Urokinase plasminogen activator (uPA) and tissue plasminogen activator (tPA) were assayed in biopsies from the base, edge and adjacent skin of ischaemic and venous ulcers using a functional bioimmunoassay and a standard immunoassay. Two series of 14 biopsies were examined: from seven venous and seven ischaemic ulcers in the first series, eight venous and six ischaemic in the second. It was found that tPA was detected in only four samples of ulcer edge using the bioimmunoassay and in no sample by the standard immunoassay. By contrast, uPA was detected in all but one ulcer edge biopsy and at a significantly lower median concentration in the adjacent skin (16.9 units/g) than the ulcer edge (26.1 units/g) or base (32.3 units/g) (both P < 0.01). Levels of uPA were greater in the edge and base of venous compared with ischaemic ulcers. The predominant plasminogen activator in chronic leg ulcers is uPA; this activator may play an important role in wound healing.
Using two-dimensional immunoelectrophoresis and an antibody to 012-antiplasmin, we assessed the plasmin generated in serum under different conditions as the plasmin-a2-antiplasmin complex. Activation in serum of human [Glu'lplasminogen ([Glu']Pg) by recombinant tissue plasminogen activator was inhibited by the normal serum levels of Cl-and was enhanced by physiological levels of fibrinogen in the presence or absence of Cl-. These results agree with the Tissue plasminogen activator (TPA) is regarded the major activator of plasminogen in human plasma and as such is an important defense via the fibrinolytic mechanism against fibrin deposition in vivo. A hypothetical mechanism by which TPA on the surface of fibrin causes dissolution of both preformed fibrin and forming fibrin has been proposed (1). This hypothesis, while explaining some phenomena seen during the classic hypercoagulable state known as disseminated intravascular coagulation, may not encompass all molecular events occurring during thrombotic episodes and use of TPA as a thrombolytic agent. Roles for Cl -and fibrinogen have been implicated in human plasminogen ([Glu1]Pg) activation in purified systems using streptokinase (2, 3), urokinase (4, 5), and TPA (6) as activators. These data show that Cl-, which exists in plasma at 90 mM, could have a significant inhibitory effect on [Glu']Pg activation and that the influence of fibrinogen on the initial rate of [Glu']Pg activation could be physiologically important during enhanced fibrinolysis in vivo. Although both effects have been shown in purified systems (2-4), this study was undertaken to assess the relevance of these effects on activation of [Glu']Pg to plasmin in its natural environment, plasma. The data suggest that anion and fibrinogen effects both need consideration when TPA is used as a thrombolytic agent; the influence of these effects on the degradation of circulating fibrinogen and the thrombus-specificity of TPA therapy must be weighed.
A previously described bioimmunoassay for tissue-type plasminogen activator (t-PA) has been modified in terms of antibody concentration and conditions of incubation to achieve a sensitivity range of approximately 5-500 IU/L of t-PA. A similar assay has been developed for urinary-type plasminogen activator (U-PA), also known as urokinase (UK), achieving a sensitivity range of approximately 5-500 X 10(-1) IU/L. The direct sensitive t-PA assay has been demonstrated to be a quantitative alternative to the traditional semiquantitative euglobulin clot lysis time (ECLT) assay, with correlation coefficients of 0.967 and 0.914, depending on the laboratory where the ECLT was carried out. This assay has the added advantages of avoiding loss of t-PA or UK during the formation of the euglobulin fraction and of calibration in terms of International Standards for both t-PA and UK. With both these assays, it has been demonstrated that the enhanced fibrinolytic activity observed after exercise results from increased levels of t-PA while the level of UK in plasma remains unaltered. Free t-PA (10-60 IU/L) has been demonstrated in resting plasma, whereas the level of free UK activity was 50-100 IU/L. Venous occlusion of matched groups of subjects with and without deep venous thrombosis (DVT) showed a wide variation of response in t-PA levels in each group and no change in UK levels. There was no difference between the DVT and non-DVT groups, and this suggests that the notion of the "nonresponder" being more prone to thrombosis needs to be reexamined.
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