In seeking more specific biomarkers of the cystic fibrosis (CF) lung inflammatory disease that would be sensitive to antibiotic therapy, we sought to evaluate the gene expression profiles of neutrophils in CF patients before treatment in comparison with non-CF healthy individuals and after antibiotic treatment. Genes involved in neutrophil-mediated inflammation, i.e. chemotaxis, respiratory burst, apoptosis, and granule exocytosis, were the targets of this study. Microarray analysis was carried out in blood and airway neutrophils from CF patients and in control subjects. A fold change (log) threshold of 1.4 and a cut-off of p,0.05 were utilized to identify significant genes. Community networks and principal component analysis were used to distinguish the groups of controls, pre- and post-therapy patients. Control subjects and CF patients before therapy were readily separated, whereas a clear distinction between patients before and after antibiotic therapy was not possible. Blood neutrophils before therapy presented 269 genes down-regulated and 56 up-regulated as compared with control subjects. Comparison between the same patients before and after therapy showed instead 44 genes downregulated and 72 up-regulated. Three genes appeared to be sensitive to therapy and returned to ‘‘healthy’’ condition: phorbol-12-myristate-13-acetate-induced protein 1 (PMAIP1), hydrogen voltage-gated channel 1 (HVCN1), and b-arrestin 1 (ARRB1). The up-regulation of these genes after therapy were confirmed by real time PCR. In airway neutrophils, 1029 genes were differentially expressed post- vs pre-therapy. Of these, 30 genes were up-regulated and 75 down-regulated following antibiotic treatment. However, biological plausibility determined that only down-regulated genes belonged to the gene classes studied for blood neutrophils. Finally, it was observed that commonly expressed genes showed a greater variability in airway neutrophils than that found in blood neutrophils, both before and after therapy. These results indicate more specific targets for future interventions in CF patients involving respiratory burst, apoptosis, and granule exocytosis.

BACKGROUND: It is not known whether antibiotic therapy for lung disease in cystic fibrosis (CF) has an influence on circulating polymorphonuclear neutrophil (PMN) function and apoptosis. PATIENTS AND METHODS: Blood PMNs were obtained from 14 CF patients before and after antibiotic treatment for an acute exacerbation, and from 10 healthy controls. PMNs were evaluated for production of reactive oxygen species (ROS) by spectrophotometry, of cytokines in the conditioned medium by ELISA, and apoptotic response by cytofluorimetry. RESULTS: ROS and interleukin (IL)-8 were produced at higher levels by CF PMNs pre-therapy than control PMNs under basal conditions. IL-8 levels further increased after therapy. Early apoptotic response was higher in CF PMNs pre-therapy than in control PMNs, and this pattern did not change after antibiotic treatment. CONCLUSIONS: Circulating PMNs are primed in CF acute patients. Further studies are needed to consider PMN-produced IL-8 as a biomarker to evaluate response to antibiotic therapy in CF patients.

The gram-positive pathogen Streptococcus pneumoniae is the most common cause of community-acquired pneumonia and is responsible for high morbidity and mortality worldwide. A major feature of pneumococcal pneumonia is an abundant neutrophil infiltration. In this work we observed that the R6 nonencapsulated S. pneumoniae strain induced a higher oxidative burst in neutrophils compared with its capsulated progenitor D39, by triggering neutrophil NADPH oxidase to produce more reactive oxygen intermediates (ROI) and by interfering with the neutrophil kinase signalling pathway. In addition, we evaluated the possibility that the capsule, lacking in R6 but present in D39, could modulate the S. pneumoniaeinduced neutrophil respiratory burst. In this respect, three knock-out isogenic mutants (D39DCPS2E, D39DCPS-R6 and R6DCPS-R6) that were unable to synthesize the capsule, were tested for their capability of inducing the release of neutrophil-ROIs. The results indicate that the mutants behaved similarly to their wild-type parental strains in enhancing respiratory burst activity, suggesting that the capsule itself is not directly involved in modulating the neutrophil oxidative burst induced by S. pneumoniae, but that other genetic differences between D39 and R6 present elsewhere in the genome could be responsible for these mechanisms.