title: Bone marrow-derived cells for cardiovascular cell therapy: an optimized GMP method based on low-density gradient improves cell purity and function.
abstract: BACKGROUND
  Cardiovascular cell therapy represents a promising field, with several approaches currently being tested. The advanced therapy medicinal product (ATMP) for the ongoing METHOD clinical study ("Bone marrow derived cell therapy in the stable phase of chronic ischemic heart disease") consists of fresh mononuclear cells (MNC) isolated from autologous bone marrow (BM) through density gradient centrifugation on standard Ficoll-Paque. Cells are tested for safety (sterility, endotoxin), identity/potency (cell count, CD45/CD34/CD133, viability) and purity (contaminant granulocytes and platelets).
  
  
  METHODS
  BM-MNC were isolated by density gradient centrifugation on Ficoll-Paque. The following process parameters were optimized throughout the study: gradient medium density; gradient centrifugation speed and duration; washing conditions.
  
  
  RESULTS
  A new manufacturing method was set up, based on gradient centrifugation on low density Ficoll-Paque, followed by 2 washing steps, of which the second one at low speed. It led to significantly higher removal of contaminant granulocytes and platelets, improving product purity; the frequencies of CD34+ cells, CD133+ cells and functional hematopoietic and mesenchymal precursors were significantly increased.
  
  
  CONCLUSIONS
  The methodological optimization described here resulted in a significant improvement of ATMP quality, a crucial issue to clinical applications in cardiovascular cell therapy.
authors.forename: Marina
authors.initials: M
authors.lastname: Radrizzani
authors.forename: Viviana
authors.initials: V
authors.lastname: Lo Cicero
authors.forename: Sabrina
authors.initials: S
authors.lastname: Soncin
authors.forename: Sara
authors.initials: S
authors.lastname: Bolis
authors.forename: Daniel
authors.initials: D
authors.lastname: Sürder
authors.forename: Tiziano
authors.initials: T
authors.lastname: Torre
authors.forename: Francesco
authors.initials: F
authors.lastname: Siclari
authors.forename: Tiziano
authors.initials: T
authors.lastname: Moccetti
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Vassalli
authors.forename: Lucia
authors.initials: L
authors.lastname: Turchetto
mesh_terms: D001854:Bone Marrow Cells / Q000166:cytology*; D002318:Cardiovascular Diseases / Q000628:therapy*; D002452:Cell Count; D002469:Cell Separation / Q000379:methods* / Q000592:standards*; D064987:Cell- and Tissue-Based Therapy; D002499:Centrifugation, Density Gradient; D006801:Humans; D016130:Immunophenotyping; D015203:Reproducibility of Results