Osteocalcin, the major noncollagenous bone protein, characterized by the vitamin K-dependent calcium-binding amino acid, Gamma-carboxyglutamic acid (G1a) has well-defined molecular and physical properties. Recently, it has been found that bone that is 99% deficient in osteocalcin and 93% deficient in Gla is poorly resorbed both in vivo, when implanted subcutaneously in a rat model, and in vitro, when incubated with human monocytes. These observations agree with previous studies implicating osteocalcin in calcium regulation. For example, the chemotactic property of osteocalcin for cells capable of resorbing bone and the specific increase in osteocalcin synthesis by 1,25(OH)2D3, a hormone which can promote bone resorption. This proposal will investigate the precise mechanism for the observed defect in the decreased resorption of osteocalcin-deficient bone. Bone from the warfarin-treated and K-deficient rat wil be chemically characterized for other protein alterations (analyses by 2D gels and HPLC profiles). The critical levels of osteocalcin and/or other Gla-containing proteins necessary to achieve the defect will be established. Experiments will compare the resorption of devitalized normal and osteocalcin-deficient bone in a model system of in vivo implant of mineralized bone particles (BP) into subcutaneous packets. In this model blood derived progenitor cells differentiate into osteoclasts that resorb control bone from 5 through 28 days after implantation. Specific cell types are identified morphologically and resorption is quantitated by hisotomorphometric analysis (number of multinucleated cells and percent mineralized matrix) and biochemically (calcium content, DNA and tartrate-resistant acid phosphatase activity). These studies will provide information on whether osteocalcin (or some other vitamin K-sensitive protein) is necessary for 1) recruitment of cells to bone; 2) attachment to mineralized surfaces; 3) promotes differentiation of a progenitor cell to a bone-resorbing cell; or 4) is involved in stimulation of activity of bone resorbing cells. The knowledge of cell-matrix interactions to be gained from the proposed studies has potential applications in defining the control mechanisms of normal bone resorption, in explaining the pathogenesis of metabolic bone diseases, inflammatory bone destruction (peridontal disease) and in pathogenesis and treatment of pathologic mineral deposition.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR035166-02
Application #
3157067
Study Section
Oral Biology and Medicine Study Section (OBM)
Project Start
1986-08-01
Project End
1989-07-31
Budget Start
1987-08-01
Budget End
1988-07-31
Support Year
2
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Children's Hospital Boston
Department
Type
DUNS #
076593722
City
Boston
State
MA
Country
United States
Zip Code
02115
Lian, J B; Stein, G S (1995) Development of the osteoblast phenotype: molecular mechanisms mediating osteoblast growth and differentiation. Iowa Orthop J 15:118-40
Heinrichs, A A; Bortell, R; Bourke, M et al. (1995) Proximal promoter binding protein contributes to developmental, tissue-restricted expression of the rat osteocalcin gene. J Cell Biochem 57:90-100
Lynch, M P; Stein, J L; Stein, G S et al. (1995) The influence of type I collagen on the development and maintenance of the osteoblast phenotype in primary and passaged rat calvarial osteoblasts: modification of expression of genes supporting cell growth, adhesion, and extracellular matrix mineralizatio Exp Cell Res 216:35-45
Shalhoub, V; Bettencourt, B; Jackson, M E et al. (1994) Abnormalities of phosphoprotein gene expression in three osteopetrotic rat mutations: elevated mRNA transcripts, protein synthesis, and accumulation in bone of mutant animals. J Cell Physiol 158:110-20
Breen, E C; Ignotz, R A; McCabe, L et al. (1994) TGF beta alters growth and differentiation related gene expression in proliferating osteoblasts in vitro, preventing development of the mature bone phenotype. J Cell Physiol 160:323-35
Bidwell, J P; van Wijnen, A J; Fey, E G et al. (1994) Subnuclear distribution of the vitamin D receptor. J Cell Biochem 54:494-500
Bidwell, J; van Wijnen, A; Banerjee, C et al. (1994) Parathyroid-responsive modifications in the nuclear matrix of ROS 17/2.8 rat osteosarcoma cells. Endocrinology 134:1738-44
Stein, G S; van Wijnen, A J; Stein, J L et al. (1994) Nuclear architecture supports integration of physiological regulatory signals for transcription of cell growth and tissue-specific genes during osteoblast differentiation. J Cell Biochem 55:4-15
Bidwell, J P; Fey, E G; van Wijnen, A J et al. (1994) Nuclear matrix proteins distinguish normal diploid osteoblasts from osteosarcoma cells. Cancer Res 54:28-32
Stein, G S; Stein, J L; van Wijnen, A J et al. (1994) Histone gene transcription: a model for responsiveness to an integrated series of regulatory signals mediating cell cycle control and proliferation/differentiation interrelationships. J Cell Biochem 54:393-404

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