Ouabain activates signaling pathways associated with cell death in human neuroblastoma.
Cardiotonic steroids (CTS) like ouabain are not only specific inhibitors of the sodium pump (Na(+),K(+)-ATPase), they also can influence various cytosolic signaling events in a hormone-like manner. In the neuroblastoma cell line SH-SY5Y ouabain triggers multiple signaling pathways. Within 30 min of incubation with 1 or 10 microM ouabain, SH-SY5Y cells generate reactive oxygen species to a level approximately 50% above control and show a modest but significant elevation in cytosolic [Ca(2+)] of about 25%. After 6 h of exposure, ouabain stimulates a series of anti-apoptotic actions in SH-SY5Y cells, including concentration-dependent phosphorylation of Erk1/2, Akt, and Bad. Nevertheless, at the same time this CTS also induces a series of events that inhibit retinoic acid-induced neuritogenesis and promote cell death. Both of these latter phenomena are possibly associated with the observed ouabain-induced reduction in the abundance of the anti-apoptotic proteins Bcl-XL and Bcl-2. In addition, ouabain treatment results in cytochrome c release into the cytosol and induces activation of caspase 3, events that point towards the stimulation of apoptotic pathways that are probably enhanced by the stimulation of p53 phosphorylation at Ser15 also observed in this study. These pathways may eventually lead to cell death: treatment with 10 nM ouabain results in a 20% decrease in cell number after 4 days of incubation and treatment with 1 microM ouabain decreases cells number by about 75%. The results obtained here emphasize the importance of further research in order to elucidate the various signalling cascades triggered by ouabain and possibly other CTS that are used in the treatment of heart failure and to identify their primary receptor(s).
Kulikov
Andrey
A
Institut für Biochemie und Endokrinologie, Fachbereich Veterinärmedizin, Justus-Liebig-Universität Frankfurter Str. 100, D-35392 Giessen, Germany.
Eva
Alexander
A
Kirch
Ulrike
U
Boldyrev
Alexander
A
Scheiner-Bobis
Georgios
G
D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D016501:Neurites / Q000187:drug effects / Q000502:physiology; D009447:Neuroblastoma / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology*; D010766:Phosphorylation; D020033:Protein Isoforms / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D000254:Sodium-Potassium-Exchanging ATPase / Q000187:drug effects / Q000378:metabolism; D014212:Tretinoin / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D051025:bcl-Associated Death Protein / Q000378:metabolism; D051020:bcl-X Protein / Q000378:metabolism
C494819:BAD protein, human; C494811:BCL2L1 protein, human; D020033:Protein Isoforms; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D016159:Tumor Suppressor Protein p53; D051025:bcl-Associated Death Protein; D051020:bcl-X Protein; D014212:Tretinoin; D010042:Ouabain; D045304:Cytochromes c; D051057:Proto-Oncogene Proteins c-akt; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D053148:Caspase 3; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium