Effects of Human Neutrophil Elastase and Pseudomonas aeruginosa Proteinases on Human Respiratory Epithelium

Abstract

It has been suggested that proteinase enzymes could play an important role in the pathogenesis of chronic bronchial infections including bronchiectasis and cystic fibrosis (CF). Because Pseudomonas aeruginosa frequently colonizes the respiratory tract in bronchiectasis and CF, we examined the in vitro effects of human neutrophil elastase (HNE) and proteinase enzymes produced by P. aeruginosa (elastase: PE; alkaline proteinase: PAP) on the ciliary beat frequency (CBF) and ultrastructure of human nasal ciliated respiratory epithelium. HNE (500 micrograms/ml) progressively reduced CBF and caused marked epithelial disruption; lower concentrations (100 and 20 micrograms/ml) also caused epithelial disruption but without slowing CBF. The effects of HNE (500 micrograms/ml) were completely abolished by adding alpha 1-antitrypsin (5 mg/ml). There was no synergy between HNE and pyocyanin, a product of P. aeruginosa which slows CBF. PE in phosphate-buffered saline also caused epithelial disruption without slowing CBF; however, PE in medium containing divalent metal ions caused CBF slowing as well as epithelial disruption at 100 micrograms/ml. PAP (500 micrograms/ml) had almost no effect on ciliated epithelium. The effects of HNE and PE on nasal and bronchial epithelium obtained from the same patient were similar. Light and transmission electron microscopy revealed that HNE and PE were cytotoxic and caused detachment of epithelial cells from neighboring cells and the basement membrane. There was cytoplasmic blebbing of the cell surface and mitochondrial damage; however, no increase of abnormalities in the ultrastructure of cilia on living cells was seen. These results support the hypothesis that HNE and PE contribute to the delayed mucociliary clearance and epithelial damage that is observed in patients with chronic bronchial infection.