|Institution:||University of Cambridge|
|Department:||Department of Medicine|
|Keywords:||Neutrophil; Degranulation; Hypoxia; COPD; Immunology|
|Full text PDF:||http://www.repository.cam.ac.uk/handle/1810/247459
Neutrophils are key effector cells of the innate immune system. They employ a number of powerful ‘weapons’ to eliminate pathogens, including an array of destructive proteins packaged into distinctive granule subsets. In addition to their microbicidal activity, these granule proteins are capable of causing substantial tissue damage if inappropriately deployed. To mitigate against this possibility, most physiological stimuli induce minimal extracellular degranulation. Sites of inflammation and infection are usually hypoxic, and it has been shown that oxygen depletion compromises neutrophil function by impairing the generation of reactive oxygen species and hence bacterial killing. The key finding reported in this thesis is that hypoxia substantially increases the release of all neutrophil granule subsets, as measured by the release of (active) hallmark proteins (elastase, myeloperoxidase, lactoferrin and matrix metalloproteinase-9). In consequence, supernatants from hypoxic neutrophils induced substantially more damage to lung epithelial cell layers than supernatants from neutrophils cultured under normoxic conditions; this damage was protein- and protease-dependent. This pattern of damage was seen consistently across lung adenocarcinoma-derived epithelial cells, primary immortalised lung epithelial cells, and primary human bronchial epithelial cells grown in physiological air-liquid interface culture. Surprisingly, the mechanism of hypoxia-augmented degranulation was found to be independent of protein synthesis and specifically, of the transcription factor HIF-1α (the ‘master-regulator’ of hypoxic responses); thus, hypoxia did not affect mRNA transcript or protein abundance of the major granule components, and hypoxia mimetics failed to recapitulate the phenotype. Inhibition of the key pathways known to be involved in neutrophil degranulation, including, phosphatidylinositol 3-kinase and phospholipase C, but not calcium flux prevented augmented granule release under hypoxia In conclusion, hypoxia induces a destructive neutrophil phenotype, with increased release of multiple histotoxic proteases. This may contribute to tissue injury and disease pathogenesis in a range of clinically important conditions.