The long-term objective of this study Is to understand the factors regulating osteoblastic cell calcium. It is becoming increasingly evident that changes In this ion are involved in the mechanism of action of several resorptive agents and that an understanding of the various calcium regulatory systems will aid in the development of potential ways to control pathological changes in bone metabolism occurring systemically or locally. Specifically, it is proposed in osteoblastic cells isolated from the calvaria of newborn rats and In the osteoblastic human osteosarcomal cell lines, SaOS-2 and G292, to: 1. to further study the nature of the regulation of mobilization of calcium from intracellular pools with particular emphasis on the role of guanine trisphosphate and Its possible interaction with inositol phosphates such as 1(1,4,5)p3; 2. to delineate the nature of regulation of phosphoinositide metabolism and cytosolic calcium levels by the bone resorptive agent and mitogen, epidermal growth factor (EGF), in order to better compare and contrast its effects with other bone resorptive agents such as thrombin; 3. to delineate the nature of regulation of phosphoinositide metabolism by calcium via experimentation with calcium channel antagonists, an ionophore and extracellular calcium levels; 4. to study the electrophysiological properties of these cells with emphasis on membrane potential and potassium, sodium and calcium currents; 5. to study the effects of EGF and thrombin on these electrophysiological properties; 6. to study the effects of inositol phosphates on the electrophysiological properties defined in aims 4 & 5; 7. to study the role of tyrosine kinase activity in the regulation of osteoblastic cell calcium, phosphoinositide metabolism as well as proliferation In response to EGF. Cytosolic calcium regulation will be studied with the fluorescent dye, Fura-2, and phosphoinositide metabolism in tritiated -inositol labelled cells by means of high performance liquid chromatography. Cell proliferation will be assessed by incorporation of tritiated thymidine. The electrophysiological properties will be studied with patch-clamp techniques. In general, the studies proposed here should provide more In depth information on the intracellular and plasma membrane regulatory mechanisms involved in the control of osteoblastic cell calcium with emphasis. on the possible diversities of these pathways involved upon activation by agents as potentially different as EGF and thrombin.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR025271-14
Application #
3155294
Study Section
General Medicine B Study Section (GMB)
Project Start
1988-04-01
Project End
1994-03-31
Budget Start
1992-04-01
Budget End
1993-03-31
Support Year
14
Fiscal Year
1992
Total Cost
Indirect Cost
Name
State University of New York at Buffalo
Department
Type
Schools of Dentistry
DUNS #
038633251
City
Buffalo
State
NY
Country
United States
Zip Code
14260
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Zhang, W; Dziak, R M; Aletta, J M (1995) EGF-mediated phosphorylation of extracellular signal-regulated kinases in osteoblastic cells. J Cell Physiol 162:348-58
Dziak, R (1993) Biochemical and molecular mediators of bone metabolism. J Periodontol 64:407-15
Zhang, W; Dziak, R (1993) Effects of interleukin-1 alpha on arachidonic acid metabolism in human osteosarcoma osteoblastic cells. Prostaglandins Leukot Essent Fatty Acids 49:521-6
Ren, W; Kinniburgh, A J; Dziak, R (1992) Regulation of C-myc protooncogene expression in osteoblastic cells by arachidonic acid metabolites: relationship to proliferation. Calcif Tissue Int 50:372-7
Kohl, A C; Tatakis, D N; Hansen, C et al. (1992) Effects of interleukin-1, tumor necrosis factor -beta, and forskolin on tissue plasminogen activator activity in human osteoblastic osteosarcoma cells. Calcif Tissue Int 50:129-33
Tatakis, D N; Schneeberger, G; Dziak, R (1991) Recombinant interleukin-1 (IL-1) stimulates prostaglandin E2 production by osteoblastic cells: role of calcium, calmodulin, and cAMP. Lymphokine Cytokine Res 10:95-9
Ren, W; Dziak, R (1991) Effects of leukotrienes on osteoblastic cell proliferation. Calcif Tissue Int 49:197-201
Falsafi, R; Tatakis, D N; Hagel-Bradway, S et al. (1991) Effects of inositol trisphosphate on calcium mobilization in bone cells. Calcif Tissue Int 49:333-9
Shapiro, S; Tatakis, D N; Dziak, R (1990) Effects of tumor necrosis factor alpha on parathyroid hormone-induced increases in osteoblastic cell cyclic AMP. Calcif Tissue Int 46:60-2

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