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Understanding the mechanisms that control the proliferation and commitment of human stem cells into cells of the osteogenic lineage for the preservation of skeletal structure is of basic importance in bone physiology. This study examines some aspects of the differentiation in vitro of human bone marrow fibroblastic cells cultured in the absence (basal media) or presence of 1nM dexamethasone and 50 micrograms/ml ascorbate for 6, 10, 14, and 21 days. Northern blot analysis and in situ hybridisation with digoxygenin-labelled riboprobes for Type I collagen, osteocalcin, bone morphogenetic proteins 2 (BMP-2), and 4 (BMP-4) and the estrogen receptor alpha (ERalpha), together with immunocytochemical analysis of ERalpha expression and histochemical staining of alkaline phosphatase was performed. In basal media, alkaline phosphatase activity and collagen expressions were detected at day 6, ERalpha from day 10 and osteocalcin from day 10. In the presence of dexamethasone and ascorbate, cell proliferation and alkaline phosphatase were markedly stimulated over 10 to 14 days with a dramatic increase in the temporal expression of Type I collagen, ERalpha, and osteocalcin mRNAs in these cultures. Northern blot analysis showed cells cultured in basal media, expressed the highest levels of the mRNA for each marker protein at day 14, whereas in the presence of ascorbate and dexamethasone, the highest levels for alkaline phosphatase, ERalpha, osteocalcin, BMP-2, and BMP-4 were observed at day 21. ERalpha, BMP-2, and BMP-4 expression were found to correlate temporally with induction of the osteoblast phenotype as determined by alkaline phosphatase, collagen, and osteocalcin expression. These results give additional information on the development of the osteoblast phenotype from early fibroblastic stem cells and on the biological factors involved in this process. These studies suggest a role for estrogen and BMP-2 and -4 in the differentiation of osteoprogenitor cells.


Journal article


J cell biochem

Publication Date





382 - 392


Aged, Aged, 80 and over, Alkaline Phosphatase, Bone Morphogenetic Protein 2, Bone Morphogenetic Protein 4, Bone Morphogenetic Proteins, Cell Differentiation, Cells, Cultured, Collagen, Estrogen Receptor alpha, Gene Expression, Hematopoietic Stem Cells, Humans, In Situ Hybridization, Osteoblasts, Osteocalcin, RNA, Messenger, Receptors, Estrogen, Transforming Growth Factor beta