Download PDF
Thromb Haemost 1989; 62(04): 1067-1073
DOI: 10.1055/s-0038-1647119
Original Article
Coagulation
Schattauer GmbH Stuttgart

Functional Properties of Factor VIIa/Tissue Factor Formed with Purified Tissue Factor and with Tissue Factor Expressed on Cultured Endothelial Cells

Fanny E Almus
The Departments of Medicine and Pathology, University of California at San Diego, La Jolla, CA, USA
,
L Vijaya Mohan Rao
The Departments of Medicine and Pathology, University of California at San Diego, La Jolla, CA, USA
,
Samuel I Rapaport
The Departments of Medicine and Pathology, University of California at San Diego, La Jolla, CA, USA
› Author Affiliations
Further Information

Publication History

Received 28 March 1989

Accepted after revision 01 September 1989

Publication Date:
24 July 2018 (online)

  PDF Download  Permissions and Reprints

Summary

Factor VIIa (F. VIIa)/tissue factor (TF) function was examined using purified human TF reconstituted into mixed phospholipid vesicles and TF expressed on cultured human umbilical vein endothelial cells (HUVEC) treated with thrombin. In reaction mixtures containing either type of TF, F. VIIa, 10 nM, either 3H-factor X or 3H-factor IX, 88 nM, and Ca2+, 5 mM, F. VIIa/TF activated factor X (F. X) several fold faster than it activated factor IX (F. IX). Adding heparin, 1 U/ml, increased rates of activation of both substrates and F. X remained the preferred substrate. Adding plasma at concentrations of 5%;or above inhibited factor VIIa/TF catalytic activity. Inhibition was shown to require F. Xa as a cofactor, was prevented by antibodies to extrinsic pathway inhibitor (EPI), and was reversible by decalcification. Thus, with factor VIIa/TF formed with both types of TF, EPI appeared responsible for inhibition induced by plasma. Our data indicate that functional properties of factor VIIa/TF as delineated in reaction mixtures made with purified TF reconstituted into mixed phospholipid vesicles also hold for factor VIIa/TF activity on the surface of cultured HUVEC.

Keywords

Tissue factor - Extrinsic pathway inhibitor - Heparin - Factor X and factor IX - Endothelial cells
 

  • References

  • 1 Østerud B, Rapaport SI. Activation of factor IX by the reaction product of tissue factor and factor VII: additional pathway for initiating blood coagulation. Proc Nat Acad Sci USA 1977; 74: 5260-5264
    CrossrefPubMedGoogle Scholar
  • 2 Jesty J, Silverberg SA. Kinetics of the tissue factor-dependent activation of coagulation factors IX and X in a bovine plasma system. J Biol Chem 1979; 254: 12337-12345
    PubMedGoogle Scholar
  • 3 Zur M, Nemerson Y. Kinetics of factor IX activation via the extrinsic pathway. J Biol Chem 1980; 255: 5703-5707
    PubMedGoogle Scholar
  • 4 Rapaport SI. Inhibition of factor VIIa/tissue factor-induced blood coagulation: with particular emphasis upon a factor Xa-dependent inhibitory mechanism. Blood 1989; 73: 359-365
    PubMedGoogle Scholar
  • 5 Rao LV M, Rapaport SI. Studies of a mechanism inhibiting the initiation of the extrinsic pathway of coagulation. Blood 1987; 69: 645-651
    PubMedGoogle Scholar
  • 6 Broze Jr GJ, Warren LA, Novotny WF, Higuchi DA, Girard JJ, Miletich JP. The lipoprotein-associated coagulation inhibitor that inhibits the factor VII-tissue factor complex also inhibits factor Xa:Insight into its possible mechanism of action. Blood 1988; 71: 335-343
    PubMedGoogle Scholar
  • 7 Rao LV M. Characterization of anti-tissue factor antibody and its use in immunoaffinity purification of human tissue factor. Thromb Res 1988; 51: 373-384
    CrossrefPubMedGoogle Scholar
  • 8 Jaffe EA, Nachman RL, Becker CG, Minick CR. Culture of human endothelial cells derived from umbilical veins. J Clin Invest 1973; 52: 2745-2756
    CrossrefPubMedGoogle Scholar
  • 9 Almus FE, Rao LV M, Rapaport SI. Decreased inducibility of tissue factor activity on human umbilical vein endothelial cells cultured with endothelial cell growth factor and heparin. Thromb Res 1988; 50: 339-344
    CrossrefPubMedGoogle Scholar
  • 10 Rao LV M, Bajaj SP. Purification of human factor VII utilizing O-(Diethylaminoethyl)-Sephadex and Sulfopropyl Sephadex Chromatography. Anal Biochem 1984; 136: 357-361
    CrossrefPubMedGoogle Scholar
  • 11 Bajaj SP, Rapaport SI, Prodanos C. A simplified procedure for purification of human prothrombin, factor IX and factor X. Prep Biochem 1981; 11: 397-412
    PubMedGoogle Scholar
  • 12 Rao LV M, Rapaport SI, Bajaj SP. Activation of human factor VII in the initiation of tissue factor-dependent coagulation. Blood 1986; 68: 685-691
    PubMedGoogle Scholar
  • 13 Van LentenL, Ashwell G. Studies on the chemical and enzymatic modification of glycoproteins. J Biol Chem 1971; 246: 1889-1894
    PubMedGoogle Scholar
  • 14 Sanders NL, Bajaj SP, Zivelin A, Rapaport SI. Inhibition of tissue factor/factor VIIa activity in plasma requires factor X and an additional plasma component. Blood 1985; 66: 204-212
    PubMedGoogle Scholar
  • 15 Warn-Cramer BL, Maki S, Zivelin A, Rapaport SI. Partial purification and characterization of extrinsic pathway inhibitor (the factor Xa dependent plasma inhibitor of factor VIIa/tissue factor). Thromb Res 1987; 48: 11-22
    CrossrefPubMedGoogle Scholar
  • 16 Laudano AP, Cottrell BA, Doolittle R. Synthetic peptides modeled on fibrin polymerization sites. Ann NY Acad Sci 1983; 408: 315-329
    CrossrefPubMedGoogle Scholar
  • 17 Silverberg SA, Nemerson Y, Zur M. Kinetics of the activation of bovine coagulation factor X by components of the extrinsic pathway. J Biol Chem 1977; 252: 8481-8488
    PubMedGoogle Scholar
  • 18 Repke DI, MacLean D, Nemerson Y. Heparin affects the substrate specificity of the tissue factor pathway of coagulation. Fed Proc 1981 40. Abstract 274
    PubMedGoogle Scholar
  • 19 Nemerson Y. Tissue factor and hemostasis. Blood 1988; 71: 1-8
    PubMedGoogle Scholar
  • 20 Born VJ J, Van Hinsbergh VW M, Reinalda-Poot JH, Mohanial RW, Bertina RM. Extrinsic activation of human coagulation factors IX and X on the endothelial surface. Thromb Haemostas 1987; 58: 522 (Abstr 1931)
    PubMedGoogle Scholar
  • 21 Bajaj SM, Ali M, Saba S, Hyers TM, Bajaj SP. Expression of tissue factor and suppression of factor VIIa-TF inhibitor production by endotoxin or phorbol ester stimulated endothelial cells. Circulation 1988; 78: 522 Abstract 1254
    CrossrefPubMedGoogle Scholar
  • 22 Rana SV, Reimers HJ, Pathikonda MS, Bajaj SP. Expression of tissue factor and factor VIIa/tissue factor inhibitor activity in endotoxin or phorbol ester stimulated U937 monocyte-like cells. Blood 1988; 71: 259-262
    PubMedGoogle Scholar
  • 23 Gramzinski RA, Broze Jr GJ, Carson S. Human fibroblast tissue factor is inhibited by lipoprotein-associated coagulation inhibitor and placental anticoagulant protein but not by apolipoprotein A-II. Blood 1989; 73: 983-989
    PubMedGoogle Scholar