Increased mitochondrial ROS formation by acetaminophen in human hepatic cells is associated with gene expression changes suggesting disruption of the mitochondrial electron transport chain

Toxicol Lett. 2015 Apr 16;234(2):139-50. doi: 10.1016/j.toxlet.2015.02.012. Epub 2015 Feb 19.


Acetaminophen (APAP) overdosage results in hepatotoxicity, but the underlying molecular mechanisms are still not completely understood. In the current study, we focused on mitochondrial-specific oxidative liver injury induced by APAP exposure. Owning to genetic polymorphisms in the CYP2E1 gene or varying inducibility by xenobiotics, the CYP2E1 mRNA level and protein activity vary extensively among individuals. As CYP2E1 is a known ROS generating enzyme, we chose HepG2 to minimize CYP2E1-induced ROS formation, which will help us better understand the APAP induced mitochondrial-specific hepatotoxicity in a subpopulation with low CYP2E1 activity. HepG2 cells were exposed to a low and toxic dose (0.5 and 10mM) of APAP and analyzed at four time points for genome-wide gene expression. Mitochondria were isolated and electron spin resonance spectroscopy was performed to measure the formation of mitochondrial ROS. The yield of ATP was measured to confirm the impact of the toxic dose of APAP on cellular energy production. Our results indicate that 10mM APAP significantly influences the expression of mitochondrial protein-encoding genes in association with an increase in mitochondrial ROS formation. Additionally, 10mM APAP affects the expression of genes encoding the subunits of electron transport chain (ETC) complexes, which may alter normal mitochondrial functions by disrupting the assembly, stability, and structural integrity of ETC complexes, leading to a measurable depletion of ATP, and cell death. The expression of mitochondrium-specific antioxidant enzyme, SOD2, is reduced which may limit the ROS scavenging ability and cause imbalance of the mitochondrial ROS homeostasis. Overall, transcriptome analysis reveals the molecular processes involved in the observed APAP-induced increase of mitochondrial ROS formation and the associated APAP-induced oxidative stress.

Keywords: Acetaminophen; ESR; HepG2; Microarray; Mitochondrial ROS; Toxicogenomics.

MeSH terms

  • Acetaminophen / toxicity*
  • Adenosine Triphosphate / metabolism
  • Cell Death / drug effects
  • Chemical and Drug Induced Liver Injury / enzymology
  • Chemical and Drug Induced Liver Injury / etiology*
  • Chemical and Drug Induced Liver Injury / genetics
  • Chemical and Drug Induced Liver Injury / pathology
  • Dose-Response Relationship, Drug
  • Electron Transport Chain Complex Proteins / genetics
  • Electron Transport Chain Complex Proteins / metabolism*
  • Energy Metabolism / drug effects
  • Gene Expression Profiling / methods
  • Gene Expression Regulation
  • Hep G2 Cells
  • Hepatocytes / drug effects*
  • Hepatocytes / metabolism
  • Hepatocytes / pathology
  • Humans
  • Mitochondria, Liver / drug effects*
  • Mitochondria, Liver / metabolism
  • Mitochondria, Liver / pathology
  • Oxidative Stress / drug effects*
  • Reactive Oxygen Species / metabolism*
  • Superoxide Dismutase / genetics
  • Superoxide Dismutase / metabolism
  • Time Factors
  • Toxicogenetics / methods


  • Electron Transport Chain Complex Proteins
  • Reactive Oxygen Species
  • Acetaminophen
  • Adenosine Triphosphate
  • Superoxide Dismutase
  • superoxide dismutase 2