Oxidative stress and genotoxicity of co-exposure to chlorpyrifos and aflatoxin B 1 in HepG2 cells

Chlorpyrifos (CPF) and aflatoxin B₁ (AFB₁) are each known to adversely affect hepatic tissue individually, but their combined hepatic effects have never been previously investigated. HepG2 cell viability, oxidative status, and genetic impairment were examined after exposing HepG2 cells to: (1) CPF alone, (2) AFB₁ alone, and (3) CPF and AFB₁ combined (20:1). CPF exposure decreased cell viability, reduced glutathione (GSH) content, and superoxide dismutase (SOD) activity but increased both glutathione peroxidase (GPx) and paraoxonase 1 activity. AFB₁ exposure decreased cell viability and GSH content but increased reactive oxygen species (ROS) production. CPF and AFB₁ combined exposure decreased GSH content (p < 0.05) further over individual CPF and AFB₁ exposures. Induction of micronucleus formation was detected in AFB₁-treated cells but undetected in both CPF and combination-treated cells. In conclusion, cytotoxic effects caused by combined exposure were antagonistic, as shown by a combination index value of 1.67. Although no change in ROS production was observed in CPF groups, the overall results confirmed the occurrence of oxidative stress through the alterations of GSH content, GPx, and SOD activity. Only intracellular GSH was evidently changed upon exposure to CPF and AFB₁ combined. Thus, this study suggested cellular GSH as a potential indicator for detecting the combined effects of CPF and AFB₁ in HepG2 cells, the detection of which could be adapted to estimate the potential toxicity of additional multiple toxicant exposures.