Regulation of neutrophilic inflammation by hypoxic signalling pathways
Item statusRestricted Access
Embargo end date06/07/2020
Watts, Emily Rose
MetadataShow full item record
Neutrophils are essential for effective innate immunity. Conversely, inappropriate or excessive neutrophil activation can result in damaging inflammation. This damage is implicated in the pathogenesis of a number of respiratory diseases including acute respiratory distress syndrome (ARDS) and chronic obstructive pulmonary disease (COPD) which are also both frequently complicated by hypoxia. Cells sense and respond to hypoxia through the activity of the transcription factor HIF (hypoxia inducible factor) and its regulatory hydroxylases, the prolyl hydroxylase domain enzymes (PHDs) 1- 3. In the presence of oxygen, PHDs hydroxylate HIF, preventing the HIF mediated transcriptional response. Close links exist between the pathways which regulate hypoxic and inflammatory responses. Our group has previously found that in mouse models of infection, acute hypoxia leads to increased sickness and that this is driven by neutrophilic inflammation. I have used a murine model of Lipopolysaccharide (LPS) -induced acute lung injury, characterised by neutrophil influx, to investigate how exposure to hypoxia alters lung inflammation. Using high-resolution mass spectrometry, I have defined the proteome of the inflammatory lung neutrophil. I have shown that hypoxia results in a distinct proteomic signature in inflammatory neutrophils. Hypoxia drives lung neutrophilic inflammation through increased neutrophil degranulation and upregulation of inflammatory receptors. I have also identified key metabolic alterations in hypoxic neutrophils. The hypoxic lung represents a low glucose, high protein environment and neutrophils adapt to exploit this. I have shown that neutrophils can scavenge proteins from their extracellular environment, catabolise these proteins in the lysosome and utilise the breakdown products for metabolism. These processes are upregulated in hypoxic lung neutrophils which show increased lysosomal protein expression, increased protein uptake and increased glutaminolysis. Utilising heavy labelled protein extracts, I have traced breakdown products from scavenged proteins into central carbon metabolism, demonstrating that extracellular protein can fuel neutrophilic inflammation. Finally, I have investigated the role of the prolyl hydroxylase PHD1 in regulating neutrophilic inflammation. Using a neutrophil specific PHD1 knockout mouse line, I have identified a specific role for PHD1 in regulating neutrophil metabolism and survival. I have found that the micro-environment, particularly oxygen availability, determines the impact of PHD1 loss with consequences for inflammation resolution in vivo. In summary, hypoxia is a key regulator of neutrophil function and is associated with increased neutrophilic inflammation. Utilising a proteomic approach, I have identified the mechanisms which drive the hyperinflammatory phenotype including the ability of neutrophils to scavenge proteins from the environment to fuel inflammation. I have also shown that PHD1, a key component of the hypoxic signalling pathway, may regulate these functions. A more complete understanding of these mechanisms will help to identify therapeutic targets for treatment of neutrophilic inflammation in the lung.