Abstract
Background Neutrophils often fail to clear a single infection source such as pneumonia, but the effect of successive infective insults is unknown.
Aim To investigate our hypothesis that neutrophil exposure to a single phagocytic stimulus or ‘primary meal’ impairs subsequent killing of a ‘secondary meal’ of live bacteria.
Methods Healthy volunteer neutrophils were exposed to different primary stimuli: killed Staphylococcus aureus (SA), zymosan, live SA or Pseudomonas aeruginosa (PA), followed by a secondary stimulus of either live SA or PA. Phagocytosis and neutrophil viability were measured using flow cytometry; bacterial killing by serial dilution, plating and colony counting.
Results Exposure to killed SA (at multiplicity of infection, MOI≥20:1) or zymosan (MOI≥4:1) impaired subsequent killing of both live SA and PA (both MOI∼10:1) in suspension (p<0.05, n=5), but not in adherent neutrophils (p=0.2, n=6). Neutrophils in suspension exposed to live SA or PA (MOI∼10:1) showed impaired subsequent killing of PA and SA (MOI∼10:1), respectively (p≤0.05, n=5). Neither a 2-hour recovery period between stimuli nor granulocyte-macrophage colony stimulating factor improved killing. Neutrophil apoptosis was ∼5% after both inert or live primary stimuli but the necrosis rate rose to ∼40% after PA exposure, ∼20% after SA, and only ∼10% after inert stimuli (n=4).
Conclusions Our study supports our hypothesis that neutrophils lose killing efficiency after a single exposure to a phagocytic stimulus, in part due to neutrophil cell death. Potential additional factors, including competition for phagocytic receptors, excessive degranulation and oxidative burst, require further study.
- © 2013 ERS