Hypoxia upregulates neutrophil degranulation and potential for tissue injury

Abstract

BackgroundThe inflamed bronchial mucosal surface is a profoundly hypoxic environment. Neutrophilic airway inflammation and neutrophil-derived proteases have been linked to disease progression in conditions such as COPD and cystic fibrosis, but the effects of hypoxia on potentially harmful neutrophil functional responses such as degranulation are unknown.Methods and resultsFollowing exposure to hypoxia (0.8% oxygen, 3 kPa for 4 h), neutrophils stimulated with inflammatory agonists (granulocyte-macrophage colony stimulating factor or platelet-activating factor and formylated peptide) displayed a markedly augmented (twofold to sixfold) release of azurophilic (neutrophil elastase, myeloperoxidase), specific (lactoferrin) and gelatinase (matrix metalloproteinase-9) granule contents. Neutrophil supernatants derived under hypoxic but not normoxic conditions induced extensive airway epithelial cell detachment and death, which was prevented by coincubation with the antiprotease α-1 antitrypsin; both normoxic and hypoxic supernatants impaired ciliary function. Surprisingly, the hypoxic upregulation of neutrophil degranulation was not dependent on hypoxia-inducible factor (HIF), nor was it fully reversed by inhibition of phospholipase C signalling. Hypoxia augmented the resting and cytokine-stimulated phosphorylation of AKT, and inhibition of phosphoinositide 3-kinase (PI3K)γ (but not other PI3K isoforms) prevented the hypoxic upregulation of neutrophil elastase release.ConclusionHypoxia augments neutrophil degranulation and confers enhanced potential for damage to respiratory airway epithelial cells in a HIF-independent but PI3Kγ-dependent fashion.

Hypoxic regulation of neutrophil function and consequences for Staphylococcus aureus infection

“…Consistent with these results, pharmacological and genetic HIF-1a knockdown decreased NET production and inhibited extracellular bacterial killing [74]. However, in vitro NET formation under true hypoxia has been shown to be diminished [71]. As NETosis is predominantly dependent on NADPH production of ROS [75], and therefore reliant on availability of molecular oxygen, it seems likely that NET production under hypoxia would be reduced in line with the ROS data, though it appears that HIF1a signalling also has a role to play.…”

“…As such, hypoxia enhanced the release of multiple granule products, including MMP-9, lactoferrin and active NE and MPO from stimulated human neutrophils [71]. HIF-1a siRNA knockdown reduced protein levels of murine b-defensins and cathelicidin-related antimicrobial peptide (analogous to human antimicrobial peptide LL-37) in a mouse model of pseudomonal infection [61].…”

Priming and de‐priming of neutrophil responses in vitro and in vivo

“…Potera et al noted involvement of p38 and ERK1/2 in TNF‐augmented granule release, but ROS suppressed the magnitude of this response, suggesting a negative feedback role. Consistent with this, our group has shown that hypoxia sufficient to suppress NADPH oxidase function enhances primed (GM‐CSF or PAF) neutrophil degranulation in a PI3Ky‐dependent fashion …”

“…Consistent with this, our group has shown that hypoxia sufficient to suppress NADPH oxidase function enhances primed (GM-CSF or PAF) neutrophil degranulation in a PI3Ky-dependent fashion. 40 Primed granule exocytosis is tightly regulated and hierarchical; in an in vivo skin blister model, complete mobilisation of secretory vesicles was accompanied by discharge of 38% of tertiary, 22% of secondary and just 7% of primary granule proteins. 41 A similar rank order of degranulation was observed in response to calcium ionophore, and calcium chelation abolishes secretion of all but secretory vesicles.…”

The interplay between neutrophils and microbiota in cancer

“…Reports have linked IL‐8 to neutrophil survival and more controversially to NETosis induction . The hypoxic environment of a tumor or the gut may also contribute to NETosis as it increases neutrophil adhesion, degranulation, and toxicity . NETs may accelerate tumorigenesis as ROS, RNS, neutrophil elastase (NE), cathepsin G, and matrix metalloproteinases (MMPs) released by NETs may promote DNA mutations, mitogenesis, inhibit T‐cell activation, and spur on tumor invasion and metastasis .…”