Abstract

The objective of this proposal is to isolate and characterize the site of mineral deposition in matrix vesicles. This will be achieved using matrix vesicles, induced-vesicles and plasma membranes isolated from epiphyseal plate chondrocytes and from chondrocytes maintained in culture. Vesicles and membranes will be fragmented by physical and chemical disruptive techniques. For some experiments, recombinant vesicles will be formed by fusing membrane fragments into liposomes. Mineralization-competent membrane fragments and recombinant vesicles will be selected by calcium-loading and density gradient separation procedures. We will then determine the mineralization potential of vesicles and fragments by calcium binding and turbidity measurements. Chemical and morphological analyses will be used to characterize the lipid, protein and mineral components of the mineralizing fragments. By modifying the lipid composition of the recombinant vesicles we will examine the role of specific phospholipids in matrix vesicle mineralization. Finally, we propose to study the role of alkaline phosphatase in mineralization by preparing alkaline phosphatase-containing liposomes and measuring their mineralization potential. These liposomes will be used in recombination experiments to investigate the interaction of the enzyme with other membrane components.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE006533-03
Application #
3220052
Study Section
Oral Biology and Medicine Study Section (OBM)
Project Start
1983-04-01
Project End
1986-03-31
Budget Start
1985-04-01
Budget End
1986-03-31
Support Year
3
Fiscal Year
1985
Total Cost
Indirect Cost

  Institution

Name
University of Pennsylvania
Department
Type
Schools of Dentistry/Oral Hygn
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104

  Publications

Kirsch, T; Harrison, G; Golub, E E et al. (2000) The roles of annexins and types II and X collagen in matrix vesicle-mediated mineralization of growth plate cartilage. J Biol Chem 275:35577-83
Kirsch, T; Harrison, G; Worch, K P et al. (2000) Regulatory roles of zinc in matrix vesicle-mediated mineralization of growth plate cartilage. J Bone Miner Res 15:261-70
Kirsch, T; Nah, H D; Demuth, D R et al. (1997) Annexin V-mediated calcium flux across membranes is dependent on the lipid composition: implications for cartilage mineralization. Biochemistry 36:3359-67
Golub, E E (1996) Enzymes in mineralizing systems: state of the art. Connect Tissue Res 35:183-8
Harrison, G; Shapiro, I M; Golub, E E (1995) The phosphatidylinositol-glycolipid anchor on alkaline phosphatase facilitates mineralization initiation in vitro. J Bone Miner Res 10:568-73
Yuan, Z A; Golub, E E; Collier, P M et al. (1995) Bovine enamel organ cells express tissue non-specific alkaline phosphatase mRNA. J Dent Res 74:1886-90
Chung, C H; Golub, E E; Forbes, E et al. (1992) Mechanism of action of beta-glycerophosphate on bone cell mineralization. Calcif Tissue Int 51:305-11
Matsumoto, H; Silverton, S F; Debolt, K et al. (1991) Superoxide dismutase and catalase activities in the growth cartilage: relationship between oxidoreductase activity and chondrocyte maturation. J Bone Miner Res 6:569-74
Oshima, O; Leboy, P S; McDonald, S A et al. (1989) Developmental expression of genes in chick growth cartilage detected by in situ hybridization. Calcif Tissue Int 45:182-92
Leboy, P S; Vaias, L; Uschmann, B et al. (1989) Ascorbic acid induces alkaline phosphatase, type X collagen, and calcium deposition in cultured chick chondrocytes. J Biol Chem 264:17281-6

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