title: Triptolide Induces hepatotoxicity via inhibition of CYP450s in Rat liver microsomes.
abstract: BACKGROUND
  Triptolide (TP), an active constituent of Tripterygium wilfordii, possesses numerous pharmacological activities. However, its effects on cytochrome P450 enzymes (CYP450s) in rats remain unexplored.
  
  
  METHODS
  In this study, the effects of triptolide on the six main CYP450 isoforms (1A2, 2C9, 2C19, 2D6, 2E1, and 3A) were investigated both in vivo and in vitro. We monitored the body weight, survival proportions, liver index, changes in pathology, and biochemical index upon TP administration, in vivo. Using a cocktail probe of CYP450 isoform-specific substrates and their metabolites, we then carried out in vitro enzymatic studies in liver microsomal incubation systems via ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Finally, we verified our results at the messenger ribonucleic acid (mRNA) and protein level through quantitative real-time polymerase chain reaction (RT-qPCR), western blotting, and immunohistochemical detection.
  
  
  RESULTS
  The in vivo toxicity study confirmed that Sprague-Dawley (SD) rats exhibited dose-dependent hepatotoxicity after intragastric administration of TP [200, 400, and 600 μg/(kg.day)] for 28 days. In case of the CYP450 isoforms 3A, 2C9, 2C19, and 2E1, the in vitro metabolic study demonstrated a decrease in the substrate metabolic rate, metabolite production rate, and Vmax, with an increase in the Km value, compared with that observed in the control group. Additionally, a TP dose-dependent decrease in the mRNA levels was observed in the four major isoforms of CYP3A subfamily (3A1/3A23, 3A2, 3A9, and 3A62) and CYP2C9. A similar effect was also observed with respect to the protein levels of CYP2C19 and CYP2E1.
  
  
  CONCLUSIONS
  This study suggests that TP can cause hepatotoxicity by reducing the substrate affinity, activity, and expression at the transcriptional and protein levels of the CYP450 isoforms 3A, 2C9, 2C19, and 2E1. TP also has the potential to cause pharmacokinetic drug interactions when co-administered with drugs metabolized by these four isoforms. However, further clinical studies are needed to evaluate the significance of this interaction.
authors.affiliation: The First Clinical College, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.|The Third Affiliated Hospital, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Lu
authors.affiliation: The First Clinical College, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Tong
authors.initials: T
authors.lastname: Xie
authors.affiliation: The Third Affiliated Hospital, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Yajie
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: The Third Affiliated Hospital, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Fuqiong
authors.initials: F
authors.lastname: Zhou
authors.affiliation: The Third Affiliated Hospital, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Ruan
authors.affiliation: The Third Affiliated Hospital, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Weina
authors.initials: W
authors.lastname: Zhu
authors.affiliation: College of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Huaxu
authors.initials: H
authors.lastname: Zhu
authors.affiliation: The First Clinical College, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China.
authors.forename: Zhe
authors.initials: Z
authors.lastname: Feng
authors.affiliation: The First Clinical College, Nanjing University of Chinese Medicine, Nanjing, Jiangsu, China. zxp@njutcm.edu.cn.
authors.forename: Xueping
authors.initials: X
authors.lastname: Zhou
mesh_terms: D000818:Animals; D065607:Cytochrome P-450 Enzyme Inhibitors / Q000737:chemistry / Q000633:toxicity*; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000378:metabolism; D004224:Diterpenes / Q000737:chemistry / Q000633:toxicity*; D004852:Epoxy Compounds / Q000737:chemistry / Q000633:toxicity; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D008297:Male; D008862:Microsomes, Liver / Q000737:chemistry / Q000187:drug effects* / Q000201:enzymology*; D010616:Phenanthrenes / Q000737:chemistry / Q000633:toxicity*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D004224:Diterpenes; D004852:Epoxy Compounds; D007527:Isoenzymes; D010616:Phenanthrenes; C001899:triptolide; D003577:Cytochrome P-450 Enzyme System
keywords: CYP450s; Hepatotoxicity; Microsomes; Triptolide