Using Sputum Proteomics to Develop a Test for Reflux Aspiration in Cystic Fibrosis Patients

Background: Reflux aspiration is a process by which gastric contents are introduced into the respiratory tract, and may lead to lung damage. Currently no method exists to identify patients with reflux aspiration. Here we describe a quantitative proteomics approach to assess differences in the sputum proteome between cystic fibrosis (CF) patients based on gastro-oesophageal reflux (GOR) measures. Methods: Spontaneous sputum samples were collected from CF subjects (n=37), and sputum was induced from healthy volunteers (HV, n=33) for comparison. The sputum proteome was analysed by bottom-up shotgun proteomic analysis. All CF patients had GOR measured using 24-hour pH-impedance; sputum samples were compared from those with the highest (n=5) and lowest (n=5) risk for reflux aspiration, based on total, proximal and/or supine reflux episodes. Results: There were significant differences between the CF and HV sputum proteome with large increases in inflammatory proteins, predominantly neutrophil granulocyte proteins. There were also significant proteome differences between CF patients with highest and lowest reflux measures. These protein differences were not indicative of inflammation but other underlying cellular processes. No proteins of gastric origin were identified. Conclusion: These data suggest the sputum proteome of CF subjects may be influenced by gastro-oesophageal reflux. Although this appears to support a relationship between reflux and CF lung disease, it is not currently possible to conclude if this is a consequence of reflux aspiration. We aim to further explore these proteome differences, with the eventual aim of developing a protein biomarker for diagnosis of reflux aspiration.