Archives of Toxicology

, Volume 92, Issue 6, pp 1939–1952 | Cite as

Omics-based responses induced by bosentan in human hepatoma HepaRG cell cultures

  • Robim M. Rodrigues
  • Laxmikanth Kollipara
  • Umesh Chaudhari
  • Agapios Sachinidis
  • René P. Zahedi
  • Albert Sickmann
  • Annette Kopp-Schneider
  • Xiaoqi Jiang
  • Hector Keun
  • Jan Hengstler
  • Marlies Oorts
  • Pieter Annaert
  • Eef Hoeben
  • Eva Gijbels
  • Joery De Kock
  • Tamara Vanhaecke
  • Vera Rogiers
  • Mathieu VinkenEmail author
Toxicokinetics and Metabolism


Bosentan is well known to induce cholestatic liver toxicity in humans. The present study was set up to characterize the hepatotoxic effects of this drug at the transcriptomic, proteomic, and metabolomic levels. For this purpose, human hepatoma-derived HepaRG cells were exposed to a number of concentrations of bosentan during different periods of time. Bosentan was found to functionally and transcriptionally suppress the bile salt export pump as well as to alter bile acid levels. Pathway analysis of both transcriptomics and proteomics data identified cholestasis as a major toxicological event. Transcriptomics results further showed several gene changes related to the activation of the nuclear farnesoid X receptor. Induction of oxidative stress and inflammation were also observed. Metabolomics analysis indicated changes in the abundance of specific endogenous metabolites related to mitochondrial impairment. The outcome of this study may assist in the further optimization of adverse outcome pathway constructs that mechanistically describe the processes involved in cholestatic liver injury.


Bosentan BSEP HepaRG Cholestasis Transcriptomics Proteomics Metabolomics Adverse outcome pathway. 



ATP-binding cassette subfamily B member 11


ATP-binding cassette subfamily C member 2


Alcohol dehydrogenase


Adverse outcome pathway


Bicinchoninic acid assay


Branched chain amino acid metabolites


Bile salt export pump


Cholic acid


Constitutive androstane receptor




Cytochrome P450 2B6


Cytochrome P450 3A4




Dimethyl sulfoxide


Filter aided sample preparation


False discovery rate


Farnesoid X receptor


Glycocholic acid


Hank’s balanced salt solution


Ingenuity pathway analysis


Interleukin 6


Inhibitory concentration of 10%


Inhibitory concentration of 50%


Interleukin 8


Isobaric tags for relative and absolute quantification


Key event(s)


Liquid chromatography–mass spectrometry/mass spectrometry


Log2-fold change


Molecular initiating event


Multidrug resistance-associated protein 2


3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide


Nuclear magnetic resonance


Sodium–taurocholate cotransporting polypeptide


Organic anion transporter 1B1/3


Organic solute transporter α andβ


Principal component analysis


Phosphatase inhibitor


Peptide-spectrum matches


Probe sliding window-analysis of variance




Pregnane X receptor


Room temperature


Reactive oxygen species


Retinoid X receptor


Sodium dodecyl sulphate


Solute carrier organic anion transporter family member 1B1


Affymetrix transcriptome analysis console




Trifluoroacetic acid


Tumor necrosis factor



This work was financially supported by the grants of European Union (FP7)/Cosmetics Europe (SEURAT-1 projects DETECTIVE (HEALTH-F5-2010-266838) and HeMiBio (HEALTH-F5-2010-266777)), the European Research Council (Starting Grant 335476), the Fund for Scientific Research-Flanders (FWO grants G009514N, G010214N, G012318N, G020018N and 12H2216N), the University Hospital of the Vrije Universiteit Brussel-Belgium (“Willy Gepts Fonds” UZ-VUB) and the Center for Alternatives to Animal Testing (CAAT) at Johns Hopkins University Baltimore-USA. The authors also gratefully acknowledge the financial support from the Ministerium für Innovation, Wissenschaft und Forschung des Landes Nordrhein-Westfalen, Senatsverwaltung für Wirtschaft, Technologie und Forschung des Landes Berlin, and the Bundesministerium für Bildung und Forschung. The authors like to thank Dr. Christophe Chesné (Biopredic) for making HepaRG cells available, Dr. Stefan Vinckier for assistance with confocal microscopy and Miss Tineke Vanhalewyn for technical assistance.

Supplementary material

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Supplementary material 1 (PPTX 3216 KB)
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Supplementary material 3 (DOCX 24 KB)
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Supplementary material 4 (DOCX 26 KB)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Robim M. Rodrigues
    • 1
  • Laxmikanth Kollipara
    • 2
  • Umesh Chaudhari
    • 3
  • Agapios Sachinidis
    • 3
  • René P. Zahedi
    • 2
  • Albert Sickmann
    • 2
    • 4
    • 5
  • Annette Kopp-Schneider
    • 6
  • Xiaoqi Jiang
    • 6
  • Hector Keun
    • 7
  • Jan Hengstler
    • 8
  • Marlies Oorts
    • 9
  • Pieter Annaert
    • 9
  • Eef Hoeben
    • 10
  • Eva Gijbels
    • 1
  • Joery De Kock
    • 1
  • Tamara Vanhaecke
    • 1
  • Vera Rogiers
    • 1
  • Mathieu Vinken
    • 1
    Email author
  1. 1.Department of In Vitro Toxicology and Dermato-CosmetologyVrije Universiteit BrusselBrusselsBelgium
  2. 2.Leibniz-Institut für Analytische Wissenschaften-ISAS-e.V.DortmundGermany
  3. 3.Institute of Neurophysiology and Center for Molecular Medicine CologneUniversity of CologneCologneGermany
  4. 4.Department of Chemistry, College of Physical SciencesUniversity of AberdeenAberdeenUK
  5. 5.Medizinische Fakultät, Medizinische Proteom-Center (MPC)Ruhr-Universität BochumBochumGermany
  6. 6.Division of BiostatisticsGerman Cancer Research CenterHeidelbergGermany
  7. 7.Computational and Systems Medicine, Department of Surgery and CancerImperial College LondonLondonUK
  8. 8.Leibniz Research Centre for Working Environment and Human Factors at the Technical University of DortmundDortmundGermany
  9. 9.Drug Delivery and Disposition, Department of Pharmaceutical and Pharmacological SciencesKatholieke Universiteit LeuvenLeuvenBelgium
  10. 10.BioNotus GCVNielBelgium

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