The plasma activity level of the recently discovered fast-acting inhibitor of tissue-type plasminogen activator (t-PA) was found to be temporarily increased after surgery, myocardial infarction and severe trauma. Detailed analysis of the postoperative period revealed simultaneously increased t-PA antigen and inhibition and decreased t-PA activity only on the first postoperative day. These changes were more rapid than those in fibrinogen and C-reactive protein. It is concluded that t-PA inhibition shows the most rapidly changing pattern observed so far in response to trauma. The postoperative fibrinolytic shutdown in blood fibrinolytic activity can be ascribed to a primary increase in t-PA inhibitor levels.

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We present the first steps in the elaboration of an approach of extracellular matrix-targeted local drug delivery (ECM-LDD), designed to provide a high concentration, ubiquitous distribution, and long-term retention of a drug within the vessel wall after local intravascular delivery. The approach is based on the concept of a bifunctional drug comprising a "therapeutic effector" and an "affinity vehicle," which should bind to an abundant component of the vessel wall. The aim of the present study was to select molecules suitable for the role of affinity vehicles for ECM-LDD and to study their intravascular delivery and retention ex vivo and in an animal model. By use of fluorescence microscopy, the following molecules were selected on the basis of strong binding to cross sections of human vessels: protamine, polylysine, polyarginine, a glycosaminoglycan-binding peptide from vitronectin, and a synthetic dendrimer. With polylysine as a prototypic affinity vehicle, we showed that after intravascular delivery, polylysine was concentrated throughout a luminal layer of the vascular wall to an extremely high concentration of 20 g/L and was retained therein for at least 72 hours of perfusion without noticeable losses. Low molecular weight (fluorescein) and high molecular weight (hirudin) compounds could be chemically conjugated to polylysine and were retained in the vessel wall after intravascular delivery of the conjugates. In conclusion, by use of the ECM-LDD method, an extremely high concentration and long-term retention of locally delivered drug can be reached. Polycationic molecules can be considered as potential affinity vehicles for ECM-LDD.

Summary. Background: The development of global tests for the fibrinolytic capacity in blood is hampered by the low base-line fibrinolytic activity in blood, by the involvement of both plasmatic components and blood cells in the fibrinolytic system and by the loss of fibrinolytic activity as a result of the action of plasminogen activator inhibitor-1 (PAI-1). Objective: To develop a new test for the global fibrinolytic capacity (GFC) of whole blood samples. Methods and results: Collection of blood in thrombin increased the subsequent generation of fibrin degradation products. This was ascribed to rapid clot formation and concomitant reduction of in vitro neutralization of tissue-type plasminogen activator (tPA) by PAI-1. On the basis of this observation, the following test was designed: blood samples were collected in thrombin with and without aprotinin and clots were incubated for 3 h at 37°C. The GFC was assessed from the difference between the fibrin degradation products in the two sera. The assay was applied to blood samples from patients and healthy subjects. Other hemostasis parameters were determined in plasma samples taken simultaneously. The GFC varied considerably (normal range 0.13-13.6 lg mL )1 ); physical exercise strongly increased the GFC. Statistically significant correlations were found with tPA activity, PAI-1 activity and fibrinogen level. A mixture of antibodies against tPA and urokinase-type plasminogen activator (uPA) completely inhibited the GFC. An inhibitor of activated thrombin-activatable fibrinolysis inhibitor (TAFI) accelerated fibrinolysis 8-fold. Conclusion: The new test represents a global assessment of the main fibrinolytic factors in plasma and potentially those associated with blood cells.

Alpha-2-antiplasmin, a major inhibitor of fibrinolysis, is synthesized in the liver and occurs in blood in two molecular forms: a very active plasminogen-binding (PB) form and a less active nonplasminogen-binding (NPB) form. This study investigates the origin and mutual relationship of these two forms in vivo and in vitro. Despite wide variation in plasma concentration of the inhibitor (16% to 138%), the ratio between the two forms in vivo was found to be, in the main, constant among healthy volunteers, heterozygotes for a congenital deficiency of alpha- 2-antiplasmin, and patients with a stable liver cirrhosis: PB/NPB = 2.41 +/- 0.34 (SD). Resynthesis after depletion or increased synthesis in the acute-phase reaction showed a specific increase of the PB form of the molecule in blood after discontinuation of L-asparaginase or streptokinase therapy and after myocardial infarction. In vitro studies demonstrated that only the PB form was present after one day in the culture medium of the human cell line Hep G2, while the NPB form appeared after 11 days. Clearance after inhibition of synthesis by L- asparaginase therapy revealed a more rapid decrease in the PB form relative to the NPB form in blood, demonstrated by a change in the PB- NPB ratio from 2.86 +/- 0.55 to 1.74 +/- 0.24 (mean of 6, SD). An apparently spontaneous first order conversion from the PB to NPB form, with an apparent half-life of about eight days, was demonstrated at 37 degrees C in plasma and serum in vitro. The conversion was found to be temperature dependent and uninfluenced by the fibrinolytic components fibrinogen, fibrin, and plasminogen. Additions of a variety of enzymes or inhibitors did not interfere with the process. These results demonstrate that the PB form of alpha-2-antiplasmin is produced by the liver and that the NPB form is formed in the circulation.