title: Culture of human mesenchymal stem cells using a candidate pharmaceutical grade xeno-free cell culture supplement derived from industrial human plasma pools.
abstract: BACKGROUND
  Fetal bovine serum (FBS) is an animal product used as a medium supplement. The animal origin of FBS is a concern if cultured stem cells are to be utilized for human cell therapy. Therefore, a substitute for FBS is desirable. In this study, an industrial, xeno-free, pharmaceutical-grade supplement for cell culture (SCC) under development at Grifols was tested for growth of human mesenchymal stem cells (hMSCs), cell characterization, and differentiation capacity.
  
  
  METHODS
  SCC is a freeze-dried product obtained through cold-ethanol fractionation of industrial human plasma pools from healthy donors. Bone marrow-derived hMSC cell lines were obtained from two commercial suppliers. Cell growth was evaluated by culturing hMSCs with commercial media or media supplemented with SCC or FBS. Cell viability and cell yield were assessed with an automated cell counter. Cell surface markers were studied by indirect immunofluorescence assay. Cells were cultured then differentiated into adipocytes, chondrocytes, osteoblasts, and neurons, as assessed by specific staining and microscopy observation.
  
  
  RESULTS
  SCC supported the growth of commercial hMSCs. Starting from the same number of seeded cells in two consecutive passages of culture with medium supplemented with SCC, hMSC yield and cell population doubling time were equivalent to the values obtained with the commercial medium and was consistent among lots. The viability of hMSCs was higher than 90%, while maintaining the characteristic phenotype of undifferentiated hMSCs (positive for CD29, CD44, CD90, CD105, CD146, CD166 and Stro-1; negative for CD14 and CD19). Cultured hMSCs maintained the potential for differentiation into adipocytes, chondrocytes, osteoblasts, and neurons.
  
  
  CONCLUSIONS
  The tested human plasma-derived SCC sustains the adequate growth of hMSCs, while preserving their differentiation capacity. SCC can be a potential candidate for cell culture supplement in advanced cell therapies.
authors.affiliation: Cell Culture and Virology Laboratory, Research & Development Biologics Industrial Group. Grifols, Carrer Llevant, 11, 08150 Parets del Vallès, Barcelona, Spain. josemaria.diez@grifols.com.
authors.forename: José M
authors.initials: JM
authors.lastname: Díez
authors.affiliation: Cell Culture and Virology Laboratory, Research & Development Biologics Industrial Group. Grifols, Carrer Llevant, 11, 08150 Parets del Vallès, Barcelona, Spain. ewa.bauman@grifols.com.
authors.forename: Ewa
authors.initials: E
authors.lastname: Bauman
authors.affiliation: Cell Culture and Virology Laboratory, Research & Development Biologics Industrial Group. Grifols, Carrer Llevant, 11, 08150 Parets del Vallès, Barcelona, Spain. rodrigo.gajardo@grifols.com.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Gajardo
authors.affiliation: Cell Culture and Virology Laboratory, Research & Development Biologics Industrial Group. Grifols, Carrer Llevant, 11, 08150 Parets del Vallès, Barcelona, Spain. juan.jorquera@grifols.com.
authors.forename: Juan I
authors.initials: JI
authors.lastname: Jorquera
mesh_terms: D017667:Adipocytes / Q000166:cytology; D050156:Adipogenesis / Q000187:drug effects / Q000502:physiology; D000328:Adult; D015415:Biomarkers / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology; D020219:Chondrogenesis / Q000187:drug effects / Q000502:physiology; D003470:Culture Media / Q000494:pharmacology*; D005260:Female; D006801:Humans; D008297:Male; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D008875:Middle Aged; D055495:Neurogenesis / Q000187:drug effects / Q000502:physiology; D009474:Neurons / Q000166:cytology; D010006:Osteoblasts / Q000166:cytology; D010012:Osteogenesis / Q000187:drug effects / Q000502:physiology; D055815:Young Adult
chemical_list: D015415:Biomarkers; D003470:Culture Media