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Background The severe high altitude illness, High Altitude Pulmonary oEdema (HAPE), occurs in some individuals exposed to hypobaric hypoxia and is characterised by neutrophil-driven lung and microvascular damage. Ultrastructural examination of lung tissue from HAPE patients shows neutrophils trapped in pulmonary capillaries. Hypoxia delays neutrophil apoptosis and increases degranulation in vitro, and delayed apoptosis is also seen in neutrophils from subjects who experienced 7-days systemic hypoxia at high altitude. However, the exact role of hypoxia in HAPE pathogenesis, and its impact on neutrophils in vivo, is poorly understood. We assessed the impact of short-term systemic hypoxia on neutrophil phenotype.Methods Venous blood was collected from healthy volunteers (n=10) pre/post exposure to normobaric hypoxia (approximately 12.8% oxygen) for two hours. Ex vivo blood/cells were maintained at venous oxygen tension (5% oxygen) throughout. Whole blood neutrophil biomechanical profiles were measured using Real-Time Deformability Cytometry (RT-DC). Gene transcription was evaluated by RT-qPCR. Apoptosis was measured via cytospin microscopy at 24 hours. Neutrophil secretion of elastase and myeloperoxidase was assessed by activity assay.Results Neutrophils exposed to pathological hypoxia (1%) in vitro are stiffer than those maintained at venous oxygen levels (p=0.045). Following systemic hypoxia, neutrophil number (p=0.0195) and porosity (membrane roughness, p=0.0273) were increased. Overall, neutrophils became slightly smaller and softer post-hypoxia (non-significant), with notable inter-donor variability. Expression of Hypoxia Inducible Factor (HIF)-dependent genes, GLUT1, BNIP3 and VEGF, was unchanged but CXCR4 gene expression was increased post-hypoxia (p=0.004), and higher CXCR4 expression was associated with smaller cell size (R2=0.71, p=0.004). Neutrophil apoptosis remained unchanged post-hypoxia, as did secretion of elastase/myeloperoxidase, although inter-donor variability was again noted.Discussion As expected, following this short in vivo hypoxic exposure, we did not see a HIF-dependent effect on neutrophil gene transcription or apoptosis. However, the blood neutrophilia and increased porosity suggest a primed (partially activated) phenotype. CXCR4 is known to be upregulated in aged pro-inflammatory neutrophils. Thus, higher CXCR4 expression in smaller cells may indicate enhanced endothelial transmigration and lung injury potential. Inter-donor variability in neutrophil biomechanical and secretion profiles in response to hypoxia may, at least partly, explain the variable individual susceptibility to HAPE.