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BMI-1 extends proliferative potential of human bronchial epithelial cells whilst retaining their mucociliary differentiation capacity.

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journal contribution
posted on 27.01.2017, 17:54 by M. M. Munye, A. Shoemark, Robert A. Hirst, J. M. Delhove, T. V. Sharp, T. R. McKay, C. O'Callaghan, D. L. Baines, S. J. Howe, S. L. Hart
Air-liquid interface (ALI) culture of primary airway epithelial cells enables mucociliary differentiation providing an in vitro model of the human airway but their proliferative potential is limited. To extend proliferation, these cells were previously transduced with viral oncogenes or mouse Bmi-1 + hTERT but the resultant cell lines did not undergo mucociliary differentiation. We hypothesised that use of human BMI-1 alone would increase the proliferative potential of bronchial epithelial cells while retaining their mucociliary differentiation potential. CF and non-CF bronchial epithelial cells were transduced by lentivirus with BMI-1 then their morphology, replication kinetics and karyotype were assessed. When differentiated at ALI, mucin production, ciliary function and transepithelial electrophysiology were measured. Finally, shRNA knockdown of DNAH5 in BMI-1 cells was used to model primary ciliary dyskinesia (PCD). BMI-1 transduced basal cells showed normal cell morphology, karyotype and doubling times despite extensive passaging. The cell lines underwent mucociliary differentiation when cultured at ALI with abundant ciliation and production of the gel-forming mucins MUC5AC and MUC5B evident. Cilia displayed a normal beat frequency and 9+2 ultrastructure. Electrophysiological characteristics of BMI-1 transduced cells were similar to un-transduced cells. shRNA knockdown of DNAH5 in BMI-1 cells produced immotile cilia and absence of DNAH5 in the ciliary axoneme as seen in cells from patients with PCD. BMI-1 delayed senescence in bronchial epithelial cells, increasing their proliferative potential but maintaining mucociliary differentiation at ALI. We have shown these cells are amenable to genetic manipulation and can be used to produce novel disease models for research and dissemination.

Funding

This study was funded by the Great Ormond Street Hospital Children’s Charity (GOSHCC), the Child Health Research Appeal Trust (CHRAT) and supported by the National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London.

History

Citation

AJP - Lung Cellular and Molecular Physiology, 2016, pp. ajplung.00471.2016-?

Author affiliation

/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/School of Medicine/Department of Infection, Immunity and Inflammation

Version

AM (Accepted Manuscript)

Published in

AJP - Lung Cellular and Molecular Physiology

Publisher

American Physiological Society

issn

1040-0605

eissn

1522-1504

Available date

15/12/2017

Publisher version

http://ajplung.physiology.org/content/early/2016/12/14/ajplung.00471.2016

Notes

One year embargo

Language

en