Neutrophils from Cystic Fibrosis Patients Are Defective in Killing of Phagocytosed Pseudomonas aeruginosa.

Cystic fibrosis (CF), the most common genetic disease in Caucasians, is caused by mutations of the gene encoding the CF transmembrane conductance regulator (CFTR), a cAMP-regulated chloride channel. CF has long been recognized as an epithelial disease whose most severe complications often occur in the lung. The clinical manifestations include persistent bacterial infection, prominent neutrophil infiltration and small airway obstruction. Even though dramatic advances have been made towards understanding of CF pathogenesis, the link between the CFTR chloride channel defect and a clinical defect in bacterial eradication has not been fully established. Our published data demonstrated that CFTR is expressed in human neutrophils and their phagolysosomes. CF neutrophils are defective in the chlorination of phagocytosed Pseudomonas aeruginosa (PAO1), indicating defective intraphagolysosomal hypochlorous acid (HOCl) production. In the current report, we assessed the bacterial killing abilities of neutrophils from CF and normal individuals. Percoll-purified peripheral blood neutrophils were incubated with opsonized PAO1 at a ratio of 1:1 or 1:50. To define the role of chloride in the killing process, two different Ringer’s buffers with either 0 mM chloride or 135 mM chloride were exploited. At various time points (0, 15, 30 and 60 minutes) after incubation of neutrophils with bacteria, samples were aliquoted to assay for viable bacteria. After correction for bacterial growth over the experimental period, the bacterial viability at each time point relative to that of the initial value was obtained. Two-way ANOVA tests indicated that CF neutrophils had a significantly lower initial rate of killing of PAO1 than that in normal controls. This defect is more pronounced under the condition of high bacterial load and low extracellular chloride. Surprisingly, the low extracellular chloride significantly affected neutrophil-mediated bacterial killing even for the normal neutrophils, suggesting the dependence of this ion in the killing process. Our data provide evidence to suggest the potential role of CFTR in supplying intraphagolysosomal chloride to produce hypochlorous acid (HOCl), an oxidant essential for the killing of HOCl-sensitive bacteria.