Archives of Toxicology

, Volume 88, Issue 7, pp 1427–1437 | Cite as

Critical differences in toxicity mechanisms in induced pluripotent stem cell-derived hepatocytes, hepatic cell lines and primary hepatocytes

  • Anna-Karin M. Sjogren
  • Maria Liljevald
  • Björn Glinghammar
  • Johanna Sagemark
  • Xue-Qing Li
  • Anna Jonebring
  • Ian Cotgreave
  • Gabriella Brolén
  • Tommy B. Andersson
Molecular Toxicology

Abstract

Human-induced pluripotent stem cell-derived hepatocytes (hiPSC-Hep) hold great potential as an unlimited cell source for toxicity testing in drug discovery research. However, little is known about mechanisms of compound toxicity in hiPSC-Hep. In this study, modified mRNA was used to reprogram foreskin fibroblasts into hiPSC that were differentiated into hiPSC-Hep. The hiPSC-Hep expressed characteristic hepatic proteins and exhibited cytochrome P450 (CYP) enzyme activities. Next, the hiPSC-Hep, primary cryopreserved human hepatocytes (cryo-hHep) and the hepatic cell lines HepaRG and Huh7 were treated with staurosporine and acetaminophen, and the toxic responses were compared. In addition, the expression of genes regulating and executing apoptosis was analyzed in the different cell types. Staurosporine, an inducer of apoptosis, decreased ATP levels and activated caspases 3 and 7 in all cell types, but to less extent in Huh7. Furthermore, a hierarchical clustering and a principal component analysis (PCA) of the expression of apoptosis-associated genes separated cryo-hHep from the other cell types, while an enrichment analysis of apoptotic pathways identified hiPSC-Hep as more similar to cryo-hHep than the hepatic cell lines. Finally, acetaminophen induced apoptosis in hiPSC-Hep, HepaRG and Huh7, while the compound initiated a direct necrotic response in cryo-hHep. Our results indicate that for studying compounds initiating apoptosis directly hiPSC-Hep may be a good alternative to cryo-hHep. Furthermore, for compounds with more complex mechanisms of toxicity involving metabolic activation, such as acetaminophen, our data suggest that the cause of cell death depends on a balance between factors controlling death signals and the drug-metabolizing capacity.

Keywords

Hepatotoxicity Drug-induced liver injury iPS cells HepaRG Acetaminophen Apoptosis 

Abbreviations

hiPSC

Human-induced pluripotent stem cells

hiPSC-Hep

Human-induced pluripotent stem cell-derived hepatocytes

CYP

Cytochrome P450

cryo-hHep

Cryopreserved human hepatocytes

PCA

Principal component analysis

NAPQI

N-acetyl-p-benzoquinone imine

PAS

Periodic acid schiff

LC/MS

Liquid chromatography/mass spectrometry

qPCR

Quantitative real-time PCR

Ct

Cycle of threshold

GSH

Glutathione

RAM

LC-/MS-radiochemical activity monitoring

HNF4α

Hepatocyte nuclear factor 4 alpha

CK18

Cytokeratin 18

AAT

Alpha-1 antitrypsin

ALB

Albumin

PXR

Pregnane X receptor

AFP

Alpha-fetoprotein

Supplementary material

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Supplementary material 1 (PDF 69395 kb)
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Supplementary material 2 (PDF 158 kb)
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Supplementary material 3 (PDF 224 kb)
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Supplementary material 5 (PDF 190 kb)
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Supplementary material 6 (DOCX 40 kb)
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Supplementary material 9 (PDF 175 kb)
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Supplementary material 10 (PDF 470 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Anna-Karin M. Sjogren
    • 1
  • Maria Liljevald
    • 2
  • Björn Glinghammar
    • 2
  • Johanna Sagemark
    • 2
  • Xue-Qing Li
    • 1
  • Anna Jonebring
    • 3
  • Ian Cotgreave
    • 4
  • Gabriella Brolén
    • 3
  • Tommy B. Andersson
    • 1
    • 5
  1. 1.Cardiovascular and Metabolic Diseases Innovative Medicines, DMPKAstraZeneca R&DMölndalSweden
  2. 2.Drug Safety and Metabolism, Discovery SafetyAstraZeneca R&DMölndalSweden
  3. 3.Discovery Sciences, Reagents and Assay DevelopmentAstraZeneca R&DMölndalSweden
  4. 4.Unit of Biochemical Toxicology, Institute of Environmental MedicineKarolinska InstituteStockholmSweden
  5. 5.Section of Pharmacogenetics, Department of Physiology and PharmacologyKarolinska InstituteStockholmSweden

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