Download PDF
Thromb Haemost 1995; 74(03): 962-968
DOI: 10.1055/s-0038-1649856
Original Article
Platelets
Schattauer GmbH Stuttgart

The Effect of Fibrin Clots and Clot-Bound Thrombin on the Development of Platelet Procoagulant Activity

Rachana Kumar
The Department of Biochemistry, Faculty of Medicine and Cardiovascular Research Institute,University of Limburg, Maastricht, The Netherlands
,
Suzette Béguin
The Department of Biochemistry, Faculty of Medicine and Cardiovascular Research Institute,University of Limburg, Maastricht, The Netherlands
,
H Coenraad Hemker
The Department of Biochemistry, Faculty of Medicine and Cardiovascular Research Institute,University of Limburg, Maastricht, The Netherlands
› Author Affiliations
Further Information

Publication History

Received 13 February 1995

Accepted after resubmission 23 May 1995

Publication Date:
09 July 2018 (online)

Also available at
  PDF Download  Permissions and Reprints

Summary

We tested different types of clot for their ability to provoke procoagulant activity in platelets: normal clots from platelet poor plasma (des AABB- or fibrin II clots), similar clots in which the adsorbed thrombin has been inhibited by hirudin, and clots obtained by the action of two snake venom enzymes that release only fibrinopeptide A (des AA- or fibrin I clots). Analogous clots from fibrinogen solutions were also tested.

In platelet rich plasma (PRP), where platelet coagulant phospholipids (PCP) are rate limiting for thrombin generation, the addition of any type of clot enhances the generation of thrombin thus it induces the appearance of PCP. Clots containing active adsorbed thrombin are the most potent ones in this respect. Lactate dehydrogenase (LDH) levels do not increase in the course of the thrombin generation so the platelets are not damaged in the process. Non-centrifugable PCP could be demonstrated to appear during the process, so the production of procoagulant phospholipid microparticles must be part of the mechanism. Membrane transbilayer phosphatidyl serine movement (flip-flop) can not be demonstrated in PRP as the activated platelets are caught in the emerging clot.

In order to demonstrate flip-flop, we tried to investigate the influence of clots on washed platelets. However, contrary to platelets in a plasma milieu, isolated platelets are damaged by fibrin clots, especially in the presence of thrombin, as can be judged from the appearance of LDH

We conclude that, in PRP, clots induce the appearance of PCP from platelets by vesiculation, possibly accompanied by flip-flop and that thrombin accelerates the process but is not an absolute requirement

 

  • References

  • 1 Hemker HC. Thrombin Generation, an Essential Step in Haemostasis and Thrombosis. In: Haemostasis and Thrombosis. 3rd ed Bloom AL, Forbes CD, Thomas DP, Tuddenham EGD. (eds) London: Churchill Livingstone: 1993. ISBN 0-443-0452lb pp 477-490
    Search in Google Scholar
  • 2 Walsh PN. Different requirements for intrinsic factor Xa forming activity and platelet factor 3 activity and their relationship to platelet aggregation and secretion. Br J Haematol 1978; 40: 311
    CrossrefPubMedSearch in Google Scholar
  • 3 Bevers EM, Comfurius P, van Rijn JLML, Hemker HC, Zwaal RFA. Generation of Prothrombin converting activity and the exposure of phosphati- dylserine at the outer surface of platelets. Eur J Biochem 1982; 122: 429-436
    CrossrefPubMedSearch in Google Scholar
  • 4 Rosing J, van Rijn JLML, Bevers EM, van Dieijen G, Comfurius P, Zwaal RFA. The role of activated human platelets in prothrombin and factor X activation. Blood 1985; 65: 319-332
    PubMedSearch in Google Scholar
  • 5 Miletich JP, Jackson CM, Majerus PW. Properties of the factor Xa binding site on platelets. J Biol Chem 1978; 253: 6908-6916
    PubMedSearch in Google Scholar
  • 6 Kane WH, Lindhout MJ, Jackson CM, Majerus PW. Factor Va-dependent binding of factor Xa to human platelets. J Biol Chem 1980; 255: 1170-1174
    PubMedSearch in Google Scholar
  • 7 Chasney CM, Pifer D, Colman RW. Subcellular localization and secretion of factor V from human platelets. Proc Natl Acad Sci USA 1981; 78: 5180-5184
    CrossrefPubMedSearch in Google Scholar
  • 8 Liu CY, Nossel HL, Kaplan KL. The binding of thrombin by fibrin. J Biol Chem 1979; 254: 10421-10425
    PubMedSearch in Google Scholar
  • 9 Kaminski M, Mcdonagh J. Inhibited thrombins, interactions with fibrinogen and fibrin. Biochem J 1987; 242: 881-887
    CrossrefPubMedSearch in Google Scholar
  • 10 Wilner GD, Danitz MP, Mudd S, Hsieh K, Fenton JW. Selective immobalization of thrombin by surface bound fibrin. J Lab Clin Med 1981; 97: 403-411
    PubMedSearch in Google Scholar
  • 11 Hogg PJ, Jackson CM. Fibrin monomer protects thrombin from inactivation by heparin-antithrombin III: Implications for heparin efficacy. Proc Natl Acad Sci USA 1989; 86: 3619-3623
    CrossrefPubMedSearch in Google Scholar
  • 12 Weitz JI, Hudoba M, Massel D, Maraganore J, Hirsh J. Clot bound thrombin is protected from inhibition by heparin-antithrombin III but is susceptible to inactivation by antithrombin III independent inhibitors. J Clin Invest 1990; 86: 385-391
    CrossrefPubMedSearch in Google Scholar
  • 13 Francis WC, Markham ER, Barlow HG, Florack MT, Dobrzynski MD, Marder JV. Thrombin activity of fibrin thrombi and soluble plasmic derivatives. J Lab Clin Med 1983; 102: 220-230
    PubMedSearch in Google Scholar
  • 14 Prins MH, Weitz JI. Heparin inhibits the amplification of coagulation mediated by clot-bound thrombin. Thromb Haemost 1991; 65: 759 abstract 319
    PubMedSearch in Google Scholar
  • 15 Bendayan P, Boccalon H, Dupouy D, Boneu B. Dermatan sulfate is a more potent inhibitor of clot-bound thrombin than unfractionated and low molecular weight heparins. Thromb Haemost 1994; 71: 576-80
    Thieme ConnectPubMedSearch in Google Scholar
  • 16 Buchanan A. Original communications on the coagulation of the blood and other fibriniferous liquids. London: Med Gaz: 1845
    Search in Google Scholar
  • 17 Béguin S, Kessels H, Hemker HC. Feedback reactions in blood coagulation are mediated by fibrin bound thrombin which is not inhibited by heparin. Thromb Haemost 1993; 69: 811 Abstract 962
    PubMedSearch in Google Scholar
  • 18 Kumar R, Béguin S, Hemker HC. The influence of fibrinogen and fibrin on thrombin generation-evidence for feedback activation of the clotting system by clot bound thrombin. Thromb Haemost 1994; 72 (05) 713-21
    Thieme ConnectPubMedSearch in Google Scholar
  • 19 Keiffer N, Phillips DR. Platelet membrane glycoproteins: Functions in cellular interactions. Annu Rev Cell Biol 1990; 6: 329-357
    CrossrefPubMedSearch in Google Scholar
  • 20 Bevers E, Comfurius P, Zwaal RFA. Changes in membrane phospholipid distribution during platelet activation. Biochim et Biophysica Acta 1983; 736: 57-66
    PubMedSearch in Google Scholar
  • 21 Discipio RG, Hermodson MA, Yates SG, Davie EW. A comparison of human prothrombin, factor IXa (Christmas factor), factor X (Stuart factor) and protein S. Biochemistry 1977; 16: 698-706
    CrossrefPubMedSearch in Google Scholar
  • 22 Rosing J, Tans G, Govers-Riemslag JW P, Zwaal RF A, Hemker HC. The role of phospholipids and factor Va in the prothrombinase complex. J Biol Chem 1980; 255: 274-283
    PubMedSearch in Google Scholar
  • 23 Chase Jr T, Shaw E. Comparison of the esterase activities of trypsin, plas- min and thrombin on guanidinobenzoate esters. Titration of the enzymes Biochemistry 1969; 8: 2212-2224
    CrossrefPubMedSearch in Google Scholar
  • 24 Fujikawa K, Legaz ME, Davie EW. Bovine factor X1(Stuart factor). Mechanism of activation by protein from Russell’s viper venom Biochemistry 1972; 11: 4892-4899
    CrossrefPubMedSearch in Google Scholar
  • 25 Smith RL. Titration of activated bovine factor X. J Biol Chem 1973; 248: 2418-2423
    PubMedSearch in Google Scholar
  • 26 Dählback B. Human coagulation factor V purification and thrombin-catalyzed activation. J Clin Invest 1980; 66: 583-551
    CrossrefPubMedSearch in Google Scholar
  • 27 Suzuki K, Dählback B, Stenflo J. Thrombin-catalyzed activation of human coagulation factor V. J Biol Chem 1982; 257: 6556-6564
    PubMedSearch in Google Scholar
  • 28 Tans G, Rosing J, Thomassen MC L G D, Heeb MJ, Zwaal RFA, Griffin P. Comparison of anticoagulant and procoagulant activities of stimulated platelets and platelet derived microparticles. Blood 1991; 77 (12) 2641-2648
    CrossrefPubMedSearch in Google Scholar
  • 29 Lindhout T, Govers-Riemslag JW P, van de Waart P, Hemker HC, Rosing J. Factor Va-factor Xa interactions. Effect of phospholipid vesicles of varying composition Biochemistry 1982; 21: 5494-54502
    CrossrefPubMedSearch in Google Scholar
  • 30 Béguin S, Lindhout T, Hemker HC. The effect of trace amounts of Tissue Factor on Thrombin Generation in Platelet Rich Plasma, its inhibition by Heparin. Thromb Haemost 1989; 61 (01) 25-29
    Thieme ConnectPubMedSearch in Google Scholar
  • 31 van Rijn JLML, Rosing J, van Dieijen G. Activity of human blood platelets in prothrombin and in factor X activation induced by lonophore A23187. Eur J Biochem 1983; 133: 1-10
    CrossrefPubMedSearch in Google Scholar
  • 32 Schroit AJ, Zwaal RFA. Transbilayer movement of phospholipids in red cell and platelet membrane. Biochim Biophys Acta 1991; 1071 (03) 313-329
    CrossrefPubMedSearch in Google Scholar
  • 33 Sandberg H, Andersson LO, Hoglund S. Isolation and characterization of lipid-protein particles containing platelet factor 3 released from human platelets. Biochem J 1982; 203 (01) 303-311
    CrossrefPubMedSearch in Google Scholar
  • 34 Agache P, Blanc D. Current status in sebum knowledge. Int J Dermatol 1982; 21 (06) 304-315
    CrossrefPubMedSearch in Google Scholar
  • 35 Wester J, Sixma JJ, Geuze JJ, van der Veen J. Morphology of the hemostatic plug in human skin wounds. Transformation of the plug Laboratory Investigation 1979; 41: 182-192
    PubMedSearch in Google Scholar