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

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 α subunit of ENaC (ENaCα), and a marker gene.

Repeated siRNA application is a precondition for successful mRNA gammaENaC knockdown in the murine airways.

The volume of the airway surface liquid is regulated by Na(+) absorption and Cl(-) secretion by the respiratory epithelium. In cystic fibrosis, Na(+) hyperabsorption caused by the absence of functional CFTR protein leads to an altered airway surface liquid composition and finally to a deteriorated mucociliary clearance. It has been suggested that down regulation or inhibition of the amiloride-sensitive epithelial Na(+) channel (ENaC) could restore the disrupted airway hydration.

Epithelial sodium channel inhibition in primary human bronchial epithelia by transfected siRNA.

Na(+) absorption and Cl(-) secretion are in equilibrium to maintain an appropriate airway surface fluid volume and ensure appropriate mucociliary clearance. In cystic fibrosis, this equilibrium is disrupted by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene resulting in the absence of functional CFTR protein, which in turn results in deficient cAMP-dependent Cl(-) secretion and predominant Na(+) absorption. It has been suggested that down-regulation of the epithelial sodium channel, ENaC, might help to restore airway hydration and reverse the airway phenotype in patients with cystic fibrosis.