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)

We carried out an immunohistochemical study of tissue-type plasminogen activator (PA) and urokinase-type PA, and their inhibitors, PA inhibitor-1 and PA inhibitor-2, using renal biopsy specimens obtained from 86 patients with various forms of glomerulonephritis. The controls were four normal renal tissue specimens. On immunofluorescent observation, granular staining for tissue-type PA was found to be distributed along the glomerular capillary walls. The fluorescence was weak in the normal renal tissue and occasionally intense in the tissues of patients with IgA nephritis, minimal change nephrotic syndrome, and lupus nephritis. PA inhibitor-1 was abundant in the glomerular epithelial cells and scarce in the mesangial area and glomerular capillary lumens of the normal renal tissues. This was confirmed by immunoelectron microscopy using gold staining. The fluorescence of PA inhibitor-1 was weaker in some specimens of nephritic tissues than in the normal renal tissues. Urokinase-type PA and PA inhibitor-2 were negative within the glomeruli in all the specimens. In the glomerulonephritic tissues which were fibrin deposition-positive, tissue-type PA expression in the glomeruli tended to be strong. An association between fibrin deposition and PA inhibitor-1 staining was not clear. These data suggest that expression of tissue-type PA in the glomeruli increases in association with fibrin deposition.
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PMID:Tissue-type plasminogen activator and its inhibitor in human glomerulonephritis. 138 27

Plasminogen activators (PAs) and/or plasmin may be involved in hematopoietic regulation. These enzymes release biologically relevant cytokines such as basic fibroblast growth factor (bFGF) from matrix and cell surfaces. In addition, transforming growth factor beta (TGF beta) and interleukin-1 beta (IL-1 beta) are converted from inactive to active forms by plasmin. Therefore, we studied the regulation of PAs and their specific inhibitors, PA inhibitor 1 (PAI-1) and PA inhibitor 2 (PAI-2), in human bone marrow stromal fibroblasts by IL-1 beta, bFGF, and TGF beta. All three cytokines stimulated PA secretion. IL-1 beta at 10(4) U/mL increased urokinase (u-PA) levels approximately 10-fold, bFGF at 0.2 ng/mL also increased production 10-fold, but increased predominantly tissue PA (t-PA) expression. TGF beta at 0.2 ng/mL increased u-PA production up to 300-fold. PAI-1 and PAI-2 are also regulated by these cytokines. IL-1 beta decreased PAI-1 levels by 50% and stimulated PAI-2 levels sixfold. bFGF had minimal effects on PAI-1 and TGF beta increased PAI-1 levels twofold. Neither of these agents had an effect on PAI-2 levels. Thus, three cytokines relevant to bone marrow physiology regulate PA and inhibitor production by human bone marrow stromal fibroblasts. In this manner PA and plasmin generation in specific microenvironments in the bone marrow may be one of the factors orchestrating the complex series of events, which results in an efficient exquisitely regulated hematopoietic process.
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PMID:Regulation of proteolytic activity in human bone marrow stromal cells by basic fibroblast growth factor, interleukin-1, and transforming growth factor beta. 153 45

Plasminogen activator inhibitor-2 (PAI-2), found in human placenta and pregnancy plasma, was prepared in a highly purified and functionally active form from human placenta. The purification was achieved by a combination of Rivanol and ammonium sulfate precipitation, followed by chromatography on DEAE Affigel Blue, hydroxylapatite and phenylalanine-Sepharose. PAI-2, which is precipitated by low Rivanol concentrations, can be selectively redissolved from the pellet by increasing the Rivanol concentration in the presence of a reducing agent, i.e. dithiothreitol. The purified protein shows a molecular mass of 45 kDa in SDS PAGE, cross-reacts with monoclonal antibodies against PAI-2 (Mab'PAI-2), and inhibits the amidolytic activity of urokinase-type plasminogen activator (u-PA) towards the chromogenic substrate Glu-Gly-Arg-pNA (S-2444). The specific activity of the purified inhibitor was 52,300 units/mg, attaining 71,000 units/mg in peak fractions. In the immunopurification of placental extract on anti-PAI-2 Sepharose, the eluate showed the expected reaction with Mab' PAI-2, and it also cross-reacted with anti-vitronectin serum. In order to complement these results, anti-vitronectin Sepharose was used for immunopurification of placenta extract. In Western Blot experiments the eluates of anti PAI-2 Sepharose and anti-vitronectin Sepharose both showed a heterogeneous pattern of high molecular weight bands recognized by either polyclonal antiserum against vitronectin or Mab'PAI-2. In either case, reduction of the eluates releases mainly a 45-kDa band, which is recognized by Mab'PAI-2, or 80-kDa and 76-kDa bands recognized by anti-serum against vitronectin. These data suggest that the predominant form of PAI-2 in placenta extract is heterogeneous and of high molecular mass, containing complexes in which vitronectin is covalently bound to PAI-2 by disulfide bridges.
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PMID:Isolation of plasminogen activator inhibitor-2 (PAI-2) from human placenta. Evidence for vitronectin/PAI-2 complexes in human placenta extract. 170 70

Expression of plasminogen activator (PA) enzyme activity is believed to be one of the mechanisms by which malignant cells cause pericellular proteolysis of stromal structures during implantation and tissue invasion. In this study, four cell lines derived from human gliomas were studied to ascertain which PA enzymes and PA inhibitors determine the level of secreted PA activity. A plasminogen-dependent esterolytic assay was used, and two lines (U251 and U373) were found to secrete high levels of PA activity, while PA activity was undetectable in the conditioned media from the remaining two lines (U138 and LM). The PA produced by U251 and U373 resolved as single bands comigrating with high molecular weight urokinase (Mr 54,000) on casein-plasminogen zymography. Northern blot analysis demonstrated high levels of mRNA for urokinase-type PA (uPA) in U251 and U373, as well as a considerably lower level of uPA message in LM. U251 and U373 also contained mRNA for tissue-type PA (tPA), although secreted tPA activity was not demonstrated by zymography. The U138 line contained essentially undetectable levels of mRNA for either uPA or tPA. U138 was also unique in secreting PA inhibitor activity and contained high levels of mRNA for PA inhibitor 2, which was not seen in any other line. All cell lines contained PA inhibitor 1 mRNA, with substantially more expression in the U138 and LM lines than in U251 and U373. None of the lines secreted measureable anti-plasmin activity. We conclude that there is considerable heterogeneity among human glioma cells in expression of PA enzymes and PA inhibitors. The coordinated regulation of these proteins likely determines secreted PA activity and the resultant role of plasminogen activation in tumor implantation and invasion.
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PMID:Expression of heterogeneous profiles of plasminogen activators and plasminogen activator inhibitors by human glioma lines. 237 61

To investigate the mechanisms by which cutaneous plasminogen activator (PA) may be regulated, we have tested cultured keratinocytes for the presence of PA inhibitors. Using biosynthetic labeling experiments with 35S-methionine in conjunction with specific antibody precipitation, we have shown that human keratinocytes in culture synthesized and secreted both PA inhibitor 1 and PA inhibitor 2. PA inhibitor 1 was present in conditioned media in the inactive form, but it could be detected with reverse phase autography. PA inhibitor 2 was detected by its ability to form complexes with 125I-uPA. Potential therapeutic relevance for cutaneous PA inhibitor 2 was suggested in skin organ culture experiments which demonstrated that purified PA inhibitor 2 from human placenta was able to prevent the acantholytic changes induced by pemphigus IgG.
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PMID:Characterization of keratinocyte plasminogen activator inhibitors and demonstration of the prevention of pemphigus IgG-induced acantholysis by a purified plasminogen activator inhibitor. 246 56

Plasminogen activator inhibitor-2 (PAI-2) can regulate the formation of plasmin by inhibiting urokinase and tissue plasminogen activator. PAI-2 is induced in monocytes and endothelium by inflammatory mediators, and it is made in the placenta during pregnancy. PAI-2 is a member of the serine protease inhibitor gene family, and it is particularly similar to chicken ovalbumin. Like ovalbumin, PAI-2 is secreted without cleavage of a signal peptide. To determine the structure of the PAI-2 gene, two bacteriophage lambda human genomic DNA libraries were screened with PAI-2 cDNA probes. Characterization of three positive clones shows that the human PAI-2 gene spans 16.5 kilobases and has eight exons. The 5'-untranslated sequence of the PAI-2 mRNA is 77 base pairs in length as suggested by primer extension and S1 nuclease mapping. The eukaryotic consensus sequence TATAAAA is found 22 base pairs 5' of the proposed cap site. The PAI-2 gene is on chromosome 18q21-23 as determined by hybridization to flow-sorted chromosomes and by in situ hybridization. There appear to be two common PAI-2 alleles that differ by six nucleotides in exons 1, 4, and 8. The structure of the PAI-2 gene is quite different from that of PAI-1 although these two inhibitors have common target protease specificity. In contrast, the structure of the PAI-2 gene is very similar to that of the chicken ovalbumin gene. When protein sequences are aligned to obtain maximal identity, six of the seven intron positions in the PAI-2 gene are identical to those in the chicken ovalbumin gene. We conclude that PAI-2 is the closest mammalian homologue of avian ovalbumin.
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PMID:Structure of the gene for human plasminogen activator inhibitor-2. The nearest mammalian homologue of chicken ovalbumin. 249 65

Macrophages have a marked capacity to invade tissue in the course of cellular immune reactions that is thought to be based on the action of urokinase (u-PA). u-PA is an ubiquitous serine protease that converts the zymogen plasminogen into the active protease plasmin. u-PA binds to specific receptors on the macrophage thereby enabling the cell to degrade interstitial tissue in the microenvironment. Two cytokines produced in the course of cellular immune reactions, IFN-gamma and TNF-alpha, increase the number of u-PA receptors on human cultured monocytes from 14,000 to 64,000 and 30,000 receptors/cell, respectively. We used an amnion invasion assay to investigate whether activated human monocytes exhibit an enhanced capacity to invade interstitial tissue in correlation to the increased numbers of u-PA receptors. We show in this study that IFN-gamma, which increases the number of endogenously occupied and saturable u-PA receptors, causes a threefold increase of monocyte invasion into amnion tissue in comparison to control cells. The anti-u-PA mAb MPW5UK, which blocks the activity of u-PA, inhibits monocyte invasiveness significantly. In contrast, TNF-alpha, which increases only the number of saturable u-PA receptors on monocytes, does not enhance their invasiveness. This finding suggests that only endogenously occupied u-PA receptors are instrumental in monocyte invasiveness. This conclusion is further supported by the findings that: 1) saturation of monocytes with u-PA does not further increase their invasiveness and that 2) plasminogen-activator inhibitor-2, a specific inhibitor of u-PA associated with endogenously occupied, but not of u-PA bound to saturable receptors, inhibits monocyte invasiveness completely.
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PMID:Endogenous receptor-bound urokinase mediates tissue invasion of human monocytes. 255 65

Plasminogen activator inhibitor-2 (PAI-2) is a serine protease inhibitor that regulates plasmin generation by inhibiting urokinase and tissue plasminogen activator. The primary structure of PAI-2 suggests that it may be secreted without cleavage of a single peptide. To confirm this hypothesis we have studied the glycosylation and secretion of PAI-2 in human monocytic U-937 cells by metabolic labeling, immunoprecipitation, glycosidase digestion, and protein sequencing. PAI-2 is variably glycosylated on asparagine residues to yield intracellular intermediates with zero, one, two, or three high mannose-type oligosaccharide units. Secretion of the N-glycosylated species began by 1 h of chase and the secreted molecules contained both complex-type N-linked and O-linked oligosaccharides. Enzymatically deglycosylated PAI-2 had an electrophoretic mobility identical to that of the nonglycosylated precursor and also to that of PAI-2 synthesized in vitro in a rabbit reticulocyte lysate from synthetic mRNA derived from full length PAI-2 cDNA. The amino-terminal protein sequence of secreted PAI-2 began with the initiator methionine residue. These results indicate that PAI-2 is glycosylated and secreted efficiently without the cleavage of a signal peptide. PAI-2 shares this property with its nearest homologue in the serine protease inhibitor family, chicken ovalbumin, and appears to be the first well characterized example of this phenomenon among natural mammalian proteins.
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PMID:Mammalian protein secretion without signal peptide removal. Biosynthesis of plasminogen activator inhibitor-2 in U-937 cells. 312 94

Baseline cellular plasminogen activator (PA) activity, and the cellular proteins responsible for variations in PA activity were evaluated in three human prostate carcinoma cell lines. Net PA activity in the cell lines PC-3, DU-145, and LNCaP was measured using a plasminogen-dependent fibrin lysis assay. These three cell lines were then analyzed to determine the specific protein(s) responsible for differences in PA activity. mRNA and protein levels of cellular urinary PA (uPA), tissue PA (tPA), PA inhibitor 1 (PAI1), PA inhibitor 2 (PAI2), and uPA receptor (uPAr) were measured using Northern analysis and ELISA assays. Net cellular PA activity in the three cell lines varied over a 3-fold range (PC3 > DU145 >> LNCaP). Net PA activity in the fibrinolysis assay demonstrated a direct correlation with mRNA transcript levels of uPA, tPA, PAI1, and uPAr (PC-3 > DU-145 > LNCaP). uPA protein was identified in both the PC-3 and the DU-145 lines. tPA, PAI1, and PAI2 proteins were identified only in PC-3 cells. In general, cellular protein levels correlated with mRNA levels. These findings demonstrate that prostate carcinoma cell lines vary in their net PA activity. This variability results from both qualitative and quantitative differences in the cellular expression of PA regulatory proteins.
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PMID:Diversity and modulation of plasminogen activator activity in human prostate carcinoma cell lines. 747 84

Effects of extracellular matrices (ECM) and the plasminogen activator (PA) system on outgrowth of sheep inner cell masses (ICM) and trophectoderm in vitro were investigated. Experiment 1 evaluated the effects of plasminogen and ECM type on ICM and trophectodermal outgrowth, on glass Lab-Tek chamber slides coated with collagen IV, fibronectin, or laminin. ICM outgrowth areas were reduced (p < 0.05) by plasminogen and were greatest (p < 0.05) on fibronectin. Trophectodermal outgrowth was not supported in this system. Experiment 2 evaluated the effects of PA inhibitor-2 (PAI-2) or antiserum to urokinase-type PA (anti-uPA) on ICM outgrowth on fibronectin. Numbers of cells in the outgrowths were increased (p < 0.05) with PAI-2, and anti-uPA had no effect (p > 0.10). Experiment 3 evaluated the relationship between PA production and ECM type on ICM and trophectodermal outgrowth in microdrop cultures. PA production by ICM was greatest (p < 0.05) on fibronectin, but no differences (p > 0.10) were observed for trophectoderm. PA production was not correlated with ICM outgrowth areas (r = -0.12; p = 0.72) or numbers of cells in the ICM outgrowths (r = 0.09; p = 0.74) but was correlated with ICM areas (r = 0.75; p < 0.01) and numbers of cells in trophectodermal outgrowths (r = 0.57; p = 0.01). These results suggest that type of ECM, culture system, and alterations in the PA system influence cellular outgrowths by ICM and trophectoderm.
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PMID:Evaluation of extracellular matrices and the plasminogen activator system in sheep inner cell mass and trophectodermal outgrowth in vitro. 763 51


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