The activation of human plasminogen (P) by two-chain tissue plasminogen activator (A) was studied in the presence of fibrin films (F) of increasing size and surface density. Initial rates of plasminogen activation (v) were determined as a function both of the plasminogen and fibrin concentration. The activation rate was strongly dependent on the presence of fibrin and plots of 1/v versus 1/ [p] or 1 /[F] yielded straight lines. The kinetic data were in agreement with the following reaction scheme.According to this model tissue plasminogen activator would bind to fibrin with a dissociation constant (KF of 0.2 µM and this complex fixes plasminogen with a Michaelis constant (Kp’) of 0.15 µM (Glu-plasminogen) or 0.02 µM (Lys-plasminogen) to form a ternary complex, converted to plasmin with a catalytic rate constant kcat = 0.05 s-1. This mechanism implies that both plasminogen and tissue plasminogen activator are concentrated on the fibrin surface through formation of a fibrin bridge. Activation of plasminogen in the absence of fibrin occurs with Km = 65 µM (Glu-plasminogen) or Km= 19 µM (Lys-plasminogen) and kcat = 0.05 s-1. Our data suggest that fibrin enhances the activation rate of plasminogen by tissue plasminogen activator by increasing the affinity of plasminogen for fibrin-bound tissue plasminogen activator and not by influencing the catalytic efficiency of the enzµMe. These data also support the hypothesis that fibrinolysis is both triggered by and directed towards fibrin.Generated plasmin was quantitated by measuring the rate of solubilization of 125I-labeled fibrin.

The potential role of the matrix metalloproteinase (MMP) system in the pathophysiology of the adipose tissue was investigated in a mouse model of nutritionally induced obesity. mRNA levels of 16 MMPs and 4 tissue inhibitors of MMPs (TIMPs) were measured by semiquantitative RT-PCR in adipose tissue isolated from mice maintained for 15 weeks on a standard or high-fat diet. In mice on standard diet, with the exception of MMP-8, all MMP and TIMP transcripts were detected in both gonadal and subcutaneous depots. In obese mice, the expression of MMP-3, -11, -12, -13, and -14 and TIMP-1 mRNAs was upregulated, whereas that of MMP-7, -9, -16, and -24 and TIMP-4 was downregulated. Most MMP and TIMP mRNAs were expressed at higher levels in stromal-vascular cells than in mature adipocytes. Analysis of adipose tissue by in situ fluorescent zymography revealed MMP-dependent proteolytic activities, demonstrating the presence of active MMPs in the intact tissue. In vitro conversion of adipogenic 3T3-F442A cells into mature adipocytes was associated with substantial modulations of MMP and TIMP expression. Moreover, this in vitro adipogenesis was reduced in the presence of a synthetic MMP inhibitor. Thus, the adipose tissue expresses a large array of MMPs and TIMPs, which modulate adipocyte differentiation.

LOOD CONTAINS an enzymatic system called the B fibrinolytic system, one of the main functions of which is the dissolution of fibrin clots in the blood vessels. The fibrinolytic system comprises a proenzyme, plasminogen, which can be converted to the active enzyme plasmin by different plasminogen activators (PAS). Two physiologic PAS have been identified, initially based on their immunologic relationship with the PA found in tissues (tissue-type PA [t-PA]) or with the PA found in urine (urokinase-type PA [u-PA]). Inhibition of fibrinolysis occurs at the level of the activators (by PA-inhibitors [PAIS]) or at the level of plasmin (mainly by a,-antiplasmin). Physiologic fibrinolysis is highly fibrin-specific as a result of specific molecular interactions between PA, plasminogen, fibrin, plasmin, and a,-antiplasmin.' Cardiovascular diseases, mainly comprising coronary artery disease leading to myocardial infarction, cerebrovascular disease causing strokes, and venous thrombosis predisposing to pulmonary embolism and the post-phlebitic syndrome, are a major cause of death and disability. The triggering event in the clinical expression of the acute ischemic event is not the underlying atherosclerotic lesion, but a thrombotic obstruction of the artery. Thus, the common cardiovascular diseases have, as their immediate underlying etiology, thrombosis of critically situated blood vessels with loss of blood flow to vital organs. One approach to the treatment of thrombosis consists of the pharmacologic dissolution of the blood clot via the intravenous infusion of PAS. Currently, five thrombolytic agents are either approved for clinical use or under clinical investigation in patients with acute myocardial infarction. These agents are streptokinase (SK), two-chain u-PA (tcu-PA; urokinase), anisoylated plasminogen streptokinase activator complex (APSAC), recombinant t-PA (rt-PA), and recombinant single-chain u-PA (rscu-PA; prourokinase).] Reduction of infarct size, preservation of ventricular function, and reduction in mortality has been obtained with SK, rt-PA, and APSAC.' Consequently, thrombolytic therapy has become standard treatment for early acute myocardial infarction. Intravenous SK recanalizes approximately 50% of occluded coronary arteries within 90 minutes and reduces mortality by 25%. rt-PA in combination with adjunctive intravenous heparin is more potent for coronary arterial thrombolysis, producing both more rapid and more frequent recanalization (patency 70% to 75% within 90 minutes). Side effects (mainly bleeding) and the incidence

Obesity is a common disorder and related diseases, such as diabetes, atherosclerosis, hypertension, cardiovascular disease and cancer, are a major cause of mortality and morbidity in Western-type societies. Development of obesity is associated with substantial modulation of adipose tissue structure.

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