT2B7.Everolimus is most likely to inhibit UGT1A3 in vivo,so it may induce metabolic diseases and drug interactions by affecting the metabolism of endogenous substances and drugs catalyzed by UGT1A3.PD has the most significant inhibitory effect on CES1,and the in vitro inhibitory potential can also be converted into the ability in vivo.It is predicted that PD may cause endogenous metabolic disorders and drug-drug interactions in vivo by inhibiting CES1.3.Bile acid metabolism plays an important regulatory role in gallstone diseaseThe levels of T-?MCA,TUDCA,THDCA and TLCA in the serum of gallstone mice show a significant decrease,indicating that in the course of gallstone disease,the disorder of bile acid metabolism may induce the occurrence of the disease.The expression of intestinal FXR in mice with gallstones is significantly up-regulated,indicating that bile acid may affect gallstone disease by regulating intestinal FXR.FXR knockout can improve the gallstone phenotype by regulating liver bile acid metabolism.Choosing representative bile acid TUDCA to treat gallstone mice,it was found that TUDCA can regulate gallstone disease by inhibiting intestinal FXR.