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Characterisation of a surrogate Type-II alveolar cell culture model for ATP binding cassette subfamily a member 3 (ABCA-3) deficiency

Shaw, N., Kicic, A., Schneider-Daum, N., Fletcher, S., Wilton, S., Huwer, H., Lehr, C., Stick, S. and Schultz, A. (2020) Characterisation of a surrogate Type-II alveolar cell culture model for ATP binding cassette subfamily a member 3 (ABCA-3) deficiency. Respirology, 25 (S1). p. 130.

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Introduction/Aim. ATP Binding Cassette Subfamily A Member 3 (ABCA-3) is a lipid transporter protein highly expressed in type-II alveolar (AT-II) cells. Mutations in ABCA3 can result in severe respiratory disease in infants and children. Cell cultures derived from patients with ABCA-3 deficiency that reflect the genetic and phenotypic features of ABCA-3 associated lung disease, would provide a means to study disease mechanisms and investigate therapeutics. Primary AT-II cells, while physiologically relevant to ABCA-3 deficiency, are not generally accessible. ABCA-3 is present in the nasopharynx, hence primary nasal epithelial cells could potentially provide a more accessible cell culture model. Our aim was to investigate the suitability of nasal epithelial cells to study ABCA-3 deficiency.

Methods. Expression of ABCA3, and AT-II cell markers, SFTPB and SFTPC, in nasal epithelial cells and primary AT-II cells, were quantified by droplet digital PCR. Protein localisation of ABCA-3 was visualised by immunofluorescent microscopy. Functionality of the ABCA-3 protein in nasal epithelial cells derived from human subjects with or without ABCA-3 deficiency was assessed by the capacity of the cells to detoxify doxorubicin.

Results. While ABCA-3 protein was localised in the cytoplasm of primary nasal epithelial cells, mRNA levels were 6.35x103-fold lower in nasal epithelial cells compared to primary AT-II cells. Gene expression of SFTPB and SFTPC was also significantly lower by 5.26 x 106- and 5.36 x 106-fold, respectively. Higher concentrations of doxorubicin reduced cell viability in ABCA-3 deficient nasal epithelial cells compared to controls in an assay-dependent manner when using an MTS assay but not with a lactate dehydrogenase assay or by calcein blue staining.

Conclusion. There may be a role for nasal epithelial cell cultures to model ABCA-3 deficiency depending on endpoints of interest. However, alternative approaches that more accurately replicate the AT-II phenotype need to be explored.

Item Type: Journal Article
Murdoch Affiliation(s): Centre for Comparative Genomics
Centre for Molecular Medicine and Innovative Therapeutics (CMMIT)
Publisher: Wiley
Copyright: © 2020 Asian Pacific Society of Respirology.
Other Information: Poster abstract given @ TSANZSRS 2020 The Australia & New Zealand Society of Respiratory Science and The Thoracic Society of Australia and New Zealand (ANZSRS/TSANZ) Annual Scientific Meeting for Leaders in Lung Health & Respiratory Science, 27–31 March 2020, Melbourne Convention and Exhibition Centre, VIC, Australia
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