Poster Presentation BACPATH 2019

Polymicrobial infection and neutrophilic disease in cystic fibrosis airways (#126)

Daniel R Laucirica 1 2 , Craig J Schofield 1 , Samantha A McLean 1 , Camilla Margaroli 3 4 , Stephen M Stick 1 2 5 , Rabindra Tirouvanziam 3 4 , Anthony Kicic 1 2 5 6 , Luke W Garratt 1 2 , AREST CF 1 5 7 8
  1. Telethon Kids Institute/ University of Western Australia, Nedlands, WA, Australia
  2. Faculty of Health and Medical Sciences, University of Western Australia, Nedlands, WA, Australia
  3. Dept of Pediatrics, Emory University, Atlanta, GA, USA
  4. Center for CF & Airways Disease Research, Children's Healthcare of Atlanta, Atlanta, GA, USA
  5. Dept of Respiratory and Sleep Medicine, Perth Children's Hospital, Nedlands, WA, Australia
  6. School of Population Health, Curtin University, Bentley, WA, Australia
  7. Murdoch Children's Research Institute, Melbourne, VIC, Australia
  8. Dept of Paediatrics, University of Melbourne, Melbourne, VIC, Australia

Cystic fibrosis (CF) lung damage is driven by a cycle of infection, pro-inflammatory signalling, and neutrophilic reprogramming. However, the mechanisms behind this process are poorly characterized. We created a multi-component in vitro system to model CF inflammatory responses and airway neutrophil recruitment, specifically in the context of polymicrobial infections. This was used to assess epithelial and neutrophil responses to rhinovirus and Pseudomonas aeruginosa infection. Submerged monolayers of primary CF airway epithelial cells (3M:1F; age≤5yr) were infected individually and in combination with rhinovirus strain RV1b (MOI 0.5) and a mucoid P. aeruginosa clinical isolate (MOI 0.001). After 48 hours, cell culture supernatants were harvested and epithelial secreted cytokines quantified by ELISA. Filtered supernatants were also applied to an in vitro model of neutrophil transmigration to the airways. Migrated neutrophils were harvested 10 hours post stimulation and assessed by flow cytometry. Both infection with RV1b or RV1b+P. aeruginosa significantly increased production of proinflammatory cytokines IL-8 and IL-1β compared to uninfected controls or bacterial infection alone (p<0.01). Production of CCL5 was significantly increased in viral infections (p<0.03). Biofilms formed upon P. aeruginosa infection, however, more non-aggregated planktonic bacteria were observed with RV1b+P. aeruginosa coinfection. In the transmigration model, neutrophils migrated in similar numbers towards all supernatants. However, neutrophils migrating towards bacterial or coinfection supernatants had significantly reduced staining of CD16, a phagocytosis marker (p<0.01). Expression of exocytosis marker CD63 was unchanged. Results highlight the role of respiratory viruses in CF as triggers of airway inflammation and promoters of secondary bacterial infection. Coinfection induced the greatest change in expression of a neutrophil phagocytosis marker, suggesting that polymicrobial infections may be implicated in CF neutrophilic reprogramming. This model permits investigation of coordinated CF airway responses to diverse pathogenic insults. Ongoing work will assess responses in non-CF airway epithelium and discern how recruited neutrophil signalling further enhances reprogramming.