Lentiviral small hairpin RNA delivery reduces apical sodium channel activity in differentiated human airway epithelial cells

J (Jamil) Aarbiou, E Copreni, RM Buijs-Offerman, P Wegen, S Castellani, A Carbone, F Tilesi, P Fradiani, PS Hiemstra, G Yueksekdag, A Diana, J Rosenecker, F Ascenzioni, M Conese, Bob Scholte

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Abstract

Background Epithelial sodium channel (ENaC) hyperactivity has been implicated in the pathogenesis of cystic fibrosis (CF) by dysregulation of fluid and electrolytes in the airways. In the present study, we show proof-of-principle for ENaC inhibition by lentiviral-mediated RNA interference. Methods Immortalized normal (H441) and CF mutant (CFBE) airway cells, and differentiated human bronchial epithelial cells in air liquid interface culture (HBEC-ALI) were transduced with a vesicular stomatitis virus G glycoprotein pseudotyped lentiviral (LV) vector expressing a short hairpin RNA (shRNA) targeting the alpha subunit of ENaC (ENaC alpha), and a marker gene. Efficacy of ENaC alpha down-regulation was assayed by the real-time polymerase chain reaction (PCR), membrane potential assay, Results Transduction to near one hundred percentage efficiency of H441, CFBE and HBEC-ALI was achieved by the addition of the LV vector before differentiation and polarization. Transduction resulted in the inhibition of ENaC alpha mRNA and antigen expression, and a proportional decrease in ENaC-dependent short circuit current and fluid transport. No effect on transepithelial resistance or cAMP-induced secretion responses was observed in HBEC-ALI. The production of interferon alpha and pro-inflam Conclusions We have established a generic method for studying the effect of RNA interference in HBEC-ALI using standard lentiviral vectors. Down-regulation of ENaC alpha by lentiviral shRNA expression vectors as shown in the absence off-target effects has potential therapeutic value in the treatment of cystic fibrosis. Copyright (C) 2012 John Wiley & Sons, Ltd.
Original languageUndefined/Unknown
Pages (from-to)733-745
Number of pages13
JournalJournal of Gene Medicine
Volume14
Issue number12
DOIs
Publication statusPublished - 2012

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