Na,K-ATPase Gene Transfer Mitigates an Oxidant-Induced Decrease of Active Sodium Transport in Rat Fetal ATII Cells

• Published in: American journal of respiratory cell and molecular biology Vol. 24; no. 3; pp. 245 - 252
• Main Authors: Thome, Ulrich, Chen, Lan, Factor, Phillip, Dumasius, Vidas, Freeman, Bruce, Iasha Sznajder, J, Matalon, Sadis
• Format: Journal Article
• Language: English
• Published: United States Am Thoracic Soc 01-03-2001; American Thoracic Society
• Subjects: Amphotericin B - pharmacology; Animals; Biological Transport, Active - drug effects; Biological Transport, Active - physiology; Cells, Cultured; DNA, Complementary; Fetus; Hydrogen Peroxide - pharmacology; Lung - cytology; Lung - physiology; Ouabain - pharmacology; Oxidants - pharmacology; Protein Subunits; Rats; Respiratory Mucosa - cytology; Respiratory Mucosa - drug effects; Respiratory Mucosa - physiology; Sodium - metabolism; Sodium-Potassium-Exchanging ATPase - genetics; Sodium-Potassium-Exchanging ATPase - metabolism; Transfection
• ISSN: 1044-1549; 1535-4989
• DOI: 10.1165/ajrcmb.24.3.3994

We investigated whether adenovirus-mediated transfer of genes encoding for subunits of the Na,K-ATPase increases transepithelial Na(+) transport in rat fetal distal lung epithelial (FDLE) monolayers and renders them more resistant to hydrogen peroxide injury. FDLE cells, isolated from rat fetuses at a gestational age of 19 to 20 d (22 d = term), were seeded on filters and infected with replication-incompetent human type 5 adenoviruses containing complementary DNAs encoding for rat Na,K-ATPase alpha(1) or beta(1) subunits (ad alpha(1) and ad beta(1), respectively). Once confluent monolayers were formed, the filters were mounted in Ussing chambers and short circuit currents (I(SC)) were measured. Increased levels of alpha(1) or beta(1) subunit proteins after infection with ad alpha(1) and ad beta(1), respectively, were confirmed by Western blot analysis. Baseline I(SC) increased after transfection with 2 plaque-forming units (pfu) of ad beta(1) from 5.1 +/- 0.3 to 6.1 +/- 0.3 microA/cm(2) (mean +/- SEM; P < 0.05). Permeabilization of the apical membrane with amphotericin B caused a large increase in I(SC); the ouabain-sensitive component of the amphotericin B-elicited I(SC) (ouab(max)) was increased from 4.0 +/- 0.2 (n = 69) in controls to 4.8 +/- 0.2 (n = 15), 5.9 +/- 0.3 (n = 53), 6.9 +/- 0.4 (n = 25), 7.7 +/- 0.9 (n = 16) in monolayers infected with 1, 2, 11, and 22 pfu of ad beta(1), respectively; transfection with ad alpha(1) had no effect on any measured variables. Further, transfection with ad beta(1) in comparison to noninfected monolayers resulted in higher baseline and ouab(max) I(SC) after injury with 500 microM H(2)O(2). We conclude that overexpression of the beta(1) subunit of the Na,K-ATPase may help maintain normal levels of vectorial Na(+) transport across ATII cell monolayers in pathologic conditions.