The interplay between tissue plasminogen activator domains and fibrin structures in the regulation of fibrinolysis: kinetic and microscopic studies

Regulation of tissue-type plasminogen activator (tPA) depends on fibrin binding and fibrin structure. tPA structure/function relationships were investigated in fibrin formed by high or low thrombin concentrations to produce a fine mesh and small pores, or thick fibers and coarse structure, respectiv...

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Bibliographic Details
Published in:Blood 2011-01, Vol.117 (2), p.661-668
Main Authors: Longstaff, Colin, Thelwell, Craig, Williams, Stella C., Silva, Marta M.C.G., Szabó, László, Kolev, Krasimir
Format: Article
Language:English
Subjects:
Biological and medical sciences
Fibrin - metabolism
Fibrin - ultrastructure
Fibrinolysis - physiology
Hematologic and hematopoietic diseases
Humans
Kinetics
Medical sciences
Microscopy, Confocal
Microscopy, Electron, Scanning
Plasminogen - metabolism
Protein Binding
Thrombin - metabolism
Thrombosis and Hemostasis
Tissue Plasminogen Activator - metabolism
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Summary:Regulation of tissue-type plasminogen activator (tPA) depends on fibrin binding and fibrin structure. tPA structure/function relationships were investigated in fibrin formed by high or low thrombin concentrations to produce a fine mesh and small pores, or thick fibers and coarse structure, respectively. Kinetics studies were performed to investigate plasminogen activation and fibrinolysis in the 2 types of fibrin, using wild-type tPA (F-G-K1-K2-P, F and K2 binding), K1K1-tPA (F-G-K1-K1-P, F binding), and delF-tPA (G-K1-K2-P, K2 binding). There was a trend of enzyme potency of tPA > K1K1-tPA > delF-tPA, highlighting the importance of the finger domain in regulating activity, but the differences were less apparent in fine fibrin. Fine fibrin was a better surface for plasminogen activation but more resistant to lysis. Scanning electron and confocal microscopy using orange fluorescent fibrin with green fluorescent protein-labeled tPA variants showed that tPA was strongly associated with agglomerates in coarse but not in fine fibrin. In later lytic stages, delF-tPA-green fluorescent protein diffused more rapidly through fibrin in contrast to full-length tPA, highlighting the importance of finger domain-agglomerate interactions. Thus, the regulation of fibrinolysis depends on the starting nature of fibrin fibers and complex dynamic interaction between tPA and fibrin structures that vary over time.
ISSN:0006-4971
1528-0020
DOI:10.1182/blood-2010-06-290338