Characterisation of an ex vivo model of human airway epithelium

Introduction: ex vivo airway epithelial cell models provide a biologically representative platform for investigating airway diseases. When cultured at the air-liquid interface (ALI), cells should mimic the physiology of in vivo conditions rendering them invaluable as diagnostic and research tools.Methods: cells from nasal brush biopsy were expanded in PneumaCult™-Ex Plus Medium (STEMCELL Technologies) for 12 days. Once confluent, the monolayer of basal cells was exposed to an ALI and fed basolaterally with PneumaCult™-ALI Medium. Epithelial tight junction formation was assessed by transepithelial electrical resistance (TEER) and cell layer integrity was measured using fluorescein sodium as a marker of paracellular permeability. Cell morphology was examined by histology, confocal and electron microscopy. Ciliary function was assessed by high speed video analysis and percentage coverage of cilial beating was quantified using a fast Fourier transform algorithm. Supernatants were analysed weekly for cytokine production.Results: motile cilia were first observed by day 7 at ALI. TEER was maintained for 4 weeks before showing signs of gradual decline. Imaging showed fully differentiated epithelium with tight junctional complexes at 12 weeks post-ALI. Ciliary beat frequency at 3-5 weeks post-ALI (mean 14.18 Hz; SD ±1.84; n=20) was within the physiologically normal range at 37°C. Percentage coverage of cilial beating at 4-7 weeks post-ALI was 41.1% (SD ±14.83; n=6).Conclusion: we report a biologically relevant model of primary human airway epithelium for the investigation of airway pathophysiology. We are currently using it to diagnose primary ciliary dyskinesia and for experimental research