Neutrophil elastase correlates with increased sphingolipid content in cystic fibrosis sputum.

Abstract

Sphingolipids are associated with the regulation of pulmonary inflammation. Although sphingolipids have been investigated in the context of cystic fibrosis (CF), the focus has been on loss of CF transmembrane conductance regulator (CFTR) function in mice, and in CF human lung epithelial cell lines. The sphingolipid content of CF sputum and the potential link between ceramide and airway inflammation in CF remain relatively unexplored. Fifteen patients with CF provided two spontaneously expectorated sputum samples, one collected during a hospitalization for an acute pulmonary exacerbation and one from an outpatient visit at a time of clinical stability. Sputum was processed, and the supernatant assessed for active neutrophil elastase (NE) using a chromogenic microplate assay and sphingolipid content using reverse phase high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). Relevant demographic data including age, sex, CF genotype, FEV1 % predicted, and sputum bacteriology were assessed as possible modifying factors that could influence the correlation between NE and sputum sphingolipids. Data were analyzed for linear correlation, with statistical significance pre-defined as P < 0.05. There was a significant association between the concentration of active NE and ceramide, sphingomyelin, and monohexosylceramide moieties as well as sphingosine-1-phosphate. The presence of Methicillin-resistant Staphylococcus aureus (MRSA), FEV1 % predicted, and female gender further strengthened the association of NE and sphingolipids, but Pseudomonas aeruginosa had no effect on the association between NE and sphingolipids. These data suggest that NE may increase pro-inflammatory sphingolipid signaling, and the association is strengthened in female patients and patients with MRSA.