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

The urokinase-type plasminogen activator receptor (u-PAR) was demonstrated on cultured smooth muscle cells (SMCs) of bovine aorta. Binding of 125I-urokinase-type plasminogen activator (u-PA) was concentration dependent and saturable within 45-60 minutes. A similar concentration and time dependence was found in functional plasminogen activation studies. Human two-chain high-molecular-weight u-PA and its proenzyme (pro-u-PA) bound specifically with identical affinity (Kd). Activation of pro-u-PA was strongly accelerated on binding to SMCs and occurred only in the presence of plasminogen on the cell surface. A 100-fold molar excess of unlabeled high-molecular-weight u-PA effectively blocked binding of the radiolabeled ligands; tissue-type plasminogen activator, plasminogen, low-molecular-weight u-PA, and unrelated proteins did not. 125I-u-PA binding was abolished by a monoclonal antibody against the specific u-PA sequence responsible for u-PAR binding. Binding of u-PA sharply decreased on SMC exposure to phosphatidylinositol-specific phospholipase C, confirming the glycan phospholipid cell anchorage of u-PAR. Bovine and human alpha-thrombin (240 nM) increased the binding of 125I-u-PA fivefold, translating into an increase in the number of sites per cell from about 10(5) to 5 x 10(5) without significant change in the Kd (1.29 +/- 0.39 nM). Active site blockade of thrombin by D-Phe-Pro-Arg-chloromethyl ketone resulted in the total loss of stimulatory activity, as did the use of the inactive active site thrombin mutant, S205A. Hirugen (100 microM), which blocks the anion-binding exosite of thrombin, blocked u-PAR stimulating activity. Thus, both the catalytic activity and integrity of the exosite are important for thrombin's stimulatory activity. Other SMC mitogens (epidermal growth factor, transforming growth factor-beta 1, basic fibroblast growth factor, platelet-derived growth factor, and phorbol 12-myristate 13-acetate) increased u-PAR expression on SMCs six- to 20-fold while concomitantly increasing Kd four- to 10-fold. In all cases the induction of u-PAR was dependent on de novo protein synthesis. These observations assign a possible role for thrombin and other mitogens in u-PAR regulation, thereby influencing the pericellular proteolysis that is important in SMC migration and atheromatous plaque development.
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PMID:Regulation of the urokinase-type plasminogen activator receptor on vascular smooth muscle cells is under the control of thrombin and other mitogens. 132 97

We have compared the cell-specific expression and regulation of the receptor for urokinase-type plasminogen activator (u-PAR) by transforming growth factor beta type 1 (TGF-beta 1) in 10 human cell lines derived from both normal and neoplastic tissues. The basal expression of u-PAR mRNA as well as its response to TGF-beta 1 varied strongly between different cell lines; however, five out of the 10 cell lines responded to TGF-beta 1 by an increase in the u-PAR mRNA level. Among these, A549 cells were selected for a detailed elucidation of the molecular mechanism involved in TGF-beta 1 regulation of u-PAR mRNA expression. TGF-beta 1 caused an early increase in u-PAR mRNA level, with a maximal 15-fold enhancement after 24 h of treatment. This was paralleled by an increase in u-PAR protein as detected by crosslinking studies with radiolabeled ligand, and also resulted in an increase in cell surface plasmin generation. The protein synthesis inhibitor cycloheximide also increased the level of u-PAR mRNA in a time-dependent fashion and when both cycloheximide and TGF-beta 1 were used, an additive effect was seen. Nuclear run-on experiments demonstrated only a moderate (3-fold) increase in the u-PAR gene transcription rate after exposure of the cells to TGF-beta 1 for 3 h compared with a 12-fold increase in the mRNA level. TGF-beta 1 also caused an increase of both u-PA and PAI-1 antigens, while there was no detectable effect on t-PA.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Urokinase-receptor biosynthesis, mRNA level and gene transcription are increased by transforming growth factor beta 1 in human A549 lung carcinoma cells. 165 20

Using immunohistochemistry and in-situ hybridization, we studied the expression of the components of the plasminogen activation system during progression to malignant melanoma with fresh melanocytic lesions. Expression of these components is confined to late stages of melanoma. t-PA expression is limited to rare cases of metastatic melanoma. The other components are frequently expressed concomitantly in the same tumour. Urokinase (u-PA) is expressed in stromal cells and only in tumour cells at invasive foci, urokinase receptor (u-PAR) in tumour cells, plasminogen activator inhibitor type I (PAI-1) in the intratumoral extracellular matrix and plasminogen activator inhibitor type II (PAI-2) in tumour cells and stromal cells. In order to investigate the role of u-PAR as a prognostic marker, we have developed an assay for quantitation of the receptor. As a first step towards structural investigations, we have determined the disulfide cross-links of the first domain of uPAR.
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PMID:Studies on functional and structural role of urokinase receptor and other components of the plasminogen activation system in malignancy. 774 86

We have analyzed the occurrence of components of the plasminogen activation system in bovine milk. Zymographic analyses showed that tissue-type plasminogen activator (t-PA) occurred in association with casein micelles, partially as a complex with type-1 plasminogen activator inhibitor (PAI-1), whereas urokinase-type plasminogen activator (u-PA) was confined to milk leukocytes. Whey contained a component with a plasminogen dependent proteolytic activity which was shown to be plasma prekallikrein (PPK). The u-PA in the milk leukocytes was shown to be bound to urokinase receptor (u-PAR). A purification to near-homogeneity of the bovine u-PAR was undertaken. Investigating the novel t-PA binding to casein micelles by ligand blotting and Sepharose immobilized casein, multimeric forms of kappa-casein and dimeric alpha s2-casein were identified as t-PA binding components. The kappa-casein gene and the fibrinogen gene are believed to have evolved from a common ancestor. Thus, the recent finding that casein enhances t-PA catalyzed plasminogen activation (Marcus, G., Hitt, S., Harvey, S.R. and Tritsch, G.L. (1993) Fibrinolysis 7, 229-236), and the observed t-PA/casein binding suggests that the casein micelle, which also contains plasminogen, may serve as a matrix for t-PA-catalyzed plasminogen activation in milk.
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PMID:The plasminogen activation system in bovine milk: differential localization of tissue-type plasminogen activator and urokinase in milk fractions is caused by binding to casein and urokinase receptor. 818 64

Proteolytic joint destruction in inflammatory and non-inflammatory arthropathy is believed to be mediated, at least in part, by the plasminogen activation (PA) system. To further investigate possible involvement of the PA system, we quantified immunoreactive urokinase-type plasminogen activator (u-PA), tissue-type plasminogen activator (t-PA), both plasminogen activator inhibitors (PAI-1 and PAI-2) and u-PA-receptor (u-PAR) in synovial tissue extracts of 14 patients with rheumatoid arthritis (RA) and 12 with osteoarthritis (OA). u-PA, PAI-1, PAI-2 and u-PAR concentrations were significantly higher in RA than in OA patients. t-PA antigen levels were significantly lower in RA than in OA synovial tissue extracts. Immunohistochemistry was performed to compare the distribution and staining intensity of these components in samples of RA and OA synovial tissue. Intense immunostaining of u-PA, u-PAR, PAI-1 and, to a lesser degree, PAI-2 was observed predominantly in the synovial lining of RA patients. In OA patients, u-PA, PAI-1, PAI-2 and u-PAR were barely detectable. t-PA immunostaining was restricted to the endothelial side of vascular walls in both groups. We conclude that the observed increase of u-PA, u-PAR and PAI expression, distributed mainly in the synovial lining area of proliferative and invasively growing synovial tissue in RA patients, supports a pathogenic role for the PA system in destructive arthritis. Depressed t-PA-mediated plasminogen activation might contribute to delayed intra-articular fibrin removal.
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PMID:Difference in expression of the plasminogen activation system in synovial tissue of patients with rheumatoid arthritis and osteoarthritis. 864 30

We investigated the effect of heat shock on the fibrinolytic potential of human umbilical vein endothelial cells (HUVECs) in culture. When cultured at 43 degrees C, the mRNA for heat shock protein 70 (HSP70) was dramatically induced within 120 min with a maximal induction of more than 90-fold compared with that in HUVECs cultured at 37 degrees C. The level of urokinase-type plasminogen activator (u-PA) receptor (u-PAR) mRNA increased up to 2.2-fold in response to heat shock, which was associated with the increased u-PA binding and cell-surface u-PA activity determined by adding exogenous u-PA to acid-treated HUVECs. The increased u-PAR mRNA returned to normal level when HUVECs were further incubated at 37 degrees C for 180 min, and this decline was not affected in the presence of actinomycin D. Though the secreted antigens for tissue-type plasminogen activator (t-PA) and type 1 plasminogen activator inhibitor (PAI-1) in the conditioned medium (CM) of HUVECs were simultaneously increased at 43 degrees C during this period, the increase in the levels of t-PA (about 26.6-fold at 120 min) was greater than that of PAI-1 (1.8-fold at 120 min). The fibrinolytic activity of CM obtained from HUVECs at 43 degrees C was significantly enhanced up to 3-fold, indicating that heat shock induced hyperfibrinolytic states in HUVECs. The secretion of u-PA into CM was also enhanced by heat shock. These results suggested that human endothelial cells respond to hyperthermia by inducing HSP70 followed by hyperfibrinolytic states with the enhanced expression of u-PAR as well as that of t-PA and u-PA.
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PMID:Effect of heat shock on the expression of urokinase-type plasminogen activator receptor in human umbilical vein endothelial cells. 881 89

We demonstrated previously that tissue-type plasminogen activator (t-PA) bound to its specific receptor (t-PAR) on human umbilical vein endothelial cells (HUVEC) in suspension and that t-PAR of mol wt. 20 kDa interacted only with t-PA to form 90 kDa complex (Fukao, H., Hagiya, Y., Nonaka, T., Okada, K., and Matsuo, O. (1992) Biochem. Biophys. Res. Commun. 187, 956-962). In the present study, 20 kDa t-PAR was purified from HUVEC and the function of the t-PAR was investigated by analyzing its effect on plasminogen activation by t-PA. About 2.2 microg t-PAR protein was purified from cell lysate of 1.0 X 10(9) HUVEC as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) by gel filtration with TSK-3000SW and reversed phase separation with high performance liquid chromatography (HPLC). 125I-t-PA but not 125I-plasminogen specifically bound to the purified t-PAR in ligand blot assay. Plasminogen activation by t-PA in the presence of purified t-PAR in solution was increased. Furthermore, t-PA bound to immobilized t-PAR efficiently expressed its plasminogen activation activity. Kinetic analysis revealed that t-PA in the presence of soluble t-PAR and t-PA bound to immobilized t-PAR exhibited 34- and 90-fold increase in plasminogen activation, respectively. The t-PAR did not interact with anti-annexin II antibody. These findings indicate that the 20 kDa t-PAR is a novel molecule which immobilizes t-PA and enhances its proteolytic activity on the cell surface of endothelial cells.
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PMID:Enhancement of tissue-type plasminogen activator (t-PA) activity by purified t-PA receptor expressed in human endothelial cells. 909 97

During activation of the fibrinolytic system plasminogen is converted to plasmin by tissue plasminogen activator (t-PA) or urokinase-type plasminogen activator (u-PA). t-PA is predominantly released from endothelial cells, u-PA primarily by renal parenchymal cells. The activation of plasminogen is regulated by plasminogen activator inhibitor-1 (PAI-1), plasmin is controlled by alpha 2-plasmin inhibitor. The fibrinolytic system is not only involved in the intravascular dissolution of fibrin (thrombi), it also plays a vital role in normal physiologic reproduction, wound repair, angiogenesis, and tissue remodeling. Fibrinolysis is also a vital component in the pathogenesis of neoplastic disease. It is essential in releasing cells from their primary site of origin, providing nutrition for neoplastic cell growth and promoting cell mobility and motility. In neoplastic cells the degradation of the extracellular matrix proteins is facilitated by excessive expression of u-PA, t-PA, and u-PAR. In many forms of carcinoma increased expression of u-PAR and u-PA is associated with significantly shorter survival. Greater expression of u-PA in breast cancer cells, for example, is associated with shorter survival and increased relapse rate. Progressively aggressive neoplastic cells evidence high expression of u-PA and u-PAR activities, variable expression of t-PA, and enhanced PAI-1 and PAI-2 activities. In acute nonlymphocytic leukemias, poor outcome correlates with high t-PA levels. In acute progranulocytic leukemia there is a high incidence of DIC. Neoplastic prostatic tissue also expresses high u-PA activity and the more aggressive the cell line, the greater the number of u-PAR and the higher the u-PA activity. In gynecologic malignancies, a greater expression of u-PA in combination with cathepsin D is associated with widespread disease and poor prognosis. High u-PA values were also seen in patients with brain, gastric, and hepatic malignancies. It is evident that the plasminogen-plasmin system is a vital component in the biology of neoplastic disease and that it is, in theses conditions, in no way beneficial to the host.
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PMID:The fibrinolytic system in neoplasia. 912 11

The expression of urokinase-type plasminogen activator (u-PA), its receptor (u-PAR) and metalloproteases activity were analyzed in 4 human gastric-cancer cell lines (AGS, Hs746T, SNU-1, and SNU-5), in an attempt to relate these activities to their invasive potential and tumorigenicity on the modified chorioallantoic membranes (CAM) of chick embryos. Only 1 of the 4 cell lines tested, Hs746T, expressed both u-PA and u-PAR as well as MMP-2, but not MMP-9. This cell line was both tumorigenic and highly invasive (51.3 +/- 13.1%) on a modified CAM. Its invasive capacity was comparable with that of a highly malignant human epidermoid-carcinoma cell line (HEp3), which usually showed 40 to 50% invasiveness. The 3 other cell lines all produced MMP-2 and MMP-9, but only AGS showed moderate invasiveness (24.2 +/- 8.8%). While antibodies to u-PA were significantly effective in reducing CAM invasiveness of Hs746T cells by approximately 40%, the invasiveness of the t-PA-expressing AGS cell line was not affected by anti-t-PA antibodies. These results suggest that when one of the components of the u-PA/u-PAR system (the enzyme and/or the receptor) is not produced and u-PA/u-PAR-dependent cell-surface proteolytic activity is thereby diminished, the malignant phenotype that can be determined by tumorigenicity and invasion of connective tissue on a CAM is compromised. Production of both type-IV collagenases (MMP-2 and MMP-9) cannot offset this deficiency.
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PMID:Co-expression of urokinase-type plasminogen activator and its receptor in human gastric-cancer cell lines correlates with their invasiveness and tumorigenicity. 918 Jan 58

VEGF has been proposed to participate in normal and pathological vessel formation. Surprisingly, lack of only a single VEGF allele resulted in embryonic lethality due to abnormal formation of intra- and extra-embryonic vessels. Homozygous VEGF-deficient embryos, generated by tetraploid aggregation, revealed an even more severe defect in vessel formation. These results (1) suggest a tight regulation of early vessel development by VEGF and, indirectly, the presence of other VEGF-like molecules; (2) reveal an unprecedented lethal phenotype associated with heterozygous deficiency of an autosomal gene, and (3) demonstrate that tetraploid aggregation was a valid and the only method to study the phenotype of the homozyogous VEGF-deficient embryos. The dominant and strict dose-dependent role of VEGF in vivo renders this molecule a desirable therapeutic target for promoting or preventing angiogenesis. Tissue factor (TF) is the principal cellular initiator of coagulation and its deregulated expression has been related to thrombogenesis in sepsis, cancer, and inflammation. However, TF appears to be also involved in a variety of non-hemostatic functions including inflammation, cancer, brain function, immune response, and tumor-associated angiogenesis. Surprisingly, TF deficiency resulted in embryonic lethality due to abnormal extra-embryonic vessel development and defective vitelloembryonic circulation. The abnormal yolk sac vasculature is reminiscent of that observed in embryos lacking VEGF, possibly suggesting that both gene functions are interconnected. These targeting studies extend the recently documented role of TF in tumor-associated angiogenesis and warrant further study of its role in angiogenesis during other pathological disorders. The plasminogen system, via its triggers, tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) and its inhibitor, plasminogen activator inhibitor-1 (PAI-1), has been implicated in thrombosis, arterial neointima formation, and atherosclerosis. Studies in mice with targeted gene inactivation of t-PA, u-PA, PAI-1, the urokinase receptor (u-PAR), and plasminogen (Plg) revealed (1) that deficiency of t-PA or u-PA increase the susceptibility to thrombosis associated with inflammation and that combined deficiency of t-PA:u-PA or deficiency of Plg induces severe spontaneous thrombosis; (2) that vascular injury-induced neointima formation is reduced in mice lacking u-PA-mediated plasmin proteolysis, unaltered in t-PA- or u-PAR-deficient mice and accelerated in PAI-1-deficient mice, but that it can be reverted by adenoviral PAI-1 gene transfer; and (3) that atherosclerosis in mice doubly deficient in apolipoprotein E (apoE) and PAI-1 is reduced after 10 weeks of cholesterol-rich diet. Thus, the plasminogen system significantly affects thrombosis, restenosis, and atherosclerosis.
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PMID:Insights in vessel development and vascular disorders using targeted inactivation and transfer of vascular endothelial growth factor, the tissue factor receptor, and the plasminogen system. 918 98


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