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The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1984 Jul;74(1):224–230. doi: 10.1172/JCI111405

Coagulation factor Va binds to human umbilical vein endothelial cells and accelerates protein C activation.

I Maruyama, H H Salem, P W Majerus
PMCID: PMC425204  PMID: 6547457

Abstract

In vitro the rate of protein C activation by thrombin is significantly accelerated by two distinct cofactors (a) the endothelial cell surface protein, thrombomodulin, and (b) human coagulation Factor Va. We have recently reported that the activity of Factor Va is contained in the 78,000-D light chain. In this study we have investigated the effects of Factor Va and its light chain on the activation of protein C in the presence of cultured endothelial cells. Thrombin-catalyzed protein C activation on human umbilical vein endothelial cells was enhanced by Factor Va. The ability of Factor Va to stimulate protein C activation on these cells was saturated at 50 nM Factor Va and was observed at several protein C concentrations. Isolated Factor Va light chain in concentrations up to 50 nM also accelerated protein C activation on endothelial cells, but higher concentrations inhibited the reaction. The effects of Factor Va or its light chain on protein C activation were also shown on a mouse hemangioma cell line but not on human fibroblasts nor on a human amelanotic melanoma cell line. Protein C activation on endothelial cells was partially inhibited by a goat anti-thrombomodulin antibody and further addition of a polyclonal rabbit anti-Factor V(Va) antibody resulted in additional inhibition. Endothelial cells grown in medium supplemented with human serum, devoid of Factor V coagulant activity, contained cell surface Factor V(Va) (approximately 15,000 molecules/cell) as measured by the binding of a monoclonal IgG directed against Factor V(Va). These cells also bound an additional 6,000-10,000 molecules Factor Va per cell as determined by direct binding studies using 125I-Factor Va. We suggest that thrombomodulin and Factor Va act in concert to regulate protein C activation on the surface of endothelial cells.

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Selected References

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  1. Broekmans A. W., Veltkamp J. J., Bertina R. M. Congenital protein C deficiency and venous thromboembolism. A study of three Dutch families. N Engl J Med. 1983 Aug 11;309(6):340–344. doi: 10.1056/NEJM198308113090604. [DOI] [PubMed] [Google Scholar]
  2. Coots M. C., Muhleman A. F., Glueck H. I. Hemorrhagic death associated with a high titer factor V inhibitor. Am J Hematol. 1978;4(2):193–206. doi: 10.1002/ajh.2830040212. [DOI] [PubMed] [Google Scholar]
  3. Esmon C. T., Owen W. G. Identification of an endothelial cell cofactor for thrombin-catalyzed activation of protein C. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2249–2252. doi: 10.1073/pnas.78.4.2249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Esmon N. L., Owen W. G., Esmon C. T. Isolation of a membrane-bound cofactor for thrombin-catalyzed activation of protein C. J Biol Chem. 1982 Jan 25;257(2):859–864. [PubMed] [Google Scholar]
  5. Fry G. L., Czervionke R. L., Hoak J. C., Smith J. B., Haycraft D. L. Platelet adherence to cultured vascular cells: influence of prostacyclin (PGI2). Blood. 1980 Feb;55(2):271–275. [PubMed] [Google Scholar]
  6. Griffin J. H., Evatt B., Zimmerman T. S., Kleiss A. J., Wideman C. Deficiency of protein C in congenital thrombotic disease. J Clin Invest. 1981 Nov;68(5):1370–1373. doi: 10.1172/JCI110385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hjelm H., Hjelm K., Sjöquist J. Protein A from Staphylococcus aureus. Its isolation by affinity chromatography and its use as an immunosorbent for isolation of immunoglobulins. FEBS Lett. 1972 Nov 15;28(1):73–76. doi: 10.1016/0014-5793(72)80680-x. [DOI] [PubMed] [Google Scholar]
  8. Jaffe E. A., Nachman R. L., Becker C. G., Minick C. R. Culture of human endothelial cells derived from umbilical veins. Identification by morphologic and immunologic criteria. J Clin Invest. 1973 Nov;52(11):2745–2756. doi: 10.1172/JCI107470. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kane W. H., Majerus P. W. Purification and characterization of human coagulation factor V. J Biol Chem. 1981 Jan 25;256(2):1002–1007. [PubMed] [Google Scholar]
  10. Kane W. H., Mruk J. S., Majerus P. W. Activation of coagulation factor V by a platelet protease. J Clin Invest. 1982 Nov;70(5):1092–1100. doi: 10.1172/JCI110697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kisiel W., Ericsson L. H., Davie E. W. Proteolytic activation of protein C from bovine plasma. Biochemistry. 1976 Nov 2;15(22):4893–4900. doi: 10.1021/bi00667a022. [DOI] [PubMed] [Google Scholar]
  12. Laposata M., Prescott S. M., Bross T. E., Majerus P. W. Development and characterization of a tissue culture cell line with essential fatty acid deficiency. Proc Natl Acad Sci U S A. 1982 Dec;79(24):7654–7658. doi: 10.1073/pnas.79.24.7654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Miletich J. P., Broze G. J., Jr, Majerus P. W. The synthesis of sulfated dextran beads for isolation of human plasma coagulation factors II, IX, and X. Anal Biochem. 1980 Jul 1;105(2):304–310. doi: 10.1016/0003-2697(80)90462-5. [DOI] [PubMed] [Google Scholar]
  14. Miletich J. P., Majerus D. W., Majerus P. W. Patients with congenital factor V deficiency have decreased factor Xa binding sites on their platelets. J Clin Invest. 1978 Oct;62(4):824–831. doi: 10.1172/JCI109194. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Owen W. G., Esmon C. T. Functional properties of an endothelial cell cofactor for thrombin-catalyzed activation of protein C. J Biol Chem. 1981 Jun 10;256(11):5532–5535. [PubMed] [Google Scholar]
  16. Owen W. G. Evidence for the formation of an ester between thrombin and heparin cofactor. Biochim Biophys Acta. 1975 Oct 20;405(2):380–387. doi: 10.1016/0005-2795(75)90103-8. [DOI] [PubMed] [Google Scholar]
  17. Rodgers G. M., Shuman M. A. Prothrombin is activated on vascular endothelial cells by factor Xa and calcium. Proc Natl Acad Sci U S A. 1983 Nov;80(22):7001–7005. doi: 10.1073/pnas.80.22.7001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Salem H. H., Broze G. J., Miletich J. P., Majerus P. W. Human coagulation factor Va is a cofactor for the activation of protein C. Proc Natl Acad Sci U S A. 1983 Mar;80(6):1584–1588. doi: 10.1073/pnas.80.6.1584. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Salem H. H., Broze G. J., Miletich J. P., Majerus P. W. The light chain of factor Va contains the activity of factor Va that accelerates protein C activation by thrombin. J Biol Chem. 1983 Jul 25;258(14):8531–8534. [PubMed] [Google Scholar]
  20. Salem H. H., Esmon N. L., Esmon C. T., Majerus P. W. Effects of thrombomodulin and coagulation Factor Va-light chain on protein C activation in vitro. J Clin Invest. 1984 Apr;73(4):968–972. doi: 10.1172/JCI111321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Stenflo J. A new vitamin K-dependent protein. Purification from bovine plasma and preliminary characterization. J Biol Chem. 1976 Jan 25;251(2):355–363. [PubMed] [Google Scholar]
  22. Suzuki K., Stenflo J., Dahlbäck B., Teodorsson B. Inactivation of human coagulation factor V by activated protein C. J Biol Chem. 1983 Feb 10;258(3):1914–1920. [PubMed] [Google Scholar]
  23. Vehar G. A., Davie E. W. Preparation and properties of bovine factor VIII (antihemophilic factor). Biochemistry. 1980 Feb 5;19(3):401–410. doi: 10.1021/bi00544a001. [DOI] [PubMed] [Google Scholar]
  24. Walker F. J., Sexton P. W., Esmon C. T. The inhibition of blood coagulation by activated Protein C through the selective inactivation of activated Factor V. Biochim Biophys Acta. 1979 Dec 7;571(2):333–342. doi: 10.1016/0005-2744(79)90103-7. [DOI] [PubMed] [Google Scholar]
  25. Wilson D. B., Salem H. H., Mruk J. S., Maruyama I., Majerus P. W. Biosynthesis of coagulation Factor V by a human hepatocellular carcinoma cell line. J Clin Invest. 1984 Mar;73(3):654–658. doi: 10.1172/JCI111256. [DOI] [PMC free article] [PubMed] [Google Scholar]

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