© 1995 by European Society of Cardiology
Copyright © 1995, European Society of Cardiology
In vitro oxidative damage to tissue-type plasminogen activator: a selective modification of the biological functions
Department of Cardiovascular Medicine, John Radcliffe Hospital, University of Oxford, Oxford OX3 9DU, UK
* Corresponding author, present address: Department of Molecular Cardiology (FF3), The Cleveland Clinic Foundation, 9500 Euclid Ave., Cleveland, OH 44195, USA. Tel. (216) 444 9331; Fax (216) 444 9263.
Objective: Tissue-type plasminogen activator (t-PA) is an important component of the blood fibrinolytic system responsible for thrombus dissolution. It is often required to function under oxidative stress, and exogenous t-PA is used clinically in the treatment of acute myocardial infarction (AMI). The aim of this study was to examine alterations in the residual activity of t-PA pre-treated with certain oxidants. Methods: Recombinant t-PA (rt-PA) and native t-PA (nt-PA) were pre-treated with freshly generated hypochlorous acid (HOCl) and chloramine T at varying concentrations. The amidolytic activity, the plasminogenolytic activity and the fibrin-binding affinity were then examined using chromogenic assays based on S-2288 and S-2251. Results: The amidolytic activity of t-PA was surprisingly found to be rather sensitive (IC50 1 and 12 µmol/l, respectively), and the plasminogenolytic activity rather resistant to pre-treatment with HOCl and chloramine T. The fibrin binding study of treated t-PA revealed substantial loss of binding to CNBr-digested fibrinogen (FDP-CNBr). The velocity of t-PA in reaction with plasminogen remained the same as non-treated t-PA. The possible mechanisms of this asymmetrical oxidative modification of the biological functions are also discussed. Conclusions: (1) The catalytic activity of t-PA and the binding affinity for its large-molecule substrate plasminogen, rather than the small-molecule substrate S-2288, are highly resistant to oxidative damage; (2) the fibrin-binding affinity of t-PA can be selectively and asymmetrically damaged by exposure to these oxidants. Thus it is possible that the characteristic advantage of thrombus selectivity of t-PA in both spontaneous thrombolysis and thrombolytic therapy may be diluted in circumstances where toxic and reactive oxidants exist.
KEYWORDS t-PA; Fibrin binding; Plasminogen; Thrombolysis
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