Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.68 (tissue plasminogen activator)
 11,311 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Slow and fast contracting muscles differ in their innervation and electrophysiological properties as well as in their regenerating potentialities. The purpose of the present work was to investigate the expression of plasminogen activators and its possible relation to each type of muscle. Slow (Soleus) and fast (Extensor Digitorum Longus) muscles were obtained from white Wistar rats. Before sectioning the muscles, the euthanized rats were perfused with cold phosphate buffer saline to avoid interference by circulating proteases and inhibitors. Muscle extracts were pounded in an ice-cold Potter tube. Plasminogen activators (PAs) were assayed by fibrin zymography and by both liquid and solid-phase fibrin spectrophotometric assays for the detection of PAs activity. Both urokinase (uPA) and tissue-type plasminogen activator (tPA) activities corresponding to proteins of 38 kDa and 65 kDa molecular masses, were detected in the extracts. Slow muscles contained higher amounts of both activators than fast muscles, but the relative amount of uPA was higher in both types of muscles. In addition, the characteristics of each type of extracts differed somewhat: the fast muscle activity curve was typical of an accelerating process, while the slow muscle curve showed an activity probably related to already formed plasmin or to some other trypsin-like enzyme. These results suggest that the amount of plasminogen activators could be a new criterion of discrimination between slow and fast skeletal muscles.
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PMID:Slow and fast rat skeletal muscles differ in their plasminogen activator activities. 211 33

Urokinase (UK, Mr 55,000) and tissue-type plasminogen activator (tPA, Mr 74,000) are serine proteinases involved in many biological processes, ie, cell migration, neoplastic transformation, and extracellular proteolysis. Cutaneous fibrinolytic activity (dependent on the activity of UK and tPA) was studied with the autohistographic fibrin film method in 40 patients affected by psoriasis vulgaris before and after topical (anthralin, betamethasone valerate, hydrocolloid occlusive dressing) or systemic psoralen-ultraviolet-light (PUVA) treatments. Autohistographic studies also were performed after apposition of monoclonal antibodies directed against the catalytic site of UK and tPA. Finally, UK and tPA were localized immunohistochemically in the psoriatic plaques and in controls using the immunoperoxidase procedure based on the biotin/avidin system. UK and tPA immunoreactivity was present in the cytoplasm and around the outlines of keratinocytes in the psoriatic patches before treatment and in the patches not cleared after treatment, while it was not detectable in normal epidermis, in the unaffected psoriatic epidermis, and in the cleared psoriatic skin. Cutaneous fibrinolytic activity was present in the cases in which UK and tPA were detected histochemically and, in the psoriatic epidermis, it was abolished by preincubation with anti-tPA but not with anti-UK antibodies. This study suggests that established topical and systemic treatments for psoriasis possess UK and tPA antagonist activity.
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PMID:Antipsoriatic therapies inhibit epidermal plasminogen activator activity. 212 55

It has previously been reported that EGF enhances uPA but not tPA in the A431 squamous carcinoma cell line. To determine whether the absence of tPA modulation by EGF reflected steady levels or the action of an anti-activator, we assayed tPA, PAI-1 and tPA/PAI-1 complexes by zymography and immunological assays. Under conditions in which EGF had no effect on tPA activity, tPA antigen paradoxically increased with a concomitant rise of tPA/PAI-1 complexes. This indicated that tPA was rapidly inactivated through the formation of a complex, immunologically and electrophoretically related to tPA/PAI-1. tPA antigen and tPA/PAI-1 complexes were modulated by EGF in a time and concentration dependent manner. PAI-1 antigen was secreted into A431 medium (CM) after a lag phase of 16 h in both control and EGF-treated cultures. Evidence is presented here that two forms of PAI-1 are present in A431 CM: an inactive form and an active form which neutralizes the tPA secreted, masking its enhancement by EGF in functional assays.
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PMID:Concomitant secretion by A431 cells of tissue plasminogen activator and a specific inhibitor masks EGF modulation of tPA activity. 212 70

The reactivity of two D-dimer assays (a latex agglutination method, D-Di test and an ELISA procedure, Asserachrom D-Di) to the various fibrin or fibrinogen degradation products generated in plasma by three different thrombolytic agents was analysed, in the presence or absence of a fibrin clot. Other assays performed in parallel were an ELISA assay for (DD)E complexes and the conventional fibrinogen degradation products (FDP) latex test on serum. The thrombolytic agents urokinase, streptokinase or tPA were added at various concentrations and incubated for different times ranging from 10 min to 24 h. The data showed that the D-dimer latex assay was always negative provided there was no fibrin in plasma and despite the presence of high FDP levels (greater than 600 micrograms/ml) in serum. In contrast, D-dimer or (DD)E complexes were measured by ELISA, but up to a given concentration (15-20 micrograms/ml) which reached a plateau and remained stable irrespective of the thrombolytic concentrations or the degradation times. In the presence of fibrin clot, fibrinolysis was extremely fast with tPA and the FDP were generated at a much higher concentration that that expected from the size of the fibrin clot. This suggests the existence of fibrinogenolysis targeted by the presence of fibrin but negative in its absence. Urokinase and streptokinase generated FDP very quickly but a much slower degradation rate of fibrin was observed. The immunoblotting confirmed these data and showed that no late FDP were formed in plasma even at high thrombolytic concentrations except when fibrin was present.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Reactivity of D-dimer assays with the fibrinogen--fibrin split products generated by thrombolytic agents. 213 30

It has been reported that EGF treatment enhances uPA but not tPA in the A431 epidermoid carcinoma cell line. To determine whether the absence of tPA modulation by EGF could be due to the action of inhibitors, we assayed tPA, PAI-1, PAI-2 and tPA/PAI-1 complexes by immunological assays and zymography in A431 serum-free medium. We found that, under conditions in which EGF had no effect on tPA activity, tPA antigen increased with a concomitant rise of tPA/PAI-1 complexes, indicating the action of an inhibitor. Both tPA antigen and tPA/PAI-1 complexes were modulated by EGF in a time and concentration dependent manner. tPA/PAI-1 complex levels were lower than tPA levels, suggesting the presence of other inhibitors. Immunological assays detected PAI-2 in addition to PAI-1 and showed a time and dose response to EGF. Modulation of tPA and the anti-activators by the growth factor was confirmed by identification of the corresponding transcripts with cDNA probes. We conclude that the net plasminogen activator activity in A431 cells is the result of a balance between activators and inhibitors.
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PMID:Modulation of tPA, PAI-1 and PAI-2 antigen and mRNA levels by EGF in the A431 cell line. 213 49

Thrombospondin (TSP), an adhesive glycoprotein found in platelets and extracellular matrix, has been shown previously to interact with plasminogen and tissue plasminogen activator, resulting in efficient plasmin generation. We now demonstrate specific complex formation of TSP with both the single-chain and two-chain forms of urokinase (scuPA and uPA). Binding of uPA and scuPA to immobilized TSP was detected and quantified using colorimetric immunoassays and a functional amidolytic assay. Binding was time and concentration dependent with apparent affinity constants of 40-50 nM. Binding was not affected by serine protease inhibitors, EDTA, or epsilon-aminocaproic acid. scUPA and uPA bound to TSP retained functional activity. Using a sensitive amidolytic assay we found that TSP. scuPA complexes were efficiently converted to TSP. uPA by catalytic plasmin concentrations. Additionally, TSP.uPA complexes were found to have plasminogen-activating activity equivalent to fluid-phase uPA and to be protected from inhibition by plasminogen activator inhibitor type 1, the major plasma and matrix plasminogen activator inhibitor. Using immunohistochemical techniques, we also demonstrated co-distribution of TSP and uPA in normal and malignant breast tissue. Complex formation of TSP with uPA may serve to localize, concentrate, and protect these enzymes on cell surfaces and within the extracellular matrix, thereby providing a reservoir of plasminogen activator activity.
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PMID:Thrombospondin forms complexes with single-chain and two-chain forms of urokinase. 214 8

Plasminogen activators (PAs), were estimated qualitatively and quantitatively in two different clonal murine skeletal muscle cell lines. Both cell lines produced the two major types of PAs found in mammalian cells, urokinase-type (uPA) and tissue type (tPA). These two lines are models for the study of myogenesis in vitro, but differ in several growth and differentiation characteristics. Because of their possible involvement in these characteristics we assayed the expression of PAs in both cell systems during development in culture. Utilizing fibrin zymography two isoforms of tPA were detected. One co-migrated with human tPA at 75 kd and another may represent a tPA:inhibitor complex at 105 Kd. Several isoenzymes of uPA were detected and these changed depending on whether cell homogenates or conditioned medium was analyzed and whether myogenic cells were at single-cell myoblast or multi-nucleated myotube stage. Species-specific antisera to mouse uPA identified 4 uPA bands in muscle cell medium and 5 in cell layers. Antigenic uPA bands also varied depending on stage of myogenesis. Quantitative amidolytic studies using chromogenic substrates showed that maximal PA activity, both uPA and tPA, occurred at the time of myoblast fusion. Furthermore, uPA activity in membranes increased during myogenesis, while both uPA and tPA in medium decreased after fusion. These studies indicate that muscle PA expression is developmentally regulated and may correlate with growth and differentiation in skeletal muscle.
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PMID:Plasminogen activators and their inhibitors in the neuromuscular system: I. Developmental regulation of plasminogen activator isoforms during in vitro myogenesis in two cell lines. 219 66

Human non-small lung cancer cell lines HS-24 (established from a primary squamous cell carcinoma) and SB-3 (established from a metastasis of a primary adenocarcinoma of the lung into the adrenal gland) were analysed for the proteinases tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator inhibitor (PAI-1). The proteinases were characterized by activity measurements, inhibition studies, enzyme-linked immunosorbent assay (ELISA), and Western blot analysis. Cell-associated proteinases were determined in cell lysates, secreted proteinases in cell conditioned culture media. Both cell lines were found to secrete uPA and PAI-1, whereas tPA could be detected only in HS-24 conditioned media. No cathepsin B activity could be detected in media of both cell lines. However, activation experiments and western blot analysis showed, that at least HS-24 secrete an inactive precursor. Cell lysates of HS-24 and SB-3 show PA activity, but on a low level. Cathepsin B activity was also found to be low in HS-24 lysates. However, SB-3 lysates show high cathepsin B activity. Further characterization of the proteinases by their sensitivity against several inhibitors suggests that they are similar to the corresponding proteinases of normal, nonmalignant cells.
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PMID:Detection of cathepsin B, plasminogen activators and plasminogen activator inhibitor in human non-small lung cancer cell lines. 222 60

Increasing evidence suggests the involvement of leukocytes in the fibrinolytic system. Monocytes secrete pro-urokinase (Grau, Thromb Res 1989; 53: 145) and it has been shown that these cells have specific receptors for urokinase and plasminogen (Miles, Thromb Haemostas 1987; 58: 936). The aim of this study was to analyse the presence of plasminogen activator inhibitor(s) in platelet-free suspensions of human peripheral blood monocytes and polymorphonuclear leukocytes (PMN). SDS-PAGE and reverse fibrin autography showed an inhibitory band of 50 kDa in the monocyte extracts (Triton X-100) but not in the PMN extracts. Urokinase (u-PA) was mixed with increasing amounts of monocyte extract for 10 min and the mixtures were added to 125I-fibrin coated wells containing plasminogen. A dose-dependent decrease in the u-PA fibrinolytic activity was observed. The amount of inhibition increased when the monocyte releasates were preincubated with u-PA (40% inhibition after 5 min preincubation and 80% after 15 min), indicating a direct interaction between this activator and an inhibitor(s). After SDS-PAGE of monocyte extracts, immunoblotting and peroxidase staining identified both PAI1 and PAI2, with an apparent molecular weight of 47-50 kDa. Monocyte-associated PAI1 formed complexes with single chain t-PA with a molecular mass 50 kDa higher than the molecular mass of the free PAI1. However, a significant amount of PAI1 remained unbound to t-PA. This inactive PAI1 could have come from a rapid inactivation of the primary active PAI1. These PAI1 and PAI2 detected in human monocytes may be transcendent in the regulation of the fibrinolytic system.
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PMID:Detection of both type 1 and type 2 plasminogen activator inhibitors in human monocytes. 233 62

The functional role of the fibrinolytic system in capillary growth was investigated using bovine capillary endothelial cells (BCEs) cultured on a Type I collagen gel matrix, into which the cells migrated to form capillary-like tubular structures. The length of the tubes formed were measured morphometrically using an image analyzer in the absence and presence of fibrinolytic proteases, namely plasminogen, plasminogen activators (PAs) and PA inhibitor (PAI). The addition of plasminogen (25 micrograms/ml) to the gel matrix significantly increased the length of BCE tubes found on the 9th day of culture (p less than 0.01), with a dose-dependent tendency. The simultaneous addition of a basic fibroblast growth factor (bFGF, 10 ng/ml) enhanced this tube formation as early as the 3rd day of culture (p less than 0.01). Cultured BCEs secreted both tissue-type and urokinase-type PAs (tPA and uPA) and PAI-1 into the culture medium, and the secretion of both PAs was enhanced by the addition of bFGF. However, the secreted tPA was composed mostly of an inactive form of tPA.PAI-1 complex, and the PA activity was derived mostly from uPA. Inhibitors of plasmin suppressed the enhancing effect of plasminogen on angiogenesis. In addition, anti-uPA IgG markedly inhibited the enhancing effect of plasminogen on the 4th and 7th days of culture (p less than 0.01), whereas anti-tPA IgG showed an inhibitory tendency only on the 4th day of culture (p less than 0.05). These findings indicate that the plasminogen-PAs system, especially uPA synthesized and secreted by BCEs, plays an important role in regulating angiogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A role of fibrinolytic activity in angiogenesis. Quantitative assay using in vitro method. 248 Nov 53


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