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|Title:||Human trabecular bone-derived osteoblasts support human osteoclast formation in vitro in a defined, serum-free medium|
|Citation:||Journal of Cellular Physiology, 2005; 203(3):573-582|
|Gerald J. Atkins, Panagiota Kostakis, Katie J. Welldon, Cristina Vincent, David M. Findlay, Andrew C.W. Zannettino|
|Abstract:||While it has been assumed that osteoblasts in the human support osteoclast formation, in vitro evidence of this is currently lacking. We tested the ability of normal human trabecular bone-derived osteoblasts (NHBCs) to support osteoclast formation from human peripheral blood mononuclear cells (PBMC) in response to treatment with either 1alpha,25-dihydroxyvitamin D3 (1,25D) or parathyroid hormone (PTH), using a serum-replete medium previously used to support human osteoclast formation on a stroma of murine ST-2 cells. Under these conditions, NHBC did not support osteoclast formation, as assessed by morphological, histochemical, and functional criteria, despite our previous results demonstrating a link between induction of RANKL mRNA expression and NHBC phenotype in these media. We next tested a defined, serum-free medium (SDM) on NHBC phenotype, their expression of RANKL and OPG, and their ability to support osteoclast formation. SDM, containing dexamethasone (DEX) and 1,25D, induced phenotypic maturation of NHBC, based on the expression of STRO-1 and the bone/liver/kidney isoform of alkaline phosphatase (AP). PTH as a single factor did not induce phenotypic change. 1,25D and DEX induced the greatest ratio of RANKL:OPG mRNA, predictive of supporting osteoclast formation. Consistent with this, co-culture of NHBC with CD14+ PBMC, or bone marrow mononuclear cell (BMMC), or CD34+ BMMC precursors in SDM + 1,25D + DEX, resulted in functional osteoclast formation. Osteoclast formation also occurred in PTH + DEX stimulated co-cultures. Interestingly, SDM supplemented with recombinant RANKL (25-100 ng/ml) and M-CSF (25 ng/ml), did not induce osteoclast formation from any of the osteoclast precursor populations in stromal-free cultures, unlike serum-replete medium. This study demonstrates that under the appropriate conditions, adult human primary osteoblasts can support de novo osteoclast formation, and this model will enable the detailed study of the role of both cell types in this process.|
|Keywords:||Bone and Bones; Bone Marrow Cells; Cells, Cultured; Cell Line; Osteoclasts; Osteoblasts; Stromal Cells; Stem Cells; Animals; Humans; Mice; Calcitriol; Dexamethasone; Parathyroid Hormone; Alkaline Phosphatase; ADP-ribosyl Cyclase; Glycoproteins; Carrier Proteins; Membrane Glycoproteins; Receptors, Tumor Necrosis Factor; Receptors, Cytoplasmic and Nuclear; RNA, Messenger; Antigens, CD; Culture Media, Serum-Free; Coculture Techniques; Bone Remodeling; Cell Communication; Cell Differentiation; Cell Lineage; RANK Ligand; Receptor Activator of Nuclear Factor-kappa B; Osteoprotegerin; GPI-Linked Proteins|
|Description:||The definitive version may be found at www.wiley.com|
|Appears in Collections:||Orthopaedics and Trauma publications|
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