2,3,7,8‐Tetrachlorodibenzo‐ p ‐dioxin induced cytochrome P450s alter the formation of reactive oxygen species in liver cells

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin (TCDD) was classified by the International Agency for Research on Cancer as a carcinogen in humans. It acts through an aryl hydrocarbon receptor‐mediated mechanism, inducing the transcription of numerous genes, including various cytochrome P450s (CYPs – CYP1A1, 1A2, 1B1). Induction of CYPs may lead to genotoxicity by generating reactive oxygen species (ROS) which can damage DNA directly and/or via the generation of reactive metabolites. We determined ROS formation with the 2′,7′‐dihydrodichlorofluorescein diacetate fluorescence assay after incubation of HepG2 hepatoma cells or primary rat hepatocytes with TCDD. The amount of 8‐oxo‐2′‐deoxyguanosine (8‐oxo‐dG) in DNA was measured using HPLC‐MS/MS, the amount of CYP1A1 protein by Western blotting. The catalytic activity of CYP1A enzymes was determined as 7‐ethoxyresorufin‐ O ‐deethylase (EROD) activity. Incubation of cells with TCDD for 48 h caused increased levels of ROS in primary rat hepatocytes as well as increased levels of 8‐oxo‐dG in DNA compared to untreated cells. In the HepG2 cell line no significant effects were observed for both ROS formation and 8‐oxo‐dG levels. Both effects were in good agreement with the extent of induction of CYP1A1 protein and EROD activity, suggesting that CYP1 induction is a major source of ROS formation in TCDD‐treated hepatocytes.