Mineralization in osteoblast cultures: a light and electron microscopic study.

Osteoblasts isolated mechanically from newborn mouse calvaria produced a calcified matrix when cultured in the presence of 10 mM beta-glycerophosphate or 3 mM inorganic phosphate. The uncalcified matrix revealed numerous matrix vesicles scattered among collagen fibrils. The calcified matrix showed mineralized collagen fibrils and calcified nodules whose underlying organic matrix was detected after decalcification. These structures resembled those described in fetal and woven bone. In partially decalcified areas, calcification was shown to spread out from these structures along collagen fibrils. Alkaline phosphatase activity was found associated with the plasma membrane and matrix vesicles. X-ray diffraction analysis demonstrated that the mineral phase deposited in culture was hydroxyapatite. These observations which demonstrate that the isolated cells elaborate in culture a mineralized matrix with chemical and ultrastructural properties of woven bone further support the osteoblastic nature of the cells.

[1]  F. Ottensmeyer,et al.  Visualization of early intramembranous ossification by electron microscopic and spectroscopic imaging , 1984, The Journal of cell biology.

[2]  A. Ornoy,et al.  Ultrastructural studies on the origin and structure of matrix vesicles in bone of young rats. , 1980, Acta anatomica.

[3]  F. Glorieux,et al.  Osteoblasts isolated from mouse calvaria initiate matrix mineralization in culture , 1983, The Journal of cell biology.

[4]  W. Neuman,et al.  Mechanisms of calcification: role of collagen, polyphosphates, and phosphatase. , 1961, The American journal of physiology.

[5]  J. Aubin,et al.  Isolation of bone cell clones with differences in growth, hormone responses, and extracellular matrix production , 1982, The Journal of cell biology.

[6]  P. Robey,et al.  Fetal bovine bone cells synthesize bone-specific matrix proteins , 1984, The Journal of cell biology.

[7]  D. V. Cohn,et al.  Collagen synthesis in cultured osteoblast-like cells. , 1980, Archives of biochemistry and biophysics.

[8]  H. Anderson,et al.  A simple and defined method to study calcification by isolated matrix vesicles. Effect of ATP and vesicle phosphatase. , 1977, Biochimica et biophysica acta.

[9]  H. Broekhuyse,et al.  Proteoglycans synthesized by cultured mouse osteoblasts. , 1987, The Journal of biological chemistry.

[10]  C. G. Groot.  An electron microscopic examination for the presence of acid groups in the organic matrix of mineralization nodules in foetal bone. , 1982, Metabolic bone disease & related research.

[11]  N. Mitchell,et al.  The use of ruthenium and p-phenylenediamine to stain cartilage simultaneously for light and electron microscopy. , 1977, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[12]  S. Rodan,et al.  Evidence for preferential effects of parathyroid hormone, calcitonin and adenosine on bone and periosteum. , 1977, Endocrinology.

[13]  S. Seibert,et al.  Characterization of Osteoblast-like Cells from Fetal Rat Calvaria , 1980, Journal of dental research.

[14]  S. Yoshiki,et al.  A light and electron microscopic study of alkaline phosphatase activity in the early stage of dentinogenesis in the young rat. , 1971, Archives of oral biology.

[15]  A. Ornoy,et al.  Ultrastructural studies of initial stages of mineralization of long bones and vertebrae in human fetuses. , 1984, Acta anatomica.

[16]  N. Mitchell,et al.  Simultaneous localization of proteoglycan by light and electron microscopy using toluidine blue O. A study of epiphyseal cartilage. , 1976, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[17]  Y. Amagai,et al.  In vitro differentiation and calcification in a new clonal osteogenic cell line derived from newborn mouse calvaria , 1983, The Journal of cell biology.

[18]  A. van der Plas,et al.  Bone formation and calcification by isolated osteoblastlike cells , 1982, The Journal of cell biology.

[19]  Howard C. Tenenbaum.  Role of Organic Phosphate in Mineralization of Bone in vitro , 1981, Journal of dental research.

[20]  E. Bonucci.  The locus of initial calcification in cartilage and bone. , 1971, Clinical orthopaedics and related research.

[21]  D. Pease,et al.  An electron microscopic study of initial intramembranous osteogenesis. , 1969, The American journal of anatomy.

[22]  S. Doty.  Problems inherent in obtaining the alkaline phosphatase reaction. , 1980, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[23]  C. G. Groot.  Electron microscopical x-ray microanalysis of mineralization nodules and several other tissue compartments in fetal bone. , 1982, Metabolic bone disease & related research.

[24]  M. Boy-Lefevre,et al.  Mineralization in vitro of matrix formed by osteoblasts isolated by collagenase digestion. , 1985, Differentiation; research in biological diversity.

[25]  P. Boonekamp,et al.  Bone formation by isolated calvarial osteoblasts in syngeneic and allogeneic transplants: light microscopic observations. , 1983, The American journal of anatomy.

[26]  D. Brunette,et al.  Parathyroid Hormone- and Prostaglandin E1-Response in a Selected Population of Bone Cells After Repeated Subculture and Storage at ‒80 C , 1977 .

[27]  R. Luben,et al.  Biochemical characterization with parathormone and calcitonin of isolated bone cells: provisional identification of osteoclasts and osteoblasts. , 1976, Endocrinology.