Effective lysis of model thrombi by a t-PA mutant (A473S) that is resistant to alpha2-antiplasmin.
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 alpha2-antiplasmin (alpha2-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 alpha2-AP. We examined these forms of t-PA in model systems that are physiologically relevant. Neutralization of alpha2-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 alpha2-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 alpha2-AP or PAI-1. A473S was more effective than wild-type t-PA without the addition of antibodies by virtue of its resistance to alpha2-AP. This resistance was remarkable, in that no complex formed between A473S t-PA and alpha2-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 alpha2-AP contribute approximately equally to the inhibition of thrombus lysis. This study underlines the functional significance of alpha2-AP as a direct inhibitor of t-PA and further explains the basis of the accepted role of alpha2-AP as a regulator of fibrin persistence and thrombus resistance to lysis.
Robbie
L A
LA
Departments of Molecular and Cell Biology and Medicine and Therapeutics, University of Aberdeen, Aberdeen, UK. l.a.robbie@abdn.ac.uk
Bennett
B
B
Keyt
B A
BA
Booth
N A
NA
D005342:Fibrinolysis / Q000187:drug effects*; D006801:Humans; D008954:Models, Biological; D009154:Mutation; D017395:Plasminogen Activator Inhibitor 1 / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D013927:Thrombosis / Q000503:physiopathology*; D010959:Tissue Plasminogen Activator / Q000235:genetics*; D000979:alpha-2-Antiplasmin / Q000494:pharmacology*
D017395:Plasminogen Activator Inhibitor 1; D011994:Recombinant Proteins; D000979:alpha-2-Antiplasmin; D010959:Tissue Plasminogen Activator