Recovery of redox homeostasis altered by CuNPs in H4IIE liver cells does not reduce the cytotoxic effects of these NPs: an investigation using aryl hydrocarbon receptor (AhR) dependent antioxidant activity

Chem Biol Interact. 2015 Feb 25;228:57-68. doi: 10.1016/j.cbi.2015.01.012. Epub 2015 Jan 21.


The generation of reactive oxygen species (ROS) and consequent oxidative stress is regarded as a relevant mechanism for nanoparticle toxicity. In cells, the activation of the aryl hydrocarbon receptor (AhR) triggers a cascade of defence responses against oxidative stress. By increasing AhR dependent cellular anti-oxidant activity, we tested the extent to which the cytotoxic effect of copper nanoparticles (CuNPs) is governed by oxidative stress. H4IIE rat hepatoma cells were challenged with high ROS levels after exposure to CuNPs, while the AhR-induced cellular anti-oxidant defence was simultaneously activated by the AhR ligand beta-Naphthoflavone (ßNF). Activation of phase II detoxification enzymes (as glutathione-S-transferases, GSTs) and anti-oxidants (glutathione, GSH) led to a complete abrogation of CuNP-induced ROS production. However, a concurrent reduction in cytotoxicity was not detected, thereby indicating that CuNPs exert non-oxidative stress mediated cytotoxic effects. Transmission electron microscopy analysis pointed to a direct physical perturbation of cellular structures by CuNPs, thus contributing to their cytotoxicity. Our observations highlight that distinct mechanisms underlie the toxicity of ions and NPs and indicate that while ROS elicitation is CuNP-specific, the cytotoxic action of these particles may not be directly related to their pro-oxidative activity. These findings have important implications with respect to the oxidative stress paradigm used to explain NP toxicity.

Keywords: Aryl hydrocarbon receptor (AhR); Co-incubation; Copper nanoparticles (CuNPs); H4IIE; Oxidative stress; ß-Naphthoflavone (ßNF).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antioxidants / metabolism
  • Antioxidants / pharmacology*
  • Cell Survival / drug effects
  • Copper / chemistry
  • Copper / pharmacology
  • Copper / toxicity*
  • Hepatocytes / cytology*
  • Hepatocytes / drug effects*
  • Hepatocytes / metabolism
  • Homeostasis / drug effects*
  • Metal Nanoparticles / chemistry*
  • Oxidation-Reduction / drug effects
  • Oxidative Stress / drug effects
  • Rats
  • Reactive Oxygen Species / metabolism
  • Receptors, Aryl Hydrocarbon / metabolism*
  • Tumor Cells, Cultured


  • Antioxidants
  • Reactive Oxygen Species
  • Receptors, Aryl Hydrocarbon
  • Copper