Lack of neutrophil degranulation in low-dose endotoxin inhalation based on a novel intracellular assay

To study the nature of endotoxin or lipopolysaccharide (LPS) induced inflammation, we developed a method of quantifying intracellular human neutrophil elastase (HNE) in lysed sputum polymorphs as a means to study the degranulation status of LPS-recruited neutrophils. Induced sputum, blood and exhaled nitric oxide (NO) were collected from 10 healthy non-atopic human subjects after inhaling a single 15 microg dose of Escherichia coil LPS in an open study. At 6 hours, LPS inhalation caused significant increase of sputum and blood neutrophils but without parallel increase in myeloperoxidase, HNE or interleukin-8 (IL-8) in sputum sol and blood, or exhaled NO. Intracellular HNE in lysed sputum polymorphs or purified blood neutrophils did not show any significant changes between inhaled LPS and saline, nor was there any appreciable change in percentage HNE release induced by N-Formyl-Met-Leu-Phe (fMLP) in vitro. We concluded that in healthy humans, the transient neutrophilic inflammation induced by a single dose of inhaled 15 microg LPS is mainly characterized by cell recruitment, not enhanced secretion of granular mediators or increased exhaled NO based on our experimental conditions.