Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

We examined the role of the plasminogen activator/plasmin system in extracellular matrix (ECM) degradation by human mesangial cells cultured on thin films of 125I-labeled ECM (Matrigel). ECM degradation (release of 125I into the medium) was dependent on exogenous plasminogen, proportional to the number of mesangial cells and amount of plasminogen added, and coincident with the appearance of plasmin in the medium. ECM degradation was completely blocked (P < 0.001) by two plasmin inhibitors, alpha-2-antiplasmin (40 micrograms/ml) and aprotinin (216 KIU/ml), and partially reduced (-33 +/- 1.8%, P < 0.01) by TIMP-1 (40 micrograms/ml), a specific inhibitor of matrix metalloproteinases. Zymography of medium obtained from cells cultured in the absence of plasminogen revealed the presence of latent matrix metalloproteinase-2 (MMP-2) which was converted to a lower molecular weight, active form in the presence of mesangial cells and plasminogen. Northern analysis of poly A+RNA prepared from cultured human mesangial cells revealed mRNA for tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor-1 (PAI-1), and uPA receptor (uPAR). The presence of uPA protein in medium obtained from cultured human mesangial cells was demonstrated by Western blotting and ELISA which revealed a large molar excess of PAI-1 (1.2 +/- 0.1 x 10(-9) M) over uPA (1.2 +/- 0.1 x 10(-12) M) and tPA (0.19 +/- 0.04 x 10(-9) M). ECM degradation was reduced by a monoclonal antibody (MAb) against human tPA (-54 +/- 8.6%) or human uPA (-39 +/- 5.2%) compared to cells treated with identical amounts of non-specific monoclonal IgG (P < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:ECM degradation by cultured human mesangial cells is mediated by a PA/plasmin/MMP-2 cascade. 754 Feb 30

We investigated in vitro chemotactic responses to fibronectin and laminin, invasion through reconstituted basement membrane (Matrigel) and secretion of matrix metalloproteinases and plasminogen activators by non-tumorigenic Mel-ab melanocytes; B16 melanoma; and the metastatic sublines, B16F1, B16F10 and B16BL6. In vitro chemotactic and invasive ability were not associated with in vivo metastatic potential. Secretion of various matrix-degrading enzymes was not related to in vitro invasion. Conditioned media from all B16 melanoma sublines, but not from Mel-ab cells, contained the M(r) 92,000 progelatinase. The activated M(r) 85,000 species was present only in conditioned media from Mel-ab, B16 and B16F1 cells. Mel-ab cells secreted copious amounts of the M(r) 72,000 progelatinase, and the M(r) 66,000 active form was also present in conditioned media. Secretion of the M(r) 72,000 progelatinase by B16 melanoma sublines was markedly lower, and only conditioned media from B16 cells contained the activated M(r) 66,000 form. Furthermore, cell lysates of Mel-ab cells contained a M(r) 67,000 metalloproteinase which was absent in the tumor cells. All cells secreted tissue plasminogen activator; however, the metastatic B16F1, B16F10 and B16-BL6 cells also secreted urokinase plasminogen activator. Our results indicate that matrix metalloproteinase secretion by itself is not associated with tumorigenicity or metastatic potential. Secretion of urokinase plasminogen activator, and not tissue plasminogen activator, reflected the metastatic characteristics of the B16 melanoma tumor sublines.
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PMID:Differences in expression of metalloproteinases and plasminogen activators in murine melanocytes and B16 melanoma variants: lack of association with in vitro invasion. 755 59

Endothelial cell differentiation into capillary structures is a complex process that requires the concerted effects of several extracellular matrix proteases, including plasminogen activators. Here, the role of tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA) was evaluated in an in vitro model of endothelial morphogenesis involving organization of human umbilical vein endothelial cells into tubular structures when they are cultured on the basement membrane preparation, Matrigel. Both uPA and tPA were detected in HUVEC cultures on Matrigel, and inhibitors of plasminogen activators or of serine proteases decreased the extent of the tube network formed by the cells. The decrease resulting from serine protease inhibitors was additive to that from matrix metalloproteinase inhibitors which have previously been shown to decrease tube formation in this model, suggesting that the two classes of proteases modulate tube formation by distinct mechanisms. Plasminogen activator inhibitor (PAI)-1 decreased tube formation by 50% when added up to 4.5 h after the initiation of an 18 h assay and caused 25% inhibition when added 9.5 h after culture initiation, indicating that the effects of plasminogen activators are not limited to an early event in the differentiation process. Steady-state expression of mRNA for uPA increased during the first several hours of culture on Matrigel, further supporting a role for PA activity throughout the process of tube formation. These findings suggested that PAs may affect multiple events during tube-forming activity. A fucosylated peptide comprising the amino-terminal domain of uPA that binds to the uPA receptor (uPAR) but lacking proteolytic activity enhanced tube formation. In contrast, a defucosylated form of the same peptide had no effect. Since fucosylation of this fragment has been shown to be essential in other models of cell stimulation by uPA-uPAR interaction, these data support the hypothesis that uPA enhances endothelial morphogenesis both through proteolytic activity and via uPAR occupancy. Plasminogen activators could facilitate angiogenesis in vivo.
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PMID:Plasminogen activators augment endothelial cell organization in vitro by two distinct pathways. 755 92

Heparin inhibits the migration and proliferation of arterial smooth muscle cells and modifies the extracellular matrix. These effects may be the result of heparin's effects on proteinases that degrade the matrix. We have previously reported that heparin inhibits the induction of tissue-type plasminogen activator and interstitial collagenase mRNA. We have investigated the possibility that heparin affects other members of the matrix metalloproteinase family. Phorbol ester increased the levels of mRNA of collagenase, 92-kD gelatinase and stromelysin as well as the synthesis of these proteins. These effects were inhibited by heparin, but not by other glycosaminoglycans, in a dose-dependent manner. The induction of these matrix metalloproteinases was also inhibited by staurosporine and pretreatment with phorbol ester indicating the involvement of the protein kinase C pathway. In contrast, the 72-kD gelatinase was expressed constitutively and was not affected by phorbol ester or heparin. Tissue inhibitor of metalloproteinase-1 was expressed constitutively and was slightly increased by phorbol ester. It was not affected by heparin. Thus, heparin inhibits the production of four proteinases (tissue plasminogen activator, collagenase, stromelysin and 92-kD gelatinase) that form an interdependent system capable of degrading all the major components of the extracellular matrix.
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PMID:Heparin inhibits the induction of three matrix metalloproteinases (stromelysin, 92-kD gelatinase, and collagenase) in primate arterial smooth muscle cells. 818 30

Constitutive overexpression of both urokinase and matrix metalloproteinase (MMP) activity is frequently observed in individual malignant tumors. In this study we describe the combined contribution of these distinct enzyme systems to the invasive phenotype of a highly metastatic human melanoma cell line (M24met). M24met cells were found to secrete a spectrum of MMPs, including interstitial collagenase, type IV collagenases (M(r) 92,000 and 72,000 progelatinases), and stromelysin. Urokinase, but not tissue-type plasminogen activator, was detected in M24met-conditioned media and on cell surfaces. The contribution of these enzymes to extracellular matrix dissolution was determined by exploiting specific inhibitors, namely tissue inhibitor of the metalloproteinases-2 and plasminogen activator inhibitor-2. Due to the coexpression of urokinase and MMP-dependent activity, M24met cells were observed to degrade multiple components of the extracellular matrix and to significantly degrade both interstitial and basement membrane matrices. Urokinase-dependent removal of matrix glycoprotein was observed to precede MMP-dependent collagenolysis as a prerequisite rate-limiting step. We present evidence which suggests that this temporal relationship is imposed by the structural architecture of the matrix such that matrix glycoprotein serves to protect associated collagen from MMP-dependent degradation. In addition to mediating significant collagenolysis, MMP activity was further implicated in the dissolution of matrix tropoelastin. Urokinase/plasmin activity was not found to be required for MMP-zymogen activation.
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PMID:Melanoma-mediated dissolution of extracellular matrix: contribution of urokinase-dependent and metalloproteinase-dependent proteolytic pathways. 842 5

Polarized secretion of matrix metalloproteinases and plasminogen activators by monkey aortic endothelial cells was studied in vitro, using transwell inserts. The endothelial cells constitutively expressed matrix metalloproteinase-2, tissue inhibitors of metalloproteinases 1 and 2, urokinase, and tissue plasminogen activator, all with basal preference. Matrix metalloproteinase-9 activity was induced by phorbol 12-myristate 13-acetate (apical), interleukin-1 alpha (basal), and by conditioned medium from DX3 human melanoma cells (basal). The DX3 melanoma conditioned medium also stimulated basal secretion of matrix metalloproteinase-2, urokinase, tissue plasminogen activator, and tissue inhibitors of metalloproteinases. The rise in proteolytic activity in the basal direction was reflected by increased capacity to degrade subendothelial basement membrane type IV collagen, shown immunohistologically, using monkey kidney tissue sections and basement membrane deposited by endothelial cells into the transwell membrane. Thus, IL-1 alpha and DX3 melanoma conditioned medium can stimulate endothelial cells in vitro to concentrate secretion of proteinases spatially onto the underlying basement membrane. We suggest that the stimulation of endothelial cell proteinase activity by tumor cells may facilitate tumor cell extravasation.
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PMID:Alterations in endothelial cell proteinase and inhibitor polarized secretion following treatment with interleukin-1, phorbol ester, and human melanoma cell conditioned medium. 882 24

The migration of arterial smooth muscle cells (SMCs) plays an important role in normal vessel development as well as the pathobiology of blood vessels. Because it is difficult to study cell migration in primates, we used ex vivo explants. The response of baboon aortic medial explants incubated in vitro in a serum-free medium with insulin and transferrin was compared with the response of whole artery injured in vivo by a balloon catheter to establish the validity of the explant model. Both the time course of entry of SMCs into the S phase and the changes in matrix metalloproteinase 9 were similar in the artery and the explants. SMCs began migrating from explants after a lag of 3 days. By day 11, > 90% of the explants exhibited SMC migration from the tissue (percent of explants with > or = 1 migrating cell). Basal migration was inhibited by antibodies to urokinase and tissue-type plasminogen activator, whereas addition of plasminogen to the explants increased migration. An inhibitor of matrix metalloproteinases. BB-94 (Batimistat), decreased migration, as did alpha 2-macroglobulin. These data demonstrate that proteinases of the matrix metalloproteinase and plasminogen/plasminogen activator families play an important role in the migration of primate arterial SMCs through the extracellular matrix.
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PMID:The role of plasminogen, plasminogen activators, and matrix metalloproteinases in primate arterial smooth muscle cell migration. 891 Dec 76

The concentrations of matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-8 (MMP-8), matrix metalloproteinase-9 (MMP-9), lactoferrin and urokinase plasminogen activator (uPA), tissue-type plasminogen activator (tPA) and the inhibitors, tissue inhibitor of metalloproteinase-1 (TIMP-1), plasminogen activator inhibitor-1 (PAI-1), plasminogen activator inhibitor (PAI-2), and alpha2-macroglobulin in the synovial fluids of patients with rheumatoid arthritis was determined before and during chemical synoviorthesis with a sodium salt of the fatty acids from cod-liver oil (Varicocid). Synovial fluids were obtained before treatment from 37 patients with rheumatoid arthritis and, in most cases, at 8 and 24 h after injection of the agent. Well-established ELISAs were used to determine the amounts of all proteins. All patients with rheumatoid arthritis revealed very high levels of metalloproteinases (about 1-15 mu g/ml) in their synovial fluids. During the inflammation inducing treatment the granulocyte enzymes increased. In contrast to this, the level of MMP-1 decreased. All granulocyte-derived enzymes were strongly correlated with each other, whereas their dependence on the granulocyte count was only weak. uPA and PAI-2 showed good correlations with the granulocytes-derived enzymes, but were also only weakly correlating with the cell counts. t-PA was not detected by the ELISA used. The proteases, MMP-8, MMP-9 and uPA were increased 8 h after the treatment, whereas the specific inhibitors TIMP-1, PAI-1 and PAI-2 showed significant changes only 24 h after the injection. Matrix metalloproteinases are important factors in the pathogenesis of rheumatoid arthritis. The inflammatory activity in the joint could be better correlated to the granulocyte enzymes than to the granulocyte counts. The levels of uPA and PAI-2 are also parallel to the granulocyte enzyme levels and might underly the same regulatory mechanism.
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PMID:Determination of metalloproteinases, plasminogen-activators and their inhibitors in the synovial fluids of patients with rheumatoid arthritis during chemical synoviorthesis. 891 99

Although heart attack is caused by occlusion of a major coronary artery, some patients have occlusion without heart attack because these patients have sufficient collateral circulation to provide an alternate pathway for blood supply to the myocardium at ischemic risk. The growth of new capillary vessels (angiogenesis) and enlargement of preexisting vessels play an important role in the collateral development. We evaluated the hypothesis that extracellular matrix metalloproteinase (MMP) expression is altered in coronary collateral arteries (0.5-1 mm o.d.) isolated from canine hearts 2-4 months after surgical placement of an ameroid occluder around the proximal left circumflex artery (n = 4), during the development of collateral vessels and restructuring new vessels. Histologic studies (hematoxylin and eosin, trichrome, and van Gieson stains) indicated cellular proliferation and increased collagen and elastin content in collateral vessels compared with comparable-sized unoccluded arterial segments of the left anterior descending (LAD) artery. In situ MMP activity of collateral vessels, measured using denatured collagen in the gel matrix, indicated an increase in total MMP activity in the intima of collateral vessels compared with normal LAD vessels. To further identify the type of MMP, tissue homogenates were prepared from collateral and LAD vessels and analyzed by SDS-PAGE zymography. The results suggest induction of gelatinase A and gelatinase B expression in collateral vessels compared with normal LAD tissue, when identical amounts of total protein were loaded onto each lane in the gel. Based on plasminogen-casein zymography, we observed the tissue plasminogen activator level to be increased in collateral vessels. On the basis of immunoblot and mRNA (Northern blot) analyses, we determined that the MMP-1 level was induced in collateral vessels 2 and 4 months after ameroid occlusion. In contrast with MMP-1, the level of TIMP-1 (tissue inhibitor of metelloproteinases) was decreased significantly (p < 0.001) in collateral compared with LAD vessels, suggesting a role for arterial TIMP in anti-angiogenic activity. Collectively, these results suggest that chronic occlusion of a major coronary artery induces upregulation of vascular remodeling mechanisms subserving collateral development. Increased MMP-2 activity in collaterals may be associated with decreased levels of tissue inhibitor of metalloproteinases and fibrous tissue remodeling following angiogenic and (or) adaptive responses of the myocardium to chronic ischemia.
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PMID:Temporal expression of extracellular matrix metalloproteinases and tissue plasminogen activator in the development of collateral vessels in the canine model of coronary occlusion. 896 Mar 89

Myoepithelial cells in situ and in vitro exert important paracrine effects on carcinoma cells which are mediated by high expression of extracellular matrix molecules, proteinase inhibitors and angiogenic inhibitors. Myoepithelial xenografts (human matrix secreting (HMS)-X, HMS-3X and HMS-4X) established from benign human salivary gland and breast myoepithelial tumors accumulate an abundant extracellular matrix which can be extracted with 6 M urea and 2 M guanidinium hydrochloride to form a gel at 25-37 degrees C. This gel, termed Humatrix, exhibits different biochemical and biological properties than the conventional non-human matrical gels in existence, i.e. Matrigel and Vitrogen 100. Whereas Matrigel consists mainly of basement membrane molecules, e.g. laminin, type IV collagen and heparan sulfate proteoglycan, and Vitrogen 100 consists mainly of non-basement membrane molecules, e.g. type I and type III collagen, Humatrix contains significant amounts of both basement membrane and non-basement membrane molecules, including large amounts of chondroitin sulfate proteoglycan. Like Matrigel, Humatrix contains bound growth factors, including epidermal growth factor (EGF) and insulin-like growth factor-I (IGF-I); unlike Matrigel, which contains predominantly significant quantities of bound proteinases, including tissue-type plasminogen activator (tPA), matrix metalloproteinase (MMP)-2 and MMP-9, and angiogenic factors, including basic fibroblast growth factor (bFGF) and transforming growth factor (TGF)-beta, Humatrix contains predominantly bound proteinase inhibitors such as protease nexin II (PN-II) and alpha1-antitrypsin and angiogenic inhibitors such as thrombospondin-1. Humatrix selectively stimulates the growth and tumorigenicity of human myoepithelial cell lines but inhibits invasion, angiogenesis and metastasis of other non-myoepithelial malignant cell lines. Because of its myoepithelial origin Humatrix represents a more natural source of extracellular matrix molecules and bound factors that carcinoma cells encounter in vivo.
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PMID:Humatrix, a novel myoepithelial matrical gel with unique biochemical and biological properties. 948 91


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