Antioxidants preserve macrophage phagocytosis of Pseudomonas aeruginosa during hyperoxia

Abstract

Pseudomonas. aeruginosa (PA) is a leading cause of nosocomial pneumonia in patients receiving mechanical ventilation with hyperoxia. Exposure to supraphysiological concentrations of reactive oxygen species during hyperoxia may result in macrophage damage that reduces their ability to phagocytose PA. We tested this hypothesis in cultured macrophage-like RAW 264.7 cells and alveolar macrophages from mice exposed to hyperoxia. Exposure to hyperoxia induced a similarly impaired phagocytosis of both the mucoid and non-mucoid forms of PA in alveolar macrophages and RAW cells. Compromised PA phagocytosis was associated with cytoskeleton disorganization and actin oxidation in hyperoxic macrophages. To test whether moderate concentrations of O 2 limit the loss of phagocytic function induced by ≥ 95% O 2 , mice and RAW cells were exposed to 65% O 2 . Interestingly, although the resulting lung injury/cell proliferation was not significant, exposure to 65% O 2 resulted in a marked reduction in PA phagocytosis that was comparable to that of ≥95% O 2 . Treatment with antioxidants, even post hyperoxic exposure, preserved actin cytoskeleton organization and phagocytosis of PA. These data suggest that hyperoxia reduces macrophage phagocytosis through effects on actin functions which can be preserved by antioxidant treatment. In addition, administration of moderate rather than higher concentrations of O 2 does not improve macrophage phagocytosis of PA.

Extracellular Superoxide Dismutase in Macrophages Augments Bacterial Killing by Promoting Phagocytosis

“…39 A more recent study found that hyperoxia reduces macrophage phagocytosis and that exogenous SOD treatment preserved actin cytoskeleton organization and phagocytosis of Pseudomonas aeruginosa. 40 These findings suggest that decreased superoxide scavenging may directly contribute to the defective phagocytosis observed in these cells. The present study further supports these previous studies in observing that regulation of oxidant production is important in promoting macrophage function.…”

Hydrogen peroxide enhances phagocytosis ofPseudomonas aeruginosain hyperoxia

“…Exposure to hyperoxia was performed in sealed humidified modular incubator chambers (BillupsRothenberg Inc., Del Mar, CA) containing 95% O 2 /5% CO 2 at 37°C, as described previously (Morrow et al, 2007). H 2 O 2 (30%) was purchased from VWR International (West Chester, PA).…”

“…The phagocytosis assay was performed as described with minor modifications (Baleeiro et al, 2003;Morrow et al, 2007). In brief, following each specific exposure, RAW 264.7 cells were treated with various concentrations of H 2 O 2 , ranging from 0 to 1000 µM for 1 h, and then incubated with PAO1 or fluorescein isothiocyanate (FITC)-labeled latex beads (Polysciences, Warrington, PA) at 100:1 [particles (beads or bacteria)/cell] at 37°C for 1 h. The macrophages were then washed with cold phosphate-buffered saline (PBS) containing 0.02% EDTA to remove the unbound particles and then treated with 0.04% trypan blue to quench the fluorescence attributable to any extracellular adherent particles.…”

“…Alveolar macrophages (AMs) are essential in lung innate immunity and play a critical role in the clearance of invading pathogens. Studies including ours indicate that AMs isolated from hyperoxic mice and cultured macrophages exposed to hyperoxia exhibit impaired ability in chemotaxis, adherence, phagocytosis, and pathogenkilling (Raffin et al, 1980;Rister, 1982;Suttorp and Simon, 1983;Crowell et al, 1995;Morrow et al, 2007). Excessive amounts of reactive oxygen species (ROS) have been shown to mediate pathogenesis of various pulmonary diseases and tissue injury (Ciencewicki et al, 2008).…”

“…Excessive amounts of reactive oxygen species (ROS) have been shown to mediate pathogenesis of various pulmonary diseases and tissue injury (Ciencewicki et al, 2008). Toxic levels of ROS generated under hyperoxic conditions can compromise phagocytic function of AMs (O'Reilly et al, 2003;Morrow et al, 2007). Besides superoxide, hydrogen peroxide (H 2 O 2 ) is another form of ROS that is generated under hyperoxic conditions (Turrens et al, 1982).…”