Factor VIII is a high molecular weight plasma glycoprotein complex which is an important regulator of blood coagulation and is also required for the recognition of damaged endothelial surfaces by blood platelets. A convenient model for the function of Factor VIII in platelet-mediated hemostasis is provided by the binding of bovine Factor VIII to human platelets. We propose to identify and to isolate the protein(s) on the platelet surface to which Factor VIII binds. The purified binding protein(s) will be characterized by biochemical techniques and the mechanism of association with Factor VIII determined. Chemical and enzymatic modification of the isolated binding protein(s) will be used to identify the regions responsible for functional activity. The effects of agents, such as ADP, which alter the responsiveness of platelets to Factor VIII, will be examined for their effects on the availability of binding sites on the platelet surface. We will also examine the effects of Factor VIII binding on the metabolic and functional responses of platelets and will study how this binding alters the responsiveness of platelets to other agonists. A congenital deficiency of the Factor VIII protein in humans (von Willebrand's disease) can cause a severe bleeding syndrome, because of the resulting defect in platelet-mediated hemostasis. Replacement therapy with concentrates of Factor VIII can correct this defect. On the other hand, a deficiency of Factor VIII (in swine) has been shown to be associated with a decreased tendency to form atherosclerotic plaques - probably because of a decreased responsiveness of the platelets to the early stages of damage to the vascular intima. These observations suggest that the interaction of Factor VIII with platelets plays a key role not only in protective hemostatic mechanisms but also in those which could lead to thrombotic conditions. A better understanding of this interaction is thus essential for a rational approach to the therapeutic management of these conditions.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL027993-03
Application #
3339402
Study Section
(EH)
Project Start
1983-05-01
Project End
1986-06-30
Budget Start
1985-05-01
Budget End
1986-06-30
Support Year
3
Fiscal Year
1985
Total Cost
Indirect Cost
Name
Temple University
Department
Type
Schools of Medicine
DUNS #
City
Philadelphia
State
PA
Country
United States
Zip Code
19122
Sinha, D; Bakhshi, M R; Vora, R K et al. (1996) Engineering DNA and protein chimeras utilizing coding sequences of restriction sites. Anal Biochem 238:205-8
Bakhshi, M R; Sinha, D; Vora, R K et al. (1996) Primary binding domain of bovine von Willebrand factor fragment expressed in E. coli. Thromb Haemost 75:196-202
Janel, N; Schwachtgen, J L; Bakhshi, M R et al. (1995) Comparison of the 5'-flanking sequences of the human and bovine von Willebrand factor-encoding genes reveals alternation of highly homologous domains with species-specific Alu-type repeats. Gene 167:291-5
Sinha, D; Bakhshi, M; Kunapuli, S et al. (1994) ASP514 within the A1 domain of bovine von Willebrand factor is required for interaction with platelet glycoprotein Ib. Biochem Biophys Res Commun 203:881-8
Sinha, D; Yang, X; Emig, F et al. (1994) Isolation and characterization of two protease inhibitors from bovine plasma. J Biochem 115:387-91
Sinha, D; Bakhshi, M; Vora, R (1994) Ligand binding assays with recombinant proteins refolded on an affinity matrix. Biotechniques 17:509-12, 514
Peng, M; Lu, W; Beviglia, L et al. (1993) Echicetin: a snake venom protein that inhibits binding of von Willebrand factor and alboaggregins to platelet glycoprotein Ib. Blood 81:2321-8
Bakhshi, M R; Myers, J C; Howard, P S et al. (1992) Sequencing of the primary adhesion domain of bovine von Willebrand factor. Biochim Biophys Acta 1132:325-8
Peng, M; Lu, W; Kirby, E P (1992) Characterization of three alboaggregins purified from Trimeresurus albolabris venom. Thromb Haemost 67:702-7
Peng, M; Lu, W; Kirby, E P (1991) Alboaggregin-B: a new platelet agonist that binds to platelet membrane glycoprotein Ib. Biochemistry 30:11529-36

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