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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Calcified tissue international 55 (1994), S. 120-127 
    ISSN: 1432-0827
    Keywords: Mineralization ; In vitro ; Stromal osteoblasts ; 3-D culture
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Physics
    Notes: Abstract The present study describes a new three-dimensional (3-D) culture system that enables the maintenance and phenotypic expression of bone marrow stromal osteoblasts. This culture substratum is advantageous in that it provides suitable conditions for attachment, growth, and differentiation of cells forming 3-D layers. The MBA-15 cell line was grown in unlimited quantities on 3-D Fibro-Cel carriers. These cells mineralized when exposed to ascorbic acid and β-glycerophosphate (βGP). Under these mineralization conditions, mRNA expressions of procollagen α2(I) and [3H]-proline-labeled protein were increased. The expression of mRNA for osteonectin and to a lesser extent, for osteopontin was increased, whereas alkaline phosphatase and biglycan remained unaffected under similar conditions. Exposure of mineralizing cultures to dexamethasone reduced mRNA of procollagen α2(I) and osteonectin to control level. Scanning electron microscopy revealed that cells were grown along the fabric's fibers and produced collagen fiberils. Under appropriate conditions, extensive mineralization had taken place. The mineralization process involves the formation of calcospherites, and correlates with an increase in calcium content. The Fibro-Cel carriers enable formation of 3-D architecture and mineralized tissue in vitro.
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  • 2
    ISSN: 0730-2312
    Keywords: vitamin A ; growth factors ; marrow stromal osteoblasts ; bone matrix proteins ; CD10/NEP ; neutral endopeptidase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: The effects of retinoic acid (RA) on the expression of osteoblastic-related cell makers was examined. A marrow and osteogenic cell line, MBA-15, was analyzed by Northern blotting for the expression of bone matrix proteins. These cells constituentively express mRNA encoding for procolllagen a2 (1), osteonectin, osteopontin, biglycan and alkaline phosphatase (ALK-P). Gene expression was unchanged in response to RA triggering for 24hr. Furthermore, cell growth and enzymatic activities of ALK-P and neutral endopeptidase (CD10/NEP) were studied. These parameters were examined in MBA-15 and clonal populations representing different stages of differentiation. The cell's growth rate was unchanged, while ALK-P activity was greatly increased during the culture period under RA treatment in MBA-15 and in the clonal cell lines examined while CD10/NEP activity dispalyed a different pattern. MBA-15.4, a presosteoblast cell ine, exhibited an inhibition in CD10/NEP activity at the beginning of the culture period, reaching basal level with time. This activity was greatly increased over control level in MBA-15.6, a mature stage of osteoblasts. Furthermore, the response of cell lines to various growth factors was tested subsequent to priming the cultures with RA. A synergistic effect was monitored for ALK-P activity in MBA-15.4 and MBA-15.6 cells under rh-bone morphogenic protein (BMP-2) and purified osteogenin (BMP-3), and an antagonist effect was measured when cells were exposed to transforming growth factor β (TGFβ). Contrarily, BMP-2 and BMP-3 inhibited the CD10/NEP activity that had remained unchanged following priming of the cell with RA. Insulin-like growth factor 1 (IGF-1) and basic fibroblast growth factors (bFGF) did not affect either ALK-P nor CD10/NEP activities in both cloned cells. Cellular response to bone-seeking hormone, parathyroid hormone (PTH), and prostaglandin E2 (PGE2) was monitored by activation of intracellular cAMP. Treatment with RA caused a dramatic increase in MBA-15.6 cell responses to PTH and PGE2 but no significant effects could be observed in other clonal lines.
    Additional Material: 6 Ill.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 62 (1996), S. 476-483 
    ISSN: 0730-2312
    Keywords: stromal osteoblasts ; dexamethasone ; attachment ; growth factors ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: The clonal subtypes of cells in the osteogenic family represented by fibroblastoid MBA-15.33, preosteoblast MBA-15.4, and mature osteoblastic MBA-15.6 cells were used to study the effects of glucocorticoid (dexamethasone). The role of dexamethasone was monitored on cell attachment when plated on various protein substrata (BSA, collagen I, and Matrigel). A 24 h exposure of the cells to 10-6 M or 10-7 M dexamethasone differential affects their attachment preference. MBA-15.33 and MBA-15.4 cells increased their attachment capability on collagen I, while MBA-15.6 cells' attachment was inhibited. Pretreatment with (10-6 M) dexamethasone caused an increase in attachment on Matrigel by MBA-15.33 cells and to less extent by MBA-15.4 cells. Additionally, measurements of two enzymatic activities were monitored; one is alkaline phosphatase (ALK-P), and the second is neutral endopeptidase (CD10/NEP). MBA-15.33, MBA-15.4, and MBA-15.6 cells were exposed to dexamethasone or to various growth factors (bone morphogenic protein (BMP-2 and BMP-3), TGFβ, and IGF-I). In some experiments, pretreatment of cells by dexamethasone was followed by exposure to the growth factors. The cells' challenged cellular responses were not uniform and revealed a differential pattern when their ALK-P and CD10/NEP enzymatic activities were measured. © 1996 Wiley-Liss, Inc.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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