Top

BMC Complementary Medicine and Therapies

Published in:

Open Access 01-12-2017 | Research article

Triptolide Induces hepatotoxicity via inhibition of CYP450s in Rat liver microsomes

Authors: Yan Lu, Tong Xie, Yajie Zhang, Fuqiong Zhou, Jie Ruan, Weina Zhu, Huaxu Zhu, Zhe Feng, Xueping Zhou

Published in: BMC Complementary Medicine and Therapies | Issue 1/2017

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.

Available only for authorised users

Kupchan SM, Court WA, Dailey Jr RG, Gilmore CJ, Bryan RF. Triptolide and tripdiolide, novel antileukemic diterpenoid triepoxides from Tripterygium wilfordii. J Am Chem Soc. 1972;94(20):7194–5.CrossRefPubMed

Qiu D, Kao PN. Immunosuppressive and anti-inflammatory mechanisms of triptolide, the principal active diterpenoid from the Chinese medicinal herb tripterygium wilfordii hook. f. Drugs R D. 2003;4(1):1–18.CrossRefPubMed

Shamon LA, Pezzuto JM, Graves JM, Mehta RR, Wangcharoentrakul S, Sangsuwan R, Chaichana S, Tuchinda P, Cleason P, Reutrakul V. Evaluation of the mutagenic, cytotoxic, and antitumor potential of triptolide, a highly oxygenated diterpene isolated from tripterygium wilfordii. Cancer Lett. 1997;112(1):113–7.CrossRefPubMed

Zheng Y, Zhang WJ, Wang XM. Triptolide with potential medicinal value for diseases of the central nervous system. CNS Neurosci Ther. 2013;19(2):76–82.CrossRefPubMed

Ziaei S, Halaby R. Immunosuppressive, anti-inflammatory and anti-cancer properties of triptolide: a mini review. Avicenna j phytomedicine. 2016;6(2):149–64.

Cao Y, Huang X, Fan Y, Chen X. Protective effect of triptolide against glomerular mesangial cell proliferation and glomerular fibrosis in rats involves the TGF- beta 1/smad signaling pathway. Evid Based Complement Alternative Med. 2015;2015:814089.

Li XJ, Jiang ZZ, Zhang LY. Triptolide: progress on research in pharmacodynamics and toxicology. J Ethnopharmacol. 2014;155(1):67–79.CrossRefPubMed

Yao J, Jiang Z, Duan W, Huang J, Zhang L, Hu L, He L, Li F, Xiao Y, Shu B, et al. Involvement of mitochondrial pathway in triptolide-induced cytotoxicity in human normal liver L-02 cells. Biol Pharm Bull. 2008;31(4):592–7.CrossRefPubMed

Cao LJ, Li HD, Yan M, Li ZH, Gong H, Jiang P, Deng Y, Fang PF, Zhang BK. The protective effects of isoliquiritigenin and glycyrrhetinic acid against triptolide-induced oxidative stress in HepG2 cells involve Nrf2 activation. Evid Based Complement Alternat Med. 2016;2016:8912184.PubMedPubMedCentral

10.

Srivastava A, Maggs JL, Antoine DJ, Williams DP, Smith DA, Park BK. Role of reactive metabolites in drug-induced hepatotoxicity. Handb Exp Pharmacol. 2010;196:165–94.CrossRef

11.

Nebert DW, Russell DW. Clinical importance of the cytochromes P450. Lancet. 2002;360(9340):1155–62.CrossRefPubMed

12.

Testa B, Pedretti A, Vistoli G. Reactions and enzymes in the metabolism of drugs and other xenobiotics. Drug Discov Today. 2012;17(11–12):549–60.CrossRefPubMed

13.

Du F, Liu Z, Li X, Xing J. Metabolic pathways leading to detoxification of triptolide, a major active component of the herbal medicine tripterygium wilfordii. J Appl Toxicol. 2014;34(8):878–84.CrossRefPubMed

14.

Zhuang XM, Shen GL, Xiao WB, Tan Y, Lu C, Li H. Assessment of the roles of P-glycoprotein and cytochrome P450 in triptolide-induced liver toxicity in sandwich-cultured rat hepatocyte model. Drug Metab Dispos. 2013;41(12):2158–65.CrossRefPubMed

15.

Shao F, Wang G, Xie H, Zhu X, Sun J, A J. Pharmacokinetic study of triptolide, a constituent of immunosuppressive chinese herb medicine, in rats. Biological & pharmaceutical bulletin.2007.30(4).702–707.

16.

Li W, Liu Y, He YQ, Zhang JW, Gao Y, Ge GB, Liu HX, Huo H, Liu HT, Wang LM, et al. Characterization of triptolide hydroxylation by cytochrome P450 in human and rat liver microsomes. Xenobiotica. 2008;38(12):1551–65.CrossRefPubMed

17.

Ye X, Li W, Yan Y, Mao C, Cai R, Xu H, Yang X. Effects of cytochrome P4503A inducer dexamethasone on the metabolism and toxicity of triptolide in rat. Toxicol Lett. 2010;192(2):212–20.CrossRefPubMed

18.

Zhang XF, Liu J, Ye F, Ji SG, Zhang N, Cao RS, He L, Wu JC, Li XF. Effects of triptolide on the pharmacokinetics of cyclophosphamide in rats: a possible role of cytochrome P3A4 inhibition. Chin J Integr Med. 2014;20(7):534–9.CrossRefPubMed

19.

Shen G, Zhuang X, Xiao W, Kong L, Tan Y, Li H. Role of CYP3A in regulating hepatic clearance and hepatotoxicity of triptolide in rat liver microsomes and sandwich-cultured hepatocytes. Food Cheml Toxicol. 2014;71:90–6.CrossRef

20.

Frye RF. Probing the world of cytochrome P450 enzymes. Mol Interv. 2004;4(3):157–62.PubMed

21.

Liu L, Jiang Z, Liu J, Huang X, Wang T, Liu J, Zhang Y, Zhou Z, Guo J, Yang L, et al. Sex differences in subacute toxicity and hepatic microsomal metabolism of triptolide in rats. Toxicology. 2010;271(1–2):57–63.CrossRefPubMed

22.

von Bahr C, Groth CG, Jansson H, Lundgren G, Lind M, Glaumann H. Drug metabolism in human liver in vitro: establishment of a human liver bank. Clin Pharmacol Ther. 1980;27(6):711–25.CrossRef

23.

Omura T, Sato R. The carbon monoxide-binding pigment of liver microsomes. Ii. Solubilization, purification, and properties. J Biol Chem. 1964;239:2379–85.PubMed

24.

Spaggiari D, Geiser L, Daali Y, Rudaz S. A cocktail approach for assessing the in vitro activity of human cytochrome P450s: an overview of current methodologies. J Pharm Biomed Anal. 2014;101:221–37.CrossRefPubMed

25.

Mahnke A, Strotkamp D, Roos PH, Hanstein WG, Chabot GG, Nef P. Expression and inducibility of cytochrome P450 3A9 (CYP3A9) and other members of the CYP3A subfamily in rat liver. Arch Biochem Biophys. 1997;337(1):62–8.CrossRefPubMed

26.

Nagata K, Murayama N, Miyata M, Shimada M, Urahashi A, Yamazoe Y, Kato R. Isolation and characterization of a new rat P450 (CYP3A18) cDNA encoding P450(6)beta-2 catalyzing testosterone 6 beta- and 16 alpha-hydroxylations. Pharmacogenetics. 1996;6(1):103–11.CrossRefPubMed

27.

Kirita S, Matsubara T. cDNA cloning and characterization of a novel member of steroid-induced cytochrome P450 3A in rats. Arch Biochem Biophys. 1993;307(2):253–8.CrossRefPubMed

28.

Matsubara T, Kim HJ, Miyata M, Shimada M, Nagata K, Yamazoe Y. Isolation and characterization of a new major intestinal CYP3A form, CYP3A62, in the rat. J Pharmacol Exp Ther. 2004;309(3):1282–90.CrossRefPubMed

29.

Martinez-Gil N, Flores-Bellver M, Atienzar-Aroca S, Lopez-Malo D, Urdaneta AC, Sancho-Pelluz J, Peris-Martinez C, Bonet-Ponce L, Romero FJ, Barcia JM. CYP2E1 in the human retinal pigment epithelium: expression, activity, and induction by ethanol. Invest Ophthalmol Vis Sci. 2015;56(11):6855–63.CrossRefPubMed

30.

Hartman JH, Martin HC, Caro AA, Pearce AR, Miller GP. Subcellular localization of rat CYP2E1 impacts metabolic efficiency toward common substrates. Toxicology. 2015;338:47–58.CrossRefPubMedPubMedCentral

31.

Lewis DF, Ito Y, Goldfarb PS. Investigating human P450s involved in drug metabolism via homology with high-resolution P450 crystal structures of the CYP2C subfamily. Curr Drug Metab. 2006;7(6):589–98.CrossRefPubMed

32.

Booth Depaz IM, Toselli F, Wilce PA, Gillam EM. Differential expression of cytochrome P450 enzymes from the CYP2C subfamily in the human brain. Drug Metab Dispos. 2015;43(3):353–7.CrossRefPubMed

33.

Kawai M, Bandiera SM, Chang TK, Bellward GD. Growth hormone regulation and developmental expression of rat hepatic CYP3A18, CYP3A9, and CYP3A2. Biochem Pharmacol. 2000;59(10):1277–87.CrossRefPubMed

34.

Zhang H, Ya G, Rui H. Inhibitory effects of triptolide on human liver cytochrome P450 enzymes and P-glycoprotein. Eur J Drug Metab Pharmacokinet. 2016.

35.

Nelson DR, Kamataki T, Waxman DJ, Guengerich FP, Estabrook RW, Feyereisen R, Gonzalez FJ, Coon MJ, Gunsalus IC, Gotoh O, et al. The P450 superfamily: update on new sequences, gene mapping, accession numbers, early trivial names of enzymes, and nomenclature. DNA Cell Biol. 1993;12(1):1–51.CrossRefPubMed

36.

Cooper KO, Reik LM, Jayyosi Z, Bandiera S, Kelley M, Ryan DE, Daniel R, McCluskey SA, Levin W, Thomas PE. Regulation of two members of the steroid-inducible cytochrome P450 subfamily (3A) in rats. Arch Biochem Biophys. 1993;301(2):345–54.CrossRefPubMed

37.

Takeuchi M, Kobayashi T, Brandao LR, Ito S. Effect of CYP2C9, VKORC1, and CYP4F2 polymorphisms on warfarin maintenance dose in children aged less than 18 years: a protocol for systematic review and meta-analysis. Syst Rev. 2016;5(1):105.CrossRefPubMedPubMedCentral

38.

Han FM, Peng ZH, Wang JJ, Chen Y. In vivo effect of triptolide combined with glycyrrhetinic acid on rat cytochrome P450 enzymes. Yao xue xue bao = Acta Pharma Sin. 2013;48(7):1136–41.

Title: Triptolide Induces hepatotoxicity via inhibition of CYP450s in Rat liver microsomes
Authors: Yan Lu
Tong Xie
Yajie Zhang
Fuqiong Zhou
Jie Ruan
Weina Zhu
Huaxu Zhu
Zhe Feng
Xueping Zhou
Publication date: 01-12-2017
Publisher: BioMed Central
Published in: BMC Complementary Medicine and Therapies / Issue 1/2017
Electronic ISSN: 2662-7671
DOI: https://doi.org/10.1186/s12906-016-1504-3

Keynote webinar | Spotlight on sleep in brain health

Springer Medicine

Triptolide Induces hepatotoxicity via inhibition of CYP450s in Rat liver microsomes

Abstract

Background

Methods

Results

Conclusions

Keynote webinar | Spotlight on sleep in brain health

Springer Medicine

Abstract

Background

Methods

Results

Conclusions

Please log in to get access to this content

Other articles of this Issue 1/2017

Psychophysical responses in patients receiving a mock laser within context of an acupuncture clinical trial: an interoceptive perspective

Case report: fainting during acupuncture stimulation at acupuncture point LI4

Scutellaria barbata D. Don extract inhibits the tumor growth through down-regulating of Treg cells and manipulating Th1/Th17 immune response in hepatoma H22-bearing mice

General perception and self-practice of complementary and alternative medicine (CAM) among undergraduate pharmacy students of Bangladesh

Erratum to: Studies on the therapeutic effect of propolis along with standard antibacterial drug in Salmonella enterica serovar Typhimurium infected BALB/c mice

Effects of a Delphi consensus acupuncture treatment protocol on the levels of stress and vascular tone in women undergoing in-vitro fertilization: a randomized clinical trial protocol