Osteoblast viability and differentiation with Me sub(2)SO as cryoprotectant compared to osteoblasts from fresh human iliac cancellous bone

The aim of this study was to compare the viability of human osteoblasts cryopreserved with Me sub(2)SO to that of fresh human iliac cancellous bone using cell culture techniques. Osteoblasts were obtained by spontaneous outgrowth of human iliac cancellous bone specimens in experiment I. In experimen...

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Veröffentlicht in:Cryobiology 2005-12, Vol.51 (3), p.311-321
Hauptverfasser: Reuther, T, Rohmann, D, Scheer, M, Kubler, A C
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Sprache:eng
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Zusammenfassung:The aim of this study was to compare the viability of human osteoblasts cryopreserved with Me sub(2)SO to that of fresh human iliac cancellous bone using cell culture techniques. Osteoblasts were obtained by spontaneous outgrowth of human iliac cancellous bone specimens in experiment I. In experiment II, human iliac cancellous bone was frozen with 10% Me sub(2)SO at -80 super(o)C for 2 weeks and osteoblasts grew spontaneously after thawing at 37 super(o)C by removing Me sub(2)SO with sucrose. The cells were grown in culture flasks containing DMEM as a culture medium, supplemented with 10% fetal calf serum. They were kept at 37 super(o)C in a humidified atmosphere of 95% air and 5% CO sub(2). Cells from the second passage were plated at a density of 5times10 super(3)cells/cm super(2) in 24-well plates. For detection of viability and differentiation, WST-1 assay, determination of alkaline phosphatase activity, concentration of procollagen I peptide, concentration of osteocalcin, and indirect immunofluorescence for osteopontin, collagen type I, integrin beta sub(1), and fibronectin were applied. Experiments were conducted at four stages of confluence (days 4, 7, 14, and 21 after plating the cells). Based on the results of this study, we conclude that osteoblast-like cells survived cryopreservation and synthesized a range of markers that were consistent with this cell type.
ISSN:0011-2240
DOI:10.1016/j.cryobiol.2005.08.007