Human thrombi vary in their susceptibility to lysis and this is clinically important. Several potential contributory factors were examined in this study by using model thrombi, created under flow; these provide a robust, reproducible and easily-manipulated system. Here we identify the plasminogen activators (PA) active in model thrombi of known age and define the cellular and plasma contribution to activity in different areas. The cell-rich head of model thrombi had strong thrombin and PA activity, with coagulant activity also at the tail. Thrombin activity decreased as model thrombi were aged. PA activity in the thrombus head also decreased on ageing of thrombi but activity emerged around the thrombi, including the tail. Activity in the head of fresh model thrombi was primarily due to uPA, with some contribution from tPA. Experiments with thrombi prepared from platelet-rich plasma and added leucocytes showed that uPA activity at the head of fresh thrombi was derived from PMN. Older thrombi had tPA activity around the tail of the thrombus; this activity occurred in the absence of cells. This study highlights the importance of PMN-derived uPA activity in the lysis of fresh thrombi, with activity originating in the leucocyte-rich head. It also shows that thrombi are dynamic structures in which fibrin can be repeatedly laid down and lysed, observations that are relevant to therapeutic lysis and potential rethrombosis.
2
Mann KG,
Jenny RJ,
Krishnaswamy S.
Cofactor proteins in the assembly and expression of blood clotting enzyme complexes. Ann Rev Biochem 1988; 266: 915-20.
5
Hogg PJ,
Jackson CM.
Fibrin monomer protects from inactivation by heparin-antithrombin III: Implications for heparin efficacy. Proc Natl Acad Sci 1989; 86: 3619-23.
6
Weitz JI,
Hudoba M,
Massel D,
Maraganore J,
Hirsh J.
Clot-bound thrombin is protected from inhibition by heparin-antithrombin III but is susceptible to inactivation by antithrombin III-independent inhibitors. J Clin Invest 1990; 86: 385-91.
8
Francis CW,
Markham Jr RE,
Barlow GH,
Florack TM,
Dobrzynski DM,
Marder VJ.
Thrombin activity of fibrin thrombi and soluble plasmic derivatives. J Lab Clin Med 1983; 102: 220-30.
11
Carmeliet P,
Schoonjans L,
Kieckens L,
Ream B,
Degen JL,
Bronson R,
de Vos R,
van den Oord JJ,
Collen D,
Mulligan RC.
Physiological consequences of loss of plasminogen activator gene function in mice. Nature 1994; 368: 419-24.
14
Brommer EJP,
van Bockel JH.
Composition and susceptibility to thrombolysis of human arterial thrombi and the influence of their age. Blood Coag Fib 1992; 03: 717-25.
15
Potter BJvan Loon,
Rijken DC,
Brommer EJP,
van der Mass APC.
The amount of plasminogen activator inhibitor type 1 in human thrombi and the relation to ex-vivo lysibility. Thromb Haemost 1992; 67: 101-5.
16
van Giezen JJJ,
Nerme V,
Abrahamsson T.
PAI-1 inhibition enhances the lysis of the platelet-rich part of arterial-like thrombi formed in vitro. A comparative study using thrombi prepared from rat and human tissue. Blood Coag Fib 1998; 09: 11-8.
17
Stringer HAR,
van Swieten P,
Heijnen FG,
Sixma JJ,
Pannekoek H.
Plasminogen activator inhibitor-1 released from activated platelets plays a key role in thrombolysis resistance: studies with thrombi generated in the Chandler loop. Arterioscler Thromb 1994; 14: 1452-8.
18
Robbie LA,
Young SP,
Bennett B,
Booth NA.
Thrombi formed in a Chandler loop mimic human arterial thrombi in structure and PAI-1 content and distribution. Thromb Haemost 1997; 77: 510-5.
20
Robbie LA,
Robbie LA,
Keyt BA,
Bennett B,
Booth NA.
Effective lysis of model thrombi by a t-PA mutant (A473S) that is resistant to α2-antiplasmin. Br J Haematol 2000; 111: 517-23.
24
Northeast ADR,
Soo KS,
Bobrow LG,
Gaffney PJ,
Burnand KG.
The tissue-plasminogen activator and urokinase response in vivo during natural resolution of venous thrombi. J Vasc Surg 1995; 22: 573-9.
26
Yamamoto K,
Loskutoff DJ.
Fibrin deposition in tissues from endotoxintreated mice correlates with decreases in the expression of urokinase-type but not tissue-type plasminogen activator. J Clin Invest 1996; 97: 2440-51.
27
Ichinose A,
Fujikawa K,
Suyuma T.
The activation of pro-urokinase by plasma kallikrein and its inactivation by thrombin. J Biol Chem 1986; 261: 2486-9.
28
Gurewich V,
Pannell R.
Inactivation of single-chain urokinase (prourokinase) by thrombin and thrombin-like enzymes: relevance of the findings to the interpretation of fibrin-binding experiments. Blood 1987; 69: 769-72.
32
Plesner T,
Ploug M,
Ellis V,
Rønne E,
Høyer-Hansen G,
Wittrup M,
Pedersen TL,
Tscherning T,
Danø K,
Hansen NE.
The receptor for urokinase-type plasminogen activator and urokinase is translocated from two distinct intracellular compartments to the plasma membrane on stimulation of human neutrophils. Blood 1994; 83: 808-15.
33
Heiple JM,
Ossowski L.
Human neutrophil plasminogen activator is localized in specific granules and is translocated to the cell surface by exocytosis. J Exp Med 1986; 164: 826-40.
34
Higazi AAR,
Barghouti I,
Abu-Much R.
Identification of an inhibitor of tissue-type plasminogen activator-mediated fibrinolysis in human neutrophils. J Biol Chem 1995; 270: 9472-7.
36
Torr SRBrown,
Sobel BE.
Attenuation of thrombolysis by release of plasminogen-activator inhibitor type-1 from platelets. Thromb Res 1993; 72: 413-21.
37
van Giezen JJ,
Nerme V,
Abrahamson T.
PAI-1 inhibition enhances the lysis of the platelet-rich part of arterial-like thrombi formed in vitro. A comparative study using thrombi prepared from rat and human blood. Blood Coag Fibrinol 1998; 09: 11-8.
