title: Acid Sphingomyelinase Inhibition Prevents Hemolysis During Erythrocyte Storage.
abstract: OBJECTIVE
  During storage, units of human red blood cells (pRBCs) experience membrane destabilization and hemolysis which may cause harm to transfusion recipients. This study investigates whether inhibition of acid sphingomyelinase could stabilize erythrocyte membranes and prevent hemolysis during storage.
  
  
  METHODS
  Human and murine pRBCs were stored under standard blood banking conditions with and without the addition of amitriptyline, a known acid sphingomyelinase inhibitor. Hemoglobin was measured with an electronic hematology analyzer and flow cytometry was used to measure erythrocyte size, complexity, phosphatidylserine externalization, and band 3 protein expression.
  
  
  RESULTS
  Cell-free hemoglobin, a marker of hemolysis, increased during pRBC storage. Amitriptyline treatment decreased hemolysis in a dose-dependent manner. Standard pRBC storage led to loss of erythrocyte size and membrane complexity, increased phosphatidylserine externalization, and decreased band 3 protein integrity as determined by flow cytometry. Each of these changes was reduced by treatment with amitriptyline. Transfusion of amitriptyline-treated pRBCs resulted in decreased circulating free hemoglobin.
  
  
  CONCLUSIONS
  Erythrocyte storage is associated with changes in cell size, complexity, membrane molecular composition, and increased hemolysis. Acid sphingomyelinase inhibition reduced these changes in a dose-dependent manner. Our data suggest a novel mechanism to attenuate the harmful effects after transfusion of aged blood products.
authors.affiliation: Department of Surgery and Institute for Military Medicine, University of Cincinnati, Cincinnati, USA.
authors.forename: Richard S
authors.initials: RS
authors.lastname: Hoehn
authors.forename: Peter L
authors.initials: PL
authors.lastname: Jernigan
authors.forename: Alex L
authors.initials: AL
authors.lastname: Chang
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Edwards
authors.forename: Charles C
authors.initials: CC
authors.lastname: Caldwell
authors.forename: Erich
authors.initials: E
authors.lastname: Gulbins
authors.forename: Timothy A
authors.initials: TA
authors.lastname: Pritts
mesh_terms: D000639:Amitriptyline / Q000494:pharmacology; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism; D001803:Blood Transfusion / Q000379:methods; D004305:Dose-Response Relationship, Drug; D004910:Erythrocyte Membrane / Q000187:drug effects / Q000378:metabolism; D004912:Erythrocytes / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D005434:Flow Cytometry; D006454:Hemoglobins / Q000378:metabolism; D006461:Hemolysis / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D008297:Male; D008810:Mice, Inbred C57BL; D010718:Phosphatidylserines / Q000378:metabolism; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014021:Tissue Preservation / Q000379:methods*
chemical_list: D001457:Anion Exchange Protein 1, Erythrocyte; D006454:Hemoglobins; D010718:Phosphatidylserines; D000639:Amitriptyline; C112980:acid sphingomyelinase-1; D013108:Sphingomyelin Phosphodiesterase