This proposal uses a cartilage cell model to address how calcifying cells modulate events at distal sites in the extracellular matrix and in particular, on the role of matrix vesicles (MV) in this process. It is based on the hypothesis that vitamin D metabolites regulate MV composition via genomic mechanisms through the classic vitamin D receptor (VDR) and membrane-mediated pathways. In addition, chondrocytes synthesize and secrete vitamin D metabolites, as well as other autocoids like prostaglandin E2, which, in turn, interact directly with the MV membrane to nongenomically regulate MV activity.
Three specific aims are proposed. The first is to examine the biochemical mechanisms involved in the production of vitamin D metabolites by chondrocyte cultures. Regulation of l,25-(OH)2 D3 (1,25) and 24,25-(OH)2 D3 (24,25) production by 1,25 and 24,25 will be examined. Changes in 1-alpha- and 24-hydroxylases, as well as gene expression of the 24-hydroxylase, will be examined as a function of hormone dose and time. The role of the VDR or membrane-initiated events will be assessed using structural analogs of 1,25 and 24,25. The second is to examine the regulation of membrane signaling systems by Vitamin D metabolites. The mechanisms for differential regulation of protein kinase C (PKC) by 1,25 and 24,25 in plasma membranes (PM) and MV and use analogues of 1,25 and 24,25 to assess the relative contributions of the VDR, as well as the role of functional moieties of the vitamin D metabolites in their nongenomic action on MV membranes will be examined. The interrelationship between the PKC and PGE2 pathways will also be determined. The third is to to characterize the membrane effects of 1,25 and 24,25 on the functional properties of MV's. The regulation of stromelysin-1 by 1,25 and 24,25 will be examined, and will focus on the role of cell maturation state and the relative contributions of the VDR and membrane-mediated pathways in regulating stromelysin-1 synthesis, secretion, and activation. Nongenomic regulation of MV's will be assessed in functional assays of calcification in proteoglycan-containing gelatin gels. This work will have major implications for our understanding of MV and how cells that calcify their matrix are able to regulate these extracellular organelles at distant sites.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
2R01DE005937-15A1
Application #
2014984
Study Section
Oral Biology and Medicine Subcommittee 1 (OBM)
Project Start
1981-03-01
Project End
2002-02-28
Budget Start
1997-05-01
Budget End
1998-02-28
Support Year
15
Fiscal Year
1997
Total Cost
Indirect Cost
Name
University of Texas Health Science Center San Antonio
Department
Orthopedics
Type
Schools of Medicine
DUNS #
800772162
City
San Antonio
State
TX
Country
United States
Zip Code
78229
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Schwartz, Z; Carney, D H; Crowther, R S et al. (2005) Thrombin peptide (TP508) treatment of rat growth plate cartilage cells promotes proliferation and retention of the chondrocytic phenotype while blocking terminal endochondral differentiation. J Cell Physiol 202:336-43
Schwartz, Z; Graham, E J; Wang, L et al. (2005) Phospholipase A2 activating protein (PLAA) is required for 1alpha,25(OH)2D3 signaling in growth plate chondrocytes. J Cell Physiol 203:54-70
Gay, I; Schwartz, Z; Sylvia, V L et al. (2004) Lysophospholipid regulates release and activation of latent TGF-beta1 from chondrocyte extracellular matrix. Biochim Biophys Acta 1684:18-28
Boyan, B D; Schwartz, Zvi (2004) Rapid vitamin D-dependent PKC signaling shares features with estrogen-dependent PKC signaling in cartilage and bone. Steroids 69:591-7
Boyan, B D; Jennings, E G; Wang, L et al. (2004) Mechanisms regulating differential activation of membrane-mediated signaling by 1alpha,25(OH)2D3 and 24R,25(OH)2D3. J Steroid Biochem Mol Biol 89-90:309-15
Boyan, Barbara D; Dean, David D; Sylvia, Victor L et al. (2003) Steroid hormone action in musculoskeletal cells involves membrane receptor and nuclear receptor mechanisms. Connect Tissue Res 44 Suppl 1:130-5
Schwartz, Z; Shaked, D; Hardin, R R et al. (2003) 1alpha,25(OH)2D3 causes a rapid increase in phosphatidylinositol-specific PLC-beta activity via phospholipase A2-dependent production of lysophospholipid. Steroids 68:423-37
Boyan, Barbara D; Sylvia, V L; McKinney, N et al. (2003) Membrane actions of vitamin D metabolites 1alpha,25(OH)2D3 and 24R,25(OH)2D3 are retained in growth plate cartilage cells from vitamin D receptor knockout mice. J Cell Biochem 90:1207-23
Lohmann, C H; Schwartz, Z; Liu, Y et al. (2003) Pulsed electromagnetic fields affect phenotype and connexin 43 protein expression in MLO-Y4 osteocyte-like cells and ROS 17/2.8 osteoblast-like cells. J Orthop Res 21:326-34

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