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)

A 64-year-old male developed pain and marked swelling of his left calf muscle in September, 1989. Deep-vein thrombophlebitis was suspected, and therapy with warfarin sodium followed by urokinase was instituted. Because of unsatisfactory effect of the therapy, the patient was referred to another hospital on January 3rd 1990. A CT scan of the left leg showed a soft-tissue mass adjacent to the fibula. A biopsy of the mass revealed non-Hodgkin's lymphoma of diffuse medium-sized cell type. Immunohistologic examination confirmed B cell type. He was admitted to our hospital on January 31st 1990. A chest roentgenogram showed right pleural effusion. A CT scan of the abdomen revealed a space-occupying lesion in the liver. A Gallium scintigraphy showed markedly increased isotope uptake in the left calf and liver. He was classed as a Stage IV B. He responded well to combination chemotherapy with cyclophosphamide, THP-adriamycin, VP-16, and prednisolone and achieved a complete remission. Although prominent infiltration of lymphoma cells in skeletal muscles is rarely reported, it is important to perform the biopsy promptly when the mass is found in a muscle. In this case report, we describe a rare case of non-Hodgkin's lymphoma with muscle invasion presented as marked calf muscle swelling.
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PMID:[Non-Hodgkin's lymphoma with marked infiltration in calf muscle]. 202 Jan 18

The aim of this study was to assess the evolution of ST segment displacement and the formation of new Q waves in relation to the late angiographic demonstration of recanalization of the infarct-related vessel in 36 patients with acute myocardial infarction admitted in our Institution within 8 hours from the onset of symptoms. Eighteen patients were treated with intravenous urokinase (200,000 + 1,200,000 UI) followed by sodium-heparin 1,000 U/ev/hour for 48 hours and calcium-heparin 3,000 UI/10 Kg/12 hours from the third day to discharge. Eighteen patients were treated from admission only with calcium-heparin as seen before. All the patients have been followed with serial electrocardiograms; we evaluated the sum of ST displacements (ST12) and Q waves score (Q12) in the 12 standard leads on admission, and on the twelfth, twenty-fourth, forty-eighth hour, and on discharge. On the basis of the late angiographic findings and apart from the treatment, we divided the patients into 2 groups: patients with recanalization of infarct-related vessel (20 patients, 55.5%) and patients with occlusion of the infarct-related vessel (16 patients, 44.6%). We did not observe statistically significant differences between the 2 groups for decrease of ST segment displacement of formation of new Q waves, even if reperfused patients showed a more rapid and complete decrease of ST segment displacement and a more rapid formation of new Q waves with respect to occluded patients. Our data indicate that from a clinical point of view electrocardiographic indices are not a sensitive marker of recanalization of infarct-related vessel.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:[Electrocardiographic indices of reperfusion and limitation of the necrotic area in acute myocardial infarct]. 209 24

Using site-directed mutagenesis, we have changed the asparagine in human single-chain urinary plasminogen activator (u-PA) at position 302 to an alanine. This alteration removes the only known amino acid residue glycosylated in the protein. The single-chain u-PA containing an alanine residue at position 302 instead of asparagine (scu-PA(N302A] cDNA gene was expressed in the yeast Saccharomyces cerevisiae. Secretion of the protein product into the culture broth was achieved by replacing the human secretion signal codons with those from yeast invertase, adding a yeast promoter from the constitutively expressed glycolytic genes triosephosphate isomerase or phosphoglycerate kinase, and integrating multiple copies of these transcriptional units into the genome of yeast strains carrying the "supersecreting" mutation ssc1. When fermented in a fed-batch mode, these recombinant baker's yeast strains secreted scu-PA(N302A) in a strongly growth-associated manner. Greater than 90% of the u-PA found in the culture broth was in the single-chain form. Scu-PA(N302A) was purified to homogeneity using two chromatography steps. The purified protein had a molecular weight of 47,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and lacked any detectable N-linked glycosylation. The in vitro fibrinolytic properties of scu-PA(N302A) were found to be essentially equivalent to those of natural single-chain u-PA derived from the human kidney cell line TCL-598. Since scu-PA(N302A) lacks the immunogenic N-linked carbohydrate pattern of yeast, it may be a useful therapeutic agent which can be produced economically by yeast fermentation.
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PMID:Characterization of a nonglycosylated single chain urinary plasminogen activator secreted from yeast. 210 31

Single-chain urokinase plasminogen activator (scu-PA) that had been modified with N-succinimidyl-3-(2-pyridyldithio)propionate was covalently linked by disulfide bonds to the Fab' of a monoclonal antibody specific for the beta-chain of fibrin (antibody 59D8). scu-PA-59D8 Fab' conjugate was separated from free scu-PA and two-chain urokinase coupled to 59D8 Fab' by two-step affinity chromatography. First, the reaction mixture was chromatographed on a column containing Sepharose linked to the peptide that had been used as immunogen for antibody 59D8; scu-PA-59D8 Fab' conjugate and unconjugated 59D8 Fab' were retained but not unconjugated scu-PA. Then, the eluate from the peptide-Sepharose column was chromatographed on a column containing Sepharose linked to benzamidine, which acts as a ligand for two-chain urokinase. The molecular weight of the scu-PA-59D8 Fab' conjugate was approximately 100 kDa when electrophoresed on a nonreducing sodium dodecylsulfate-polyacrylamide gel. Enzymatic assay after purification revealed that more than 97% of the scu-PA present in the conjugate retained the single-chain form. The Fab' portion of the conjugate functioned in a manner indistinguishable from that of native antibody 59D8. In an in vitro assay for lysis of fibrin monomer, the fibrinolytic potency of scu-PA-59D8 Fab' was 33-fold more than that of tissue plasminogen activator (p less than 0.001), 230-fold more than that of unconjugated scu-PA (p less than 0.0001), and 420-fold more than that of urokinase (p less than 0.0001).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Conjugation to antifibrin Fab' enhances fibrinolytic potency of single-chain urokinase plasminogen activator. 211 44

ES-1 cells, which showed a higher sensitivity to the cytocidal action of estradiol were isolated from a human breast cancer MCF-7 cell line. Growth of ES-1 cells was inhibited by a dose of 17-beta estradiol that stimulated the growth of the parental MCF-7 cells. Proteins secreted from MCF-7 and ES-1 cells when cultured with 17-beta estradiol were compared by sodium dodecyl sulfate-containing polyacrylamide gel electrophoresis (SDS-PAGE). Addition of estradiol to culture medium enhanced secretion of a protein of molecular mass of 52 kDa in media for both MCF-7 and ES-1 cell lines, but the secretion of a second 67 kDa protein was enhanced about 10-fold only in ES-1 cells. The analysis by SDS-PAGE of culture medium immunoprecipitated with anti-tissue-type plasminogen activator (t-PA) antibody demonstrated that the band of 67 kDa protein specifically secreted from estradiol-treated ES-1 cells contained t-PA. Zymography assays, quantitative immunoreactive assays, and Northern analysis showed about 5-fold specific increase by estradiol of t-PA with molecular mass of 65-70 kDa in ES-1 but not in its parental MCF-7 cells. Cellular level of the plasminogen activity was also specifically enhanced in ES-1 cells by estradiol, but only a slightly in MCF-7 cells. By contrast, another urokinase-type PA (u-PA) with molecular weight of 55 kDa showed very low level activity in both MCF-7 and ES-1 cell lines in the presence of estradiol. Formation of t-PA mRNA was specifically enhanced in ES-1 cells when ES-1 cells were treated for more than 12 h with 10(-8) M 17-beta estradiol. Estradiol did not elongate the lifetime of t-PA mRNA in ES-1 cells. A unique phenotype of ES-1 cells in response to estradiol is discussed in relation to activating expression of the t-PA gene.
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PMID:Enhanced production of tissue-type plasminogen activator by estradiol in a novel type variant of human breast cancer MCF-7 cell line. 211 58

Increasing attention is being paid to alterations of the hemostatic balance in tumors, in general, and brain tumors, in particular. Apparently divergent results, showing excess fibrinolysis (i.e., increased plasminogen activator activity) or its inhibition (i.e., increased inhibitor activity), have been reported. The 9L rat brain tumor is a gliosarcoma and a model used to study treatment paradigms for human gliomas. To study the roles of fibrin and fibrinolysis in this brain tumor model, we used these features to investigate the nature of the plasminogen activator (PA) and thrombin inhibitors in normal rat brain and in the 9L rat brain tumor, growing both in vitro and in vivo in rat brain. The results indicate that cells cultured from the tumor in vitro express PA inhibitory activity which is both of the protease nexin I and PA inhibitor 1 types. However, the serpin PA inhibitory activity in extracts of both the normal brain and tumor is of the protease nexin I/PA inhibitor 3 type. This activity is higher in the tumor than in the surrounding "normal" tissue. In addition, we present evidence for a novel thrombin inhibitor which (a) is present only in the tumor growing in rat brain and undetectable either in the normal brain tissue or in vitro, (b) is in a latent, but sodium dodecyl sulfate-activatable, state, and (c) does not bind urokinase. In current studies, investigators are exploring the roles of these molecules and the target serine proteases they inhibit in the pathogenesis of gliomas.
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PMID:Serpin inhibitors of urokinase and thrombin in normal rat brain and the 9L brain tumor: evidence for elevated expression of protease nexin I-like inhibitor and a novel sodium dodecyl sulfate-activated tumor antithrombin. 211 23

Changes in plasminogen activator (PA) and PA inhibitor (PAI) activities were measured during follicular development in granulosa cells (GC) and theca tissue (TT) isolated from the six largest yolk-filled preovulatory follicles (F1, F2, F3, F4, F5, F6) and large white follicles (LWF) of the domestic hen. PA activity increased and PAI activity decreased during follicular development, with the peak PA value and minimum activity for PAI observed in the largest preovulatory follicle (F1) 12-14 h before expected time of ovulation. The PA activity in GC and TT appears to be principally of the tissue (t)-PA type judging from its substrate specificity and biochemical characteristics. The enzyme cleaved the chromogenic substrate specific for t-PA (Spectrozyme TM t-PA; CH3SO2-D-CHT-Gly-Arg-p-nitroanilide) more efficiently (4-6 x) than that for u-PA (Spectrozyme TM UK; Cbo-L-Glu-(alpha-t-BuO)-Gly-Arg-p-nitroanilide), suggesting that t-PA may be the predominant PA in the chicken preovulatory follicle. Determination of PA activity following sodium dodecyl sulphate-polyacrylamide gel electrophoresis and isoelectric focussing suggested the presence of two forms of the enzyme in GC and TT. The predominant form of PA had a molecular weight of 75,000 and an isoelectric point (pI) of 7.7, characteristics similar to those reported for t-PA in humans, pigs, and rodents. The other form of PA had a molecular weight of 35,000 and pI of 8.4. PAI present in GC and TT had a molecular weight of 50,000 and pI of 4.7. In GC, an acid-labile PAI was detected with biochemical characteristics similar to those of the protease, nexin I.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Changes in tissue-type plasminogen activator-like and plasminogen activator inhibitor activities in granulosa and theca layers during ovarian follicle development in the domestic hen. 211 20

The interaction of urokinase-type plasminogen activators with receptors on the surface of endothelial cells may play an important role in the regulation of fibrinolysis and cell migration. Therefore, we investigated whether human umbilical vein endothelial cells (HUVEC) express receptors for single-chain urokinase (scu-PA) on the cell surface and examined the effect of such binding on plasminogen activator activity. Binding of 125I-labeled scu-PA to HUVEC, performed at 4 degrees C, was saturable, reversible, and specific (k+1 4 +/- 1 X 10(6) min-1 M-1, k-1 6.2 +/- 1.4 X 10(-3) min-1, Kd 2.8 +/- 0.1 nM; Bmax 2.2 +/- 0.1 X 10(5) sites/cell; mean +/- S.E.). Binding of radiolabeled scu-PA was inhibited by both natural and recombinant wild-type scu-PA, high molecular weight two-chain u-PA (tcu-PA), catalytic site-inactivated tcu-PA, an amino-terminal fragment of u-PA (amino acids 1-143), and a smaller peptide (amino acids 4-42) corresponding primarily to the epidermal growth factor-like domain. Binding was not inhibited by low molecular weight urokinase or by a recombinant scu-PA missing amino acids 9-45. Cell-bound scu-PA migrated at its native molecular mass on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the presence of plasminogen, scu-PA bound to endothelial cells generated greater plasmin activity than did scu-PA in the absence of cells. In contrast, when tcu-PA was added directly to HUVEC, sodium dodecyl sulfate-stable complexes formed with cell or matrix-associated plasminogen activator inhibitors with a loss of plasminogen activator activity. These studies suggest that endothelial cells in culture express high affinity binding sites for the epidermal growth factor domain of scu-PA. Interaction of scu-PA with these receptors may permit plasminogen activator activity to be expressed at discrete sites on the endothelial cell membrane.
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PMID:Interaction of single-chain urokinase-type plasminogen activator with human endothelial cells. 215 62

The receptor for human urokinase-type plasminogen activator (u-PA) was purified from phorbol 12-myristate 13-acetate-stimulated U937 cells by temperature-induced phase separation of detergent extracts, followed by affinity chromatography with immobilized diisopropyl fluorophosphate-treated u-PA. The purified protein shows a single 55-60 kDa band after sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining. It is a heavily glycosylated protein, the deglycosylated polypeptide chain comprising only 35 kDa. The glycosylated protein contains N-acetyl-D-glucosamine and sialic acid, but no N-acetyl-D-galactosamine. Glycosylation is responsible for substantial heterogeneity in the receptor on phorbol ester-stimulated U937 cells, and also for molecular weight variations among various cell lines. The amino acid composition and the NH2-terminal amino acid sequence are reported. The protein has a high content of cysteine residues. The NH2-terminal sequence is not closely related to any known sequence. The identification of the purified and sequenced protein with the human u-PA receptor is based on the following findings: 1) the ability of the purified protein to bind u-PA and its amino-terminal fragment; 2) the identical electrophoretic mobilities observed for cross-linked conjugates, formed between either the purified protein or the u-PA receptor on intact U937 cells and the above ligands; 3) the identity of the apparent molecular weight of the purified protein to that predicted for the u-PA receptor in the same cross-linking studies; 4) the identical extent of glycosylation of the purified protein and of the u-PA receptor in crude membrane fractions, as detected after cross-linking; 5) the ability of antibodies raised against the purified protein to inhibit cellular binding of the amino-terminal fragment of u-PA.
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PMID:The human receptor for urokinase plasminogen activator. NH2-terminal amino acid sequence and glycosylation variants. 215 52

A recombinant form of plasminogen activator inhibitor-1 (rPAI-1) has been purified from lysates of pCE1200, a bacterial expression vector containing the full length PAI-1 gene, by utilizing sequential anion exchange and cation exchange chromatography on Q-Sepharose and S-Sepharose columns. Approximately 140 mg of rPAI-1, estimated at 98% purity on the basis of analytical high performance liquid chromatography, could be obtained from 200 g wet weight of cells. The purified protein exhibited a single Coomassie Blue-stainable band at the region of Mr = 42,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an NH2-terminal amino acid sequence consistent with the expected translation product of the pCE1200 PAI-1 insert. The rPAI-1 rapidly inhibited single- and two-chain tissue plasminogen activators, as well as urokinase, with apparent second order rate constants in the range of 2-5 x 10(7) M-1 s-1. A specific activity measurement of 250,000 units/mg was calculated for the rPAI-1 based on its ability to inhibit the enzymatic activity of a single-chain tissue plasminogen activator. Stability studies showed that the activity of the rPAI-1 was very stable when stored at temperatures of 25 degrees C or lower, but decayed within hours when stored at 37 degrees C. Sodium dodecyl sulfate treatment, which partially activates the latent form of natural PAI-1, inactivated rPAI-1. These results show that the purified rPAI-1 produced from pCE1200 displays many of the properties associated with the biologically active form of natural PAI-1.
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PMID:Purification and characterization of recombinant plasminogen activator inhibitor-1 from Escherichia coli. 218 80


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