Using an in vitro model of human cystic fibrosis airways to investigate bacterial strategies which inactivate defence mechanisms and increase infection

Pallett, Ross (2019). Using an in vitro model of human cystic fibrosis airways to investigate bacterial strategies which inactivate defence mechanisms and increase infection. PHD thesis, Aston University.

Abstract

Cystic fibrosis (CF) is the most common inherited genetic condition amongst Caucasians and arises due to mutations in the cystic fibrosis transmembrane conductance regulator, a chloride channel expressed upon the apical surface of epithelia. Whilst CF is a multi-organ disease, the inability to clear dehydrated mucus from the airways predisposes individuals to the development of chronic bacterial infections, the main cause of morbidity and mortality in CF. Infection of CF airways is highly ordered, with Staphylococcus aureus predominating in the first decade of life, followed by Pseudomonas aeruginosa during adulthood. Two obstacles to the development of better treatments stem from an incomplete understanding of thepolymicrobial nature of CF airway infection and its impact upon interspecies and host-pathogen interactions, alongside the need for models which more closely mimic the CF lung and its unique environment. After characterising a panel of P. aeruginosa CF clinical isolates, this study sought to determine the impact of oxygen availability upon S. aureus-P. aeruginosa interspecies interactions, in light of evidence that mucus plugging within CF airways leads to regions of anoxia. Anoxia was shown to modulate S. aureus-P. aeruginosa community composition in planktonic co-culture and mixed species biofilms in an isolate-dependent manner. Further investigations into the mechanisms facilitating P. aeruginosa dominance suggest that the anti-staphylococcal agentis extracellular, >3 kDa in size and heat-resistant. Whilst pulmonary inflammation is a hallmark of CF, how airways respond to stimuli received during polymicrobial airway infections is poorly understood. Monolayers of CF and non-CF bronchial epithelia were challenged with S. aureus and/or P. aeruginosa extracellular products.CF airway epithelia exhibited a hyper-inflammatory phenotype at baseline compared to non-CF epithelia. Furthermore, only co-stimulation of non-CF epithelia with both pathogens, enhanced the IL-6 and IL-8 response beyond that measured following single bacterial challenges. Finally, CF and non-CF airway epithelia grown at air-liquid interface in the presence of fibroblasts were used to mimic the sequential nature of CF infection. Binding studies demonstrated that prior infection with S. aureus enhanced P. aeruginosa binding to the CF airway model in an isolate-specific manner, a finding not seen in the non-CF airway model. These studies demonstrate that S. aureus-P. aeruginosa interactions are likely to influence the CF microbiome, airway inflammation, airway colonisation and ultimately, disease progression.It is hoped that the models used here can be employed in future studies to understand the complex interspecies and host-pathogen interactions that occur in CF, with the aim to identify novel targets and treatments to combat these life limiting infections.

Divisions: Life & Health Sciences > Biosciences
Aston University (General)
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Institution: Aston University
Uncontrolled Keywords: cystic fibrosis,in vitro,pathogen-pathogen,air liquid interface,host-pathogen
Completed Date: 2019-03-05
Authors: Pallett, Ross

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