MRC DiMeN Doctoral Training Partnership: Widening the ‘NET’ to identify new treatments for bronchiectasis

Bronchiectasis (NCFB) is a chronic lung condition that affects >200,000 people in the UK. Acute exacerbations of NCFB require intense treatment and carry significant implications for morbidity and mortality particularly from cardiovascular events. The precise pathophysiological changes leading up to exacerbation are poorly understood. However, some disease characteristics share striking similarities to that of inflammatory diseases including rheumatoid arthritis (RA).

Neutrophils and neutrophil extracellular traps (NETs) drive inflammation in RA and are elevated in NCFB sputum. However, little is known about the role of neutrophils/NETs in the clinical transition from stable to exacerbation to resolution of NCFB. This new project between the Universities of Liverpool and Newcastle will accelerate impact and translation of two longitudinal NCFB studies, POSTED and BronchUK, to define the relationship between the sputum proteome, systemic inflammation, neutrophils/NETs and clinical outcomes.

Objectives

1. Determine the temporal dynamics of the sputum proteome and NETs in NCFB exacerbation and resolution.

2. Determine the relationship between NETs/sputum proteome, serum markers of inflammation (including NETs and platelet dysfunction) and cardiovascular outcomes.

3. Investigate the effect of NET-targeting therapeutics on the activation of NCFB neutrophils.

Novelty and Timeliness

NETs decrease in response to antibiotics in a manner that correlates with clinical outcomes. However the temporal dynamics/stability of NETs and how these relate to exacerbations is not known. We recently identified NETs as a potential therapeutic target in RA. This study will, for the first time, define the potential for NETs to serve as biomarkers for exacerbations in NCFB, and their potential as a novel therapeutic target.

Experimental Approach

Sputum proteomics will be performed before and during exacerbation in derivation (POSTED) and validation (BronchUK) samples. NETs (sputum/serum) and platelet activation markers (serum) will be measured by ELISA. Linked clinical metadata including detailed respiratory and cardiovascular disease metrics will be explored to determine relationships with disease outcomes. Neutrophils from people with NCFB and healthy controls will be activated using lipopolysaccharide and crude bacterial lysates corresponding to NCFB lung microbiome (measured in POSTED). Neutrophil activation will be measured and NETs imaged using fluorescence microscopy. Experiments will be performed in the absence and presence of potential small molecule therapeutics.

Training

The student will be trained in both quantitative and interdisciplinary skills. These will including immuno-assays including tissue culture of primary immune cells, molecular biology, microscopy, proteomics and training in handling of human tissue. The student will be embedded in Dr Emmott’s group for 6-months to learn how to process sputum samples for proteomics as well as analyse proteomic datasets. These skills will include the bioinformatic processing of raw data, as well as statistical analysis and pathway modelling via attendance on R-based courses provided by the Computational Biology Facility. The student will also benefit from a 3-month research visit to Prod De Soyza’s lab in Newcastle to train in the preparation of crude bacterial lysates for use in NET assays. They will have opportunities to take part in patient involvement presentations, and public engagement activities. 

Supervisor Weblinks

https://www.liverpool.ac.uk/people/helen-wright

https://www.liverpool.ac.uk/people/frederick-frost

https://www.liverpool.ac.uk/people/edward-emmott

https://www.ncl.ac.uk/medical-sciences/people/profile/anthonyde-soyza.html

Benefits of being in the DiMeN DTP:

This project is part of the Discovery Medicine North Doctoral Training Partnership (DiMeN DTP), a diverse community of PhD students across the North of England researching the major health problems facing the world today. Our partner institutions (Universities of Leeds, Liverpool, Newcastle, York and Sheffield) are internationally recognised as centres of research excellence and can offer you access to state-of-the-art facilities to deliver high impact research.

We are very proud of our student-centred ethos and committed to supporting you throughout your PhD. As part of the DTP, we offer bespoke training in key skills sought after in early career researchers, as well as opportunities to broaden your career horizons in a range of non-academic sectors.

Being funded by the MRC means you can access additional funding for research placements, training opportunities or internships in science policy, science communication and beyond. Further information on the programme and how to apply can be found on our website:

https://www.dimen.org.uk/