Absence of Synergism Between Tissue-Type Plasminogen Activator (t-PA), Single-Chain UrokinaseType Plasminogen Activator (scu-PA) and Urokinase on Clot Lysis in a Plasma Milieu In Vitro

Summary

A potential synergic effect of tissue-type plasminogen activator (t-PA), single-chain urokinase-type plasminogen activator (scuPA) or urokinase on clot lysis was investigated in a whole human plasma system in vitro. The system consisted of a human plasma clot labeled with ¹²⁵I-fibrinogen, immersed in titrated whole human plasma, to which the thrombolytic agents were added. Clot lysis was quantitated by measurement of released ¹²⁵I, and activation of the fibrinolytic system in the surrounding plasma by measurements of fibrinogen and α₂-antiplasmin.

t-PA, scu-PA and urokinase induced a dose-dependent and time-dependent clot lysis; 50 percent lysis after 2 h was obtained with 5 nM t-PA, 20 nM scu-PA and 12 nM urokinase. At these concentrations no significant activation of the fibrinolytic system in the plasma was observed with t-PA and scu-PA, whereas urokinase caused significant α₂-antiplasmin consumption and concomitant fibrinogen degradation. The shape of the dose-response curves was different; t-PA and urokinase showed a log linear dose-response whereas that of scu-PA was sigmoidal.

Keywords

t-PA - scu-PA - Urokinase - Synergism - Thrombolysis

• References

• 1 Report of the Subcommittee on Fibrinolysis, San Diego, CA, USA, July 13, 1985. Thromb Haemostas 1985; 54: 893
• 2 Hoylaerts M, Rijken DC, Lijnen HR, Collen D. Kinetics of the activation of plasminogen by human tissue-plasminogen activator. Role of fibrin. J Biol Chem 1982; 257: 2912-2919
• 3 Suenson O, Lützen O, Thorsen S. Initial plasmin-degradation of fibrin as the basis of a positive feed-back mechanism in fibrinolysis. Eur J Biochem 1984; 140: 513-522
• 4 Norrman B, Wallen P, Ranby M. Fibrinolysis mediated by tissue plasminogen activator. Disclosure of a kinetic transition. Eur J Biochem 1985; 149: 193-200
• 5 Husain S, Gurewich V, Lipinski B. Purification and partial characterization of a single-chain high-molecular-weight form of urokinase from human urine. Arch Biochem Biophys 1983; 220: 31-38
• 6 Stump DC, Thienpont M, Collen D. Urokinase-related proteins in human urine. Isolation and characterization of single-chain urokinase (pro-urokinase) and urokinase-inhibitor complex. J Biol Chem 1986; 261: 1267-1273
• 7 Wun TC, Schleuning WD, Reich E. Isolation and characterization of urokinase from human plasma. J Biol Chem 1982; 257: 3276-3283
• 8 Collen D, Zamarron C, Lijnen HR, Hoylaerts M. Activation of plasminogen by pro-urokinase. II. Kinetics. J Biol Chem 1986; 261: 1259-1266
• 9 Lijnen HR, Zamarron C, Blaber M, Winkler ME, Collen D. Activation of plasminogen by pro-urokinase. I. Mechanism. J Biol Chem 1986; 261: 1253-1258
• 10 Lijnen HR, Van Hoef B, Collen D. Influence of cyanogen-bromide digested fibrinogen on the kinetics of plasminogen activation by urokinase. Eur J Biochem 1984; 144: 541-544
• 11 Matsuo O, Rijken DC, Collen D. Comparison of the relative fibrinogenolytic, fibrinolytic and thrombolytic properties of tissue plasminogen activator and urokinase in vitro. Thromb Haemostas 1981; 45: 225-229
• 12 Zamarron C, Lijnen HR, Van Hoef B, Collen D. Biological and thrombolytic properties of proenzyme and active forms of human urokinase. I. Fibrinolytic and fibrinogenolytic properties in human plasma in vitro of urokinases obtained from human urine or by recombinant DNA technology. Thromb Haemostas 1984; 52: 19-23
• 13 Collen D, Stassen JM, Stump DC, Verstraete M. In vivo synergism of thrombolytic agents. Circulation (in press)
• 14 Rijken DC, Collen D. Purification and characterization of the plasminogen activator secreted by human melanoma cells in culture. J Biol Chem 1981; 256: 7035-7041
• 15 Stump DC, Lijnen HR, Collen D. Purification and characterization of single-chain urokinase-type plasminogen activator (scu-PA) from human cell cultures. J Biol Chem 1986; 261: 1274-1278
• 16 Berenbaum MC. Synergy, additivism and antagonism in immunosuppression. A critical review. Clin Exp Immunol 1977; 28: 01-18
• 17 Gurewich V, Pannell R, Louie S, Kelley P, Suddith RL, Greenlee R. Effective and fibrin-specific clot lysis by a zymogen precursor form of urokinase (pro-UK). A study in vitro and in two animal species. J clin Invest 1984; 73: 1731-1739
• 18 Korninger C, Collen D. Neutralization of human extrinsic (tissue-type) plasminogen activator in human plasma: no evidence for a specific inhibitor. Thromb Haemostas 1981; 46: 662-665
• 19 Collen D, Stassen JM, Blaber M, Winkler M, Verstraete M. Biological and thrombolytic properties of proenzyme and active forms of urokinase. III. Thrombolytic properties of natural and recombinant urokinase in rabbits with experimental jugular vein thrombosis. Thromb Haemostas 1984; 52: 27-30
• 20 Colucci M, Paramo JA, Stassen JM, Collen D. Influence of the fast-acting inhibitor of plasminogen activator (PA-inhibitor) on in vivo thrombolysis induced by tissue-type plasminogen activator (t-PA). J Clin Invest 1986 (in press)
• 21 Collen D, Bounameaux H, De Cock F, Lijnen HR, Verstraete M. Analysis of coagulation and fibrinolysis during intravenous infusion of recombinant human tissue-type plasminogen activator (rt-PA) in patients with acute myocardial infarction. Circulation 1986; 73: 511-517
• 22 Van de Werf F, Nobuhara M, Collen D. Coronary thrombolysis with human single-chain urokinase-type plasminogen activator (scu-PA) in patients with acute myocardial infarction. Ann Int Med 1986; 104: 345-348