title: Stimulation of suicidal erythrocyte death by PRIMA-1.
abstract: BACKGROUND
  The anticarcinogenic drug PRIMA-1 (p53 reactivation and induction of massive apoptosis 1) induces suicidal death of tumor cells, an effect in large part attributed to the up-regulation of the proapoptotic transcription factor p53. Erythrocytes are lacking gene transcription but are nevertheless able to enter eryptosis, a suicidal erythrocyte death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Stimulators of eryptosis include increase of cytosolic Ca(2+)-activity ([Ca(2+)]i) and ceramide formation. The present study tested whether PRIMA-1 stimulates eryptosis.
  
  
  METHODS
  Phosphatidylserine exposure at the cell surface was estimated from annexin V binding, cell volume from forward scatter, [Ca(2+)]i from Fluo3-fluorescence, ceramide abundance from binding of specific antibodies, and ROS formation from DCFDA fluorescence.
  
  
  RESULTS
  A 48 h exposure of human erythrocytes to PRIMA-1 (25 µM) significantly increased the percentage of annexin-V-binding cells without significantly influencing [Ca(2+)]i or forward scatter. PRIMA-1 (100 µM) induced annexin-V-binding was not significantly blunted by removal of extracellular Ca(2+) or by the caspase-3 inhibitor zVAD. PRIMA-1 (100 µM) further increased the ceramide abundance at the cell surface and ROS formation.
  
  
  CONCLUSIONS
  PRIMA-1 stimulates phosphatidylserine translocation at the erythrocyte cell membrane, an effect at least partially due to up-regulation of ceramide abundance and ROS formation.
authors.affiliation: Department of Biological and Environmental Sciences, University of Messina, Agata-Messina, Italy.
authors.forename: Caterina
authors.initials: C
authors.lastname: Faggio
authors.forename: Kousi
authors.initials: K
authors.lastname: Alzoubi
authors.forename: Salvatrice
authors.initials: S
authors.lastname: Calabrò
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D001372:Aza Compounds / Q000494:pharmacology*; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D002518:Ceramides / Q000378:metabolism; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D010718:Phosphatidylserines / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D017304:Annexin A5; D001372:Aza Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002518:Ceramides; D010718:Phosphatidylserines; D017382:Reactive Oxygen Species; C451219:2,2-bis(hydroxymethyl)-1-azabicyclo(2,2,2,)octan-3-one; D002118:Calcium