Cycloheximide and lipopolysaccharide downregulate αENaC mRNA via different mechanisms in alveolar epithelial cells

• Published in: American journal of physiology. Lung cellular and molecular physiology Vol. 305; no. 10; p. L747
• Main Authors: Migneault, Francis, Boncoeur, Emilie, Morneau, Frédéric, Pascariu, Mihai, Dagenais, André, Berthiaume, Yves
• Format: Journal Article
• Language: English
• Published: United States 15-11-2013
• Subjects: Animals; Blotting, Western; Cells, Cultured; Cycloheximide - pharmacology; Epithelial Cells - cytology; Epithelial Cells - drug effects; Epithelial Cells - metabolism; Epithelial Sodium Channels - genetics; Epithelial Sodium Channels - metabolism; Extracellular Signal-Regulated MAP Kinases - genetics; Extracellular Signal-Regulated MAP Kinases - metabolism; Lipopolysaccharides - pharmacology; Luciferases - metabolism; Male; p38 Mitogen-Activated Protein Kinases - genetics; p38 Mitogen-Activated Protein Kinases - metabolism; Promoter Regions, Genetic - genetics; Protein Synthesis Inhibitors - pharmacology; Pulmonary Alveoli - cytology; Pulmonary Alveoli - drug effects; Pulmonary Alveoli - metabolism; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; Signal Transduction - drug effects; messenger ribonucleic acid; epithelial sodium channel; inflammation; signal transduction; cellular stress
• ISSN: 1522-1504
• DOI: 10.1152/ajplung.00023.2013

Active Na(+) transport mediated by epithelial Na(+) channel (ENaC) is vital for fetal lung fluid reabsorption at birth and pulmonary edema resolution. Previously, we demonstrated that αENaC expression and activity are downregulated in alveolar epithelial cells by cycloheximide (Chx) and Pseudomonas aeruginosa. The regulatory mechanisms of αENaC mRNA expression by Chx and lipopolysaccharide (LPS) from P. aeruginosa were further studied in the present work. Both agents decreased αENaC mRNA expression to 50% of control values after 4 h. Chx repressed αENaC expression in a dose-dependent manner independently of protein synthesis. Although extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) pathways were activated by the two treatments, their mechanisms of ENaC mRNA modulation were different. First, activation of the signaling pathways was sustained by Chx but only transiently by LPS. Second, ERK1/2 or p38 MAPK inhibition attenuated the effects of Chx on αENaC mRNA, whereas suppression of both signaling pathways was necessary to alleviate the outcome of LPS on αENaC mRNA. The molecular mechanisms involved in the decrease of αENaC expression were investigated in both conditions. LPS, but not Chx, significantly reduced αENaC promoter activity via the ERK1/2 and p38 MAPK pathways. These results suggest that LPS attenuates αENaC mRNA expression via diminution of transcription, whereas Chx could trigger some posttranscriptional mechanisms. Although LPS and Chx downregulate αENaC mRNA expression similarly and with similar signaling pathways, the mechanisms modulating ENaC expression are different depending on the nature of the cellular stress.