Differential sensitivity of metabolically competent and non-competent HepaRG cells to apoptosis induced by diclofenac combined or not with TNF-α

Toxicol Lett. 2016 Sep 6;258:71-86. doi: 10.1016/j.toxlet.2016.06.008. Epub 2016 Jun 14.


The role of reactive metabolites and inflammatory stress has been largely evoked in idiosyncratic hepatotoxicity of diclofenac (DCF); however mechanisms remain poorly understood. We aimed to evaluate the influence of liver cell phenotype on the hepatotoxicity of DCF combined or not with TNF-α using differentiated and undifferentiated HepaRG cells, and for comparison, HepG2 cells. Our results demonstrate that after a 24h-treatment metabolizing HepaRG cells were less sensitive to DCF than their undifferentiated non-metabolizing counterparts as shown by lower oxidative and endoplasmic reticulum stress responses and lower activation of caspase 9. Differentiated HepaRG cells were also less sensitive than HepG2 cells. Their lower sensitivity to DCF was related to their high content in glutathione transferases. DCF-induced apoptotic effects were potentiated by TNF-α only in death receptor-expressing differentiated HepaRG and HepG2 cells and were associated with marked activation of caspase 8. TNF-α co-treatment did not aggravate DCF-induced cholestatic features. Altogether, our results demonstrate that (i) lower sensitivity to DCF of differentiated HepaRG cells compared to their non-metabolically active counterparts was related to their high detoxifying capacity, giving support to the higher sensitivity of nonhepatic tissues than liver to this drug; (ii) TNF-α-potentiation of DCF cytotoxicity occurred only in death receptor-expressing cells.

Keywords: Caspases; Cholestasis; Differentiation status; Drug metabolism; Endoplasmic reticulum stress; Glutathione transferases; HepG2 cells; Primary human hepatocytes; Reactive oxygen species; Tumor necrosis factor α.

Publication types

  • Comparative Study

MeSH terms

  • Anti-Inflammatory Agents, Non-Steroidal / agonists
  • Anti-Inflammatory Agents, Non-Steroidal / metabolism
  • Anti-Inflammatory Agents, Non-Steroidal / pharmacology*
  • Apoptosis / drug effects*
  • Biotransformation / drug effects
  • Cell Differentiation
  • Cell Line
  • Cells, Cultured
  • Diclofenac / agonists
  • Diclofenac / metabolism
  • Diclofenac / pharmacology*
  • Drug Resistance*
  • Endoplasmic Reticulum Stress / drug effects
  • Enzyme Inhibitors
  • Glutathione Transferase / antagonists & inhibitors
  • Glutathione Transferase / metabolism
  • Hep G2 Cells
  • Hepatocytes / cytology
  • Hepatocytes / drug effects*
  • Hepatocytes / metabolism
  • Humans
  • Inhibitory Concentration 50
  • Kinetics
  • Microscopy, Phase-Contrast
  • Oxidative Stress / drug effects
  • Tumor Necrosis Factor-alpha / metabolism*
  • fas Receptor / agonists*
  • fas Receptor / metabolism


  • Anti-Inflammatory Agents, Non-Steroidal
  • Enzyme Inhibitors
  • Tumor Necrosis Factor-alpha
  • fas Receptor
  • Diclofenac
  • Glutathione Transferase