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)

The endothelial lining of blood vessels is constantly exposed to fluid mechanical forces generated by flowing blood. In vitro application of fluid shear stresses to cultured endothelial cells influences the expression of multiple genes, as reflected by changes in their steady-state mRNA levels. We have utilized the B chain of platelet-derived growth factor (PDGF-B) as a model to investigate the mechanisms of shear-stress-induced gene regulation in cultured bovine aortic endothelial cells (BAECs). Northern blot analysis revealed elevated endogenous PDGF-B transcript levels in BAECs, after exposure to a physiological level of laminar shear stress (10 dynes/cm2; 1 dyne = 100 mN) for 4 h. A transfected reporter gene, consisting of a 1.3-kb fragment of the human PDGF-B promoter coupled to chloramphenicol acetyltransferase (CAT), indicated a direct effect on transcriptional activity. Transfection of a series of PDGF-B-CAT deletion mutants led to the characterization of a cis-acting component within the PDGF-B promoter that was necessary for shear-stress responsiveness. In gel-shift assays, overlapping oligonucleotide probes of this region formed several protein-DNA complexes with nuclear extracts prepared from both static and shear-stressed BAECs. A 12-bp component (CTCTCAGAGACC) was identified that formed a distinct pattern of complexes with nuclear proteins extracted from shear-stressed BAECs. This shear-stress-responsive element does not encode binding sites for any known transcription factor but does contain a core binding sequence (GAGACC), as defined by deletion mutation in gel-shift assays. Interestingly, this putative transcription factor binding site is also present in the promoters of certain other endothelial genes, including tissue plasminogen activator, intercellular adhesion molecule 1, and transforming growth factor beta 1, that also are induced by shear stress. Thus, the expression of PDGF-B and other pathophysiologically relevant genes in vascular endothelium appears to be regulated, in part, by shear-stress-induced transcription factors interacting with a common promoter element.
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PMID:Platelet-derived growth factor B chain promoter contains a cis-acting fluid shear-stress-responsive element. 835

The present study was undertaken to evaluate in vitro the relative importance of tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) in the mitogenic and chemotactic potential of bovine fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF)-BB for smooth muscle cells (SMC). Aortic SMC were isolated from transgenic mice showing single inactivations of the t-PA, u-PA, plasminogen activator inhibitor-1, or urokinase-type plasminogen activator receptor (u-PAR) genes. With regard to serum-induced proliferation, all cell types showed similar responses. However, SMC isolated from t-PA-deficient mice did not proliferate or migrate in response to PDGF, whereas SMC isolated from u-PA-deficient animals appeared to be much less sensitive to bFGF than the cells isolated from the other animals. Supplementation of cells from deficient animals with exogenous murine t-PA or u-PA restored the normal response of the growth factors with regard to both migration and proliferation. The mitogenic and chemotactic responses of bFGF were specifically inhibited in u-PAR-deficient cells or in wild-type SMC, cultured in the presence of antibodies to u-PAR. The role of u-PA and t-PA in bFGF and PDGF-induced growth and migration of SMC was not dependent on plasmin generation and activity as demonstrated by the inactivity of epsilon-aminocaproic acid and aprotinin. A 4-5-fold increase in the steady-state levels of u-PA and t-PA mRNA and proteins were observed after 24 h of incubation of the cell cultures with bFGF and PDGF-BB, respectively. These results therefore indicate that, at least in vitro, t-PA is an important element of the activity of PDGF-BB with regard to the proliferation and migration of SMC whereas u-PA is a key factor in the effect of bFGF on SMC.
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PMID:Urokinase and tissue-type plasminogen activator are required for the mitogenic and chemotactic effects of bovine fibroblast growth factor and platelet-derived growth factor-BB for vascular smooth muscle cells. 929 97

Human omental microvascular endothelial (HOME) and mesothelial (MESO) cells share many phenotypic properties, but can be characterized from one another based upon a comprehensive panel of endothelial and mesothelial markers. Traditional cell markers such as von-Willebrand factor, DiI-Ac-LDL, and Ulex europaeus I lectin are not sufficient to distinguish between HOME and MESO cells. Furthermore, immunoreactivity to a panel of endothelial cell-specific monoclonal antibodies, including representatives from the known clusters of differentiation (CD), indicate that some of these antigens are coexpressed in HOME and MESO cells. In distinguishing between the two cell types, HOME and not MESO cells express E-selectin, E/P-selectin, P-selectin (CD62), Le-y, and VLA-6 (CDw49f*). Moreover, HOME cells and not MESO cells form tube-like structures when cultured on Matrigel. MESO cells differ from HOME cells based upon (1) the expression of cytokeratins; (2) their rapid proliferation in response to platelet-derived growth factor; and (3) a change from an epitheliod to fibroblast-like morphology in response to tumor necrosis factor and epidermal growth factor. Both HOME and MESO cells express tissue plasminogen activator and plasminogen activator inhibitor, but urokinase activity is only expressed by MESO cells. As there is no one universal endothelial or mesothelial cell marker that can specifically confirm the identity of these cells, it appears necessary to employ a comprehensive panel of cell markers to rule out the possibility of misidentifying a cell culture.
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PMID:Human omental microvascular endothelial and mesothelial cells: characterization of two distinct mesodermally derived epithelial cells. 932 82

Repair of demyelination in the CNS requires that oligodendrocyte precursors (OPs) migrate, divide and then myelinate. Repair of axon damage requires axonal regeneration. Limited remyelination and axon regeneration occurs soon after injury, but usually ceases in a few days. In vivo and in vitro experiments have shown that astrocytic environments are not very permissive for migration of OPs or for axonal re-growth. Yet remyelination and axon sprouting early after injury occurs in association with astrocytes, while later astrocytes can exclude remyelination and prevent axon regeneration. A large and changing cast of cytokines are released following CNS injury, so we investigated whether some of these alone or in combination can affect the ability of astrocytes to support migration of OPs and neuritic outgrowth. Interleukin (IL) 1alpha, tumour necrosis factor alpha, transforming growth factor (TGF) beta, basic fibroblast growth factor (bFGF), platelet-derived growth factor and epidermal growth factor alone exerted little or no effect on migration of OPs on astrocytes, whereas interferon (IFN) gamma was inhibitory. The combination of IL-1alpha + bFGF was found to be pro-migratory, and this effect could be neutralized by TGFbeta. We also examined neuritic outgrowth from dorsal root ganglion explants in three-dimensional astrocyte cultures treated with cytokines and found that IL-1alpha + bFGF greatly increased axon outgrowth and that this effect could be blocked by TGFbeta and IFNgamma. All these effects were absent or much smaller when OP migration or axon growth was tested on laminin, so the main effect of the cytokines was via astrocytes. The cytokine effects did not correlate with expression on astrocytes of laminin, fibronectin, tenascin, chondroitin sulphate proteoglycan, N-cadherin, polysialyated NCAM (PSA-NCAM), tissue plasminogen activator (tPA) or urokinase (uPA).
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PMID:Cytokine-induced changes in the ability of astrocytes to support migration of oligodendrocyte precursors and axon growth. 974 68

The objective of this study was to establish a technique to isolate porcine mesothelial cells (PMCs) from omental tissue and to compare them to human mesothelial cells (HMCs). The PMCs were dispersed by collagenase digestion and isolated on a Ficoll layer. Their morphologic and ultrastructural features were assessed at confluence by light and electronic microscopy, and they were characterized by immunohistochemistry using specific HMC markers. PMC proliferation was studied in the presence of growth factors platelet-derived growth factor (PDGF), epidermal growth factor (EGF) or transforming growth factors beta1, beta2, or beta3 (TGF). Fibrinolytic PMC activity was detected by zymography for tissue plasminogen activator (tPA) and by reverse zymography for plasminogen activator inhibitor-1 (PAI-1). The recalcification time of cell lysates was used to define PMC procoagulant activity, and gelatinase zymography was used to detect metalloproteinase production. At confluence, PMCs formed typical cobblestone monolayers and exhibited structural features characteristic of HMCs. Weibel Palade bodies were never seen. Specific HMC markers (HBME1, ME1, WT1) cross-reacted with PMCs. As HMCs and PMCs coexpressed cytokeratin and vimentin, and also expressed vinculin and alpha-actin. Addition of PDGF or EGF to the culture medium stimulated PMC proliferation. PMCs constitutively expressed fibrinolytic and procoagulant activity and secreted MMP9 and MMP2. The technique described in this study allows isolation of mesothelial cells from porcine omental tissue. These porcine cells exhibit a mesothelial phenotype and functional properties similar to those of HMCs. Our data warrant an evaluation of mesothelial cells as targets in several therapeutic strategies with porcine models.
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PMID:Phenotypic and functional characteristics of porcine peritoneal mesothelial cells. 1061 73

To investigate developmental differences in the wound repair process between fetal and adult skin fibroblasts, we studied the expression of plasminogen activator, plasminogen activator inhibitor, matrix metalloproteinase, and tissue inhibitor of metalloproteinase in E-15, E-17, newborn and adult mouse skin fibroblasts cultured within three dimensional matrices of either collagen or fibrin. Fibrin overlay and reverse overlay analyses revealed that mouse skin fibroblasts secreted tissue plasminogen activator and type1 plasminogen activator inhibitor. However, only E-15 and E-17 fibroblasts secreted the active form of tissue plasminogen activator, while in newborn and adult fibroblasts tissue plasminogen activator was conjugated to type1 plasminogen activator inhibitor. Only adult fibroblasts expressed a high level of active type1 plasminogen activator inhibitor. Gelatin zymography revealed that the predominant matrix metalloproteinase secreted by all the mouse fibroblasts was gelatinase A (matrix metalloproteinase -2). Matrix metalloproteinase -2 was partially activated in the adult fibroblasts cultured within a collagen matrix. The tissue inhibitor of metalloproteinase-2 was expressed by all fibroblasts, but levels were highest in the newborn and adult fibroblasts. When E-15 fibroblasts were cultured within a fibrin matrix, tissue plasminogen activator was downregulated. Transforming growth factor-betadownregulated tissue plasminogen activator while upregulating type1 plasminogen activator inhibitor, and platelet-derived growth factor enhanced tissue plasminogen activator expression in E-15 fibroblasts. Therefore, plasminogen activator and its inhibitor, and matrix metalloproteinase and its associated tissue inhibitor are differentially expressed in fetal and adult fibroblasts, and their expression is controlled by extracellular matrix components and growth factors present in wounds.
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PMID:Developmental differences in the expression and modulation of extracellular matrix proteases and inhibitors in mouse skin fibroblasts. 1063 6

The clinical benefit of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) may derive from a qualitative, functional change in atherosclerotic lesions in addition to their lipid-lowering properties. We examined whether statins altered expression of the major determinants of fibrinolytic balance, plasminogen activator inhibitor-1 (PAI-1), and tissue-type plasminogen activator (tPA) in human vascular smooth muscle (SMC) and endothelial (EC) cells. Simvastatin reduced levels of PAI-1 antigen released from SMCs and ECs stimulated with platelet-derived growth factor or transforming growth factor-beta (IC(50) approximately 1 micromol/L). Levels of EC-derived tPA increased 2-fold over the same concentrations of simvastatin that inhibited release of PAI-1. Simvastatin's inhibitory effect was mimicked by C3 exoenzyme and prevented by geranylgeranyl pyrophosphate, but not by farnesyl pyrophosphate, suggesting the involvement of geranylgeranyl-modified intermediates. Decreased PAI-1 antigen was correlated with reduced mRNA transcription and activity of the PAI-1 promoter. By inhibiting expression of PAI-1 from SMCs and ECs while increasing expression of tPA from ECs, simvastatin may alter the local fibrinolytic balance within the vessel wall toward increased fibrinolytic capacity that, in turn, would reduce thrombotic risk after plaque rupture.
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PMID:HMG CoA reductase inhibitors reduce plasminogen activator inhibitor-1 expression by human vascular smooth muscle and endothelial cells. 1066 56

Antisense therapy with phosphorothioate oligodeoxynucleotides (PS oligos) has emerged as a potentially useful strategy for inhibiting angioplasty restenosis. Several groups have reported that PS oligos inhibit in vitro vascular smooth muscle cell (SMC) proliferation as well as in vivo neointimal formation after rat carotid artery balloon injury. More recent studies have revealed the presence of a PS oligo G-quartet inhibitory effect, reflecting binding of oligonucleotides to cellular proteins, which is distinct from a hybridization-dependent antisense effect. Studies with the 28-mer phosphorothioate cytidine homopolymer S-dC28 have demonstrated the presence of a non-G-quartet, non-sequence-specific inhibitory effect on SMC proliferation and migration in vitro and neointimal hyperplasia after rat carotid balloon injury in vivo. These effects are the result, in part, of the avid binding of the polyanion PS oligos to heparin-binding growth factors, such as platelet-derived growth factor (PDGF) and basic fibroblast growth factor. Moreover, S-dC28 attenuates PDGF-induced SMC tissue plasminogen activator antigen levels without affecting SMC plasminogen activator inhibitor-1 levels, thereby suggesting a PS oligo net antifibrinolytic effect that would impede SMC migration. Therefore, the further development of these drugs to inhibit angioplasty restenosis must consider the hybridization-specific antisense effects, the non-G-quartet inhibitory effects, and the non-G-quartet, non-sequence-specific inhibitory effects of the pleiotropic PS oligos.
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PMID:Phosphorothioate Oligodeoxynucleotide Inhibition of Restenosis. 1076 6

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

Thrombolytic treatment of ischemic stroke with tissue plasminogen activator (tPA) is markedly limited owing to concerns about hemorrhagic complications and the requirement that tPA be administered within 3 h of symptoms. Here we report that tPA activation of latent platelet-derived growth factor-CC (PDGF-CC) may explain these limitations. Intraventricular injection of tPA or active PDGF-CC, in the absence of ischemia, leads to significant increases in cerebrovascular permeability. In contrast, co-injection of neutralizing antibodies to PDGF-CC with tPA blocks this increased permeability, indicating that PDGF-CC is a downstream substrate of tPA within the neurovascular unit. These effects are mediated through activation of PDGF-alpha receptors (PDGFR-alpha) on perivascular astrocytes, and treatment of mice with the PDGFR-alpha antagonist imatinib after ischemic stroke reduces both cerebrovascular permeability and hemorrhagic complications associated with late administration of thrombolytic tPA. These data demonstrate that PDGF signaling regulates blood-brain barrier permeability and suggest potential new strategies for stroke treatment.
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PMID:Activation of PDGF-CC by tissue plasminogen activator impairs blood-brain barrier integrity during ischemic stroke. 1860 66


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