title: [Na+]i/[K+]i -independent death of ouabain-treated renal epithelial cells is not mediated by Na+,K+ -ATPase internalization and de novo gene expression.
abstract: The cytotoxic effect of long-term exposure of renal epithelial cells to ouabain and other cardiotonic steroids (CTS) is mediated by the interaction of these compounds with Na(+),K(+)-ATPase but is independent of the inhibition of Na(+),K(+)-ATPase-mediated ion fluxes. Sustained application of CTS also leads to Na(+),K(+)-ATPase endocytosis and its translocation into the nuclei that might trigger the cell death machinery via the regulation of gene expression. This study examines the role of Na(+),K(+)-ATPase internalization and de novo gene expression in the death of ouabain-treated C7-Madin-Darby canine kidney (MDCK) cells derived from distal tubules of the MDCK. In these cells, 6-h exposure to 3 microM ouabain led to the internalization of approximately 50% of plasmalemmal Na(+),K(+)-ATPase. Prolonged incubation in a K(+)-free medium abolished ouabain-induced Na(+),K(+)-ATPase internalization but did not affect the cytotoxic action of ouabain seen after 18-h incubation. Previously, it was shown that CTS-induced Na(+),K(+)-ATPase internalization is mediated by its interaction with Src within caveolae. Neither caveolae damage by cholesterol depletion with methyl-beta-cyclodextrin nor Src inhibition with 4-amino-5(4-chlorophenyl)-7-(t-butyl)pyrazol[3,4-d]pyridine affected the death of ouabain-treated C7-MDCK cells. Actinomycin D at the 0.1-microg/ml concentration almost completely abolished ribonucleic acid synthesis but did not protect C7-MDCK cells from the cytotoxic action of ouabain. Our results show that neither Na(+),K(+)-ATPase endocytosis nor de novo gene expression contributes to Na(+)(i), K(+)(i)-independent cell death signaling evoked by prolonged exposure to CTS.
authors.affiliation: Research Centre, Centre Hospitalier de l'Université de Montréal, Technôpole Angus, 2901 Rachel East, Montreal, Quebec H1W 4A4, Canada.
authors.forename: Olga A
authors.initials: OA
authors.lastname: Akimova
authors.forename: Pavel
authors.initials: P
authors.lastname: Hamet
authors.forename: Sergei N
authors.initials: SN
authors.lastname: Orlov
mesh_terms: D000818:Animals; D002316:Cardiotonic Agents / Q000633:toxicity*; D021941:Caveolae / Q000187:drug effects / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002460:Cell Line; D002784:Cholesterol / Q000172:deficiency; D003609:Dactinomycin / Q000494:pharmacology; D004285:Dogs; D004705:Endocytosis / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000633:toxicity*; D004847:Epithelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D015870:Gene Expression / Q000187:drug effects; D007668:Kidney / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010042:Ouabain / Q000633:toxicity*; D011188:Potassium / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013997:Time Factors; D047392:beta-Cyclodextrins / Q000494:pharmacology; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: C412373:AG 1879; D002316:Cardiotonic Agents; D004791:Enzyme Inhibitors; D019384:Nucleic Acid Synthesis Inhibitors; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D003609:Dactinomycin; D010042:Ouabain; D002784:Cholesterol; D012964:Sodium; D019061:src-Family Kinases; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium