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Thromb Haemost 2001; 86(02): 660-667
DOI: 10.1055/s-0037-1616110
Scientific and Standardization Committee Communications
Schattauer GmbH

Platelets and Phospholipids in Tissue Factor-initiated Thrombin Generation

Saulius Butenas
1   University of Vermont, Departments of Biochemistry and Medicine, Burlington, Vermont, USA
,
Richard F Branda
1   University of Vermont, Departments of Biochemistry and Medicine, Burlington, Vermont, USA
,
Cornelis van ’t Veer
1   University of Vermont, Departments of Biochemistry and Medicine, Burlington, Vermont, USA
,
Kevin M Cawthern
1   University of Vermont, Departments of Biochemistry and Medicine, Burlington, Vermont, USA
,
Kenneth G Mann
1   University of Vermont, Departments of Biochemistry and Medicine, Burlington, Vermont, USA
› InstitutsangabenThis study was supported by Program Project Grant HL 46703 from the National Institutes of Health (KGM) and by a TALENT stipend of the Netherlands Organization of Scientific Research (CvV).
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Publikationsverlauf

Received 28. Dezember 2000

Accepted after revision 16. März 2001

Publikationsdatum:
12. Dezember 2017 (online)

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Summary

The influence of platelets on tissue factor (TF)-initiated thrombin generation in a reconstituted model of blood coagulation and in whole blood was evaluated. No thrombin generation was observed over 15 min in the reconstituted model when either TF or platelets and phospholipids were omitted. At 25 pM TF, the rates of thrombin generation were platelet and PCPS concentration-dependent and achieved maximum (1.0 nM/s) in the physiological range of platelet concentration. Similar rates were achieved in the absence of platelets when 1-2 μM phospholipid was used. However, the maximum rates of thrombin generation (5.2-6.0 nM/s) and the shortest initiation phase (1 min) were attained between 25 and 100 μM phospholipid. In the reconstituted model, an increase in platelet concentration from 0.125 × 108/ml to 0.5 × 108/ml decreased the duration of the initiation phase (in the absence of phospholipids) from 4.3 min to 2 min. Further increases in platelet concentration did not affect this phase. Sequential whole blood studies were conducted in blood of a chemotherapy patient who developed reduced platelet counts. The TF (12.5 pM) initiated clotting of patient’s blood was accelerated from ~10 min to 5 min when the platelet concentration increased from 0.05 × 108/ml to 0.11 × 108/ml. Clotting times were essentially unchanged for platelet concentrations exceeding 0.5 × 108/ml (range 0.5-3.1 × 108/ml). Similarly, clotting of whole blood obtained from healthy volunteers was not affected by the platelet count, which varied from 1.5 × 108/ml to 3.1 × 108/ml (4.0 ± 0.5 min). The data obtained in both models are consistent with in vivo observations that clinical bleeding is most likely to occur at platelet counts <0.1 × 108/ml.

Portions of this work were presented at the 40th Annual Meeting of the American Society of Hematology, December 4-8, 1998, Miami Beach, Florida (abstracts #140 and #738).

Keywords

Platelets - phospholipids - thrombocytopenia - tissue factor - factor V
 

  • References

  • 1 Jenny NS, Mann KG. Coagulation cascade: an overview. In: Loscalzo J, Schafer AI. eds. Throm Hemorrhage. Baltimore, MD: Williams and Wilkins; 1998: 3-27.
    Suche in Google Scholar
  • 2 Bevers EM, Comfinius P, van Rijn JLML, Hemker HC, Zwaal RFA. Generation of prothrombin converting activity and the exposure of phosphatidylserine at the outer surface of platelets. Eur J Biochem 1982; 122: 429-36.
    CrossrefPubMedSuche in Google Scholar
  • 3 Rosing J, van Rijn JLML, Bevers EM, van Dieijen G, Comfurius P, Zwaal RF. The role of activated human platelets in prothrombin and factor X activation. Blood 1985; 65: 319-32.
    PubMedSuche in Google Scholar
  • 4 Gailani D, Broze Jr GJ. Factor XI activation in a revised model of blood coagulation. Science 1991; 253: 909-12.
    CrossrefPubMedSuche in Google Scholar
  • 5 Pieters J, Lindhout T, Hemker HC. In situ-generated thrombin is the only enzyme that effectively activates factor VIII and factor V in thromboplastin-activated plasma. Blood 1989; 74: 1021-4.
    PubMedSuche in Google Scholar
  • 6 Butenas S, van ’t Veer C, Mann KG. Evaluation of the initiation phase of blood coagulation using ultrasensitive assays for serine proteases. J Biol Chem 1997; 272: 21527-33.
    CrossrefPubMedSuche in Google Scholar
  • 7 Brummel KE, Butenas S, Mann KG. An integrated study of fibrinogen during blood coagulation. J Biol Chem 1999; 274: 22862-70.
    CrossrefPubMedSuche in Google Scholar
  • 8 Tans G, Rosing J, Thomassen MC, Heeb MJ, Zwaal RF, Griffin JH. Comparison of anticoagulant and procoagulant activities of stimulated platelets and platelet-derived microparticles. Blood 1991; 77: 2641-8.
    PubMedSuche in Google Scholar
  • 9 Hugel B, Socie G, Vu T, Toti F, Gluckman E, Freyssinet JM, Scrobohaci ML. Elevated levels of circulating procoagulant microparticles in patients with paroxysmal nocturnal hemoglobinuria and aplastic anemia. Blood 1999; 93: 3451-6.
    CrossrefPubMedSuche in Google Scholar
  • 10 Moyer MP, Tracy RP, Tracy PB, van ’t Veer C, Sparks CE, Mann KG. Plasma lipoproteins support prothrombinase and other procoagulant enzymatic complexes. Arterioscler Thromb Vasc Biol 1998; 18: 458-65.
    CrossrefPubMedSuche in Google Scholar
  • 11 Stern D, Nawroth P, Handley D, Kisiel W. An endothelial cell-dependent pathway of coagulation. Proc Natl Acad Sci USA 1985; 82: 2523-7.
    CrossrefPubMedSuche in Google Scholar
  • 12 Tracy PB, Eide LL, Mann KG. Human prothrombinase complex assembly and function on isolated peripheral blood cell populations. J Biol Chem 1985; 260: 2119-24.
    PubMedSuche in Google Scholar
  • 13 Broze Jr GJ. Tissue factor pathway inhibitor and the revised theory of coagulation. Annu Rev Med 1995; 46: 103-12.
    CrossrefPubMedSuche in Google Scholar
  • 14 Bevers EM, Comfurius P, Zwaal RF. Changes in membrane phospholipid distribution during platelet activation. Biochim Biophys Acta 1983; 736: 57-66.
    CrossrefPubMedSuche in Google Scholar
  • 15 Mann KG, Bovill EG, Krishnaswamy S. Surface-dependent reactions in the propagation phase of blood coagulation. Ann NY Acad Sci 1991; 614: 63-75.
    CrossrefPubMedSuche in Google Scholar
  • 16 Krishnaswamy S, Field KA, Edgington TS, Morrissey JH, Mann KG. Role of the membrane surface in the activation of human coagulation factor X. J Biol Chem 1992; 267: 26110-20.
    PubMedSuche in Google Scholar
  • 17 Neuenschwander PF, Morrissey JH. Roles of the membrane-interactive regions of factor VIIa and tissue factor. The factor VIIa Gla domain is dispensable for binding to tissue factor but important for activation of factor X. J Biol Chem 1994; 269: 8007-13.
    PubMedSuche in Google Scholar
  • 18 Smirnov MD, Ford DA, Esmon CT, Esmon NL. The effect of membrane composition on the hemostatic balance. Biochemistry 1999; 38: 3591-8.
    CrossrefPubMedSuche in Google Scholar
  • 19 van ’t Veer C, Butenas S, Golden NJ, Mann KG. Regulation of prothrombinase activity by protein S. Thromb Haemost 1999; 82: 80-7.
    Thieme ConnectPubMedSuche in Google Scholar
  • 20 Butenas S, Mann KG. Kinetics of human factor VII activation. Biochemistry 1996; 35: 1904-10.
    CrossrefPubMedSuche in Google Scholar
  • 21 Lawson JH, Kalafatis M, Stram S, Mann KG. A model for the tissue factor pathway to thrombin. I. An empirical study. J Biol Chem 1994; 269: 23357-66.
    PubMedSuche in Google Scholar
  • 22 Rand MD, Lock JB, van ’t Veer C, Gaffney DP, Mann KG. Blood clotting in minimally altered whole blood. Blood 1996; 88: 3432-45.
    PubMedSuche in Google Scholar
  • 23 Buteanas S, Cawthern KM, van’t Veer C, DiLorenzo ME, Lock JB, Mann KG. Antiplatelet agents in tissue factor-induced blood coagulation. Blood 2001; 97: 2314-22.
    CrossrefPubMedSuche in Google Scholar
  • 24 van ’t Veer C, Mann KG. Regulation of tissue factor initiated thrombin generation by the stoichiometric inhibitors tissue factor pathway inhibitor, antithrombin-III, and heparin cofactor-II. J Biol Chem 1997; 272: 4367-77.
    CrossrefPubMedSuche in Google Scholar
  • 25 Cawthern KM, van ’t Veer C, Lock JB, DiLorenzo M, Branda RF, Mann KG. Blood coagulation in hemophilia A and hemophilia C. Blood 1998; 91: 4581-92.
    PubMedSuche in Google Scholar
  • 26 Higgins DL, Mann KG. The interaction of bovine factor V and factor V-derived peptides with phospholipid vesicles. J Biol Chem 1983; 258: 6503-8.
    PubMedSuche in Google Scholar
  • 27 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.
    PubMedSuche in Google Scholar
  • 28 Katzmann JA, Nesheim ME, Hibbard LS, Mann KG. Isolation of functional human coagulation factor V by using a hybridoma antibody. Proc Natl Acad Sci USA 1981; 78: 162-6.
    CrossrefPubMedSuche in Google Scholar
  • 29 Griffith MJ, Noyes CM, Church FC. Reactive site peptide structural similarity between heparin cofactor II and antithrombin III. J Biol Chem 1985; 260: 2218-25.
    PubMedSuche in Google Scholar
  • 30 Dahlback B. Purification of human vitamin K-dependant protein S and its limited proteolysis by thrombin. Biochem J 1983; 209: 837-46.
    CrossrefPubMedSuche in Google Scholar
  • 31 Mustard JF, Perry DW, Ardlie NG, Packham MA. Preparation of suspension of washed platelets from humans. Br J Haematol 1972; 22: 193-204.
    CrossrefPubMedSuche in Google Scholar
  • 32 Nesheim ME, Tracy RP, Mann KG. “Clotspeed”, a mathematical simulation of the functional properties of prothrombinase. J Biol Chem 1984; 259: 1447-53.
    PubMedSuche in Google Scholar
  • 33 Rosing J, Tans G, Govers-Riemslag JWP, Zwaal RFA, Hemker HC. The role of phospholipids and factor Va in the prothrombinase complex. J Biol Chem 1980; 255: 274-83.
    PubMedSuche in Google Scholar
  • 34 van Dieijen G, Tans G, Rosing J, Hemker HC. The role of phospholipid and factor VIIIa in the activation of bovine factor X. J Biol Chem 1981; 256: 3433-42.
    PubMedSuche in Google Scholar
  • 35 Jones KC, Mann KG. A model of the tissue factor pathway to thrombin. J Biol Chem 1994; 269: 23367-73.
    PubMedSuche in Google Scholar
  • 36 Schafer AI. Approach to bleeding. In: Loscalzo J, Schafer AI. eds. Thrombosis and Hemorrhage. Baltimore, MD: Williams and Wilkins; 1998: 459-75.
    Suche in Google Scholar
  • 37 Miescher PA. Drug-induced thrombocytopenia. Semin Hematol 1973; 10: 311-25.
    PubMedSuche in Google Scholar
  • 38 Panella TJ, Peters W, White JG, Hannun YA, Greenberg CS. Platelets acquire a secretion defect after high-dose chemotherapy. Cancer 1990; 65: 1711-6.
    CrossrefPubMedSuche in Google Scholar
  • 39 Beutler E. Platelet transfusions: the 20,000/microL trigger. Blood 1993; 81: 1411-3.
    PubMedSuche in Google Scholar
  • 40 Gordon-Smith EC. Aplastic anaemia – aetiology and clinical features. Baillieres Clin Haematol 1989; 2: 1-18.
    CrossrefPubMedSuche in Google Scholar
  • 41 Roy AJ, Jaffe N, Djerassi I. Prophylactic platelet transfusions in children with acute leukemia: a dose response study. Transfusion 1973; 13: 283-90.
    CrossrefPubMedSuche in Google Scholar
  • 42 Lind SE. The bleeding time does not predict surgical bleeding. Blood 1991; 77: 2547-52.
    PubMedSuche in Google Scholar
  • 43 Rodgers RP, Levin J. A critical reappraisal of the bleeding time. Semin Thromb Hemost 1990; 16: 1-20.
    Thieme ConnectPubMedSuche in Google Scholar
  • 44 Walsh PN. Platelet coagulant activities and hemostasis: a hypothesis. Blood 1974; 43: 597-605.
    PubMedSuche in Google Scholar
  • 45 Bouchard BA, Catcher CS, Thrash BR, Adida C, Tracy PB. Effector cell protease receptor-1, a platelet activation-dependent membrane protein, regulates prothrombinase-catalyzed thrombin generation. J Biol Chem 1997; 272: 9244-51.
    CrossrefPubMedSuche in Google Scholar
  • 46 Hartwig JH. Platelet morphology. In: Loscalzo J, Schafer AI. eds. Thrombosis and Hemorrhage. Baltimore, MD: Williams and Wilkins; 1998: 207-28.
    Suche in Google Scholar
  • 47 Miletich JP, Kane WH, Hofmann SL, Stanford N, Majerus PW. Deficiency of factor Xa-factor Va binding sites on the platelets of a patient with a bleeding disorder. Blood 1979; 54: 1015-22.
    PubMedSuche in Google Scholar
  • 48 Tracy PB, Giles AR, Mann KG, Eide LL, Hoogendoom H, Rivard GE. Factor V (Quebec): a bleeding diathesis associated with a qualitative platelet factor V deficiency. J Clin Invest 1984; 74: 1221-8.
    CrossrefPubMedSuche in Google Scholar
  • 49 Nesheim ME, Nichols WL, Cole TL. et al. Isolation and study of an acquired inhibitor of human coagulation factor V. J Clin Invest 1986; 77: 405-15.
    CrossrefPubMedSuche in Google Scholar
  • 50 Tracy PB, Mann KG. Abnormal formation of the prothrombinase complex: factor V deficiency and related disorders. Hum Pathol 1987; 18: 162-9.
    CrossrefPubMedSuche in Google Scholar
  • 51 Monkovic DD, Tracy PB. Functional characterization of human platelet-released factor V and its activation by factor Xa and thrombin. J Biol Chem 1990; 265: 17132-40.
    PubMedSuche in Google Scholar