title: Three-dimensional culture and characterization of mononuclear cells from human bone marrow.
abstract: OBJECTIVE
  The diverse phenotypic changes and clinical and economic disadvantages associated with the monolayer expansion of bone marrow-derived mesenchymal stromal cells (MSCs) have focused attention on the development of one-step intraoperative cells therapies and homing strategies. The mononuclear cell fraction of bone marrow, inclusive of discrete stem cell populations, is not well characterized, and we currently lack suitable cell culture systems in which to culture and investigate the behavior of these cells.
  
  
  METHODS
  Human bone marrow-derived mononuclear cells were cultured within fibrin for 2 weeks with or without fibroblast growth factor-2 supplementation. DNA content and cell viability of enzymatically retrieved cells were determined at days 7 and 14. Cell surface marker profiling and cell cycle analysis were performed by means of multi-color flow cytometry and a 5-ethynyl-2'-deoxyuridine incorporation assay, respectively.
  
  
  RESULTS
  Total mononuclear cell fractions, isolated from whole human bone marrow, was successfully cultured in fibrin gels for up to 14 days under static conditions. Discrete niche cell populations including MSCs, pericytes and hematopoietic stem cells were maintained in relative quiescence for 7 days in proportions similar to that in freshly isolated cells. Colony-forming unit efficiency of enzymatically retrieved MSCs was significantly higher at day 14 compared to day 0; and in accordance with previously published works, it was fibroblast growth factor-2-dependant.
  
  
  CONCLUSIONS
  Fibrin gels provide a simple, novel system in which to culture and study the complete fraction of bone marrow-derived mononuclear cells and may support the development of improved bone marrow cell-based therapies.
authors: AO Research Institute Davos, Davos Platz, Switzerland.
authors: Jennifer J
authors: JJ
authors: Bara
authors: AO Research Institute Davos, Davos Platz, Switzerland.
authors: Marietta
authors: M
authors: Herrmann
authors: AO Research Institute Davos, Davos Platz, Switzerland.
authors: Ursula
authors: U
authors: Menzel
authors: Department of Orthopaedic Surgery, Inselspital, Bern University Hospital and University of Bern, Bern, Switzerland.
authors: Lorin
authors: L
authors: Benneker
authors: AO Research Institute Davos, Davos Platz, Switzerland.
authors: Mauro
authors: M
authors: Alini
authors: AO Research Institute Davos, Davos Platz, Switzerland. Electronic address: martin.stoddart@aofoundation.org.
authors: Martin J
authors: MJ
authors: Stoddart
mesh_terms: D001854:Bone Marrow Cells / Q000166:cytology*; D018929:Cell Culture Techniques; D002469:Cell Separation; D002470:Cell Survival; D002478:Cells, Cultured; D005337:Fibrin / Q000378:metabolism*; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D009000:Monocytes / Q000166:cytology*; D020286:Pericytes / Q000166:cytology
chemical_list: D005337:Fibrin
keywords: FGF(2); bone marrow; bone marrow niche; cell transplantation; mesenchymal stem cells; mononuclear cells