Oxidative stress in fish cells:In vitro studies

Bluegill sunfish BF-2 fibroblasts were used in the neutral red (NR) cytotoxicity assay to discern the toxicities of hydrogen peroxide (H₂O₂) and paraquat as indicated by their abilities to induce oxidative stress. The toxicity of H₂O₂ was markedly enhanced in BF-2 cells treated with the glutathione depleting agents, buthionine sulfoximine (BSO), maleic acid, and chlorodinitrobenzene; similar treatments did not sensitize the BF-2 cells to paraquat, a redox cycling xenobiotic. BSO treated BF-2 cells, however, were sensitized to nitrofurantoin, also a redox cycling chemical. Diethyldithiocarbamate, an ihibitor of superoxide dismutase, only weakly enhanced the sensitivity of the BF-2 cells to H₂O₂ and paraquat. 1,10-Phenanthroline, a chelator of Fe²⁺, reduced the cytotoxicity of H₂O₂ and paraquat, presumably by preventing hydroxyl radical formation in the Fenton reaction. Quin 2 AM, an intracellular chelator of Ca²⁺, markedly lessened the toxicity of H₂O₂, but not of paraquat; EGTA, an extracellular chelator of Ca²⁺, had no effect on the toxicity of H₂O₂ or paraquat. Apparently, perturbation of intracellular Ca²⁺ homeostasis is involved in H₂O₂ toxicity. For comparative purposes, some studies were performed with fathead minnow FHM epithelioid cells, BALB/c mouse 3T3 fibroblasts, and human HepG2 hepatoma cells. The BF-2 fibroblast/NR cytotoxicity red assay was shown to be a suitable model to study oxidative stress in fish.