Optimisation of growth conditions for ovine airway epithelial cell differentiation at an air-liquid interface

O'Boyle, N., Sutherland, E., Berry, C. C. and Davies, R. L. (2018) Optimisation of growth conditions for ovine airway epithelial cell differentiation at an air-liquid interface. PLoS ONE, 13(3), e0193998. (doi: 10.1371/journal.pone.0193998) (PMID:29518140) (PMCID:PMC5843276)

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Abstract

Respiratory tract infections are of significant concern in the agriculture industry. There is a requirement for the development of well-characterised in vitro epithelial cell culture models in order to dissect the diverse molecular interactions occurring at the host-pathogen interface in airway epithelia. We have analysed key factors that influence growth and differentiation of ovine tracheal epithelial cells in an air-liquid interface (ALI) culture system. Cellular differentiation was assessed at 21 days post-ALI, a time-point which we have previously shown to be sufficient for differentiation in standard growth conditions. We identified a dose-dependent response to epidermal growth factor (EGF) in terms of both epithelial thickening and ciliation levels. Maximal ciliation levels were observed with 25 ng ml-1 EGF. We identified a strict requirement for retinoic acid (RA) in epithelial differentiation as RA exclusion resulted in the formation of a stratified squamous epithelium, devoid of cilia. The pore-density of the growth substrate also had an influence on differentiation as high pore-density inserts yielded higher levels of ciliation and more uniform cell layers than low pore-density inserts. Differentiation was also improved by culturing the cells in an atmosphere of sub-ambient oxygen concentration. We compared two submerged growth media and observed differences in the rate of proliferation/expansion, barrier formation and also in terminal differentiation. Taken together, these results indicate important differences between the response of ovine tracheal epithelial cells and other previously described airway epithelial models, to a variety of environmental conditions. These data also indicate that the phenotype of ovine tracheal epithelial cells can be tailored in vitro by precise modulation of growth conditions, thereby yielding a customisable, potential infection model.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Berry, Dr Catherine and Davies, Dr Robert and O'Boyle, Dr Nicky and Sutherland, Miss Erin
Creator Roles:
O'Boyle, N.Data curation, Formal analysis, Investigation, Methodology, Validation, Visualization, Writing – original draft, Writing – review and editing
Sutherland, E.Data curation, Investigation
Berry, C. C.Project administration, Supervision, Writing – review and editing
Davies, R. L.Conceptualization, Funding acquisition, Project administration, Resources, Software, Supervision, Writing – review and editing
Authors: O'Boyle, N., Sutherland, E., Berry, C. C., and Davies, R. L.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:PLoS ONE
Publisher:Public Library of Science
ISSN:1932-6203
ISSN (Online):1932-6203
Copyright Holders:Copyright © 2018 O'Boyle et al.
First Published:First published in PLoS ONE 13(3): e0193998
Publisher Policy:Reproduced under a Creative Commons License

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
637741Molecular interactions of Mannheimia haemolytica with the bovine and ovine respiratory tracts using three-dimensional tissue engineering approachesRobert DaviesBiotechnology and Biological Sciences Research Council (BBSRC)BB/L010534/1III - BACTERIOLOGY