Hydrogen Peroxide Release from Alveolar Macrophages and Alveolar Type II Cells During Adaptation to Hyperoxia in Vivo

Abstract

The effect of hyperoxia (1–14 days, 85% O₂) on rat alveolar macrophage and alveolar type II cell oxidant and antioxidant characteristics was investigated. Unstimulated control macrophages (2 h ex vivo) released hydrogen peroxide at a rate of 3.5 ± 1.3 nmol/min mg protein⁻¹ which was a cyanide-sensitive process. H₂O₂ release from alveolar macrophages decreased slightly but not significantly after 1 day in hyperoxia and increased significantly after 3 days (180%, p < .05) and 14 days (380%, p < .01). When H₂O₂ release was expressed as nmol from total macrophages per animal, the increase after 14 days in hyperoxia was 760%. H₂O₂ generation by hyperoxic macrophages was cyanide resistant, indicating the involvement of active NADPH oxidase. In both control and hyperoxic macrophages H₂O₂ release could be significantly stimulated with phorbol myristate acetate (PMA). Comparisons of H₂O₂ release by freshly isolated alveolar macrophages and alveolar type II cells must be cautiously interpreted because some cell functions may change during the isolation procedure. Freshly isolated (6 h ex vivo) control alveolar type II cells were found to generate H₂O₂ at a rate of 0.26 ± 0.05 nmol/min mg protein⁻¹. In type II cells H₂O₂ release, calculated as nmol/mg protein, decreased during the first 7 days of hyperoxia to 10% (p < .01) of the control value and then returned back up to the control level after 14 days. A similar decrease was observed if H₂O₂ release was calculated as nmol/cell number. H₂O₂ release from control and hyperoxic type II cells was cyanide sensitive. The decrease in H₂O₂ release in type II cells was associated with cell membrane injury (as assessed by electron microscopy), while biochemical markers of cellular injury (trypan blue exclusion and cellular high-energy phosphates ATP, ADP) were unchanged. The ability of type II cells to scavenge extracellular H₂O₂ did not change in acute hyperoxia, but it increased significantly during the second week in hyperoxia. These results indicate that macrophages but not type II cells are stimulated to produce H₂O₂ during prolonged exposure to hyperoxia