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Characterization of chemical-induced sterile inflammation in vitro: application of the model compound ketoconazole in a human hepatic co-culture system

  • Immunotoxicology
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Abstract

Liver injury as a result of a sterile inflammation is closely linked to the activation of immune cells, including macrophages, by damaged hepatocytes. This interaction between immune cells and hepatocytes is as yet not considered in any of the in vitro test systems applied during the generation of new drugs. Here, we established and characterized a novel in vitro co-culture model with two human cell lines, HepG2 and differentiated THP-1. Ketoconazole, an antifungal drug known for its hepatotoxicity, was used as a model compound in the testing of the co-culture. Single cultures of HepG2 and THP-1 cells were studied as controls. Different metabolism patterns of ketoconazole were observed for the single and co-culture incubations as well as for the different cell types. The main metabolite N-deacetyl ketoconazole was found in cell pellets, but not in supernatants of cell cultures. Global proteome analysis showed that the NRF2-mediated stress response and the CXCL8 (IL-8) pathway were induced by ketoconazole treatment under co-culture conditions. The upregulation and ketoconazole-induced secretion of several pro-inflammatory cytokines, including CXCL8, TNF-α and CCL3, was observed in the co-culture system only, but not in single cell cultures. Taking together, we provide evidence that the co-culture model applied might be suitable to serve as tool for the prediction of chemical-induced sterile inflammation in liver tissue in vivo.

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Abbreviations

ATP:

Adenosine triphosphate

CCL3:

Chemokine (C–C-motif) ligand 3

CD54:

Cluster of differentiation 54

CXCL8:

C–X–C-motif ligand 8, interleukin 8

DAK:

N-deacetyl ketoconazole

GM-CSF:

Granulocyte macrophage colony-stimulating factor

HMGB1:

High-mobility group box 1

IL-1rα:

Interleukin 1 receptor-α

KTZ:

Ketoconazole

LC50 :

Half maximal toxic concentration

MIF:

Macrophage migration inhibitory factor

TNF-α:

Tumor necrosis factor-α

Serpin E1:

Serpin peptide inhibitor clade E (nexin, plasminogen activator inhibitor type 1)

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Acknowledgments

We would like to thank Barbara Gerding for excellent technical assistance and S. Weichenhain for discussions. The financial support of the BfR through intramural Grants SFP 1322-530 and 1329-529 is gratefully acknowledged. Florent Jouy and Scarlett Gebauer acknowledge network funding through Helmholtz Interdisciplinary Graduate School for Environmental Research (HIGRADE). We also acknowledge the ProVis platform in general and in particular Benjamin Scheer for excellent technical support.

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Author notes

  1. Dirk K. Wissenbach

    Present address: Institute of Forensic Medicine, Jena University Hospital, 07743, Jena, Germany

  2. Stefan Kalkhof

    Present address: Department of Bioanalytics, University of Applied Sciences and Arts of Coburg, 96450, Coburg, Germany

Authors and Affiliations

  1. Department of Chemical and Product Safety, German Federal Institute for Risk Assessment (BfR), Max-Dohrn-Strasse 8-10, 10589, Berlin, Germany

    Franziska Wewering, Ralph Pirow, Andreas Luch & Sebastian Zellmer

  2. Department of Proteomics, UFZ, Helmholtz-Centre for Environmental Research, Permoserstrasse 15, 04318, Leipzig, Germany

    Florent Jouy, Martin von Bergen & Stefan Kalkhof

  3. Department of Metabolomics, UFZ, Helmholtz-Centre for Environmental Research, Permoserstrasse 15, 04318, Leipzig, Germany

    Dirk K. Wissenbach, Scarlett Gebauer & Martin von Bergen

  4. Clinic for Endocrinology and Nephrology, Faculty of Medicine, University of Leipzig, Liebigstrasse 20, 04103, Leipzig, Germany

    Matthias Blüher

  5. Institute of Biochemistry, Faculty of Medicine, University of Leipzig, Johannisallee 30, 04103, Leipzig, Germany

    Rolf Gebhardt

  6. Department of Life Sciences and Chemistry, Centre for Microbial Communities, University of Aalborg, Sohngaardsholmsvej 49, 9000, Aalborg, Denmark

    Martin von Bergen

  7. Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, University of Leipzig, Brüderstrasse 34, 04103, Leipzig, Germany

    Martin von Bergen

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  1. Franziska Wewering
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Correspondence to Franziska Wewering.

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Franziska Wewering, Florent Jouy and Dirk K. Wissenbach have contributed equally to this work.

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Wewering, F., Jouy, F., Wissenbach, D.K. et al. Characterization of chemical-induced sterile inflammation in vitro: application of the model compound ketoconazole in a human hepatic co-culture system. Arch Toxicol 91, 799–810 (2017). https://doi.org/10.1007/s00204-016-1686-y

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