title: Optimized Cryopreservation and Banking of Human Bone-Marrow Fragments and Stem Cells.
abstract: Adult mesenchymal stem cells are a promising source for cell therapies and tissue engineering applications. Current procedures for banking of human bone-marrow mesenchymal stem cells (hBM-MSCs) require cell isolation and expansion, and thus the use of large amounts of animal sera. However, animal-derived culture supplements have the potential to trigger infections and severe immune reactions. The aim of this study was to investigate an optimized method for cryopreservation of human bone-marrow fragments for application in cell banking procedures where stem-cell expansion and use are not immediately needed. Whole trabecular fragments enclosing the bone marrow were stored in liquid nitrogen for 1 year in a cryoprotective solution containing a low concentration of dimethyl sulfoxide and a high concentration of human serum (HuS). After thawing, the isolation, colony-forming-unit ability, proliferation, morphology, stemness-related marker expression, cell senescence, apoptosis, and multi-lineage differentiation potential of hBM-MSCs were tested in media containing HuS compared with hBM-MSCs isolated from fresh fragments. Human BM-MSCs isolated from cryopreserved fragments expressed MSC markers until later passages, had a good proliferation rate, and exhibited the capacity to differentiate toward osteogenic, adipogenic, and myogenic lineages similar to hBM-MSCs isolated from fresh fragments. Moreover, the cryopreservation method did not induce cell senescence or cell death. These results imply that minimal processing may be adequate for the banking of tissue samples with no requirement for the immediate isolation and use of hBM-MSCs, thus limiting cost and the risk of contamination, and facilitating banking for clinical use. Furthermore, the use of HuS for cryopreservation and expansion/differentiation has the potential for clinical application in compliance with good manufacturing practice standards.
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .|2 Dipartimento Sperimentale Interaziendale, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Gianluca
authors.initials: G
authors.lastname: Carnevale
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Alessandra
authors.initials: A
authors.lastname: Pisciotta
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Massimo
authors.initials: M
authors.lastname: Riccio
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Sara
authors.initials: S
authors.lastname: De Biasi
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Lara
authors.initials: L
authors.lastname: Gibellini
authors.affiliation: 2 Dipartimento Sperimentale Interaziendale, University of Modena and Reggio Emilia , Modena, Italy .|3 Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Adriano
authors.initials: A
authors.lastname: Ferrari
authors.affiliation: 2 Dipartimento Sperimentale Interaziendale, University of Modena and Reggio Emilia , Modena, Italy .|4 Department of Medical and Surgical Sciences for Children and Adults, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Giovanni B
authors.initials: GB
authors.lastname: La Sala
authors.affiliation: 5 Oro-Maxillo-Facial Department, AUSL Baggiovara , Modena, Italy .
authors.forename: Giacomo
authors.initials: G
authors.lastname: Bruzzesi
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .|2 Dipartimento Sperimentale Interaziendale, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Andrea
authors.initials: A
authors.lastname: Cossarizza
authors.affiliation: 1 Department of Surgery, Medicine, Dentistry and Morphological Sciences, University of Modena and Reggio Emilia , Modena, Italy .|2 Dipartimento Sperimentale Interaziendale, University of Modena and Reggio Emilia , Modena, Italy .
authors.forename: Anto
authors.initials: A
authors.lastname: de Pol
mesh_terms: D017209:Apoptosis; D018070:Biological Specimen Banks; D001853:Bone Marrow; D049109:Cell Proliferation; D002478:Cells, Cultured; D016922:Cellular Senescence; D015925:Cryopreservation / Q000379:methods*; D003470:Culture Media; D006412:Hematopoietic Stem Cells; D006801:Humans; D016130:Immunophenotyping; D008297:Male
chemical_list: D003470:Culture Media