Download PDF
Thromb Haemost 2000; 84(03): 492-498
DOI: 10.1055/s-0037-1614050
Commentary
Schattauer GmbH

GPIIb-IIIa Antagonist-induced Reduction in Platelet Surface Factor V/Va Binding and Phosphatidylserine Expression in Whole Blood

Mark I. Furman
1   From the Center for Platelet Function Studies, Worcester, MA
2   Division of Cardiovascular Medicine, Departments of Medicine, Worcester, MA
,
Lori A. Krueger
1   From the Center for Platelet Function Studies, Worcester, MA
3   Pediatrics, Worcester, MA
,
A. L. Frelinger III
1   From the Center for Platelet Function Studies, Worcester, MA
3   Pediatrics, Worcester, MA
,
Marc R. Barnard
1   From the Center for Platelet Function Studies, Worcester, MA
3   Pediatrics, Worcester, MA
,
Mary Ann Mascelli
5   Centocor, Inc., Malvern PA, USA
,
Marian T. Nakada
5   Centocor, Inc., Malvern PA, USA
,
Alan D. Michelson
1   From the Center for Platelet Function Studies, Worcester, MA
3   Pediatrics, Worcester, MA
4   Surgery, University of Massachusetts Medical School, Worcester, MA
› Author AffiliationsFunded in part by Centocor, Inc., Malvern, PA.
Further Information

Publication History

Received 07 January 2000

Accepted after resubmission 14 April 2000

Publication Date:
14 December 2017 (online)

Also available at
    Permissions and Reprints

Summary

In addition to inhibition of platelet aggregation, GPIIb-IIIa antagonists may reduce thrombotic events via other mechanisms. In a novel whole blood flow cytometric system, we investigated the effects of GPIIb-IIIa antagonists, in the presence or absence of thrombin inhibitors, on platelet surface-bound factor V/Va and platelet surface phospholipids. Diluted venous blood was incubated with either buffer or a GPIIb-IIIa antagonist (abciximab, tirofiban, or eptifibatide). Some samples were pre-incubated with clinically relevant concentrations of unfractionated heparin (UFH), a low molecular weight heparin, a direct thrombin inhibitor, or buffer only. Platelets were then activated and labeled with mAb V237 (factor V/Va-specific) or annexin V (binds phosphatidylserine), fixed, and analyzed by flow cytometry. In the absence of thrombin inhibitors, GPIIb-IIIa antagonists (especially abciximab) significantly reduced agonist-induced platelet procoagulant activity, as determined by reduced binding of V237 and annexin V. At high pharmacologic concentrations, unfractionated heparin and enoxaparin, but not hirudin, further reduced factor V/Va binding to the surface of activated platelets in the presence of GPIIb-IIIa antagonists. Agonist-induced platelet procoagulant activity was reduced in a patient with Glanzmann’s thrombasthenia. We conclude that GPIIb-IIIa antagonists reduce platelet procoagulant activity in whole blood and heparin and enoxaparin augment this reduction. Fibrinogen binding to GPIIb-IIIa is important in the generation of platelet procoagulant activity.

Key words

Platelets - coagulation - glycoproteins - platelet aggregation inhibitors - anticoagulants
 

  • References

  • 1 Fuster V, Badimon L, Badimon JJ, Chesebro JH. The pathogenesis of coronary artery disease and the acute coronary syndromes. N Engl J Med 1992; 326: 242-50.
    CrossrefPubMedSearch in Google Scholar
  • 2 The Antiplatelet Trialist Collaboration. Collaborative overview of randomised trials of antiplatelet therapy. Prevention of death, myocardial infarction, and stroke by prolonged antiplatelet therapy in various categories of patients. Br Med J 1994; 308: 1540.
    CrossrefPubMedSearch in Google Scholar
  • 3 The PURSUIT Trial Investigators. Inhibition of platelet glycoprotein IIb/ IIIa with eptifibatide in patients with acute coronary syndromes. The PURSUIT Trial Investigators. Platelet Glycoprotein IIb/IIIa in Unstable Angina: Receptor Suppression Using Integrilin Therapy. N Engl J Med 1998; 339: 436-43.
    CrossrefPubMedSearch in Google Scholar
  • 4 The PRISM STUDY Investigators. A comparison of aspirin plus tirofiban with aspirin plus heparin for unstable angina. Platelet Receptor Inhibition in Ischemic Syndrome Management (PRISM) Study Investigators. N Engl J Med 1998; 338: 1498-505.
    CrossrefPubMedSearch in Google Scholar
  • 5 The PRISM-PLUS Study Investigators. Inhibition of the platelet glycoprotein IIb/IIIa receptor with tirofiban in unstable angina and non-Q-wave myocardial infarction. Platelet Receptor Inhibition in Ischemic Syndrome Management in Patients Limited by Unstable Signs and Symptoms (PRISM-PLUS) Study Investigators [published erratum appears in N Engl J Med 1998 Aug 6; 339 (6): 415]. N Engl J Med 1998; 338: 1488-97.
    CrossrefPubMedSearch in Google Scholar
  • 6 The EPIC Investigators. Use of monoclonal antibody directed against the platelet glycoprotein IIb/IIIa receptor in high risk angioplasty. N Engl J Med 1994; 330: 956-61.
    CrossrefPubMedSearch in Google Scholar
  • 7 The EPILOG Investigators. Platelet glycoprotein IIb/IIIa receptor blockade and low-dose heparin during percutaneous coronary revascularization. N Engl J Med 1997; 336: 1689-96.
    CrossrefPubMedSearch in Google Scholar
  • 8 The CAPTURE Study Investigators. Randomised placebo-controlled trial of abciximab before and during coronary intervention in refractory unstable angina: the CAPTURE Study [published erratum appears in Lancet 1997 Sep 6; 350 (9079): 744]. Lancet 1997; 349: 1429-35.
    CrossrefPubMedSearch in Google Scholar
  • 9 The EPISTENT Investigators. Randomised placebo-controlled and balloonangioplasty-controlled trial to assess safety of coronary stenting with use of platelet glycoprotein-IIb/IIIa blockade. The EPISTENT Investigators. Evaluation of Platelet IIb/IIIa Inhibitor for Stenting. Lancet 1998; 352: 87-92.
    CrossrefPubMedSearch in Google Scholar
  • 10 Dachary-Prigent J, Freyssinet JM, Pasquet JM, Carron JC, Nurden AT. Annexin V as a probe of aminophospholipid exposure and platelet membrane vesiculation: a flow cytometry study showing a role for free sulfhydryl groups. Blood 1993; 81: 2554-65.
    PubMedSearch in Google Scholar
  • 11 Sandberg H, Andersson LO, Hoglund S. Isolation and characterization of lipid-protein particles containing platelet factor III released from human platelets. Biochem J 1982; 203: 303-11.
    CrossrefPubMedSearch in Google Scholar
  • 12 Rosing J, van Rijn JL, 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.
    PubMedSearch in Google Scholar
  • 13 Sims PJ, Wiedmer T, Esmon CT, Weiss HJ, Shattil SJ. Assembly of the platelet prothrombinase complex is linked to vesiculation of the platelet plasma membrane. Studies in Scott syndrome: an isolated defect in platelet procoagulant activity. J Biol Chem 1989; 264: 17049-57.
    PubMedSearch in Google Scholar
  • 14 Chang CP, Zhao J, Wiedmer T, Sims PJ. Contribution of platelet microparticle formation and granule secretion to the transmembrane migration of phosphatidylserine. J Biol Chem 1993; 268: 7171-8.
    PubMedSearch in Google Scholar
  • 15 Thiagarajan P, Tait JF. Collagen-induced exposure of anionic phospholipid in platelets and platelet-derived microparticles. J Biol Chem 1991; 266: 24302-7.
    PubMedSearch in Google Scholar
  • 16 Mann KG. Prothrombin and thrombin. In: Colman WR, Hirsh J, Marder JV, Salzman WE. eds. Hemostasis and Thrombosis. Basic Principles and Clinical Practice. 3rd ed. Philadelphia: J.P. Lippincott Company; 1994: 184-200.
    Search in Google Scholar
  • 17 Rosing J, Tans G. Coagulation factor V: an old star shines again. Thromb Haemost 1997; 78: 427-33.
    Thieme ConnectPubMedSearch in Google Scholar
  • 18 Weng G, Bhalla US, Iyengar R. Complexity in biological signaling systems. Science 1999; 284: 92-6.
    CrossrefPubMedSearch in Google Scholar
  • 19 Ammar T, Scudder LE, Coller BS. In vitro effects of the platelet glycoprotein IIb/IIIa receptor antagonist c7E3 Fab on the activated clotting time. Circulation 1997; 95: 614-7.
    CrossrefPubMedSearch in Google Scholar
  • 20 Moliterno DJ, Califf RM, Aguirre FV. et al. Effect of platelet glycoprotein IIb/IIIa integrin blockade on activated clotting time during percutaneous transluminal coronary angioplasty or directional atherectomy (the EPIC trial). Am J Cardiol 1995; 75: 559-62.
    CrossrefPubMedSearch in Google Scholar
  • 21 Dangas G, Badimon JJ, Coller BS. et al. Administration of abciximab during percutaneous coronary intervention reduces both ex vivo platelet thrombus formation and fibrin deposition: implications for a potential anticoagulant effect of abciximab. Arterioscler Thromb Vasc Biol 1998; 18: 1342-9.
    CrossrefPubMedSearch in Google Scholar
  • 22 Pedicord DL, Thomas BE, Mousa SA, Dicker IB. Glycoprotein IIb/IIIa receptor antagonists inhibit the development of platelet procoagulant activity. Thromb Res 1998; 90: 247-58.
    CrossrefPubMedSearch in Google Scholar
  • 23 Keularts IM, Beguin S, de Zwaan C, Hemker HC. Treatment with a GPIIb/IIIa antagonist inhibits thrombin generation in platelet rich plasma from patients. Thromb Haemost 1998; 80: 370-1.
    Thieme ConnectPubMedSearch in Google Scholar
  • 24 Cheresh DA, Spiro RC. Biosynthetic and functional properties of an Arg-Gly-Asp-directed receptor involved in human melanoma cell attachment to vitronectin, fibrinogen, and von Willebrand factor. J Biol Chem 1987; 262: 17703-11.
    PubMedSearch in Google Scholar
  • 25 Cheresh DA. Human endothelial cells synthesize and express an Arg-Gly-Asp-directed adhesion receptor involved in attachment to fibrinogen and von Willebrand factor. Proc Natl Acad Sci USA 1987; 84: 6471-5.
    CrossrefPubMedSearch in Google Scholar
  • 26 Coller BS, Peerschke EI, Scudder LE, Sullivan CA. A murine monoclonal antibody that completely blocks the binding of fibrinogen to platelets produces a thrombasthenic-like state in normal platelets and binds to glycoproteins IIb and/or IIIa. J Clin Invest 1983; 72: 325-38.
    CrossrefPubMedSearch in Google Scholar
  • 27 Gralnick HR, Williams SB, McKeown L. et al. Endogenous platelet fibrinogen expression on activated platelets. J Lab Clin Med 1991; 118: 604-13.
    PubMedSearch in Google Scholar
  • 28 Gralnick HR, Williams SB, McKeown L. et al. Endogenous platelet fibrinogen: its modulation after surface expression is related to sizeselective access to and conformational changes in the bound fibrinogen. Br J Haematol 1992; 80: 347-57.
    CrossrefPubMedSearch in Google Scholar
  • 29 Michelson AD. Flow cytometric analysis of platelet surface glycoproteins: Phenotypically distinct subpopulations of platelets in children with chronic myeloid leukemia. J Lab Clin Med 1987; 110: 346-54.
    PubMedSearch in Google Scholar
  • 30 Michelson AD, Ellis PA, Barnard MR, Matic GB, Viles AF, Kestin AS. Downregulation of the platelet surface glycoprotein Ib-IX complex in whole blood stimulated by thrombin, ADP or an in vivo wound. Blood 1991; 77: 770-9.
    PubMedSearch in Google Scholar
  • 31 Thiagarajan P, Tait JF. Binding of annexin V/placental anticoagulant protein I to platelets. Evidence for phosphatidylserine exposure in the procoagulant response of activated platelets. J Biol Chem 1990; 265: 17420-3.
    PubMedSearch in Google Scholar
  • 32 Andree HA, Reutelingsperger CP, Hauptmann R, Hemker HC, Hermens WT, Willems GM. Binding of vascular anticoagulant alpha (VAC alpha) to planar phospholipid bilayers. J Biol Chem 1990; 265: 4923-8.
    PubMedSearch in Google Scholar
  • 33 Raynal P, Pollard HB. Annexins: the problem of assessing the biological role for a gene family of multifunctional calciumand phospholipid-binding proteins. Biochim Biophys Acta 1994; 1197: 63-93.
    CrossrefPubMedSearch in Google Scholar
  • 34 Tam SH, Sassoli PM, Jordan RE, Nakada MT. Abciximab (ReoPro, chimeric 7E3 Fab) demonstrates equivalent affinity and functional blockade of glycoprotein IIb/IIIa and αvβ3 integrins. Circulation 1998; 98: 1085-91.
    CrossrefPubMedSearch in Google Scholar
  • 35 Reverter JC, Beguin S, Kessels H, Kumar R, Hemker HC, Coller BS. Inhibition of platelet-mediated, tissue factor-induced thrombin generation by the mouse/human chimeric 7E3 antibody. Potential implications for the effect of c7E3 Fab treatment on acute thrombosis and “clinical restenosis”. J Clin Invest 1996; 98: 863-74.
    CrossrefPubMedSearch in Google Scholar
  • 36 Gemmell CH, Sefton MV, Yeo EL. Platelet-derived microparticle formation involves glycoprotein IIb-IIIa. Inhibition by RGDS and a Glanzmann’s thrombasthenia defect. J Biol Chem 1993; 268: 14586-9.
    PubMedSearch in Google Scholar
  • 37 Michelson AD. Flow cytometry: A clinical test of platelet function. Blood 1996; 87: 4925-36.
    PubMedSearch in Google Scholar
  • 38 Santos MT, Valles J, Marcus AJ. et al. Enhancement of platelet reactivity and modulation of eicosanoid production by intact erythrocytes. A new approach to platelet activation and recruitment. J Clin Invest 1991; 87: 571.
    CrossrefPubMedSearch in Google Scholar
  • 39 LaRosa CA, Rohrer MJ, Benoit SE, Rodino LJ, Barnard MR, Michelson AD. Human neutrophil cathepsin G is a potent platelet activator. J Vasc Surg 1994; 19: 306-18.
    CrossrefPubMedSearch in Google Scholar
  • 40 Udis BJ, Hickerson D, Bode AP. Vesiculation versus light scatter shifts in activated platelets. Cytometry 1994; (Suppl. 07) 54.
    PubMedSearch in Google Scholar
  • 41 Nesheim ME, Furmaniak-Kazmierczak E, Henin C, Cote G. On the existence of platelet receptors for factor V(a) and factor VIII(a). Thromb Haemost 1993; 70: 80-6.
    Thieme ConnectPubMedSearch in Google Scholar
  • 42 Alberio L, Safa O, Clemetson KJ, Esmon CT, Dale GL. Surface expression and functional characterization of alpha-granule factor V in human platelets: effects of ionophore A23187, thrombin, collagen, and convulxin (In Process Citation). Blood 2000; 95: 1694-702.
    CrossrefPubMedSearch in Google Scholar
  • 43 Hayward CP, Furmaniak-Kazmierczak E, Cieutat AM. et al. Factor V is complexed with multimerin in resting platelet lysates and colocalizes with multimerin in platelet alpha-granules. J Biol Chem 1995; 270: 19217-24.
    CrossrefPubMedSearch in Google Scholar
  • 44 Coller BS, Cheresh DA, Asch E, Seligsohn U. Platelet vitronectin receptor expression differentiates Iraqi-Jewish from Arab patients with Glanzmann’s thrombasthenia in Israel. Blood 1991; 77: 75-83.
    PubMedSearch in Google Scholar
  • 45 Agnelli G. Thrombin plays a pivotal role in vascular re-occlusion after PTCA and coronary thrombolysis. Cardiovasc Res 1996; 31: 232-4.
    CrossrefPubMedSearch in Google Scholar
  • 46 Gallo R, Padurean A, Toschi V. et al. Prolonged thrombin inhibition reduces restenosis after balloon angioplasty in porcine coronary arteries. Circulation 1998; 97: 581-8.
    CrossrefPubMedSearch in Google Scholar
  • 47 Mascelli MA, Lance ET, Damaraju L, Wagner CL, Weisman HF, Jordan RE. Pharmacodynamic profile of short-term abciximab treatment demonstrates prolonged platelet inhibition with gradual recovery from GP IIb/IIIa receptor blockade. Circulation 1998; 97: 1680-8.
    CrossrefPubMedSearch in Google Scholar
  • 48 Topol EJ, Ferguson JJ, Weisman HF. et al. Long-term protection from myocardial ischemic events in a randomized trial of brief integrin beta3 blockade with percutaneous coronary intervention. EPIC Investigator Group. Evaluation of Platelet IIb/IIIa Inhibition for Prevention of Ischemic Complication. JAMA 1997; 278: 479-84.
    CrossrefPubMedSearch in Google Scholar
  • 49 Madan M, Berkowitz SD, Tcheng JE. Glycoprotein IIb/IIIa integrin blockade. Circulation 1998; 98: 2629-35.
    CrossrefPubMedSearch in Google Scholar
  • 50 Beguin S, Kumar R, Keularts I, Seligsohn U, Coller BS, Hemker HC. Fibrin-dependent platelet procoagulant activity requires GPIb receptors and von Willebrand factor. Blood 1999; 93: 564-70.
    PubMedSearch in Google Scholar
  • 51 Fox JE, Reynolds CC, Austin CD. The role of calpain in stimulus-response coupling: evidence that calpain mediates agonist-induced expression of procoagulant activity in platelets. Blood 1990; 76: 2510-9.
    PubMedSearch in Google Scholar
  • 52 Fox JE, Austin CD, Reynolds CC, Steffen PK. Evidence that agonistinduced activation of calpain causes the shedding of procoagulant-containing microvesicles from the membrane of aggregating platelets. J Biol Chem 1991; 266: 13289-95.
    PubMedSearch in Google Scholar
  • 53 Felsenfled DP, Schwartzberg PL, Venegas A, Tse R, Sheetz MP. Selective regulation of integrin-cytoskeletal interactions by the tyrsoine kinase Src. Nature Cell Biol 1999; 01: 200-6.
    CrossrefPubMedSearch in Google Scholar
  • 54 Defilippi P, Venturino M, Gulino D. et al. Dissection of pathways implicated in integrin-mediated actin cytoskeleton assembly. J Biol Chem 1997; 272: 21726-34.
    CrossrefPubMedSearch in Google Scholar
  • 55 Furman MI, Gardner TM, Goldschmidt-Clermont PJ. Mechanisms of cytoskeletal reorganization during platelet activation. Thromb Haemost 1993; 70: 229-32.
    Thieme ConnectPubMedSearch in Google Scholar
  • 56 Byzova TV, Plow EF. Networking in the hemostatic system. Integrin alpha IIb β3 binds prothrombin and influences its activation. J Biol Chem 1997; 272: 27183-8.
    CrossrefPubMedSearch in Google Scholar
  • 57 Byzova TV, Plow EF. Activation of αvβ3 on vascular cells controls recognition of prothrombin. J Cell Biol 1998; 143: 2081-92.
    CrossrefPubMedSearch in Google Scholar
  • 58 Furman MI, Benoit SE, Barnard MR. et al. Increased platelet reactivity and circulating monocyte-platelet aggregates in patients with stable coronary artery disease. J Am Coll Cardiol 1998; 31: 352-8.
    PubMedSearch in Google Scholar
  • 59 Feng D, Lindpaintner K, Larson MG. et al. Increased platelet aggregability associated with platelet GPIIIa PlA2 polymorphism: the Framingham Offspring Study. Arterioscler Thromb Vasc Biol 1999; 19: 1142-7.
    CrossrefPubMedSearch in Google Scholar
  • 60 Michelson AD, Furman MI, Goldschmidt-Clermont P. et al. Platelet GP IIIa Pl(A) polymorphisms display different sensitivities to agonists. Circulation 2000; 101: 1013-8.
    CrossrefPubMedSearch in Google Scholar