title: Cell isolation procedures matter: a comparison of different isolation protocols of bone marrow mononuclear cells used for cell therapy in patients with acute myocardial infarction.
abstract: OBJECTIVE
  The recently published REPAIR-AMI and ASTAMI trial showed differences in contractile recovery of left ventricular function after infusion of bone marrow-derived cells in acute myocardial infarction. Since the trials used different protocols for cell isolation and storage (REPAIR-AMI: Ficoll, storage in X-vivo 10 medium plus serum; ASTAMI: Lymphoprep, storage in NaCl plus plasma), we compared the functional activity of BMC isolated by the two different protocols.
  
  
  RESULTS
  The recovery of total cell number, colony-forming units (CFU), and the number of mesenchymal stem cells were significantly reduced to 77 +/- 4%, 83 +/- 16%, and 65 +/- 15%, respectively, when using the ASTAMI protocol compared with the REPAIR protocol. The capacity of the isolated BMC to migrate in response to stromal cell-derived factor 1 (SDF-1) was profoundly reduced when using the ASTAMI cell isolation procedure (42 +/- 8% and 78 +/- 3% reduction in healthy and CAD-patient cells, respectively). Finally, infusion of BMC into a hindlimb ischaemia model demonstrated a significantly blunted blood-flow-recovery by BMC isolated with the ASTAMI protocol (54 +/- 6% of the effect obtained by REPAIR cells). Comparison of the individual steps identified the use of NaCl and plasma for cell storage as major factors for functional impairment of the BMC.
  
  
  CONCLUSIONS
  Cell isolation protocols have a major impact on the functional activity of bone marrow-derived progenitor cells. The assessment of cell number and viability may not entirely reflect the functional capacity of cells in vivo. Additional functional testing appears to be mandatory to assure proper cell function before embarking on clinical cell therapy trials.
authors.affiliation: Department of Molecular Cardiology, University of Frankfurt, Theodor-Stern-Kai 7, Frankfurt am Main, Germany.
authors.forename: Florian H
authors.initials: FH
authors.lastname: Seeger
authors.forename: Torsten
authors.initials: T
authors.lastname: Tonn
authors.forename: Nicola
authors.initials: N
authors.lastname: Krzossok
authors.forename: Andreas M
authors.initials: AM
authors.lastname: Zeiher
authors.forename: Stefanie
authors.initials: S
authors.lastname: Dimmeler
mesh_terms: D000818:Animals; D001783:Blood Flow Velocity; D001854:Bone Marrow Cells; D016026:Bone Marrow Transplantation / Q000379:methods*; D002469:Cell Separation / Q000379:methods*; D002985:Clinical Protocols; D005434:Flow Cytometry; D019650:Hematopoietic Stem Cell Mobilization / Q000379:methods; D006614:Hindlimb / Q000098:blood supply; D006801:Humans; D007511:Ischemia / Q000209:etiology; D051379:Mice; D009203:Myocardial Infarction / Q000628:therapy*; D010641:Phenotype