Pulmonary expression of granulocyte/macrophage colony-stimulating factor (GM-CSF) is critically important for normal functional maturation of alveolar macrophages. We found previously that lung GM-CSF is dramatically suppressed in mice exposed to hyperoxia. Alveolar epithelial cells (AEC) are a major source of GM-CSF in the peripheral lung, and in vivo hyperoxia resulted in greatly reduced expression of GM-CSF protein by AEC ex vivo. We now explore the mechanisms responsible for this effect, using primary cultures of murine AEC exposed to hyperoxia in vitro. Exposure of AEC to 80% oxygen/5% CO₂ for 48 h did not induce overt toxicity, but resulted in significantly decreased GM-CSF protein and mRNA expression compared with cells in normoxia. Similar effects were seen when AEC were stressed with serum deprivation, an alternative inducer of oxidative stress. The effects in AEC were opposite those in a murine lung epithelial cell line (MLE-12 cells), in which hyperoxia induced GM-CSF expression. Both hyperoxia and serum deprivation resulted in increased intracellular reactive oxygen species (ROS) in AEC. Hyperoxia and serum deprivation induced significantly accelerated turnover of GM-CSF mRNA. Treatment of AEC with catalase during oxidative stress preserved GM-CSF protein and mRNA and was associated with stabilization of GM-CSF mRNA. We conclude that hyperoxia-induced suppression of AEC GM-CSF expression is a function of ROS-induced destabilization of GM-CSF mRNA. We speculate that AEC oxidative stress results in significantly impaired pulmonary innate immune defense due to effects on local GM-CSF expression in the lung.