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)

Biopsies of involved and uninvolved skin from psoriatic patients and of normal skin were stained immunocytochemically with monoclonal antibodies against urokinase-type (u-PA) and tissue-type (t-PA) plasminogen activator using a multilayer peroxidase technique. Epidermis from psoriatic lesions showed focal staining for u-PA in and between the basal keratinocytes in the suprapapillary epidermal areas, while t-PA was found in the superficial keratinizing cells, including both stratum spinosum and the parakeratotic layer. No staining of keratinocytes was observed in uninvolved and normal skin. The specificity of the staining was supported by the finding that 3 different monoclonal antibodies and polyclonal antibodies against each of the plasminogen activators gave identical staining, while monoclonal antibodies of irrelevant specificity gave no staining. The present findings suggest abnormalities in the regulation of both types of plasminogen activators in psoriatic epidermis.
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PMID:Immunohistochemical localization of urokinase- and tissue-type plasminogen activators in psoriatic skin. 309 60

An enzyme-linked immunosorbent assay (ELISA) for the measurement of urokinase-type plasminogen activator (u-PA) was developed. Three murine monoclonal antibodies to single chain urokinase-type plasminogen activator (scu-PA) were isolated and shown to react with non-overlapping epitopes in scu-PA. Two of the three antibodies were coated onto microtiter plates and bound u-PA was quantitated with the third antibody conjugated to horseradish peroxidase. The assay was equally sensitive to Mr 54,000 u-PA in the single chain or two-chain form but did not respond to Mr 33,000 urokinase. The lower limit of sensitivity of the assay was 0.1 ng/ml in buffer and 1 ng/ml in plasma. Coefficients of variation of the assay at physiological levels of u-PA were 6.5 percent within assays and 13 percent between assays. The level of u-PA in normal resting plasma was 1.9 +/- 0.66 ng/ml (mean +/- SD, n = 54). The assay can be performed within one working day and provides an efficient, reproducible, and stable means for the measurement of u-PA in biological fluids. As such it may facilitate physiological and pharmacological studies of urokinase-type plasminogen activators in man.
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PMID:Measurement of urokinase-type plasminogen activator (u-PA) with an enzyme-linked immunosorbent assay (ELISA) based on three murine monoclonal antibodies. 310 11

An enzyme-linked immunosorbent assay (ELISA) was developed for the measurement of human urokinase-type plasminogen activator (u-PA) in plasma and serum. Microtiter plates were coated with a monoclonal antibody and incubated with standard or sample. Bound u-PA was quantitated with polyclonal antibodies conjugated with biotin, followed by avidin-peroxidase. The assay was 10 times as sensitive as previously reported immunoassays, the detection limit being approximately 1 pg u-PA in a volume of 100 microliter, with a linear dose-response up to 15 pg u-PA. The assay detected active u-PA and its inactive proenzyme form equally well, and the recovery of both forms was higher than 90% in plasma. It also detected u-PA complexed with plasminogen activator inhibitor type 1. Various structurally related proteins, including t-PA, were tested, but no reaction was observed with proteins other than u-PA and its amino-terminal fragment. The intra-assay and interassay coefficients of variation for determination of u-PA in plasma were 7.6% and 8.4%, respectively. The ELISA was used to measure the concentration of u-PA in plasma from 34 healthy donors and 92 patients with breast cancer with a varying extent of disease. The mean value for the healthy donors was 1.1 +/- 0.3 ng/ml (SD) of u-PA in plasma. This value is substantially lower than those previously reported. The mean value for the patients with breast cancer was 1.3 +/- 0.4 ng/ml. This moderate increase was statistically significant at the 1% level. Approximately one quarter of the patients had plasma u-PA concentrations above the range observed for the healthy controls. There was a positive correlation between the mean u-PA plasma concentration and the extent of disease in different groups of patients.
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PMID:Sensitive and specific enzyme-linked immunosorbent assay for urokinase-type plasminogen activator and its application to plasma from patients with breast cancer. 312 72

The immunoperoxidase technique, using antibodies against human urinary urokinase (Mr 55,000), was used for the localization of this enzyme in histological preparations of human colon tumors and normal colon tissue. The localization of tissue (vascular) activator was also investigated using antibodies against enzyme purified from human malignant melanoma. Both the "indirect method" and the peroxidase-antiperoxidase technique were found to be useful. Urokinase-reactive material was found in all tissues examined (33 primary cancers, 11 metastases, and 8 adenomas). In the normal colon, urokinase was found only in some of the goblet cells of the mucosal epithelium. In colon cancer, diffuse specific staining was observed in the cytoplasm, but the most intense staining was localized at the edge of the cancer cells bordering the lumen of the glands. In some cases, intense supranuclear staining could be observed in a location corresponding to the Golgi apparatus. In a few instances, urokinase could be seen associated with fibroblasts near the advancing front of an invading tumor. Adenoma, a benign tumor but often a precursor of cancer, also showed the presence of urokinase. Most significant were the observations showing that, in regions of the mucosal glands where normal epithelial cells were abruptly replaced by cancer cells, the appearance of cytoplasmic urokinase showed strict and exclusive association with the malignant cells, and the same was the case in transitions from normal epithelium to adenoma. In contrast to urokinase, tissue plasminogen activator was not associated with cancer cells, but was consistently present in the stroma which separates the cancer glands and was localized in the endothelium of the blood vessels. This visual evidence was supported by results of extraction of plasminogen activators from tumors, and from the separated mucosal and submucosal layers of the normal colon of the same patients, which showed that urokinase is most abundant in the tumor tissue and least abundant in the submucosa, while tissue activator is most prevalent in the well-vascularized mucosa and submucosa and scarce in the usually poorly vascularized adenocarcinomas.
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PMID:Localization of plasminogen activators in human colon cancer by immunoperoxidase staining. 388 45

The subcellular localization of glandular kallikrein in ducts and tubules of the rat submandibular gland was determined using a postembedding immunostaining (peroxidase--antiperoxidase) technique on thin sections of Eponembedded tissue. Kallikrein was found in large granules of granular convoluted tubule cells and in small, apical granules of striated duct cells. It was also present in patchy aggregates along the surface of striated duct cells and intercalated duct cells, but not in granules of the latter. Basal dense bodies (lysosomes?) of granular tubules also stained for kallikrein. Absorption of kallikrein antiserum with rat urinary kallikrein, but not with rat urinary esterase A, abolished the specific immunostaining in these sites.
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PMID:Electron microscopic immunostaining of kallikrein in rat submandibular glands. 633 7

The immobilization of some medically useful enzymes were studied by means of radiation-induced polymerization at -78 degrees C. Glucose oxidase and glucose peroxidase were immobilized in the form of thin membranes inside polyvinyl chloride tubes and on polyethylene films; these membranes showed considerable activity yield, as well as good activity retention. Two effective methods were adopted to improve the surface properties of the base materials and to facilitate firm immobilization by coating: that is, an undercoating method followed by radiation curing of the undercoating and an irradiation grafting method with a monomer. Both were tested with good results. An immobilization of urokinase was also carried out successfully by similar methods. The thrombogenicity of the immobilized urokinase showed a remarkable effect on thombus formation.
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PMID:Immobilization of enzymes for medical uses on plastic surfaces by radiation-induced polymerization at low temperatures. 736 85

The serine protease inhibitor protein C inhibitor is present in semen at a relatively high concentration and forms in vivo complexes with two plasminogen activators also present in semen, urokinase-type and tissue-type plasminogen activators. Therefore, the fact that prostate-specific antigen (PSA), a major prostate enzyme, complexes and inactivates protein C inhibitor (PCI) in semen could have implications in human reproduction. The present study was undertaken to develop an enzyme-linked immunosorbent assay (ELISA) for complexes of PSA with PCI (PSA:PCI) with purified PSA:PCI complexes as a standard. Seminal plasma was utilized as the starting material for purification of complexes by affinity chromatography on heparin-Sepharose and gel filtration. The final preparation contained equimolar concentrations of PSA and PCI and was used for calibration of an ELISA for PSA:PCI complexes involving polyclonal anti-PSA and horseradish peroxidase-labeled anti-PCI antibodies. The ELISA had a detection limit of about 0.2 ng/ml of complex and was specific for PSA:PCI complexes because no color was developed at PSA or PCI concentrations up to 100 microgram/ml. Normal plasma or plasma from patients with prostate carcinoma who had high PSA levels had no detectable PSA:PCI complexes. Seminal plasma from voluntary donors collected in the absence of inhibitors and incubated at room temperature for at least 3 hours had PSA:PCI complex levels ranging from 30 to 46 micrograms/ml, accounting for up to 34% of the total PCI in seminal plasma.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A quantitative ELISA for the measurement of complexes of prostate-specific antigen with protein C inhibitor when using a purified standard. 750 42

The enzymatic activities of uPA, and a collagenase-like proteinase in the post-nuclear fraction of cell homogenates of a metastatic carcinomatous cell line following X-ray irradiation were examined by the use of chromogenic substrates and by casein- or gelatin-containing zymographies and electrophoretic gel stained with avidin-conjugated peroxidase. Enhanced activities were observed in these cells, while those of 5'nucleotidase and Na(+)-K(+)-ATPase were attenuated. A partial purification and characterization of the collagenase showed that it was able to hydrolyze the heat-denatured type-I collagen more efficiently than the native one. The activation of both uPA and collagenase enables an efficient degradation of matrix barrier proteins. These findings suggest that following a certain dose range of X-ray irradiation, tumor cells may increase their ability to migrate and invade through the enhancement of uPA and collagenase activities.
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PMID:The concomitant augmentation of urokinase-type plasminogen activator and collagenase-like proteinase activities in X-ray irradiated cells of a human metastatic carcinomatous line. 809

Two immunoassays for quantitation of the biological markers uPA and PAI-1 were evaluated for their use with detergent extracts of breast cancer tissue. Both assays were based on murine monoclonal capture antibodies and rabbit polyclonal detector antibodies. Horseradish peroxidase-conjugated goat anti-rabbit antibodies enabled measurement of the bound antigen. The detection limit of the uPA assay was 13 pg/ml, with a linear dose-response relationship up to 350 pg/ml. The assay detected free uPA as well as uPA in complex with PAI-1 and/or with its receptor. The detection limit of the PAI-1 assay was 50 pg/ml, with a linear dose-response relationship up to 1500 pg/ml. The assay detected both free PAI-1 and uPA:PAI-1 complex. Both assays were validated for detergent extracts using immunoabsorption and recovery tests. Highly significant associations between tumour tissue uPA and PAI-1 levels and prognosis were verified in a cohort of 164 lymph node-negative primary breast cancer patients. It is concluded that the two immunoassays are well-suited for the quantitation of uPA and PAI-1 in detergent extracts of breast cancer tissues.
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PMID:Immunoassays of urokinase (uPA) and its type-1 inhibitor (PAI-1) in detergent extracts of breast cancer tissue. 1270 58

A water-soluble, ligand-bound polymer has been synthesized for the purpose of isolation of urokinase, an important plasminogen activator. The affinity polymer was formed by copolymerizing N-acryloyl-m-aminobenza-midicine and acrylamide in the absence of oxygen. An affinity ultrafiltration process was then developed for isolating urokinase from an artificial solution containing peroxidase and urokinase and from a crude urine source. The process yields were determined to be 86% and 49%, respectively. The recovered urokinase exhibited a specific activity close to that of the highest commercial grade. This article also presents a new technique for assaying urokinase by coupling plasminogen with L-benzoyl arginine-p-nitroanilide (L-BAPNA), an inexpensive chromogenic substrate.
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PMID:Isolation of urokinase by affinity ultrafiltration. 1858 35


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