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  • 1
    Electronic Resource
    Electronic Resource
    A scanning electron microscopic investigation of in vitro osteogenesis (1980)
    Springer
    Calcified tissue international 30 (1980), S. 43-50 
    Details
    ISSN: 1432-0827
    Keywords: Osteogenesis ; In vitro ; Electron microscopy ; Mineralization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Physics
    Notes: Summary Chick limb mesenchymal cells differentiate into muscle, cartilage, fibrous, and bone tissue. Previous reports show that when stage 24 limb mesenchymal cells are cultured in vitro, chondrocytes, myocytes, fibrocytes, and osteoblasts can be identified on the basis of morphological and biochemical parameters. The study reported here demonstrates that phenotypic expression in culture seems to be dependent on the initial plating density, Scanning electron microscopic observations indicate that when stage 24 limb mesenchymal cells are initially seeded at high densities (5 × 106 cells per 35 mm culture dish), mounds of cells appear in culture. These mounds represent cartilage nodules composed of a fine fibrous matrix and chondrocytes, surrounded by a loose fibrous connective tissue matrix. Cultures initially plated at intermediate densities (2.0–2.5 × 106 cells/35 mm culture dish) produce a flattened layer of fibrocytes overlying a matrix of collagen fibers and calcium phosphate deposits as determined by electron-microprobe analysis; these observations are indicative of osteoblast expression. Cells seeded at this intermediate density appear larger and possess greater surface area than cells seeded at high density. It is suggested that conditions that permit such increased cell surface area coupled with a relative compaction due to cell crowding may provide conditions permissive for osteogenesis. Based on morphological criteria, it appears that chick limb mesenchymal cell osteogenesis in vitro is not associated with chondrogenesis but represents a separate route of phenotypic expression.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF02408605
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  • 2
    Electronic Resource
    Electronic Resource
    Avian osteoblast conditioned media stimulate bone resorption by targeting multinucleating osteoclast precursors (1992)
    Springer
    Calcified tissue international 51 (1992), S. 317-323 
    Details
    ISSN: 1432-0827
    Keywords: Osteoblast conditioned medium ; Osteoclast precursors ; Bone resorption
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Physics
    Notes: Summary Osteoblasts are thought to secrete factors that regulate the rate of osteoclastic bone resorption. We studied the effect of osteoblast conditioned medium on bone degradation by multinucleated osteoclast-like cells generated in vitro from mononuclear precursors and found that the medium stimulates bone degradation primarily through interactions with osteoclast precursors. The conditioned medium also stimulates expression of the osteoclast-specific antigen 121F. The increased bone degradation, but not increased 121F expression, is due to the conditioned medium maintaining activity of the osteoclast precursors. Although the osteoclast precursors exhibit the DNA fragmentation characteristic of apoptosis, the osteoblast conditioned medium does not prevent such fragmentation. Chicken macrophage growth factor neither mimics nor augments the ability of the conditioned medium to stimulate bone degradation. Studies of osteoclast generation or function should carefully consider whether the effects are dependent on the viability of the resorbing cells.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00334494
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  • 3
    Electronic Resource
    Electronic Resource
    Evidence for an immunological and functional relationship between superoxide dismutase and a high molecular weight osteoclast plasma membrane glycoprotein (1991)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 46 (1991), S. 331-344 
    Details
    ISSN: 0730-2312
    Keywords: bone resorption ; osteoclast ; superoxide dismutase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Large multinucleated osteoclasts are the major cells responsible for bone breakdown and have been reported to produce high levels of superoxides which may contribute to the process of bone resorption (Key et al.: J Bone and Mineral Res 4 [suppl. 1]:S206, 1989). Osteoclasts also possess high levels of superoxide dismutase, a protective enzyme capable of converting toxic superoxides to less toxic H2O2 (Fridovich: J Biol Chem 264:7761-7764, 1989). The amino acid sequence of manganese and/or iron superoxide dismutase has a conserved region which exhibits substantial homology with a fragment obtained from a high molecular weight osteoclast surface marker glycoprotein which is reactive with monoclonal antibody 121F. In this report, evidence is presented substantiating immunological, biochemical, and functional similarities between the osteoclast membrane antigen recognized by the 121F monoclonal antibody and superoxide dismutase. Western blot and immunoprecipitation studies show that a monospecific polyclonal antibody generated against immunoaffinity purified antigen is cross-reactive with superoxide dismutase. Both the antigen and a high molecular weight superoxide dismutase activity have been detected in osteoclast plasma membrane preparations. The levels of superoxide dismutase activity and the membrane antigen have been found to correlate in antigen depletion studies and in western blots probing osteoclasts and closely related marrow-derived giant cells. Moreover, regions of osteoclast superoxide dismutase activity identified by electrophoretic zymogram analysis have been shown by gel electrophoresis and western blots to contain the high molecular weight antigen, or complexes of the antigen with the 121F monoclonal antibody when these were premixed prior to nondenaturing electrophoresis. It is proposed that the osteoclast plasma membrane possesses a high molecular weight superoxide dismutase activity. Furthermore, it appears that this activity is associated with the osteoclast antigen recognized by the 121F monoclonal antibody.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcb.240460408
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  • 4
    Electronic Resource
    Electronic Resource
    Bone cell function, regulation, and communication: A role for nitric oxide (1995)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 57 (1995), S. 399-408 
    Details
    ISSN: 0730-2312
    Keywords: nitric oxide ; bone remodeling ; free radicals ; osteoclasts ; osteoblasts ; cytokines ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: A large array of factors serve as vital communication links between cells and the characterization, regulation, and mechanisms of action of such factors are topics of intense research efforts. Most intercellular messenger molecules which have been described over the years are represented by proteins, small peptides, amino acids or their derivatives, ions, lipid metabolites, or steroids. However, a small uncharged free radical, nitric oxide, has recently garnered much attention as a potent multifunctional signal molecule with widespread actions within and between diverse tissues. Biochemical, molecular, and regulatory studies of the family of enzymes responsible for nitric oxide synthesis, nitric oxide synthases, have established that there are at least three distinct isoforms of this enzyme which are differentially expressed and regulated in various cells or tissues. Modulation of these isoenzyme levels or activities by diverse signals is mediated via transcriptional, translational, and/or post-translational mechanisms, and consequently, alterations in such control may influence normal or pathological processes. Nitric oxide appears to exert pronounced effects on skeletal physiology and its production by various bone cells, elicited target cell responses, modulation by other signalling molecules (e.g., cytokines, hormones, fatty acid derivatives), and chemical interactions with other free radicals (e.g., superoxide anions, hydroxyl radicals) may form one important facet of the many complicated communication pathways controlling bone cell physiology and remodeling. Further cell and molecular studies are needed to address the precise roles that nitric oxide plays in bone development and in the formation and degradation of bone during ordinary bone metabolism. In addition, alterations in the regulation and action of the bone nitric oxide system as a function of certain bone disorders may be manifested by perturbations in bone integrity or mineral homeostasis. In this article, we review the current evidence implicating nitric oxide as an important messenger molecule in bone intercellular communication, speculate on potential roles for this radical in bone biology, and discuss possible future directions for advanced research into the function of nitric oxide in skeletal physiology.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcb.240570305
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  • 5
    Electronic Resource
    Electronic Resource
    Spectrin localization in osteoclasts: Immunocytochemistry, cloning, and partial sequencing (1998)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 71 (1998), S. 204-215 
    Details
    ISSN: 0730-2312
    Keywords: osteoclast ; spectrin ; membrane skeleton ; bone ; bone resorption ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: The presence of spectrin was demonstrated in chick osteoclasts by Western blotting and light and electron microscopic immunolocalization. Additionally, screening of a chick osteoclast cDNA library revealed the presence of α-spectrin. Light microscope level immunocytochemical staining of osteoclasts in situ revealed spectrin staining throughout the cytoplasm with heavier staining found at the marrow-facing cell margin and around the nuclei. Confocal microscopy of isolated osteoclasts plated onto a glass substrate showed that spectrin encircled the organelle-rich cell center. Nuclei and cytoplasmic inclusions were also stained and the plasma membrane was stained in a nonuniform, patchy distribution corresponding to regions of apparent membrane ruffling. Ultracytochemical localization showed spectrin to be found at the plasma membrane and distributed throughout the cytoplasm with especially intense staining of the nuclear membrane and filaments within the nuclear compartment. J. Cell. Biochem. 71:204-215, 1998. © 1998 Wiley-Liss, Inc.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Proinflammatory agents, IL-8 and IL-10, upregulate inducible nitric oxide synthase expression and nitric oxide production in avian osteoclast-like cells (1996)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 60 (1996), S. 469-483 
    Details
    ISSN: 0730-2312
    Keywords: lipopolysaccharide ; interleukin-1 ; tumor necrosis factor ; interferon ; transforming growth factor β ; dexamethasone ; RT-PCR ; NADPH diaphorase ; bone ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Nitric oxide synthase (NOS) isoenzymes generate nitric oxide (NO), a sensitive multifunctional intercellular signal molecule. High NO levels are produced by an inducible NOS (iNOS) in activated macrophages in response to proinflammatory agents, many of which also regulate local bone metabolism. NO is a potent inhibitor of osteoclast bone resorption, whereas inhibitors of NOS promote bone resorption both in vitro and in vivo. The possibility that osteoclasts, like macrophages, express a regulated iNOS and produce NO as a potential autocrine signal following inflammatory stimulation was investigated in well-characterized avian marrow-derived osteoclast-like cells. NO production (reflected by medium nitrite levels) was markedly elevated in these cells by the proinflammatory agents lipopolysaccharide (LPS) and the synergistic action of IL-1α, TNFα, and IFNγ. Inhibitors of NOS activity (aminoguanidine, L-NAME) or iNOS induction (dexamethasone, TGFβ) reduced LPS-stimulated nitrite production. LPS also increased the NOS-associated diaphorase activity of these cells and their reactivity with anti-iNOS antibodies. RT-PCR cloning, using avian osteoclast-like cell RNA and human iNOS primers, yielded a novel 900 bp cDNA with high sequence homology (76%) to human, rat, and mouse iNOS genes. In probing osteoclast-like cell RNA with the PCR-derived iNOS cDNA, a 4.8 kb mRNA species was detected whose levels were greatly increased by LPS. Induction of iNOS mRNA by LPS, or by proinflammatory cytokines, occurred prior to the rise of medium nitrite in time course studies and was diminished by dexamethasone. Moreover, osteoclast-like cells demonstrated an upregulation of NO production and iNOS mRNA by IL-8 and IL-10, regulatory mechanism's not previously described. It is concluded that osteoclast-like cells express a novel iNOS that is upregulated by inflammatory mediators, leading to NO production. Therefore, NO may serve as both a paracrine and autocrine signal for modulating osteoclast bone resorption. © 1996 Wiley-Liss, Inc.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
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  • 7
    Electronic Resource
    Electronic Resource
    Purification and characterization of an osteoclast membrane glycoprotein with homology to manganese superoxide dismutase (1991)
    New York, N.Y. : Wiley-Blackwell
    Journal of Cellular Biochemistry 46 (1991), S. 219-233 
    Details
    ISSN: 0730-2312
    Keywords: osteoclast ; membrane glycoprotein ; superoxide dismutase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: The osteoclast is the specialized multinucleated cell primarily responsible for the degradation of the inorganic and organic components of bone matrix. Isolated avian osteoclasts have been used to immunize mice and generate an osteoclast-directed monoclonal antibody library (J. Cell Biology, 100:1592). A subset of these monoclonal antibodies recognizes antigens which are expressed on osteoclasts and which are absent or nearly so on multinucleated giant cells formed in vitro from monocyte or marrow mononuclear cells. One of these antibodies, designated 121F, has been used to identify and purify an osteoclast plasma membrane-associated glycoprotein. Western blot analysis on disulfide bond-reduced extracts from osteoclasts or multinucleated giant cells formed in vitro demonstrates that the 121F antibody recognizes a 150 kDa protein detectable only in osteoclasts. This high molecular weight protein has been purified by a combination of immunoaffinity and gel filtration chromatography procedures, in conjunction with electroelution of a single band from SDS-polyacrylamide gels. Silver staining of the purified antigen on SDS-polyacrylamide gels has revealed a single protein species larger than 200 kDa in its unreduced form and 150 kDa when disulfides are reduced. Isoelectric focusing of the purified antigen reveals a single species, having a neutral pl point of 6.95. Whereas endoglycosidase treatment and lectin affinity chromatographic analyses demonstrate that the antigen recognized by the 121F antibody possesses complex N-linked sugars, trifluoromethanesulfonic acid treatment indicates there are no additional O-linked carbohydrate components. Periodate oxidation and monosaccharide hapten inhibition studies provide no evidence for the antigenic epitope bound by the 121F antibody being carbohydrate in nature. Although the native antigen is blocked at its N-terminus, amino acid analysis of a hydroxylamine generated peptide disclosed a striking relationship between the osteoclast antigen recognized by the 121F monoclonal antibody and manganese and iron superoxide dismutase. Therefore, in addition to serving as a distinguishing cell type-specific marker for osteoclasts, this cell surface glycoprotein may function directly in osteoclast-mediated bone resorption.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcb.240460305
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  • 8
    Electronic Resource
    Electronic Resource
    The origin, development and regulation of osteoclasts (1987)
    New York, NY : Wiley-Blackwell
    BioEssays 7 (1987), S. 30-34 
    Details
    ISSN: 0265-9247
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Osteoclasts, the multinucleated cells primarily responsible for dissolution of bone tissue, form by fusion of precursors that circulate in the bloodstream. A variety of factors have been shown to affect the mature osteoclast and its progenitor cell, such as calcium-regulating hormones, products of the immune system, and constituents of the arachidonic acid cascade. To date, however, the osteoclast precursor has not been identified. Furthermore, there is limited information on the factors that influence osteoclast development and regulation, reflecting in part the paucity of data on the osteoclast cell surface. Recent progress in understanding osteoclasts formation and function is discussed.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/bies.950070107
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