Although the effect of
hyperoxia on
antioxidant enzymes is well known, the effect of subtoxic levels of
hyperoxia on
gamma-glutamyltransferase (gamma-GT), involved in the degradation and uptake of extracellular GSH for intracellular GSH synthesis, is unknown. The aim of the study was to investigate (1) the effects of in vitro
hyperoxia on gamma-GT activity of type II cells and (2) the effects of the lazaroid
U-74389G and
N-acetylcysteine (NAC) on the
hyperoxia-induced changes in gamma-GT and
antioxidant enzyme activities. At 48 h after isolation, rat type II cells were exposed for 2 days to air, 60% O2 or 85% O2 with or without 30 microM
U-74389G or 100 microM NAC. After the exposure, the cells were harvested and assayed for
superoxide dismutase (SOD),
glutathione peroxidase (GPx), gamma-GT activity, and GSH levels. In another series of experiments 85% O2-exposed cells, with or without
U-74389G, were used for Northern blotting of gamma-GT
mRNA. Exposure to 60% O2 decreased gamma-GT and GSH by -47 and -34%, respectively, while SOD and GPx activities remained unchanged. After 85% O2-exposure gamma-GT decreased by -55%, SOD and GPx increased by +55 and +87%, respectively, while GSH decreased by -35%. NAC treatment decreased gamma-GT activity by -42% in the air-exposed cells. After 60% O2,
U-74389G led to significantly higher gamma-GT (+117%) and GSH (+26%) while NAC only led to higher GSH (+28%) compared to the
oxygen-exposed cells not treated with NAC or
U-74389G. After 85% O2
U-74389G increased gamma-GT, SOD, and GSH by +72, +58, and +68%, respectively, while NAC only increased SOD (+49%) and GSH (+26%) compared to the
oxygen-exposed cells not treated with NAC or
U-74389G. The 85% O2 exposure, with or without
U-74389G, had no effect on gamma-GT
mRNA levels. The results show that
hyperoxia decreases rat type II cell gamma-GT activity in vitro. This effect was not related to an altered regulation at
mRNA level and it was not associated with the
hyperoxia-induced decrease in intracellular GSH, since restoration of the GSH levels by NAC did not restore gamma-GT activity. The lazaroid
U-74389G with
vitamin E-like properties effectively prevented the decrease in gamma-GT and GSH, so that direct inactivation of the membrane-bound gamma-GT by
hyperoxia is the most likely mechanism.