title: Regulation of myofibroblast differentiation by cardiac glycosides.
abstract: Myofibroblast differentiation is a key process in pathogenesis of fibrotic diseases. Cardiac glycosides (ouabain, digoxin) inhibit Na(+)-K(+)-ATPase, resulting in increased intracellular [Na(+)]-to-[K(+)] ratio in cells. Microarray analysis suggested that increased intracellular [Na(+)]/[K(+)] ratio may promote the expression of cyclooxygenase-2 (COX-2), a critical enzyme in the synthesis of prostaglandins. Given antifibrotic effects of prostaglandins through activation of protein kinase A (PKA), we examined if cardiac glycosides stimulate COX-2 expression in human lung fibroblasts and how they affect myofibroblast differentiation. Ouabain stimulated a profound COX-2 expression and a sustained PKA activation, which was blocked by COX-2 inhibitor or by COX-2 knockdown. Ouabain-induced COX-2 expression and PKA activation were abolished by the inhibitor of the Na(+)/Ca(2+) exchanger, KB-R4943. Ouabain inhibited transforming growth factor-β (TGF-β)-induced Rho activation, stress fiber formation, serum response factor activation, and the expression of smooth muscle α-actin, collagen-1, and fibronectin. These effects were recapitulated by an increase in intracellular [Na(+)]/[K(+)] ratio through the treatment of cells with K(+)-free medium or with digoxin. Although inhibition of COX-2 or of the Na(+)/Ca(2+) exchanger blocked ouabain-induced PKA activation, this failed to reverse the inhibition of TGF-β-induced Rho activation or myofibroblast differentiation by ouabain. Together, these data demonstrate that ouabain, through the increase in intracellular [Na(+)]/[K(+)] ratio, drives the induction of COX-2 expression and PKA activation, which is accompanied by a decreased Rho activation and myofibroblast differentiation in response to TGF-β. However, COX-2 expression and PKA activation are not sufficient for inhibition of the fibrotic effects of TGF-β by ouabain, suggesting that additional mechanisms must exist.
authors: Section of Pulmonary and Critical Care Medicine, Department of Medicine, the University of Chicago, Chicago, Illinois;
authors: Jennifer
authors: J
authors: La
authors: Section of Pulmonary and Critical Care Medicine, Department of Medicine, the University of Chicago, Chicago, Illinois;
authors: Eleanor B
authors: EB
authors: Reed
authors: Laboratory of Biomembranes, Faculty of Biology, Lomonosov Moscow State University, Moscow, Russian Federation; and.
authors: Svetlana
authors: S
authors: Koltsova
authors: Laboratory of Biomembranes, Faculty of Biology, Lomonosov Moscow State University, Moscow, Russian Federation; and.
authors: Olga
authors: O
authors: Akimova
authors: Section of Pulmonary and Critical Care Medicine, Department of Medicine, the University of Chicago, Chicago, Illinois;
authors: Robert B
authors: RB
authors: Hamanaka
authors: Section of Pulmonary and Critical Care Medicine, Department of Medicine, the University of Chicago, Chicago, Illinois;
authors: Gökhan M
authors: GM
authors: Mutlu
authors: Laboratory of Biomembranes, Faculty of Biology, Lomonosov Moscow State University, Moscow, Russian Federation; and Siberian State Medical University, Tomsk, Russian Federation.
authors: Sergei N
authors: SN
authors: Orlov
authors: Section of Pulmonary and Critical Care Medicine, Department of Medicine, the University of Chicago, Chicago, Illinois; ndulin@medicine.bsd.uchicago.edu.
authors: Nickolai O
authors: NO
authors: Dulin
mesh_terms: D002301:Cardiac Glycosides / Q000494:pharmacology*; D002454:Cell Differentiation; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism; D004077:Digoxin / Q000494:pharmacology*; D004789:Enzyme Activation; D015870:Gene Expression; D006801:Humans; D054990:Idiopathic Pulmonary Fibrosis / Q000473:pathology; D008168:Lung / Q000473:pathology; D058628:Myofibroblasts / Q000187:drug effects / Q000502:physiology*; D010042:Ouabain / Q000494:pharmacology*
chemical_list: D002301:Cardiac Glycosides; D010042:Ouabain; D004077:Digoxin; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: cyclooxygenase-2; digoxin; intracellular sodium-to-potassium ion ratio; ouabain