title: Circulating angiogenic cells can be derived from cryopreserved peripheral blood mononuclear cells.
abstract: BACKGROUND
  Cell transplantation for regenerative medicine has become an appealing therapeutic method; however, stem and progenitor cells are not always freshly available. Cryopreservation offers a way to freeze cells as they are generated, for storage and transport until required for therapy. This study was performed to assess the feasibility of cryopreserving peripheral blood mononuclear cells (PBMCs) for the subsequent in vitro generation of their derived therapeutic population, circulating angiogenic cells (CACs).
  
  
  METHODS
  PBMCs were isolated from healthy human donors. Freshly isolated cells were either analyzed immediately or cryopreserved in media containing 6% plasma serum and 5% dimethyl sulfoxide. PBMCs were thawed after being frozen for 1 (early thaw) or 28 (late thaw) days and analyzed, or cultured for 4 days to generate CACs. Analysis of the cells consisted of flow cytometry for viability and phenotype, as well as functional assays for their adhesion and migration potential, cytokine secretion, and in vivo angiogenic potential.
  
  
  RESULTS
  The viability of PBMCs and CACs as well as their adhesion and migration properties did not differ greatly after cryopreservation. Phenotypic changes did occur in PBMCs and to a lesser extent in CACs after freezing; however the potent CD34(+)VEGFR2(+)CD133(+) population remained unaffected. The derived CACs, while exhibiting changes in inflammatory cytokine secretion, showed no changes in the secretion of important regenerative and chemotactic cytokines, nor in their ability to restore perfusion in ischemic muscle.
  
  
  CONCLUSIONS
  Overall, it appears that changes do occur in cryopreserved PBMCs and their generated CACs; however, the CD34(+)VEGFR2(+)CD133(+) progenitor population, the secretion of pro-vasculogenic factors, and the in vivo angiogenic potential of CACs remain unaffected by cryopreservation.
authors.affiliation: Division of Cardiac Surgery, University of Ottawa Heart Institute, Ottawa, Canada.
authors.forename: Tanja
authors.initials: T
authors.lastname: Sofrenovic
authors.forename: Kimberly
authors.initials: K
authors.lastname: McEwan
authors.forename: Suzanne
authors.initials: S
authors.lastname: Crowe
authors.forename: Jenelle
authors.initials: J
authors.lastname: Marier
authors.forename: Robbie
authors.initials: R
authors.lastname: Davies
authors.forename: Erik J
authors.initials: EJ
authors.lastname: Suuronen
authors.forename: Drew
authors.initials: D
authors.lastname: Kuraitis
mesh_terms: D000071916:AC133 Antigen; D015703:Antigens, CD / Q000096:biosynthesis; D018952:Antigens, CD34 / Q000096:biosynthesis; D002448:Cell Adhesion; D002465:Cell Movement; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods*; D005434:Flow Cytometry / Q000379:methods; D006023:Glycoproteins / Q000096:biosynthesis; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D037102:Lectins / Q000737:chemistry; D007963:Leukocytes, Mononuclear / Q000166:cytology*; D008077:Lipoproteins, LDL / Q000378:metabolism; D018919:Neovascularization, Physiologic; D010455:Peptides; D010641:Phenotype; D013997:Time Factors; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000096:biosynthesis
chemical_list: D000071916:AC133 Antigen; D015703:Antigens, CD; D018952:Antigens, CD34; D006023:Glycoproteins; D007074:Immunoglobulin G; D037102:Lectins; D008077:Lipoproteins, LDL; C000606023:PROM1 protein, human; D010455:Peptides; D040301:Vascular Endothelial Growth Factor Receptor-2