Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
 10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

It is well known that cultured Sertoli cells secrete plasminogen activators (Lacroix et al., Mol Cell Endocrinol 1977; 9:227-236; Hettle et al., Biol Reprod 1986; 34:895-904). We now show that testicular cells in culture also secrete gelatinolytic metalloproteinases. Gelatin zymographic analysis of concentrated culture medium proteins reveals that Sertoli cells secrete gelatinases of 185 kDa, 110 kDa, 83 kDa, 76 kDa, and 72 kDa in addition to plasminogen activators (PAs). Gelatinase 185 kDa is induced by FSH. Media from Sertoli (epithelial)/peritubular (mesenchymal) cell cocultures contain the Sertoli cell gelatinases and one FSH-stimulated gelatinase of 50 kDa, indicating that gelatinase 50 kDa is regulated by both FSH and cell-cell interactions. A 50-kDa fibronectinolytic activity is also present in the coculture medium from cells grown in the presence of FSH. Casein zymography demonstrates a prominent 30-kDa protease only in media from cocultures. Peritubular cells secrete urokinase-type plasminogen activator (u-PA) and exhibit slight degrading activity at 86 kDa and 74 kDa. The gelatinases are most active in the pH range 7.3-8.5 and are completely or partially inhibited by metal ion chelators indicating that they are metalloproteinases. Our data demonstrate that testicular cells in culture secrete several gelatinases in addition to PAs, and that FSH and coculture conditions regulate some of these secreted proteases. We suggest that the highly regulated secretion of these proteases may well be of physiological importance during testicular development and spermatogenesis.
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PMID:Secreted metalloproteinases in testicular cell culture. 212 32

The presence of urokinase-type plasminogen activator (uPA) in human eccrine sweat has not been reported previously. Clean sweat was obtained from the upper trunk and arms of subjects which had been painted with white petrolatum to minimize epidermal contamination. Sweat was concentrated x 50 by ultrafiltration, and its PA activity determined by the two-step assay method (conversion of plasminogen to plasmin with the subsequent assay of plasmin activity using the substrate S-2251). PA activity was detectable in nine of 17 subjects by this method, which probably represents an underestimate of the true activity because of possible loss of the enzyme during concentration. Scraped (crude) sweat samples contained less PA activity. Sephacryl S-200 gel chromatography of the PA-positive pooled sweat showed a major peak of PA activity at M(r) 55,000. Gelatin-polyacrylamide enzymography revealed a major PA band at M(r) 55,000 and a minor band at 33,000. Sweat PA activity was 94% inhibited by epidermal PA inhibitor and anti-uPA IgG, but not by anti-tPA IgG. We conclude that the PA activity in sweat is derived from the sweat gland and is most likely of the urokinase type. The physiological significance of sweat uPA remains to be determined.
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PMID:Urokinase-type plasminogen activator in human eccrine sweat. 845 52

The regulation of microvessel formation and the expression of CNS-specific endothelial properties are attributed to perivascular astroglia. Specific proteolytic pathways mediate processes such as tissue remodeling, differentiation, invasion, and metastasis. We used a co-culture system in which C6 glial cells induce CNS microvascular endothelial cells to form capillary-like structures to examine the role of plasminogen activators and collagenases in CNS microvessel morphogenesis. Fibrin zymography revealed the presence of high-molecular weight urokinase-type plasminogen activator (uPA), low-molecular weight uPA, and uPA/inhibitor complexes within endothelial cultures and cocultures. Gelatin zymography revealed the presence of 92-, 72-, and 62-kDa type IV collagenases within endothelial cultures and cocultures. uPA activity was confirmed by incubating the extracts with amiloride, an inhibitor of uPA. Collagenase activity was confirmed by incubating the gels with EDTA, an inhibitor of metalloproteinases. Quantitative densitometry showed a six- to eightfold decrease in coculture uPA during capillary-like structure formation. Substantially less change in type IV 72-kDa procollagenase activity was seen in cocultures during capillary-like structure formation, but active type IV 62-kDa collagenase activity was significantly increased during capillary-like structure formation. These findings establish that uPA and activated type IV collagenase activity specifically regulates morphogenic endothelial responses to glial interactions and suggest mechanisms by which microvessels respond within the CNS.
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PMID:Modulation of serine proteinases and metalloproteinases during morphogenic glial-endothelial interactions. 862 23

In the present study, the time-dependent collagenolytic potential and mRNA transcription of extracellular matrix (ECM)-degrading components, transforming growth factor beta1 (TGFbeta1), and both basic fibroblast growth factor (bFGF) and FGF receptors (FGFR) in bovine cumulus-oocyte complexes (COCs) were investigated during 24 h of in vitro maturation (IVM). COCs were collected from 2- to 6-mm follicles, cultured in maturation medium, and sequentially removed at 3-h intervals for analysis. From 285 oocytes matured under these conditions, 114 (40.0%) developed to blastocysts on Day 9 after fertilization. Gelatin zymograms revealed protease activity at about 55 kDa for COCs matured for at least 3 h and two additional proteolytic zones at about 70 kDa after at least 9 h of IVM. The mRNAs of ECM-degrading components urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor 1 (PAI1), matrix metalloproteinase 1 (MMP1), and tissue inhibitor of metalloproteinases 1 (TIMP1), as well as TGFbeta1, bFGF, and FGFR, were detected during IVM in a factor-specific pattern: all transcript levels found at COC 0 generally increased after 3 h of maturation and either remained high or decreased thereafter. On the basis of the chosen reverse transcription-polymerase chain reaction technique, one could suggest relative higher mRNA concentrations for TIMP1, PAI1, and both growth factors compared to uPA, MMP1, and FGFR. Our results suggest a finely tuned extracellular proteolysis during IVM of bovine COCs, for which the concerted action of modulating growth factors like bFGF and TGFbeta1 may be essential.
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PMID:Growth factors and components for extracellular proteolysis are differentially expressed during in vitro maturation of bovine cumulus-oocyte complexes. 974 28

We and other researchers have previously found that colony-stimulating factors (CSFs), which generally include granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF), promote invasion by lung cancer cells. In the present study, we studied the effects of these CSFs on gelatinase production, urokinase plasminogen activator (uPA) production and their activity in human lung cancer cells. Gelatin zymographs of conditioned media derived from human lung adenocarcinoma cell lines revealed two major bands of gelatinase activity at 68 and 92 kDa, which were characterized as matrix metalloproteinase (MMP)-2 and MMP-9 respectively. Treatment with CSFs increased the 68- and 92-kDa activity and converted some of a 92-kDa proenzyme to an 82-kDa enzyme that was consistent with an active form of the MMP-9. Plasminogen activator zymographs of the conditioned media from the cancer cells showed that CSF treatment resulted in an increase in a 48-55 kDa plasminogen-dependent gelatinolytic activity that was characterized as human uPA. The conditioned medium from the cancer cells treated with CSFs stimulated the conversion of plasminogen to plasmin, providing a direct demonstration of the ability of enhanced uPA to increase plasmin-dependent proteolysis. The enhanced invasive behaviour of the cancer cells stimulated by CSFs was well correlated with the increase in MMPs and uPA activities. These data suggest that the enhanced production of extracellular matrix-degrading proteinases by the cancer cells in response to CSF treatment may represent a biochemical mechanism which promotes the invasive behaviour of the cancer cells.
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PMID:Granulocyte, granulocyte-macrophage, and macrophage colony-stimulating factors can stimulate the invasive capacity of human lung cancer cells. 1040 91

The in vitro release of matrix-degrading proteinases from breast cancer cells is associated in part with shed membrane vesicles. To determine whether shed vesicles might play a similar role in ovarian cancer cells, we analyzed the shedding phenomenon in vivo and in vitro as well as the enzymatic content of their vesicles. This is the first time that an immunoelectron microscopical analysis revealed membrane vesicles carrying tumor-associated antigen alpha-Folate Receptor (alpha-FR), circulating in biological fluids (ascites and serum) of an ovarian carcinoma patient. These vesicles were trapped in a fiber network with characteristic fibrin periodicity. An ovarian cancer cell line (CABA I) established from ascitic fluid cells of this patient, grew in Matrigel and formed tubular structures suggesting invasive capability. Immunofluorescence analysis demonstrated strong cytoplasmic staining of CABA I cells with anti-matrix metalloproteinase-9 (MMP-9) and anti-urokinase-type plasminogen activator (uPA) antibodies. CABA I cells shed membrane vesicles, which were morphologically similar to those identified in vivo, as determined by electron microscopy. Gelatin zymography of vesicles isolated both in vivo and in vitro revealed major gelatinolytic bands of the MMP family, identified as the zymogen and active forms of gelatinase B (MMP-9) and gelatinase A (MMP-2). By casein-plasminogen zymography we observed high-molecular weight (HMW)-uPA and plasmin bands. Incubation of purified vesicles from CABA I cells with Matrigel led to cleavage of Matrigel components. Taken together, our results point to a possible role of shed vesicles, both in vivo and in vitro, in proteolysis that mediates invasion and spread of ovarian epithelial carcinoma cells.
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PMID:Matrix-degrading proteinases are shed in membrane vesicles by ovarian cancer cells in vivo and in vitro. 1041 Nov 5

A prior study has reported that a rapid recanalization therapy of cerebral embolism, using liquid jet impacts generated by the interaction of gas bubbles with shock waves, can potentially penetrate through thrombi in as little as a few microseconds with very efficient ablation (Kodama et al. 1997). The present study was undertaken to examine the liquid jet impact effect on fibrinolysis in a tube model of an internal carotid artery. First, the conditions for generating the maximum penetration depth of liquid jets in the tube were investigated. Gelatin was used to mimic thrombi. The shock wave was generated by detonating a silver azide pellet weighing about a few micrograms located in a balloon catheter. The collapse of the inserted gas bubbles and the subsequent liquid jet formation were recorded with high-speed photography. Second, thrombi were formed using fresh human blood from healthy volunteers. The fibrinolysis induced by the liquid jet impact with urokinase was explored. This was conducted under selected conditions based on the experiment using the gelatin. Fibrinolysis was calculated as the percentage of the weight loss of the thrombus. Fibrinolysis with urokinase alone and with a single liquid jet impact with urokinase was 1.9 +/- 3.7% (n = 16) and 20.0 +/- 9.0% (n = 35), respectively, for an incubation time of 60 min. Statistical differences were obtained between all groups (ANOVA). These results suggest that liquid jet impact thrombolysis has the potential to be a rapid and effective therapeutic modality in recanalization therapy for patients with cerebral embolism and other clinical conditions of intra-arterial thrombosis.
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PMID:Liquid jets, accelerated thrombolysis: a study for revascularization of cerebral embolism. 1046 27

Wound healing is a complex process involving the interactions of many different cell types, matrix components and biological factors, including proteinases and cytokines. This study compared the levels of proteinases (matrix metalloproteinases and plasminogen activators), proteinase inhibitors (tissue inhibitors of metalloproteinases and plasminogen activator inhibitors), inflammatory cytokines and growth factors in acute wound fluid samples collected from the surgical drains of elective breast (n = 24) and colorectal (n = 26) patients on the first postoperative day. Gelatin zymography was used to determine matrix metalloproteinase-2 and -9 levels, quenched fluorescence substrate hydrolysis was applied for total MMP activity and enzyme-linked immunoassays were used to quantitate other factors. Colorectal wound fluid samples showed significantly (p < 0.05) greater levels of the matrix metalloproteinases (MMP-1, 2, 3, and 9), tissue inhibitor of metalloproteinases-1, urokinase plasminogen activator receptor and the inflammatory cytokines (interleukin-1beta, -6, and tumor necrosis factor-alpha); e.g., matrix metalloproteinase-3 colon; median 275 (range 11-2.530) ng/ml; breast; 530-400. However, tissue plasminogen activator and growth factor levels (epidermal growth factor, platelet-derived growth factor, basic fibroblast growth factor, transforming growth factor-beta1) were significantly greater in breast samples; e.g., epidermal growth factor breast 468 (103-1, 444) pg/ml; colon 57(1-573). There was no difference in the levels of urokinase type plasminogen activator, plasminogen activator inhibitor-1 and -2, tissue inhibitor of metalloproteinases -2 or vascular endothelial growth factor. Acute wound fluid from different surgical wounds showed different profiles of proteinases, proteinase inhibitors, and cytokines. This may lead to differences in the rate of tissue remodeling and therefore healing in these two wounds in vivo.
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PMID:Proteinases, their inhibitors, and cytokine profiles in acute wound fluid. 1111 51

Components of the extracellular matrix take part in tissue rebuilding as well as activating surface-bound growth factors. In the present study, expression and selected activities of urokinase-type plasminogen activator (uPA), matrix metalloproteinases (MMPs), their inhibitors (plasminogen activator inhibitor 1 (PAI-1) and tissue inhibitors of metalloproteinases (TIMPs)) were examined in bovine oviducts by RT--PCR, ribonuclease protection assay and activity assays. A high content of mRNA encoding for uPA was detected before ovulation with a three-fold decrease after ovulation. In contrast, PAI-1 expression appeared to be stable during the oestrous cycle. Oviductal flushings produced caseinolytic zones in zymograms containing plasminogen at approximately 50 kDa and 28 kDa. An activity assay for uPA showed highest net activity during the early to mid-luteal phase. Increased TIMP-1 and MMP-2 mRNA concentrations were found around the time of ovulation compared with the luteal phase. In contrast, MMP-1 mRNA transcripts were enriched during the early to mid-luteal phase. Gelatin zymograms detected a 70--72 kDa protease activity showing an oestrous cycle-dependent activity with highest activity before ovulation. Reverse zymography detecting TIMPs revealed proteins between 21 kDa and 24 kDa. Only for the smallest (21 kDa) protein were amounts increased around the time of ovulation compared with the luteal phase. The observation that several extracellular matrix components were regulated distinctly in bovine oviducts indicates that local interactions between these components, growth factors, gametes and the embryo are possible and may influence fertilization and early embryonic development.
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PMID:Differential expression of extracellular matrix components in the bovine oviduct during the oestrous cycle. 1142 36

Although it has been shown that the cross-talk between osteoblasts and tumor cells stimulates proliferation and invasion of prostate carcinoma (PCa) cells, the molecular mechanisms underlying this event are largely unknown. In this study, we demonstrated that the PCa cells, PC3, derived from bone metastasis, undergo changes of their invasive capability if grown in the presence of osteoblast-derived conditioned media (OBCM). Specifically, they were able to organize tridimensional structures in Matrigel, such as large branching colonies, tube-like structures and clusters of proliferating cells, after treatment. At the ultrastructural level, we observed that PC3 cells grown in the presence of OBCM presented an increment of membrane activity with a blast of shed membrane vesicles from the cell surface. After 6 h of incubation, protein content was approximately 5-fold more elevated in vesicles isolated from PC3 cells cultured in OBCM than in unstimulated cultures. Gelatin zymography of vesicles collected from OBCM-treated PC3 cells showed an increment of lytic bands of MMP family members identified as pro-enzymatic and active forms of gelatinase A (MMP-2) and gelatinase B (MMP-9). By casein-plasminogen zymography, this latter culture also presented an elevated level of high-molecular weight urokinase plasminogen activator (HMW-uPA). Purified vesicles from OBCM-treated PC3 cells incubated with Matrigel cleaved its components more efficiently than vesicles from untreated PC3 cells. Collectively, these findings indicate that osteoblasts produce factor/s able to modify the invasive capability of prostate cancer cells, increasing the amount of shed vesicles and of their associated lytic enzymes.
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PMID:Osteoblast-conditioned media stimulate membrane vesicle shedding in prostate cancer cells. 1652 40


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