Measurement of Thrombin Generation in Whole Blood - The Effect of Heparin and Aspirin

 

PDF Download Buy Article Permissions and Reprints

Summary

A technique has been developed to monitor the development of thrombin in freshly collected whole blood in the absence of anticoagulants. It is based on the centrifugal separation of the cellular components from subsamples of blood drawn from non-anticoagulated clotting whole blood which are diluted in buffer containing a chromogenic substrate.

It is shown that the burst of thrombin generation after triggering coagulation with trace amounts of tissue thromboplastin occurs sooner in non-anticoagulated whole blood than in citrated whole blood. Heparin is shown to prolong the lag-time of thrombin generation more in native blood than in recalcified citrated blood.

It is also demonstrated that intake of 500 mg of aspirin significantly delays and inhibits thrombin generation in non-anticoagulated, thromboplastin triggered whole blood, whereas it has no effect on the coagulation in citrated plasma. The effect of aspirin intake on thrombin generation in blood is roughly equal to that of 0.03 U/ml of unfractionated heparin. This demonstrates that platelet reactions and the coagulation system are closely linked processes. It further lends support to the hypothesis that inhibition of thrombin generation is a common denominator of antithrombotic therapy.

• References

• 1 Biggs R, Macfarlane RG. Human blood coagulation and its disorders. Oxford: Blackwell Scientific Publications; 1953
• 2 Béguin S, Lindhout MT, Hemker HC. The effect of trace amounts of tissue thromboplastin on thrombin generation in platelet rich plasma, its inhibition by heparin. 1989; 60: 25-29
  PubMedGoogle Scholar
• 3 Béguin S, Lindhout T, Hemker HC. The mode of action of heparin in plasma. Thromb Haemost 1988; 60: 457-462
  Article in Thieme ConnectPubMedGoogle Scholar
• 4 Frank HLL, Hemker HC. The influence of glycerol on the Ca²⁺ flux in the isolated perfused rat heart. Basic Res Cardiol 1976; 71: 252-262
  CrossrefPubMedGoogle Scholar
• 5 Bevers E, Comfurius P, van Rijn JMML, 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: 429-436
  CrossrefPubMedGoogle Scholar
• 6 Roth GJ, Majerus PW. The mechanism of the effect of aspirin on human platelets. J Clin Invest 1975; 56: 624-632
  CrossrefPubMedGoogle Scholar
• 7 Rodgers RPC, Levin J. A critical reappraisal of the bleeding time. Semin Thromb Haemost 1990; 16: 1-20
  Article in Thieme ConnectPubMedGoogle Scholar
• 8 Chesebro JH, Clements IP, Fuster V, Elveback LR, Smith HC, Bardsley WT. et al. A platelet-inhibitor-drug trial in coronary-artery bypass operations. N Engl J Med 1982; 307: 73-78
  CrossrefPubMedGoogle Scholar
• 9 Lewis DH, Davis J, Archibald DG, Steinke WE, Smitherman TC, Doherty EJ. et al. Protective effects of aspirin against acute myocardial infarction and death in men with unstable angina. N Engl J Med 1983; 309: 396-403
  CrossrefPubMedGoogle Scholar
• 10 Cairns JA, Gent M, Singer J, Finnie KJ, Froggatt GM, Holder DA. et al. Aspirin, sulfinpyrazone, or both in unstable angina. N Engl J Med 1985; 313: 1369-1375
  CrossrefPubMedGoogle Scholar
• 11 UK TIA Study Group. United Kingdom transient ischaemic attack (UK-TIA) aspirin trial: interim results. Br Med J 1988; 296: 316-320
  CrossrefPubMedGoogle Scholar
• 12 Owren PA, Aas K. The control of dicumarol therapy and the quantitative determination of prothrombin and proconvertin. Scand J Clin Lab Invest 1951; 3: 201-218
  CrossrefPubMedGoogle Scholar
• 13 Hemker HC, Willems GM, Beguin SA. A computer assisted method to obtain the prothrombin activation velocity in whole plasma independent of thrombin decay processes. Thromb Haemost 1986; 56: 9-17
  Article in Thieme ConnectPubMedGoogle Scholar
• 14 Documenta Geigy. Wissenschaftliche Tabelle, 7th edt 1968
  PubMedGoogle Scholar
• 15 Bendetowicz AV, Beguin S, Caplain H, Hemker HC. Pharmacokinetics and pharmacodynamics of a low molecular weight heparin (Enoxaparin) after subcutaneous injection, comparison with unfractionated heparin. A three way cross over study in human volunteers. Thromb Haemost 1994; 71: 305-313
  Article in Thieme ConnectPubMedGoogle Scholar
• 16 Weiss HJ, Lages B. Evidence for tissue factor-dependent activation of the classic extrinsic coagulation mechanism in blood obtained from bleeding time wounds. Blood 1988; 71: 629-635
  PubMedGoogle Scholar
• 17 Heras M, Chesebro JH, Webster MWI, Mruk JS, Grill DE, Penny WJ. et al. Hirudin, heparin, and placebo during deep arterial injury in the pig. Circulation 1990; 82: 1476-1484
  CrossrefPubMedGoogle Scholar
• 18 The sixty plus reinfarction study research group. A double-blind trial to assess long term oral anticoagulant therapy in elderly patients after myocardial infarction. Lancet 1980; 2: 989-994
  PubMedGoogle Scholar
• 19 Turpie AG, Levine MN, Hirsh J, Carter CJ, Jay RM, Powers PJ. et al. A randomized controlled trial of a low-molecular-weight heparin (enoxaparin) to prevent deep-vein thrombosis in patients undergoing elective hip surgery. N Engl J Med 1986; 315: 925-929
  CrossrefPubMedGoogle Scholar
• 20 Theroux P, Quimet H, McCans J, Latppr JQ, Jojy P, Levy G. et al. Aspirin, heparin, or both to treat acute unstable angina. N Engl J Med 1988; 319: 1105-1111
  CrossrefPubMedGoogle Scholar
• 21 Pedersen AK, Fitzgerald GA. Dose-related kinetics of aspirin. New Engl J Med 1984; 311 (19) 1206-1211
  CrossrefPubMedGoogle Scholar
• 22 Basic-Micic M, Breddin HK. PITT and detection of a risk of venous and arterial thrombosis. Abstract, XHIth congress ISTH, Amsterdam. Thromb Haemost 1991; 65: 1318-1325
  PubMedGoogle Scholar
• 23 Kyrle PA, Westwick J, Scully MF, Kakkar VV, Lewis GP. Investigation of the interaction of blood platelets with the coagulation system at the site of plug formation in vivo in man - Effect of low-dose aspirin. Thromb Haemost 1987; 57 (01) 62-66
  Article in Thieme ConnectPubMedGoogle Scholar
• 24 Baruch D, Hemker HC, Lindhout T. Kinetics of thrombin-induced release and activation of platelet factor V. Eur J Bioch 1986; 154: 213-218
  CrossrefPubMedGoogle Scholar