Effective lysis of model thrombi by a t-PA mutant (A473S) that is resistant to alpha(2)-antiplasmin

L A Robbie, B Bennett, B A Keyt, N A Booth

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Abstract

This study used two mutants of tissue-type plasminogen activator (t-PA) with resistance to inhibitors of fibrinolysis to define the contribution of plasminogen activator inhibitor (PAI)-1 and alpha (2)-antiplasmin (alpha (2)-AP) to the control of fibrin lysis. Wild-type t-PA was compared with KHRR296-299AAAA, which is resistant to PAI-1, and with A473S, which is resistant to alpha (2)-AP. We examined these forms of t-PA in model systems that are physiologically relevant. Neutralization of alpha (2)-AP was essential for lysis of plasma clots, irrespective of their platelet content, by either wild-type t-PA or KHRR296-299AAAA. In marked contrast, A473S lysed plasma clots without neutralization of alpha (2)-AP. Model thrombi, with structures similar to in vivo thrombi, were lysed slowly by wild-type t-PA; the rate and extent of lysis were enhanced by the addition of antibodies to alpha (2)-AP or PAI-1. A473S was more effective than wild-type t-PA without the addition of antibodies by virtue of its resistance to alpha (2)-AP. This resistance was remarkable, in that no complex formed between A473S t-PA and alpha (2)-AP, even after extended incubation, when 50% of wild-type t-PA could be converted to complex. Comparison of A473S and KHRR296-299AAAA mutants showed their similar effectiveness in lysis of platelet-rich model thrombi. Thus, PAI-1 and alpha (2)-AP contribute approximately equally to the inhibition of thrombus lysis. This study underlines the functional significance of alpha (2)-AP as a direct inhibitor of t-PA and further explains the basis of the accepted role of alpha (2)-AP as a regulator of fibrin persistence and thrombus resistance to lysis.

Original languageEnglish
Pages (from-to)517-523
Number of pages7
JournalBritish Journal of Haematology
Volume111
Publication statusPublished - 2000

Keywords

  • alpha(2)-antiplasmin
  • plasminogen activator inhibitor-1
  • tissue-type plasminogen activator
  • fibrin lysis
  • plasmin
  • PLASMINOGEN-ACTIVATOR INHIBITOR-1
  • HUMAN-PLASMA
  • ALPHA-2-PLASMIN INHIBITOR
  • CROSS-LINKING
  • CLOT LYSIS
  • PROTEINASE-INHIBITORS
  • CHANDLER LOOP
  • FIBRINOLYSIS
  • PURIFICATION
  • DEFICIENCY

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