Neutrophil elastase contributes to activated human neutrophil-induced rat small airway dysfunction

Abstract

Small airways (SA) are the main site of airflow obstruction in COPD. Neutrophil products, including proteases and oxidants, damage lung tissue and are implicated in COPD pathophysiology. We studied the effects of protease inhibitors or an antioxidant ( N -acetylcysteine, NAC) on COPD neutrophil supernatant-induced SA dysfunction in ex vivo rat precision-cut lung slices (PCLS) using videomicroscopy. fMLP-stimulated neutrophil supernatants were pre-treated with either neutrophil elastase (NE) inhibitor (10μM, Calbiochem 324745) or MMP-9 inhibitor (10μM, Calbiochem 444278), or NAC (1mM) prior to overnight incubation with rat PCLS, and SA contraction to carbachol (CCh, 10nM-10μM) assessed. Compared with untreated (UT) airways, there was a 33% reduction in maximal airway closure in SA incubated with COPD neutrophil supernatants (UT 86±2, n=10; COPD 53±9% closure, n=7), which was attenuated by 24% by NE inhibitor (77±8%, n=7), but with no effect on PCLS viability (COPD 42±15, n=7; COPD+inhibitor 47±16%UT viability, n=7). MMP-9 inhibitor had no effect on maximal airway closure (UT 83±4, n=8; COPD 36±2, n=4, COPD+inhibitor 24±13% closure, n=5) or viability (COPD 59±13, n=4; COPD+MMP-9 inhibitor 48±12 %UT viability, n=4). NAC restored maximal airway closure by 16% in 1mM H 2 O 2 -treated SA (UT 91±4, n=8; H 2 O 2 70±8, n=8; H 2 O 2 +NAC 86±4%closure, n=6), but not in SA treated with COPD neutrophil supernatants (UT 78±6, n=6; COPD 28±7, n=4; COPD+NAC 7±9% closure, n=4). Therefore, NE was involved in COPD neutrophil supernatant-induced rat SA dysfunction. NE inhibition may protect against elastolytic damage, or inhibit tissue mediator release and therefore could be a target for SA dysfunction in COPD.