PCBs and PCB metabolites have been suggested to cause cytotoxicity by inducing oxidative stress, but the effectiveness of antioxidant intervention after exposure has not been established. Exponentially growing MCF-10A human breast and RWPE-1 human prostate epithelial cells continuously exposed for 5 days to 3 microM PCBs [Aroclor 1254 (Aroclor), PCB153, and the 2-(4-chlorophenyl)-1,4-benzoquinone metabolite of PCB3 (4ClBQ)] were found to exhibit growth inhibition and clonogenic cell killing, with 4ClBQ having the most pronounced effects. These PCBs were also found to increase steady-state levels of intracellular O(2)(*-) and H(2)O(2) (as determined by dihydroethidium, MitoSOX red, and 5-(and 6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate oxidation). These PCBs also caused 1.5- to 5.0-fold increases in MnSOD activity in MCF-10A cells and 2.5- to 5-fold increases in CuZnSOD activity in RWPE-1 cells. Measurement of MitoSOX red oxidation with confocal microscopy coupled with colocalization of MitoTracker green in MCF-10A and RWPE-1 cells supported the hypothesis that PCBs caused increased steady-state levels of O(2)(*-) in mitochondria. Finally, treatment with either N-acetylcysteine (NAC) or the combination of polyethylene glycol (PEG)-conjugated CuZnSOD and PEG-catalase added 1 h after PCBs significantly protected these cells from PCB toxicity. These results support the hypothesis that exposure of exponentially growing human breast and prostate epithelial cells to PCBs causes increased steady-state levels of intracellular O(2)(*-) and H(2)O(2), induction of MnSOD or CuZnSOD activity, and clonogenic cell killing that could be inhibited by a clinically relevant thiol antioxidant, NAC, as well as by catalase and superoxide dismutase after PCB exposure.