Human Neutrophils Couple Nitric Oxide Production and Extracellular Trap Formation in Allergic Asthma

Rationale: Nitric oxide (NO) is increased in the airways and serum of patients with allergic asthma, suggesting an important role in asthma. NO production has been widely attributed to the canonical inducible NO synthase. Much effort has been made to inhibit this enzyme, with two outcomes: no asthma improvement and partial NO reduction, suggesting the involvement of an inducible NO synthase-independent source. Objectives: Neutrophils produce NO under inflammatory conditions, and their role in asthma has been overlooked. The present study analyzes their possible role as sources of NO. Methods: Our hypothesis was tested in 99 allergic patients with intermittent bronchial asthma and 26 healthy donors. NO production by blood and sputum neutrophils in response to allergens, anti-IgE, and anti-IgE receptor antibodies was assessed by Griess reagent, flow cytometry, and confocal microscopy. The formation of extracellular traps (ETs) as a possible consequence of NO production was quantified by Western blot and confocal microscopy, and reactive oxygen species were assessed with luminol-enhanced chemiluminescence. Measurements and Main Results: Among blood and sputum granulocytes from patients with allergic asthma, only neutrophils produce NO by an IgE-dependent mechanism. This production is independent of NO synthase, but dependent on a reaction between L-arginine and reactive oxygen species from NOX2 (NADPH oxidase). NO and ETosis are induced in parallel, and NO amplifies ET formation, which is a key mediator in asthma. Conclusions: Our findings reveal a novel role of neutrophils as the unique allergen/IgE-dependent NO source in allergic asthma, enhancing ET formation. These results suggest that NO produced by neutrophils needs further consideration in the treatment of allergic asthma.