title: Isoform specificity of cardiac glycosides binding to human Na+,K+-ATPase alpha1beta1, alpha2beta1 and alpha3beta1.
abstract: Cardiac glycosides inhibit the Na(+),K(+)-ATPase and are used for the treatment of symptomatic heart failure and atrial fibrillation. In human heart three isoforms of Na(+),K(+)-ATPase are expressed: alpha(1)beta(1), alpha(2)beta(1) and alpha(3)beta(1). It is unknown, if clinically used cardiac glycosides differ in isoform specific affinities, and if the isoforms have specific subcellular localization in human cardiac myocytes. Human Na(+),K(+)-ATPase isoforms alpha(1)beta(1), alpha(2)beta(1) and alpha(3)beta(1) were expressed in yeast which has no endogenous Na(+),K(+)-ATPase. Isoform specific affinities of digoxin, digitoxin, beta-acetyldigoxin, methyldigoxin and ouabain were assessed in [(3)H]-ouabain binding assays in the absence or presence of K(+) (each n=5). The subcellular localizations of the Na(+),K(+)-ATPase isoforms were investigated in isolated human atrial cardiomyocytes by immunohistochemistry. In the absence of K(+), methyldigoxin (alpha(1)>alpha(3)>alpha(2)) and ouabain (alpha(1)=alpha(3)>alpha(2)) showed distinct isoform specific affinities, while for digoxin, digitoxin and beta-acetyldigoxin no differences were found. In the presence of K(+), also digoxin (alpha(2)=alpha(3)>alpha(1)) and beta-acetyldigoxin (alpha(1)>alpha(3)) had isoform specificities. A comparison between the cardiac glycosides demonstrated highly different affinity profiles for the isoforms. Immunohistochemistry showed that all three isoforms are located in the plasma membrane and in intracellular membranes, but only alpha(1)beta(1) and alpha(2)beta(1) are located in the T-tubuli. Cardiac glycosides show distinct isoform specific affinities and different affinity profiles to Na(+),K(+)-ATPase isoforms which have different subcellular localizations in human cardiomyocytes. Thus, in contrast to current notion, different cardiac glycoside agents may significantly differ in their pharmacological profile which could be of hitherto unknown clinical relevance.
authors.affiliation: Laboratory of Muscle Research and Molecular Cardiology, Department of Internal Medicine III, University of Cologne, Cologne, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Hauck
authors.forename: Tatjana
authors.initials: T
authors.lastname: Potter
authors.forename: Michaela
authors.initials: M
authors.lastname: Bartz
authors.forename: Thorsten
authors.initials: T
authors.lastname: Wittwer
authors.forename: Thorsten
authors.initials: T
authors.lastname: Wahlers
authors.forename: Uwe
authors.initials: U
authors.lastname: Mehlhorn
authors.forename: Georgios
authors.initials: G
authors.lastname: Scheiner-Bobis
authors.forename: Alicia A
authors.initials: AA
authors.lastname: McDonough
authors.forename: Wilhelm
authors.initials: W
authors.lastname: Bloch
authors.forename: Robert H G
authors.initials: RH
authors.lastname: Schwinger
authors.forename: Jochen
authors.initials: J
authors.lastname: Müller-Ehmsen
mesh_terms: D000818:Animals; D002301:Cardiac Glycosides / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D017506:H(+)-K(+)-Exchanging ATPase / Q000096:biosynthesis / Q000378:metabolism; D006333:Heart Failure / Q000378:metabolism / Q000628:therapy; D006801:Humans; D007150:Immunohistochemistry; D042541:Intracellular Space / Q000378:metabolism; D007527:Isoenzymes / Q000096:biosynthesis / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology; D010042:Ouabain / Q000378:metabolism; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: C488756:ATP1A3 protein, human; D002301:Cardiac Glycosides; D007527:Isoenzymes; D010042:Ouabain; C499076:ATP1A1 protein, human; C088641:ATP12A protein, human; D017506:H(+)-K(+)-Exchanging ATPase; D000254:Sodium-Potassium-Exchanging ATPase