Subunit-specific coordinate upregulation of sodium pump activity in alveolar epithelial cells by lentivirus-mediated gene transfer

Resolution of alveolar edema depends on active ion transport by sodium pumps located on the basolateral surface of alveolar epithelial cells (AECs), suggesting that upregulation of sodium pump activity may facilitate clearance of edema fluid. We have investigated the use of lentiviral vectors to augment sodium pump activity via gene transfer of sodium pump subunits to AECs. Full-length cDNA for the α₁ or β₁ subunit of rat Na⁺,K⁺-ATPase was cloned into the lentiviral vector pRRLsin.hCMV.IRES.EGFP. Rat AECs in primary culture were transduced on day 4 with lentiviral vectors pseudotyped with vesicular stomatitis virus glycoprotein G. Transduction with lentiviral vectors encoding either α₁ subunit (Lenti-α₁-EGFP) or β₁ subunit (Lenti-β₁-EGFP) led to dose-dependent increases in mRNA and protein for the corresponding subunit. Transduction with Lenti- β₁-EGFP was accompanied by coordinate upregulation of endogenous α₁ expression, whereas endogenous β₁ expression was unchanged after transduction with Lenti-α₁-EGFP. Consistent with these findings, transduction with Lenti-β₁- EGFP, but not Lenti-α₁-EGFP, led to augmentation of sodium pump activity as a result of increases in Na⁺,K⁺-ATPase holoenzyme. Sodium pump α₂ subunit and sodium channel protein did not change after Lenti-β₁-EGFP transduction. These results demonstrate that overall sodium pump activity can be efficiently upregulated in AECs specifically via gene transfer of the sodium pump β₁ subunit and support the feasibility of lentivirus-mediated gene transfer to augment alveolar fluid clearance.