title: Stimulation of Suicidal Erythrocyte Death by the CDC25 Inhibitor NSC-95397.
abstract: OBJECTIVE
  The CDC25B inhibitor NSC-95397 triggers apoptosis of tumor cells and is thus considered for the treatment of malignancy. The substance is effective in part by modification of gene expression. Similar to apoptosis of nucleated cells erythrocytes may undergo eryptosis, the suicidal erythrocyte death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Eryptosis may be triggered by increase of cytosolic Ca2+ activity ([Ca2+]i), oxidative stress, ceramide, as well as activation of protein kinases. The present study explored, whether NSC-95397 induces eryptosis and, if so, to shed some light on the mechanisms involved.
  
  
  METHODS
  Phosphatidylserine exposure at the cell surface was estimated from annexin-V-binding, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, ROS formation from DCFDA dependent fluorescence, and ceramide abundance utilizing specific antibodies.
  
  
  RESULTS
  A 48 hours exposure of human erythrocytes to NSC-95397 significantly increased the percentage of annexin-V-binding cells (≥ 1 µM), significantly decreased forward scatter (≥ 2.5 µM), and significantly increased Fluo3-fluorescence (≥ 1 µM), DCFDA fluorescence (5 µM) and ceramide abundance (≥ 5 µM). The effect of NSC-95397 (5 µM) on annexin-V-binding was slightly, but significantly blunted by removal of extracellular Ca2+ and by addition of the protein kinase C inhibitor staurosporine (1 µM).
  
  
  CONCLUSIONS
  NSC-95397 triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part requiring entry of Ca2+ and activation of staurosporine sensitive kinase(s).
authors.affiliation: Department of Cardiology, Vascular Medicine and Physiology, Eberhard-Karls-University of Tuebingen, Tuebingen, Germany.
authors.forename: Mohamed
authors.initials: M
authors.lastname: Jemaà
authors.forename: Morena
authors.initials: M
authors.lastname: Mischitelli
authors.forename: Myriam
authors.initials: M
authors.lastname: Fezai
authors.forename: Mustafa
authors.initials: M
authors.lastname: Almasry
authors.forename: Caterina
authors.initials: C
authors.lastname: Faggio
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D002118:Calcium / Q000378:metabolism; D002518:Ceramides / Q000378:metabolism; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D000072817:Eryptosis / Q000187:drug effects*; D006801:Humans; D009285:Naphthoquinones / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects; D010718:Phosphatidylserines / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D012542:Scattering, Radiation; D020687:cdc25 Phosphatases / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: C451261:2,3-bis(2-hydroxyethylsulfanyl)-(1,4)naphthoquinone; D002518:Ceramides; D004791:Enzyme Inhibitors; D009285:Naphthoquinones; D010718:Phosphatidylserines; D047428:Protein Kinase Inhibitors; D017382:Reactive Oxygen Species; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D011493:Protein Kinase C; D020687:cdc25 Phosphatases; D002118:Calcium