Participation of active oxygen species in 6-hydroxydopamine toxicity to a human neuroblastoma cell line

Abstract

Catalase, Superoxide dismutase, and dimethylsulfoxide were tested for their ability to prevent the cytotoxic effect of 6-hydroxydopamine (6-OHDA) on the human neuroblastoma line SY5Y. Viability was measured at two time points after 6-OHDA treatment: at 3 hr by means of amino acid incorporation and at 24 hr by trypan blue dye exclusion. Survival of cells treated concomitantly with catalase (50 μg/ml) and 6-OHDA was at least 90 per cent that of untreated controls. Cells receiving 6-OHDA alone showed less than 30 per cent survival relative to untreated controls. Superoxide dismutase (50 μg/ml) temporarily protected cells from a high concentration of 6-OHDA. Dimethylsulfoxide treatment increased survival from the control level 24 hr after treatment with 6-OHDA. Two other cell lines (A 1B 1 human glial cells and CHO fibroblasts) had intermediate and high resistance to the drug, respectively, compared to the low resistance of SY5Y cells. CHO and SY5Y cells had similar responses to 6-OHDA and to H 2O 2 when tested at twice the molarity of 6-OHDA. Specific activities of three enzymes known to detoxify H 2O 2 or H 2O 2-generated organic hydroperoxides (catalase, glutathione S-transferase, and glutathione peroxidase) were compared in the three cell lines. Catalase activity was 2.5 times as high in A 1B 1 and CHO cells as in SY5Y cells when expressed as units/mg protein and 7 times as high in units/culture dish. Other enzyme activities showed no correlation to 6-OHDA resistance.