title: Epidermal growth factor, basic fibroblast growth factor and platelet-derived growth factor-bb can substitute for fetal bovine serum and compete with human platelet-rich plasma in the ex vivo expansion of mesenchymal stromal cells derived from adipose tissue.
abstract: OBJECTIVE
  Human mesenchymal stromal cells (MSC) are multipotent cells possessing self-renewal capacity, long-term viability and multilineage potential. We analyzed the effect of four different medium supplements on the expansion and differentiation of adipose tissue-derived MSC (ADSC) in order to avoid the use of xenogeneic serum.
  
  
  METHODS
  We compared fetal bovine serum (FBS) with 10% human platelet-rich plasma (hPRP), 3% human platelet-poor plasma (hPPP) and with a cytokine cocktail composed of epidermal growth factor (EGF), basic fibroblast growth factor (bFGF) and platelet-derived growth factor-bb (PDGFbb) added to 3% hPPP. This mixture was developed testing EGF, bFGF, granulocyte-colony-stimulating factor (G-CSF), hepatocyte growth factor (HGF), insulin-like growth factor (IGF-I), PDGFbb and transforming growth factor (TGF)-β1 added alone or in combination with hPPP.
  
  
  RESULTS
  Our data demonstrate that the addition of EGF, bFGF and PDGFbb, in a medium supplemented with hPPP, obtainable from 150-200 mL whole autologous blood, supports ADSC expansion better than FBS, as confirmed by cumulative population doublings (cPD; 15.0 ± 0.5 versus 9.4 ± 2.8). The addition of human platelet-rich plasma (hPRP) further improved ADSC proliferation (cPD 20.0 ± 1.2), but the achievement of hPRP presented a major drawback, requiring 1000-1200 mL autologous or donor whole blood. The medium supplements did not influence ADSC phenotype: they expressed CD105, CD90 and CD44 lacking hematopoietic antigens. The exposure to the proposed cocktail or to hPRP increased adipogenic and osteogenic differentiation.
  
  
  CONCLUSIONS
  The addition of EGF, bFGF and PDGFbb to hPPP could ensure a sufficient number of ADSC for clinical applications, avoiding the use of animal serum and representing a novel approach in regenerative medicine.
authors.affiliation: Department of Cell Therapy and Hematology, San Bortolo Hospital, Vicenza, Italy.
authors.forename: Katia
authors.initials: K
authors.lastname: Chieregato
authors.forename: Silvia
authors.initials: S
authors.lastname: Castegnaro
authors.forename: Domenico
authors.initials: D
authors.lastname: Madeo
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Astori
authors.forename: Maurizio
authors.initials: M
authors.lastname: Pegoraro
authors.forename: Francesco
authors.initials: F
authors.lastname: Rodeghiero
mesh_terms: D050156:Adipogenesis / Q000187:drug effects; D000273:Adipose Tissue / Q000166:cytology*; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D002417:Cattle; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D004815:Epidermal Growth Factor / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010012:Osteogenesis / Q000187:drug effects; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D053657:Platelet-Rich Plasma / Q000378:metabolism*; D044968:Regenerative Medicine / Q000639:trends
chemical_list: D015703:Antigens, CD; D010982:Platelet-Derived Growth Factor; D016222:Fibroblast Growth Factor 2; D004815:Epidermal Growth Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor