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In vitro and in vivo regulation of transepithelial lung alveolar sodium transport by serine proteases
American Journal of Physiology Lung Cellular and Molecular Physiology
In Vitro Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. --- Old month value: Jun
The amiloride-sensitive epithelial sodium channel (ENaC) constitutes a rate-limiting step for sodium (Na+) and water absorption across lung alveolar epithelium. Recent reports suggested that ENaC is regulated by membrane-bound extracellular serine proteases, such as channel-activating proteases (CAPs). The objectives of this study were to examine the role of serine proteases in the regulation of transepithelial alveolar Na+ and water transport in vitro and in vivo and the expression of CAPs in rodent distal lung. In vitro experiments showed that inhibition of endogenous serine proteases by apical aprotinin 1) decreased ENaC-mediated currents in primary cultures of rat and mouse alveolar epithelial cells without affecting the abundance nor the electrophoretic migration pattern of biotinylated alpha- and beta-ENaC expressed at the cell surface and 2) suppressed the increase in amiloride-sensitive short-circuit current induced by the beta2-agonist terbutaline. RT-PCR experiments indicated that CAP1, CAP2, and CAP3 mRNAs were expressed in mouse alveolar epithelial cells, whereas CAP1 was also expressed in alveolar macrophages recovered by bronchoalveolar lavage. CAP1 protein was detected by Western blotting in rat and mouse alveolar epithelial cells, alveolar macrophages and bronchoalveolar lavage fluid. Finally, in vivo experiments revealed that intra-alveolar treatment with aprotinin abolished the increase in Na+-driven alveolar fluid clearance (AFC) induced by terbutaline in an in situ mouse lung model, whereas trypsin potentiated it. These results show that endogenous membrane-bound and/or secreted serine proteases such as CAPs regulate alveolar Na+ and fluid transport in vitro and in vivo in rodent lung.
Adrenergic beta-Agonists/pharmacology Animals Aprotinin/pharmacology Biological Transport/drug effects/*physiology Bronchoalveolar Lavage Carrier Proteins/genetics/metabolism Cytoskeletal Proteins Epithelial Cells/cytology/drug effects/*metabolism Epithelial Sodium Channel Female Macrophages, Alveolar/cytology/metabolism Male Membrane Proteins/genetics/metabolism Mice Mice, Inbred C57BL Proteins/genetics/metabolism Pulmonary Alveoli/*metabolism RNA, Messenger/genetics/metabolism Rats Reverse Transcriptase Polymerase Chain Reaction Serine Endopeptidases/genetics/metabolism/*pharmacology Serine Proteinase Inhibitors/pharmacology Serpins/genetics/metabolism Sodium/*metabolism Sodium Channels/genetics/*metabolism Terbutaline/pharmacology Trypsin/pharmacology Water/metabolism
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