Cystic fibrosis (CF) airway disease is characterized by chronic microbial infections and infiltration of inflammatory polymorphonuclear (PMN) granulocytes. Staphylococcus aureus (S. aureus) is a major lung pathogen in CF that persists despite the presence of PMNs and has been associated with CF lung function decline. While PMNs represent the main mechanism of the immune system to kill S. aureus, it remains largely unknown why PMNs fail to eliminate S. aureus in CF. The goal of this study was to observe how the CF airway environment affects S. aureus killing by PMNs. PMNs were isolated from the blood of healthy volunteers and CF patients. Clinical isolates of S. aureus were obtained from the airways of CF patients. The results show that PMNs from healthy volunteers were able to kill all CF isolates and laboratory strains of S. aureus tested in vitro. The extent of killing varied among strains. When PMNs were pretreated with supernatants of CF sputum, S. aureus killing was significantly inhibited suggesting that the CF airway environment compromises PMN antibacterial functions. CF blood PMNs were capable of killing S. aureus. Although bacterial killing was inhibited with CF sputum, PMN binding and phagocytosis of S. aureus was not diminished. The S. aureus-induced respiratory burst and neutrophil extracellular trap release from PMNs also remained uninhibited by CF sputum. In summary, our data demonstrate that the CF airway environment limits killing of S. aureus by PMNs and provides a new in vitro experimental model to study this phenomenon and its mechanism.
Cystic fibrosis (CF) airway disease is characterized by chronic microbial infections and infiltration of polymorphonuclear neutrophil granulocytes (PMNs). In CF, PMNs are unable to clear lung pathogens resulting in lung damage. Neutrophil extracellular traps (NETs) and superoxide production represent defense mechanisms of PMNs to kill extracellular pathogens. NETs and superoxide in the CF lung, however, seem unable to clear Pseudomonas aeruginosa and Staphylococcus aureus, the two main lung pathogens in CF. Our goal of this study was to determine antimicrobial functions of PMNs to S. aureus and P. aeruginosa CF isolates and to observe how the CF airway environment influences them. Human PMNs were isolated from healthy donors. Isolates of S. aureus were obtained from adult CF patients: four strains were methicillin-resistant (MRSA), four were methicillin-sensitive S. aureus (MSSA), and ten CF isolates of P. aeruginosa. PMN effector functions were measured: killing was assessed by bacterial growth assay, NET formation was measured by Sytox fluorescence and superoxide was monitored by Diogenes chemiluminescence. Results were subjected to paired t-test, significant if p<0.05. PMNs from healthy volunteers were able to kill all CF isolates. The extent of killing varied among strains, however, P. aeruginosa was killed more efficiently by blood PMNs than S. aureus clinical isolates. The enhanced killing of P. aeruginosa is correlated with enhanced NET release and superoxide production compared to S. aureus. Killing of all isolates was inhibited by CF sputum pretreatment. Overall, the results show that CF bacterial clinical isolates stimulate PMN effector responses that are impaired by the CF airway environment.
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