Comparative Study of the Activity of High and Low Molecular Weight Urokinase in the Presence of Fibrin

Summary

The fibrinolytic or thrombolytic activity of low molecular weight urokinase (LMW-UK) and high molecular weight urokinase (HMW-UK) is not significantly different when measured in a bovine fibrin plate method, in a circulating plasma system containing a ¹²⁵I-labelled human fibrin clot, or on ¹²⁵I-fibrin films in culture plates using normal or α₂-antiplasmin depleted human plasma.

In a human fibrin plate method however HMW-UK was found to be more active than LMW-UK. In a purified system on human ¹²⁵I-fibrin films the activation of native or modified human plasminogen by HMW-UK was also found to be more effective than by LMW-UK.

Using a clot lysis test system we did not observe a different inhibition of LMW-UK and HMW-UK upon incubation in human plasma. This is in contrast with previous reports that HMW-UK is inhibited more rapidly in human plasma than LMW-UK.

In a purified system the inhibition rate of LMW-UK and HMW-UK by α₂-antiplasmin is the same (rate constants at 25ΰC of 167 ± 9 M⁻¹s⁻¹ and 171 ± 5 M⁻¹s⁻¹ respectively).

The clinical trials available at present used doses of urokinase which were in excess of those required to obtain a maximal fibrinolytic effect. This might explain why in these trials no difference was observed between the thrombolytic effect of LMW-UK and HMW-UK, while in vitro HMW-UK appeared to be more effective. However, one should always be careful to extrapolate in vitro observations as such to the in vivo situation encountered during thrombolytic therapy.

Key words

High and low molecular weight urokinase - Comparison of the fibrinolytic activity - Inhibition in human plasma - Inhibition by α₂-antiplasmin

• References

• 1 Lormeau JC, Goulay J, Vairel EG, Choay J. A comment on the activities of high and low molecular weight urokinase. In: Fibrinolysis. Current Fundamental and Clinical Concepts. Gaffney PJ, Balkuv-Ulutin S. (Eds). 1978: 77-82 Academic Press; London:
• 2 Samama M, Cazenave B, Otero AM. Urokinase I and II activity. Thromb Haemostas 1978; 40: 578-580
• 3 Urokinase Pulmonary Embolism Trial. A national cooperative study. American Heart Association Monograph No 39. Circulation 1973; 47: 1-108
• 4 Urokinase-Streptokinase Pulmonary Embolism Trial. Phase 2, Results. A cooperative study. J Am Med Assoc 1974; 229: 1606-1613
• 5 Marder VJ, Donahoe JF, Bell WR, Cranley JJ, Kwaan HC, Sasahara AA, Barlow GH. Changes in the plasma fibrinolytic system during urokinase therapy: comparison of tissue culture urokinase with urinary source urokinase in patients with pulmonary embolism. J Lab Clin Med 1978; 92: 721-729
• 6 Murano G, Aronson DL. High and low molecular weight urokinase. Thromb Haemostas 1979; 42: 1066-1068
• 7 Murano G, Aronson D, Williams L, Brown L. The inhibition of high and low molecular weight urokinase in plasma. Blood 1980; 55: 430-436
• 8 Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography. Science 1970; 170: 1095-1096
• 9 Claeys H, Vermylen J. Physicochemical and proenzyme properties of NH₂-terminal glutamic acid and NH₂-terminal lysine human plasminogen. Influence of 6-aminohexanoic acid. Biochim Biophys Acta 1974; 342: 351-359
• 10 Blombäck B, Blombäck M. Purification of human and bovine fibrinogen. Ark Kemi 1956; 10: 415-443
• 11 Wiman B. Affinity-chromatographic purification of human α₂-antiplasmin. Biochem J 1980; 191: 229-232
• 12 Fenton II JW, Fasco MJ, Stackrow AB, Aronson DL, Young AM, Finlayson JS. Human thrombins. Production, evaluation, and properties of α-thrombin. J Biol Chem 1977; 252: 3587-3598
• 13 Wiman B, Collen D. On the kinetics of the reaction between human antiplasmin and plasmin. Eur J Biochem 1978; 84: 573-578
• 14 Edy J, De Cock F, Collen D. Inhibition of plasmin by normal and antiplasmin depleted human plasma. Thrombos Res 1976; 8: 513-518
• 15 Astrup T, Müllertz S. The fibrin plate method for estimating fibrinolytic activity. Arch Biochem Biophys 1952; 40: 346-351
• 16 Hoylaerts M, Lijnen HR, Collen D. Studies on the mechanism of the antifibrinolytic action of tranexamic acid. Biochim Biophys Acta 1981; 673: 75-85
• 17 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
• 18 Vermylen C, De Vreker R, Verstraete M. A rapid enzymatic method for assay of fibrinogen: the fibrin polymerization time (FPT-test). Clin Chim Acta 1963; 8: 418-424
• 19 Friberger P, Knös M. Plasminogen determination in human plasma. In: Chromogenic Peptide Substrates. Scully MF, Kakkar VV. (Eds). 128-140 Churchill Livingstone; Edinburgh: 1979
• 20 Suyama T, Nishida M, Iga Y, Naito R. Difference in thrombolytic effect between higher and lower molecular weight forms of urokinase. Thromb Haemostas 1977; 38: 48 (Abstr)
• 21 Philo RD, Gaffney PJ. Assay methodology for urokinase: its use in assessing the composition of mixtures of high- and low-molecular weight urokinase. Thromb Res 1981; 21: 81-88
• 22 Nobuhara M, Sakamaki M, Ohnishi H, Suzuki Y. A comparative study of high molecular weight urokinase and low molecular weight urokinase. J Biochem 1981; 90: 225-232
• 23 Ogston D, Bennett B, Herbert RJ, Douglas AS. The inhibition of urokinase by α₂-macroglobulin. Clin Sci 1973; 44: 73-79
• 24 Clemmensen I, Christensen F. Inhibition of urokinase by complex formation with human alpha-1-antitrypsin. Biochim Biophys Acta 1976; 429: 591-599
• 25 Clemmensen I. Inhibition of urokinase by complex formation with human antithrombin III in absence and presence of heparin. Thromb Haemostas 1978; 39: 616-623
• 26 Moroi M, Aoki N. Isolation and characterization of α₂-plasmin inhibitor from human plasma. A novel proteinase inhibitor which inhibits activator-induced clot lysis. J Biol Chem 1976; 251: 5956-5965
• 27 Matsuo O, Kosugi T, Mihara H. Urokinase inactivation rate in the rabbit: effect of circulatory isolation of the liver, spleen and kidneys. Haemostasis 1978; 7: 367-372
• 28 Barlow GH, Marder VJ. Determination of plasma urokinase levels by chromogenic assay: comparison of levels attained during infusion of tissue culture and urinary source urokinase preparations. Thromb Haemostas 1979; 42: 295 (Abstr)
• 29 Barlow GH, Francis CW, Marder VJ. On the conversion of high molecular weight urokinase to the low molecular weight form. Thromb Haemostas 1981; 46: 11 (Abstr)