WorldWideScience
 
 
1

Insulin and insulin-like growth factor I exert different effects on plasminogen activator production or cell growth in the ovine thyroid cell line OVNIS.  

Science.gov (United States)

Insulin and Insulin-like Growth Factor I (IGF-I) are evaluated for their capacity to affect cell proliferation and plasminogen activator (PA) activity production in an ovine thyroid cell line OVNIS. Insulin at physiological and supraphysiological doses induces cell proliferation and increases PA activity. IGF-I, which is also clearly mitogenic for these cells, surprisingly does not modulate PA activity. The results indicate that the growth promoting effect is mediated through the insulin and IGF-I receptors whereas PA activity is solely regulated via the insulin receptors. PMID:1802921

Degryse, B; Maisonobe, F; Hovsépian, S; Fayet, G

1991-11-01

2

Plasminogen activation in cancer  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The subject of this thesis focusses on the role of the plasminogen activation system in angiogenesis and cancer. The plasminogen activation system regulates fibrinolysis and controls cell migration and invasion by plasmin-mediated matrix proteolysis. Plasmin is formed upon cleavage of the zymogen plasminogen by plasminogen activators, urokinase-type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA). In contrast to uPA, plasminogen activation by tPA requires a cofactor, s...

2004-01-01

3

Plasminogen activation and cancer  

DEFF Research Database (Denmark)

Breakdown of the extracellular matrix is crucial for cancer invasion and metastasis. It is accomplished by the concerted action of several proteases, including the serine protease plasmin and a number of matrix metalloproteases.The activity of each of these proteases is regulated by an array of activators, inhibitors and cellular receptors.Thus, the generation of plasmin involves the pro-enzyme plasminogen, the urokinase type plasminogen activator uPA and its pro-enzyme pro-uPA, the uPA inhibitor PAI-1, the cell surface uPA receptor uPAR, and the plasmin inhibitor a2 -antiplasmin. Furthermore, the regulation of extracellular proteolysis in cancer involves a complex interplay between cancer cells and non-malignant stromal cells in the expression of the molecular components involved. For some types of cancer, this cellular interplay mimics that observed in the tissue of ori- gin during non-neoplastic tissue remodelling processes.We propose that cancer invasion can be considered as uncontrolled tissue remodelling. Inhibition of extracellular proteases is an attractive approach to cancer therapy. Because proteases have many different functions in the normal organism, efficient inhibition will have toxic side effects.In cancer invasion, like in normal tissue remodelling processes, there appears to be a functional overlap between different extracellular proteases.This redundancy means that combinations of protease inhibitors must be used. Such combination therapy, however, is also likely to increase toxicity.Therefore for each type of cancer, a combination of protease inhibitors that is optimised with respect to both maximal therapeutic effect and minimal toxic side effects need to be identified.

Danø, Keld; Behrendt, N.

2005-01-01

4

Characterization of plasmin, plasminogen and plasminogen activator in goat milk  

Directory of Open Access Journals (Sweden)

Full Text Available Plasmin (PL, a serine-proteinase, appears to be the predominant native proteinase in milk and it is mainly associated to casein micelles which represent its substrate (Bastian and Brown, 1996. Plasmin occurs in milk together with its inactive zymogene, plasminogen (PG (Bastian et al. 1991. The cascade of reactions leading to plasminogen activation is regulated by a complex network of molecular interactions between plasminogen activators (PA (tissue-type and urokinase-type and at least three types of specific PA inhibitors (Politis, 1996. Stage of lactation affects PL and PA activities: late lactation is associated with higher activity of PL and PA (Baldi et al., 1996. Plasmin in milk is responsible for the hydrolysis of ?- and ?-caseins (Aslam and Hurley, 1997; Trujillo et al., 1997...

F. Polidori

2011-03-01

5

The effect of anti-human plasminogen monoclonal antibodies on Glu-plasminogen activation by plasminogen activators  

Directory of Open Access Journals (Sweden)

Full Text Available Background: Human plasminogen is a plasma glycoprotein synthesized mainly in the liver. Conversion of plasminogen to plasmin by plasminogen activators is a key event in the fibrinolytic system. In this study, we investigated the effects of two anti-human plasminogen monoclonal antibodies, A1D12 and MC2B8 on Glu-plasminogen activation in presence of u-PA, t-PA and streptokinase. Methods: Producing of Hybridoma antibodies was performed by fusion of spleen cells from BALB/C mice immunized with Glu-plasminogen and NS1 myeloma cells. Antibody binding to Human Glu-plasminogen was assessed using an ELISA assay. Activation of plasminogen was determined by measuring plasmin generation using the chromogenic substrate S-2251 and the effect of monoclonal antibodies, A1D12 and MC2B8 on plasminogen activation in solution was then evaluated. Initial rates and kinetic parameters of plasminogen activation in the presence of monoclonal antibodies were calculated. The effect of the monoclonal antibody MC2B8 on the rate of plasmin hydrolysis was measured. The effect of F(ab'2 fragment of A1D12 on u-PA catalyzed-plasminogen activation also compared with the effect of the whole antibody in this reaction. Results: ELISA assay showed that the antibodies reacted well with antigens. A1D12 increased the maximum velocity (Vmax of plasminogen activation by each of the three plasminogen activators and MC2B8 decreased it. In all activation reactions, the KM value of plasminogen activation did not significantly change in the presence of antibody A1D12 whereas antibody MC2B8 increased the KM value of plasminogen activation by u-PA, fibrin monomer dependent t-PA and streptokinase. Monoclonal antibody MC2B8 had no significant effect on plasmin hydrolysis rate of synthetic substrate S-2251. Activation rate of plasminogen by u-PA in the lower concentration of F (ab2 fragment of A1D12 was identical to activation in the presence of the whole antibody. Conclusion: The binding of the A1D12 F(ab region to Glu-plasminogen increases the catalytic efficiency of plasminogen activation by plasminogen activators. Therefore, it may be useful to apply clinically A1D12 for the therapy of thromboembolic events such as myocardial infarction by humanizing the F(ab fragment of the A1D12 antibody. Inhibition pattern of antibody MC2B8 obey the mixed type of enzyme inhibition by binding the antibody probably at, or near, the cleavage site of Glu-plasminogen.

M. Akrami

2006-07-01

6

Fibrin-targeted plasminogen activation by plasminogen activator, PadA, from Streptococcus dysgalactiae.  

Science.gov (United States)

Bacterial plasminogen activators differ from each other in their mechanism of plasminogen activation besides their host specificity. Three-domain streptokinase (SK) and two-domain PauA generate nonproteolytic active site center in their cognate partner plasminogen but their binary activator complexes are resistant to ?2-antiplasmin (a2AP) inhibition causing nonspecific plasminogen activation in plasma. In contrast, single-domain plasminogen activator, staphylokinase (SAK), requires proteolytic cleavage of human plasminogen into plasmin for the active site generation, and this activator complex is inhibited by a2AP. The single-domain plasminogen activator, PadA, from Streptococcus dysgalatiae, having close sequence and possible structure homology with SAK, was recently reported to activate bovine Pg in a nonproteolytic manner similar to SK. We report hereby that the binary activator complex of PadA with bovine plasminogen is inhibited by a2AP and PadA is recycled from this complex to catalyze the activation of plasminogen in the clot environment, where it is completely protected from a2AP inhibition. Catalytic efficiency of the activator complex formed by PadA and bovine plasminogen is amplified several folds in the presence of cyanogen bromide digested fibrinogen but not by intact fibrinogen indicating that PadA may be highly efficient at the fibrin surface. The present study, thus, demonstrates that PadA is a unique single-domain plasminogen activator that activates bovine plasminogen in a fibrin-targeted manner like SAK. The sequence optimization by PadA for acquiring the characteristics of both SK and SAK may be exploited for the development of efficient and fibrin-specific plasminogen activators for thrombolytic therapy. PMID:24639287

Singh, Satish; Bhando, Timsy; Dikshit, Kanak L

2014-06-01

7

Does plasminogen activator inhibitor-1 drive lymphangiogenesis?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The purpose of this study is to explore the function of plasminogen activator inhibitor-1 (PAI-1) during pathological lymphangiogenesis. PAI-1, the main physiological inhibitor of plasminogen activators is involved in pathological angiogenesis at least by controlling extracellular proteolysis and by regulating endothelial cell survival and migration. Protease system's role in lymphangiogenesis is unknown yet. Thus, based on its important pro-angiogenic effect, we hypothesized that PAI-1 may r...

2010-01-01

8

Does Plasminogen Activator Inhibitor-1 Drive Lymphangiogenesis?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The purpose of this study is to explore the function of plasminogen activator inhibitor-1 (PAI-1) during pathological lymphangiogenesis. PAI-1, the main physiological inhibitor of plasminogen activators is involved in pathological angiogenesis at least by controlling extracellular proteolysis and by regulating endothelial cell survival and migration. Protease system's role in lymphangiogenesis is unknown yet. Thus, based on its important pro-angiogenic effect, we hypothesized that PAI-1 may r...

2010-01-01

9

Thrombolysis with low dose tissue plasminogen activator.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Two cases of vena caval thrombosis in infants were successfully treated with low dose (0.01-0.05 mg/kg/hour) local infusions of tissue plasminogen activator after conventional anticoagulant treatment had been unsuccessful. This approach is useful for clots associated with indwelling intravascular catheters, and a low dose infusion of tissue plasminogen activator as a regional application is recommended to achieve clot lysis with minimal systemic effects.

Doyle, E.; Britto, J.; Freeman, J.; Munro, F.; Morton, N. S.

1992-01-01

10

Plasminogen activator activity of rat hepatomas  

International Nuclear Information System (INIS)

Plasminogen activators (PA) of urokinase or tissue type have been associated with fibrinolysis, cell differentiation, and cancer growth. In the present study, PA activity has been measured in rat hepatomas having various growth rates, ranging from fast growth to slow growth. The tumor tissues were separated into homogenate, cytosol, and membrane fractions by centrifugation. The PA activity for each fraction was measured as plasminogen dependent hydrolysis of 125I labelled fibrin or D-Val-L-Leu-L-Lys-pNA. In addition, an immunobinding assay was performed on the homogenate fractions using anti-urokinase and 125I labelled protein A. Results have indicated that urokinase type PA is the dominant type found in these hepatomas and is mainly active in the homogenate and membrane fraction. The activity of PA was also found to be much greater in the faster growing hepatomas, in contrast to the moderate and slow growth tumors. These studies suggest that PA activity may be important in the growth of rat hepatomas

1987-05-01

11

Plasminogen activator activity of rat hepatomas  

Energy Technology Data Exchange (ETDEWEB)

Plasminogen activators (PA) of urokinase or tissue type have been associated with fibrinolysis, cell differentiation, and cancer growth. In the present study, PA activity has been measured in rat hepatomas having various growth rates, ranging from fast growth to slow growth. The tumor tissues were separated into homogenate, cytosol, and membrane fractions by centrifugation. The PA activity for each fraction was measured as plasminogen dependent hydrolysis of SVI labelled fibrin or D-Val-L-Leu-L-Lys-pNA. In addition, an immunobinding assay was performed on the homogenate fractions using anti-urokinase and SVI labelled protein A. Results have indicated that urokinase type PA is the dominant type found in these hepatomas and is mainly active in the homogenate and membrane fraction. The activity of PA was also found to be much greater in the faster growing hepatomas, in contrast to the moderate and slow growth tumors. These studies suggest that PA activity may be important in the growth of rat hepatomas.

Katzenberger, G.J.; Lea, M.A.; Kumar, S.

1987-05-01

12

Structural Basis for Recognition of Urokinase-type Plasminogen Activator by Plasminogen Activator Inhibitor-1*  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator inhibitor-1 (PAI-1), together with its physiological target urokinase-type plasminogen activator (uPA), plays a pivotal role in fibrinolysis, cell migration, and tissue remodeling and is currently recognized as being among the most extensively validated biological prognostic factors in several cancer types. PAI-1 specifically and rapidly inhibits uPA and tissue-type PA (tPA). Despite extensive structural/functional studies on these two reactions, the underlying structura...

2011-01-01

13

Zinc Inhibits Tumor Metastasis by Regulating Plasminogen Activation  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In this study, we found that zinc exerted a dual inhibition effect on plasminogen activator/plasmin system by inhibiting plasminogen activator activation and down-regulating plasmin activity. Zinc demonstrated significant inhibition effects on plasminogen activator and plasmin induced cell morphology change and movement. These results explain the anti-tumor and metastasis blocking effects of zinc observed in clinical studies. In addition, the strong down-regulation effects of zinc on...

2006-01-01

14

Oxidative Stress, Plasminogen Activator Inhibitor 1, and Lung Fibrosis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Fibrosis is characterized by excessive accumulation of extracellular matrix (ECM) in basement membranes and interstitial tissues, resulting from increased synthesis or decreased degradation of ECM or both. The plasminogen activator/plasmin system plays an important role in ECM degradation, whereas the plasminogen activator inhibitor 1 (PAI-1) is a physiologic inhibitor of plasminogen activators. PAI-1 expression is increased in the lung fibrotic diseases and in experimental fibrosis models. T...

Liu, Rui-ming

2008-01-01

15

Plasminogen activator system, vascular endothelial growth factor, and colorectal cancer progression  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Aims—The plasminogen activator system (PAS) consists of the plasminogen activators (urokinase (uPA) and tissue-type (tPA) plasminogen activators), the uPA receptor (uPAR), and the plasminogen activator inhibitors (PAI-1 and PAI-2). Plasminogen activators activate plasminogen to plasmin, which can break down extracellular matrix (ECM) components. Vascular endothelial growth factor (VEGF) is a mitogen for endothelial cells and is involved in angiogenesis. VEGF has been shown to upregulate uPA...

Baker, E. A.; Bergin, F. G.; Leaper, D. J.

2000-01-01

16

Does plasminogen activator inhibitor-1 drive lymphangiogenesis?  

Science.gov (United States)

The purpose of this study is to explore the function of plasminogen activator inhibitor-1 (PAI-1) during pathological lymphangiogenesis. PAI-1, the main physiological inhibitor of plasminogen activators is involved in pathological angiogenesis at least by controlling extracellular proteolysis and by regulating endothelial cell survival and migration. Protease system's role in lymphangiogenesis is unknown yet. Thus, based on its important pro-angiogenic effect, we hypothesized that PAI-1 may regulate lymphangiogenesis associated at least with metastatic dissemination of cancer cells. To address this issue, we studied the impact of PAI-1 deficiency in various murine models of tumoral lymphangiogenesis. Wild-type PAI-1 proficient mice were used as controls. We provide for the first time evidence that PAI-1 is dispensable for tumoral lymphangiogenesis associated with breast cancers either induced by mammary carcinoma cell injection or spontaneously appearing in transgenic mice expressing the polyomavirus middle T antigen (PymT) under the control of a mouse mammary tumor virus long-terminal repeat promoter (MMTV-LTR). We also investigated inflammation-related lymphatic vessel recruitment by using two inflammatory models. PAI-1 deficiency did neither affect the development of lymphangioma nor burn-induced corneal lymphangiogenesis. These novel data suggest that vascular remodelling associated with lymphangiogenesis and angiogenesis involve different molecular determinants. PAI-1 does not appear as a potential therapeutic target to counteract pathological lymphangiogenesis. PMID:20300183

Bruyère, Françoise; Melen-Lamalle, Laurence; Blacher, Silvia; Detry, Benoît; Masset, Anne; Lecomte, Julie; Lambert, Vincent; Maillard, Catherine; Høyer-Hansen, Gunilla; Lund, Leif R; Foidart, Jean-Michel; Noël, Agnès

2010-01-01

17

A miniaturized fibrinolytic assay for plasminogen activators  

Science.gov (United States)

This report describes a micro-clot lysis assay (MCLA) for evaluating fibrinolytic activity of plasminogen activators (PA). Fibrin clots were formed in wells of microtiter plates. Lysis of the clots by PA, indicated by change in turbidity (optical density, OD), was monitored with a microplate reader at five minutes intervals. Log-log plots of PA dilution versus endpoint, the time at which the OD value was halfway between the maximum and minimum value for each well, were linear over a broad range of PA concentrations (2-200 International units/ml). The MCLA is a modification and miniaturization of well established fibrinolytic methods. The significant practical advantages of the MCLA are that it is a simple, relatively sensitive, non-radioactive, quantitative, kinetic, fibrinolytic micro-technique which can be automated.

Lewis, M. L.; Nachtwey, D. S.; Damron, K. L.

1991-01-01

18

Tissue Plasminogen Activator Reduces Neurological Damage after Cerebral Embolism  

Science.gov (United States)

Intravenous administration of tissue plasminogen activator immediately after the injection of numerous small blood clots into the carotid circulation in rabbit embolic stroke model animals caused a significant reduction in neurological damage. In vitro studies indicate that tissue plasminogen activator produced substantial lysis of clots at concentrations comparable to those expected in vivo, suggesting that this may be the mechanism of action of this drug. Drug-induced hemorrhages were not demonstrable. Tissue plasminogen activator may be of value for the immediate treatment of embolic stroke.

Zivin, Justin A.; Fisher, Marc; Degirolami, Umberto; Hemenway, Carl C.; Stashak, Joan A.

1985-12-01

19

Reduction of Steroid-Induced Intraocular Pressure Elevation in Sheep by Tissue Plasminogen Activator  

Science.gov (United States)

Purpose. To investigate whether tissue plasminogen activator (tPA) can prevent and/or reverse steroid-induced IOP elevation in an ovine model. Methods. Three animal groups were subjected to bilateral steroid-induced IOP elevation using thrice daily topical ocular prednisolone administration. In the first group (N = 8), one eye each of two sheep was injected intravitreally with 100 ?g, 200 ?g, 500 ?g, or 1 mg human recombinant tPA, while contralateral eyes received vehicle. In the second group (N = 2), one eye was injected intravitreally with tPA (100 ?g), while contralateral eyes received vehicle containing L-arginine. In the third group (N = 4), each animal received intravitreal tPA in one eye concurrently with initiation of bilateral steroid administration. IOP was monitored for the duration of the experiment. Tissues from eyes of the third group were used to determine relative gene expression. Results. In the first and second groups, IOP decreased by 9.7 (±2.8) and 9.7 (±1.6) mm Hg, respectively, 24 hours after tPA administration. In the third group, tPA-treated eyes did not develop IOP elevation with ?IOP of 11.8 (±1.3) mm Hg 8 days later. In all tPA-treated eyes, IOP remained low until the end of the study. mRNA levels in the trabecular meshwork were decreased for plasminogen activator tissue (PLAT), increased for matrix-metalloproteinase 1 (MMP-1), and stable for plasminogen activator inhibitor 1 (PAI-1), MMP-2, MMP-9, and MMP-13 in tPA-treated eyes compared with contralateral controls. PAI-1 mRNA levels in ciliary processes also remained similar. Conclusions. Recombinant human tPA is effective in both preventing and reversing steroid-induced IOP elevation in sheep. Tissue plasminogen activator may be useful as a therapeutic agent in steroid-induced glaucoma.

Gerometta, Rosana; Kumar, Sandeep; Shah, Shaily; Alvarez, Larry; Candia, Oscar; Danias, John

2013-01-01

20

Inactivation of plasminogen activator inhibitor by oxidants  

Energy Technology Data Exchange (ETDEWEB)

The rapidly acting plasminogen activator inhibitor (PAI) purified from cultured bovine endothelial cells (BAEs) was inactivated during iodination with chloramine T and other oxidizing iodination systems. Inactivation was observed in the absence of iodine, suggesting that the loss of activity resulted from the oxidizing conditions employed. In an attempt to further study the nature of this inactivation, the PAI was treated with chloramine T under conditions that specifically oxidize methionine and cystein residues. Both PAI inhibitory activity and the ability of the PAI to form complexes with tissue-type PA were decreased in a dose-dependent manner by such treatment. PAI activity was measured with the lysis of /sup 125/I-labelled fibrin. The reductase is a DTT-dependent enzyme that specifically converts methionine sulfoxide to methionine. Little activity was restored by either the reductase or DTT alone. These results indicate that the oxidation of at least one critical methionine residue is responsible for the loss of PAI activity upon iodination. In this respect, the BAE PAI resembles ..cap alpha../sub 1/-protease inhibitor, a well-characterized elastase inhibitor that also is inactivated by oxidants. Both inhibitors are members of the serine protease inhibitor superfamily (Serpins), and both have a methionine residue in their reactive center.

Lawrence, D.A.; Loskutoff, D.J.

1986-10-21

 
 
 
 
21

Inactivation of plasminogen activator inhibitor by oxidants  

International Nuclear Information System (INIS)

The rapidly acting plasminogen activator inhibitor (PAI) purified from cultured bovine endothelial cells (BAEs) was inactivated during iodination with chloramine T and other oxidizing iodination systems. Inactivation was observed in the absence of iodine, suggesting that the loss of activity resulted from the oxidizing conditions employed. In an attempt to further study the nature of this inactivation, the PAI was treated with chloramine T under conditions that specifically oxidize methionine and cystein residues. Both PAI inhibitory activity and the ability of the PAI to form complexes with tissue-type PA were decreased in a dose-dependent manner by such treatment. PAI activity was measured with the lysis of "1"2"5I-labelled fibrin. The reductase is a DTT-dependent enzyme that specifically converts methionine sulfoxide to methionine. Little activity was restored by either the reductase or DTT alone. These results indicate that the oxidation of at least one critical methionine residue is responsible for the loss of PAI activity upon iodination. In this respect, the BAE PAI resembles ?_1-protease inhibitor, a well-characterized elastase inhibitor that also is inactivated by oxidants. Both inhibitors are members of the serine protease inhibitor superfamily (Serpins), and both have a methionine residue in their reactive center

1986-10-21

22

Influence of Plasminogen Activator Inhibitor Type 1 on Choroidal Neovascularization  

Digital Repository Infrastructure Vision for European Research (DRIVER)

High levels of the plasminogen activators, but also their inhibitor, plasminogen activator inhibitor 1 (PAI-1), have been documented in neovascularization of severe ocular pathologies such as diabetic retinopathy or age-related macular degeneration (AMD). AMD is the primary cause of irreversible photoreceptors loss, and current therapies are limited. PAI-1 has recently been shown to be essential for tumoral angiogenesis. We report here that deficient PAI-1 expression in mice prevented the dev...

2001-01-01

23

Tissue Plasminogen Activator Induced Delayed Edema in Experimental Porcine Intracranial Hemorrhage: Reduction with Plasminogen Activator Inhibitor-1 Administration  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Hematoma puncture and subsequent clot lysis with recombinant tissue plasminogen activator (rtPA) emerged as an alternative therapy for spontaneous intracerebral hemorrhage (ICH) and is associated with delayed edema possibly counteracting the beneficial effects of hematoma volume reduction. We hypothesized that immediate reversal of rtPA activity after clot lysis and hematoma drainage diminishes edema formation. To test this hypothesis, we administered plasminogen activator inhibitor (PAI)-1 a...

Keric, Naureen; Maier, Gerrit; Samadani, Uzma; Kallenberg, Kai; Dechent, Peter; Brueck, Wolfgang; Heuer, Jan; Rohde, Veit

2012-01-01

24

The Glycosylation of Plasminogen Activator Inhibitor-1  

DEFF Research Database (Denmark)

Plasminogen activator inhibitor type-1 (PAI-1) has three potential sites for N-linked glycosylation, including Asn209Tyr210Thr211, Asn265Met266Thr267, and Asn329Glu330Ser331. Using a HEK293 expression system, we have made mutants with Asp or Gln substitutions of the Asn residue in each of these sequences. Analyses of these mutants for the content of N-acetyl glucosamine showed that Asn209 and Asn265, but not Asn329, are glycosylated, in agreement with previous suggestions made on the basis of X-ray crystal structure analysis of PAI-1 expressed in CHO cells (Xue et al. (1998) Structure 6, 627-636). In contrast, PAI-1, containing a total of 26 Ser and 26 Thr residues, which are potential targets for O-linked glycosylation, was found to be devoid of N-acetyl-galactosamine, demonstrating the absence of O-linked glycosylation. Analysis of PAI-1 variants with mutational inactivation of each of the sequences utilized for N-linked glycosylation by Fluorophore Assisted Carbohydrate Electrophoresis (FACE), showed a different N-linked glycosylation profile of the glycans at each of the 2 sites. The exact structure of the carbohydrate chains at each of these 2 sequences are being determined using MALDI mass spectrometry and monosaccharide composition analysis and compared to that of natural and recombinant PAI-1 from other sources. These results contribute to a structural basis for previous observations of a different functional importance of the N-linked glycosylation at each of the 2 sequences.

Skottrup, Peter Durand; Pedersen, Katrine Egelund

25

Plasmin-mediated fibrinolysis by variant recombinant tissue plasminogen activators.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A rapid and quantitative fibrinolytic assay has been used to measure the overall activity of a recombinant tissue plasminogen activator (rTPA) preparation for dissolution of a fibrin clot by its ability to activate [Glu1]plasminogen (containing glutamic acid at position 1) to plasmin. A standard curve constructed for wild-type two-chain rTPA that contains, from the amino terminus, the finger (F)-growth factor (E)-kringle 1 (K1)-kringle 2 (K2)-serine protease (P) domains was used to assess the...

Urano, S.; Metzger, A. R.; Castellino, F. J.

1989-01-01

26

Structural basis for recognition of urokinase-type plasminogen activator by plasminogen activator inhibitor-1  

DEFF Research Database (Denmark)

Plasminogen activator inhibitor-1 (PAI-1), together with its physiological target urokinase-type plasminogen activator (uPA), plays a pivotal role in fibrinolysis, cell migration, and tissue remodeling and is currently recognized as being among the most extensively validated biological prognostic factors in several cancer types. PAI-1 specifically and rapidly inhibits uPA and tissue-type PA (tPA). Despite extensive structural/functional studies on these two reactions, the underlying structural mechanism has remained unknown due to the technical difficulties of obtaining the relevant structures. Here, we report a strategy to generate a PAI-1·uPA(S195A) Michaelis complex and present its crystal structure at 2.3-� resolution. In this structure, the PAI-1 reactive center loop serves as a bait to attract uPA onto the top of the PAI-1 molecule. The P4-P3' residues of the reactive center loop interact extensively with the uPA catalytic site, accounting for about two-thirds of the total contact area. Besides the active site, almost all uPA exosite loops, including the 37-, 60-, 97-, 147-, and 217-loops, are involved in the interaction with PAI-1. The uPA 37-loop makes an extensive interaction with PAI-1 β-sheet B, and the 147-loop directly contacts PAI-1 β-sheet C. Both loops are important for initial Michaelis complex formation. This study lays down a foundation for understanding the specificity of PAI-1 for uPA and tPA and provides a structural basis for further functional studies.

Lin, Zhonghui; Jiang, Longguang

2011-01-01

27

Limited cleavage of cellular fibronectin by plasminogen activator purified from transformed cells.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The substrate specificity and direct catalytic activity of plasminogen activator (PA) was examined under conditions where its natural substrate, plasminogen, was missing or inhibited. PA, purified from cultures of transformed chicken fibroblasts, was incubated with purified preparations of potential substrates. The adhesive glycoprotein fibronectin, isolated from normal chicken fibroblast extracellular matrix, underwent limited but specific cleavage by PA in the absence of plasminogen. Analys...

Quigley, J. P.; Gold, L. I.; Schwimmer, R.; Sullivan, L. M.

1987-01-01

28

The tissue-type plasminogen activator-plasminogen activator inhibitor 1 complex promotes neurovascular injury in brain trauma: evidence from mice and humans  

Science.gov (United States)

The neurovascular unit provides a dynamic interface between the circulation and central nervous system. Disruption of neurovascular integrity occurs in numerous brain pathologies including neurotrauma and ischaemic stroke. Tissue plasminogen activator is a serine protease that converts plasminogen to plasmin, a protease that dissolves blood clots. Besides its role in fibrinolysis, tissue plasminogen activator is abundantly expressed in the brain where it mediates extracellular proteolysis. However, proteolytically active tissue plasminogen activator also promotes neurovascular disruption after ischaemic stroke; the molecular mechanisms of this process are still unclear. Tissue plasminogen activator is naturally inhibited by serine protease inhibitors (serpins): plasminogen activator inhibitor-1, neuroserpin or protease nexin-1 that results in the formation of serpin:protease complexes. Proteases and serpin:protease complexes are cleared through high-affinity binding to low-density lipoprotein receptors, but their binding to these receptors can also transmit extracellular signals across the plasma membrane. The matrix metalloproteinases are the second major proteolytic system in the mammalian brain, and like tissue plasminogen activators are pivotal to neurological function but can also degrade structures of the neurovascular unit after injury. Herein, we show that tissue plasminogen activator potentiates neurovascular damage in a dose-dependent manner in a mouse model of neurotrauma. Surprisingly, inhibition of activity following administration of plasminogen activator inhibitor-1 significantly increased cerebrovascular permeability. This led to our finding that formation of complexes between tissue plasminogen activator and plasminogen activator inhibitor-1 in the brain parenchyma facilitates post-traumatic cerebrovascular damage. We demonstrate that following trauma, the complex binds to low-density lipoprotein receptors, triggering the induction of matrix metalloproteinase-3. Accordingly, pharmacological inhibition of matrix metalloproteinase-3 attenuates neurovascular permeability and improves neurological function in injured mice. Our results are clinically relevant, because concentrations of tissue plasminogen activator: plasminogen activator inhibitor-1 complex and matrix metalloproteinase-3 are significantly elevated in cerebrospinal fluid of trauma patients and correlate with neurological outcome. In a separate study, we found that matrix metalloproteinase-3 and albumin, a marker of cerebrovascular damage, were significantly increased in brain tissue of patients with neurotrauma. Perturbation of neurovascular homeostasis causing oedema, inflammation and cell death is an important cause of acute and long-term neurological dysfunction after trauma. A role for the tissue plasminogen activator–matrix metalloproteinase axis in promoting neurovascular disruption after neurotrauma has not been described thus far. Targeting tissue plasminogen activator: plasminogen activator inhibitor-1 complex signalling or downstream matrix metalloproteinase-3 induction may provide viable therapeutic strategies to reduce cerebrovascular permeability after neurotrauma.

Sashindranath, Maithili; Sales, Eunice; Daglas, Maria; Freeman, Roxann; Samson, Andre L.; Cops, Elisa J.; Beckham, Simone; Galle, Adam; McLean, Catriona; Morganti-Kossmann, Cristina; Rosenfeld, Jeffrey V.; Madani, Rime; Vassalli, Jean-Dominique; Su, Enming J.; Lawrence, Daniel A.

2012-01-01

29

Diagnostic value of plasminogen activity level in acute mesenteric ischemia  

Directory of Open Access Journals (Sweden)

Full Text Available AIM: To investigate the changes in plasminogen activity level during mesenteric ischemia.METHODS: We performed laparotomy in 90 female Wistar-Albino rats (average weight 230 g. In sham groups (SL (GroupsIand II the superior mesenteric artery (SMA and vein (SMV were explored, but not tied. In SMA groups (Groups III and IV the SMA was ligated, and in SMV groups (Groups V and VI the SMV was ligated. On re-laparotomy 2 mL of blood was drawn at 1 h in groupsI, III and V, and at 3 h in groups II, IV and VI. Plasminogen levels were assessed and comparisons were made between groups and within each group.RESULTS: The mean plasminogen activity in the SL group was significantly higher than SMA (25.1 ± 10.8 vs 11.8 ± 4.6, P < 0.001 or SMV (25.1 ± 10.8 vs 13.7 ± 4.4, P < 0.001 groups both at 1 h and at 3 h (29.8 ± 8.9 vs 15.1 ± 5.7, P < 0.0001; 29.8 ± 8.9 vs 14.2 ± 2.9, P < 0.0001. There were no significant differences between the values of SMA and SMV groups at 1 h (P = 0.28 and at 3 h (P = 0.71. In each group, plasminogen activity levels did not change significantly between the two measurements performed at 1 h and 3 h.CONCLUSION: We conclude that blood plasminogen activities decrease during early phases of both arterial and venous mesenteric ischemia which may be a useful marker for early diagnosis.

Yusuf Gunerhan, Neset Koksal, Munire Kayahan, Yavuz Eryavuz, Hilal Sekban

2008-04-01

30

A simple quantitative assay of tissue plasminogen activator.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A simple method for the quantitative assay of tissue plasminogen activator is described. Human veins and uterus obtained at operation are disintegrated in a membrane disintegrator at -70 degrees C and a known weight of the powder, suspended in buffered saline and thoroughly mixed. Assay of the dilutions of this homogenate on isotope-labelled fibrin clots gives straight line plots of log weight against log activity of the dilution and the sample activity is calculated from this. The method has...

Cossart, L.; Marcuson, R. W.

1982-01-01

31

Immunomodulatory effects of plasminogen activators on hepatic fibrogenesis.  

Science.gov (United States)

Tissue-type plasminogen activators (tPA) and urokinase-type plasminogen activators (uPA) are involved in liver repair. We examined the potential immunomodulatory actions of uPA, tPA and uPA-receptor (uPAR) in carbon-tetrachloride-induced hepatic fibrosis in wild-type (WT), tPA-/-, uPA-/- and uPAR-/- mice. Carbon-tetrachloride treatment increased fibrosis in four groups but significantly less in three knock-out models. Serum cytokines and intrahepatic T cells elevated significantly following fibrosis process in WT animals but not in the knock-out groups. In culture, uPA increased lymphocyte proliferation significantly in WT and uPA-/- but not uPAR-/- animals. Following uPA exposure in vivo, there was CD8 predominance. To isolate uPA's effect on lymphocytes, WT mice were irradiated sublethally and then reconstituted with WT or uPA-/- lymphocytes. In these animals fibrosis was decreased and T cells were reduced in the uPA-/- recipients. Based on these data we postulate that plasminogen activators affect fibrosis in part by liver-specific activation of CD8 subsets that govern the fibrogenic activity of hepatic stellate cells. PMID:18279442

Higazi, A A; El-Haj, M; Melhem, A; Horani, A; Pappo, O; Alvarez, C E; Muhanna, N; Friedman, S L; Safadi, R

2008-04-01

32

Mechanism of the action of SMTP-7, a novel small-molecule modulator of plasminogen activation.  

Science.gov (United States)

SMTP-7 is a small molecule that promotes the proteolytic activation of plasminogen by relaxing its conformation. SMTP-7 has excellent therapeutic activities against thrombotic stroke in several rodent models. The objective of this study was to elucidate detailed mechanism of the action of SMTP-7 in vitro. We report here that the action of SMTP-7 requires a cofactor with a long-chain alkyl or alkenyl group, and that the fifth kringle domain (kringle 5) of plasminogen is involved in the SMTP-7 action. In this study, we found that the SMTP-7 action to enhance plasminogen activation depended on the presence of a certain type of surfactant, and we screened biologically relevant molecules for their cofactor activity for the SMTP action. As a result, phospholipids, sphingolipids, and oleic acid were found to be active in assisting the SMTP-7 action. On the contrary, stearic acid and bile acids were inactive. Thus, a certain structural element, not only the surface-activating potential, is required for a compound to act as a cofactor for the SMTP-7 action. The plasminogen molecule consists of a PAN domain, five kringle domains, and a serine protease domain. The cofactor-dependent effects of SMTP-7 was observed with plasminogen species including kringle 5 such as intact plasminogen (Glu-plasminogen), des-PAN plasminogen (Lys-plasminogen), and des-[PAN?-?(kringles 1-4)] plasminogen (mini-plasminogen). However, SMTP-7 effect was not observed with the smallest plasminogen species des-[PAN?-?(kringles 1-4) and a half of kringle 5)] plasminogen (micro-plasminogen). Thus, kringle 5 is crucial for the action of SMTP-7. PMID:24784315

Koyanagi, Keiji; Narasaki, Ritsuko; Yamamichi, Shingo; Suzuki, Eriko; Hasumi, Keiji

2014-06-01

33

Simplex optimization of acoustic assay for plasminogen activators.  

Science.gov (United States)

This article discusses the optimization of a newly developed method for measuring the activity of plasminogen activators using a thickness-shear-mode acoustic sensor. A variable-size simplex algorithm was used for optimization. Preliminary tests were performed to design the first simplex. A desirability function was defined to translate each performance value to a membership value of 0 to 1. If there was more than one performance variable, their membership values were translated to an aggregated membership value using another function that considers their individual influence on sensor performance. Two rounds of optimization were carried out for streptokinase followed by a single optimization for tissue-type plasminogen activator. In the last optimization, ratios of control variables were used in order to reduce the number of parameters and to formulate easily adjustable assay conditions. The results showed the usefulness of the simplex method for optimizing this type of assay, and the importance of preliminary tests and prior knowledge in providing rapid convergence using fewer experiments. The optimized plasminogen activator assay can be considered a reference method for measurement of all members of this drug class. PMID:19023567

Ghazali, Mirnader; Hayward, Gordon L

2009-01-01

34

Telephone consultations for tissue plasminogen activator administration in acute stroke.  

Science.gov (United States)

Effective treatment for acute ischemic stroke has been available for 17 years, but wide geographic variability remains in timely access to neurologic expertise and other components of stroke systems of care. Telemedical technology can be used to improve such access, but it is debatable whether neurologists have an ethical obligation to provide consultation regarding tissue plasminogen activator use via the telephone. This article examines whether neurologists are ethically obligated to provide telephone-mediated acute stroke consultation. PMID:24699491

Sattin, Justin A

2014-04-01

35

Immunohistochemical localisation of tissue plasminogen activator in human brain tumours.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The distribution of tissue plasminogen activator (t-PA) has been studied in a series of 38 human brain tumours and two specimens of cerebral cortex, using the monoclonal antibody ESP6. t-PA was localised in vascular endothelium in the majority of tumours and both the cortical specimens, confirmed by double staining with Ulex europaeus lectin (Uel) and Factor 8-related antigen. Nineteen out of 22 high grade astrocytomas showed strong endothelial staining whereas staining was weak or absent in ...

Franks, A. J.; Ellis, E.

1989-01-01

36

Plasminogen Activation–Induced Pericellular Fibronectin Proteolysis Promotes Fibroblast Apoptosis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Apoptosis of fibroblasts/myofibroblasts is a critical event in the resolution of tissue repair responses; however, mechanisms for the regulation of (myo)fibroblast apoptosis/survival remain unclear. In this study, we demonstrate counter-regulatory interactions between the plasminogen activation system and transforming growth factor-?1 (TGF-?1) in the control of fibroblast apoptosis. Plasmin treatment induced fibroblast apoptosis in a time- and dose-dependent manner in association with prote...

Horowitz, Jeffrey C.; Rogers, David S.; Simon, Richard H.; Sisson, Thomas H.; Thannickal, Victor J.

2008-01-01

37

Intraventricular recombinant tissue plasminogen activator for lysis of intraventricular haemorrhage.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A prospective series of 20 patients with moderate to severe intraventricular haemorrhage (IVH) was studied for the effect of intraventricular administration of recombinant tissue plasminogen activator (rt-PA) on reduction of haematoma volume and prognosis. On the day of haemorrhage ventriculostomy was performed and 2 to 5 mg of rt-PA were injected via the external ventricular drainage, followed by drainage closure for two hours. In 14 patients rt-PA treatment was repeated. Computed tomography...

Rohde, V.; Schaller, C.; Hassler, W. E.

1995-01-01

38

Role of Plasminogen Activator in Spinal Cord Remodeling after Spinal Cord Injury  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activators play an active role in synaptic plasticity associated with the crossed phrenic phenomenon (CPP) and recovery of respiratory function following spinal cord injury. A genetic approach has been used to identify molecular mechanisms underlying this synaptic plasticity. Knockout mice lacking different genes in the plasminogen activator/plasmin system demonstrate that expression of urokinase plasminogen activator (uPA) is required during the critical 1-2h delay period followi...

Seeds, Nicholas W.; Akison, Lisa; Minor, Kenneth

2009-01-01

39

Adenosine potentiates human lung mast cell tissue plasminogen activator activity.  

Science.gov (United States)

We investigated whether adenosine, a potent contributor to the regulation of pulmonary function, can modulate human lung mast cell (HLMC) fibrinolytic activity. Tissue plasminogen activator (tPA) activity and tPA transcript expression levels from a human mast cell line (HMC-1) and HLMC were monitored following adenosine application. Adenosine potentiated mast cell tPA activity and tPA gene expression in a dose-dependent manner. Adenosine effects were abolished in the presence of adenosine deaminase. HMC-1 cells and HLMC predominantly expressed adenosine A(2A) and A(2B) receptor transcripts (A(2B) ? A(2A) > A(3) > A(1)). Pharmacological and signaling studies suggest that the A(2A) receptor is the major subtype accounting for adenosine-induced mast cell tPA activity. Finally, the supernatant from HMC-1 cells and HLMC treated with adenosine (for 24 h) significantly increased fibrin clot lysis, whereas ZM241385, an A(2A) receptor antagonist, abolished this effect. To our knowledge, this study provides the first data to demonstrate the potentiating effect of adenosine on mast cell tPA activity and fibrin clot lysis. PMID:21149610

Sereda, Michal J; Bradding, Peter; Vial, Catherine

2011-01-15

40

Immunoradiometric determination of the blood/tissue plasminogen activator in thrombophilia  

Energy Technology Data Exchange (ETDEWEB)

Immunoradiometric determination of the blood/tissue plasminogen activator was performed in plasma from patients before and after response to venous occlusion, infusion of 1-desamino-8-D-arginine-vasopressin (DDAVP) or exercise. The raise in the level of plasminogen activator was most pronounced after venous occlusion. In patients who earlier had had verified thrombosis the levels of plasminogen activator compared to normals did not show any significant difference.

Astedt, B.; Fagner, U. (Lund Univ. (Sweden))

1984-01-01

 
 
 
 
41

Immunoradiometric determination of the blood/tissue plasminogen activator in thrombophilia  

International Nuclear Information System (INIS)

Immunoradiometric determination of the blood/tissue plasminogen activator was performed in plasma from patients before and after response to venous occlusion, infusion of 1-desamino-8-D-arginine-vasopressin (DDAVP) or exercise. The raise in the level of plasminogen activator was most pronounced after venous occlusion. In patients who earlier had had verified thrombosis the levels of plasminogen activator compared to normals did not show any significant difference. (author)

1983-05-14

42

Degradation of human plasma and extracellular matrix fibronectin by tissue type plasminogen activator and urokinase.  

Science.gov (United States)

Fibronectins and plasminogen activators, both tissue and urokinase types, are involved in the physiopathological degradation of the extracellular matrix. Previous reports indicate that fibronectin can be degraded by urokinase without plasminogen. Also, we have shown that tissue-type plasminogen activator can cleave fibronectin, without plasminogen, generating fragments of 30 and 220-250 kDa detectable by immunoblotting analysis. A comparison with urokinase-induced degradation indicates that the cleavage sites are the same for both plasminogen activators. One is close to the carboxyl-terminal, disrupting the fibronectin dimeric structure, and one is near the amino-terminal, generating a 30 kDa fragment. In solution, the activity of tissue-type plasminogen activator was prevalent on the amino-terminal site, while urokinase activity was prevalent on the carboxyl-terminal site. On fibronectin immobilized onto a gelatin coated surface, only the 30 kDa fragment was released when treated with both plasminogen activators. Plasminogen activators also were active on fibronectin assembled into the extracellular matrix of cultured fibroblasts. Urokinase caused the complete disappearance of extracellular matrix fibronectin, together with the release of the 30 and the 220-250 kDa fibronectin fragments, but left a flat morphology, while tissue-type plasminogen activator induced the release of the 30 kDa fragment associated with changes in cellular morphology. The plasminogen-independent fibronectin degradation exerted by tissue-type plasminogen activator and urokinase is 100 times lower than that exerted by plasmin. This may provide a mechanism for localized and limited degradation of fibronectin preventing the generalized proteolysis associated with plasminogen activation. PMID:8930138

Marchina, E; Barlati, S

1996-10-01

43

Cellular and extracellular plasminogen activator and inhibitor in an experimental tumour.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We determined plasminogen activator (PA) and PA inhibitor (PAI) activities in the intra- and extracellular compartments of an experimental pancreatic ascites tumour with indirect and direct functional assays, and partially characterized these activities on SDS-polyacrylamide gels coupled with fibrin and reverse fibrin autography. Intact tumour cells caused lysis of plasminogen-rich but not plasminogen-free fibrin clots, and the extent of lysis of the former was related to tumour cell count. D...

Dong, Q.; Zhou, M. H.; Subbarao, V.; Ts Ao, C. H.

1988-01-01

44

Secretion of platelet-activating factor by periovulatory ovine follicles  

International Nuclear Information System (INIS)

Secretion of platelet-activating factor (PAF) in vitro by ovine follicles and ovarian interstitium obtained at various times before, during and after the endogenous preovulatory surge of luteinizing hormone (LH) and ovulation was quantified by radioimmunoassay. Release of PAF by the preovulatory follicle increased within 2 h after initiation of the surge of LH. Capacity for secretion of PAF was greatest at the time of ovulation, then declined thereafter. Production of PAF by ovarian interstitium throughout the periovulatory period was relatively low and did not change with time. It appears that PAF could act as an intrafollicular mediator in the mechanisms of ovulation and(or) luteinization

1990-01-01

45

Plasminogen- and colony-stimulating factor-1-associated markers in bladder carcinoma: diagnostic value of urokinase plasminogen activator receptor and plasminogen activator inhibitor type-2 using immunocytochemical analysis.  

Science.gov (United States)

The expression of plasminogen- and colony-stimulating factor-1-associated markers was first investigated in seven bladder carcinoma cell lines and in 15 primary bladder tumors using RT-PCR (mRNAs), zymography (protein activity), ELISA and immunocytochemistry analysis (ICC) (protein levels). The mRNAs expression, the activity and the levels of the secreted proteins were not informative. Only urokinase plasminogen activator receptor (uPA-R/CD87) and possibly plasminogen activator inhibitor type-2 (PAI2) antigen expression at the cellular levels seem to be useful markers. uPA-R antigen expression correlated with the secretion of hepatocyte growth factor (HGF) ( P=0.016) and the motility of the bladder tumor cells ( P=0.014), two markers associated with a poor prognosis in bladder carcinoma. To validate our technique and confirm these preliminary results, uPA-R and PAI2 antigen expression was determined in the imprints from 129 resected bladder carcinoma fragments. uPA-R correlated with the grade ( P=0.002), tumor invasion ( P=0.003) and the ploidy ( P=0.05) of the bladder carcinomas and with the low overall survival ( P=0.045) of the patients. PAI2 correlated only with the stage ( P=0.02) and low overall survival ( P=0.038). We conclude that in bladder carcinomas, studying the transcripts of PAs, PAIs, CSF-1 and its receptor, as well as measuring their concentration or activity in culture supernatants was of no clinical interest in terms of diagnostic or prognostic value. Only the ICC of uPA-R, which correlated with the major histopathological parameters of tumors and the low overall survival, proved to be a diagnostic and prognostic marker. PMID:12389118

Champelovier, Pierre; Boucard, Nathalie; Levacher, Geraldine; Simon, Annick; Seigneurin, Daniel; Praloran, Vincent

2002-10-01

46

Maspin increases extracellular plasminogen activator activity associated with corneal fibroblasts and myofibroblasts.  

Science.gov (United States)

Maspin, an inhibitor of cell migration and a stimulator of adhesion of cells to the ECM, is synthesized and released by corneal keratocytes into the extracellular matrix. When the cornea is wounded, the quiescent stromal keratocytes underlying the wound undergo apoptosis and cells adjacent to this apoptotic area convert to fibroblasts or myofibroblasts. This study explores the effect of extracellular maspin on the plasminogen-plasminogen activator system of corneal stromal cells following wounding. Treatment of corneal fibroblasts and myofibroblasts with r-maspin increased extracellular but not cell-associated tissue-type plasminogen activator (tPA), urinary-type plasminogen activator (uPA) or plasminogen activator inhibitor-1 (PAI-1). Despite the extracellular increase in PAI-1, the net effect of maspin treatment was an increase in plasminogen activation. At physiological levels, maspin did not alter uPA or tPA mRNA levels, in these cells. The increase in pro and active uPA was due to decreased clearance in the presence of maspin for myofibroblasts but not for fibroblasts. The clearance of pro and active tPA was normal in fibroblasts indicating different mechanisms for the increase of these homologous enzymes in the two cell types. Increased generation of plasmin by maspin treated corneal stromal fibroblasts and myofibroblasts led to conversion of plasminogen to active plasmin degradation products and angiostatin-like molecules. This study suggests that extracellular maspin increased pro and active uPA and tPA released by corneal fibroblasts and myofibroblasts on the short time scale of 1-4 h, but by 24 h there was no increase over the levels produced without maspin. This augmentation of plasminogen activator activity increases plasmin activation and angiostatin generation. It further indicates that the effect of maspin on uPA and tPA levels is cell type dependent. PMID:21810423

Warejcka, Debra J; Narayan, Malathi; Twining, Sally S

2011-11-01

47

Method and tool for prognosticating HIV infection in a subject by measuring soluble urokinase plasminogen activator receptor, degradation products thereof, and urokinase plasminogen activator receptor  

DEFF Research Database (Denmark)

Method of diagnosing and/or prognosticating HIV infection in a subject comprising the steps of: (a) performing in vitro a measurement of the level of a marker in the form of (i) urokinase plasminogen activator receptor (uPAR), (ii) soluble urokinase plasminogen activator receptor (suPAR), (iii) urokinase-type plasminogen activator (uPA), (iv) one or more degradation products of (i), (ii), or (iii), and/or (v) an mRNA for (i), (ii) or (iii), in a biological fluid sample from a subject, and (b) using the measurement value obtained to evaluate the state of the subject.

Eugen-Olsen, Jesper Hvidovre Hospital,

48

Differences between neonates and adults in tissue-type-plasminogen activator (t-PA)-catalyzed plasminogen activation with various effectors and in carbohydrate sequences of fibrinogen chains.  

Science.gov (United States)

Our study investigates the effect of fetal and adult soluble fibrin (SF), fetal and adult fibrinogen Aalpha- and gamma-chains, as well as adult CNBr-fibrinogen fragments on tissue-type plasminogen activator (t-PA)-catalyzed plasminogen activation of both fetal and adult Glu-plasminogen types 1 and 2. In addition, we determined carbohydrate sequences of fetal and adult Bbeta- and gamma-chains by mass spectrometric analysis. In the absence of an effector, no substantial differences in the rate of plasmin formation could be seen between the fetal and adult plasminogen types. In the presence of an effector, both fetal Glu-plasminogen types revealed lower values for k(cat app) than the respective adult types. No differences could be seen in the values for K(m app). The resulting differences in catalytic efficiencies between the fetal and adult plasminogen types were much less than previously reported. No differences could be seen between fetal and adult effectors in stimulating t-PA-catalyzed plasminogen activation. Detailed analyses of the activation kinetics revealed a longer initial phase of slow plasmin formation of both fetal Glu-plasminogen types compared to their respective adult types, indicating a slower plasmin-induced modification of CNBr-fibrinogen fragments or SF by fetal plasmin. Mass spectrometric analysis of the N-glycans present on adult and fetal Bbeta- and gamma-fibrinogen chains showed the presence of a major monosialylated biantennary structure with lesser amounts of the disialylated form. In contrast to previous data, we conclude that catalytic efficiency of t-PA-catalyzed plasminogen activation in neonates is only slightly lower than in adults. PMID:11672579

Ries, M; Easton, R L; Longstaff, C; Zenker, M; Corran, P H; Morris, H R; Dell, A; Gaffney, P J

2001-08-01

49

Stimulation of radiation-impaired plasminogen activator release by phorbol ester in aortic endothelial cells  

International Nuclear Information System (INIS)

Ionizing radiation has been reported to affect the fibrinolytic activity of exposed tissue. With cultured bovine aortic endothelial cells, radiation suppresses the release of plasminogen activator to the conditioned media, with a concomitant increase in intracellular plasminogen activator. Thus study was undertaken to determine whether radiation-impaired plasminogen activator release can be modified by phorbol ester. We exposed cultured bovine aortic endothelial cells to a sterilizing dose of 10 Gy of gamma-rays and found the treatment led to cell injury, as evidenced by an increased release of prelabeled chromium, and to a reduction of plasminogen activator in the conditioned media with elevated intracellular plasminogen activator in irradiated cells. Phorbol ester enhanced plasminogen activator activity in both sham-irradiated and irradiated endothelial cells. It was interesting to note that the increased plasminogen activator in phorbol ester-stimulated sham-irradiated cells was largely retained inside the cell, while it was released to the conditioned media in irradiated cells. Apparently, altered plasminogen activator activity of radiation-sterilized endothelial cells can be modified by exogenous stimuli

1990-01-01

50

Extending the capabilities of targeted molecular dynamics: Simulation of a large conformational transition in plasminogen activator inhibitor 1  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator inhibitor type 1 (PAI-1) is an inhibitor of plasminogen activators such as tissue-type plasminogen activator or urokinase-type plasminogen activator. For this molecule, different conformations are known. The inhibiting form that interacts with the proteinases is called the active form. The noninhibitory, noncleavable form is called the latent form. X-ray and modeling studies have revealed a large change in position of the reactive center loop (RCL), responsible for the i...

2001-01-01

51

Measurement of human tissue-type plasminogen activator by a two-site immunoradiometric assay  

International Nuclear Information System (INIS)

A two-site immunoradiometric assay for human extrinsic (tissue-type) plasminogen activator was developed by using rabbit antibodies raised against plasminogen activator purified from human melanoma cell culture fluid. Samples of 100 ?l containing 1 to 100 ng/ml plasminogen activator were incubated in the wells of polyvinyl chloride microtiter plates coated with antibody. The amount of bound extrinsic plasminogen activator was quantitated by the subsequent binding of 125I-labeled affinospecific antibody. The mean level of plasma samples taken at rest was 6.6 +/- 2.9 ng/ml (n = 54). This level increased approximately threefold by exhaustive physical exercise, venous occlusion, or infusion of DDAVP. Extrinsic plasminogen activator in plasma is composed of a fibrin-adsorbable and active component (1.9 +/- 1.1 ng/ml, n = 54, in resting conditions) and an inactive component that does not bind to a fibrin clot (probably extrinsic plasminogen activator-proteinase inhibitor complexes). The fibrin-adsorbable fraction increased approximately fivefold to eightfold after physical exercise, venous occlusion, or DDAVP injections. Potential applications of the immunoradiometric assay are illustrated by the measurement of extrinsic plasminogen activator in different tissue extracts, body fluids, and cell culture fluids and in oocyte translation products after injection with mRNA for plasminogen activator

1983-01-01

52

Measurement of human tissue-type plasminogen activator by a two-site immunoradiometric assay  

Energy Technology Data Exchange (ETDEWEB)

A two-site immunoradiometric assay for human extrinsic (tissue-type) plasminogen activator was developed by using rabbit antibodies raised against plasminogen activator purified from human melanoma cell culture fluid. Samples of 100 ..mu..l containing 1 to 100 ng/ml plasminogen activator were incubated in the wells of polyvinyl chloride microtiter plates coated with antibody. The amount of bound extrinsic plasminogen activator was quantitated by the subsequent binding of /sup 125/I-labeled affinospecific antibody. The mean level of plasma samples taken at rest was 6.6 +/- 2.9 ng/ml (n = 54). This level increased approximately threefold by exhaustive physical exercise, venous occlusion, or infusion of DDAVP. Extrinsic plasminogen activator in plasma is composed of a fibrin-adsorbable and active component (1.9 +/- 1.1 ng/ml, n = 54, in resting conditions) and an inactive component that does not bind to a fibrin clot (probably extrinsic plasminogen activator-proteinase inhibitor complexes). The fibrin-adsorbable fraction increased approximately fivefold to eightfold after physical exercise, venous occlusion, or DDAVP injections. Potential applications of the immunoradiometric assay are illustrated by the measurement of extrinsic plasminogen activator in different tissue extracts, body fluids, and cell culture fluids and in oocyte translation products after injection with mRNA for plasminogen activator.

Rijken, D.C. (Univ. of Leuven, Belgium); Juhan-Vague, I.; De Cock, F.; Collen, D.

1983-02-01

53

Maspin increases extracellular plasminogen activator activity associated with corneal fibroblasts and myofibroblasts  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Maspin, an inhibitor of cell migration and a stimulator of adhesion of cells to the ECM, is synthesized and released by corneal keratocytes into the extracellular matrix. When the cornea is wounded, the quiescent stromal keratocytes underlying the wound undergo apoptosis and cells adjacent to this apoptotic area convert to fibroblasts or myofibroblasts. This study explores the effect of extracellular maspin on the plasminogen-plasminogen activator system of corneal stromal cells following wou...

2011-01-01

54

Binding and activation of plasminogen at the surface of Fusobacterium necrophorum.  

Science.gov (United States)

Fusobacterium necrophorum is a gram-negative, anaerobic bacterium, which has been suggested to be a normal inhabitant of the oral flora. In rare cases, it can invade the tonsils and deeper tissues, causing the serious condition Lemièrre's syndrome. Recruitment of host plasminogen is a well-known bacterial virulence mechanism, and plasmin activity at the bacterial surface is thought to be important for bacterial invasion. Herein we show that plasminogen can be recruited to the surface of F. necrophorum, that surface-bound plasminogen is more easily converted to active plasmin than plasminogen in buffer, and that bound plasminogen is protected against inactivation by ?2-antiplasmin. These findings add to the understanding of the pathogenesis of Lemièrre's syndrome. PMID:23583624

Holm, Karin; Rasmussen, Magnus

2013-01-01

55

Tetranectin binds hepatocyte growth factor and tissue-type plasminogen activator.  

Science.gov (United States)

In the search for new ligands for the plasminogen kringle 4 binding-protein tetranectin, it has been found by ligand blot analysis and ELISA that tetranectin specifically bound to the plasminogen-like hepatocyte growth factor and tissue-type plasminogen activator. The dissociation constants of these complexes were found to be within the same order of magnitude as the one for the plasminogen-tetranectin complex. The study also revealed that tetranectin did not interact with the kindred proteins: macrophage-stimulating protein, urokinase-type plasminogen activator and prothrombin. In order to examine the function of tetranectin, a kinetic analysis of the tPA-catalysed plasminogen activation was performed. The kinetic parameters of the tetranectin-stimulated enhancement of tPA were comparable to fibrinogen fragments, which are so far the best inducer of tPA-catalysed plasminogen activation. The enhanced activation was suggested to be caused by tetranectin's ability to bind and accumulate tPA in an active conformation. PMID:12694198

Westergaard, Uffe B; Andersen, Mikkel H; Heegaard, Christian W; Fedosov, Sergey N; Petersen, Torben E

2003-04-01

56

Structural and functional peculiarities of plasminogen activator inhibitor PAI-1  

Directory of Open Access Journals (Sweden)

Full Text Available PAI-1, an important component of the hemostasis system, is a specific inhibitor of both urokinase type and tissue type plasminogen activators. PAI-1 belongs to the serpin family. The interaction between somatomedin-like domain of vitronectin and PAI-1 leads to stabilization of the latter. PAI-1 latency transition is related to the conformational changes in the reactive central loop. The inhibitory mechanism of PAI-1 is in accordance with the classic scheme of serpin action. PAI-1 blocks the adhesion mediated by UPA and integrins, so this inhibitor plays an important role in adhesion process and angiogenesis. An altered PAI-1level is associated with the development of cardiovascular diseases, kidney fibrosis, diabetis, cancerogenesis.

Kondratuk A. S.

2010-07-01

57

Simvastatin suppresses dexamethasone-induced secretion of plasminogen activator inhibitor-1 in human bone marrow adipocytes  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Background Osteonecrosis of the femoral head is a common complication of high-dose glucocorticoid treatment. Intravascular thrombosis is thought to be associated with the ischemic state of the femoral head. Plasminogen activator inhibitor-1 (PAI-1) is an adipokine, which are physiologically active substances secreted from visceral and subcutaneous adipocytes. PAI-1 suppresses fibrinolysis by binding tissue-type plasminogen activator. Several reports have described th...

2011-01-01

58

Analysis of five streptokinase formulations using the euglobulin lysis test and the plasminogen activation assay  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Streptokinase, a 47-kDa protein isolated and secreted by most group A, C and G ß-hemolytic streptococci, interacts with and activates human protein plasminogen to form an active complex capable of converting other plasminogen molecules to plasmin. Our objective was to compare five streptokinase formulations commercially available in Brazil in terms of their activity in the in vitro tests of euglobulin clot formation and of the hydrolysis of the plasmin-specific substrate S-2251™. Euglo...

Couto, L. T.; Donato, J. L.; Nucci, G.

2004-01-01

59

Soluble urokinase plasminogen activator receptor is associated with subclinical organ damage and cardiovascular events  

DEFF Research Database (Denmark)

The soluble urokinase plasminogen activator receptor (suPAR) is a plasma marker of low grade inflammation and has been associated with cardiovascular risk. We wanted to investigate whether suPAR was associated with markers of subclinical organ damage.

Sehestedt, T; Lyngbæk, S

2011-01-01

60

A Plasminogen Activator Inhibitor-1 Promoter Polymorphism and Idiopathic Interstitial Pneumonia  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The normal fibrinolytic activity within the alveolar space is suppressed in fibrotic lung diseases in part because of increased levels of plasminogen activator inhibitor-1 (PAI-1). Studies with animals have shown that inhibition of the plasminogen system by PAI-1 increases the generation of pulmonary fibrosis. To determine if a similar relationship occurs in human fibrotic lung diseases, we took advantage of a polymorphism (4G/5G) that occurs in the promoter region of the human PAI-1 gene and...

Kim, Kevin K.; Flaherty, Kevin R.; Long, Qi; Hattori, Noboru; Sisson, Thomas H.; Colby, Thomas V.; Travis, William D.; Martinez, Fernando J.; Murray, Susan; Simon, Richard H.

2003-01-01

 
 
 
 
61

Increased expression of urokinase plasminogen activator and its cognate receptor in human seminomas  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Background The urokinase plasminogen activating system (uPAS) is implicated in neoplastic progression and high tissue levels of uPAS components correlate with a poor prognosis in different human cancers. Despite that, relative few studies are available on the expression and function of the uPAS components in human seminomas. In the present study we characterized the expression of the urokinase plasminogen activator (uPA), its cognate receptor (uPAR) and the uPA inhib...

Ulisse Salvatore; Baldini Enke; Mottolese Marcella; Sentinelli Steno; Gargiulo Patrizia; Valentina Brancato; Sorrenti Salvatore; Di Benedetto Anna; De Antoni Enrico; Armiento Massimino, D.

2010-01-01

62

New role of plasminogen activator inhibitor-1 in alcohol-induced liver injury  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator inhibitor-1 (PAI-1) is the main inhibitor of plasminogen activators, thereby playing a major role in fibrinolysis. Whereas hyperfibrinolysis is common in alcoholic cirrhosis, hypofibrinolysis (driven mostly by elevated levels of PAI-1) is common during the development of alcoholic liver disease (ALD). However, whether or not PAI-1 plays a causal role in the development of ALD has been unclear. The role of PAI-1 was therefore investigated in models of early (steatosis), i...

Arteel, Gavin E.

2008-01-01

63

Targeting the urokinase plasminogen activator receptor enhances gene transfer to human airway epithelia  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Developing gene therapy for cystic fibrosis has been hindered by limited binding and endocytosis of vectors by human airway epithelia. Here we show that the apical membrane of airway epithelia express the urokinase plasminogen activator receptor (uPAR). Urokinase plasminogen activator (uPA), or a 7-residue peptide derived from this protein (u7-peptide), bound the receptor and stimulated apical endocytosis. Both ligands enhanced gene transfer by nonspecifically bound adenovirus and adeno-assoc...

Drapkin, Paola T.; O’riordan, Catherine R.; Yi, Su Min; Chiorini, John A.; Cardella, Jonathan; Zabner, Joseph; Welsh, Michael J.

2000-01-01

64

Synergistic fibrinolysis: combined effects of plasminogen activators and an antibody that inhibits alpha 2-antiplasmin.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

To improve the efficacy of plasminogen activators, we produced a monoclonal antibody (RWR) that inhibits human alpha 2-antiplasmin (alpha 2AP). In addition to inhibiting alpha 2AP in plasma, RWR binds to and inhibits fibrin cross-linked alpha 2AP and reproduces the "spontaneous" clot lysis that is the hallmark of human alpha 2AP deficiency. By inhibiting the inactivation of plasmin by alpha 2AP, RWR interacts synergistically with plasminogen activators to increase the potency (for 50%...

Reed, G. L.; Matsueda, G. R.; Haber, E.

1990-01-01

65

Simple and sensitive assay employing stable reagents for quantification of plasminogen activator  

Energy Technology Data Exchange (ETDEWEB)

A simple and sensitive indirect assay for quantifying plasminogen activator using (/sup 3/H)-casein as substrate is described. The assay has been used to measure plasminogen activators from various sources including bacteria, cultured cells, and human plasma. The assay is linear with respect to concentration of plasminogen activator and time of incubation and is independent of concentration of plasminogen. The assay can routinely be used to quantify as few as 8 X 10(-3) international units ml-1 of streptokinase. Results obtained with the (/sup 3/H)-casein assay correlate well with those obtained by the fibrin plate method (R . 0.83 by Spearman's ranked co-efficient of correlation). The reagents are extremely stable and easily stored. The ease with which the assay can be performed gives the clinical laboratory the potential to test large numbers of patients upon short notice and within a short period of time.

Lewis, J.G.; Pizzo, S.V.; Adams, D.O.

1981-01-01

66

Simple and sensitive assay employing stable reagents for quantification of plasminogen activator  

International Nuclear Information System (INIS)

A simple and sensitive indirect assay for quantifying plasminogen activator using ["3H]-casein as substrate is described. The assay has been used to measure plasminogen activators from various sources including bacteria, cultured cells, and human plasma. The assay is linear with respect to concentration of plasminogen activator and time of incubation and is independent of concentration of plasminogen. The assay can routinely be used to quantify as few as 8 X 10(-3) international units ml-1 of streptokinase. Results obtained with the ["3H]-casein assay correlate well with those obtained by the fibrin plate method (R . 0.83 by Spearman's ranked co-efficient of correlation). The reagents are extremely stable and easily stored. The ease with which the assay can be performed gives the clinical laboratory the potential to test large numbers of patients upon short notice and within a short period of time

1981-01-01

67

Immunologic detection of the cellular receptor for urokinase plasminogen activator.  

Science.gov (United States)

The cellular receptor for urokinase plasminogen activator (uPA-R) is a monomeric phosphatidylinositol-linked glycoprotein (gp40-65) that may contribute to the invasive capacity of tumor and inflammatory cells by focusing the activity of urokinase (uPA) in converting plasminogen to plasmin, a serine protease capable of degrading extracellular matrix proteins. The further characterization of uPA-R has been facilitated by our recent development of a monoclonal antibody, anti-Mo3f, specific for uPA-R. This mAb bound to uPA-R expressed by phorbol myristate acetate-stimulated U-937 cells and by NIH-3T3 cells permanently transfected with uPA-R cDNA. In competitive binding assays, anti-Mo3f inhibited the binding of fluorescein-conjugated uPA ligand to uPA-R expressed by U-937 cells and uPA-R transfectants; conversely, preexposure of cells to saturating quantities of exogenous uPA partially blocked the subsequent binding of anti-Mo3f mAb to uPA-R. Anti-Mo3f mAb was employed as the capture reagent in an ELISA for the quantitation of soluble forms of uPA-R (derived from U-937 cells and recombinant uPA-R) which had a sensitivity of approximately 4-12 ng/ml. Anti-Mo3f mAb was also applied as a serologic probe for the detection of uPA-R expressed by human tumor tissues. By immunoperoxidase staining, anti-Mo3f demonstrated positive tumor cell staining in 4 of 16 breast and 7 of 31 prostate carcinomas in formalin-fixed, paraffin-embedded specimens. These data indicate that the anti-Mo3f mAb detects an epitope proximate to or within the ligand binding domain (domain 1) of uPA-R and may be useful as a tool for the serologic detection of uPA-R in soluble form or associated with human tumors. PMID:8137563

Mizukami, I F; Garni-Wagner, B A; DeAngelo, L M; Liebert, M; Flint, A; Lawrence, D A; Cohen, R L; Todd, R F

1994-04-01

68

Tissue plasminogen activator (tPA) inhibits plasmin degradation of fibrin. A mechanism that slows tPA-mediated fibrinolysis but does not require alpha 2-antiplasmin or leakage of intrinsic plasminogen.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Thrombolysis is dramatically slower when high concentrations of lytic agent are used. This paradoxical observation, first described as "plasminogen steal," was originally believed to be due to depletion of extrinsic plasminogen and consequent leaching of clot-bound plasminogen. We report that administration of increasing concentrations of recombinant human tissue plasminogen activator (tPA) to fibrin gels resulted in lysis rates that displayed a maximum, with significantly slower rate...

Wu, J. H.; Diamond, S. L.

1995-01-01

69

Enhanced expression of plasminogen activator inhibitor-1 by dedifferentiated thyrocytes.  

Science.gov (United States)

Porcine thyrocytes in vitro in the presence of TSH adopt follicular-like morphology. Epidermal growth factor, phorbol esters or transforming growth factor beta-1 (TGFbeta-1) induce a rapid spreading of the cells and dedifferentiation. In addition to thyroglobulin, dedifferentiated thyrocytes secreted into the culture medium three proteins in abundant quantities. Two of them have been previously identified as thrombospondin-1 and clusterin, respectively. Using the microsequencing method we identified the third one, a M(r) 45,000 glycosylated protein, as plasminogen activator inhibitor-1 (PAI-1). EGF, phorbol esters or TGF-beta1 predominantly increased PAI-1 protein expression in TSH-treated cells. The maximal increase of PAI-1 mRNA steady-state level was observed 6 h after EGF treatment and sustained up to 48 h. Recombinant PAI-1 inhibited cell-associated plasmin activity and delayed cell spreading. Enhanced synthesis and secretion of PAI-1 upon treatment with different growth factors during dedifferentiation process and spreading may be considered a feed-back defence mechanism of the cells to harmful extracellular stimuli. PMID:12099701

Kotlarz, Grazyna; Wegrowski, Yanusz; Martiny, Laurent; Declerck, Paul J; Bellon, Georges

2002-07-19

70

Tumour cell thrombospondin-1 regulates tumour cell adhesion and invasion through the urokinase plasminogen activator receptor  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We have previously shown that platelet-produced thrombospondin-1 up-regulates the urokinase plasminogen activator and its receptor and promotes tumour cell invasion. Although tumour cells produce thrombospondin-1 in vivo, they produce only minimal amounts of thrombospondin-1 in vitro. To determine the effect of tumour cell-produced thrombospondin-1 in the regulation of the plasminogen/plasmin system and tumour cell invasion, we studied THBS-1 -transfected MDA-MB-435 breast cancer cells that o...

Albo, D.; Rothman, V. L.; Roberts, D. D.; Tuszynski, G. P.

2000-01-01

71

Isolation from human plasma of a plasminogen activator identical to urinary high molecular weight urokinase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Two different plasmatic plasminogen activators (PA) can be demonstrated after sodium dodecyl sulfate polyacrylamide gel electrophoresis of plasma freshly collected from resting volunteers, followed by transfer of the gels onto plasminogen-rich fibrin-agarose plates. These two PA are also present in plasmas deficient in coagulation Factor XI, Factor XII, prekallikrein, or high molecular weight-kininogen. The slower-moving PA has an apparent 85,000 Mr and is immunologically unrelated to urokina...

Tissot, J. D.; Schneider, P.; Hauert, J.; Ruegg, M.; Kruithof, E. K.; Bachmann, F.

1982-01-01

72

Pneumatic displacement without tissue plasminogen activator in premacular subhyaloid hemorrhage  

Directory of Open Access Journals (Sweden)

Full Text Available To assess the efficacy and safety of intravitreal injection of Sulfur Hexafluoride (SF6 gas without the use of tissue Plasminogen Activator (tPA in premacular Subhyaloid Hemorrhage (SHH, 5 eyes of 5 patients with premacular SHH were enrolled. After performing paracentesis of the anterior chamber, 0.3 ml pure SF6 gas was injected through pars plana with a 30 gauge needle. Facedown position was maintained for 5 days. Subhyaloid Hemorrhage was displaced in 4/5 (80% eyes with a duration of SHH less than 2 weeks. The pre-injection visual acuity of all 5 eyes was finger counting and improved in 4/5 ( 80% eyes within 3 days to 7 days post-injection to 6/20 - 6/6. The underlying disease was hypercoagulation in 1 patient, diabetes mellitus in 2 patients, hypertension in 1 patient and unknown in 1 patient. No complications were encountered. In conclusion, SF6 gas injected into the vitreous without the use of tPA, can displace SHH if performed within 14 days of duration, and results in rapid visual recovery. This procedure is proven to be safe. (Med J Indones 2007; 16:104-7 Keywords: subhyaloid hemorrhage, pneumatic displacement, sulfur hexafluoride gas

Rumita S. Kadarisman

2007-06-01

73

Biochemical Importance of Glycosylation of Plasminogen Activator Inhibitor-1  

DEFF Research Database (Denmark)

The serpin plasminogen activator inhibitor-1 (PAI-1) is a potential target for anti-thrombotic and anti-cancer therapy. PAI-1 has 3 potential sites for N-linked glycosylation. We demonstrate here that PAI-1 expressed recombinantly or naturally by human cell lines display a heterogeneous glycosylation pattern of the sites at N209 and N265, while that at N329 is not utilised. The IC(50)-values for inactivation of PAI-1 by 4 monoclonal antibodies differed strongly between glycosylated PAI-1 and non-glycosylated PAI-1 expressed in E. coli. For 3 antibodies, an overlap of the epitopes with the glycosylation sites could be excluded as explanation for the differential reactivity. The latency transition of non-glycosylated, but not of glycosylated PAI-1, was strongly accelerated by a non-ionic detergent. The different biochemical properties of glycosylated and non-glycosylated PAI-1 depended specifically on glycosylation of either one or the other of the utilised sites. The PAI-1-binding protein vitronectin reversed the changes associated with the lack of glycosylation at one of the sites. Our results stress the importance of the source of PAI-1 when studying the mechanisms of action of PAI-1-inactivating compounds of potential clinical importance.

Gils, Ann; Pedersen, Katrine Egelund

2003-01-01

74

Early increases in plasminogen activator activity following partial hepatectomy in humans  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Background Increases in urokinase-like plasminogen activator (uPA) activity are reported to be amongst the earliest events occurring in remnant liver following partial hepatectomy in rats, and have been proposed as a key component of the regenerative response. Remodelling of the extracellular matrix, conversion of single chain hepatocyte growth factor to the active two-chain form and a possible activation of a mitogenic signalling pathway have all been ascribed to th...

Mangnall David; Smith Kirsty; Bird Nigel C; Majeed Ali W

2004-01-01

75

Breast cancer prognosis is poor when total plasminogen activator activity is low.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator (PA) is a serine protease which exists in two forms: tissue-type (t-PA) and urokinase-type (u-PA). The total PA activity was measured in tumour extracts of 235 breast cancer patients who were followed for a median of 8.5 years after surgery. Patients were initially divided into three groups with low ( 300 unit mg-1 protein) total PA activity in tumour extracts. The PA activity was not significan...

Yamashita, J.; Ogawa, M.; Inada, K.; Yamashita, S.; Nakashima, Y.; Saishoji, T.; Nomura, K.

1993-01-01

76

Beta subunits of human choriogonadotropin and ovine lutropin are biologically active.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Highly purified beta subunits of human choriogonadotropin and ovine lutropin were found to possess measurable steroidogenic activity in the in vitro rat Leydig cell assay. In addition, beta subunits of these two gonadotropins were able to inhibit the biological activity of human choriogonadotropin in the rat Leydig cell assay.

1982-01-01

77

Complexes between tissue-type plasminogen activator and proteinase inhibitors in human plasma, identified with an immunoradiometric assay  

International Nuclear Information System (INIS)

Extrinsic (tissue-type) plasminogen activator antigen in human plasma, as measured by a two-site immunoradiometric assay, is composed of a fibrin-adsorbable and a nonadsorbable fraction. Gel filtration on Ultrogel AcA 44 in 1.6M KSCN of the fibrin-adsorbable fraction showed a peak with M/sub r/ approx. =70,000, which contained plasminogen activator activity and was assumed to represent free extrinsic plasminogen activator. The nonadsorbable fraction showed a broad peak with M/sub r/ approx. =140,000 without plasminogen activator activity. Overnight incubation at 370C of postexercise plasma revealed a shift of the M/sub r/ approx. =70,000 peak to the M/sub r/ approx. =140,000 position, suggesting that the M/sub r/ approx. =140,000 peak consists of extrinsic plasminogen activator-protease inhibitor complex(es). ?2-Antiplasmin is the main inhibitor of extrinsic plasminogen activator in plasma and is probably responsible for the generation of the M/sub r/ approx. =140,000 component. A possible involvement of other plasma proteinase inhibitors was explored by incubation of 125I-labeled extrinsic plasminogen activator in ?2-antiplasmin-depleted plasma. A complex was formed with a t1/2 of about 1 hr, which was identified by immunoprecipitation as extrinsic plasminogen activator-?2-antiplasmin complex. Additional evidence for the presence of extrinsic plasminogen activator complexes with ?2-antiplasmin and ?1-antitrypsin in plasma was obtained from two-site immunoradiometric assays. It was concluded that plasma contains both free extrinsic plasminogen activator and plasminogen activator complexes with ?2-antiplasmin and ?1-antitrypsin. These complexes are also present in plasma collected on the active site inhibitor, D-Phe-Pro-Arg-CH2Cl, at rest and after exercise and are therefore assumed to circulate in vivo

1983-01-01

78

Plasminogen activator activity during growth and differentiation of human HL-60 promyelocytes  

International Nuclear Information System (INIS)

The involvement of plasminogen activators (PA) of urokinase or tissue type which activate plasminogen to plasmin have been implicated in fibrinolysis, differentiation and cancer growth. In the present study, alteration of PA activity of human HL-60 promyelocytes during growth and differentiation has been investigated. HL-60 promyelocytes seeded at a density of 1 x 105 cells/ml, were examined at different densities of growth (0.2-2.0 x 106 cells/ml for PA activity, measured as plasminogen dependent hydrolysis of 125I labelled fibrin or D-Val-L-Leu-L-Lys-pNA. 125I-fibrin is hydrolyzed at a linear rate between 2-6 hours and at cell concentration of .2-1 x 106 per ml. The cellular PA activity per 106 cells at different growth densities decreases by 50% between .2-2 x 106 cells/ml. Similarly, when cells are differentiated to monocytes-macrophages by 1,25-dihydroxyvitamin D3 (as determined by NBT reduction), a decrease in PA activity is observed which parallels the degree of differentiation. The major alteration of PA activity is in the heavy membrane fraction of the differentiated cells. These studies suggest that PA levels may be important in the growth and differentiation of HL-60 cells

1986-05-01

79

Plasminogen activator activity during growth and differentiation of human HL-60 promyelocytes  

Energy Technology Data Exchange (ETDEWEB)

The involvement of plasminogen activators (PA) of urokinase or tissue type which activate plasminogen to plasmin have been implicated in fibrinolysis, differentiation and cancer growth. In the present study, alteration of PA activity of human HL-60 promyelocytes during growth and differentiation has been investigated. HL-60 promyelocytes seeded at a density of 1 x 10/sup 5/ cells/ml, were examined at different densities of growth (0.2-2.0 x 10/sup 6/ cells/ml for PA activity, measured as plasminogen dependent hydrolysis of /sup 125/I labelled fibrin or D-Val-L-Leu-L-Lys-pNA. /sup 125/I-fibrin is hydrolyzed at a linear rate between 2-6 hours and at cell concentration of .2-1 x 10/sup 6/ per ml. The cellular PA activity per 10/sup 6/ cells at different growth densities decreases by 50% between .2-2 x 10/sup 6/ cells/ml. Similarly, when cells are differentiated to monocytes-macrophages by 1,25-dihydroxyvitamin D/sub 3/ (as determined by NBT reduction), a decrease in PA activity is observed which parallels the degree of differentiation. The major alteration of PA activity is in the heavy membrane fraction of the differentiated cells. These studies suggest that PA levels may be important in the growth and differentiation of HL-60 cells.

Kumar, S.; Russo, M.S.

1986-05-01

80

Plasminogen activator inhibitor-1 regulates myoendothelial junction formation  

Science.gov (United States)

Rationale Plasminogen activator inhibitor-1 (PAI-1) is a biomarker for several vascular disease states; however, its target of action within the vessel wall is undefined. Objective Determine the ability of PAI-1 to regulate myoendothelial junction (MEJ) formation. Methods and Results Myoendothelial junctions are found throughout the vasculature linking endothelial cells (EC) and vascular smooth muscle cells (VSMC). Using a vascular cell co-culture (VCCC) we isolated MEJ fractions and performed two-dimensional differential gel electrophoresis. Mass spectrometry identified PAI-1 as being enriched within MEJ fractions, which we confirmed in vivo. In the VCCC, recombinant PAI-1 (rPAI-1) added to the EC monolayer significantly increased MEJs. Conversely, addition of a PAI-1 monoclonal antibody to the EC monolayer reduced the number of MEJs. This was also observed in vivo where mice fed a high fat diet had increased PAI-1 and MEJs and the number of MEJs in coronary arterioles of PAI-1?/? mice was significantly reduced when compared to C57Bl/6 mice. The presence of MEJs in PAI-1?/? coronary arterioles was restored when their hearts were transplanted into and exposed to the circulation of C57Bl/6 mice. Application of biotin-conjugated PAI-1 to the EC monolayer in vitro confirmed the ability of luminal PAI-1 to translocate to the MEJ. Functionally, phenylephrine-induced heterocellular calcium communication in the VCCC was temporally enhanced when rPAI-1 was present, and prolonged when PAI-1 was absent. Conclusion Our data implicate circulating PAI-1 as a key regulator of MEJ formation and a potential target for pharmacological intervention in diseases with vascular abnormalities (e.g., diabetes mellitus).

Heberlein, Katherine; Straub, Adam C.; Best, Angela K.; Greyson, Mark A.; Looft-Wilson, Robin C.; Sharma, Poonam R.; Meher, Akshaya; Leitinger, Norbert; Isakson, Brant E

2010-01-01

 
 
 
 
81

Stereotactic evacuation of hypertensive cerebellar hemorrhage using plasminogen activator.  

Science.gov (United States)

Treatment for hypertensive cerebellar hemorrhage still remains controversial as to whether direct surgical procedure is indicated or not. This is so even after the introduction of CT scan which easily demonstrates the location and size of the hematoma and the presence of hydrocephalus. In this paper, we present our experience of 20 patients with cerebellar hemorrhage treated by stereotactic evacuation using Komai's CT-stereotactic apparatus. All the patients had vertigo, cerebellar symptoms, dysfunction of brain stem or consciousness disturbance. The hematomas on CT scan were more than 28 mm in diameter. Acute obstructive hydrocephalus occurred in 90% of the patients with hematoma 40 mm or larger in size. The patients with consciousness disturbance were immediately operated on after the attack, and a drainage tube was placed in the hematoma cavity to drain cerebrospinal fluid and liquefied hematoma for one to eight days. On the other hand, when patients with hematoma around 30 mm in diameter complained vertigo for about two weeks, they also were operated on stereotactically. After the operation, their symptoms improved rapidly. The stereotactic operation could aspirate about 85% of the estimated hematoma volume and improved the hydrocephalus, except in one case in which the patient rapidly deteriorated to coma level with a large cerebellar hemorrhage and brain stem damage. This stereotactic evacuation of cerebellar hematoma using a plasminogen activator is effective for not only the removal of hematoma, but also for the treatment of secondary hydrocephalus following obstruction of the fourth ventricle by cerebellar hemorrhage.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2674756

Yokote, H; Komai, N; Nakai, E; Ueno, M; Hayashi, S; Terashita, T

1989-05-01

82

Recombinant tissue plasminogen activator in the treatment of suprachoroidal hemorrhage  

Directory of Open Access Journals (Sweden)

Full Text Available Nancy Kunjukunju1, Christine R Gonzales2, William S Rodden21Ochsner Medical Center, New Orleans, Louisiana; 2Retina and Vitreous Center of Southern Oregon, Ashland, Oregon, USABackground: Suprachoroidal hemorrhages are a vision-threatening complication, and poor visual outcome is correlated with increasing hemorrhage complexity. The recommended time of surgical drainage is 10–14 days after the hemorrhage begins to liquefy. We describe a case in which recombinant tissue plasminogen activator (r-tPA, alteplase, is injected within the suprachoroidal space before surgery to assist in the drainage of an organized clot prior to liquefaction. This is a report of a technique in which r-tPA is used in the intrachoroidal space to target the organized clot of suprachoroidal hemorrhage prior to drainage.Case report: A 62-year-old male presented 12 days after retinal detachment repair with sudden ocular pain and vision loss after a Valsalva maneuver. Vision was light perception only, and intraocular pressure was 43 mmHg. Diagnosed with hyphema and suprachoroidal hemorrhage, the patient underwent surgery the following day. An injection of r-tPA 100 µg was given intracamerally, and an additional dose of r-tPA 100 µg was injected into the suprachoroidal space prior to surgery. Liquified by r-tPA, the clot was expressed through the sclerotomies. Best corrected vision in the eye eight months after the drainage procedure was 20/40.Conclusion: To the author’s knowledge, this is the first reported case in which r-tPA was successfully injected in the suprachoroidal space to liquefy and drain a suprachoroidal hemorrhage prior to natural dissolution.Keywords: tPA, suprachoroidal hemorrhage, vision loss

Nancy Kunjukunju

2011-02-01

83

Hormone control of total plasminogen activator activity is specific to malignant DMBA-induced rat mammary tumours.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Hormonal regulation of plasminogen activator expression in 7,12-dimethylbenz[a]anthracene (DMBA)--induced rat mammary carcinomas was studied both in vivo and in vitro and was compared to that in DMBA-mammary dysplasia induced in neonatally androgenised rats. The plasminogen activator activity in DMBA-mammary carcinomas, but not in DMBA-mammary dysplasia, was regulated by oestrogen. This suggests that expression of this enzyme is hormonally regulated in carcinoma cells. Furthermore, in two of ...

Inada, K.; Yamashita, J.; Matsuo, S.; Nakashima, Y.; Yamashita, S.; Ogawa, M.

1992-01-01

84

Post-Transcriptional Regulation of Plasminogen Activator Inhibitor Type–1 Expression in Human Pleural Mesothelial Cells  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The plasminogen activator inhibitor type–1 (PAI-1) effectively blocks the activities of free and receptor-bound urokinase-type plasminogen activator. Incubation of cultured human pleural mesothelial (Met5A) cells with TGF-? increased PAI-1 protein. TGF-?, phorbol myristate acetate, and the translation inhibitor cycloheximide induced PAI-1 mRNA and slowed its degradation, suggesting that PAI-1 mRNA could be regulated by interaction of a PAI-1 binding protein (PAI-1 mRNABp) with PAI-1 mRNA....

2010-01-01

85

Inhibitors of Urokinase Type Plasminogen Activator and Cytostatic Activity from Crude Plants Extracts  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In view of the clear evidence that urokinase type plasminogen activator (uPA) plays an important role in the processes of tumor cell metastasis, aortic aneurysm, and multiple sclerosis, it has become a target of choice for pharmacological intervention. The goal of this study was thus to determine the presence of inhibitors of uPA in plants known traditionally for their anti-tumor properties. Crude methanol extracts were prepared from the leaves of plants (14) collected from the subtropical dr...

Xueqiang Zha; Ricardo Diaz; Jose Javier Rosado Franco; Veronica Forbes Sanchez; Ezio Fasoli; Gabriel Barletta; Augusto Carvajal; Vibha Bansal

2013-01-01

86

A molecular variant of plasminogen activator inhibitor of rat decidual tissue.  

Science.gov (United States)

Artificially induced rat decidual tissue expresses plasminogen activator inhibitor (PAI). This PAI, isolated and purified employing chromatographic techniques, is a low molecular weight protein unlike the known PAIs. The final purified preparation resolves into a single band following SDS-PAGE and has an approximate molecular weight of 29 kDa. The properties studied include specificity for urokinase-type (uPA) and tissue-type (tPA) plasminogen activators, binding to conA and heparin, inhibition of thrombin, plasmin and trypsin. Decidual PAI is immunogenic in rabbit and a monospecific antiserum raised against the decidual inhibitor cross reacts with an extract of human placenta. PMID:8747533

Tarachand, U; Pawse, A R

1995-12-01

87

Therapeutic Value of Small Molecule Inhibitor to Plasminogen Activator Inhibitor–1 for Lung Fibrosis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Fibrosis is a final stage of many lung diseases, with no effective treatment. Plasminogen activator inhibitor–1 (PAI-1), a primary inhibitor of tissue-type and urokinase-type plasminogen activators (tPA and uPA, respectively), plays a critical role in the development of fibrosis. In this study, we explored the therapeutic potential of an orally effective small molecule PAI-1 inhibitor, TM5275, in a model of lung fibrosis induced by transforming growth factor–?1 (TGF-?1), the most potent...

Huang, Wen-tan; Vayalil, Praveen K.; Miyata, Toshio; Hagood, James; Liu, Rui-ming

2012-01-01

88

Interaction study between nifedipine and recombinant tissue-type plasminogen activator in healthy subjects.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

1. In a previous study it was demonstrated that a decrease in liver blood flow produced a decrease in clearance of recombinant human tissue-type plasminogen activator (rt-PA). 2. The purpose of this randomized, double-blind, placebo-controlled, three-way, cross-over investigation was to determine the effect of nifedipine (20 mg orally), a compound that increases liver blood flow, on plasma concentrations of steady state endogenous and recombinant tissue-type plasminogen activator (t-PA and rt...

Boer, A.; Kluft, C.; Kasper, F. J.; Kroon, J. M.; Schoemaker, H. C.; Breimer, D. D.; Soons, P. A.; Cohen, A. F.

1993-01-01

89

Phenotypic overlap between MMP-13 and the plasminogen activation system during wound healing in mice  

DEFF Research Database (Denmark)

Proteolytic degradation of extracellular matrix is a crucial step in the healing of incisional skin wounds. Thus, healing of skin wounds is delayed by either plasminogen-deficiency or by treatment with the broad-spectrum metalloproteinase (MP) inhibitor Galardin alone, while the two perturbations combined completely prevent wound healing. Both urokinase-type plasminogen activator and several matrix metallo proteinases (MMPs), such as MMP-3, -9 and -13, are expressed in the leading-edge keratinocytes of skin wounds, which may account for this phenotypic overlap between these classes of proteases.

Juncker-Jensen, Anna; Lund, Leif R

2011-01-01

90

Plasminogen activator inhibitor type 1 regulates microglial motility and phagocytic activity  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Plasminogen activator inhibitor type 1 (PAI-1 is the primary inhibitor of urokinase type plasminogen activators (uPA and tissue type plasminogen activators (tPA, which mediate fibrinolysis. PAI-1 is also involved in the innate immunity by regulating cell migration and phagocytosis. However, little is known about the role of PAI-1 in the central nervous system. Methods In this study, we identified PAI-1 in the culture medium of mouse mixed glial cells by liquid chromatography and tandem mass spectrometry. Secretion of PAI-1 from glial cultures was detected by ELISA and western blotting analysis. Cell migration was evaluated by in vitro scratch-wound healing assay or Boyden chamber assay and an in vivo stab wound injury model. Phagocytic activity was measured by uptake of zymosan particles. Results The levels of PAI-1 mRNA and protein expression were increased by lipopolysaccharide and interferon-? stimulation in both microglia and astrocytes. PAI-1 promoted the migration of microglial cells in culture via the low-density lipoprotein receptor-related protein (LRP 1/Janus kinase (JAK/signal transducer and activator of transcription (STAT1 axis. PAI-1 also increased microglial migration in vivo when injected into mouse brain. PAI-1-mediated microglial migration was independent of protease inhibition, because an R346A mutant of PAI-1 with impaired PA inhibitory activity also promoted microglial migration. Moreover, PAI-1 was able to modulate microglial phagocytic activity. PAI-1 inhibited microglial engulfment of zymosan particles in a vitronectin- and Toll-like receptor 2/6-dependent manner. Conclusion Our results indicate that glia-derived PAI-1 may regulate microglial migration and phagocytosis in an autocrine or paracrine manner. This may have important implications in the regulation of brain microglial activities in health and disease.

Jeon Hyejin

2012-06-01

91

t-plasminogen activator inhibitor-1 polymorphism in idiopathic pulmonary arterial hypertension  

Directory of Open Access Journals (Sweden)

Full Text Available Aim: The aim of the present study was to identify the possible genotypic association of 3?UTR Hind III polymorphism of Plasminogen activator Inhibitor-1 (PAI-1 gene with idiopathic pulmonary arterial hypertension (IPAH. Background: IPAH is a disorder with abnormally raised mean pulmonary arterial pressure and increase in the resistance to blood flow in pulmonary artery. One of the pathological features seen is development of intraluminal thrombin deposition leading to thrombosis. Plasminogen activator inhibitor-1 is an important inhibitor of the fibrinolytic system; its up-regulation may suppress fibrinolysis and result in an increased risk of thrombosis. Method: Blood samples from 54 IPAH patients and 100 healthy voluntary donors were analyzed by PCR-RFLP method for 3?UTR Hind III polymorphism. Results and Disscussion: A significant association of Hd2 allele with the disease was observed. Raised mean level of right ventricular systolic pressure was observed in the Hd2/Hd2 genotypic patients, strengthening the role of Hd2 allele in the disease progression. Our data suggests an association of Hd2/Hd2 genotype, which may lead to the up-regulation of PAI-1 gene leading to increased levels of PAI-1, which is seen in IPAH. PAI-1 competes with plasminogen activators and hinders the normal mechanism of plasminogen activation system and leads to thrombosis and formation of plexiform lesions in the lung tissue, further strengthening its role in tissue remodeling and disease progression.

Katta Sujana

2008-01-01

92

Relationship between plasminogen activator inhibitor type-1 (PAI-1) gene polymorphisms and osteoporosis in Turkish women  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english OBJECTIVE: The development of osteoporosis is associated with several risk factors, such as genetic structures that affect bone turnover and bone mass. The impact of genetic structures on osteoporosis is not known. Plasminogen activator inhibitor type-1 regulates the bone matrix and bone balance. Th [...] is study assessed the correlation between plasminogen activator inhibitor type-1 gene 4G/5G polymorphisms and osteoporosis in a population of Turkish women. METHODS: A total of 195 postmenopausal female patients who were diagnosed with osteoporosis (Group I) based on bone mineral density measurements via dual-energy x-ray absorptiometry and 90 females with no osteoporosis (Group II) were included in this study. Correlations between PAI-1 gene 4G/5G polymorphisms and osteoporosis were investigated through the identification of PAI-1 gene 4G/5G polymorphism genotypes using the polymerase chain reaction. RESULTS: No significant differences in the genotype and allele frequency of 4G/5G plasminogen activator inhibitor type-1 polymorphisms were observed between the two groups, and both groups exhibited the most frequently observed 4G5G genotype. CONCLUSION: No correlation between the development of osteoporosis in the female Turkish population and 4G/5G plasminogen activator inhibitor type-1 gene polymorphisms was observed.

Merih, Ozgen; Didem Turgut, Cosan; Fulya, Doganer; Ahu, Soyocak; Onur, Armagan; Hasan Veysi, Gunes; Irfan, Degirmenci; Gulsah Ogutler, Ozkara; Fezan Sahin, Mutlu.

93

On the activity and specificity of cardosin B, a plant proteinase, on ovine caseins  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The proteolytic activity of cardosin B, an aspartic proteinase from the thistle, Cynara cardunculus, on ovine ?s-caseins and ?-caseins (independently or present together in sodium caseinate) was followed by urea polyacrylamide gel electrophoresis and reversed phase high performance liquid chromatography. This enzyme degraded both types of caseins, but not to the same degree. In sodium-caseinate, by 10 h at 30°C, ?s-caseins were more susceptible to proteolysis by cardosin B than ?-casein ...

Silva, Sofia V.; Malcata, F. Xavier

1999-01-01

94

NMR secondary structure of the plasminogen activator protein staphylokinase  

Energy Technology Data Exchange (ETDEWEB)

Staphylokinase (Sak) is a 15.5 kDa protein secreted by several strains of Staphylococcusaureus. Due to its ability to convert plasminogen, the inactive proenzyme of the fibrinolytic system, into plasmin, Sak is presently undergoing clinical trials for blood clot lysis in the treatment of thrombovascular disorders. With a view to developing a better understanding of the mode of action of Sak, we have initiated a structural investigation of Sak via multidimensional heteronuclear NMR spectroscopy employing uniformly 15N- and 15N, 13C-labelled Sak. Sequence-specific resonance assignments have been made employing 15N-edited TOCSY and NOE experiments and from HNCACB, CBCA(CO)NH, HBHA(CBCACO)NH and CC(CO)NH sets of experiments. From an analysis of the chemical shifts, 3JHNH{alpha} scalar coupling constants, NOEs and HN exchange data, the secondary structural elements of Sak have been characterized.

Ohlenschlaeger, O.; Ramachandran, R.; Flemming, J.; Guehrs, K.-H.; Schlott, B.; Brown, L.R. [Institute of Molecular Biotechnology, Department of Molecular Biophysics/NMR Spectroscopy (Germany)

1997-04-15

95

NMR secondary structure of the plasminogen activator protein staphylokinase  

International Nuclear Information System (INIS)

Staphylokinase (Sak) is a 15.5 kDa protein secreted by several strains of Staphylococcusaureus. Due to its ability to convert plasminogen, the inactive proenzyme of the fibrinolytic system, into plasmin, Sak is presently undergoing clinical trials for blood clot lysis in the treatment of thrombovascular disorders. With a view to developing a better understanding of the mode of action of Sak, we have initiated a structural investigation of Sak via multidimensional heteronuclear NMR spectroscopy employing uniformly 15N- and 15N, 13C-labelled Sak. Sequence-specific resonance assignments have been made employing 15N-edited TOCSY and NOE experiments and from HNCACB, CBCA(CO)NH, HBHA(CBCACO)NH and CC(CO)NH sets of experiments. From an analysis of the chemical shifts, 3JHNH? scalar coupling constants, NOEs and HN exchange data, the secondary structural elements of Sak have been characterized

1997-04-01

96

Inhibition of plasminogen activator inhibitor type-1 activity enhances rapid and sustainable hematopoietic regeneration.  

Science.gov (United States)

The prognosis of patients undergoing hematopoietic stem cell transplantation (HSCT) depends on the rapid recovery and sustained life-long hematopoiesis. The activation of the fibrinolytic pathway promotes hematopoietic regeneration; however, the role of plasminogen activator inhibitor-1 (PAI-1), a negative regulator of the fibrinolytic pathway, has not yet been elucidated. We herein demonstrate that bone marrow (BM) stromal cells, especially osteoblasts, produce PAI-1 in response to myeloablation, which negatively regulates the hematopoietic regeneration in the BM microenvironment. Total body irradiation in mice dramatically increased the local expression levels of fibrinolytic factors, including tissue-type plasminogen activator (tPA), plasmin, and PAI-1. Genetic disruption of the PAI-1 gene, or pharmacological inhibition of PAI-1 activity, significantly improved the myeloablation-related mortality and promoted rapid hematopoietic recovery after HSCT through the induction of hematopoiesis-promoting factors. The ability of a PAI-1 inhibitor to enhance hematopoietic regeneration was abolished when tPA-deficient mice were used as recipients, thus indicating that PAI-1 represses tPA-dependent hematopoietic regeneration. The PAI-1 inhibitor not only accelerated the expansion of the donor HSCs during the early-stage of regeneration, but also supported long-term hematopoiesis. Our results indicate that the inhibition of PAI-1 activity could be a therapeutic approach to facilitate the rapid recovery and sustained hematopoiesis after HSCT. PMID:24155177

Ibrahim, Abd Aziz; Yahata, Takashi; Onizuka, Makoto; Dan, Takashi; Van Ypersele De Strihou, Charles; Miyata, Toshio; Ando, Kiyoshi

2014-04-01

97

The voltage-dependent anion channel (VDAC) binds tissue-type plasminogen activator and promotes activation of plasminogen on the cell surface.  

Science.gov (United States)

The voltage-dependent anion channel (VDAC), a major pore-forming protein in the outer membrane of mitochondria, is also found in the plasma membrane of a large number of cells where in addition to its role in regulating cellular ATP release and volume control it is important for maintaining redox homeostasis. Cell surface VDAC is a receptor for plasminogen kringle 5, which promotes partial closure of the channel. In this study, we demonstrate that VDAC binds tissue-type plasminogen activator (t-PA) on human neuroblastoma SK-N-SH cells. Binding of t-PA to VDAC induced a decrease in K(m) and an increase in the V(max) for activation of its substrate, plasminogen (Pg). This resulted in accelerated Pg activation when VDAC, t-PA, and Pg were bound together. VDAC is also a substrate for plasmin; hence, it mimics fibrin activity. Binding of t-PA to VDAC occurs between a t-PA fibronectin type I finger domain located between amino acids Ile(5) and Asn(37) and a VDAC region including amino acids (20)GYGFG(24). These VDAC residues correspond to a GXXXG repeat motif commonly found in amyloid ? peptides that is necessary for aggregation when these peptides form fibrillar deposits on the cell surface. Furthermore, we also show that Pg kringle 5 is a substrate for the NADH-dependent reductase activity of VDAC. This ternary complex is an efficient proteolytic complex that may facilitate removal of amyloid ? peptide deposits from the normal brain and cell debris from injured brain tissue. PMID:23161549

Gonzalez-Gronow, Mario; Ray, Rupa; Wang, Fang; Pizzo, Salvatore V

2013-01-01

98

Analysis of five streptokinase formulations using the euglobulin lysis test and the plasminogen activation assay  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Streptokinase, a 47-kDa protein isolated and secreted by most group A, C and G ß-hemolytic streptococci, interacts with and activates human protein plasminogen to form an active complex capable of converting other plasminogen molecules to plasmin. Our objective was to compare five streptokinase form [...] ulations commercially available in Brazil in terms of their activity in the in vitro tests of euglobulin clot formation and of the hydrolysis of the plasmin-specific substrate S-2251™. Euglobulin lysis time was determined using a 96-well microtiter plate. Initially, human thrombin (10 IU/ml) and streptokinase were placed in individual wells, clot formation was initiated by the addition of plasma euglobulin, and turbidity was measured at 340 nm every 30 s. In the second assay, plasminogen activation was measured using the plasmin-specific substrate S-2251™. Streptase™ was used as the reference formulation because it presented the strongest fibrinolytic activity in the euglobulin lysis test. The Unitinase™ and Solustrep™ formulations were the weakest, showing about 50% activity compared to the reference formulation. All streptokinases tested activated plasminogen but significant differences were observed. In terms of total S-2251™ activity per vial, Streptase™ (75.7 ± 5.0 units) and Streptonase™ (94.7 ± 4.6 units) had the highest activity, while Unitinase™ (31.0 ± 2.4 units) and Strek™ (32.9 ± 3.3 units) had the weakest activity. Solustrep™ (53.3 ± 2.7 units) presented intermediate activity. The variations among the different formulations for both euglobulin lysis test and chromogenic substrate hydrolysis correlated with the SDS-PAGE densitometric results for the amount of 47-kDa protein. These data show that the commercially available clinical streptokinase formulations vary significantly in their in vitro activity. Whether these differences have clinical implications needs to be investigated.

L.T., Couto; J.L., Donato; G. de, Nucci.

99

Analysis of five streptokinase formulations using the euglobulin lysis test and the plasminogen activation assay  

Directory of Open Access Journals (Sweden)

Full Text Available Streptokinase, a 47-kDa protein isolated and secreted by most group A, C and G ß-hemolytic streptococci, interacts with and activates human protein plasminogen to form an active complex capable of converting other plasminogen molecules to plasmin. Our objective was to compare five streptokinase formulations commercially available in Brazil in terms of their activity in the in vitro tests of euglobulin clot formation and of the hydrolysis of the plasmin-specific substrate S-2251™. Euglobulin lysis time was determined using a 96-well microtiter plate. Initially, human thrombin (10 IU/ml and streptokinase were placed in individual wells, clot formation was initiated by the addition of plasma euglobulin, and turbidity was measured at 340 nm every 30 s. In the second assay, plasminogen activation was measured using the plasmin-specific substrate S-2251™. Streptase™ was used as the reference formulation because it presented the strongest fibrinolytic activity in the euglobulin lysis test. The Unitinase™ and Solustrep™ formulations were the weakest, showing about 50% activity compared to the reference formulation. All streptokinases tested activated plasminogen but significant differences were observed. In terms of total S-2251™ activity per vial, Streptase™ (75.7 ± 5.0 units and Streptonase™ (94.7 ± 4.6 units had the highest activity, while Unitinase™ (31.0 ± 2.4 units and Strek™ (32.9 ± 3.3 units had the weakest activity. Solustrep™ (53.3 ± 2.7 units presented intermediate activity. The variations among the different formulations for both euglobulin lysis test and chromogenic substrate hydrolysis correlated with the SDS-PAGE densitometric results for the amount of 47-kDa protein. These data show that the commercially available clinical streptokinase formulations vary significantly in their in vitro activity. Whether these differences have clinical implications needs to be investigated.

L.T. Couto

2004-12-01

100

Immunoradiometric quantification of tissue plasminogen activator secreted by fetal organs. Comparison with urokinase  

International Nuclear Information System (INIS)

Explants of fetal tissues were maintained in organ cultures for 2-3 weeks and the conditioned culture media analysed for the two main plasminogen activators, tissue activator and urokinase. A new immunoradiometric method was used for determining the tissue activator. The method is based on "1"2"5I-labelled antibodies to a tissue plasminogen activator which was purified from the culture medium of an established melanoma cell line. It detected tissue activator in a concentration of 1 ?g/l or even less. Urokinase was measured with a RIA. Explants of kidney as well as thyroid gland and thymus released very substantial amounts of urokinase and smaller amounts of tissue activator. Urokinase was also detected in media conditioned by skin, spleen and pancreas. Aorta explants released only the tissue activator. The capacity of many tissues to release urokinase indicates that this is a more significant plasminogen activator in extrarenal organs than hitherto realized. The tissue activator is probably confined to vascular structures. (author)

1982-01-01

 
 
 
 
101

Immunoradiometric quantification of tissue plasminogen activator secreted by fetal organs. Comparison with urokinase  

Energy Technology Data Exchange (ETDEWEB)

Explants of fetal tissues were maintained in organ cultures for 2-3 weeks and the conditioned culture media analysed for the two main plasminogen activators, tissue activator and urokinase. A new immunoradiometric method was used for determining the tissue activator. The method is based on /sup 125/I-labelled antibodies to a tissue plasminogen activator which was purified from the culture medium of an established melanoma cell line. It detected tissue activator in a concentration of 1 ..mu..g/l or even less. Urokinase was measured with a RIA. Explants of kidney as well as thyroid gland and thymus released very substantial amounts of urokinase and smaller amounts of tissue activator. Urokinase was also detected in media conditioned by skin, spleen and pancreas. Aorta explants released only the tissue activator. The capacity of many tissues to release urokinase indicates that this is a more significant plasminogen activator in extrarenal organs than hitherto realized. The tissue activator is probably confined to vascular structures.

Holmberg, L.; Kristoffersson, A.C.; Lecander, I.; Wallen, P.; Aestedt, B. (Malmoe Allmaenna Sjukhus (Sweden). Dept. of Paediatrics; Lund Univ. (Sweden). Dept. of Obstetrics and Gynaecology; Umeaa Univ. (Sweden))

1982-06-01

102

An enzyme-immunobinding assay for fast screening of expression of tissue plasminogen activator cDNA in E. coli  

International Nuclear Information System (INIS)

Tissue plasminogen activator (TPA) has been isolated from normal human tissues and certain human cell lines in culture. The enzyme is a serine protease which converts an inactive zymogen, plasminogen to plasmin, and causes lysis of fibrin clots. The high affinity of TPA for fibrin indicates that it is a potential thrombolytic agent and is superior to urokinase-like plasminogen activators. Recently, TPA has been cloned and expressed in E. coli. Using TPA as a model protein, the authors report here the development of a direct, sensitive enzyme-immunoassay for the screening of a cDNA expression library using specific antibodies and peroxidase-labeled second antibody

1984-01-01

103

Plasminogen activator inhibitor-1, free fatty acids, and insulin resistance in patients with myocardial infarction  

Directory of Open Access Journals (Sweden)

Full Text Available Olga Gruzdeva, Evgenya Uchasova, Yulia Dyleva, Ekaterina Belik, Ekaterina Shurygina, Olga Barbarash Research Institute for Complex Issues of Cardiovascular Diseases under the Siberian Branch of the Russian Academy of Medical Sciences, Kemerovo, Russian Federation Background: Insulin resistance is known to be a common feature of type 2 diabetes mellitus and is regarded as an important mechanism in the pathogenesis of this disease. The key pathogenetic mechanisms of insulin resistance progression are free fatty acids metabolism impairment and enhanced activity of plasminogen activator inhibitor 1. Both free fatty acids and plasminogen activator inhibitor 1 are recognized as risk factors for coronary heart disease. Methods: The patients were divided into two groups: group 1 included 65 non-diabetic myocardial infarction patients and group 2 enrolled 60 diabetic myocardial infarction patients. The control group consisted of 30 sex- and age-matched volunteers. The concentration of serum free fatty acids, glucose, C-peptide, and insulin were measured on the 1st and 12th days of the study. All the patients had their postprandial glycemia, insulin, and C-peptide concentrations measured 2 hours after a standard carbohydrate breakfast containing 360 kcal (protein 20 g, carbohydrate 57 g, and fat 9 g. Results: Free fatty acids levels in group 1 and in group 2 exceeded the control group values by 7-fold and 11-fold, respectively. Plasminogen activator inhibitor 1 concentration was 2.5-fold higher in group 1 and 4.6-fold higher in group 2 compared to the control group on the 1st day from the myocardial infarction onset. In addition, plasminogen activator inhibitor 1 concentration was significantly reduced in both groups on the 12th day from the myocardial infarction onset; however, it did not achieve the control group values. Conclusion: Increased postprandial glucose level, insulinemia, and elevated levels of free fatty acids and plasminogen activator inhibitor are associated with myocardial infarction-associated progression of insulin resistance. However, insulin resistance metabolic markers are of great predictive capacity in the assessment of risk of acute coronary events. Keywords: free fatty acids, type 2 diabetes mellitus, myocardial infarction, insulin resistance, plasminogen activator inhibitor 1

Gruzdeva O

2013-08-01

104

Induction of macrophage plasminogen activator by asbestos is independent of PKC activation  

Energy Technology Data Exchange (ETDEWEB)

This study was undertaken to assess whether plasminogen activator (PA) induction in macrophages exposed to chrysotile fibers is mediated by protein kinase C (PKC) activation. In PKC depleted J774 cells, PA induction could be elicited by chrysotile whereas, as expected, the response to phorbol myristate acetate (PMA) was abolished. The effect of PMA and chrysotile on the distribution of PKC activity in the J774 cell line was also compared by measuring the enzyme catalytic activity and phorbol dibutyrate (PDBu) binding sites. No redistribution of PKC was observed after simulation with PA inducing doses of chrysotile, whereas a clear translocation was observed with PMA. It is concluded that the mechanism of PA induction by chrysotile in this macrophage-like cell line is independent of PKC activation. (orig.).

Lison, D.; Raguzzi, F.; Lauwerys, R. (Louvain Univ., Brussels (Belgium). Industrial Toxicology and Occupational Medicine Unit)

1991-07-01

105

A nonantigenic covalent streptokinase-polyethylene glycol complex with plasminogen activator function.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A series of new, covalent polyethylene glycol (PEG)-streptokinase adducts were synthesized and characterized. PEGs with average molecular weights of 2,000, 4,000, and 5,000 were activated with carbonyldiimidazole and coupled to the protein under standardized reaction conditions. Steady-state kinetic analysis demonstrated comparable Km values for the activation of plasminogen by streptokinase, PEG-2-streptokinase, and PEG-4-streptokinase. The kcat values were somewhat decreased when PEG-2 or P...

Rajagopalan, S.; Gonias, S. L.; Pizzo, S. V.

1985-01-01

106

The Plasminogen Activation System Modulates Differently Adipogenesis and Myogenesis of Embryonic Stem Cells  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Regulation of the extracellular matrix (ECM) plays an important functional role either in physiological or pathological conditions. The plasminogen activation (PA) system, comprising the uPA and tPA proteases and their inhibitor PAI-1, is one of the main suppliers of extracellular proteolytic activity contributing to tissue remodeling. Although its function in development is well documented, its precise role in mouse embryonic stem cell (ESC) differentiation in vitro is unknown. We found that...

Hadadeh, Ola; Barruet, Emilie; Peiretti, Franck; Verdier, Monique; Bernot, Denis; Hadjal, Yasmine; Yazidi, Claire El; Robaglia-schlupp, Andre?e; Paula, Andre Maues; Ne?gre, Didier; Iacovino, Michelina; Kyba, Michael; Alessi, Marie-christine; Bine?truy, Bernard

2012-01-01

107

Tissue-type plasminogen activator is a target of the tumor suppressor?gene?maspin  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The maspin protein has tumor suppressor activity in breast and prostate cancers. It inhibits cell motility and invasion in vitro and tumor growth and metastasis in nude mice. Maspin is structurally a member of the serpin (serine protease inhibitors) superfamily but deviates somewhat from classical serpins. We find that single-chain tissue plasminogen activator (sctPA) specifically interacts with the maspin reactive site loop peptide and forms a stable complex with recombinant maspin [rMaspin(...

1998-01-01

108

2-Amidino Analogs of Glycine-Amiloride Conjugates: Inhibitors of Urokinase-type Plasminogen Activator  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The relative non-toxicity of the diuretic amiloride, coupled with its selective inhibition of the protease urokinase plasminogen activator (uPA), makes this compound class attractive for structure-activity studies. Herein we substituted the C(2)-acylguanidine of C(5)-glycyl-amiloride with amidine and amidoxime groups. The data show the importance of maintaining C(5)-hydrophobicity. The C(5)-benzylglycine analogs containing either C(2)-acylguanidine or amidine inhibited uPA with an IC50 rangin...

Massey, Archna P.; Harley, William R.; Pasupuleti, Nagarekha; Gorin, Fredric A.; Nantz, Michael H.

2012-01-01

109

Plasminogen activator inhibitor type I stabilizes vitronectin-dependent adhesions in HT-1080 cells  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Polyclonal antibodies against plasminogen activator inhibitor type-I (PAI-1) caused rapid retraction and rounding of substrate-attached HT- 1080 cells. The kinetics and extent of antibody-mediated cell rounding were not dependent on either urokinase or plasmin activity. Cells adherent to vitronectin-coated substrates detached within 2 h of antibody addition. Cells adherent to fibronectin were unaffected by the antibodies. Immunoblotting of substrate-attached material indicated that HT-1080 ce...

1990-01-01

110

Urokinase plasminogen activator system in humans with stable coronary artery disease.  

Science.gov (United States)

1. The present study compares plasma urokinase plasminogen activator (uPA) peptide levels, plasma plasminogen inhibitor (PAI-1) activity and urokinase receptors (uPAR) on peripheral blood monocytes of patients with stable coronary artery disease (SCAD) and healthy volunteers. 2. Urokinase plasminogen activator levels were analysed by ELISA and PAI-1 activity was determined by a plasmin generation method using the chromogenic substrate S2390. Relative uPAR numbers and the adhesion molecules CD11b/CD18 on peripheral blood monocytes were estimated using specific antibodies and flow cytometry. 3. Patients with SCAD were found to have higher plasma uPA peptide levels than age-matched healthy subjects (10.40 +/- 0.99 vs 8.25 +/- 0.53 pmol/L, respectively; P < 0.05). 4. Plasma PAI-1 activity was also higher in patients with SCAD than in healthy subjects (13.6 +/- 2.5 vs 5.2 +/- 1.0 IU/mL, respectively; P < 0.05). 5. Relative uPAR and CD11b/CD18 adhesion molecules were similar on peripheral blood monocytes of patients with SCAD and in healthy subjects. 6. The data indicate a pattern of expression/activity of uPA and PAI-1 in patients with SCAD suggestive of an impaired fibrinolytic ability. PMID:10225148

Krasnikova, T L; Parfyonova, Y; Alekseeva, I A; Arefieva, T I; Mukhina, S A; Dobrovolsky, A B; Titaeva, Y; Lyakishev, A A; Resink, T J; Erne, P; Tkachuk, V A

1999-04-01

111

Expression of Active Recombinant Human Tissue-Type Plasminogen Activator by Using In Vivo Polyhydroxybutyrate Granule Display?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Recombinant human tissue plasminogen activator (rPA) is a truncated version of tissue plasminogen activator (tPA), which contains nine disulfide bonds and is prone to forming inactive inclusion bodies when expressed in bacteria. To obtain functional rPA expression, we displayed the rPA on the surface of polyhydroxybutyrate (PHB) granules using phasin as the affinity tag. rPA was fused to the N terminus of the phasin protein with a thrombin cleavage site as the linker. Sodium dodecyl sulfate-p...

Geng, Yanping; Wang, Shengjun; Qi, Qingsheng

2010-01-01

112

Increased Plasminogen Activator Inhibitor-1 in Keloid Fibroblasts May Account for their Elevated Collagen Accumulation in Fibrin Gel Cultures  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Proteolytic degradation of the provisional fibrin matrix and subsequent substitution by fibroblast-produced collagen are essential features of injury repair. Immunohistochemical studies revealed that although dermal fibroblasts of normal scars and keloids expressed both urokinase type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1), keloid fibroblasts had a much higher PAI-1 expression. In long-term three-dimensional fibrin gel cultures (the in vitro fibroplasia mode...

Tuan, Tai-lan; Wu, Huayang; Huang, Eunice Y.; Chong, Sheree S. N.; Laug, Walter; Messadi, Diana; Kelly, Paul; Le, Anh

2003-01-01

113

Purification and activation of caprine and canine plasminogens: Comparison with human plasminogen / Purificación y activación de los plasminógenos caprino y canino: comparación con el plasminógeno humano  

Scientific Electronic Library Online (English)

Full Text Available SciELO Colombia | Language: English Abstract in spanish Objetivo: unificar la purificación y activación de los plasminógenos de tres especies diferentes, a saber: humana, caprina y canina. Materiales y métodos: se usaron Lysina-Sefarosa 4B y Sefacel DEAE para las cromatografías de afinidad y de intercambio iónico, respectivamente. Se determinó la secuenc [...] ia terminal-N tanto de los plasminógenos intactos como de los degradados. Resultados: en las tres especies se identificaron bandas de 92 kDa correspondientes a los plasminógenos nativos. Se halló que sus secuencias terminales-N eran EPLDDY, DPLDDY y XXLDDY para los plasminógenos humano, caprino y canino, respectivamente. Además, se purificaron los plasminógenos degradados circulantes, cuyas secuencias terminales-N fueron, en el mismo orden, KVYLSE, RITLL Y RIYSL. Conclusión: la activación de los tres plasminógenos confirmó la formación de las bandas electroforéticas típicas de la plasmina humana correspondientes a las cadenas pesada A y liviana B, que también se identificaron en las plasminas caprina y canina. Este nuevo método de purificación facilita la comparación y el esclarecimiento de los sistemas fibrinolíticos de los mamíferos. Abstract in english Objective: To unify the purification and activation of plasminogens from three different species, namely: human, caprine and canine. Materials and methods: Lysine-Sepharose 4B and sephacel DEAE were used, for affinity and ion-exchange chromatography, respectively. The N-terminal sequence was determi [...] ned for both the intact and degraded plasminogens. Results: Bands of 92 kDa corresponding to native plasminogens were identified in the three species. Their N-terminal sequences were found to be EPLDDY, DPLDDY and XXLDDY for human, caprine and canine plasminogen, respectively. Furthermore, the degraded in vivo circulating plasminogens from the three species were purified and their N-terminal sequences were KVYLSE, RITLL and RIYLS for the human, caprine and canine, in that order. Conclusion: Activation of the three plasminogens confirmed the formation of the typical electrophoretic bands for human plasmin corresponding to the heavy A and the light B chains which were also identified in the caprine and canine plasmins. This new purification methodology facilitates the comparison and further elucidation of the fibrinolytic systems in mammals.

Omaira, Cañas Bermúdez; Alfonso, Quijano Parra; Luis Fernando, Arbeláez Ramírez.

114

Longistatin, a plasminogen activator, is key to the availability of blood-meals for ixodid ticks.  

Science.gov (United States)

Ixodid ticks are notorious blood-sucking ectoparasites and are completely dependent on blood-meals from hosts. In addition to the direct severe effects on health and productivity, ixodid ticks transmit various deadly diseases to humans and animals. Unlike rapidly feeding vessel-feeder hematophagous insects, the hard ticks feed on hosts for a long time (5-10 days or more), making a large blood pool beneath the skin. Tick's salivary glands produce a vast array of bio-molecules that modulate their complex and persistent feeding processes. However, the specific molecule that functions in the development and maintenance of a blood pool is yet to be identified. Recently, we have reported on longistatin, a 17.8-kDa protein with two functional EF-hand Ca(++)-binding domains, from the salivary glands of the disease vector, Haemaphysalis longicornis, that has been shown to be linked to blood-feeding processes. Here, we show that longistatin plays vital roles in the formation of a blood pool and in the acquisition of blood-meals. Data clearly revealed that post-transcriptional silencing of the longistatin-specific gene disrupted ticks' unique ability to create a blood pool, and they consequently failed to feed and replete on blood-meals from hosts. Longistatin completely hydrolyzed ?, ? and ? chains of fibrinogen and delayed fibrin clot formation. Longistatin was able to bind with fibrin meshwork, and activated fibrin clot-bound plasminogen into its active form plasmin, as comparable to that of tissue-type plasminogen activator (t-PA), and induced lysis of fibrin clot and platelet-rich thrombi. Plasminogen activation potentiality of longistatin was increased up to 4 times by soluble fibrin. Taken together, our results suggest that longistatin may exert potent functions both as a plasminogen activator and as an anticoagulant in the complex scenario of blood pool formation; the latter is critical to the feeding success and survival of ixodid ticks. PMID:21423674

Anisuzzaman; Islam, M Khyrul; Alim, M Abdul; Miyoshi, Takeharu; Hatta, Takeshi; Yamaji, Kayoko; Matsumoto, Yasunobu; Fujisaki, Kozo; Tsuji, Naotoshi

2011-03-01

115

Expression, purification and characterization of recombinant plasminogen activator from Gloydius brevicaudus venom in Escherichia coli.  

Science.gov (United States)

The plasminogen activator (PA) in snake venom, a serine protease, can convert plasminogen to active plasmin, indirectly causing the degradation of fibrin. It is difficult to purify sufficient snake venom PA (SV-PA) for clinical applications due to the low SV-PA content in venom. The gene encoding PA was obtained from the venom gland of Gloydius brevicaudus using RT-PCR with primers designed according to the N-terminal amino acids of G. brevicaudus venom PA (GBV-PA), was cloned into the prokaryotic expression vector pET-42a, and recombinant GBV-PA (rGBV-PA) was expressed via Isopropyl-?-d-1-Thiogalactopyranoside (IPTG) induction. Like human tissue PA, the purified renatured rGBV-PA could significantly reduce the rabbit plasma euglobulin lysis time (ELT) and prevent thrombus formation in the inferior vena cava of rats. Within the dosage range, the dosage and effects were positively correlated. PMID:23891573

Zhang, Jinhua; Meng, Wenli; Wang, Chunhua; Wu, Zhiqiang; Wu, Guotu; Xu, Yunlu

2013-09-01

116

Structural insight into inactivation of plasminogen activator inhibitor-1 by a small-molecule antagonist.  

DEFF Research Database (Denmark)

Plasminogen activator inhibitor-1 (PAI-1), a serpin, is the physiological inhibitor of tissue-type and urokinase-type plasminogen activators and thus also an inhibitor of fibrinolysis and tissue remodeling. It is a potential therapeutic target in many pathological conditions, including thrombosis and cancer. Several types of PAI-1 antagonist have been developed, but the structural basis for their action has remained largely unknown. Here we report X-ray crystal structure analysis of PAI-1 in complex with a small-molecule antagonist, embelin. We propose a mechanism for embelin-induced rapid conversion of PAI-1 into a substrate for its target proteases and the subsequent slow conversion of PAI-1 into an irreversibly inactivated form. Our work provides structural clues to an understanding of PAI-1 inactivation by small-molecule antagonists and an important step toward the design of drugs targeting PAI-1.

Lin, Zhonghui; Jensen, Jan Kristian

2013-01-01

117

Structural insight into inactivation of plasminogen activator inhibitor-1 by a small-molecule antagonist.  

Science.gov (United States)

Plasminogen activator inhibitor-1 (PAI-1), a serpin, is the physiological inhibitor of tissue-type and urokinase-type plasminogen activators and thus also an inhibitor of fibrinolysis and tissue remodeling. It is a potential therapeutic target in many pathological conditions, including thrombosis and cancer. Several types of PAI-1 antagonist have been developed, but the structural basis for their action has remained largely unknown. Here we report X-ray crystal structure analysis of PAI-1 in complex with a small-molecule antagonist, embelin. We propose a mechanism for embelin-induced rapid conversion of PAI-1 into a substrate for its target proteases and the subsequent slow conversion of PAI-1 into an irreversibly inactivated form. Our work provides structural clues to an understanding of PAI-1 inactivation by small-molecule antagonists and an important step toward the design of drugs targeting PAI-1. PMID:23438754

Lin, Zhonghui; Jensen, Jan K; Hong, Zebin; Shi, Xiaoli; Hu, Lihong; Andreasen, Peter A; Huang, Mingdong

2013-02-21

118

Acute myocardial infarction following intravenous tissue plasminogen activator for acute ischemic stroke: An unknown danger  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Thrombolysis with intravenous tissue (IV) plasminogen activator (tPA) is considered for patients with acute ischemic stroke falling within the described inclusion criteria defined by The National Institute of Neurological Disorders and Stroke (NINDS) rtPA trial. Complications of IV thrombolysis with tPA are commonly related to hemorrhage, anaphylaxis, or arterial occlusion. We describe two cases of acute myocardial infarction (MI) following IV tPA infusion for acute stroke. One of the patient...

Adatia Sweta; Sanghani Sejal; Sanzgiri Prakash; Chauhan Vinay; Hastak Shirish

2010-01-01

119

Reduction of brain metastases in plasminogen activator inhibitor-1-deficient mice with transgenic ocular tumors  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator inhibitor-1 is known to play a paradoxical positive role in tumor angiogenesis, but its contribution to metastatic spread remains unclear. We studied the impact of PAI-1 deficiency in a transgenic mouse model of ocular tumors originating from retinal epithelial cells and leading to brain metastasis (TRP-1/SV40 Tag mice). PAI-1 deficiency did not affect primary tumor growth or vascularization, but was associated with a smaller number of brain metastases. Brain metastases ...

Maillard, Catherine; Bouquet, C.; Petitjean, Marie; Mestdagt, Me?lanie; Frau, E.; Jost, Maud; Masset, Anne; Opolon, P. H.; Beerman, F.; Abitbol, M.; Foidart, Jean-michel; Perricaudet, M.; Noe?l, Agne?s

2008-01-01

120

Aggregation and retention of human urokinase type plasminogen activator in the yeast endoplasmic reticulum  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Background Secretion of recombinant proteins in yeast can be affected by their improper folding in the endoplasmic reticulum and subsequent elimination of the misfolded molecules via the endoplasmic reticulum associated protein degradation pathway. Recombinant proteins can also be degraded by the vacuolar protease complex. Human urokinase type plasminogen activator (uPA) is poorly secreted by yeast but the mechanisms interfering with its secretion are largel...

2002-01-01

 
 
 
 
121

Polymorphism in methylenetetrahydrofolate reductase, plasminogen activator inhibitor-1, and apolipoprotein E in hemodialysis patients  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The methylenetetrahydrofolate reductase (MTHFR) gene polymorphism, apolipoprotein E (apo s4) gene polymorphism and polymorphism of plasminogen activator inhibitor-1 (PAI-1) have been shown to be associated with end-stage renal disease (ESRD). To determine the prevalence of these mutations in Saudi patients with ESRD on hemodialysis, we studied the allelic frequency and genotype distribution in patients receiving hemodialysis and in a control group, all residing in the Eastern Province of Saud...

Al-Muhanna Fahad; Al-Mueilo Samir; Al-Ali Amein; Larbi Emmanuel; Rubaish Abdullah; Abdulmohsen Mohammed; Al-Zahrani Alhussain; Al-Ateeq Suad

2008-01-01

122

t-plasminogen activator inhibitor-1 polymorphism in idiopathic pulmonary arterial hypertension  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Aim: The aim of the present study was to identify the possible genotypic association of 3?UTR Hind III polymorphism of Plasminogen activator Inhibitor-1 (PAI-1) gene with idiopathic pulmonary arterial hypertension (IPAH). Background: IPAH is a disorder with abnormally raised mean pulmonary arterial pressure and increase in the resistance to blood flow in pulmonary artery. One of the pathological features seen is development of intraluminal thrombin deposition le...

Katta Sujana; Vadapalli Shivani; Bks, Sastry; Nallari Pratibha

2008-01-01

123

Increased expression of urokinase plasminogen activator in Quebec platelet disorder is linked to megakaryocyte differentiation  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Quebec platelet disorder (QPD) is an inherited bleeding disorder associated with increased urokinase plasminogen activator (uPA) in platelets but not in plasma, intraplatelet plasmin generation, and ?-granule protein degradation. These abnormalities led us to investigate uPA expression by QPD CD34+ progenitors, cultured megakaryocytes, and platelets, and whether uPA was stored in QPD ?-granules. Although QPD CD34+ progenitors expressed normal amounts of uPA, their differentiation into megak...

Veljkovic, D. Kika; Rivard, Georges E.; Diamandis, Maria; Blavignac, Jessica; Cramer-borde?, Elisabeth M.; Hayward, Catherine P. M.

2009-01-01

124

Decreased serotonin levels associated with behavioral disinhibition in tissue plasminogen activator deficient (tPA?/?) mice  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Tissue Plasminogen Activator (tPA) is a serine protease expressed in different areas of the mammalian brain. It has been used clinically to dissolve clots and shown to have a role in neurodegeneration. Early studies suggested that tPA plays an important role in the processes of learning and memory, demonstrated at the level of behavior and synaptic plasticity. Herein, we extend the behavioral characterization of these mice to the related dimension of exploratory-related behavior using an exte...

2010-01-01

125

Tryptophan Properties in Fluorescence and Functional Stability of Plasminogen Activator Inhibitor 1  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator inhibitor 1 harbors four tryptophan residues at positions 86, 139, 175, and 262. To investigate the contribution of each tryptophan residue to the total fluorescence and to reveal the mutual interactions of the tryptophan residues and interactions with the other amino acids, 15 mutants in which tryptophan residues have been replaced by phenylalanines were constructed, purified, and characterized. Conformational distribution analysis revealed that the tryptophan mutants h...

2003-01-01

126

Recombinant tissue-type plasminogen activator and immediate angioplasty in acute myocardial infarction.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

BACKGROUND. The European Cooperative Study Group conducted two randomized trials in patients with suspected myocardial infarction to assess the effect of 100 mg single-chain recombinant tissue-type plasminogen activator (rt-PA, alteplase) on enzymatic infarct size, left ventricular function, morbidity and mortality relative to placebo (alteplase/placebo trial) and to assess the effect of immediate percutaneous transluminal coronary angioplasty (PTCA) in addition to alteplase (alteplase/PTCA t...

Arnold, A. E. R.; Simoons, M. L.; Bono, D. P.; Tijssen, J. G. P.; Serruys, P. W. J. C.; Verstraete, M.; Lubsen, J.; Werf, F. J. J.

1992-01-01

127

The roles of the plasminogen activator and matrix metalloproteinase systems in ovulation and corpus luteum formation  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Proteases of the plasminogen activator (PA) and the matrix metalloproteinase (MMP) enzyme systems are expressed in the ovulatory follicle and in the developing corpus luteum (CL). However, the functional role of these extracellular degrading protease systems in the ovulatory and CL development processes remains elusive. The first aim of this thesis was to develop a mouse model to study gonadotropin-induced CL formation. The second aim was to study the involvement of the PA and the MMP systems...

Bode?n, Ida

2004-01-01

128

Modulation of the plasminogen activation system by inflammatory cytokines in human colon carcinoma cells.  

Science.gov (United States)

Inflammation may promote malignant invasion by enhancing cancer cell-associated proteolysis. Here we present the effect of inflammatory cytokines on the plasminogen activation system of eight human colon carcinoma cell lines. Tumour necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) increased in several, but not all, cell lines the production of urokinase-type plasminogen activator (uPA), tissue-type PA (tPA) and plasminogen activator inhibitor type 1 (PAI-1) as analysed by zymography, enzyme immunoassays and Northern analysis. Interleukin 6 (IL-6) had no effect. uPA receptor (uPAR) mRNA levels were also upregulated. However, each individual cell line responded differently following exposure to TNF-alpha or IL-1 beta. For example, there was a dose-dependent up-regulation of uPA and PAI-1 in SW 620 cells, whereas increased uPA production in SW 1116 cells was not accompanied by an increase in PAI-1. The TNF-alpha stimulatory effect was blocked by anti-TNF-alpha Fab fragments. All cell lines expressed both types of TNF receptor mRNAs, whereas no transcript for TNF-alpha, IL-1 beta, IL-6, IL-6 receptor or the IL-1 receptors was found. Our results demonstrate that TNF-alpha and IL-1 beta stimulate the plasminogen activation system in tumour cell but the responses differed even in cells derived from the same tissue origin. Images Figure 1 Figure 2 Figure 3 Figure 5

Tran-Thang, C.; Kruithof, E.; Lahm, H.; Schuster, W. A.; Tada, M.; Sordat, B.

1996-01-01

129

Tissue Plasminogen Activator Concentration Dependence of 120 kHz Ultrasound Enhanced Thrombolysis-Revised #1  

Digital Repository Infrastructure Vision for European Research (DRIVER)

It has been known for some time that the application of ultrasound can enhance the efficacy of thrombolytic medications such as recombinant tissue plasminogen activator (rt-PA). Potential clinical applications of this ultrasound enhanced thrombolysis (UET) include the treatment of myocardial infarction, acute ischemic stroke, deep venous thrombosis, and other thrombotic disorders. It may be possible to reduce the dose of rt-PA while maintaining lytic efficacy, however there is little data on ...

Shaw, George J.; Meunier, Jason M.; Lindsell, Christopher J.; Holland, Christy K.

2008-01-01

130

Tissue plasminogen activator (alteplase) treatment for femoral artery thrombosis after cardiac catheterisation in infants and children.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

OBJECTIVE--To determine the efficacy of fibrinolytic therapy with tissue plasminogen activator (alteplase) in infants and children with arterial thrombosis after cardiac catheterisation. DESIGN--Use of alteplase (Actilyse) in a protocol with prospective data collection. Alteplase was administered to infants and children with arterial thrombosis after cardiac catheterisation. A dose of 0.5 mg/kg/h was given continuously via a peripheral vein for the first hour followed by 0.25 mg/kg/h till clo...

Zenz, W.; Muntean, W.; Beitzke, A.; Zobel, G.; Riccabona, M.; Gamillscheg, A.

1993-01-01

131

Inhibitors of urokinase type plasminogen activator and cytostatic activity from crude plants extracts.  

Science.gov (United States)

In view of the clear evidence that urokinase type plasminogen activator (uPA) plays an important role in the processes of tumor cell metastasis, aortic aneurysm, and multiple sclerosis, it has become a target of choice for pharmacological intervention. The goal of this study was thus to determine the presence of inhibitors of uPA in plants known traditionally for their anti-tumor properties. Crude methanol extracts were prepared from the leaves of plants (14) collected from the subtropical dry forest (Guanica, Puerto Rico), and tested for the presence of inhibitors of uPA using the fibrin plate assay. The extracts that tested positive (6) were then partitioned with petroleum ether, chloroform, ethyl acetate and n-butanol, in a sequential manner. The resulting fractions were then tested again using the fibrin plate assay. Extracts from leaves of Croton lucidus (C. lucidus) showed the presence of a strong uPA inhibitory activity. Serial dilutions of these C. lucidus partitions were performed to determine the uPA inhibition IC?? values. The chloroform extract showed the lowest IC?? value (3.52 µg/mL) and hence contained the most potent uPA inhibitor. Further investigations revealed that the crude methanol extract and its chloroform and n-butanol partitions did not significantly inhibit closely related proteases such as the tissue type plasminogen activator (tPA) and plasmin, indicating their selectivity for uPA, and hence superior potential for medicinal use with fewer side effects. In a further evaluation of their therapeutic potential for prevention of cancer metastasis, the C. lucidus extracts displayed cytostatic activity against human pancreatic carcinoma (PaCa-2) cells, as determined through an MTS assay. The cytostatic activities recorded for each of the partitions correlated with their relative uPA inhibitory activities. There are no existing reports of uPA inhibitors being present in any of the plants reported in this study. PMID:23896619

Zha, Xueqiang; Diaz, Ricardo; Franco, Jose Javier Rosado; Sanchez, Veronica Forbes; Fasoli, Ezio; Barletta, Gabriel; Carvajal, Augusto; Bansal, Vibha

2013-01-01

132

Inhibitors of Urokinase Type Plasminogen Activator and Cytostatic Activity from Crude Plants Extracts  

Science.gov (United States)

In view of the clear evidence that urokinase type plasminogen activator (uPA) plays an important role in the processes of tumor cell metastasis, aortic aneurysm, and multiple sclerosis, it has become a target of choice for pharmacological intervention. The goal of this study was thus to determine the presence of inhibitors of uPA in plants known traditionally for their anti-tumor properties. Crude methanol extracts were prepared from the leaves of plants (14) collected from the subtropical dry forest (Guanica, Puerto Rico), and tested for the presence of inhibitors of uPA using the fibrin plate assay. The extracts that tested positive (6) were then partitioned with petroleum ether, chloroform, ethyl acetate and n-butanol, in a sequential manner. The resulting fractions were then tested again using the fibrin plate assay. Extracts from leaves of Croton lucidus (C. lucidus) showed the presence of a strong uPA inhibitory activity. Serial dilutions of these C. lucidus partitions were performed to determine the uPA inhibition IC50 values. The chloroform extract showed the lowest IC50 value (3.52 ?g/mL) and hence contained the most potent uPA inhibitor. Further investigations revealed that the crude methanol extract and its chloroform and n-butanol partitions did not significantly inhibit closely related proteases such as the tissue type plasminogen activator (tPA) and plasmin, indicating their selectivity for uPA, and hence superior potential for medicinal use with fewer side effects. In a further evaluation of their therapeutic potential for prevention of cancer metastasis, the C. lucidus extracts displayed cytostatic activity against human pancreatic carcinoma (PaCa-2) cells, as determined through an MTS assay. The cytostatic activities recorded for each of the partitions correlated with their relative uPA inhibitory activities. There are no existing reports of uPA inhibitors being present in any of the plants reported in this study.

Zha, Xueqiang; Diaz, Ricardo; Franco, Jose Javier Rosado; Sanchez, Veronica Forbes; Fasoli, Ezio; Barletta, Gabriel; Carvajal, Augusto; Bansal, Vibha

2014-01-01

133

Inhibitors of Urokinase Type Plasminogen Activator and Cytostatic Activity from Crude Plants Extracts  

Directory of Open Access Journals (Sweden)

Full Text Available In view of the clear evidence that urokinase type plasminogen activator (uPA plays an important role in the processes of tumor cell metastasis, aortic aneurysm, and multiple sclerosis, it has become a target of choice for pharmacological intervention. The goal of this study was thus to determine the presence of inhibitors of uPA in plants known traditionally for their anti-tumor properties. Crude methanol extracts were prepared from the leaves of plants (14 collected from the subtropical dry forest (Guanica, Puerto Rico, and tested for the presence of inhibitors of uPA using the fibrin plate assay. The extracts that tested positive (6 were then partitioned with petroleum ether, chloroform, ethyl acetate and n-butanol, in a sequential manner. The resulting fractions were then tested again using the fibrin plate assay. Extracts from leaves of Croton lucidus (C. lucidus showed the presence of a strong uPA inhibitory activity. Serial dilutions of these C. lucidus partitions were performed to determine the uPA inhibition IC50 values. The chloroform extract showed the lowest IC50 value (3.52 µg/mL and hence contained the most potent uPA inhibitor. Further investigations revealed that the crude methanol extract and its chloroform and n-butanol partitions did not significantly inhibit closely related proteases such as the tissue type plasminogen activator (tPA and plasmin, indicating their selectivity for uPA, and hence superior potential for medicinal use with fewer side effects. In a further evaluation of their therapeutic potential for prevention of cancer metastasis, the C. lucidus extracts displayed cytostatic activity against human pancreatic carcinoma (PaCa-2 cells, as determined through an MTS assay. The cytostatic activities recorded for each of the partitions correlated with their relative uPA inhibitory activities. There are no existing reports of uPA inhibitors being present in any of the plants reported in this study.

Augusto Carvajal

2013-07-01

134

Hormonal regulation of plasminogen activator production in porcine kidney cells (LLC-PK_1)  

International Nuclear Information System (INIS)

An attempt is made to study the molecular events regulating and accompanying the hormonally modulated plasminogen activator (PA) gene expression. The model system of interest is a porcine kidney cell line, LLC-PK_1, responding to calcitonin in the PA production. The plasminogen activator secreted by calcitonin treated pig kidney cells has been purified, characterized, and compared with human urinary urokinase. The purified enzyme resembles the 53 k MW components of human urokinase. PA induction in LLC-PK_1 cells is sensitive to inhibition by actinomycin D, suggesting the enhanced transcription of PA-mRNA. This hypothesis was tested by measuring PA-mRNA sequences in Xenopus oocyte system which showed a 15-20 fold enhanced PA synthesis when supplied with poly (A)"+ RNA from induced cells, above that obtained from uninduced cell RNA. A pleiotropic response to calcitonin in LLC-PK_1 cells has been examined using two-dimensional gel electrophoresis of "3"5S methionine labeled polypeptides. A set of twelve intracellular polypeptides, ten induced and two repressed, has been identified in calcitonin stimulated cells. One of the induced polypeptides has been identified as plasminogen activator by two dimensional tryptic peptide mapping. Other calcitonin induced polypeptides have been identified as cytokeratins by their solubility properties and cross-reactivity with antiserum against human keratin. The results indicate that the experimental system presented here is a useful and valid one for the study of molecular events regulating and accompanying the hormonally modulated PA gene expression

1983-01-01

135

Coordinate regulation of fibronectin matrix assembly by the plasminogen activator system and vitronectin in human osteosarcoma cells  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Background Plasminogen activators are known to play a key role in the remodeling of bone matrix which occurs during tumor progression, bone metastasis and bone growth. Dysfunctional remodeling of bone matrix gives rise to the osteoblastic and osteolytic lesions seen in association with metastatic cancers. The molecular mechanisms responsible for the development of these lesions are not well understood. Studies were undertaken to address the role of the plasminogen ac...

Vial Daniel; Monaghan-Benson Elizabeth; McKeown-Longo Paula J

2006-01-01

136

[Plasminogen activator and PAI. Detection in aqueous humor of the human eye].  

Science.gov (United States)

Tissue-type plasminogen activator (t-PA), a serine protease that catalyzes the conversion from plasminogen to plasmin, plays an important role in the fibrinolytic system and has also in recent years attracted attention in the field of ophthalmology. In order to examine the role of t-PA in the physiology of the anterior segment, we detected t-PA in aqueous humor by using immunoassays. The samples were taken by keratocentesis prior to cataract or glaucoma surgery. The sample volumes ranged from 50-200 microliters. The quantities of t-PA Ag and plasminogen-activator inhibitor (PAI) Ag were determined by using enzyme-linked immunosorbent assays. We determined t-PA and PAI in the aqueous humor of 54 patients between 32 and 87 years of age. The t-PA Ag levels ranged from 0.2 to 1.9 ng/ml (average 0.8 ng/ml), PAI-Ag from 0.2 to 1.7 ng/ml (average 0.9 ng/ml). The values measured in men were slightly higher than those measured in women. No association between t-PA and PAI levels and accompanying diseases or metabolic disorders was noted. Precise knowledge about the presence of t-PA in aqueous humor is a prerequisite for the recognition of pathological events following intraocular fibrin formation and may be an important basis for the therapeutic use of rt-PA in the intraocular inflammatory process. PMID:8443455

Steinkamp, G W; Hattenbach, L O; Heider, H W; Scharrer, I

1993-02-01

137

Relation of depressive mood to plasminogen activator inhibitor, tissue plasminogen activator, and fibrinogen levels in patients with versus without coronary heart disease.  

Science.gov (United States)

The increased risk of coronary heart disease (CHD) associated with depression is well documented. We hypothesized that impaired fibrinolysis is involved in this link. To explore the association of depressive mood and/or vital exhaustion with various measurements of fibrinolysis activity, 231 men (40 to 65 years old; 123 without CHD and taking no medication and 108 with documented CHD), completed the Center of Epidemiologic Studies Depression Scale and the Maastricht Questionnaire for vital exhaustion. Using classic cut-off points (Center of Epidemiologic Studies Depression Scale score >or=17, Maastricht Questionnaire score >or=8), 6.5% and 9.8% of subjects without CHD and 38% and 48.1% of those with CHD were classified as depressed and exhausted, respectively. Patients with CHD were older, had a higher body mass index, and higher levels of total cholesterol, glucose, plasminogen activator inhibitor 1 (PAI-1), tissue plasminogen activator (t-PA) antigen, and fibrinogen; 47% were treated for hypertension. Depressed subjects had higher levels of PAI-1 activity (p = 0.006) and exhausted patients had higher levels of PAI-1 activity (p = 0.011) and fibrinogen (p = 0.009). After adjusting for clinical condition (with or without CHD), smoking, hypertension, triglyceride concentration, and body mass index, PAI-1 activity remained higher in depressed subjects (p = 0.03). This association persisted after further adjustment for vital exhaustion or for t-PA antigen and fibrinogen levels. t-PA antigen and fibrinogen levels were not associated with depressive mood in multivariate analyses. No fibrinolytic variable was associated with vital exhaustion in multivariate analyses. In conclusion, depressive mood, but not vital exhaustion, is associated with higher levels of PAI-1 activity, suggesting a possible impairment of fibrinolysis and indicating a potential additional mechanism by which depressive mood may act as a cardiovascular risk factor. PMID:16635597

Lahlou-Laforet, Khadija; Alhenc-Gelas, Martine; Pornin, Maurice; Bydlowski, Sarah; Seigneur, Etienne; Benetos, Athanase; Kierzin, Jean-Michel; Scarabin, Pierre-Yves; Ducimetiere, Pierre; Aiach, Martine; Guize, Louis; Consoli, Silla M

2006-05-01

138

Activators of plasminogen and the progression of small abdominal aortic aneurysms  

DEFF Research Database (Denmark)

The aim of this study was to examine the role of activating pathways of plasminogen in the natural history of abdominal aortic aneurysms (AAA). To fulfill this objective 70 male patients with small AAA (> 3 cm) were interviewed and examined. Their blood samples were taken at diagnosis. The patients were scanned annually for a minimum period of 1 year and a maximum of 5 years (mean 2.5 years), and referred for surgery if the AAA exceeded 5 cm in diameter. Plasma levels of urokinase-like plasminogen activator (uPA), tissue-type plasminogen activator (tPA), plasminogen-activator-inhibitor-1 (PAI-1), macrophage-inhibiting factor (MIF), transforming-growth-factor-beta1 (TGF-beta1), homocysteine, and serum levels of IgA-antibodies against Chlamydia pneumoniae (IgA-CP) and cotinine (a nicotine metabolite) were measured. The annual expansion rate correlated positively with tPA, IgA-CP, and S-cotinine; rho = 0.37 (P = 0.004), 0.28 (P = 0.01), and 0.24 (P = 0.04), while PAI-1, uPA, TGF-beta1, homocysteine, and MIF did not. S-cotinine and PAI-1 also correlated positively with tPA, rho = 0.24 (P = 0.04), and 0.33 (P = 0.005). IgA-CP did not correlate with tPA. By receiver operating characteristics (ROC) curve analysis, tPA showed to be predictive of cases expanding to above 5 cm within the first 5 years with an optimal sensitivity and specificity of 0.73 and 0.71, respectively (P = 0.015). The aortic matrix degradation in AAA may be partly caused by an activation of plasminogen by tPA, but not by uPA, which usually dominates matrix degradation. Smoking seems to be an important factor for this pathway, while the pathway of IgA-CP seems different.

Lindholt, Jes Sanddal

2006-01-01

139

Plasminogen Activator Inhibitor-1 is an Aggregate Response Factor with Pleiotropic Effects on Cell Signaling in Vascular Disease and the Tumor Microenvironment  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In hemostasis, the serine protease inhibitor (serpin) plasminogen activator inhibitor-1 (PAI-1) functions to stabilize clots via inhibition of tissue plasminogen activator (tPA) with subsequent inhibition of fibrinolysis. In tissues, PAI-1 functions to inhibit extracellular matrix degradation via inhibition of urokinase plasminogen activator (uPA). Elevated levels of PAI-1 in the vasculature and in tissues have long been known to be associated with thrombosis and fibrosis, respectively. Howev...

2010-01-01

140

Plasminogen interaction with Trypanosoma cruzi  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english The ability of Trypanosoma cruzi to interact with plasminogen, the zimogenic form of the blood serin protease plasmin, was examined. Immunohistochemistry studies revealed that both forms, epimastigotes and metacyclic trypomastigotes, were able to fix plasminogen in a lysine dependant manner. This in [...] teraction was corroborated by plasminogen activation studies. Both forms of the parasite enhanced the plasminogen activation by tissue-type plasminogen activator.The maximal enhancements obtained were 15-fold and 3.4-fold with epimastigotes and metacyclic trypomastigotes, respectively, as compared to plasminogen activation in absence of cells. Ligand-blotting analysis of proteins extracted with Triton X-114 from a microsomal fraction of epimastigotes revealed at least five soluble proteins and one hydrophobic protein able to bind plasminogen.

Laura, Almeida; Gilmer, Vanegas; Marina, Calcagno; Juan Luis, Concepción; Luisana, Avilan.

 
 
 
 
141

Secretion of Active Recombinant Human Tissue Plasminogen Activator Derivatives in Escherichia coli  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The DNA fragment coding for kringle 2 plus serine protease domains (K2S) of tissue plasminogen activator (tPA) was inserted into a phagemid vector, pComb3HSS. In the recombinant vector, pComb3H-K2S, the K2S gene was fused to gpIII of ?M13 and linked to the OmpA signal sequence. The resulting gene, rK2S-gpIII, was inducibly expressed in Escherichia coli XL-1 Blue. The protein was presented on the phage particle. To stop the expression of gpIII, a stop codon between K2S and the gpIII gene was ...

Manosroi, Jiradej; Tayapiwatana, Chatchai; Go?tz, Friedrich; Werner, Rolf G.; Manosroi, Aranya

2001-01-01

142

Electrocortical activity in the near-term ovine fetus: automated analysis using amplitude frequency components.  

Science.gov (United States)

We have designed an automated method for analyzing electrocortical (ECoG) activity in the near-term ovine fetus to process and quantitatively classify large amounts of data rapidly and objectively. Seven chronically catheterized fetal sheep were studied for 8h each at ~0.9 of gestation with continuous recording of ECoG activity using a computerized data acquisition system. Multiple ECoG amplitude and frequency parameters were scored from which we established animal specific parameter cut-off values as well as population based duration cut-off values to distinguish low-voltage/high frequency (LV/HF) and high-voltage/low frequency (HV/LF) state epochs, and indeterminate voltage/frequency (IV/F) and transition period activities. We have shown that the incidence of the predominant LV/HF and HV/LF activity states at 45% and 36% of the time, respectively, is comparable to that previously reported using semi-quantitative techniques with visual analysis. However, the duration of these state epochs is considerably shorter due to the detection of brief periods of IV/F activity which would be difficult to capture using visual analysis. Importantly, our findings in the healthy ovine fetus near-term using this automated ECoG scoring methodology now provide a framework from which to study maturational events in younger animals, and under adverse pregnancy conditions. PMID:21665193

Keen, Ashley E; Frasch, Martin G; Sheehan, Melissa A; Matushewski, Brad J; Richardson, Bryan S

2011-07-21

143

Stability of Recombinant Tissue Plasminogen Activator at ?30 °C over One Year  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Recombinant tissue plasminogen activator (rt-PA) is used to restore patency and avoid inadvertent removal of peripheral and central venous catheters. rt-PA was reconstituted (1 mg/mL) then cryopreserved at ?30 °C for 1, 2, 3, 6, 8, and 12 months and, then its stability was determined. After cryopreservation for one and two months, rt-PA kept more than 95% of its activity compared to standard samples, while cryopreservation for three months caused 8% loss of activity. However, after...

2013-01-01

144

A catalytic switch and the conversion of streptokinase to a fibrin-targeted plasminogen activator  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen (Pg) activators such as streptokinase (SK) save lives by generating plasmin to dissolve blood clots. Some believe that the unique ability of SK to activate Pg in the absence of fibrin limits its therapeutic utility. We have found that SK contains an unusual NH2-terminal “catalytic switch” that allows Pg activation through both fibrin-independent and fibrin-dependent mechanisms. Unlike SK, a mutant (rSK?59) fusion protein lacking the 59 NH2-terminal residues was no longer capa...

Reed, Guy L.; Houng, Aiilyan K.; Liu, Lin; Parhami-seren, Behnaz; Matsueda, Lee H.; Wang, Shunguang; Hedstrom, Lizbeth

1999-01-01

145

Construction and expression of a recombinant antibody-targeted plasminogen activator  

Energy Technology Data Exchange (ETDEWEB)

Covalent linkage of tissue-type plasminogen activator (t-PA) to a monoclonal antibody specific for the fibrin ..beta.. chain (anti-fibrin 59D8) results in a thrombolytic agent that is more specific and more potent that t-PA alone. To provide a ready source of this hybrid molecule and to allow tailoring of the active moieties for optimal activity, the authors have engineered a recombinant version of the 59D8-t-PA conjugate. The rearranged 59D8 heavy chain gene was cloned and combined in the expression vector pSV2gpt with sequence coding for a portion of the ..gamma..2b constant region and the catalytic ..beta.. chain of t-PA. This construct was transfected into heavy chain loss variant cells derived form the 59D8 hybridoma. Recombinant protein was purified by affinity chromatography and analyzed with electrophoretic transfer blots and radioimmunoassay. These revealed a 65-kDa heavy chain-t-PA fusion protein that is secreted in association with the 59D8 light chain in the form of a 170-kDa disulfide-linked dimer. Chromogenic substrate assays showed the fusion protein to have 70% of the peptidolytic activity of native t-PA and to activate plasminogen as efficiently as t-PA. IN a competitive binding assay, reconstituted antibody was shown to have a binding profile similar to that of native 59D8. Thus, by recombinant techniques, they have produced a hybrid protein capable of high affinity fibrin binding and plasminogen activation.

Schnee, J.M.; Runge, M.S.; Matsueda, G.A.; Hudson, N.W.; Seidman, J.G.; Haber, E.; Quertermous, T.

1987-10-01

146

Metals affect the structure and activity of human plasminogen activator inhibitor-1. I. Modulation of stability and protease inhibition  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Human plasminogen activator inhibitor type 1 (PAI-1) is a serine protease inhibitor with a metastable active conformation. Under physiological conditions, half of the inhibitor transitions to a latent state within 1–2 h. The interaction between PAI-1 and the plasma protein vitronectin prolongs this active lifespan by ?50%. Previously, our group demonstrated that PAI-1 binds to resins using immobilized metal affinity chromatography (Day, U.S. Pat. 7,015,021 B2, March 21, 2006). In this stu...

Thompson, Lawrence C.; Goswami, Sumit; Ginsberg, David S.; Day, Duane E.; Verhamme, Ingrid M.; Peterson, Cynthia B.

2011-01-01

147

Metals affect the structure and activity of human plasminogen activator inhibitor-1. II. Binding affinity and conformational changes  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Human plasminogen activator inhibitor type 1 (PAI-1) is a serine protease inhibitor with a metastable active conformation. The lifespan of the active form of PAI-1 is modulated via interaction with the plasma protein, vitronectin, and various metal ions. These metal ions fall into two categories: Type I metals, including calcium, magnesium, and manganese, stabilize PAI-1 in the absence of vitronectin, whereas Type II metals, including cobalt, copper, and nickel, destabilize PAI-1 in the absen...

Thompson, Lawrence C.; Goswami, Sumit; Peterson, Cynthia B.

2011-01-01

148

von Willebrand factor antigen and plasminogen activator inhibitor in giant cell arteritis.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Sixty three patients (51 women, 12 men) with giant cell arteritis were studied by serial analyses of von Willebrand factor antigen (vWF: Ag) concentration and plasminogen activator inhibitor activity. Their mean age at the time of diagnosis was 71 years. Two hundred and one randomly selected subjects from the general population, aged 75 years, served as controls. The mean concentration of vWF:Ag in the patients with giant cell arteritis before the start of corticosteroid treatment was 2.63 (S...

Nordborg, E.; Andersson, R.; Tengborn, L.; Ede?n, S.; Bengtsson, B. A.

1991-01-01

149

Localization of tissue plasminogen activator in the endothelium of a limited number of vessels.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The immunolocalization of tissue plasminogen activator (tPA) was assessed in vessels of various sizes from baboons. Femoral artery and vein, carotid artery, aorta, and sections from basal ganglia and cerebral cortex were stained for tPA and CD31, an endothelial cell-specific surface antigen. In each case, the endothelium of the large vessel stained positively for anti-CD31 but not for tPA. However, vascular structures in the adventitia corresponding to the vasa vasorum were found to be associ...

Levin, E. G.; Del Zoppo, G. J.

1994-01-01

150

Management of plastic bronchitis with nebulized tissue plasminogen activator: another brick in the wall  

Science.gov (United States)

Plastic bronchitis is a rare complication of a variety of respiratory diseases and congenital heart disease surgery, particularly Fontan procedure. Bronchial casts with rubber-like consistency develop acutely and may cause severe life-threatening respiratory distress. The management of plastic bronchitis is yet not well defined. Early intermittent, self-administered nebulization of tissue plasminogen activator was found to be effective in preventing deterioration of acute respiratory symptoms in a patient with primary ciliary dyskinesia and recurrent cast formation. Further investigation into new therapeutic strategies for this devastating disease is advocated.

2014-01-01

151

Expression of Plasminogen Activator Pla of Yersinia pestis Enhances Bacterial Attachment to the Mammalian Extracellular Matrix  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The effect of the plasminogen activator Pla of Yersinia pestis on the adhesiveness of bacteria to the mammalian extracellular matrix was determined. Y. pestis KIM D27 harbors the 9.5-kb plasmid pPCP1, encoding Pla and pesticin; the strain efficiently adhered to the reconstituted basement membrane preparation Matrigel, to the extracellular matrix prepared from human lung NCI-H292 epithelial cells, as well as to immobilized laminin. The isogenic strain Y. pestis KIM D34 lacking pPCP1 exhibited ...

La?hteenma?ki, Kaarina; Virkola, Ritva; Sare?n, Anne; Emo?dy, Levente; Korhonen, Timo K.

1998-01-01

152

Soluble fibrin degradation products potentiate tissue plasminogen activator-induced fibrinogen proteolysis.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Despite its affinity for fibrin, tissue plasminogen activator (t-PA) administration causes systemic fibrinogenolysis. To investigate the mechanism, t-PA was incubated with plasma in the presence or absence of a fibrin clot, and the extent of fibrinogenolysis was determined by measuring B beta 1-42. In the presence of fibrin, there is a 21-fold increase in B beta 1-42 levels. The potentiation of fibrinogenolysis in the presence of fibrin is mediated by soluble fibrin degradation products becau...

Weitz, J. I.; Leslie, B.; Ginsberg, J.

1991-01-01

153

Interconversion of active and inactive conformations of urokinase-type plasminogen activator  

DEFF Research Database (Denmark)

The catalytic activity of serine proteases depends on a salt-bridge between the amino group of residue 16 and the side chain of Asp194. The salt-bridge stabilizes the oxyanion hole and the S1 specificity pocket of the protease. Some serine proteases exist in only partially active forms, in which the amino group of residue 16 is exposed to the solvent. Such a partially active state is assumed by a truncated form of the murine urokinase-type plasminogen activator (muPA), consisting of residues 16-243. Here we investigated the allosteric interconversion between partially active states and the fully active state. Both a monoclonal antibody (mU3) and a peptidic inhibitor (mupain-1--16) stabilize the active state. The epitope of mU3 is located in the 37- and 70-loops at a site homologous to exosite I of thrombin. The N-terminus((Ile16)) of muPA((16--243)) was less exposed upon binding of mU3 or mupain-1--16. In contrast, introduction of the mutations F40Y or E137A into muPA((16--243)) increased exposure of the N-terminus((Ile16)) and resulted in large changes in the thermodynamic parameters for mupain-1--16 binding. We conclude that the distorted state of muPA((16--243)) is conformationally ordered upon binding of ligands to the active site and upon binding of mU3 to the 37- and 70-loops. Our study establishes the 37- and 70-loops as a unique site for binding to compounds stabilizing the active state of serine proteases.

Liu, Zhuo; Kromann-Hansen, Tobias

2012-01-01

154

Interconversion of Active and Inactive Conformations of Urokinase-Type Plasminogen Activator  

DEFF Research Database (Denmark)

The catalytic activity of serine proteases depends on a salt-bridge between the amino group of residue 16 and the side chain of Asp194. The salt-bridge stabilizes the oxyanion hole and the S1 specificity pocket of the protease. Some serine proteases exist in only partially active forms, in which the amino group of residue 16 is exposed to the solvent. Such a partially active state is assumed by a truncated form of the murine urokinase-type plasminogen activator (muPA), consisting of residues 16-243. Here we investigated the allosteric interconversion between partially active states and the fully active state. Both a monoclonal antibody (mU3) and a peptidic inhibitor (mupain-1-16) stabilize the active state. The epitope of mU3 is located in the 37- and 70-loops at a site homologous to exosite I of thrombin. The N-terminus((Ile16)) of muPA((16-243)) was less exposed upon binding of mU3 or mupain-1-16. In contrast, introduction of the mutations F40Y or E137A into muPA((16-243)) increased exposure of the N-terminus((Ile16)) and resulted in large changes in the thermodynamic parameters for mupain-1-16 binding. We conclude that the distorted state of muPA((16-243)) is conformationally ordered upon binding of ligands to the active site and upon binding of mU3 to the 37- and 70-loops. Our study establishes the 37- and 70-loops as a unique site for binding to compounds stabilizing the active state of serine proteases.

Liu, Zhuo; Kromann-Hansen, Tobias

2012-01-01

155

Biological mechanisms underlying the clinical effects of RU 486: modulation of cultured endometrial stromal cell plasminogen activator and plasminogen activator inhibitor expression.  

Science.gov (United States)

The abortifacient and menstrual effects of the potent antiprogestin, RU 486, are associated with both endometrial hemorrhage and extracellular matrix (ECM) degradation. Such processes reflect reduced perivascular decidual cell hemostatic and increased ECM-degrading protease activity. Therefore, we assessed the effects of RU 486 administration on the expression of immunoreactive (ir) endometrial stromal cell urokinase-type (uPA) and tissue-type (tPA) plasminogen activator and their activities as well as levels of ir type 1 plasminogen activator inhibitor (PAI-1) using a well characterized in vitro model of decidualization. Thus, confluent stromal cell cultures were exposed to vehicle control, 10(-8) mol/L estradiol (E2), 10(-7)-10(-8) mol/L medroxyprogesterone acetate (MPA), E2 plus MPA, or 10(-6)-10(-7) mol/L RU 486 alone or in combination with MPA or E2 plus MPA for 3-4 days. Compared to the vehicle control, E2 and RU 486 used alone had no effect on levels of ir PAI-1, uPA, or tPA or on PA activity in the conditioned medium. In contrast, MPA and E2 plus MPA decreased ir uPA and tPA levels and their corresponding activities, whereas MPA increased, and E2 plus MPA further increased ir PAI-1 release. These effects of progestin were blocked by a log higher concentration of RU 486. Similar results were obtained for steady state PAI-1 messenger ribonucleic acid levels. To determine if RU 486 reversed progestin-inhibited stromal cell uPA and tPA release and progestin-enhanced PAI-1 expression, confluent cultures were exposed to 10(-8) mol/L E2 plus 10(-7) mol/L MPA for 10 days, washed, and reexposed to E2 plus MPA, steroid-free medium, or RU 486 for 3-5 or 9-11 days. Compared with cultures maintained in E2 plus MPA for 3-5 days, withdrawal to a steroid-free medium failed to affect stromal cell ir PAI-1, uPA, or tPA levels. In contrast, exposure to RU 486 for 3-5 days increased ir uPA and tPA levels 5- to 8-fold (P dissolution. PMID:7714076

Lockwood, C J; Krikun, G; Papp, C; Aigner, S; Schatz, F

1995-04-01

156

Plasminogen activator inhibitor-1 polymers, induced by inactivating amphipathic organochemical ligands.  

DEFF Research Database (Denmark)

Negatively charged organochemical inactivators of the anti-proteolytic activity of plasminogen activator inhibitor-1 (PAI-1) convert it to inactive polymers. As investigated by native gel electrophoresis, the size of the PAI-1 polymers ranged from dimers to multimers of more than 20 units. As compared with native PAI-1, the polymers exhibited an increased resistance to temperature-induced unfolding. Polymerization was associated with specific changes in patterns of digestion with non-target proteases. During incubation with urokinase-type plasminogen activator, the polymers were slowly converted to reactive centre-cleaved monomers, indicating substrate behaviour of the terminal PAI-1 molecules in the polymers. A quadruple mutant of PAI-1 with a retarded rate of latency transition also had a retarded rate of polymerization. Studying a number of serpins by native gel electrophoresis, ligand-induced polymerization was observed only with PAI-1 and heparin cofactor II, which were also able to copolymerize. On the basis of these results, we suggest that the binding of ligands in a specific region of PAI-1 leads to so-called loop-sheet polymerization, in which the reactive centre loop of one molecule binds to beta-sheet A in another molecule. Induction of serpin polymerization by small organochemical ligands is a novel finding and is of protein chemical interest in relation to pathological protein polymerization in general. Udgivelsesdato: 2003-Jun-15

Pedersen, Katrine E; Einholm, Anja P

2003-01-01

157

Serum-stable RNA aptamers to urokinase-type plasminogen activator blocking receptor binding  

DEFF Research Database (Denmark)

The serine proteinase urokinase-type plasminogen activator (uPA) is widely recognized as a potential target for anticancer therapy. Its association with cell surfaces through the uPA receptor (uPAR) is central to its function and plays an important role in cancer invasion and metastasis. In the current study, we used systematic evolution of ligands by exponential enrichment (SELEX) to select serum-stable 2'-fluoro-pyrimidine-modified RNA aptamers specifically targeting human uPA and blocking the interaction to its receptor at low nanomolar concentrations. In agreement with the inhibitory function of the aptamers, binding was found to be dependent on the presence of the growth factor domain of uPA, which mediates uPAR binding. One of the most potent uPA aptamers, upanap-12, was analyzed in more detail and could be reduced significantly in size without severe loss of its inhibitory activity. Finally, we show that the uPA-scavenging effect of the aptamers can reduce uPAR-dependent endocytosis of the uPA-PAI-1 complex and cell-surface associated plasminogen activation in cell culture experiments. uPA-scavenging 2'-fluoro-pyrimidine-modified RNA aptamers represent a novel promising principle for interfering with the pathological functions of the uPA system.

Dupont, Daniel Miotto; Madsen, Jeppe Buur

2010-01-01

158

Cleaved intracellular plasminogen activator inhibitor 2 in human myeloleukaemia cells is a marker of apoptosis  

DEFF Research Database (Denmark)

The proteolytic modification of plasminogen activator inhibitor 2 (PAI-2) was studied during apoptosis in the human promyelocytic leukaemic NB4 cell line during treatment with the phosphatase inhibitors okadaic acid and calyculin A as well as the protein synthesis inhibitor cycloheximide. The apoptic type of cell death was ascertained by morphological and biochemical criteria. In cell homogenates PAI-2 was probed by [125I]urokinase plasminogen activator (uPA) and detected as a sodium dodecyl sulphate-stable M(r) 80,000 complex after reducing sodium dodecyl sulphate-polyacrylamide gel electrophoresis and autoradiography. During apoptosis a smaller (M(r) 70,000) uPA-PAI-2 complex was consistently detected. The modification was in the PAI-2 moiety, as the [125I]uPA tracer could be extracted in its intact form from the complex. Thus the cleaved PAI-2 isoform is a biochemical marker of apoptosis in the promyelocytic NB4 cell line. The modified PAI-2 isoform was also detected in homogenates made from purified humanmononuclear leukaemic cells aspirated from the bone marrow of patients suffering from acute and chronic myeloid leukaemia.

Jensen, P H; Cressey, L I

1994-01-01

159

Tissue plasminogen activator for the treatment of parapneumonic effusions in pediatric patients.  

Science.gov (United States)

Intrapleural fibrinolysis has been investigated for the treatment of pleural effusion for several decades. Fibrinolytics have the ability to break up fibrin and loculations that characterize complicated pleural effusions, facilitating drainage. Older fibrinolytics such as urokinase and streptokinase have been replaced by tissue plasminogen activator (tPA) for this indication due to product availability and a more favorable safety profile. The literature supports tPA as a treatment approach for this indication in adult patients, and the use of tPA has become a standard management approach in this population. Over the past decade, data on the efficacy of intrapleural fibrinolytic therapy in children have also been generated, which now support the use of fibrinolysis as a treatment alternative to more invasive therapeutic options such as surgical intervention. In this review, we discuss the pathophysiology, diagnosis, and treatment of parapneumonic effusion and empyema, with a focus on intrapleural fibrinolysis, specifically tissue plasminogen activator, in the pediatric population. Recent articles provide sufficient evidence to support the use of this drug in pediatric patients for the management of pleural effusions; however, due to study heterogeneity, questions remain that may be addressed in future studies. PMID:24390857

Israel, Emily N; Blackmer, Allison B

2014-05-01

160

Increased expression of urokinase plasminogen activator and its cognate receptor in human seminomas  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background The urokinase plasminogen activating system (uPAS is implicated in neoplastic progression and high tissue levels of uPAS components correlate with a poor prognosis in different human cancers. Despite that, relative few studies are available on the expression and function of the uPAS components in human seminomas. In the present study we characterized the expression of the urokinase plasminogen activator (uPA, its cognate receptor (uPAR and the uPA inhibitors PAI-1 and PAI-2 in normal human testis and seminomas. Methods The expression of the above genes was evaluated by means of quantitative RT-PCR, western blot, zymographic analysis and immunohistochemistry. Results Quantitative RT-PCR analysis of 14 seminomas demonstrated that uPA and uPAR mRNAs were, with respect to control tissues, increased in tumor tissues by 3.80 ± 0.74 (p Conclusions We demonstrated the increased expression of uPA and uPAR in human seminomas with respect to normal testis tissues, which may be relevant in testicular cancer progression.

Sorrenti Salvatore

2010-04-01

 
 
 
 
161

Regulation of programmed cell death by plasminogen activator inhibitor type 1 (PAI-1)  

DEFF Research Database (Denmark)

Et forhøjet niveau af plasminogen activator inhibitor-1 (PAI-1) er forbundet med dårlig prognose i kræft. En forklaring på det forhøjede niveau af PAI-1 kunne være et beskyttende respons på en forøget proteolytisk aktivitet forårsaget af et forhøjet niveau af urokinase-type plasminogen activator (uPA) som er observeret i tumorer. En lang række af videnskabelige arbejder peger derimod i retning af at PAI-1 i sig selv bidrager til sydomsudviklingen. PAI-1 er blevet rapporteret at ahve indvikning på de fleste basale cellulære mekanismer herunder, celle-adhæsion, celle-migration, celle-proliferation og et stigernde antal af videnskabelige artikler indikere at PAI-1 også kan regulere programmeret celle-død (PCD) in kræft-celler og normale celler. I en række videnskabelige arbejder indikere, at PAI-1 can hæmme PCD gennem PAI-1's pro-adhæsive/anti-proteolytiske egenskab, hvorimod andre videnskabelige arbejder indikere at PAI-1 enten kan inducere PCD gennem PAI-1's anti-adhæsive egenskab.

Lademann, Ulrik Axel; Rømer, Maria Unni Koefoed

2008-01-01

162

Expression of biologically active kringle 5 domain of human plasminogen in Escherichia coli.  

Science.gov (United States)

The kringle 5 domain of plasminogen exhibits potent inhibitory effect on endothelial cell proliferation. It can also cause cell cycle arrest and apoptosis of endothelial cell specifically, and shows promise in anti-angiogenic therapy. It has been prepared via both proteolysis of native plasminogen and recombinant DNA methodologies. When previously expressed in Escherichia coli, recombinant kringle 5 mainly deposited as inactive, insoluble inclusion bodies and the refolding yield was low. In the present study, human kringle 5 was fusion-expressed with GST (gluthathione-S-transferase) under the control of T7 promoter in E. coli. The IPTG-induced GST-kringle 5 was about 20% of the total cellular proteins and, among the expressed GST-kringle 5 proteins, 80% was present in the supernatant. The GST-kringle 5 fusion protein exhibited some anti-proliferation activity towards bovine capillary endothelial cells. After GST-kringle 5 purification, subsequent enterokinase release of intact kringle 5 from the fusion protein and further purification by gluthathione-Sepharose 4B affinity chromatography, the recombinant kringle 5, with a yield of 10.5 mg/L culture, displayed apparent inhibition of endothelial cell proliferation in a dose-dependent manner with ED50 about 20 nM. PMID:15704494

Zhang, Hong-Xia; Fang, Lei; Cheng, Jian; Wei, Zheng; Hua, Zi-Chun

2005-01-01

163

Both u-PA inhibition and vitronectin binding by plasminogen activator inhibitor 1 regulate HT1080 fibrosarcoma cell metastasis.  

Science.gov (United States)

Overexpression of plasminogen activator inhibitor 1 (PAI-1) reduces tumor cell migration in vitro and metastasis in mice in vivo by mechanisms involving either inhibition of urokinase plasminogen activator (u-PA) activity or competition for an integrin binding site on vitronectin. To analyze the effects of PAI-1 on tumor cell migration in vitro and metastasis in vivo, recombinant adenoviral vectors expressing wild-type or mutant PAI-1 proteins were constructed. The mutant PAI-1 proteins were defective in either vitronectin binding (PAI-1(VN-)), plasminogen activator inhibition (PAI-1(INH-)) or both (PAI-1(VN-,INH-)). In vitro, migration of HT1080 human fibrosarcoma cells through a reconstituted extracellular matrix (ECM) was reduced 73% by overexpression of wild-type PAI-1 and 65% by PAI-1(VN-) compared with control virus-infected cells. Migration of cells infected by virus expressing either PAI-1(INH-) or PAI-1(VN-,INH-) was unaffected, indicating a requirement for plasminogen activator inhibitory activity. In vivo, however, only overexpression of wild-type PAI-1 reduced the burden of metastasis by 68% compared with the control group. This indicates that both u-PA inhibition and PAI-1 ECM interactions contribute to the mechanism of PAI-1-mediated regulation of cell migration. PMID:12447999

Praus, Michael; Collen, Désiré; Gerard, Robert D

2002-12-20

164

Expression of plasminogen activators in preimplantation rat embryos developed in vivo and in vitro  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Embryo implantation plays a major role in embryogenesis and the outcome of pregnancy. Plasminogen activators (PAs have been implicated in mammalian fertilization, early stages of development and embryo implantation. The invasion of trophoblast cells into the endometrium during the implantation process can be blocked by inhibitors of serine proteases, illustrating the role of these enzymes in the invasion process. As in vitro developing embryos resulted in lower implantation rate than those developed in vivo we assume that a reduced PAs activity may lead to it. There is hardly any information regarding qualitative or quantitative differences in expression of PAs in preimplantation embryos, or comparisons between in vivo and in vitro developed embryos. The purpose of this study was to assess the expression of urokinase type (uPA and tissue type (tPA plasminogen activators in in vivo and in vitro preimplantation development in rat embryos using immunofluorescence confocal microscopy and computerized image analysis. Methods Zygotes, 2-cell, 4-cell, 8-cell, morula and blastocyst stages of development were flushed from the reproductive tract (control groups of Wistar rats. Zygotes were flushed and grown in vitro to the above mentioned developmental stages and comprised the experimental groups. Immunofluorescence microscopy and computerized image analysis were used to evaluate both qualitative (localization and quantitative expression of plasminogen activators. Results uPA and tPA were found to be expressed in rat embryos throughout their preimplantation development, both in vivo and in vitro. While uPA was localized mainly in the cell cytoplasm, the tPA was detected mainly on cell surface and in the perivitelline space. In blastocysts, both in vivo and in vitro, uPA and tPA were localized in the trophectoderm cells. Total uPA content per embryo was higher in the in vivo as compared with the in vitro developed embryos at all stages measured. Blastocyst uPA content was significantly low as compared with the four-cell, eight-cell, and morula stages. Total tPA content was higher in embryos developed in vivo than those developed in vitro except for the 4-cell and 8-cell stages. Conclusion In vitro embryo development leads to lower PAs expression in a stage dependent manner as compared with in vivo developing controls. The enzymes studied vary probably in the ratio of their active and inactive forms as there is no correlation between their content and the activity observed in our previous study. The localization of both PAs in the blastocysts' trophectoderm supports the assumption that PAs plays a role in the implantation process in rats.

Har-Vardi Iris

2005-02-01

165

Clinical significance of plasminogen activator inhibitor activity in patients with exercise-induced ischemia  

International Nuclear Information System (INIS)

To assess the fibrinolytic system in patients with exercise-induced ischemia and its relation to ischemia and severity of coronary artery disease (CAD), 47 patients with CAD confirmed by results of coronary angiography underwent symptom-limited multistage exercise thallium-201 emission computed tomography. All patients with CAD had exercise-induced ischemia as assessed from thallium-201 images. Pre- and peak exercise blood samples from each patient and preexercise blood samples from control subjects were assayed for several fibrinolytic components and were also assayed for plasma adrenaline. The extent of ischemia was defined as delta visual uptake score (total visual uptake score in delayed images minus total visual uptake score in initial images) and the severity of CAD as the number of diseased vessels. In the basal condition, plasminogen activator inhibitor (PAI) activity was significantly higher in patients with exercise-induced ischemia as compared to control subjects (p less than 0.01), although there were no significant differences in other fibrinolytic variables between the two groups. Moreover, PAI activity in the basal condition displayed a significantly positive correlation with the extent of ischemia (r = 0.47, p less than 0.01). Patients with exercise-induced ischemia were divided into two groups (24 with single-vessel disease and 23 with multivessel disease). There were no significant differences in coronary risk factors, hemodynamics, or plasma adrenaline levels during exercise between single-vessel and multivessel disease except that delta visual uptake score was significantly higher in multivessel disease (p less than 0.01)

1990-01-01

166

Clinical significance of plasminogen activator inhibitor activity in patients with exercise-induced ischemia  

Energy Technology Data Exchange (ETDEWEB)

To assess the fibrinolytic system in patients with exercise-induced ischemia and its relation to ischemia and severity of coronary artery disease (CAD), 47 patients with CAD confirmed by results of coronary angiography underwent symptom-limited multistage exercise thallium-201 emission computed tomography. All patients with CAD had exercise-induced ischemia as assessed from thallium-201 images. Pre- and peak exercise blood samples from each patient and preexercise blood samples from control subjects were assayed for several fibrinolytic components and were also assayed for plasma adrenaline. The extent of ischemia was defined as delta visual uptake score (total visual uptake score in delayed images minus total visual uptake score in initial images) and the severity of CAD as the number of diseased vessels. In the basal condition, plasminogen activator inhibitor (PAI) activity was significantly higher in patients with exercise-induced ischemia as compared to control subjects (p less than 0.01), although there were no significant differences in other fibrinolytic variables between the two groups. Moreover, PAI activity in the basal condition displayed a significantly positive correlation with the extent of ischemia (r = 0.47, p less than 0.01). Patients with exercise-induced ischemia were divided into two groups (24 with single-vessel disease and 23 with multivessel disease). There were no significant differences in coronary risk factors, hemodynamics, or plasma adrenaline levels during exercise between single-vessel and multivessel disease except that delta visual uptake score was significantly higher in multivessel disease (p less than 0.01).

Sakata, K.; Kurata, C.; Taguchi, T.; Suzuki, S.; Kobayashi, A.; Yamazaki, N.; Rydzewski, A.; Takada, Y.; Takada, A. (Hamamatsu Univ. School of Medicine, Shizuoka (Japan))

1990-10-01

167

Specific interaction of tissue-type plasminogen activator (t-PA) with annexin II on the membrane of pancreatic cancer cells activates plasminogen and promotes invasion in vitro  

Science.gov (United States)

Background: Overexpression of tissue plasminogen activator (t-PA) in pancreatic cancer cells promotes invasion and proliferation in vitro and tumour growth and angiogenesis in vivo. Aims: To understand the mechanisms by which t-PA favours cancer progression, we analysed the surface membrane proteins responsible for binding specifically t-PA and studied the contribution of this interaction to the t-PA promoted invasion of pancreatic cancer cells. Methods: The ability of t-PA to activate plasmin and a fluorogenic plasmin substrate was used to analyse the nature of the binding of active t-PA to cell surfaces. Specific binding was determined in two pancreatic cancer cell lines (SK-PC-1 and PANC-1), and complex formation analysed by co-immunoprecipitation experiments and co-immunolocalisation in tumours. The functional role of the interaction was studied in Matrigel invasion assays. Results: t-PA bound to PANC-1 and SK-PC-1 cells in a specific and saturable manner while maintaining its activity. This binding was competitively inhibited by specific peptides interfering with the interaction of t-PA with annexin II. The t-PA/annexin II interaction on pancreatic cancer cells was also supported by co-immunoprecipitation assays using anti-t-PA antibodies and, reciprocally, with antiannexin II antibodies. In addition, confocal microscopy showed t-PA and annexin II colocalisation in tumour tissues. Finally, disruption of the t-PA/annexin II interaction by a specific hexapeptide significantly decreased the invasive capacity of SK-PC-1 cells in vitro. Conclusion: t-PA specifically binds to annexin II on the extracellular membrane of pancreatic cancer cells where it activates local plasmin production and tumour cell invasion. These findings may be clinically relevant for future therapeutic strategies based on specific drugs that counteract the activity of t-PA or its receptor annexin II, or their interaction at the surface level.

Diaz, V M; Hurtado, M; Thomson, T M; Reventos, J; Paciucci, R

2004-01-01

168

Urokinase Plasminogen Activator Receptor (uPAR) and Plasminogen Activator Inhibitor-1 (PAI-1) Are Potential Predictive Biomarkers in Early Stage Oral Squamous Cell Carcinomas (OSCC)  

Science.gov (United States)

Oral squamous cell carcinoma (OSCC) is often associated with metastatic disease and a poor 5 year survival rate. Patients diagnosed with small tumours generally have a more favourable outcome, but some of these small tumours are aggressive and lead to early death. To avoid harmful overtreatment of patients with favourable prognosis, there is a need for predictive biomarkers that can be used for treatment stratification. In this study we assessed the possibility to use components of the plasminogen activator (PA) system as prognostic markers for OSCC outcome and compared this to the commonly used biomarker Ki-67. A tissue-micro-array (TMA) based immunohistochemical analysis of primary tumour tissue obtained from a North Norwegian cohort of 115 patients diagnosed with OSCC was conducted. The expression of the biomarkers was compared with clinicopathological variables and disease specific death. The statistical analyses revealed that low expression of uPAR (p?=?0.031) and PAI-1 (p?=?0.021) in the tumour cells was significantly associated with low disease specific death in patients with small tumours and no lymph node metastasis (T1N0). The commonly used biomarker, Ki-67, was not associated with disease specific death in any of the groups of patients analysed. The conclusion is that uPAR and PAI-1 are potential predictive biomarkers in early stage tumours and that this warrants further studies on a larger cohort of patients.

Hadler-Olsen, Elin; Uhlin-Hansen, Lars; Svineng, Gunbj?rg

2014-01-01

169

Radioimmunoreactivity and receptor-binding activity of the recombined molecule obtained by complementation of two fibrinolysin fragments of ovine prolactin.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The recombined molecule obtained by complementation of two fibrinolysin fragments of ovine prolactin (oPRL) has been characterized by radioimmunoassay and radioreceptor assay. The recombinant alone exhibits very low radioimmunoreactivity and radioreceptor activity. However, in the presence of excess fragment oPRL-(1-53), which does not compete with oPRL in either assay, the recombinant has 101% of the radioimmunoreactivity and only 13% of the radioreceptor activity. The result shows that the ...

Wong, T. M.; Cheng, C. H.; Li, C. H.

1981-01-01

170

Reduced cardiac expression of plasminogen activator inhibitor 1 and transforming growth factor ?1 in obese Zucker rats by perindopril  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Objective: To determine whether angiotensin converting enzyme inhibition by perindopril can reduce cardiac transforming growth factor ?1 (TGF?1) and plasminogen activator inhibitor 1 (PAI-1) and therefore control collagen accumulation in an animal model with the metabolic syndrome such as the obese Zucker rat (OZR).

Toblli, J. E.; Cao, G.; Derosa, G.; Forcada, P.

2005-01-01

171

Xanthoangelols isolated from Angelica keiskei inhibit inflammatory-induced plasminogen activator inhibitor 1 (PAI-1) production.  

Science.gov (United States)

The folk medicine Angelica keiskei (Ashitaba) exhibits antitumor, antioxidant and antidiabetic activities and it has recently attracted attention as a health food. Ashitaba is thought to have antithrombotic properties, but this has not yet been scientifically proven. The elevation of plasma plasminogen activator inhibitor 1 (PAI-1), an inhibitor of fibrinolysis results in a predisposition to the risk of thrombosis. The present study showed that Ashitaba exudates injected intraperitoneally and orally administered over long-term suppressed the lipopolysaccharide (LPS) induced PAI-1 increase in mouse plasma. We also found that xanthoangelol, xanthoangelols B and D, the components of Ashitaba exudates, significantly inhibited TNF?-induced PAI-1 production from human umbilical vein endothelial cells (HUVECs). These findings suggest that Ashitaba can decrease elevated PAI-1 production, and that daily consumption of Ashitaba product might maintain anticoagulant status by inhibiting elevations in PAI-1 under inflammatory conditions. PMID:22038782

Ohkura, Naoki; Nakakuki, Yoshitaka; Taniguchi, Masahiko; Kanai, Shiho; Nakayama, Akiko; Ohnishi, Katsunori; Sakata, Toshiyuki; Nohira, Tomoyoshi; Matsuda, Juzo; Baba, Kimiye; Atsumi, Gen-Ichi

2011-01-01

172

A faster-acting and more potent form of tissue plasminogen activator.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Current treatment with tissue plasminogen activator (tPA) requires an intravenous infusion (1.5-3 h) because the clearance of tPA from the circulation is rapid (t 1/2 approximately 6 min). We have developed a tPA variant, T103N,N117Q, KHRR(296-299)AAAA (TNK-tPA) that has substantially slower in vivo clearance (1.9 vs. 16.1 ml per min per kg for tPA in rabbits) and near-normal fibrin binding and plasma clot lysis activity (87% and 82% compared with wild-type tPA). TNK-tPA exhibits 80-fold high...

Keyt, B. A.; Paoni, N. F.; Refino, C. J.; Berleau, L.; Nguyen, H.; Chow, A.; Lai, J.; Pen?a, L.; Pater, C.; Ogez, J.

1994-01-01

173

Plasminogen activator inhibitor-1 suppresses endogenous fibrinolysis in a canine model of pulmonary embolism  

Energy Technology Data Exchange (ETDEWEB)

Plasminogen activator inhibitor-1 (PAI-1), the specific, fast-acting inhibitor of tissue-type plasminogen activator (t-PA), binds to fibrin and has been found in high concentrations within arterial thrombi. These findings suggest that the localization of PAI-1 to a thrombus protects that same thrombus from fibrinolysis. In this study, clot-bound PAI-1 was assessed for its ability to suppress clot lysis in vivo. Autologous, canine whole blood clots were formed in the presence of increasing amounts of activated PAI-1 (0-30 micrograms/ml). Approximately 6-8% of the PAI-1 bound to the clots under the experimental conditions. Control and PAI-1-enriched clots containing iodine-125-labeled fibrin (ogen) were homogenized, washed to remove nonbound elements, and delivered to the lungs of anesthetized dogs where the homogenates subsequently underwent lysis by the endogeneous fibrinolytic system. 125I-labeled fibrin degradation products appeared in the blood of control animals within 10 minutes and were maximal by 90 minutes. PAI-1 reduced fibrin degradation product release in a dose-responsive manner at all times between 30 minutes and 5 hours (greater than or equal to 76% inhibition at 30 minutes, PAI-1 greater than or equal to 6 micrograms/ml). PAI-1 also suppressed D-dimer release from clots containing small amounts of human fibrin (ogen). t-PA administration attenuated the effects of PAI-1, whereas latent PAI-1 (20 micrograms/ml) had no effect on clot lysis. Blood levels of PA and PAI activity remained unaltered during these experiments. The results indicate that PAI-1 markedly inhibits endogenous fibrinolysis in vivo and, moreover, suggest that the localization of PAI-1 to a forming thrombus is an important physiological mechanism for subsequent thrombus stabilization.

Reilly, C.F.; Fujita, T.; Hutzelmann, J.E.; Mayer, E.J.; Shebuski, R.J. (Department of Pharmacology, Merck Sharp Dohme Research Laboratories, West Point, PA (USA))

1991-07-01

174

Plasminogen activator inhibitor-1 suppresses endogenous fibrinolysis in a canine model of pulmonary embolism  

International Nuclear Information System (INIS)

Plasminogen activator inhibitor-1 (PAI-1), the specific, fast-acting inhibitor of tissue-type plasminogen activator (t-PA), binds to fibrin and has been found in high concentrations within arterial thrombi. These findings suggest that the localization of PAI-1 to a thrombus protects that same thrombus from fibrinolysis. In this study, clot-bound PAI-1 was assessed for its ability to suppress clot lysis in vivo. Autologous, canine whole blood clots were formed in the presence of increasing amounts of activated PAI-1 (0-30 micrograms/ml). Approximately 6-8% of the PAI-1 bound to the clots under the experimental conditions. Control and PAI-1-enriched clots containing iodine-125-labeled fibrin (ogen) were homogenized, washed to remove nonbound elements, and delivered to the lungs of anesthetized dogs where the homogenates subsequently underwent lysis by the endogeneous fibrinolytic system. 125I-labeled fibrin degradation products appeared in the blood of control animals within 10 minutes and were maximal by 90 minutes. PAI-1 reduced fibrin degradation product release in a dose-responsive manner at all times between 30 minutes and 5 hours (greater than or equal to 76% inhibition at 30 minutes, PAI-1 greater than or equal to 6 micrograms/ml). PAI-1 also suppressed D-dimer release from clots containing small amounts of human fibrin (ogen). t-PA administration attenuated the effects of PAI-1, whereas latent PAI-1 (20 micrograms/ml) had no effect on clot lysis. Blood levels of PA and PAI activity remained unaltered during these experiments. The results indicate that PAI-1 markedly inhibits endogenous fibrinolysis in vivo and, moreover, suggest that the localization of PAI-1 to a forming thrombus is an important physiological mechanism for subsequent thrombus stabilization

1991-01-01

175

Urokinase-type Plasminogen Activator-like Proteases in Teleosts Lack Genuine Receptor-binding Epidermal Growth Factor-like Domains  

DEFF Research Database (Denmark)

Plasminogen activation catalyzed by urokinase-type plasminogen activator (uPA) plays an important role in normal and pathological tissue remodeling processes. Since its discovery in the mid-1980s, the cell membrane-anchored urokinase-type plasminogen activator receptor (uPAR) has been believed to be central to the functions of uPA, as uPA-catalyzed plasminogen activation activity appeared to be confined to cell surfaces through the binding of uPA to uPAR. However, a functional uPAR has so far only been identified in mammals. We have now cloned, recombinantly produced, and characterized two zebrafish proteases, zfuPA-a and zfuPA-b, which by several criteria are the fish orthologs of mammalian uPA. Thus, both proteases catalyze the activation of fish plasminogen efficiently and both proteases are inhibited rapidly by plasminogen activator inhibitor-1 (PAI-1). But zfuPA-a differs from mammalian uPA by lacking the exon encoding the uPAR-binding epidermal growth factor-like domain; zfuPA-b differs from mammalian uPA by lacking two cysteines of the epidermal growth factor-like domain and a uPAR-binding sequence comparable with that found in mammalian uPA. Accordingly, no zfuPA-b binding activity could be found in fish white blood cells or fish cell lines. We therefore propose that the current consensus of uPA-catalyzed plasminogen activation taking place on cell surfaces, derived from observations with mammals, is too narrow. Fish uPAs appear incapable of receptor binding in the manner known from mammals and uPA-catalyzed plasminogen activation in fish may occur mainly in solution. Studies with nonmammalian vertebrate species are needed to obtain a comprehensive understanding of the mechanism of plasminogen activation.

Bager, René; Kristensen, Thomas Kielsgaard

2012-01-01

176

Urokinase-type plasminogen activator-like proteases in teleosts lack genuine receptor-binding epidermal growth factor-like domains  

DEFF Research Database (Denmark)

Plasminogen activation catalyzed by urokinase-type plasminogen activator (uPA) plays an important role in normal and pathological tissue remodeling processes. Since its discovery in the mid-1980s, the cell membrane-anchored urokinase-type plasminogen activator receptor (uPAR) has been believed to be central to the functions of uPA, as uPA-catalyzed plasminogen activation activity appeared to be confined to cell surfaces through the binding of uPA to uPAR. However, a functional uPAR has so far only been identified in mammals. We have now cloned, recombinantly produced, and characterized two zebrafish proteases, zfuPA-a and zfuPA-b, which by several criteria are the fish orthologs of mammalian uPA. Thus, both proteases catalyze the activation of fish plasminogen efficiently and both proteases are inhibited rapidly by plasminogen activator inhibitor-1 (PAI-1). But zfuPA-a differs from mammalian uPA by lacking the exon encoding the uPAR-binding epidermal growth factor-like domain; zfuPA-b differs from mammalian uPA by lacking two cysteines of the epidermal growth factor-like domain and a uPAR-binding sequence comparable with that found in mammalian uPA. Accordingly, no zfuPA-b binding activity could be found in fish white blood cells or fish cell lines. We therefore propose that the current consensus of uPA-catalyzed plasminogen activation taking place on cell surfaces, derived from observations with mammals, is too narrow. Fish uPAs appear incapable of receptor binding in the manner known from mammals and uPA-catalyzed plasminogen activation in fish may occur mainly in solution. Studies with nonmammalian vertebrate species are needed to obtain a comprehensive understanding of the mechanism of plasminogen activation.

Bager, René; Kristensen, Thomas K

2012-01-01

177

Urokinase-like plasminogen activator as a marker of endometrial neoplasia.  

Science.gov (United States)

Urokinase-like plasminogen activator (ULPA) was determined with a radioimmunoassay in uterine aspirates from 67 patients prior to curettage. In women with normal endometrial histology the mean concentration of ULPA was 5.4 +/- 2.5 microgram per liter. Only one of this aspirates contained more than 8 microgram per liter. In cases with malignant histology ULPA was above this level in 10 of 11 with a mean concentration of 31.2 +/- 31.4 microgram per liter. In cases with glandular cystic hyperplasia and/or adenomatous hyperplasia 6 of 15 had values above 8 microgram per liter with a mean of 10.3 +/- 6.1 microgram per liter. In addition to other parameters of malignancy such as cytology, detection of ULPA in uterine fluid might prove useful in screening examinations for endometrial neoplasia. PMID:7196798

Niklasson, O; Svanberg, L; Thorell, J; Lecander, I; Astedt, B

1981-09-15

178

Ischaemia-reperfusion injury impairs tissue plasminogen activator release in man  

DEFF Research Database (Denmark)

AimsIschaemia-reperfusion (IR) injury causes endothelium-dependent vasomotor dysfunction that can be prevented by ischaemic preconditioning. The effects of IR injury and preconditioning on endothelium-dependent tissue plasminogen activator (t-PA) release, an important mediator of endogenous fibrinolysis, remain unknown.Methods and resultsIschaemia-reperfusion injury (limb occlusion at 200 mmHg for 20 min) was induced in 22 healthy subjects. In 12 subjects, IR injury was preceded by local or remote ischaemic preconditioning (three 5 min episodes of ipsilateral or contralateral limb occlusion, respectively) or sham in a randomized, cross-over trial. Forearm blood flow (FBF) and endothelial t-PA release were assessed using venous occlusion plethysmography and venous blood sampling during intra-arterial infusion of acetylcholine (5-20 µg/min) or substance P (2-8 pmol/min). Acetylcholine and substance P caused dose-dependent increases in FBF (P

Pedersen, Christian Møller; Barnes, Gareth

2011-01-01

179

Ovine rotaviruses  

Directory of Open Access Journals (Sweden)

Full Text Available Rotavirus has been recognized as a predominant cause of acute diarrhea in young animals and humans. Rotavirus has segmented genome composed of 11 segments of double stranded RNA. The virus has a triple layered protein shell consisting of a core, an inner capsid and an outer capsid. The inner capsid protein is responsible for group specificity and based on it rotaviruses are classified into seven groups. Ovine rotavirus strains have only been identified into two serogroups (A and B. The two outer capsid proteins (VP7 and VP4 are responsible for G and P typing of rotavirus, respectively. Although rotavirus has been frequently reported in many animal species, data regarding ovine rotavirus strains is very scanty and limited. Only a few ovine rotaviruses have been isolated and characterized so far. Recently, the G and P types circulating in ovines have been identified. The ovine rotavirus strain NT isolated from a diarrheic lamb in China is being considered as a promising vaccine candidate for human infants.

M.A. Bhat

2011-12-01

180

Gastric Expression of Plasminogen Activator Inhibitor (PAI)-1 Is Associated with Hyperphagia and Obesity in Mice  

Science.gov (United States)

The adipokine plasminogen activator inhibitor (PAI)-1 is increased in plasma of obese individuals and exhibits increased expression in the stomachs of individuals infected with Helicobacter. To investigate the relevance of gastric PAI-1, we used 1.1 kb of the H+/K+? subunit promoter to overexpress PAI-1 specifically in mouse gastric parietal cells (PAI-1-H/K? mice). We studied the physiological, biochemical, and behavioral characteristics of these and mice null for PAI-1 or a putative receptor, urokinase plasminogen activator receptor (uPAR). PAI-1-H/K? mice had increased plasma concentrations of PAI-1 and increased body mass, adiposity, and hyperphagia compared with wild-type mice. In the latter, food intake was inhibited by cholecystokinin (CCK)8s, but PAI-1-H/K? mice were insensitive to the satiating effects of CCK8s. PAI-1-H/K? mice also had significantly reduced expression of c-fos in the nucleus tractus solitarius in response to CCK8s and refeeding compared with wild-type mice. Exogenous PAI-1 reversed the effects of CCK8s on food intake and c-fos levels in the nucleus tractus solitarius of wild-type mice, but not uPAR-null mice. Infection of C57BL/6 mice with Helicobacter felis increased gastric abundance of PAI-1 and reduced the satiating effects of CCK8s, whereas the response to CCK8s was maintained in infected PAI-1–null mice. In cultured vagal afferent neurons, PAI-1 inhibited stimulation of neuropeptide Y type 2 receptor (Y2R) expression by CCK8s. Thus, gastric expression of PAI-1 is associated with hyperphagia, moderate obesity, and resistance to the satiating effects of CCK indicating a new role in suppressing signals from the upper gut that inhibit food intake.

Kenny, Susan; Gamble, Joanne; Lyons, Suzanne; Vlatkovic, Nikolina; Dimaline, Rod; Varro, Andrea

2013-01-01

 
 
 
 
181

RNA aptamers as conformational probes and regulatory agents for plasminogen activator inhibitor-1  

DEFF Research Database (Denmark)

The hallmark of serpins is the ability to undergo the so-called "stressed-to-relaxed" switch during which the surface-exposed reactive center loop (RCL) becomes incorporated as strand 4 in central beta-sheet A. RCL insertion drives not only the inhibitory reaction of serpins with their target serine proteases but also the conversion to the inactive latent state. RCL insertion is coupled to conformational changes in the flexible joint region flanking beta-sheet A. One interesting serpin is plasminogen activator inhibitor-1 (PAI-1), a fast and specific inhibitor of the serine proteases tissue-type and urokinase-type plasminogen activator. Via its flexible joints' region, native PAI-1 binds vitronectin and relaxed, protease-complexed PAI-1 certain endocytosis receptors. From a library of 35-nucleotides long 2'-fluoropyrimidine-containing RNA oligonucleotides, we have isolated two aptamers binding PAI-1 by the flexible joint region with low nanomolar K(D) values. One of the aptamers exhibited measurable binding to native PAI-1 only, while the other also bound relaxed PAI-1. While none of the aptamers inhibited the antiproteolytic effect of PAI-1, both aptamers inhibited vitronectin binding and the relaxed PAI-1-binding aptamer also endocytosis receptor binding. The aptamer binding exclusively to native PAI-1 increased the half-life for the latency transition to more than 6 h, manyfold more than vitronectin. Contact with Lys124 in the flexible joint region was critical for strong inhibition of the latency transition and the lack of binding to relaxed PAI-1. We conclude that aptamers yield important information about the serpin conformational switch and, because they can compete with high-affinity protein-protein interactions, may provide leads for pharmacological intervention.

Madsen, Jeppe B; Dupont, Daniel Miotto

2010-01-01

182

Solution structure of the kringle domain from urokinase-type plasminogen activator.  

Science.gov (United States)

The solution structure of the kringle domain from urokinase-type plasminogen activator (u-PA) has been determined using 1H nuclear magnetic resonance spectroscopy and dynamical simulated annealing calculations. A total of 35 structures, 20 generated using a distance geometry method prior to simulated annealing and 15 generated using initial random phi, psi values, have been calculated based on 946 experimental nuclear Overhauser effect distance constraints and 48 dihedral angle constraints. Excluding the N- and C-terminal residues (-1 to 12, 77 to 82) and a number of surface residues (M18, G19, S42, D55 to R60, G67) that are disordered or flexible, the root mean square deviation values from the mean structure are 0.49(+/- 0.14) A and 0.65(+/- 0.16) A for the backbone atoms, and 1.03(+/- 0.21) A and 1.39(+/- 0.24) A for all heavy atoms, for the two sets of structures, respectively. An extended binding site for anionic polysaccharides such as heparin has been located on a relatively flat facet of the molecule, involving three consecutive arginines, R57, R58 and R60 (there is a deletion at site 59 of the consensus sequence), which form a cationic triad facing the solvent, and two histidines, H37 and H40, at the opposite end of the molecule. Comparison between the u-PA kringle structure and the crystal and NMR solution structures of tissue-type plasminogen activator kringle 2 has shown that the two proteins have similar global folds but demonstrate a number of local differences. PMID:8107091

Li, X; Bokman, A M; Llinás, M; Smith, R A; Dobson, C M

1994-02-01

183

Ex vivo bubble production from ovine large blood vessels: Size on detachment and evidence of "active spots".  

Science.gov (United States)

Nanobubbles formed on the hydrophobic silicon wafer were shown to be the source of gas micronuclei from which bubbles evolved during decompression. Bubbles were also formed after decompression on the luminal surface of ovine blood vessels. Four ovine blood vessels: aorta, pulmonary vein, pulmonary artery, and superior vena cava, were compressed to 1013kPa for 21h. They were then decompressed, photographed at 1-s intervals, and bubble size was measured on detachment. There were certain spots at which bubbles appeared, either singly or in a cluster. Mean detachment diameter was between 0.7 and 1.0mm. The finding of active spots at which bubbles nucleate is a new, hitherto unreported observation. It is possible that these are the hydrophobic spots at which bubbles nucleate, stabilise, and later transform into the gas micronuclei that grow into bubbles. The possible neurological effects of these large arterial bubbles should be further explored. PMID:24933644

Arieli, R; Marmur, A

2014-08-15

184

The Conversion of Active to Latent Plasminogen Activator Inhibitor-1 Is an Energetically Silent Event  

Digital Repository Infrastructure Vision for European Research (DRIVER)

PAI-1 is a proteinase inhibitor, which plays a key role in the regulation of fibrinolysis. It belongs to the serpins, a family of proteins that behave either as proteinase inhibitors or proteinase substrates, both reactions involving limited proteolysis of the reactive center loop and insertion of part of this loop into ?-sheet A. Titration calorimetry shows that the inhibition of tissue-type plasminogen and pancreatic trypsin are exothermic reactions with ?H = ?20.3, and ?22.5 kcal.mol...

Boudier, Christian; Gils, Ann; Declerck, Paul J.; Bieth, Joseph G.

2005-01-01

185

The Single Substitution I259T, Conserved in the Plasminogen Activator Pla of Pandemic Yersinia pestis Branches, Enhances Fibrinolytic Activity ?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The outer membrane plasminogen activator Pla of Yersinia pestis is a central virulence factor in plague. The primary structure of the Pla ?-barrel is conserved in Y. pestis biovars Antiqua, Medievalis, and Orientalis, which are associated with pandemics of plague. The Pla molecule of the ancestral Y. pestis lineages Microtus and Angola carries the single amino acid change T259I located in surface loop 5 of the ?-barrel. Recombinant Y. pestis KIM D34 or Escherichia coli XL1 expressing Pla T2...

Haiko, Johanna; Kukkonen, Maini; Ravantti, Janne J.; Westerlund-wikstro?m, Benita; Korhonen, Timo K.

2009-01-01

186

Expression in Escherichia coli of biologically active enzyme by a DNA sequence coding for the human plasminogen activator urokinase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We have isolated clones of Escherichia coli strain K-12 that contain a hybrid pBR322 plasmid having a 4.2-kilobase insert of a DNA transcript of the mRNA of human plasminogen activator, urokinase. The bacterially produced enzyme has properties similar to those of urokinase from human fetal kidney cells. Both enzymes occur in discrete forms ranging from 32,000 to 150,000 daltons in size. They react with antibody to purified urokinase from human kidney cells, bind to a benzamidine-Sepharose col...

Ratzkin, B.; Lee, S. G.; Schrenk, W. J.; Roychoudhury, R.; Chen, M.; Hamilton, T. A.; Hung, P. P.

1981-01-01

187

Candesartan reduces the hemorrhage associated with delayed tissue plasminogen activator treatment in rat embolic stroke.  

Science.gov (United States)

We have previously reported that angiotensin receptor blockade reduces reperfusion hemorrhage in a suture occlusion model of stroke, despite increasing matrix metalloproteinase (MMP-9) activity. We hypothesized that candesartan will also decrease hemorrhage associated with delayed (6 h) tissue plasminogen activator (tPA) administration after embolic stroke, widening the therapeutic time window of tPA. Adult male Wistar rats were subjected to embolic middle cerebral artery occlusion (eMCAO) and treated with either candesartan (1 mg/kg) alone early at 3 h, delayed tPA (10 mg/kg) alone at 6 h, the combination of candesartan and tPA, or vehicle control. Rats were sacrificed at 24 and 48 h post-eMCAO and brains perfused for evaluation of neurological deficits, cerebral hemorrhage in terms of hemoglobin content, occurrence rate of hemorrhage, infarct size, tissue MMP activity and protein expression. The combination therapy of candesartan and tPA after eMCAO reduced the brain hemorrhage, and improved neurological outcome compared with rats treated with tPA alone. Further, candesartan in combination with tPA increased activity of MMP-9 but decreased MMP-3, nuclear factor kappa-B and tumor necrosis factor-? expression and enhanced activation of endothelial nitric oxide synthase. An activation of MMP-9 alone is insufficient to cause increased hemorrhage in embolic stroke. Combination therapy with acute candesartan plus tPA may be beneficial in ameliorating tPA-induced hemorrhage after embolic stroke. PMID:24194350

Ishrat, Tauheed; Pillai, Bindu; Ergul, Adviye; Hafez, Sherif; Fagan, Susan C

2013-12-01

188

Plasminogen acquisition and activation at the surface of leptospira species lead to fibronectin degradation.  

Science.gov (United States)

Pathogenic Leptospira species are the etiological agents of leptospirosis, a widespread disease of human and veterinary concern. In this study, we report that Leptospira species are capable of binding plasminogen (PLG) in vitro. The binding to the leptospiral surface was demonstrated by indirect immunofluorescence confocal microscopy with living bacteria. The PLG binding to the bacteria seems to occur via lysine residues because the ligation is inhibited by addition of the lysine analog 6-aminocaproic acid. Exogenously provided urokinase-type PLG activator (uPA) converts surface-bound PLG into enzymatically active plasmin, as evaluated by the reaction with the chromogenic plasmin substrate d-Val-Leu-Lys 4-nitroanilide dihydrochloridein. The PLG activation system on the surface of Leptospira is PLG dose dependent and does not cause injury to the organism, as cellular growth in culture was not impaired. The generation of active plasmin within Leptospira was observed with several nonvirulent high-passage strains and with the nonpathogenic saprophytic organism Leptospira biflexa. Statistically significant higher activation of plasmin was detected with a low-passage infectious strain of Leptospira. Plasmin-coated virulent Leptospira interrogans bacteria were capable of degrading purified extracellular matrix fibronectin. The breakdown of fibronectin was not observed with untreated bacteria. Our data provide for the first time in vitro evidence for the generation of active plasmin on the surface of Leptospira, a step that may contribute to leptospiral invasiveness. PMID:19581392

Vieira, Monica L; Vasconcellos, Silvio A; Gonçales, Amane P; de Morais, Zenaide M; Nascimento, Ana L T O

2009-09-01

189

Plasminogen Acquisition and Activation at the Surface of Leptospira Species Lead to Fibronectin Degradation ?  

Science.gov (United States)

Pathogenic Leptospira species are the etiological agents of leptospirosis, a widespread disease of human and veterinary concern. In this study, we report that Leptospira species are capable of binding plasminogen (PLG) in vitro. The binding to the leptospiral surface was demonstrated by indirect immunofluorescence confocal microscopy with living bacteria. The PLG binding to the bacteria seems to occur via lysine residues because the ligation is inhibited by addition of the lysine analog 6-aminocaproic acid. Exogenously provided urokinase-type PLG activator (uPA) converts surface-bound PLG into enzymatically active plasmin, as evaluated by the reaction with the chromogenic plasmin substrate d-Val-Leu-Lys 4-nitroanilide dihydrochloridein. The PLG activation system on the surface of Leptospira is PLG dose dependent and does not cause injury to the organism, as cellular growth in culture was not impaired. The generation of active plasmin within Leptospira was observed with several nonvirulent high-passage strains and with the nonpathogenic saprophytic organism Leptospira biflexa. Statistically significant higher activation of plasmin was detected with a low-passage infectious strain of Leptospira. Plasmin-coated virulent Leptospira interrogans bacteria were capable of degrading purified extracellular matrix fibronectin. The breakdown of fibronectin was not observed with untreated bacteria. Our data provide for the first time in vitro evidence for the generation of active plasmin on the surface of Leptospira, a step that may contribute to leptospiral invasiveness.

Vieira, Monica L.; Vasconcellos, Silvio A.; Goncales, Amane P.; de Morais, Zenaide M.; Nascimento, Ana L. T. O.

2009-01-01

190

Species Specificity of Plasminogen Activation and Acquisition of Surface-Associated Proteolytic Activity by Group C Streptococci Grown in Plasma  

Science.gov (United States)

Our laboratory previously demonstrated that group C streptococcal isolates from humans and horses secrete streptokinases that preferentially activate plasminogens reflecting the origin of the isolates. To analyze the significance of these findings, series of streptokinase-producing Streptococcus equisimilis isolates recovered from humans and horses were examined. Southern blot analysis revealed that chromosomal DNA of the streptococcal isolates from humans reacted exclusively with a skchu probe and that chromosomal DNA of streptococcal isolates from horses reacted preferentially with an skceq probe in a distinct pattern. The streptococcal isolates were examined for the ability to acquire surface-bound plasmin-like activity when grown in the presence of human or equine plasma. Each of eight isolates from humans acquired significant enzymatic activity only when grown in the presence of human plasma, while each of eight isolates from horses acquired activity only when grown in the presence of equine plasma. Analysis of bacterial and host protein requirements indicated critical roles for streptokinase, activatable plasminogen, and fibrinogen. These requirements may explain why certain streptococcal isolates cause disease only in a limited number of mammalian hosts.

Schroeder, Brett; Boyle, Michael D. P.; Sheerin, Barbara R.; Asbury, Atwood C.; Lottenberg, Richard

1999-01-01

191

Critical role of type 1 plasminogen activator inhibitor (PAI-1) in early host defense against Nontypeable Haemophilus influenzae (NTHi) infection  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Respiratory systems are constantly being challenged by pathogens. Lung epithelial cells serve as a first line of defense against microbial pathogens by detecting pathogen-associated molecular patterns (PAMPs) and activating downstream signaling pathways, leading to a plethora of biological responses required for shaping both the innate and adaptive arms of the immune response. Acute-phase proteins (APPs), such as type 1 plasminogen activator inhibitor (PAI-1), play important roles in immune/i...

Lim, Jae Hyang; Woo, Chang-hoon; Li, Jian-dong

2011-01-01

192

Polyoma Middle T-induced Vascular Tumor Formation: The Role of the Plasminogen Activator/Plasmin System  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The middle T antigen of murine Polyomavirus (PymT) rapidly transforms endothelial cells, leading to the formation of vascular tumors in newborn mice. Transformed endothelial (End.) cell lines established from such tumors exhibit altered proteolytic activity as a result of increased expression of urokinase-type plasminogen activator (uPA) and are capable of inducing vascular tumors efficiently when injected into adult mice. In this study we have used mice lacking components of the PA/pla...

Sabapathy, Kanaga T.; Pepper, Michael S.; Kiefer, Friedemann; Mo?hle-steinlein, Uta; Tacchini-cottier, Fabienne; Fetka, Ingrid; Breier, Georg; Risau, Werner; Carmeliet, Peter; Montesano, Roberto; Wagner, Erwin F.

1997-01-01

193

A novel inhibitor of plasminogen activator inhibitor-1 provides antithrombotic benefits devoid of bleeding effect in nonhuman primates  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Inhibition of plasminogen activator inhibitor (PAI)-1 is useful to treat several disorders including thrombosis. An inhibitor of PAI-1 (TM5275) was newly identified by an extensive study of structure-activity relationship based on a lead compound (TM5007) which was obtained through virtual screening by docking simulations. Its antithrombotic efficacy and adverse effects were tested in vivo in rats and nonhuman primates (cynomolgus monkey). TM5275, administered orally in rats (1 to 10 mg/kg), ...

Izuhara, Yuko; Yamaoka, Nagahisa; Kodama, Hidehiko; Dan, Takashi; Takizawa, Shunya; Hirayama, Noriaki; Meguro, Kanji; Ypersele Strihou, Charles; Miyata, Toshio

2010-01-01

194

Plasminogen activator inhibitor 1 is an intracellular inhibitor of furin proprotein convertase.  

Science.gov (United States)

Proprotein convertases (PCs) are a family of serine proteases that are involved in the post-translational processing and activation of a wide range of regulatory proteins. The upstream role of PCs in the control of many physiological and pathological processes generates a growing interest in understanding their regulation. Here, we demonstrate that the serine protease inhibitor plasminogen activator inhibitor 1 (PAI-1) forms an SDS-stable complex with the PC furin, which leads to the inhibition of the intra-Golgi activity of furin. It is known that elevated PAI-1 plasma levels are correlated with the occurrence of the metabolic syndrome and type 2 diabetes, and we show that PAI-1 reduces the furin-dependent maturation and activity of the insulin receptor and ADAM17: two proteins involved in the onset of these metabolic disorders. In addition to demonstrating that PAI-1 is an intracellular inhibitor of furin, this study also provides arguments in favor of an active role for PAI-1 in the development of metabolic disorders. PMID:21406565

Bernot, Denis; Stalin, Jimmy; Stocker, Pierre; Bonardo, Bernadette; Scroyen, Ilse; Alessi, Marie-Christine; Peiretti, Franck

2011-04-15

195

Modulation of staphylokinase-dependent plasminogen activation by mono- and divalent ions  

Directory of Open Access Journals (Sweden)

Full Text Available The effect of several ions (Cl-, Na+, K+, Ca2+ on the rate of plasminogen (Pg activation by recombinant staphylokinase (rSTA is reported. Both monovalent and divalent ions affect the rate at which Pg is activated by rSTA, in a concentration-dependent manner (range 0-100 mM. In almost all cases, a decrease of the initial velocity of activation was observed. Cl- showed the most striking inhibitory effect at low concentrations (64% at 10 mM. However, in the presence of a fibrin surface, this inhibition was attenuated to 38%. Surprisingly, 10 mM Ca2+ enhanced the Pg activation rate 21% when a polymerized fibrin matrix was present. These data support the idea that ions can modulate the rate of Pg activation through a mechanism that may be associated with changes in the molecular conformation of the zymogen. This effect is strongly dependent on the presence of a fibrin clot.

Yarzábal A.

1999-01-01

196

A novel plasminogen activator from Agkistrodon blomhoffii Ussurensis venom (ABUSV-PA): Purification and characterization  

International Nuclear Information System (INIS)

A plasminogen activator with arginine ester hydrolysis activity (ABUSV-PA) has been identified and purified to homogeneity from Chinese Agkistrodon blomhoffii Ussurensis snake venom. ABUSV-PA, a monomeric protein with molecular mass of 27815.2 Da, was purified 180-fold with 0.02% recovery for protein and 3.6% recovery for esterase activity. ABUSV-PA reacts optimally with its substrate N ?-tosyl-L-arginine-methyl ester (TAME) at ?pH 7.5 and at 51 oC. Measurement from inductively coupled plasma-atomic emission spectroscopy (ICP-AES) reveals that ABUSV-PA is a Zn2+-containing protein with a stoichiometry of 1:1 [Zn2+]:[ABUSV-PA]. Analyses of esterase hydrolysis and UV absorption and CD spectra indicate that Zn2+ plays an important role in maintaining the structural integrity rather than the esterase activity of ABUSV-PA. Divalent metal ions, including Ca2+, Mg2+, Cu2+, Ni2+, Mn2+, and Co2+, increase the TAME hydrolysis activity of ABUSV-PA. A red-shift of the emission wavelengths of the synchronous fluorescence of ABUSV-PA, compared to those of free Tyr and Trp, indicates a conformation where the Tyr and Trp residues are in exposed hydrophilic environments. The presence of zinc increases the hydrophobicity of the conformational environments surrounding the Trp residues of ABUSV-PA and affects the secondary structure of ABUSV-PA, as proved by UV absorption and CD spectroscopy

2006-10-06

197

Functional role of proteolytic cleavage at arginine-275 of human tissue plasminogen activator as assessed by site-directed mutagenesis  

International Nuclear Information System (INIS)

Activation of the zymogen form of a serine protease is associated with a conformational change that follows proteolysis at a specific site. Tissue-type plasminogen activator (t-PA) is homologous to mammalian serine proteases and contains an apparent activation cleavage site at arginine-275. To clarify the functional consequences of cleavage at arginine-275 of t-PA, site-specific mutagenesis was performed to convert arginine-275 to a glutamic acid. The mutant enzyme (designated Arg-275 ? Glu t-PA) could be converted to the two-chain form by Staphylococcus aureus V8 protease but not by plasmin. The one-chain form was 8 times less active against the tripeptide substrate H-D-isoleucyl-L-prolyl-L-arginine-rho-nitroanilide (S-2288), and the ability of the enzyme to activate plasminogen in the absence of fibrinogen was reduced 20-50 times compared to the two-chain form. In contrast, one-chain Arg-275 ? Glu t-PA has equal activity to the two-chain form when assayed in the presence of physiological levels of fibrinogen and plasminogen. Fibrin bound significantly more of the one-chain form of t-PA than the two-chain form for both the wild-type and mutated enzymes. One- and two-chain forms of the wild-type and mutated plasminogen activators slowly formed complexes with plasma protease inhibitors, although the one-chain forms showed decreased complex formation with ?_2-macroglobulin. The one-chain form of t-PA therefore is fully functional under physiologic conditions and has a increased fibrin binding compared to the two-chain form

1987-01-27

198

Partial purification and characterization of proteins with growth promoting activities from ovine mammary gland secretions.  

Science.gov (United States)

Developmental regulation of growth promoting activities in mammary secretions of pregnant Awassi ewes was defined, and growth factors contained in these secretions were partially purified and characterised. Mammary secretions from pregnant ewes enhanced fibroblast cell (AKR-2B) and mammary cell (CID-9 cell strain) proliferation to levels comparable to that induced by 10% Foetal calf serum. Major milk proteins in mammary secretions collected from pregnant ewes one month prior to lambing up to one week after lambing, were resolved by SDS-PAGE, while gelatinases were resolved by zymography. Gelatinase activity was noted prior to P134 and decreased thereafter to reach a minimum during lactation. This decrease was concomitant with the onset of casein production. It is during this critical developmental period that highest growth promoting activity in mammary secretions was detected. Secretions with highest growth promoting activity were fractionated by ion exchange and gel filtration chromatography. Two heat-resistant, trypsin/chymotrypsin sensitive, growth-promoting activities were characterised. The first, designated ovine mammary derived growth factor-1 (oMDGF-1), had around a 30 kDa molecular weight and eluted at 0.65 M NaCl gradient on cation ion exchange chromatography. The second, oMDGF-2, eluted under gel filtration conditions at a molecular weight of 50 kDa and 150 kDa. oMDGF-1 induced changes in Connexin 43, but not in beta-casein mRNA expression by CID-9 mammary cells. In conclusion, growth factor activities in ewe mammary secretions peak during gestation at a period that overlaps maximal gelatinase expression and precedes milk protein synthesis. The factors modulate mammary cell function and may play a role in mammary gland development. PMID:11707361

Talhouk, R S; Maa'ni, F A; Kalaa'ni, N; Zoubian, G S; Simaa'n, C J; Abi-Sai'd, M; Hamadeh, S; Barbour, E; El-Sabban, M E

2001-10-01

199

Biological and binding activities of ovine and porcine prolactins in porcine mammary tissue  

International Nuclear Information System (INIS)

The concentration of prolactin receptors may play a critical role in regulating growth and development of the mammary gland during gestation and tumor development; however, the discrepancy between specific binding of ovine prolactin (oPRL) and porcine prolactin (pPRL) in porcine mammary tissue was disturbing. It was possible that 125I-oPRL may be an unsuitable ligand for the procine prolactin receptor. The validate the use of oPRL in binding assays, the biological and binding activities of oPRL and pPRL were compared. A lactogenic bioassay of pPRL was developed using porcine mammary explants cultured in Medium 199 containing insulin, cortisol, and pPRL. The potencies of oPRL and pPRL were compared using this bioassay. Oxidation of glucose and incorporation of glucose into lipids were similarly enhanced by physiological concentrations of both oPRL and pPRL. However, specific binding of 125I-oPRL was 20%, while less than 1% of 125I-pPRL was bound. 125I-oPRL bound to high affinity sites

1987-01-01

200

Differential effects of tissue plasminogen activator and streptokinase on infarct size and on rate of enzyme release: influence of early infarct related artery patency  

Digital Repository Infrastructure Vision for European Research (DRIVER)

BACKGROUND: The recent international GUSTO trial of 41,021 patients with acute myocardial infarction demonstrated improved 90-min infarct related artery patency as well as reduced mortality in patients treated with an accelerated regimen of tissue plasminogen activator, compared to patients treated with streptokinase. A regimen combining tissue plasminogen activator and streptokinase yielded intermediate results. The present study investigated the effects of treatment on infarct size and enzy...

Baardman, T.; Hermens, W. T.; Molhoek, G. P.; Grollier, G.; Pfisterer, M. E.; Simoons, M. L.; Lenderink, T.

1996-01-01

 
 
 
 
201

Plasminogen activator inhibitor type 1 derived peptide, EEIIMD, diminishes cortical infarct but fails to improve neurological function in aged rats following middle cerebral artery occlusion  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Age is a primary risk factor in stroke that is often overlooked in animal studies. We contend that using aged animals yields insight into aspects of stroke injury and recovery that are masked, or not elicited, in younger animals. In this study, we examined effects of co-administration of a plasminogen activator inhibitor type 1 derived peptide, EEIIMD, with tissue plasminogen activator (tPA) on infarct volume and functional outcome in aged rats following a transient middle cerebral artery occ...

Tan, Zhenjun; Li, Xinlan; Kelly, Kimberly A.; Rosen, Charles L.; Huber, Jason D.

2009-01-01

202

Engineering streptokinase for generation of active site-labeled plasminogen analogs.  

Science.gov (United States)

We previously demonstrated that streptokinase (SK) can be used to generate active site-labeled fluorescent analogs of plasminogen (Pg) by virtue of its nonproteolytic activation of the zymogen. The method is versatile and allows stoichiometric and active site-specific incorporation of any one of many molecular probes. The limitation of the labeling approach is that it is both time-consuming and low yield. Here we demonstrate an improved method for the preparation of labeled Pg analogs by the use of an engineered SK mutant fusion protein with both COOH- and NH(2)-terminal His(6) tags. The NH(2)-terminal tag is followed by a tobacco etch virus proteinase cleavage site to ensure that the SK Ile(1) residue, essential for conformational activation of Pg, is preserved. The SK COOH-terminal Lys(414) residue and residues Arg253-Leu260 in the SK ?-domain were deleted to prevent cleavage by plasmin (Pm) and to disable Pg substrate binding to the SK·Pg(?)/Pm catalytic complexes, respectively. Near elimination of Pm generation with the SK?(R253-L260)?K414-His(6) mutant increased the yield of labeled Pg 2.6-fold and reduced the time required more than 2-fold. The versatility of the labeling method was extended to the application of Pg labeled with a near-infrared probe to quantitate Pg receptors on immune cells by flow cytometry. PMID:21570944

Laha, Malabika; Panizzi, Peter; Nahrendorf, Matthias; Bock, Paul E

2011-08-15

203

Engineering streptokinase for generation of active site-labeled plasminogen analogs*  

Science.gov (United States)

We previously demonstrated that streptokinase (SK) can be used to generate active site-labeled fluorescent analogs of plasminogen (Pg) by virtue of its non-proteolytic activation of the zymogen. The method is versatile and allows for stoichiometric and active site-specific incorporation of any one of many molecular probes. The limitation of the labeling approach is that it is both time-consuming and low yield. Here we demonstrate an improved method for the preparation of labeled Pg analogs by the use of an engineered SK mutant fusion protein with both COOH- and NH2-terminal His6-tags. The NH2-terminal tag is followed by a tobacco etch virus proteinase cleavage site to ensure that the SK Ile1 residue, essential for conformational activation of Pg, is preserved. The SK COOH-terminal Lys414 residue and residues Arg253-Leu260 in the SK ?-domain were deleted to prevent cleavage by plasmin (Pm), and to disable Pg substrate binding to the SK·Pg*/Pm catalytic complexes, respectively. Near-elimination of Pm generation with the SK?(R253-L260)?K414-His6 mutant increased the yield of labeled Pg 2.6-fold and reduced the time required >2-fold. The versatility of the labeling method was extended to the application of Pg labeled with a near-infrared probe to quantitate Pg receptors on immune cells by flow cytometry.

Laha, Malabika; Panizzi, Peter; Nahrendorf, Matthias; Bock, Paul E.

2011-01-01

204

Targeted delivery of tissue plasminogen activator by binding to silica-coated magnetic nanoparticle  

Directory of Open Access Journals (Sweden)

Full Text Available Jyh-Ping Chen,1 Pei-Ching Yang,1 Yunn-Hwa Ma,2 Su-Ju Tu,3 Yu-Jen Lu1,41Department of Chemical and Materials Engineering, 2Department of Physiology and Pharmacology, 3Department of Medical Imaging and Radiological Sciences, Chang Gung University, Kwei-San, Taoyuan, Taiwan, Republic of China; 4Department of Neurosurgery, Chang Gung Memorial Hospital, Kwei-San, Taoyuan, Taiwan, Republic of ChinaBackground and methods: Silica-coated magnetic nanoparticle (SiO2-MNP prepared by the sol-gel method was studied as a nanocarrier for targeted delivery of tissue plasminogen activator (tPA. The nanocarrier consists of a superparamagnetic iron oxide core and an SiO2 shell and is characterized by transmission electron microscopy, Fourier transform infrared spectroscopy, X-ray diffraction, superconducting quantum interference device, and thermogravimetric analysis. An amine-terminated surface silanizing agent (3-aminopropyltrimethoxysilane was used to functionalize the SiO2 surface, which provides abundant —NH2 functional groups for conjugating with tPA.Results: The optimum drug loading is reached when 0.5 mg/mL tPA is conjugated with 5 mg SiO2-MNP where 94% tPA is attached to the carrier with 86% retention of amidolytic activity and full retention of fibrinolytic activity. In vitro biocompatibility determined by lactate dehydrogenase release and cell proliferation indicated that SiO2-MNP does not elicit cytotoxicity. Hematological analysis of blood samples withdrawn from mice after venous administration indicates that tPA-conjugated SiO2-MNP (SiO2-MNP-tPA did not alter blood component concentrations. After conjugating to SiO2-MNP, tPA showed enhanced storage stability in buffer and operation stability in whole blood up to 9.5 and 2.8-fold, respectively. Effective thrombolysis with SiO2-MNP-tPA under magnetic guidance is demonstrated in an ex vivo thrombolysis model where 34% and 40% reductions in blood clot lysis time were observed compared with runs without magnetic targeting and with free tPA, respectively, using the same drug dosage. Enhanced penetration of SiO2-MNP-tPA into blood clots under magnetic guidance was confirmed from microcomputed tomography analysis.Conclusion: Biocompatible SiO2-MNP developed in this study will be useful as a magnetic targeting drug carrier to improve clinical thrombolytic therapy.Keywords: magnetic nanoparticles, drug delivery, thrombolysis, tissue plasminogen activator, silica

Chen JP

2012-09-01

205

The effect of 40 kHz ultrasound on tissue plasminogen activator-induced clot lysis in three in vitro models  

Science.gov (United States)

In previous work from the same laboratory, high-frequency ultrasound (US) (3 MHz) was shown to promote in vitro fibrinolysis through enhanced supply of plasminogen to the clot surface. The application of high-frequency US is limited in vivo due to tissue heating. Low-frequency US, however, has less tissue heating and improved penetration. Internal plasma clot lysis and external lysis with compacted and non-compacted plasma clots were used to determine the magnitude of the effect of low-frequency US (40 kHz; 0.5 W/cm2) on tissue plasminogen activator-induced lysis and to elucidate the mechanisms behind the effect. Ultrasound enhanced lysis in all three models, with the largest effects (4-fold) in the external lysis model with compacted plasminogen-poor clots. The acceleration effect of ultrasound in this model decreased with increasing t-PA-and decreasing plasminogen concentrations. Ultrasound had a much smaller effect in this model when compacted plasminogen-rich clots were used. In the external lysis, non-compacted clot model, ultrasound resulted in consistently higher lysis rates. The acceleration effect of lysis, increased slightly (1.3 to 1.8-fold) with increasing t-PA-and decreasing plasminogen concentrations. Plasminogen supply to the clot surface was again shown to be an important contributor to ultrasound-enhanced lysis. [M. Pieters, R. T. Hekkenberg, M. Barrett-Bergshoeff, and D. C. Rijken. Ultrasound in Med & Biol 30, 1545-1552 (2004).

Pieters, Marlien; Hekkenberg, Rob T.; Barrett-Bergshoeff, Marrie; Rijken, Dingeman C.

2005-04-01

206

Cpa, the Outer Membrane Protease of Cronobacter sakazakii, Activates Plasminogen and Mediates Resistance to Serum Bactericidal Activity?  

Science.gov (United States)

Cronobacter spp. are emerging neonatal pathogens in humans, associated with outbreaks of meningitis and sepsis. To cause disease, they must survive in blood and invade the central nervous system by penetrating the blood-brain barrier. C. sakazakii BAA-894 possesses an ?131-kb plasmid (pESA3) that encodes an outer membrane protease (Cpa) that has significant identity to proteins that belong to the Pla subfamily of omptins. Members of this subfamily of proteins degrade a number of serum proteins, including circulating complement, providing protection from the complement-dependent serum killing. Moreover, proteins of the Pla subfamily can cause uncontrolled plasmin activity by converting plasminogen to plasmin and inactivating the plasmin inhibitor ?2-antiplasmin (?2-AP). These reactions enhance the spread and invasion of bacteria in the host. In this study, we found that an isogenic cpa mutant showed reduced resistance to serum in comparison to its parent C. sakazakii BAA-894 strain. Overexpression of Cpa in C. sakazakii or Escherichia coli DH5? showed that Cpa proteolytically cleaved complement components C3, C3a, and C4b. Furthermore, a strain of C. sakazakii overexpressing Cpa caused a rapid activation of plasminogen and inactivation of ?2-AP. These results strongly suggest that Cpa may be an important virulence factor involved in serum resistance, as well as in the spread and invasion of C. sakazakii.

Franco, A. A.; Kothary, M. H.; Gopinath, G.; Jarvis, K. G.; Grim, C. J.; Hu, L.; Datta, A. R.; McCardell, B. A.; Tall, B. D.

2011-01-01

207

Cpa, the outer membrane protease of Cronobacter sakazakii, activates plasminogen and mediates resistance to serum bactericidal activity.  

Science.gov (United States)

Cronobacter spp. are emerging neonatal pathogens in humans, associated with outbreaks of meningitis and sepsis. To cause disease, they must survive in blood and invade the central nervous system by penetrating the blood-brain barrier. C. sakazakii BAA-894 possesses an ~131-kb plasmid (pESA3) that encodes an outer membrane protease (Cpa) that has significant identity to proteins that belong to the Pla subfamily of omptins. Members of this subfamily of proteins degrade a number of serum proteins, including circulating complement, providing protection from the complement-dependent serum killing. Moreover, proteins of the Pla subfamily can cause uncontrolled plasmin activity by converting plasminogen to plasmin and inactivating the plasmin inhibitor ?2-antiplasmin (?2-AP). These reactions enhance the spread and invasion of bacteria in the host. In this study, we found that an isogenic cpa mutant showed reduced resistance to serum in comparison to its parent C. sakazakii BAA-894 strain. Overexpression of Cpa in C. sakazakii or Escherichia coli DH5? showed that Cpa proteolytically cleaved complement components C3, C3a, and C4b. Furthermore, a strain of C. sakazakii overexpressing Cpa caused a rapid activation of plasminogen and inactivation of ?2-AP. These results strongly suggest that Cpa may be an important virulence factor involved in serum resistance, as well as in the spread and invasion of C. sakazakii. PMID:21245266

Franco, A A; Kothary, M H; Gopinath, G; Jarvis, K G; Grim, C J; Hu, L; Datta, A R; McCardell, B A; Tall, B D

2011-04-01

208

Elevated levels of plasminogen activators in the pathogenesis of delayed radiation damage in rat cervical spinal cord in vivo  

International Nuclear Information System (INIS)

The pathophysiology of the cellular basis of radiation-induced demyelination and white-matter necrosis of the central nervous system (CNS) is poorly understood. Preliminary data suggest that tissue damage is partly mediated through changes in the proteolytic enzymes. In this study, we irradiated rat cervical spinal cords with single doses of 24 Gy of 18 MV photons or 20 MeV electrons and measured the levels of plasminogen activators at days 2, 7, 30, 60, 90, 120, 130 and 145 after irradiation, using appropriate controls at each time. Fibrin zymography revealed fibrinolytic bands representing molecular weights of 68,000 and 48,000 in controls and irradiated samples; these bands increased significantly at days 120, 130 and 145 after irradiation. Inhibition of these enzymatic bands with specific antibodies against tissue-type plasminogen activator (tPA) and amiloride, an inhibitor for urokinase plasminogen activator (uPA), confirmed that these bands were tPA and uPA. Enzymatic levels quantified by densitometry showed a twofold elevation in the levels of tPA and more than a tenfold increase in uPA after 120 days' irradiation. Activity of uPA was increased threefold by day 2 and increased steadily with time compared to nonirradiated control samples. Enzyme-linked immunosorbent assay (ELISA) also showed a threefold increase in the tPA content in the extracts of irradiated rat cervical spinal cords at days 120, 130 and 145. This study adds additional information to the proposed role of plasminogen activators in the pathogenic pathways of radiation damage in the CNS. 38 refs., 6 figs

1994-06-01

209

Bioconjugation of recombinant tissue plasminogen activator to magnetic nanocarriers for targeted thrombolysis  

Directory of Open Access Journals (Sweden)

Full Text Available Hung-Wei Yang,1,* Mu-Yi Hua,1,* Kun-Ju Lin,2,* Shiaw-Pyng Wey,3 Rung-Ywan Tsai,4 Siao-Yun Wu,5 Yi-Ching Lu,5 Hao-Li Liu,6 Tony Wu,7 Yunn-Hwa Ma5 1Chang Gung Molecular Medicine Research Center, Department of Chemical and Materials Engineering, 2Molecular Imaging Center, Department of Nuclear Medicine, Chang Gung Memorial Hospital, Kuei-Shan, Tao-Yuan, Taiwan, Republic of China; 3Department of Medical Imaging and Radiological Sciences, 4Electronics and Optoelectronics Research Laboratories, Industrial Technology Research Institute, Hsin-chu, Taiwan, Republic of China; 5Department of Physiology and Pharmacology and Healthy Aging Research Center, 6Department of Electrical Engineering, Chang Gung University, Kuei-Shan, Tao-Yuan, Taiwan, Republic of China; 7Department of Neurology, Chang Gung University College of Medicine and Memorial Hospital, Tao-Yuan, Taiwan, Republic of China*These authors contributed equally to this workAbstract: Low-toxicity magnetic nanocarriers (MNCs composed of a shell of poly [aniline-co-N-(1-one-butyric acid aniline] over a Fe3O4 magnetic nanoparticle core were developed to carry recombinant tissue plasminogen activator (rtPA in MNC-rtPA for targeted thrombolysis. With an average diameter of 14.8 nm, the MNCs exerted superparamagnetic properties. Up to 276 µg of active rtPA was immobilized per mg of MNCs, and the stability of the immobilized rtPA was greatly improved during storage at 4°C and 25°C. In vitro thrombolysis testing with a tubing system demonstrated that magnet-guided MNC-rtPA showed significantly improved thrombolysis compared with free rtPA and reduced the clot lysis time from 39.2 ± 3.2 minutes to 10.8 ± 4.2 minutes. In addition, magnet-guided MNC-rtPA at 20% of the regular rtPA dose restored blood flow within 15–25 minutes of treatment in a rat embolism model without triggering hematological toxicity. In conclusion, this improved system is based on magnetic targeting accelerated thrombolysis and is potentially amenable to therapeutic applications in thromboembolic diseases.Keywords: thrombolysis, recombinant tissue plasminogen activator, magnetic nanocarriers, magnetic targeting, targeting therapy

Yang HW

2012-10-01

210

Expression of urokinase-type plasminogen activator/urokinase-type plasminogen activator receptor and maspin in oral squamous cell carcinoma: Association with mode of invasion and clinicopathological factors.  

Science.gov (United States)

It is well documented that the binding of urokinase-type plasminogen activator (uPA) to its receptor (uPAR), which has been implicated in cancer invasion and metastasis, is regulated by several inhibitors such as maspin. In this study, we investigated the interrelationship between clinicopathologic findings and expression of uPA, uPAR and maspin in oral squamous cell carcinoma (OSCC) to elucidate the participation of maspin in the uPA/uPAR system in the malignant behavior of OSCC. Using immunohistochemical techniques to examine the expression levels of uPA, uPAR and maspin in 54 cases of OSCC, we also compared the clinicopathologic features of OSCC with the expression levels of each. Moreover, we examined the expression of uPA, uPAR and maspin in six cell lines derived from OSCC using reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting. uPA and uPAR showed a positive correlation with the mode of cancer invasion; conversely maspin showed a negative correlation with the mode of invasion. Multivariate analysis revealed that only two factors (N-category and uPA+/uPAR+/maspin- expression pattern) were significant and independent variables with relative risks of 3.84 and 2.52, respectively. In particular, tumors exhibiting an expression pattern of uPA+/uPAR+/maspin- were highly malignant and were associated with the worst survival rate (5-year overall survival rate, 29.4%), while tumors with an expression pattern, uPA-/uPAR-/?aspin+, showed the most favorable survival rate (5-year overall survival rate, 77.8%). In vitro, lower expression of maspin was also noted in the cell lines derived from grade 4D OSCC, which exhibited a stronger invasive potential than the cells lines derived from the other grades of OSCC, while uPA and uPAR demonstrated an expression trend opposite to maspin. These results indicate that uPA, uPAR and maspin expression patterns may be useful markers for evaluating the clinical course or prognosis of OSCC patients. PMID:21833477

Yoshizawa, Kunio; Nozaki, Shinichi; Kitahara, Hiroko; Kato, Koroku; Noguchi, Natsuyo; Kawashiri, Shuichi; Yamamoto, Etsuhide

2011-12-01

211

Diverse inhibition of plasminogen activator inhibitor type 1 by theaflavins of black tea.  

Science.gov (United States)

Fruits, vegetables, spices and a variety of teas are suggested for the prevention of many diseases. They encompass active, non-nutritional ingredients called nutraceuticals which are defined as food products that provide health benefits. Many nutraceuticals have been tested to identify inhibitors of plasminogen activator inhibitor (PAI-1). PAI-1 is the major and fast acting physiological inhibitor of fibrinolysis. However, preclinical studies of PAI-1 inhibitors have revealed an additional role of PAI-1 in the pathogenesis of vascular remodeling, renal injury, diabetes, obesity, Alzheimer's disease and cancer. Thus PAI-1 is a potential therapeutic target in some of these diseases. Our previous study revealed that a black tea extract (containing mostly theaflavins) inhibits PAI-1. In this study we report results for four pure (>98%) theaflavins. Inactivation of PAI-1 was tested by clot formation and by its lysis using thromboelastometry and measurements of human plasma turbidity. Among four tested theaflavins, theaflavin-3'-gallate was the most potent in PAI-1 inhibition trailed by theaflavin-3,3'-digallate, while the other two i.e., theaflavin and theaflavin-3-gallate did not show inhibitory activity. PMID:21308350

Jankun, Jerzy; Skotnicka, Magdalena; ?ysiak-Szyd?owska, Wies?awa; Al-Senaidy, Abdulrahman; Skrzypczak-Jankun, Ewa

2011-04-01

212

Terminalia catappa attenuates urokinase-type plasminogen activator expression through Erk pathways in Hepatocellular carcinoma  

Science.gov (United States)

Background The survival rate of malignant tumors, and especially hepatocellular carcinoma (HCC), has not improved primarily because of uncontrolled metastasis. In our previous studies, we have reported that Terminalia catappa leaf extract (TCE) exerts antimetastasis effects on HCC cells. However, the molecular mechanisms of urokinase-type plasminogen activator (u-PA) in HCC metastasis have not been thoroughly investigated, and remain poorly understood. Methods The activities and protein levels of u-PA were determined by casein zymography and western blotting. Transcriptional levels of u-PA were detected by real-time PCR and promoter assays. Results We found that treatment of Huh7 cells with TCE significantly reduced the activities, protein levels and mRNA levels of u-PA. A chromatin immunoprecipitation (ChIP) assay showed that TCE inhibited the transcription protein of nuclear factors SP-1 and NF-?B. TCE also did inhibit the effects of u-PA by reducing the phosphorylation of ERK1/2 pathway. Conclusions These results show that u-PA expression may be a potent therapeutic target in the TCE-mediated suppression of HCC metastasis.

2014-01-01

213

Tissue plasminogen activator alters intracellular sequestration of zinc through interaction with the transporter ZIP4.  

Science.gov (United States)

Glutamatergic neurons contain free zinc packaged into neurotransmitter-loaded synaptic vesicles. Upon neuronal activation, the vesicular contents are released into the synaptic space, whereby the zinc modulates activity of postsynaptic neurons though interactions with receptors, transporters and exchangers. However, high extracellular concentrations of zinc trigger seizures and are neurotoxic if substantial amounts of zinc reenter the cells via ion channels and accumulate in the cytoplasm. Tissue plasminogen activator (tPA), a secreted serine protease, is also proepileptic and excitotoxic. However, tPA counters zinc toxicity by promoting zinc import back into the neurons in a sequestered form that is nontoxic. Here, we identify the zinc influx transporter, ZIP4, as the pathway through which tPA mediates the zinc uptake. We show that ZIP4 is upregulated after excitotoxin stimulation of the mouse, male and female, hippocampus. ZIP4 physically interacts with tPA, correlating with an increased intracellular zinc influx and lysosomal sequestration. Changes in prosurvival signals support the idea that this sequestration results in neuroprotection. These experiments identify a mechanism via which neurons use tPA to efficiently neutralize the toxic effects of excessive concentrations of free zinc. PMID:20463217

Emmetsberger, Jaime; Mirrione, Martine M; Zhou, Chun; Fernandez-Monreal, Monica; Siddiq, Mustafa M; Ji, Kyungmin; Tsirka, Stella E

2010-05-12

214

Sustained thrombolysis with DNA-recombinant tissue type plasminogen activator in rabbits  

International Nuclear Information System (INIS)

Tissue type plasminogen activator (t-PA) is an effective thrombolytic agent in experimental animals. The duration of the thrombolytic effect of infused t-PA is unknown. The authors compared the duration of the thrombolytic effect of t-PA with streptokinase by measuring the lysis of "1"2"5I-fibrin-labeled thrombi in rabbit jugular veins at different times after a bolus injection of the fibrinolytic agents. The pharmacodynamics of both thrombolytic agents were determined in rabbits using a sensitive ex vivo fibrinolytic assay. Streptokinase and t-PA were given as a bolus dose of 15,000 U/kg. There was no detectable circulating fibrinolytic activity 30 minutes after the bolus dose of t-PA and 120 minutes after the bolus dose of streptokinase. The t-PA injection produced 34% thrombolysis at 30 minutes, 90% thrombolysis at 120 minutes, and 96% thrombolysis at 240 minutes. The streptokinase injection produced 17% thrombolysis at 30 minutes, 34% at 120 minutes, and 34% at 240 minutes. These observations indicate that the thrombolytic effect of t-PA is sustained beyond its time of clearance from the circulation whereas the thrombolytic effect of streptokinase closely parallels its activity in the circulation

1985-01-01

215

Activities of Acyclic Nucleoside Phosphonates against Orf Virus in Human and Ovine Cell Monolayers and Organotypic Ovine Raft Cultures  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Orf virus, a member of the Parapoxvirus genus, causes a contagious pustular dermatitis in sheep, goats, and humans. Previous studies have demonstrated the activity of (S)-1-[3-hydroxy-2-(phosphonomethoxy)propyl]cytosine (HPMPC; cidofovir; Vistide) against orf virus in cell culture and humans. We have evaluated a broad range of acyclic nucleoside phosphonates (ANPs) against several orf virus strains in primary lamb keratinocytes (PLKs) and human embryonic lung (HEL) monolayers. HPMPC, (S)-9-[3...

Dal Pozzo, F.; Andrei, G.; Holy?, A.; Den Oord, J.; Scagliarini, A.; Clercq, E.; Snoeck, R.

2005-01-01

216

Subacute Intranasal Administration of Tissue Plasminogen Activator Increases Functional Recovery and Axonal Remodeling after Stroke in Rats  

Digital Repository Infrastructure Vision for European Research (DRIVER)

As a thrombolytic agent, application of recombinant tissue plasminogen activator (tPA) to ischemic stroke is limited by the narrow time window and side effects on brain edema and hemorrhage. This study examined whether tPA, administered by intranasal delivery directly targeting the brain and spinal cord, provides therapeutic benefit during the subacute phase after stroke. Adult male Wistar rats were subjected to permanent right middle cerebral artery occlusion (MCAo). Animals were treated int...

Liu, Zhongwu; Li, Yi; Zhang, Li; Xin, Hongqi; Cui, Yisheng; Hanson, Leah R.; Frey, William H.; Chopp, Michael

2012-01-01

217

Successful thrombolysis using recombinant tissue plasminogen activator in cases of severe pulmonary embolism with mobile thrombi in the right atrium.  

Science.gov (United States)

Hereby, we report two cases of acute pulmonary embolism with concomitant right-sided thrombus, which were successfully treated using recombinant tissue plasminogen activator (rtPA). These patients had life-threatening acute right ventricular failure, which dramatically improved within hours following thrombolysis. These cases emphasize the clinical utility of rtPA for the treatment of life-threatening pulmonary embolism. PMID:24936311

Satiro?lu, Omer; Durako?lugil, Murtaza Emre; U?urlu, Yavuz; Sahin, Ismail; Do?an, Sitki; Ergül, Elif; Karada?, Zakir; Bostan, Mehmet

2014-06-01

218

Cloning and Expression of a Human Tissue Plasminogen Activator Variant:K2S in Escherichia coli  

Directory of Open Access Journals (Sweden)

Full Text Available The DNA sequence of Kringle-2 and serine protease domains of the human tissue plasminogen activator (reteplase, K2S was PCR amplified. This product was then cloned into the expression vector pET15b plasmid. The presence of the insert was confirmed by restriction digestion, PCR and determination of the nucleotide sequence. By using isopropyl ?-D thiogalactopyranoside (IPTG, reteplase was induced in E. coli BL21 cells and analyzed using polyacrylamide gel electrophoresis (PAGE.

Hamid Mir Mohammad Sadeghi

2007-01-01

219

Variants of human tissue-type plasminogen activator that lack specific structural domains of the heavy chain.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The heavy chain of tissue plasminogen activator (t-PA) consists of four domains [finger, epidermal-growth-factor (EGF)-like, kringle 1 and kringle 2] that are homologous to similar domains present in other proteins. To assess the contribution of each of the domains to the biological properties of the enzyme, site-directed mutagenesis was used to generate a set of mutants lacking sequences corresponding to the axons encoding the individual structural domains. The mutant proteins were assayed f...

1988-01-01

220

Alpha 2-antiplasmin supplementation inhibits tissue plasminogen activator-induced fibrinogenolysis and bleeding with little effect on thrombolysis.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Tissue plasminogen activator (t-PA) causes fibrinogen proteolysis when alpha 2-antiplasmin levels fall, and this may contribute to t-PA-induced hemorrhage. Because clot-bound plasmin is protected from alpha 2-antiplasmin inhibition, we tested the possibility that alpha 2-antiplasmin supplementation would block t-PA-induced fibrinogenolysis and bleeding without affecting thrombolysis. When added to human or rabbit plasma, alpha 2-antiplasmin inhibits t-PA-induced fibrinogenolysis, but hat litt...

Weitz, J. I.; Leslie, B.; Hirsh, J.; Klement, P.

1993-01-01

 
 
 
 
221

Targeting of peptide conjugated magnetic nanoparticles to urokinase plasminogen activator receptor (uPAR) expressing cells  

Science.gov (United States)

Ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles are currently being used as a magnetic resonance imaging (MRI) contrast agent in vivo, mainly by their passive accumulation in tissues of interest. However, a higher specificity can ideally be achieved when the nanoparticles are targeted towards cell specific receptors and this may also facilitate specific drug delivery by an enhanced target-mediated endocytosis. We report efficient peptide-mediated targeting of magnetic nanoparticles to cells expressing the urokinase plasminogen activator receptor (uPAR), a surface biomarker for poor patient prognosis shared by several cancers including breast, colorectal, and gastric cancers. Conjugation of a uPAR specific targeting peptide onto polyethylene glycol (PEG) coated USPIO nanoparticles by click chemistry resulted in a five times higher uptake in vitro in a uPAR positive cell line compared to nanoparticles carrying a non-binding control peptide. In accordance with specific receptor-mediated recognition, a low uptake was observed in the presence of an excess of ATF, a natural ligand for uPAR. The uPAR specific magnetic nanoparticles can potentially provide a useful supplement for tumor patient management when combined with MRI and drug delivery.Ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles are currently being used as a magnetic resonance imaging (MRI) contrast agent in vivo, mainly by their passive accumulation in tissues of interest. However, a higher specificity can ideally be achieved when the nanoparticles are targeted towards cell specific receptors and this may also facilitate specific drug delivery by an enhanced target-mediated endocytosis. We report efficient peptide-mediated targeting of magnetic nanoparticles to cells expressing the urokinase plasminogen activator receptor (uPAR), a surface biomarker for poor patient prognosis shared by several cancers including breast, colorectal, and gastric cancers. Conjugation of a uPAR specific targeting peptide onto polyethylene glycol (PEG) coated USPIO nanoparticles by click chemistry resulted in a five times higher uptake in vitro in a uPAR positive cell line compared to nanoparticles carrying a non-binding control peptide. In accordance with specific receptor-mediated recognition, a low uptake was observed in the presence of an excess of ATF, a natural ligand for uPAR. The uPAR specific magnetic nanoparticles can potentially provide a useful supplement for tumor patient management when combined with MRI and drug delivery. Electronic supplementary information (ESI) available. See DOI: 10.1039/c3nr32922d

Hansen, Line; Unmack Larsen, Esben Kjær; Nielsen, Erik Holm; Iversen, Frank; Liu, Zhuo; Thomsen, Karen; Pedersen, Michael; Skrydstrup, Troels; Nielsen, Niels Chr.; Ploug, Michael; Kjems, Jørgen

2013-08-01

222

Age-dependent neonatal intracerebral hemorrhage in plasminogen activator inhibitor 1 knockout mice.  

Science.gov (United States)

Intracerebral-intraventricular hemorrhages (ICH/IVH) in very preterm neonates are responsible for high mortality and subsequent disabilities. In humans, tissue plasminogen activator (t-PA) initiates fibrinolysis and activates endoluminal-endothelial receptors; dysfunction of the t-PA inhibitor (PAI-1) results in recurrent hemorrhages. We used PAI-1 knockout (PAI-1) mice to examine the role of t-PA in age-dependent intracranial hemorrhages as a possible model of preterm ICH/IVH. Intracortical injection of 2 ?L of phosphate-buffered saline produced a small traumatic injury and a high rate of hemorrhage in PAI-1 pups at postnatal day 3 (P3) or P5, whereas it had no effect in wild-type neonates. This resulted in white matter and cortical lesions, ventricle enlargement, hyperlocomotion, and altered cortical levels of serotonin and dopamine in the adult PAI mice. N-methyl-D-aspartate receptor blockers, plasmin- and matrix metalloproteinases inhibitors reduced hemorrhage and tissue lesions. In contrast to P3 to P5, no significant hemorrhages were induced in P10 PAI-1 pups and there were no behavioral or neurochemical alterations in adulthood. These data suggest that microvascular immaturity up to P5 in mice is a determinant factor required for t-PA-dependent vascular rupture. Neonatal PAI-1 mice could be a useful ICH/IVH model for studying the ontogenic window of vascular immaturity and vascular protection against later neurodisabilities. PMID:24709679

Leroux, Philippe; Omouendze, Priscilla L; Roy, Vincent; Dourmap, Nathalie; Gonzalez, Bruno J; Brasse-Lagnel, Carole; Carmeliet, Peter; Leroux-Nicollet, Isabelle; Marret, Stéphane

2014-05-01

223

Hepatitis B virus inhibits liver regeneration via epigenetic regulation of urokinase-type plasminogen activator.  

Science.gov (United States)

Liver regeneration after liver damage caused by toxins and pathogens is critical for liver homeostasis. Retardation of liver proliferation was reported in hepatitis B virus (HBV) X protein (HBx)-transgenic mice. However, the underlying mechanism of the HBx-mediated disturbance of liver regeneration is unknown. We investigated the molecular mechanism of the inhibition of liver regeneration using liver cell lines and a mouse model. The mouse model of acute HBV infection was established by hydrodynamic injection of viral DNA. Liver regeneration after partial hepatectomy was significantly inhibited in the HBV DNA-treated mice. Mechanism studies have revealed that the expression of urokinase-type plasminogen activator (uPA), which regulates the activation of hepatocyte growth factor (HGF), was significantly decreased in the liver tissues of HBV or HBx-expressing mice. The down-regulation of uPA was further confirmed using liver cell lines transiently or stably transfected with HBx and the HBV genome. HBx suppressed uPA expression through the epigenetic regulation of the uPA promoter in mouse liver tissues and human liver cell lines. Expression of HBx strongly induced hypermethylation of the uPA promoter by recruiting DNA methyltransferase (DNMT) 3A2. Conclusion: Taken together, these results suggest that infection of HBV impairs liver regeneration through the epigenetic dysregulation of liver regeneration signals by HBx. PMID:23483589

Park, Eun-Sook; Park, Yong Kwang; Shin, Chan Young; Park, Seung Hwa; Ahn, Sung Hyun; Kim, Doo Hyun; Lim, Keo-Heun; Kwon, So Young; Kim, Kwang Pyo; Yang, Sung-Il; Seong, Baik L; Kim, Kyun-Hwan

2013-08-01

224

Cinaciguat, a soluble guanylate cyclase activator, causes potent and sustained pulmonary vasodilation in the ovine fetus.  

Science.gov (United States)

Impaired nitric oxide-cGMP signaling contributes to severe pulmonary hypertension after birth, which may in part be due to decreased soluble guanylate cyclase (sGC) activity. Cinaciguat (BAY 58-2667) is a novel sGC activator that causes vasodilation, even in the presence of oxidized heme or heme-free sGC, but its hemodynamic effects have not been studied in the perinatal lung. We performed surgery on eight fetal (126 +/- 2 days gestation) lambs (full term = 147 days) and placed catheters in the main pulmonary artery, aorta, and left atrium to measure pressures. An ultrasonic flow transducer was placed on the left pulmonary artery to measure blood flow, and a catheter was placed in the left pulmonary artery for drug infusion. Cinaciguat (0.1-100 microg over 10 min) caused dose-related increases in pulmonary blood flow greater than fourfold above baseline and reduced pulmonary vascular resistance by 80%. Treatment with 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), an sGC-oxidizing inhibitor, enhanced cinaciguat-induced pulmonary vasodilation by >120%. The pulmonary vasodilator effect of cinaciguat was prolonged, decreasing pulmonary vascular resistance for >1.5 h after brief infusion. In vitro stimulation of ovine fetal pulmonary artery smooth muscle cells with cinaciguat after ODQ treatment resulted in a 14-fold increase in cGMP compared with non-ODQ-treated cells. We conclude that cinaciguat causes potent and sustained fetal pulmonary vasodilation that is augmented in the presence of oxidized sGC and speculate that cinaciguat may have therapeutic potential for severe neonatal pulmonary hypertension. PMID:19465519

Chester, Marc; Tourneux, Pierre; Seedorf, Greg; Grover, Theresa R; Gien, Jason; Abman, Steven H

2009-08-01

225

Fibrinogen Nijmegen: congenital dysfibrinogenemia associated with impaired t-PA mediated plasminogen activation and decreased binding of t-PA.  

Science.gov (United States)

Congenital dysfibrinogenemia was found in a patient with venous thrombosis. Blood clot lysis was prolonged and suggested an impairment of fibrinolysis. We investigated whether this was related to the fibrinogen abnormality. Fibrinopeptide release was normal but fibrin polymerization was defective in the patient. The stimulating effect of the patient's fibrin on t-PA mediated plasminogen activation was impaired. This could not be attributed to defective binding of plasminogen. However, the binding of t-PA to the patient's fibrin was about 16% less than to normal fibrin. A variant t-PA (G K1 K2 P), which contained only one of the two fibrin binding sites, i.e. the kringle-2 domain, was bound to the abnormal fibrin for only 50% of normal. We conclude that the prolongation of blood clot lysis and the impaired stimulation of t-PA mediated plasminogen activation are related to the defective binding of the kringle-2 domain of t-PA onto the fibrin moiety of the abnormal fibrinogen. The impairment of fibrinolysis might explain the occurrence of thrombosis in the patient. PMID:3142089

Engesser, L; Koopman, J; de Munk, G; Haverkate, F; Nováková, I; Verheijen, J H; Briët, E; Brommer, E J

1988-08-30

226

Angiotensin I-converting-enzyme (ACE)-inhibitory activity of tryptic peptides of ovine $\\beta$-lactoglobulin and of milk yoghurts obtained by using different starters  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The aim of this study was to investigate the angiotensin I-converting-enzyme (ACE)-inhibitory activity of tryptic hydrolysates of ovine $\\beta$-lactoglobulin, and of yoghurts made by using different starters. Ovine $\\beta$-lactoglobulin (a mixture of variants A and B at a ratio of 50/50) was subjected to trypsin activity. The degree of hydrolysis of native whole $\\beta$-lactoglobulin reached 56, 72, 93 and 95% after 1, 2, 8 and 24 h, respectively. ACE-inhibitory activity of tryptic hydrolysat...

2005-01-01

227

Adipocyte-derived factors potentiate nutrient-induced production of plasminogen activator inhibitor-1 by macrophages.  

Science.gov (United States)

Macrophages are more abundant in adipose tissue from obese individuals than from those of normal weight and may contribute to the metabolic consequences of obesity by producing various circulating factors. One of these factors is plasminogen activator inhibitor-1 (PAI-1), which contributes to both atherosclerosis and insulin resistance. Because nutritional factors appear to regulate PAI-1 expression, we hypothesized that exposure to fatty acids and adipocyte secretory products could stimulate production of PAI-1 by adipose macrophages. Increased free fatty acid (FFA) concentrations in blood for 5 hours in nondiabetic, overweight subjects markedly suppressed insulin-stimulated glucose uptake and raised circulating PAI-1 concentrations, with a concomitant increase in the expression of the PAI-1 gene in adipose tissue. FFAs also rapidly increased PAI-1 gene expression in adipose macrophages and PAI-1 protein immunofluorescence surrounding these cells. By contrast, PAI-1 expression in circulating monocytes was very low and was not affected by raising the concentration of FFAs. Medium from cultured adipocytes stimulated PAI-1 expression in cultured macrophages and potentiated the increase in PAI-1 messenger RNA expression in response to FFAs. Together, our data suggest that adipocyte-derived factors prime adipose macrophages so that they respond to nutritional signals (FFAs) by releasing a key inflammatory adipokine, PAI-1. PMID:20371491

Kishore, Preeti; Li, Weijie; Tonelli, Julia; Lee, Do-Eun; Koppaka, Sudha; Zhang, Kehao; Lin, Ying; Kehlenbrink, Sylvia; Scherer, Philipp E; Hawkins, Meredith

2010-02-24

228

Bicyclic peptide inhibitor of urokinase-type plasminogen activator : mode of action  

DEFF Research Database (Denmark)

The development of protease inhibitors for pharmacological intervention has taken a new turn with the use of peptide-based inhibitors. Here, we report the rational design of bicyclic peptide inhibitors of the serine protease urokinase-type plasminogen activator (uPA), based on the established monocyclic peptide, upain-2. It was successfully converted to a bicyclic peptide, without loss of inhibitory properties. The aim was to produce a peptide cyclised by an amide bond with an additional stabilising across-the-ring covalent bond. We expected this bicyclic peptide to exhibit a lower entropic burden upon binding. Two bicyclic peptides were synthesised with affinities similar to that of upain-2, and their binding energetics were evaluated by isothermal titration calorimetry. Indeed, compared to upain-2, the bicyclic peptides showed reduced loss of entropy upon binding to uPA. We also investigated the solution structures of the bicyclic peptide by NMR spectroscopy to map possible conformations. An X-ray structureof the bicyclic-peptide-uPA complex confirmed an interaction similar to that for the previous upain-1/upain-2-uPA complexes. These physical studies of the peptide-protease interactions will aid future designs of bicyclic peptide protease inhibitors.

Roodbeen, Renée; Jensen, Berit Paaske

2013-01-01

229

Factors predicting intracerebral hemorrhage of patients treated with intravenous recombinant tissue plasminogen activator  

International Nuclear Information System (INIS)

The use of recombinant tissue plasminogen activator (rt-PA) was approved in Japan in October 2005, and has had a marked effect on the treatment of patients presenting with acute ischemic stroke. Since the administration of rt-PA might cause intracerebral hemorrhage (ICH) and a poor prognosis, it is necessary to identify predictors of ICH after treatment with rt-PA. In this article, we examined 58 consecutive patients with acute ischemic stroke treated with intravenous rt-PA within 3 hours of symptom onset for 45 months, March 2006 to November 2009. In principle, we evaluated patients before and one day after rt-PA with MRI. We made a retrospective comparison of 21 patients with hemorrhagic change on CT and MRI T2* within 36 hours and 37 patients without hemorrhagic change. The rate of ICH with or without symptoms was increased with a higher National Institutes of Health Stroke Scale (NIHSS) and infarction range, defined by diffusion weighted imaging (DWI) Alberta Stroke Programme Early CT Score (ASPECTS). Major artery occlusion and reperfusion, including partial recanalization in MR angiography (MRA), were taken as factors in the hemorrhage group. In conclusion, DWI ASPECTS and NIHSS were useful predictors of ICH after rt-PA administration. (author)

2010-09-01

230

Acute myocardial infarction following intravenous tissue plasminogen activator for acute ischemic stroke: An unknown danger  

Directory of Open Access Journals (Sweden)

Full Text Available Thrombolysis with intravenous tissue (IV plasminogen activator (tPA is considered for patients with acute ischemic stroke falling within the described inclusion criteria defined by The National Institute of Neurological Disorders and Stroke (NINDS rtPA trial. Complications of IV thrombolysis with tPA are commonly related to hemorrhage, anaphylaxis, or arterial occlusion. We describe two cases of acute myocardial infarction (MI following IV tPA infusion for acute stroke. One of the patients had underlying ischemic heart disease (IHD while the other did not have any prior IHD. Both had presented with acute ischemic stroke within the window period of thrombolysis and had no contraindications for thrombolysis. Both the patients succumbed due to myocardial infarction and cardiovascular collapse due to new onset arrhythmias. Acute MI immediately following IV tPA for stroke is a rare but serious complication. The disruption of intracardiac thrombus and subsequent embolization to coronary arteries may be an important mechanism in the occurrence of MI after administration of tPA for acute ischemic stroke. As both the patients succumbed before the arrangement for coronary angiography, the demonstration of intracardiac or intracoronary thrombus was not possible. But clinically, the presence of chest pain with elevated troponin levels and ST segment elevation pointed to MI. We suspect that fragmentation and lysis of intracardiac thrombus may result in MI after use of tPA for acute ischemic stroke, though the remote possibility of simultaneous occurrence of two atherosclerotic events MI and stroke exists.

Adatia Sweta

2010-01-01

231

Acute myocardial infarction following intravenous tissue plasminogen activator for acute ischemic stroke: An unknown danger.  

Science.gov (United States)

Thrombolysis with intravenous tissue (IV) plasminogen activator (tPA) is considered for patients with acute ischemic stroke falling within the described inclusion criteria defined by The National Institute of Neurological Disorders and Stroke (NINDS) rtPA trial. Complications of IV thrombolysis with tPA are commonly related to hemorrhage, anaphylaxis, or arterial occlusion. We describe two cases of acute myocardial infarction (MI) following IV tPA infusion for acute stroke. One of the patients had underlying ischemic heart disease (IHD) while the other did not have any prior IHD. Both had presented with acute ischemic stroke within the window period of thrombolysis and had no contraindications for thrombolysis. Both the patients succumbed due to myocardial infarction and cardiovascular collapse due to new onset arrhythmias. Acute MI immediately following IV tPA for stroke is a rare but serious complication. The disruption of intracardiac thrombus and subsequent embolization to coronary arteries may be an important mechanism in the occurrence of MI after administration of tPA for acute ischemic stroke. As both the patients succumbed before the arrangement for coronary angiography, the demonstration of intracardiac or intracoronary thrombus was not possible. But clinically, the presence of chest pain with elevated troponin levels and ST segment elevation pointed to MI. We suspect that fragmentation and lysis of intracardiac thrombus may result in MI after use of tPA for acute ischemic stroke, though the remote possibility of simultaneous occurrence of two atherosclerotic events MI and stroke exists. PMID:20436751

Sweta, Adatia; Sejal, Sanghani; Prakash, Sanzgiri; Vinay, Chauhan; Shirish, Hastak

2010-01-01

232

Angiotensinogen and Plasminogen Activator Inhibitor-1 Gene Polymorphism in Relation to Renovascular Disease  

International Nuclear Information System (INIS)

The present study was designed to evaluate angiotensinogen (AGT) M235T and plasminogen activator inhibitor-1 (PAI-1) (4G/5G) polymorphisims in relation to the occurrence of atherosclerotic renal artery stenosis (ARAS) and recurrent stenosis. In this study, 30 patients were enrolled after angiographic demonstration of ARAS; 100 healthy subjects for AGT polymorphism and 80 healthy subjects for PAI-1 polymorphism were considered the control group. The patients were followed for a mean 46.1 ± 9.2 months. The patients had significantly higher frequencies of the MT genotype and the T allele than control group (?2 = 18.2, p 2 = 11.5 p 2= 2.45, p = 0.29 and ?2 = 0.019, p = 0.89). There were no significant differences in the genotype and allele findings for the patients with and without restenosis (p > 0.05). The C-reactive protein (CRP) level was higher in the patients with restenosis than in the patients without restenosis (7.694 ± 0.39 mg/L and 1.56 ± 1.08 mg/L) (p = 0.001). Our results suggest that the M235T MT genotype and T allele might be associated with increased risk of atherosclerotic renal artery stenosis. The CRP level might be an independent predictor for recurrent stenosis

2006-02-01

233

TFRC and ACTB as the best reference genes to quantify Urokinase Plasminogen Activator in breast cancer  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Biomedical researchers have long looked for ways to diagnose and treat cancer patients at the early stages through biomarkers. Although conventional techniques are routinely applied in the detection of biomarkers, attitudes towards using Real-Time PCR techniques in detection of many biomarkers are increasing. Normalization of quantitative Real-Time PCR is necessary to validate non-biological alteration occurring during the steps of RNA quantification. Selection of variably expressed housekeeping genes (HKs will affect the validity of the data. The aim of the present study was to identify uniformly expressed housekeeping genes in order to use in the breast cancer gene expression studies. Urokinase Plasminogen Activator was used as a gene of interest. Findings The expression of six HKs (TFRC, GUSB, GAPDH, ACTB, HPRT1 and RPLP0 was investigated using geNorm and NormFinder softwares in forty breast tumor, four normal and eight adjacent tissues. RPLP0 and GAPDH revealed maximum M value, while TFRC demonstrated lowest M value. Conclusions In the present study the most and the least stable genes were TFRC and RPLP0 respectively. TFRC and ACTB were verified as the best combination of two genes for breast cancer quantification. The result of this study shows that in each gene expression analysis HKs selection should be done based on experiment conditions.

Abdoli Nasrin

2011-06-01

234

Aggregation and retention of human urokinase type plasminogen activator in the yeast endoplasmic reticulum  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Secretion of recombinant proteins in yeast can be affected by their improper folding in the endoplasmic reticulum and subsequent elimination of the misfolded molecules via the endoplasmic reticulum associated protein degradation pathway. Recombinant proteins can also be degraded by the vacuolar protease complex. Human urokinase type plasminogen activator (uPA is poorly secreted by yeast but the mechanisms interfering with its secretion are largely unknown. Results We show that in Hansenula polymorpha overexpression worsens uPA secretion and stimulates its intracellular aggregation. The absence of the Golgi modifications in accumulated uPA suggests that aggregation occurs within the endoplasmic reticulum. Deletion analysis has shown that the N-terminal domains were responsible for poor uPA secretion and propensity to aggregate. Mutation abolishing N-glycosylation decreased the efficiency of uPA secretion and increased its aggregation degree. Retention of uPA in the endoplasmic reticulum stimulates its aggregation. Conclusions The data obtained demonstrate that defect of uPA secretion in yeast is related to its retention in the endoplasmic reticulum. Accumulation of uPA within the endoplasmic reticulum disturbs its proper folding and leads to formation of high molecular weight aggregates.

Smirnov Vladimir N

2002-10-01

235

Role of plasminogen activator inhibitor type-1 in radiation-induced normal tissues injury  

International Nuclear Information System (INIS)

Radiotherapy is an essential tool for cancer treatment, but there is a balance between benefits and risks related to the use of ionizing radiation: the objective is to deliver a maximum dose to the tumour to destroy or to sterilize it while protecting surrounding normal tissues. Radio-induced damages to normal tissues are therefore a limiting factor when increasing the dose delivered to the tumour. One of the objectives of this research thesis is to bring to the fore a relationship between the initiation of lesions and the development of late damages, more particularly in the intestine, and to identify the involved molecular actors and their inter-connectivity. After a first part presenting ionizing radiation, describing biological effects of ionizing radiation and their use in radiotherapy, presenting the intestine and the endothelium and discussing the intestine radio-sensitivity, discussing the radio-induced intestine damages and radiotherapy-induced complications, and presenting the plasminogen activator inhibitor (PAI-1) and its behaviour in presence of ionizing radiation, two articles are reproduced. The first one addresses the effect of a pharmacological inhibition and of genetic deficiency in PAI-1 on the evolution of radio-induced intestine lesions. The second one discusses the fact that radio-induced PAI-1-related death of endothelial cells determines the severity of early radio-induced intestine lesions

2010-01-01

236

DNA repair and induction of plasminogen activator in human fetal cells treated with ultraviolet light  

International Nuclear Information System (INIS)

Human fetal fibroblasts have been tested for development associated changes in DNA repair by utilizing nucleoid sedimentation as an assay for excision repair. Among skin fibroblasts the rate of excision repair was significantly higher in non-fetal cells than in fibroblasts derived from an 8 week fetus. Skin fibroblasts derived at 12 week gestation were more repair proficient than at 8 weeks. However, they exhibited a lower rate of repair than non-fetal cells. Enhancement of protease plasminogen activator (PA) was higher after u.v. irradiation in skin fibroblasts derived at 8 weeks than at 12 weeks gestation and was absent in non-fetal skin fibroblasts. Excision repair and PA inducibility depended on the tissue of origin in addition to gestational stage, as shown for skin and lung fibroblasts from the same 12 week fetus. The sedimentation velocity of nucleoids, prepared from unirradiated fibroblasts, in neutral sucrose gradients with or without ethidium bromide, indicated the presence of DNA strand breaks in fetal cells. It is proposed that reduced DNA repair in fetal cells may result from alterations in DNA supercoiling, and that persistent DNA strand breaks enhance transcription of PA gene(s). (author)

1984-01-01

237

Plasma soluble urokinase plasminogen activator receptor in children with urinary tract infection  

DEFF Research Database (Denmark)

OBJECTIVE: In this prospective study we investigated the role of plasma levels of soluble urokinase plasminogen activator receptor (suPAR) in children with urinary tract infection. MATERIAL AND METHODS: We measured the levels of plasma suPAR during admission in 42 children with suspected acute pyelonephritis and compared the results to acute DMSA scintigraphy. RESULTS: The mean level of plasma suPAR at admission was significantly elevated in children with renal involvement (7.3 ng/ml) assessed by the DMSA scintigraphy compared to children without renal involvement (4.4 ng/ml, P = 0.010). The positive predictive value of suPAR seems high, since all patients without renal involvement had low suPAR values. During treatment the mean level of plasma suPAR decreased. CONCLUSION: We conclude that plasma suPAR could be of clinical use for the diagnosis of acute pyelonephritis and that high levels of plasma suPAR might reflect the level of renal involvement and could therefore be a new indicator for renal scarring.

Wittenhagen, Per; Andersen, Jesper Brandt

2011-01-01

238

Saturated fatty acid intake can influence increase in plasminogen activator inhibitor-1 in obese adolescents.  

Science.gov (United States)

The aim of this study was to verify if saturated fatty acid intake adjusted by tertiles can influence metabolic, inflammation, and plasminogen activator inhibitor-1 (PAI-1) in obese adolescents. Body mass, height, body mass index, waist circumference, blood pressure, and body composition of 108 obese adolescents were obtained. Fasting glucose, insulin, PAI-1, and CRP were determined. Insulin resistance was assessed by Homeostasis Model Assessment (HOMA-IR) and insulin sensitivity by Quantitative Insulin Sensitivity Check Index (QUICKI). Dietetic intake was estimated by a 3-day dietary record, and volunteers were divided according to consumption of saturated fatty acids: tertile 1 [Low Saturated Fatty Acid Intake (Low-SFA): ?12.14?g], tertile 2 [Moderate Saturated Fatty Intake (Moderate SFA intake): 12.15-20.48?g], and tertile 3 [High Saturated Fatty Acid Intake (High-SFA Intake); >20.48?g]. Statistical analysis was performed using STATISTICA 7.0 software and the significance level was set at pinfluenced cardiovascular disease risks, since it increased PAI-1 and insulin resistance, and decreased insulin sensibility, leading to vicious cycle among food ingestion, pro-thrombotic state, and cardiovascular risks in obese adolescents. PMID:24619821

Masquio, D C L; de Piano, A; Campos, R M S; Sanches, P L; Corgosinho, F C; Carnier, J; Oyama, L M; do Nascimento, C M P O; de Mello, M T; Tufik, S; Dâmaso, A R

2014-04-01

239

Bicyclic Peptide Inhibitor of Urokinase-Type Plasminogen Activator : Mode of Action  

DEFF Research Database (Denmark)

The development of protease inhibitors for pharmacological intervention has taken a new turn with the use of peptidebased inhibitors. Here, we report the rational design of bicyclic peptide inhibitors of the serine protease urokinase-type plasminogen activator (uPA), based on the established monocyclic peptide, upain-2. It was successfully converted to a bicyclic peptide, without loss of inhibitory properties. The aim was to produce a peptide cyclised by an amide bond with an additional stabilising across-the-ring covalent bond. We expected this bicyclic peptide to exhibit a lower entropic burden upon binding. Two bicyclic peptides were synthesised with affinities similar to that of upain-2, and their binding energetics were evaluated by isothermal titration calorimetry. Indeed, compared to upain-2, the bicyclic peptides showed reduced loss of entropy upon binding to uPA. We also investigated the solution structures of the bicyclic peptide by NMR spectroscopy to map possible conformations. An X-ray structure of the bicyclicpeptideâ?? uPA complex confirmed an interaction similar to that for the previous upain-1/upain-2â??uPA complexes. These physical studies of the peptideâ??protease interactions will aid future designs of bicyclic peptide protease inhibitors

Roodbeen, Renée; Paaske, Berit

2013-01-01

240

The myofibroblast is the predominant plasminogen activator inhibitor-1-expressing cell type in human breast carcinomas  

DEFF Research Database (Denmark)

The tumor level of plasminogen activator inhibitor-1 (PAI-1) is an informative biochemical marker of a poor prognosis in several cancer types. However, the tumor biological functions of PAI-1 and the identity of PAI-1-expressing cells are controversial. With the aim of immunohistochemically localizing PAI-1 in formalin-fixed, paraffin-embedded invasive ductal breast carcinoma samples, we raised new polyclonal antibodies against PAI-1 from different expression systems. The antibodies were affinity purified by absorption on immobilized preparations of PAI-1 different from those used for immunization. The specificity of the antibodies was ensured by immunoblotting analysis. In immunohistochemistry, the staining pattern obtained with the antibodies showed a good correlation with the PAI-1 mRNA expression pattern. In all 25 cases analyzed, PAI-1 immunoreactivity was predominantly localized in fibroblast-like cells. Double-immunofluorescence analyses showed co-expression of PAI-1 and alpha-smooth muscle actin in these cells, suggesting that they are myofibroblasts. PAI-1 was also seen in some myoepithelial cells surrounding occasional foci of ductal carcinoma in situ (9 of 25), some endothelial cells (8 of 25), some cancer cells (3 of 25), and some mast cells (6 of 25). In conclusion, we have provided a robust immunohistochemical procedure for detection of PAI-1 and shown that the majority of the PAI-1-expressing cells in invasive ductal breast carcinomas are myofibroblasts.

Offersen, Birgitte Vrou; Nielsen, Boye Schnack

2003-01-01

 
 
 
 
241

Fibrinolytic therapy with low-dose recombinant tissue plasminogen activator in retinal vein occlusion.  

Science.gov (United States)

Fibrinolytic therapy aimed at early restoration of blood flow appears to be a promising therapeutic approach in haemorrhagic retinopathy. The risk of bleeding complications, a major problem with fibrinolysis, can be reduced by the use of low-dose thrombolytic regimens. In our study, 14 patients with ischaemic central (CRVO) or branch (BRVO) retinal vein occlusion who presented with severe visual loss and recent onset of symptoms were treated with a low dose (50 mg) of recombinant tissue plasminogen activator (rt-PA) and intravenous heparin. In 10 of 14 patients (7 CRVO, 3 BRVO), an increase in visual acuity of one line or more on the logarithmic visual acuity chart was noted and in 8 patients (6 CRVO, 2 BRVO) a reduction of areas of capillary non-perfusion was observed, suggesting that a restoration of retinal capillary blood flow can be achieved if fibrinolysis is initiated in the early phase of haemorrhagic retinopathy. In view of the poor prognosis in the natural course of haemorrhagic retinopathy and the potential haemorrhagic risk in fibrinolysis, the use of low-dose rt-PA appears to constitute an encouraging approach in the management of this disease. PMID:9787229

Hattenbach, L O; Steinkamp, G; Scharrer, I; Ohrloff, C

1998-01-01

242

Induction of plasminogen activator by UV light in normal and xeroderma pigmentosum fibroblasts  

Energy Technology Data Exchange (ETDEWEB)

Normal and DNA repair-deficient human fibroblasts have been used to study induction of plasminogen activator (PA) by DNA damage. UV light induced the synthesis of PA in skin fibroblasts of all types of xeroderma pigmentosum (XP) in XP heterozygotes and in human amniotic cells. Enzyme induction was, however, not observed in fibroblasts of normal adults. In classical XP, which are deficient in excision repair, PA synthesis occurred in a narrow range of low-UV fluences. In such strains, the level of enzyme produced was correlated with the extent of repair deficiency. UV fluences required for PA induction in XP variants and XP heteozygotes were at least 10 times those inducing enzyme synthesis in excision-deficient XP. Maximum enzyme induction occurred 48 hr after irradiation, and the highest levels of enzyme produced were 15-20 times those of PA baseline levels. Electrophoretic analysis showed that UV irradiation enhances the synthesis of the M/sub r/ 60,000 human urokinase-type PA, which is present in low amounts in untreated cells. Our results suggest that PA induction in human cells is caused by unrepaired DNA damage and represents a eukaryotic SOS-like function. In addition, PA induction may provide a sensitive assay for detection of cellular DNA repair deficiencies and identification of XP heterozygotes.

Miskin, R. (Weizmann Inst. of Science, Rehovot, Israel); Ben-Ishai, R.

1981-10-01

243

Induction of plasminogen activator by UV light in normal and xeroderma pigmentosum fibroblasts  

International Nuclear Information System (INIS)

Normal and DNA repair-deficient human fibroblasts have been used to study induction of plasminogen activator (PA) by DNA damage. UV light induced the synthesis of PA in skin fibroblasts of all types of xeroderma pigmentosum (XP) in XP heterozygotes and in human amniotic cells. Enzyme induction was, however, not observed in fibroblasts of normal adults. In classical XP, which are deficient in excision repair, PA synthesis occurred in a narrow range of low-UV fluences. In such strains, the level of enzyme produced was correlated with the extent of repair deficiency. UV fluences required for PA induction in XP variants and XP heteozygotes were at least 10 times those inducing enzyme synthesis in excision-deficient XP. Maximum enzyme induction occurred 48 hr after irradiation, and the highest levels of enzyme produced were 15-20 times those of PA baseline levels. Electrophoretic analysis showed that UV irradiation enhances the synthesis of the M/sub r/ 60,000 human urokinase-type PA, which is present in low amounts in untreated cells. Our results suggest that PA induction in human cells is caused by unrepaired DNA damage and represents a eukaryotic SOS-like function. In addition, PA induction may provide a sensitive assay for detection of cellular DNA repair deficiencies and identification of XP heterozygotes

1981-01-01

244

Biochemical mechanism of action of a diketopiperazine inactivator of plasminogen activator inhibitor-1.  

DEFF Research Database (Denmark)

XR5118 [(3 Z,6 Z )-6-benzylidine-3-(5-(2-dimethylaminoethyl-thio-))-2-(thienyl)methylene-2,5-dipiperazinedione hydrochloride] can inactivate the anti-proteolytic activity of the serpin plasminogen activator inhibitor-1 (PAI-1), a potential therapeutic target in cancer and cardiovascular diseases. Serpins inhibit their target proteases by the P(1) residue of their reactive centre loop (RCL) forming an ester bond with the active-site serine residue of the protease, followed by insertion of the RCL into the serpin's large central beta-sheet A. In the present study, we show that the RCL of XR5118-inactivated PAI-1 is inert to reaction with its target proteases and has a decreased susceptibility to non-target proteases, in spite of a generally increased proteolytic susceptibility of specific peptide bonds elsewhere in PAI-1. The properties of XR5118-inactivated PAI-1 were different from those of the so-called latent form of PAI-1. Alanine substitution of several individual residues decreased the susceptibility of PAI-1 to XR5118. The localization of these residues in the three-dimensional structure of PAI-1 suggested that the XR5118-induced inactivating conformational change requires mobility of alpha-helix F, situated above beta-sheet A, and is in agreement with the hypothesis that XR5118 binds laterally to beta-sheet A. These results improve our understanding of the unique conformational flexibility of serpins and the biochemical basis for using PAI-1 as a therapeutic target. Udgivelsesdato: 2003-Aug-1

Einholm, Anja P; Pedersen, Katrine E

2003-01-01

245

Oviduct-specific expression of tissue plasminogen activator in laying hens  

Directory of Open Access Journals (Sweden)

Full Text Available Egg-laying hens are important candidate bioreactors for pharmaceutical protein production because of the amenability of their eggs for protein expression. In this study, we constructed an oviduct-specific vector containing tissue plasminogen activator (tPA protein and green fluorescent protein (pL-2.8OVtPAGFP and assessed its expression in vitro and in vivo. Oviduct epithelial and 3T3 cells were cultured and transfected with pL-2.8OVtPAGFP and pEGP-N1 (control vector, respectively. The pL-2.8OVtPAGFP vector was administered to laying hens via a wing vein and their eggs and tissues were examined for tPA expression. The oviduct-specific vector pL-2.8OVtPAGFP was expressed only in oviduct epithelial cells whereas pEGP-N1 was detected in oviduct epithelial and 3T3 cells. Western blotting detected a 89 kDa band corresponding to tPA in egg white and oviduct epithelial cells, thus confirming expression of the protein. The amount of tPAGFP in eggs ranged 9 to 41 ng/mL on the third day after vector injection. The tPA expressed in egg white and oviduct epithelial cells showed fibrinolytic activity, indicating that the protein was expressed in active form. GFP was observed only in oviducts, with no detection in heart, muscle, liver and intestine. This is the first study to report the expression of tPA in egg white and oviduct epithelial cells using an oviduct-specific vector.

Hubdar Ali Kaleri

2011-01-01

246

Oviduct-specific expression of tissue plasminogen activator in laying hens  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Egg-laying hens are important candidate bioreactors for pharmaceutical protein production because of the amenability of their eggs for protein expression. In this study, we constructed an oviduct-specific vector containing tissue plasminogen activator (tPA) protein and green fluorescent protein (pL- [...] 2.8OVtPAGFP) and assessed its expression in vitro and in vivo. Oviduct epithelial and 3T3 cells were cultured and transfected with pL-2.8OVtPAGFP and pEGP-N1 (control vector), respectively. The pL-2.8OVtPAGFP vector was administered to laying hens via a wing vein and their eggs and tissues were examined for tPA expression. The oviduct-specific vector pL-2.8OVtPAGFP was expressed only in oviduct epithelial cells whereas pEGP-N1 was detected in oviduct epithelial and 3T3 cells. Western blotting detected a 89 kDa band corresponding to tPA in egg white and oviduct epithelial cells, thus confirming expression of the protein. The amount of tPAGFP in eggs ranged 9 to 41 ng/mL on the third day after vector injection. The tPA expressed in egg white and oviduct epithelial cells showed fibrinolytic activity, indicating that the protein was expressed in active form. GFP was observed only in oviducts, with no detection in heart, muscle, liver and intestine. This is the first study to report the expression of tPA in egg white and oviduct epithelial cells using an oviduct-specific vector.

Hubdar Ali, Kaleri; Liu, Xiang; Jueken, Aniwashi; Shiyong, Xu.

247

Anti-sense inhibition of tissue plasminogen activator production in differentiated F9 teratocarcinoma cells.  

Science.gov (United States)

F9 teratocarcinoma cells secrete the serine protease, tissue plasminogen activator (t-PA), upon differentiation induced in vitro by retinoic acid (RA) or RA and dibutyryl cAMP (RA/dbcAMP). A recombinant plasmid capable of directing the production of t-PA anti-sense RNA was constructed and transfected into F9 stem cells in an attempt to create a hypomorphic phenotype for t-PA synthesis. Several colonies were isolated which contained anti-sense RNA and which showed greater than a 50% reduction in t-PA activity upon differentiation. One such colony, 3b4, exhibited a 75% reduction in t-PA activity and was analyzed further. Large quantities of t-PA anti-sense transcript were expressed in the stem cells which are characterized by the absence of t-PA gene expression. In the induced cells, which normally express t-PA, the amount of detectable anti-sense transcript was significantly decreased. The amount of t-PA mRNA in differentiated cells containing t-PA anti-sense RNA was comparable to that in differentiated control cells. Subcellular localization of the mRNA in induced 3b4 cells appeared to be the same as induced control cells. Expression of collagen type IV, another marker of differentiation, was also monitored and was unaffected by the presence of t-PA anti-sense RNA in RA/dbcAMP-treated cells. The inhibition of differentiation-specific gene expression by anti-sense RNA may be useful for further studies of developmentally regulated genes. PMID:2458288

Pecorino, L T; Rickles, R J; Strickland, S

1988-10-01

248

High molecular weight kininogen binds phosphatidylserine and opsonizes urokinase plasminogen activator receptor-mediated efferocytosis.  

Science.gov (United States)

Phagocytosis of apoptotic cells (efferocytosis) is essential for regulation of immune responses and tissue homeostasis and is mediated by phagocytic receptors. In this study, we found that urokinase plasminogen activator receptor (uPAR) plays an important role in internalization of apoptotic cells and also characterized the underlying mechanisms. In a flow cytometry-based phagocytic assay, uPAR-deficient macrophages displayed significant defect in internalization but not tethering of apoptotic cells. When uPAR-deficient mice were challenged with apoptotic cells, they exhibited pronounced splenomegaly resulting from accumulation of abundant apoptotic cells in spleen. Overexpression of uPAR in HEK-293 cells enhanced efferocytosis, which was inhibited by Annexin V and phosphatidylserine (PS) liposome, suggesting that uPAR-mediated efferocytosis is dependent on PS. In serum lacking high m.w. kininogen (HK), a uPAR ligand, uPAR-mediated efferocytosis was significantly attenuated, which was rescued by replenishment of HK. As detected by flow cytometry, HK selectively bound to apoptotic cells, but not viable cells. In purified systems, HK was specifically associated with PS liposome. HK binding to apoptotic cells induced its rapid cleavage to the two-chain form of HK (HKa) and bradykinin. Both the H chain and L chain of HKa were associated with PS liposome and apoptotic cells. HKa has higher binding affinity than HK to uPAR. Overexpression of Rac1/N17 cDNA inhibited uPAR-mediated efferocytosis. HK plus PS liposome stimulated a complex formation of CrkII with p130Cas and Dock-180 and Rac1 activation in uPAR-293 cells, but not in control HEK-293 cells. Thus, uPAR mediates efferocytosis through HK interaction with PS on apoptotic cells and activation of the Rac1 pathway. PMID:24688027

Yang, Aizhen; Dai, Jihong; Xie, Zhanli; Colman, Robert W; Wu, Qingyu; Birge, Raymond B; Wu, Yi

2014-05-01

249

Minocycline and tissue plasminogen activator for stroke: Assessment of interaction potential  

Science.gov (United States)

Background and purpose New treatment strategies for acute ischemic stroke must be evaluated in the context of effective reperfusion. Minocycline is a neuroprotective agent that inhibits proteolytic enzymes and therefore could potentially both inactivate the clot lysis effect and decrease the damaging effects of tissue plasminogen activator. This study aimed to determine the effect of minocycline on tPA clot lysis and tPA induced hemorrhage formation after ischemia. Methods Fibrinolytic and amidolytic activities of tPA were investigated in vitro over a range of clinically relevant minocycline concentrations. A suture occlusion model of three hour temporary cerebral ischemia in rats treated with tPA and two different minocycline regimens was used. Blood brain barrier basal lamina components, MMPs, hemorrhage formation, infarct size, edema and behavior outcome were assessed. Results Minocycline did not affect tPA fibrinolysis. However, minocycline 3 mg/kg intravenous treatment decreased total protein expression of both MMP-2 (p=0.0034) and MMP-9 (p = 0.001 for 92 kDa and p=0.0084 for 87 kDa). It also decreased incidence of hemorrhage (p = 0.019), improved neurological outcome (p=0.0001 for Bederson score and p=0.0391 for paw grasp test) and appeared to decrease mortality. MMP inhibition was associated with decreased degradation in collagen IV and laminin-?1 (p=0.0001). Conclusions Combination treatment with minocycline is beneficial in tPA-treated animals and does not compromise clot lysis. These results also suggest minocycline neurovascular protection after stroke may involve the direct protection of the blood brain barrier during thrombolysis with tPA.

Machado, Livia S.; Sazonova, Irina; Kozak, Anna; Wiley, Daniel C.; El-Remessy, Azza B.; Ergul, Adviye; Hess, David C.; Waller, Jennifer L.; Fagan, Susan C.

2009-01-01

250

Uncharged isocoumarin-based inhibitors of urokinase-type plasminogen activator  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Urokinase-type plasminogen activator (uPA plays a major role in extracellular proteolytic events associated with tumor cell growth, migration and angiogenesis. Consequently, uPA is an attractive target for the development of small molecule active site inhibitors. Most of the recent drug development programs aimed at nonpeptidic inhibitors targeted at uPA have focused on arginino mimetics containing amidine or guanidine functional groups attached to aromatic or heterocyclic scaffolds. There is a general problem of limited bioavailability of these charged inhibitors. In the present study, uPA inhibitors were designed on an isocoumarin scaffold containing uncharged substituents. Results 4-Chloro-3-alkoxyisocoumarins were synthesized in which the 3-alkoxy group contained a terminal bromine; these were compared with similar inhibitors that contained a charged terminal functional group. Additional variations included functional groups attached to the seven position of the isocoumarin scaffold. N- [3-(3-Bromopropoxy-4-chloro-1-oxo-1H-isochromen-7-yl]benzamide was identified as an uncharged lead inhibitor of uPA, Ki = 0.034 ?M. Molecular modeling of human uPA with these uncharged inhibitors suggests that the bromine occupies the same position as positively charged arginino mimetic groups. Conclusion This study demonstrates that potent uncharged inhibitors of uPA can be developed based upon the isocoumarin scaffold. A tethered bromine in the three position and an aromatic group in the seven position are important contributors to binding. Although the aim was to develop compounds that act as mechanism-based inactivators, these inhibitors are competitive reversible inhibitors.

Deck Lorraine M

2006-02-01

251

The miR-143/-145 cluster regulates plasminogen activator inhibitor-1 in bladder cancer  

DEFF Research Database (Denmark)

Background:Upregulation of the proto-oncogene plasminogen activator inhibitor-1 (PAI-1) is a common hallmark of various solid tumours, but the mechanisms controlling its expression are not fully understood.Methods:We investigate microRNAs (miRNAs) regulating PAI-1 in a panel of normal bladder urothelial biopsies, superficial Ta bladder tumours and invasive T1-T4 tumours using expression microarrays and qRT-PCR. The prognostic implications of PAI-1 deregulation are established by tissue microarray staining of non-muscle-invasive bladder tumours. MicroRNA repression of PAI-1 is assayed by ectopic miRNA expression, argonaute immunoprecipitation and luciferase assays.Results:We found that the miR-143/-145 cluster is downregulated in all stages of bladder cancer and inversely correlated with PAI-1 expression. Mature miR-143 and miR-145 are coordinately expressed, and both directly target the PAI-1 3'UTR, leading to reduced PAI-1 mRNA and protein levels. Furthermore, we show that PAI-1 and miR-145 levels may serve as useful prognostic markers for non-muscle-invasive bladder tumours for which accurate progressive outcome is currently difficult to predict.Conclusion:This report provides the first evidence for direct miRNA regulation of PAI-1 in bladder cancer. We also demonstrate mRNA co-targeting by a cluster of non-family miRNAs, and suggest miR-145 and PAI-1 as clinically relevant biomarkers in bladder cancer.British Journal of Cancer advance online publication, 22 November 2011; doi:10.1038/bjc.2011.520 www.bjcancer.com

Villadsen, S B; Bramsen, Jesper Bertram

2012-01-01

252

Recombinant human erythropoietin reduces plasminogen activator inhibitor and ameliorates pro-inflammatory responses following trauma  

Directory of Open Access Journals (Sweden)

Full Text Available "n  "n Background and the purpose of the study: Besides its hematopoietic effects, erythropoietin (EPO by mobilization of iron and modulation of some inflammatory cytokines has antioxidant and anti-inflammatory properties. The purpose of this study was to evaluate these effects of erythropoietin and its impact on organ function in traumatized patients. "n Methods: Twenty-six ICU-admitted traumatized patients within 24 hrs after trauma were randomly assigned to the EPO (received EPO, 300 units/Kg/day and Control (not received EPO groups. The inflammatory biomarkers including Tumor Necrosis Factor alpha (TNF-?, Interleukin 1 (IL-1, Plasminogen Activator Inhibitor 1 (PAI-1 and Nitrotyrosine were recorded at the admission, 3, 6 and 9 days thereafter. Acute Physiology and Chronic Health Evaluation (APACHE II and Sequential Organ Failure Assessment (SOFA scores were also recorded. "n Results: Among 12 patients (EPO group TNF-? level at the day of 9 (P=0.046, and within EPO group at the days of 3 (P=0.026 ameliorate, 6 (P=0.016, and 9 (P=0.052 were significantly lowered. Level of IL-1 and PAI-1 decreased significantly at days of 3, 6 and 9 post intervention. Also there were significant differences between two groups in the SOFA score during three measured time intervals (the first, third and seventh days. "n Conclusion: From the results of this study it seems that injection of erythrocyte stimulating agent is well tolerated and inhibits the inflammatory response and oxidative stress following trauma.

M Mojtahedzadeh

2011-05-01

253

DNA repair and induction of plasminogen activator in human fetal cells treated with ultraviolet light  

International Nuclear Information System (INIS)

We have tested human fetal fibroblasts for development associated changes in DNA repair by utilizing nucleoid sedimentation as an assay for excision repair. Among skin fibroblasts the rate of excision repair was significantly higher in non-fetal cells than in fibroblasts derived from an 8 week fetus; this was evident by a delay in both the relaxation and the restoration of DNA supercoiling in nucleoids after irradiation. Skin fibroblasts derived at 12 week gestation were more repair proficient than those derived at 8 week gestation. However, they exhibited a somewhat lower rate of repair than non-fetal cells. The same fetal and non-fetal cells were also tested for induction of the protease plasminogen activator (PA) after u.v. irradiation. Enhancement of PA was higher in skin fibroblasts derived at 8 week than in those derived at 12 week gestation and was absent in non-fetal skin fibroblasts. These results are consistent with our previous findings that in human cells u.v. light-induced PA synthesis is correlated with reduced DNA repair capacity. Excision repair and PA inducibility were found to depend on tissue of origin in addition to gestational stage, as shown for skin and lung fibroblasts from the same 12 week fetus. Lung compared to skin fibroblasts exhibited lower repair rates and produced higher levels of PA after irradiation. The sedimentation velocity of nucleoids, prepared from unirradiated fibroblasts, in neutral sucrose gradients with or without ethidium bromide, indicated the presence of DNA strand breaks in fetal cells. It is proposed that reduced DNA repair in fetal cells may result from alterations in DNA supercoiling, and that persistent DNA strand breaks enhance transcription of PA gene(s)

1984-01-01

254

Polymorphism in methylenetetrahydrofolate reductase, plasminogen activator inhibitor-1, and apolipoprotein E in hemodialysis patients  

Directory of Open Access Journals (Sweden)

Full Text Available The methylenetetrahydrofolate reductase (MTHFR gene polymorphism, apolipoprotein E (apo s4 gene polymorphism and polymorphism of plasminogen activator inhibitor-1 (PAI-1 have been shown to be associated with end-stage renal disease (ESRD. To determine the prevalence of these mutations in Saudi patients with ESRD on hemodialysis, we studied the allelic frequency and genotype distribution in patients receiving hemodialysis and in a control group, all residing in the Eastern Province of Saudi Arabia. The genotypes were determined using allele specific hybridization procedures and were confirmed by restriction fragment length polymorphism. The T allele frequency and homozygous genotype of MTHFR in ESRD patients were 14% and 2.4%, respectively compared to 13.4% and 0%, respectively in the control group. The allele frequency and homozygous genotype of 4G/4G PAI-1 gene polymorphism were 46.4% and 4.8% respectively in ESRD patients compared to 57.1% and 32% respectively in the control group. The apo s4 allele frequency and homozygous genotype distribution in hemodialysis patients were 7% and 2.4%, respectively compared to 13% and 2% in the control group. Although allele frequency of C677T of MTHFR was statistically similar in the hemodialysis patients and in the control group, the homozygotes T allele genotype was over repre-sented in the hemodialysis group compared to normal. The prevalence of PAI-1 4G/4G polymorphism in ESRD patients was lower when com-pared to the control group. The prevalence of apo s4 allele did not differ significantly between the two groups. The present results demonstrate that all three studied polymorphic mutations are present in our population and that they may contribute to the etiology of the disease in our area.

Al-Muhanna Fahad

2008-01-01

255

The hepatic clearance of recombinant tissue-type plasminogen activator decreases after an inflammatory stimulus  

Directory of Open Access Journals (Sweden)

Full Text Available We have shown that tissue-type plasminogen activator (tPA and plasma kallikrein share a common pathway for liver clearance and that the hepatic clearance rate of plasma kallikrein increases during the acute-phase (AP response. We now report the clearance of tPA from the circulation and by the isolated, exsanguinated and in situ perfused rat liver during the AP response (48-h ex-turpentine treatment. For the sake of comparison, the hepatic clearance of a tissue kallikrein and thrombin was also studied. We verified that, in vivo, the clearance of 125I-tPA from the circulation of turpentine-treated rats (2.2 ± 0.2 ml/min, N = 7 decreases significantly (P = 0.016 when compared to normal rats (3.2 ± 0.3 ml/min, N = 6. The AP response does not modify the tissue distribution of administered 125I-tPA and the liver accounts for most of the 125I-tPA (>80% cleared from the circulation. The clearance rate of tPA by the isolated and perfused liver of turpentine-treated rats (15.5 ± 1.3 µg/min, N = 4 was slower (P = 0.003 than the clearance rate by the liver of normal rats (22.5 ± 0.7 µg/min, N = 10. After the inflammatory stimulus and additional Kupffer cell ablation (GdCl3 treatment, tPA was cleared by the perfused liver at 16.2 ± 2.4 µg/min (N = 5, suggesting that Kupffer cells have a minor influence on the hepatic tPA clearance during the AP response. In contrast, hepatic clearance rates of thrombin and pancreatic kallikrein were not altered during the AP response. These results contribute to explaining why the thrombolytic efficacy of tPA does not correlate with the dose administered.

Nagaoka M.R.

2000-01-01

256

Dopamine D3 receptor deletion increases tissue plasminogen activator (tPA) activity in prefrontal cortex and hippocampus.  

Science.gov (United States)

Considerable evidence indicates that dopamine (DA) influences tissue plasminogen activator (tPA)-mediated proteolytic processing of the precursor of brain-derived neurotrophic factor (proBDNF) into mature BDNF (mBDNF). However, specific roles in this process for the dopamine D3 receptor (D3R) and the underlying molecular mechanisms are yet to be fully characterized. In the present study, we hypothesized that D3R deletion could influence tPA activity in the prefrontal cortex and hippocampus. Using D3R knockout (D3(-/-)) mice, we show that receptor inactivation is associated with increased tPA expression/activity both in the prefrontal cortex and, to a greater extent, in the hippocampus. Augmented tPA expression in D3(-/-) mice correlated with increased BDNF mRNA levels, plasmin/plasminogen protein ratio and the conversion of proBDNF into mBDNF, as well as enhanced tPA and mBDNF immunoreactivity, as determined by quantitative real time polymerase chain reaction (qRT-PCR), immunoblot and immunohistochemistry. In addition, when compared to wild-type controls, D3(-/-) mice exhibited increased basal activation of the canonical cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)-driven Akt/cAMP-response element-binding protein (CREB) signaling cascade, as determined by the increased Akt phosphorylation both at Thr304 and Ser473 residues, of DA and cAMP-regulated protein of 32kDa (DARPP-32) at Thr34 and a phosphorylation state-dependent inhibition of glycogen synthetase kinase-3? (GSK-3?) at Ser9, a substrate of Akt whose constitutive function impairs normal CREB transcriptional activity through phosphorylation at its Ser129 residue. Accordingly, CREB phosphorylation at Ser133 was significantly increased in D3(-/-) mice, whereas the GSK-3?-dependent phosphorylation at Ser129 was diminished. Altogether, our finding reveals that mice lacking D3Rs show enhanced tPA proteolytic activity on BDNF which may involve, at least in part, a potentiated Akt/CREB signaling, possibly due to hindered GSK-3? activity. PMID:23906635

Castorina, A; D'Amico, A G; Scuderi, S; Leggio, G M; Drago, F; D'Agata, V

2013-10-10

257

Acute radiation effects on the content and release of plasminogen activator activity in cultured aortic endothelial cells  

International Nuclear Information System (INIS)

Confluent monolayers from three lines of bovine aortic endothelial cells were exposed to a single dose of 10 Gy of 60Co ? rays. Seventy-two hours later, the morphology of the irradiated and sham-irradiated monolayers was examined, and cellular DNA and protein contents were determined. In addition, the release of plasminogen activator (PA) activity into the culture media and PA activity in the cell lysates were assayed. DNA and protein contents in the irradiated monolayers were reduced to 43-50% and 72-95% of the control levels, respectively. These data indicate that radiation induced cell loss (detachment and/or lysis) from the monolayer, with hypertrophy of surviving (attached) cells to preserve the continuity of the monolayer surface. Total PA activity (lysate plus medium) in the irradiated dishes was reduced to 50-75% of the control level. However, when endothelial PA activity was expressed on the basis of DNA content, the irradiated monolayers from two of the three cell lines contained significantly more PA activity than did sham-irradiated monolayers. These data suggest that fibrinolytic defects observed in irradiated tissues in situ may be attributable at least in part to a radiation-induced inhibition of PA release by vascular endothelial cells

1985-01-01

258

Biochemical characterization and molecular cloning of a plasminogen activator proteinase (LV-PA) from bushmaster snake venom.  

Science.gov (United States)

The protein (LV-PA) from bushmaster (Lachesis muta muta) venom is a serine proteinase which specifically activates the inactive proenzyme plasminogen. LV-PA is a single chain glycoprotein with an apparent molecular mass of 33 kDa that fell to 28 kDa after treatment with N-Glycosidase F (PNGase F). Approximately 93% of its protein sequence was determined by automated Edman degradation of various fragments derived from a digestion with trypsin. A cDNA library of L. m. muta was constructed to generate expressed sequence tags (ESTs) and the plasminogen activator precursor cDNA was sequenced. The complete amino acid sequence of the enzyme was deduced from the cDNA sequence. LV-PA is composed of 234 residues and contains a single asparagine-linked glycosylation site, Asn-X-Ser, bearing sugars that account for approximately 10% of the enzyme's total molecular mass of 33 kDa. The sequence of LV-PA is highly similar to the plasminogen activators (PAs) TSV-PA from Trimeresurus stejnegeri venom and Haly-PA from Agkistrodon halys. Furthermore, the mature protein sequence of LV-PA exhibits significant similarity with other viperidae venom serine proteinases which affect many steps of hemostasis, ranging from the blood coagulation cascade to platelet function. The Michaelis constant (Km) and the catalytic rate constant (kcat) of LV-PA on four chromogenic substrates were obtained from Lineweaver-Burk plots. In addition, we used an indirect enzyme-linked immunoabsorbent assay (ELISA) to explore the phylogenetic range of immunological cross-reactivity (using antibodies raised against LV-PA) with analogous serine proteinases from two viperidae venoms and mammals. PMID:17034951

Sanchez, Eladio F; Felicori, Liza F; Chavez-Olortegui, Carlos; Magalhaes, Henrique B P; Hermogenes, Ana L; Diniz, Marcelo V; Junqueira-de-Azevedo, Inacio de L M; Magalhaes, Arinos; Richardson, Michael

2006-12-01

259

Plasminogen activation independent of uPA and tPA maintains wound healing in gene-deficient mice  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Simultaneous ablation of the two known activators of plasminogen (Plg), urokinase-type (uPA) and the tissue-type (tPA), results in a substantial delay in skin wound healing. However, wound closure and epidermal re-epithelialization are significantly less impaired in uPA;tPA double-deficient mice than in Plg-deficient mice. Skin wounds in uPA;tPA-deficient mice treated with the broad-spectrum matrix metalloproteinase (MMP) inhibitor galardin (N-[(2R)-2-(hydroxamido-carbonylmethyl)-4-methylpent...

Lund, Leif R.; Green, Kirsty A.; Stoop, Allart A.; Ploug, Michael; Almholt, Kasper; Lilla, Jennifer; Nielsen, Boye S.; Christensen, Ib J.; Craik, Charles S.; Werb, Zena; Danø, Keld; Rømer, John

2006-01-01

260

Dose-dependent modulation of choroidal neovascularization by plasminogen activator inhibitor type 1: Implications for clinical trials  

Digital Repository Infrastructure Vision for European Research (DRIVER)

PURPOSE. To explain the conflicting reports about the influence of plasminogen activator inhibitor type (PAI-1) on pathologic angiogenesis, such as occurs during the exudative form of age-related macular degeneration. METHODS. The expression of PAI-1 mRNA was analyzed in human and murine choroidal neovascularization (CNV) by RTPCR. The influences of increasing doses of recombinant PAI-1 were evaluated by daily intraperitoneal injections in PAI-1-1-and wild-type animals with a model of laser-i...

2003-01-01

 
 
 
 
261

Current use of intrapleural tissue plasminogen activator in the treatment of complex pleural processes: a review of the literature and report of 9 cases.  

Science.gov (United States)

Intrapleural tissue plasminogen activator is increasingly being utilized to treat complex pleural processes, such as complicated pleural effusions and empyemas, without surgical intervention. This technique is especially useful for patients with numerous co-morbidities or who are poor surgical candidates. We present our experience in treating nine adult patients with intrapleural tissue plasminogen activator for complex pleural processes. Patients were treated with one to eight doses until their condition resolved or surgical intervention was necessary. Seven patients had complete resolution, two patients required surgical intervention, and there were no complications from therapy. A review of all available literature on the use of intrapleural tissue plasminogen activator in adults is presented, comparing the various methods and techniques used by others. PMID:23066590

Dulaney, Caleb R; Merrill, Walter H; Tesseneer, Stephanie; Jenkins-Lonidier, Lora; Tribble, Curtis G

2012-07-01

262

Plasminogen in periodontitis and wound repair  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The plasminogen activator (PA) system plays a critical role in many physiological and pathological processes, such as fibrinolysis, extracellular matrix (ECM) degradation, wound healing, inflammation, and cancer. The key component of the PA system is plasmin, a broad-spectrum serine protease that is derived from its inactive form, plasminogen. The first aim of this thesis research was to determine the role of plasminogen in periodontitis, an inflammatory oral disease. The second aim was to ex...

2013-01-01

263

Plasminogen Acquisition and Activation at the Surface of Leptospira Species Lead to Fibronectin Degradation ?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Pathogenic Leptospira species are the etiological agents of leptospirosis, a widespread disease of human and veterinary concern. In this study, we report that Leptospira species are capable of binding plasminogen (PLG) in vitro. The binding to the leptospiral surface was demonstrated by indirect immunofluorescence confocal microscopy with living bacteria. The PLG binding to the bacteria seems to occur via lysine residues because the ligation is inhibited by addition of the lysine analog 6-ami...

2009-01-01

264

Plasminogen carbohydrate side chains in receptor binding and enzyme activation: A study of C6 glioma cells and primary cultures of rat hepatocytes  

International Nuclear Information System (INIS)

The human [Glu1]-plasminogen carbohydrate isozymes, plasminogen type I (Pg 1) and plasminogen type II (Pg 2), were separated by chromatography and studied in cell binding experiments at 4 degrees C with primary cultures of rat hepatocytes and rat C6 glioma cells. In both cell systems, Pg 1 and Pg 2 bound to an equivalent number of receptors, apparently representing the same population of surface molecules. The affinity for Pg 2 was slightly higher. With hepatocytes, the KD for Pg 1 was 3.2 +/- 0.2 microM, and the KD for Pg 2 was 1.9 +/- 0.1 microM, as determined from Scatchard transformations of the binding isotherms. The Bmax was approximately the same for both isozymes. With C6 cells, the KD for Pg 1 was 2.2 +/- 0.1 microM vs. 1.5 +/- 0.2 microM for Pg 2. Again, the Bmax was similar with both isozymes. 125I-Pg 1 and 125I-Pg 2 were displaced from specific binding sites by either nonradiolabeled isozyme. The KI for Pg 2 was slightly lower than the KI for Pg 1 with hepatocytes (0.9 vs. 1.3 microM) and with C6 cells (0.6 vs. 1.1 microM). No displacement was detected with miniplasminogen at concentrations up to 5.0 microM. Activation of Pg 1 and Pg 2 by recombinant two-chain tissue-plasminogen activator (rt-PA) was enhanced by hepatocyte cultures. The enhancing effect was greater with Pg 2. Hepatocyte cultures did not affect the activation of miniplasminogen by rt-PA or the activation of plasminogen by streptokinase. Unlike the hepatocytes, C6 cells did not enhance the activation of plasminogen by rt-PA or streptokinase; however, plasmin generated in the presence of C6 cells reacted less readily with alpha 2-antiplasmin

1990-01-01

265

Plasminogen carbohydrate side chains in receptor binding and enzyme activation: A study of C6 glioma cells and primary cultures of rat hepatocytes  

Energy Technology Data Exchange (ETDEWEB)

The human (Glu1)-plasminogen carbohydrate isozymes, plasminogen type I (Pg 1) and plasminogen type II (Pg 2), were separated by chromatography and studied in cell binding experiments at 4{degrees}C with primary cultures of rat hepatocytes and rat C6 glioma cells. In both cell systems, Pg 1 and Pg 2 bound to an equivalent number of receptors, apparently representing the same population of surface molecules. The affinity for Pg 2 was slightly higher. With hepatocytes, the KD for Pg 1 was 3.2 +/- 0.2 microM, and the KD for Pg 2 was 1.9 +/- 0.1 microM, as determined from Scatchard transformations of the binding isotherms. The Bmax was approximately the same for both isozymes. With C6 cells, the KD for Pg 1 was 2.2 +/- 0.1 microM vs. 1.5 +/- 0.2 microM for Pg 2. Again, the Bmax was similar with both isozymes. 125I-Pg 1 and 125I-Pg 2 were displaced from specific binding sites by either nonradiolabeled isozyme. The KI for Pg 2 was slightly lower than the KI for Pg 1 with hepatocytes (0.9 vs. 1.3 microM) and with C6 cells (0.6 vs. 1.1 microM). No displacement was detected with miniplasminogen at concentrations up to 5.0 microM. Activation of Pg 1 and Pg 2 by recombinant two-chain tissue-plasminogen activator (rt-PA) was enhanced by hepatocyte cultures. The enhancing effect was greater with Pg 2. Hepatocyte cultures did not affect the activation of miniplasminogen by rt-PA or the activation of plasminogen by streptokinase. Unlike the hepatocytes, C6 cells did not enhance the activation of plasminogen by rt-PA or streptokinase; however, plasmin generated in the presence of C6 cells reacted less readily with alpha 2-antiplasmin.

Hall, S.W.; VandenBerg, S.R.; Gonias, S.L. (Univ. of Virginia Health Sciences Center, Charlottesville (USA))

1990-07-01

266

Plasminogen activator inhibitor-1 removal using dextran sulphate columns. Evidence of PAI-1 homeostasis.  

LENUS (Irish Health Repository)

Patients with high plasma plasminogen activator inhibitor-1 (PAI-1) antigen levels are prone to develop thrombosis. Lowering PAI-1 levels may offer a therapeutic option and help to better understand PAI-1 metabolism. We examined the effect on plasma PAI-1 levels of LDL-apheresis using dextran sulphate (DS) columns in 12 patients (9 male, 3 female, 49 +\\/- 10 years) with heterozygous familial hypercholesterolaemia and coronary artery disease. One plasma volume equivalent (2.3-4.0 l) was treated during each procedure (at flow rates of 23 +\\/- 2 ml\\/min). Lipids and PAI-1 antigen levels were measured in plasma before and immediately after 19 aphereses (once in 7 patients, twice in 3 patients and three times in 2 patients) and also at 3 and 7 days post apheresis in five of these patients and in the column eluates from 8 of these patients. DS-apheresis reduced plasma cholesterol (50 +\\/- 8%), triglyceride (45 +\\/- 27%), apolipoprotein B (59 +\\/- 10%) and PAI-1 antigen levels from 10.2 +\\/- 5.2 to 6.0 +\\/- 3.1 ng\\/ml (P = 0.005). The PAI-I changes were independent of circadian variation. PAI-I bound to the DS-columns (3.51 +\\/- 1.03 ng\\/ml filtered plasma) and the percent of filtered PAI-1 that was bound correlated inversely (r = -0.81, P < 0.02) with basal PAI-1 levels indicating a high affinity saturable binding process. In four patients, plasma PAI-1 levels post-apheresis were higher than expected based on the amount of PAI-removed by the DS columns. The difference between the expected and actual PAI-1 level post apheresis, reflecting PAI-1 secretion or extracellular redistribution, correlated inversely with basal PAI-1 levels (r = -0.83, P = 0.01). PAI-1 levels returned to baseline pre-apheresis values 7 days post apheresis. PAI-1 antigen may be removed from plasma without adverse effect, resulting temporarily in its extracellular redistribution and restoration to baseline levels over one week. PAI-1 redistribution particularly when baseline pre-apheresis values were low may reflect a homeostatic mechanism to maintain sufficient PAI-1 levels. Procedures that could selectively remove PAI-1 from plasma may offer a treatment option for those with very high plasma PAI-1 levels and thrombosis.

Maher, Vincent M G

2009-08-01

267

Thrombolysis by intravenous tissue plasminogen activator (t-PA). Current status and future direction  

International Nuclear Information System (INIS)

In Japan, the intravenous tissue plasminogen activator (t-PA) Alteplase (0.6 mg/kg) administration of the within 3 h of the onset of acute ischemic stroke was approved for therapeutic use in the year 2006. t-PA induces thrombolysis in patients with acute ischemic stroke, and this method has gradually gained recognition among physicians and the general population. However, the number of patients who were treated using Alteplase is low (4,000-5,000 patients/year), and this figure accounts for only 2-3% of the annual number of cases of ischemic stroke. There is little doubt that Alteplase treatment is a potentially effective modality for some patients with acute ischemic stroke. The post-marketing surveillance of 4,749 Japanese patients treated using Alteplase showed that 33% of the patients had modified Rankin scale (mRS) scores of 0-1, 17% of patients died and 4.5% presented with symptomatic intracerebral hemorrhage (ICH); these results were comparable to those from other countries. The expansion of the therapeutic time window has been a matter of concern. The investigators of the European Cooperative Acute Stroke Study (ECASS) have reported that there was significant improvement in the clinical outcomes of patients with acute ischemie stroke when Alteplase was administered 3-4.5 h after the onset of the symptoms. Mismatches in perfusion- and diffusion-weighted (DW) magnetic resonance imaging (MRI) images have been used for selecting patients 3 h after the onset of symptoms, and the findings from MRI, dwimages (DWI) and MR angiography are practical predictors of t-PA therapy within 3 h of onset. The Middle Cerebral Artery Embolism Local Fibrinolytic Intervention Trial (MELT) Japan study showed that local intra-arterial fibrinolysis is effective in patients with embolic MCA occlusion within 6 h of the onset of symptoms. Combining the initiation of intravenous t-PA administration with further intra-arterial fibrinolysis or mechanical thrombolectomy may improve the recanalization rate. Thrombolysis in combination with ultrasound-enhanced clot lysis is another attractive therapy. In Japan the neuroprotective agent edaravone (radical scavenger) is commonly used in combination with t-PA, and it is expected to decrease the hemorrhagic transformation after t-PA administration. Acute cerebral ischemic symptoms may occasionally precede thoracic aortic dissection. Thoracic aortic dissection after t-PA administration may prove to be fatal, and it is an important disorder that must be differentially diagnosed. (author)

2009-01-01

268

The hepatic clearance of recombinant tissue-type plasminogen activator decreases after an inflammatory stimulus  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english We have shown that tissue-type plasminogen activator (tPA) and plasma kallikrein share a common pathway for liver clearance and that the hepatic clearance rate of plasma kallikrein increases during the acute-phase (AP) response. We now report the clearance of tPA from the circulation and by the isol [...] ated, exsanguinated and in situ perfused rat liver during the AP response (48-h ex-turpentine treatment). For the sake of comparison, the hepatic clearance of a tissue kallikrein and thrombin was also studied. We verified that, in vivo, the clearance of 125I-tPA from the circulation of turpentine-treated rats (2.2 ± 0.2 ml/min, N = 7) decreases significantly (P = 0.016) when compared to normal rats (3.2 ± 0.3 ml/min, N = 6). The AP response does not modify the tissue distribution of administered 125I-tPA and the liver accounts for most of the 125I-tPA (>80%) cleared from the circulation. The clearance rate of tPA by the isolated and perfused liver of turpentine-treated rats (15.5 ± 1.3 µg/min, N = 4) was slower (P = 0.003) than the clearance rate by the liver of normal rats (22.5 ± 0.7 µg/min, N = 10). After the inflammatory stimulus and additional Kupffer cell ablation (GdCl3 treatment), tPA was cleared by the perfused liver at 16.2 ± 2.4 µg/min (N = 5), suggesting that Kupffer cells have a minor influence on the hepatic tPA clearance during the AP response. In contrast, hepatic clearance rates of thrombin and pancreatic kallikrein were not altered during the AP response. These results contribute to explaining why the thrombolytic efficacy of tPA does not correlate with the dose administered.

M.R., Nagaoka; M., Kouyoumdjian; D.R., Borges.

269

Urokinase-type plasminogen activator receptor (uPAR) as a promising new imaging target : potential clinical applications  

DEFF Research Database (Denmark)

Urokinase-type plasminogen activator receptor (uPAR) has been shown to be of special importance during cancer invasion and metastasis. However, currently, tissue samples are needed for measurement of uPAR expression limiting the potential as a clinical routine. Therefore, non-invasive methods are needed. In line with this, uPAR has recently been identified as a very promising imaging target candidate. uPAR consists of three domains attached to the cell membrane via a glycosylphosphatidylinositol (GPI) anchor and binds it natural ligand uPA with high affinity to localize plasminogen activation at the cell surface. Due to the importance of uPAR in cancer invasion and metastasis, a number of high-affinity ligands have been identified during the last decades. These ligands have recently been used as starting point for the development of a number of ligands for imaging of uPAR using various imaging modalities such as optical imaging, magnetic resonance imaging, single photon emission computer tomography (SPECT) and positron emission topography (PET). In this review, we will discuss recent advances in the development of uPAR-targeted imaging ligands according to imaging modality. In addition, we will discuss the potential future clinical application for uPAR imaging as a new imaging biomarker.

Persson, Morten; Kjaer, Andreas

2013-01-01

270

The use of recombinant tissue-type plasminogen activator for the treatment of sudden and chronic hearing loss.  

Science.gov (United States)

We treated 80 patients with sudden hearing loss and 70 patients with chronic decreasing cochlear function to date with intravenous infusion of a glycoprotein analogous to recombinant tissue-type plasminogen activator (rt-PA): 3 mg dissolved in 250 mg of physiological saline given every 12 hours intravenously. Specifically excluded were patients with known abnormal coagulation. We treated no patient for longer than 20 days. Before therapy, 6 months therapy began, and at the end of treatment, all patients underwent the following instrumental examinations: prothrombin and fibrinogen level measurements, liminal tonal audiometry, tympanometry, and assessment of otoacoustic emissions and otoacoustic products of distortion. Daily during the treatment, we monitored the patients by liminal tonal audiometry and assessment of otoacoustic emissions with linear click emission and otoacoustic products of distortion. We discharged patients when their audibility threshold stabilized. No patient experienced side effects due to the treatment, and functional results were excellent in all (even more so if compared with the protocols of therapy previously used for these kinds of diseases). The tissue plasminogen activator was used with remarkable success for the treatment of sudden hearing loss; this study shows remarkable success with a very low dose of rt-PA as compared to the standard dosage used for treatment of myocardial infarction. PMID:16639920

Mora, Renzo; Dellepiane, Massimo; Mora, Francesco; Jankowska, Barbara

2005-01-01

271

D-glucose increases the synthesis of tissue-type plasminogen activator (t-PA) in human peritoneal mesothelial cells.  

Science.gov (United States)

Physical and chemical irritation of the peritoneum through glucose-based hyperosmolar dialysis solutions results in a nonbacterial serositis with fibrinous exudation. Thereby, human peritoneal mesothelial cells (HMC) play an important role in maintaining the balance between the peritoneal generation and degradation of fibrin by expressing the fibrinolytic enzyme tissue-type plasminogen activator (t-PA) as well as the specific plasminogen activator inhibitor-1 (PAI-1). In this study, we analyzed the effect of D-glucose and metabolically inert monosaccharides on the synthesis of t-PA and PAI-1 in cultured HMC. Incubation of HMC with D-glucose or the metabolically inert monosaccharides mannitol and L-glucose (5-90 mM) resulted in a time- and concentration-dependent increase in t-PA mRNA expression and antigen secretion without affecting PAI-1 synthesis. A similar effect was evident when HMC were first exposed sequentially to pooled spent peritoneal dialysis effluent for up to 4 hours, and subsequently incubated for 20 hours in control medium. The stimulating effect of high D-glucose on t-PA expression in HMC was prevented by treating the cells with different protein kinase C (PKC) inhibitors (Ro 31-8220, Gö 6976), but could not be mimicked by the PKC-activating phorbol ester PMA, indicating that this effect of high glucose is dependent on PKC activity, but not mediated through PKC activation. Also, using specific inhibitors (PD 98059, SB 203580) and activators (PMA, anisomycin, IL-1alpha) of the major routes of the mitogen-activated protein kinases (MAPKs) cascade, we found no evidence for a role of this cascade in regulating t-PA expression in HMC. We conclude that hyperosmolarity induces t-PA (but not PAI-1) in HMC via a regulatory mechanism that requires active PKC, but that does not involve a major pathway in the MAPK cascade. PMID:10494783

Sitter, T; Mandl-Weber, S; Wörnle, M; Haslinger, B; Goedde, M; Kooistra, T

1999-09-01

272

Complementary modes of action of tissue-type plasminogen activator and pro-urokinase by which their synergistic effect on clot lysis may be explained.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Tissue plasminogen activator (t-PA) and/or pro-urokinase (pro-UK) induced lysis of standard 125I-fibrin clots suspended in plasma was studied. Doses were kept below the concentration at which a nonspecific effect was seen, i.e., where fibrinogenolysis and major plasminogen consumption were observed. Small amounts of t-PA potentiated clot lysis by pro-UK by attenuating the lag phase characteristic of pro-UK, and causing a much earlier transition to the rapid phase of lysis. Similar promotion o...

Pannell, R.; Black, J.; Gurewich, V.

1988-01-01

273

Genetic and metabolic determinants of increased plasma plasminogen activator inhibitor-1 activity in children with renal transplants.  

Science.gov (United States)

Recent studies have shown that activity of plasminogen activator inhibitor-1 (PAI-1), a prothrombotic protein, may be increased in transplanted patients. The aim of the present investigation was to determine PAI-1 activity in pediatric recipients of renal transplants and to establish the relative contribution of both genetic and metabolic factors. In 29 children and adolescents with stable renal transplants, we related plasma PAI-1 activity to an indicator of inflammatory status [plasma concentration of C-reactive protein (CRP)] and to elements of the insulin resistance syndrome [body mass index (BMI), fasting insulinemia, HOMA index and plasma triglyceride, HDL-cholesterol, apolipoproteins A-1 and B concentrations]. Polymorphisms of PAI-1, apolipoprotein E (apoE) and angiotensin-converting enzyme (ACE) genes were also investigated. In all patients the study was repeated 1 year later. PAI-1 activity remained constantly elevated (23.4+/-22.8 and 18.6+/-7.8 U/ml in the first and second study, respectively, P=NS). Plasma PAI-1 activity correlated positively with CRP ( P=0.001), BMI z score ( P=0.02), fasting insulinemia ( P=0.009), and HOMA index ( P=0.006). No significant correlations were found in this population between plasma PAI-1 activity and age, gender, time elapsed after transplantation and plasma homocysteine, total cholesterol, LDL-cholesterol, HDL-cholesterol, apolipoprotein B, and apolipoprotein A-1. Plasma PAI-1 activity was not related to the cumulative dose of prednisone, cyclosporin A, or tacrolimus. Plasma PAI-1 activity was significantly higher in 5 children with apoE3/apoE4 genotype. No apparent influences of the PAI-1 4G/4G and ACE I/D genotypes were observed. In a multiple stepwise regression model, fasting insulinemia and apoE3/apoE4 genotype accounted for 45% of the observed plasma PAI-1 variability. We conclude that increased PAI-1 activity in children with stable renal transplants is determined both by genetic factors and by metabolic factors, the latter mainly linked to the insulin resistance syndrome. PMID:12761670

Aldámiz-Echevarría, Luis; Sanjurjo, Pablo; Vallo, Alfredo; Aguirre, Mireia; Pérez-Nanclares, Gustavo; Gimeno, Pilar; Rueda, Miguel; Ruiz, José Ignacio; Rodríguez-Soriano, Juan

2003-08-01

274

Binding areas of urokinase-type plasminogen activator-plasminogen activator inhibitor-1 complex for endocytosis receptors of the low-density lipoprotein receptor family, determined by site-directed mutagenesis  

DEFF Research Database (Denmark)

Some endocytosis receptors related to the low-density lipoprotein receptor, including low-density lipoprotein receptor-related protein-1A, very-low-density lipoprotein receptor, and sorting protein-related receptor, bind protease-inhibitor complexes, including urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor-1 (PAI-1), and the uPA-PAI-1 complex. The unique capacity of these receptors for high-affinity binding of many structurally unrelated ligands renders mapping of receptor-binding surfaces of serpin and serine protease ligands a special challenge. We have mapped the receptor-binding area of the uPA-PAI-1 complex by site-directed mutagenesis. Substitution of a cluster of basic residues near the 37-loop and 60-loop of uPA reduced the receptor-binding affinity of the uPA-PAI-1 complex approximately twofold. Deletion of the N-terminal growth factor domain of uPA reduced the affinity 2-4-fold, depending on the receptor, and deletion of both the growth factor domain and the kringle reduced the affinity sevenfold. The binding affinity of the uPA-PAI-1 complex to the receptors was greatly reduced by substitution of basic and hydrophobic residues in alpha-helix D and alpha-helix E of PAI-1. The localization of the implicated residues in the 3D structures of uPA and PAI-1 shows that they form a continuous receptor-binding area spanning the serpin as well as the A-chain and the serine protease domain of uPA. Our results suggest that the 10-100-fold higher affinity of the uPA-PAI-1 complex compared with the free components depends on the bonus effect of bringing the binding areas on uPA and PAI-1 together on the same binding entity.

Skeldal, Sune; Larsen, Jakob Vejby

2006-01-01

275

Evaluation of Serum Fibrinogen, Plasminogen, ? 2-Anti-Plasmin, and Plasminogen Activator Inhibitor Levels (PAI) and Their Correlation with Presence of Retinopathy in Patients with Type 1 DM.  

Science.gov (United States)

Background. Diabetic retinopathy (DR) is the leading cause of blindness in the world. Retinopathy can still progress despite optimal metabolic control. The aim of the study was to determine whether different degrees of DR (proliferative or nonproliferative) were associated with abnormally modulated hemostatic parameters in patients with T1DM. Method. 52 T1DM patients and 40 healthy controls were enrolled in the study. Patients were subdivided into three categories. Group I was defined as those without retinopathy, group II with NPRP, and group III with PRP. We compared these subgroups with each other and the control group (Group IV) according to the serum fibrinogen, plasminogen, alpha2-anti-plasmin ( ? 2-anti-plasmin), and PAI. Results. We detected that PAI-1, serum fibrinogen, and plasminogen levels were similar between the diabetic and control groups (P = 0.209, P = 0.224, and P = 0.244, resp.), whereas ? 2-anti-plasmin was higher in Groups I, II, and III compared to the control group (P < 0.01, P < 0.05, and P < 0.001, resp.). There was a positive correlation between serum ? 2-anti-plasmin and HbA1c levels (r = 0,268, P = 0.031). Conclusion. To our knowledge there is scarce data in the literature about ? 2-anti-plasmin levels in type 1 diabetes. A positive correlation between ? 2-anti-plasmin with HbA1c suggests that fibrinolytic markers may improve with disease regulation and better glycemic control. PMID:24818165

Polat, Sefika Burcak; Ugurlu, Nagihan; Yulek, Fatma; Simavli, Huseyin; Ersoy, Reyhan; Cakir, Bekir; Erel, Ozcan

2014-01-01

276

Maspin, the molecular bridge between the plasminogen activator system and beta1 integrin that facilitates cell adhesion.  

Science.gov (United States)

Maspin is a non-inhibitory serine protease inhibitor (serpin) that influences many cellular functions including adhesion, migration, and invasion. The underlying molecular mechanisms that facilitate these actions are still being elucidated. In this study we determined the mechanism by which maspin mediates increased MCF10A cell adhesion. Utilizing competition peptides and mutation analyses, we discovered two unique regions (amino acid residues 190-202 and 260-275) involved in facilitating the increased adhesion function of maspin. In addition, we demonstrate that the urokinase-type plasminogen activator (uPA)/uPA receptor (uPAR) complex is required for the localization and adhesion function of maspin. Finally, we showed that maspin, uPAR, and ?1 integrin co-immunoprecipitate, suggesting a novel maspin-uPA-uPAR-?1 integrin mega-complex that regulates mammary epithelial cell adhesion. PMID:21606500

Endsley, Michael P; Hu, Yanqiu; Deng, Yong; He, Xiaolin; Warejcka, Debra J; Twining, Sally S; Gonias, Steven L; Zhang, Ming

2011-07-15

277

Cryopreserved recombinant tissue plasminogen activator for the restoration of occluded central venous access devices in pediatric oncology patients  

International Nuclear Information System (INIS)

Thrombolytic therapy with urokinase 5000 units has been the standard therapy for restoration of thrombosed central catheters. However, with the decreased availability of urokinase, alternatives needed to be sought. The aim of the study was to determine the efficacy, bioactivity, dwell time and cost of cryopreserved recombinant tissue plasminogen activator (rTPA) in the restoration of occluded central venous access devices. For children 10kg, a dose of 1 mg was used. The dwell time was 1-2 hours. Of the 40 courses of rTPA, 39 fully restored central venous line patency (97%). Successful courses were instilled for an average of 1 hour. Cryopreserved rTPA appears to be safe and effective in the dose used to restore the patency of occluded central venous access devices in pediatric oncology patients. (author)

2002-01-01

278

Cerebrospinal fluid u-plasminogen activator and matrix metalloproteinase-9 levels in human eosinophilic meningitis associated with angiostrongyliasis.  

Science.gov (United States)

Cerebrospinal fluid (CSF) urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-9 (MMP-9) levels were measured in patients with eosinophilic meningitis associated with angiostrongyliasis (EOMA) by quantitative sandwich enzyme immunoassays. The CSF concentrations of uPA and MMP-9 were evaluated in 30 EOMA patients and 10 controls. The CSF uPA and MMP-9 levels of the EOMA patients were significantly higher than those of the controls (pstiff neck (p=0.048), while the MMP-9 was in correlation with CSF total protein (p=0.006), CSF leukocytosis (p=0.004) and CSF eosinophil numbers (p=0.02). CSF uPA and MMP-9 levels are potentially useful for the understanding of immunologic pathogenesis, for therapeutic targets, for the diagnosis of EOMA and for monitoring treatment efficacy. PMID:20584616

Sanpool, Oranuch; Intapan, Pewpan M; Thanchomnang, Tongjit; Santivarangkana, Taweesak; Niwattayakul, Kanigar; Chotmongkol, Verajit; Maleewong, Wanchai

2010-09-01

279

Activation of cAMP-dependent protein kinase alters the chromatin structure of the urokinase-type plasminogen activator gene promoter.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In LLC-PK1 cells, the urokinase-type plasminogen activator (uPA) gene is induced by two of the major signal transduction pathways, the protein kinase C (PKC) and the cAMP-dependent protein kinase (PKA) pathways. We have analyzed the chromatin structure of 26 kb of the uPA gene locus and have shown that PKA activation but not PKC activation induce major chromatin structural alterations in the uPA gene promoter. In uninduced cells, several DNase I hypersensitive (HS) sites were detected in the ...

Lee, J. S.; Catanzariti, L.; Hemmings, B. A.; Kiefer, B.; Nagamine, Y.

1994-01-01

280

Cloning and expression of a cDNA coding for a human monocyte-derived plasminogen activator inhibitor  

Energy Technology Data Exchange (ETDEWEB)

Human monocyte-derived plasminogen activator inhibitor (mPAI-2) was purified to homogeneity from the U937 cell line and partially sequenced. Oligonucleotide probes derived from this sequence were used to screen a cDNA library prepared from U937 cells. One positive clone was sequenced and contained most of the coding sequence as well as a long incomplete 3' untranslated region (1112 base pairs). This cDNA sequence was shown to encode mPAI-2 by hybrid-select translation. A cDNA clone encoding the remainder of the mPAI-2 mRNA was obtained by primer extension of U937 poly(A)/sup +/ RNA using a probe complementary to the mPAI-2 coding region. The coding sequence for mPAI-2 was placed under the control of the lambda P/sub L/ promoter, and the protein expressed in Escherichia coli formed a complex with urokinase that could be detected immunologically. By nucleotide sequence analysis, mPAI-2 cDNA encodes a protein containing 415 amino acids with a predicted unglycosylated M/sub r/ of 46,543. The predicted amino acid sequence of mPAI-2 is very similar to placental PAI-2 and shows extensive homology with members of the serine protease inhibitor (serpin) superfamily. mPAI-2 was found to be more homologous to ovalbumin (37%) than the endothelial plasminogen activator inhibitor, PAI-1 (26%). The 3' untranslated region of the mPAI-2 cDNA contains a putative regulatory sequence that has been associated with the inflammatory mediators.

Antalis, T.M.; Clark, M.A.; Barnes, T.; Lehrbach, P.R.; Devine, P.L.; Schevzov, G.; Goss, N.H.; Stephens, R.W.; Tolstoshev, P.

1988-02-01

 
 
 
 
281

Cloning and expression of a cDNA coding for a human monocyte-derived plasminogen activator inhibitor  

International Nuclear Information System (INIS)

Human monocyte-derived plasminogen activator inhibitor (mPAI-2) was purified to homogeneity from the U937 cell line and partially sequenced. Oligonucleotide probes derived from this sequence were used to screen a cDNA library prepared from U937 cells. One positive clone was sequenced and contained most of the coding sequence as well as a long incomplete 3' untranslated region (1112 base pairs). This cDNA sequence was shown to encode mPAI-2 by hybrid-select translation. A cDNA clone encoding the remainder of the mPAI-2 mRNA was obtained by primer extension of U937 poly(A)+ RNA using a probe complementary to the mPAI-2 coding region. The coding sequence for mPAI-2 was placed under the control of the ? P/sub L/ promoter, and the protein expressed in Escherichia coli formed a complex with urokinase that could be detected immunologically. By nucleotide sequence analysis, mPAI-2 cDNA encodes a protein containing 415 amino acids with a predicted unglycosylated M/sub r/ of 46,543. The predicted amino acid sequence of mPAI-2 is very similar to placental PAI-2 and shows extensive homology with members of the serine protease inhibitor (serpin) superfamily. mPAI-2 was found to be more homologous to ovalbumin (37%) than the endothelial plasminogen activator inhibitor, PAI-1 (26%). The 3' untranslated region of the mPAI-2 cDNA contains a putative regulatory sequence that has been associated with the inflammatory mediators

1988-01-01

282

Significant association of urokinase plasminogen activator Pro141Leu with serum lipid profiles in a Japanese population.  

Science.gov (United States)

Urokinase plasminogen activator (uPA) plays important physiological and pathological roles in fibrinolysis, cancer metastasis, and atherosclerosis. One study suggested that uPA also has a major role in cholesterol biosynthesis in humans via its receptor uPAR. Thus, we investigated the associations of functional uPA polymorphism (plasminogen activator, urokinase; PLAU Pro141Leu, rs2227564) with serum lipid profiles in a Japanese cohort. The study included 5152 participants (1465 male, 3687 female; age range, 35-69 years) of the Daiko Study, a part of the Japan Multi-Institutional Collaborative Cohort Study (J-MICC Study). Subjects were enrolled at the Daiko Medical Center from June 2008 to May 2010. Low-density lipoprotein cholesterol (LDL-C) and non-HDL-C (subtraction of high-density lipoprotein cholesterol from total cholesterol) in fasting blood of participants were each classified into two groups, < or ? 140 mg/dL, and < or ? 170 mg/dL, respectively. Genotype frequencies of PLAU Pro141Leu (rs2227564) were 59.1% for ProPro, 35.6% for ProLeu, and 5.3% for LeuLeu, and were in Hardy-Weinberg equilibrium (p=0.789). The allele frequencies were 0.769 for Pro and 0.231 for Leu. The multivariate-adjusted odd ratios (ORs) and 95% confidence intervals (CIs) for high LDL-C and non-HDL-C were 1.11 (95%CI; 1.00-1.23) and 1.16 (95%CI; 1.03-1.30) for those with Leu allele relative to ProPro. This study suggested that PLAU Pro141Leu (rs2227564) is significantly associated with serum lipid levels in a Japanese population. PMID:23628797

Tamura, Takashi; Morita, Emi; Kawai, Sayo; Okada, Rieko; Naito, Mariko; Wakai, Kenji; Hori, Yoko; Kondo, Takaaki; Hamajima, Nobuyuki

2013-07-25

283

Mannose 6-Phosphate/Insulin-like Growth Factor 2 Receptor Limits Cell Invasion by Controlling ?V?3 Integrin Expression and Proteolytic Processing of Urokinase-type Plasminogen Activator Receptor  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The multifunctional mannose 6-phosphate/insulin-like growth factor 2 receptor (M6P/IGF2R) is considered a tumor suppressor. We report here that RNA interference with M6P/IGF2R expression in urokinase-type plasminogen activator (uPA)/urokinase-type plasminogen activator receptor (uPAR) expressing human cancer and endothelial cells resulted in increased pericellular plasminogen activation, cell adhesion, and higher invasive potential through matrigel. M6P/IGF2R silencing led also to the cell su...

Schiller, Herbert B.; Szekeres, Andreas; Binder, Bernd R.; Stockinger, Hannes; Leksa, Vladimir

2009-01-01

284

Monocyte procoagulant activity and plasminogen activator. Role in human renal allograft rejection  

International Nuclear Information System (INIS)

Currently the mechanism of renal allograft rejection is not well understood. This study was designed to determine whether induction of monocyte procoagulant activity (MCPA) is important in the pathogenesis of renal allograft rejection. The MPCA assay was performed utilizing a one stage clotting assay both in normal and in factor-VII-deficient plasma. There was no increase in spontaneous MPCA in 20 patients with endstage renal failure and in 10 patients following abdominal or orthopedic operation, as compared with 20 normal controls. MPCA was assessed daily in 18 patients who had received renal allografts. Rejection episodes (RE) were predicted on the basis of persistent elevation in MPCA as compared with pretransplant levels. Rejection was diagnosed clinically and treated on the basis of standard criteria. Treated RE were compared with those predicted by elevated MPCA, and 3 patients were assessed as having no RE by MPCA and by standard criteria. In 8 RE, MPCA correlated temporally with RE (same day) when compared with standard criteria. In 12 RE, MPCA was predictive of rejection preceding standard criteria by at least 24 hr. There were 7 false-positive predictions on the basis of MPCA; however, there was only 1 false negative. MPCA was shown to be a prothrombinase by its dependence only on prothrombin and fibrinogen for full activity. MPCA may be important in the pathogenesis of allograft rejection, and additionally it may be a useful adjunct in the clinical management of this disease

1985-01-01

285

Activation of tissue plasminogen activator gene transcription by Neovastat, a multifunctional antiangiogenic agent  

International Nuclear Information System (INIS)

We recently reported that Neovastat, an antiangiogenic drug that is currently undergoing Phase III clinical trials for the treatment of non-small cell lung cancer, may inhibit angiogenesis through an increase in tPA activity. Here, we show that Neovastat also stimulates tPA gene transcription in endothelial cells, in a TNF?-like manner. RT-PCR analysis and gene reporter assays using the human tPA promoter indicated that upregulation of the tPA gene transcription by both Neovastat and TNF? was correlated with the phosphorylation of JNK1/2 and of I?B and that SP600125 and BAY11-7082, inhibitors of JNK and I?K, respectively, inhibit the increase of tPA gene transcription induced by Neovastat and TNF?. These results suggest that Neovastat induces tPA gene transcription through activation of the JNK and NF?B signaling pathways, leading to an increase of tPA secretion by endothelial cells. This may lead to the localized destruction of the fibrin provisional matrix that is necessary for neovessel formation and thus contribute to the reported antiangiogenic properties of this compound

2004-07-16

286

Induction of urokinase-type plasminogen activator by UV light in human fetal fibroblasts is mediated through a UV-induced secreted protein  

International Nuclear Information System (INIS)

Plasminogen activator was previously shown to be induced by UV light in human cells with low capacity to repair UV-induced DNA lesions. We now show that in human fetal fibroblasts UV light enhanced the two mRNA species coding for the urokinase-type plasminogen activator (uPA) and the tissue-type plasminogen activator, but immunological analysis revealed exclusively uPA activity. Several independent and complementary experiments indicated that induction of uPA was mediated, apparently entirely, through a UV-induced, secreted protein (UVIS) in the growth medium of irradiated cells. First, elevation of uPA mRNA after irradiation was severely blocked by cycloheximide. Second, replacement of conditioned medium in irradiated cells while the rate of plasminogen activator induction was maximal rapidly and completely stopped any further increase in uPA activity. Third, addition of the same removed conditioned medium to nonirradiated cells mimicked UV light in enhancing the level of uPA activity as well as that of uPA mRNA. Fourth, UVIS activity was completely lost by treating the conditioned medium with trypsin but not with nucleases. Kinetic measurements indicated that the accumulation of UVIS rather than the induction of uPA by UVIS conferred the rate-limiting step in the overall process of uPA induction. Both UV light and UVIS acted synergistically with inhibitors of DNA repair for uPA induction. Based on these results, a model is proposed implicating relaxation of DNA torsional stress of an as yet undefined DNA sequence(s) in the induction of UVIS, which is then responsible for activation of the uPA gene

1987-01-01

287

Soluble urokinase plasminogen activator receptor is a marker of dysmetabolism in HIV-infected patients receiving highly active antiretroviral therapy  

DEFF Research Database (Denmark)

Circulating soluble urokinase plasminogen activator receptor (suPAR) reflects the immune and pro-inflammatory status of the HIV-infected patient. Highly active antiretroviral therapy (HAART) suppresses suPAR. Independent of the immune response to HAART, suPAR remains elevated in some HIV-infected patients, reflecting possibly a low-grade pro-inflammatory state. Low-grade inflammation has been implicated in insulin resistance and other features of dysmetabolism. Accordingly it is hypothesized that circulating suPAR is associated with the metabolic status of HIV-infected patients on HAART. Fasting plasma suPAR was determined in 36 normoglycaemic HIV-infected patients on HAART (n = 18 lipodystrophic, and n = 18 non-lipodystrophic) who had estimated insulin sensitivity (Rd) and non-oxidative glucose disposal (NOGM) by euglycaemic hyperinsulinaemic clamps, indirect calorimetry, and glucose tracer infusion. Five patients had circadian suPAR concentrations measured (24 hr, 20 min-intervals). suPAR and non-HDL-cholesterol were higher and Rd, NOGM, and limb fat were lower in lipodystrophic patients than in non-lipodystrophic patients (P < 0.05). suPAR correlated positively with non-HDL-cholesterol and inversely with Rd, NOGM and limb fat (P < 0.005, n = 36). suPAR also correlated positively with leukocyte count and TNF-alpha (P < 0.01, n = 36) but not with IL-6. In multiple regression analyses suPAR was a stronger predictor of dysmetabolism than TNF-alpha and IL-6. Circadian suPAR did not systematically fluctuate. In conclusion, suPAR may reflect the metabolic status of the HIV-infected patient on HAART, thus linking low-grade inflammation, immune constitution, lipid and glucose metabolism, and fat redistribution. Circadian suPAR concentration appeared stable, suggesting that sampling schedule does not affect measurement. Further studies addressing whether suPAR predicts lipodystrophy and dysmetabolism in HIV-infected patients are warranted. J. Med. Virol. 80:209-216, 2008. (c) 2007 Wiley-Liss, Inc.

Andersen, Ove; Eugen-Olsen, Jesper

2008-01-01

288

Different radiolabelling methods alter the pharmacokinetic and biodistribution properties of Plasminogen Activator Inhibitor Type 2 (PAI-2) forms  

International Nuclear Information System (INIS)

Introduction: Tumour-associated urokinase plasminogen activator (uPA) is a critical marker of invasion and metastasis, and it is recognised as having strong prognostic relevance as well as being a therapeutic target. The specific uPA inhibitor plasminogen activator inhibitor type-2 (PAI-2, SerpinB2) specifically targets cell bound uPA and is internalised. Furthermore, preclinical studies have established the “proof-of-principle” of uPA-targeting by PAI-2-cytotoxin conjugates in human carcinoma models. However, these studies also suggest that PAI-2 is rapidly cleared via the renal system with low total dose reaching the tumour. In this study, a comparative single photon emission computed tomography (SPECT) and biodistribution (BD) analysis of different forms of PAI-2 labelled with the radioisotopes iodine-123 (123I) and technetium-99m (99mTc) was undertaken. Methods: The pharmacokinetic (PK) properties and BD of wild-type, ?CD-loop and PEGylated ?CD-loop PAI-2 labelled with the commonly used diagnostic SPECT radioisotopes 99mTc or 123I were compared in mouse models of human prostate carcinoma. Whole body SPECT imaging was also performed. Results: Both wild-type and the shorter but active ?CD-loop form of PAI-2 123I-labelled indirectly via conjugation to free amine groups (termed 123I-Bn-PAI-2) exhibited low tumour uptake, rapid excretion and similar PK profiles. Preliminary studies with a short branched-chain PEGylated 123I-Bn-PAI-2 ?CD-loop indicated an increase in blood retention time and tumour uptake. All 123I-Bn-labelled radiotracers were largely excreted through the kidneys. By comparison, both wild-type 123I-PAI-2 (labelled directly via tyrosine residues) and 99mTc-PAI-2 displayed different PK/BD patterns compared to 123I-Bn-PAI-2, suggesting greater liver based catabolism and thus slower elimination. SPECT imaging mimicked the BD results of all radiotracers. Conclusion: The different labelling methods gave distinct PAI-2 BD and tumour uptake profiles, with radioiodination resulting in the best non-tumour organ clearance profiles. Preliminary analyses with short branched-chain PEGylated 123I-Bn-PAI-2 ?CD-loop suggest that further investigations with other PEGylation reagents are required to optimise this approach for tumour imaging. These findings impact on the use of PAI-2 for drug delivery and/or diagnostic development.

2012-08-01

289

Expression of the plasminogen activator system and the inhibitors PAI-1 and PAI-2 in posttraumatic lesions of the CNS and brain injuries following dramatic circulatory arrests: an immunohistochemical study.  

Science.gov (United States)

Plasminogen activators as inducible extracellular serine proteases are involved in a variety of processes, such as the degradation of brain structures. In regions of brain degradation, an increase in the expression of genes encoding cytokines and proteinases has recently been demonstrated. We tested the hypothesis, whether the plasminogen activator system as well as the plasminogen activator inhibitors are expressed and possibly involved in a proteolytic cascade that breaks down the extracellular matrix as a result of ischemic or posttraumatic brain destructions. To study this supposition, we investigated immunohistochemically the expression of tPA, uPA and its receptor, the plasminogen activator inhibitors PAI-1 and PAI-2, tetranectin as well as the laminin breakdown as an event of secondary brain injury. Brain tissue from 21 autopsy cases with severe brain injuries, material from 14 ischemic infarcts and 11 controls with acute hypoxia were used. All components of the plasminogen activator system studied were over-expressed immunohistochemically in reactive astrocytes, microglia and endothelial cells around the lesion zone. Tetranectin showed an analogous distribution to the plasminogen activator system. A reduced immunoreactivity of laminin within the identical region of destruction was detected concomitant with laminin remnants in perivascular macrophages, so that a remarkable role of the plasmin cascade in the degradation of extracellular matrix proteins in the brain is taken into consideration. PMID:10674268

Dietzmann, K; von Bossanyi, P; Krause, D; Wittig, H; Mawrin, C; Kirches, E

2000-01-01

290

Impact of early accelerated dose tissue plasminogen activator on in-hospital patency of the infarcted vessel in patients with acute right ventricular infarction.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

OBJECTIVE: To assess the efficacy of early accelerated dose tissue plasminogen activator on in-hospital patency of the infarct related artery in patients with inferior myocardial infarction with and without right ventricular involvement. DESIGN: Single centre prospective assessment before discharge of infarct related vessel patency after early thrombolysis. SETTING: Tertiary cardiac referral centre at a university hospital. PATIENTS AND METHODS: 90 consecutive unselected patients with acute m...

Giannitsis, E.; Potratz, J.; Wiegand, U.; Stierle, U.; Djonlagic, H.; Sheikhzadeh, A.

1997-01-01

291

Quantitative PET of human urokinase-type plasminogen activator receptor with 64Cu-DOTA-AE105 : implications for visualizing cancer invasion  

DEFF Research Database (Denmark)

Expression levels of the urokinase-type plasminogen activator receptor (uPAR) represent an established biomarker for poor prognosis in a variety of human cancers. The objective of the present study was to explore whether noninvasive PET can be used to perform a quantitative assessment of expression levels of uPAR across different human cancer xenograft models in mice and to illustrate the clinical potential of uPAR PET in future settings for individualized therapy.

Persson, Morten; Madsen, Jacob

2012-01-01

292

Effects of Different Protein Sources on Plasminogen Inhibitor-1 and Factor VII Coagulant Activity Added to a Fat-Rich Meal in Type 2 Diabetes  

Digital Repository Infrastructure Vision for European Research (DRIVER)

BACKGROUND: Exaggerated postprandial triglyceride concentration is believed to be atherogenic, and to influence the risk of thrombosis. Both elevated plasminogen inhibitor 1 (PAI-1) and increased factor VII coagulant activity (FVIIc) are potential important contributors to the increased risk of cardiovascular disease in type 2 diabetes. AIM: We aimed to investigate the effect of adding four different protein types (i.e. casein, whey, cod, and gluten) to a fat-rich meal on postprandial respon...

Mortensen, Lene S.; Thomsen, Claus; Hermansen, Kjeld

2010-01-01

293

Cloning and Expression of a Human Tissue Plasminogen Activator Variant:K2S in Escherichia coli  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The DNA sequence of Kringle-2 and serine protease domains of the human tissue plasminogen activator (reteplase, K2S) was PCR amplified. This product was then cloned into the expression vector pET15b plasmid. The presence of the insert was confirmed by restriction digestion, PCR and determination of the nucleotide sequence. By using isopropyl ?-D thiogalactopyranoside (IPTG), reteplase was induced in E. coli BL21 cells and analyzed using polyacrylamide gel electrophoresis (PAGE).

2007-01-01

294

Increased soluble urokinase plasminogen activator receptor (suPAR) is associated with thrombosis and inhibition of plasmin generation in paroxysmal nocturnal hemoglobinuria (PNH) patients  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired genetic disorder of the bone marrow that produces intravascular hemolysis, proclivity to venous thrombosis, and hematopoietic failure. Mutation in the PIG-A gene of a hematopoietic stem cell abrogates synthesis of glycosylphosphoinositol (GPI) anchors and expression of all GPI-anchored proteins on the surface of progeny erythrocytes, leukocytes, and platelets. Urokinase plasminogen activator receptor (uPAR), a GPI-linked protein express...

Sloand, Elaine M.; Pfannes, Loretta; Scheinberg, Phillip; More, Kenneth; Wu, Colin O.; Horne, Mcdonald; Young, Neal S.

2008-01-01

295

Soluble urokinase plasminogen activator receptor in plasma is associated with incidence of CVD. Results from the Malmö Diet and Cancer Study.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

BACKGROUND: Soluble urokinase plasminogen activator receptor (suPAR) is a highly sensitive marker that reflects increased inflammation and is positively correlated with pro-inflammatory biomarkers. The aim of this study was to explore the relationship between suPAR, cardiovascular disease (CVD) risk factors and incidence of CVD. METHODS: suPAR was assessed in a random sample of participants (N=569), aged 63-68 years (mean age 65.5), from the Malmö Diet and Cancer Study (MDCS) cardiovascular ...

2012-01-01

296

Transconjunctival sutureless vitrectomy with tissue plasminogen activator, gas and intravitreal bevacizumab in the management of predominantly hemorrhagic age-related macular degeneration  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Luis Arias1,2, Jordi Monés11Institut de la Màcula i de la Retina, Centro Médico Teknon, Barcelona; 2Hospital Universitari de Bellvitge, L’Hospitalet de Llobregat, BarcelonaPurpose: To determine the efficacy and safety of treating predominantly hemorrhagic age-related macular degeneration (AMD) with transconjunctival sutureless vitrectomy (TSV), tissue plasminogen activator (tPA), sulphur hexafluoride (SF6), and intravitreal bevacizumab.Methods: Retrospective study, ...

Luis Arias; Jordi Monés

2010-01-01

297

Elevated urinary levels of urokinase-type plasminogen activator receptor (uPAR) in pancreatic ductal adenocarcinoma identify a clinically high-risk group  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Background The urokinase plasminogen activator receptor is highly expressed and its gene is amplified in about 50% of pancreatic ductal adenocarcinomas; this last feature is associated with worse prognosis. It is unknown whether the level of its soluble form (suPAR) in urine may be a diagnostic-prognostic marker in these patients. Methods The urinary level of suPAR was measured in 146 patients, 94 pancreatic ductal adenocarcinoma and 52 chronic pancreat...

Sorio Claudio; Mafficini Andrea; Furlan Federico; Barbi Stefano; Bonora Antonio; Brocco Giorgio; Blasi Francesco; Talamini Giorgio; Bassi Claudio; Scarpa Aldo

2011-01-01

298

Enhanced hippocampal long-term potentiation and learning by increased neuronal expression of tissue-type plasminogen activator in transgenic mice.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Adult cortical neurons can produce tissue-type plasminogen activator (tPA), an extracellular protease that plays a critical role in fibrinolysis and tissue remodelling processes. There is growing evidence that extracellular proteolysis may be involved in synaptic plasticity, axonal remodelling and neurotoxicity in the adult central nervous system. Here we show that transgenic mice overexpressing tPA in post-natal neurons have increased and prolonged hippocampal long-term potentiation (LTP), a...

Madani, R.; Hulo, S.; Toni, N.; Madani, H.; Steimer, T.; Muller, D.; Vassalli, J. D.

1999-01-01

299

Stereotactic fibrinolysis of spontaneous intracerebral hematoma using infusion of recombinant tissue plasminogen activator Fibrinólise com infusão de rtPA e drenagem estereotáxica de hematoma intracerebral espontâneo profundo  

Digital Repository Infrastructure Vision for European Research (DRIVER)

PURPOSE: The authors present a prospective study on 10 patients with stereotactic infusion of tissue plasminogen activator (rtPA) intraparenchimal hemorrhage. METHODS: Between 1999 and 2000, 10 patients with deep seated hematomas in the basal ganglia were selected for stereotactic infusion of rtPA and spontaneous clot drainage. RESULTS: All cases had about 80% reduction of the hematoma volume in the CT scan at the third day. The intracranial pressure was normalized by the third day too. There...

José Augusto Nasser; Asdrubal Falavigna; Márcio Bezerra; Victor Martinez; Gabriel Freitas; Armando Alaminos; Antônio Bonatelli; Fernando Ferraz

2002-01-01

300

Impact of the -675 4G/5G polymorphism of the plasminogen activator inhibitor-1 gene on childhood IgA nephropathy  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen activator inhibitor-1 (PAI-1) is an important regulator of the fibrinolytic pathway and extracellular matrix (ECM) turnover. The -675 4G/5G polymorphism in the PAI-1 promoter is associated with altered PAI-1 transcription, suggesting that this polymorphism may be a candidate risk factor for diseases characterized by ECM accumulation, such as immunoglobulin A nephropathy (IgAN) and mesangial proliferative glomerulonephritis (MesPGN). We genotyped childhood patients with biopsy-conf...

Han, Su-ryun; Kim, Cheon-jong; Lee, Byung-cheol

2012-01-01

 
 
 
 
301

Overexpression of SERBP1 (Plasminogen activator inhibitor 1 RNA binding protein in human breast cancer is correlated with favourable prognosis  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Plasminogen activator inhibitor 1 (PAI-1 overexpression is an important prognostic and predictive biomarker in human breast cancer. SERBP1, a protein that is supposed to regulate the stability of PAI-1 mRNA, may play a role in gynaecological cancers as well, since upregulation of SERBP1 was described in ovarian cancer recently. This is the first study to present a systematic characterisation of SERBP1 expression in human breast cancer and normal breast tissue at both the mRNA and the protein level. Methods Using semiquantitative realtime PCR we analysed SERBP1 expression in different normal human tissues (n?=?25, and in matched pairs of normal (n?=?7 and cancerous breast tissues (n?=?7. SERBP1 protein expression was analysed in two independent cohorts on tissue microarrays (TMAs, an initial evaluation set, consisting of 193 breast carcinomas and 48 normal breast tissues, and a second large validation set, consisting of 605 breast carcinomas. In addition, a collection of benign (n?=?2 and malignant (n?=?6 mammary cell lines as well as breast carcinoma lysates (n?=?16 were investigated for SERBP1 expression by Western blot analysis. Furthermore, applying non-radioisotopic in situ hybridisation a subset of normal (n?=?10 and cancerous (n?=?10 breast tissue specimens from the initial TMA were analysed for SERBP1 mRNA expression. Results SERBP1 is not differentially expressed in breast carcinoma compared to normal breast tissue, both at the RNA and protein level. However, recurrence-free survival analysis showed a significant correlation (P?=?0.008 between abundant SERBP1 expression in breast carcinoma and favourable prognosis. Interestingly, overall survival analysis also displayed a tendency (P?=?0.09 towards favourable prognosis when SERBP1 was overexpressed in breast cancer. Conclusions The RNA-binding protein SERBP1 is abundantly expressed in human breast cancer and may represent a novel breast tumour marker with prognostic significance. Its potential involvement in the plasminogen activator protease cascade warrants further investigation.

Serce Nuran Bektas

2012-12-01

302

Accelerating thrombolysis with chitosan-coated plasminogen activators encapsulated in poly-(lactide-co-glycolide) (PLGA) nanoparticles.  

Science.gov (United States)

Accelerating thrombolysis using plasminogen activators (PAs) encapsulated liposomes or PEG microparticles by pressure-driven permeation have been demonstrated in vitro and in vivo in animal models. However, designing and delivering PA-encapsulated nanoparticles (NPs) to enhance thrombolysis by applying electrostatic forces or ligand-receptor interactions between the NPs and blood clots has not been proposed. Therefore, without a pressure-driving factor, tissue-plasminogen activator (t-PA) encapsulated in PLGA NPs with chitosan (CS) and CS-GRGD coating and their thrombolysis capabilities in a blood clot-occluded tube model were evaluated by determining clot lysis times and the masses of the digested clots. The characteristics and release profiles of t-PA-encapsulated PLGA, PLGA/CS and PLGA/CS-GRGD NPs are determined by FT-IR, a laser particle/zeta potential analyzer and HPLC. Additionally, the permeation capacities of the NPs into flat blood clots were examined. For example, the mean particle sizes and encapsulation efficacies of t-PA for the NPs are in the ranges 260-320 nm and 65.5-70.5%, respectively. The results reveal that the NPs for the shortest clot lysis time and the highest weight percentages of digested clot are PLGA/CS (20.7 +/- 0.7 min) and PLGA/CS-GRGD (25.7 +/- 1.3 wt%), respectively. Compared with t-PA solution, the NPs can significantly shorten clot lysis times in the following order: PLGA/CS NPs (38.8 +/- 1.5%) > PLGA/CS-GRGD NPs (16.3 +/- 1.0%) > PLGA NPs (7.7 +/- 1.2%). Compared with t-PA solution, the NPs significantly increase the weight of digested clots in the order, PLGA/CS-GRGD (40.9 +/- 1.5%) > PLGA/CS (27.8 +/- 1.2%) > PLGA (8.6 +/- 0.6%). The highest release rate of t-PA in the fast release phase and the highest permeability into intra-clots of PLGA/CS and PLGA/CS-GRGD NPs, respectively, correspond to the shortest clot lysis time and the largest increase in weight of the digested clots among the NP system. In conclusion, the NPs designed based on new concepts significantly accelerate thrombolysis in vitro in this model, and may be useful in clinical study. PMID:17953984

Chung, Tze-Wen; Wang, Shoei-Shen; Tsai, Wei-Jain

2008-01-01

303

Circulating soluble urokinase plasminogen activator receptor predicts cancer, cardiovascular disease, diabetes and mortality in the general population  

DEFF Research Database (Denmark)

Abstract. Eugen-Olsen J, Andersen O, Linneberg A, Ladelund S, Hansen TW, Langkilde A, Petersen J, Pielak T, Møller LN, Jeppesen J, Lyngbaek S, Fenger M, Olsen MH, Hildebrandt PR, Borch-Johnsen K, Jørgensen T, Haugaard SB (Copenhagen University, Hvidovre Hospital, Hvidovre; Copenhagen University Hospital, Glostrup; Copenhagen University Hospital, Copenhagen; Copenhagen University Hospital, Glostrup; Copenhagen University, Hvidovre Hospital, Hvidovre; Steno Diabetes Center, Gentofte; University of Aarhus, Aarhus; University of Copenhagen, Copenhagen; Copenhagen University, Hvidovre Hospital, Hvidovre, Denmark). Circulating soluble urokinase plasminogen activator receptor predicts cancer, cardiovascular disease, diabetes and mortality in the general population. J Intern Med 2010; doi: 10.1111/j.1365-2796.2010.02252.x. Background. Low-grade inflammation is thought to contribute to the development of cardiovascular disease (CVD), type-2 diabetes mellitus (T2D), cancer and mortality. Biomarkers of inflammation may aid in risk prediction and enable early intervention and prevention of disease. Objective. The aim of this study was to investigate whether plasma levels of the inflammatory biomarker soluble urokinase plasminogen activator receptor (suPAR) are predictive of disease and mortality in the general population. Design. This was an observational prospective cohort study. Cohort participants were included from June 1993 to December 1994 and followed until the end of 2006. Setting. General adult Caucasian population. Participants. The MONICA10 study, a population-based cohort recruited from Copenhagen, Denmark, included 2602 individuals aged 41, 51, 61 or 71 years. Measurements. Blood samples were analysed for suPAR levels using a commercially available enzyme-linked immunosorbent assay. Risk of cancer (n = 308), CVD (n = 301), T2D (n = 59) and mortality (n = 411) was assessed with a multivariate proportional hazards model using Cox regression. Results. Elevated baseline suPAR level was associated with an increased risk of cancer, CVD, T2D and mortality during follow-up. suPAR was more strongly associated with cancer, CVD and mortality in men than in women, and in younger compared with older individuals. suPAR remained significantly associated with the risk of negative outcome after adjustment for a number of relevant risk factors including C-reactive protein levels. Limitation. Further validation in ethnic populations other than Caucasians is needed. Conclusion. The stable plasma protein suPAR may be a promising biomarker because of its independent association with incident cancer, CVD, T2D and mortality in the general population.

Eugen-Olsen, J; Andersen, O

2010-01-01

304

The prognostic value of plasma soluble urokinase plasminogen activator receptor (suPAR) levels in stage III ovarian cancer patients.  

Science.gov (United States)

The level of the urokinase plasminogen activator receptor (uPAR) is elevated in tumor tissue from several forms of cancer. uPAR is shed from the cell surface and the soluble form, soluble urokinase plasminogen activator receptor (suPAR), has been detected in several body fluids. High plasma levels of suPAR in patients with colorectal cancer and high serum levels of suPAR in patients with recurrent metastatic breast cancer have been associated with poor prognosis. In patients with ovarian cancer (OC) it has been shown that the level of suPAR is very high in ascites and cystic fluid and that high serum levels of suPAR were associated with shorter survival of the patients. We evaluated suPAR preoperatively in plasma from primary OC stage III patients and tested for association with prognosis. The prognostic significance of suPAR was also compared to two biochemical markers; cancer antigen 125 (CA125) and tetranectin (TN). No significant differences were found between patients who died of OC compared to patients still alive regarding median plasma suPAR levels (p=0.62) and median serum CA125 levels (p=0.26). In contrast, a significant difference was found between dead and alive OC patients for the median serum TN level (p<0.0001). Dividing the patients into two groups, corresponding to preoperative plasma suPAR levels below or equal to 2.0 ng/ml and higher than 2.0 ng/ml, no significant difference in survival was found between the two groups (p=0.49). When different cut-off levels of plasma suPAR were considered (2.74 ng/ml, 3.25 ng/ml and 4.18 ng/ml), no significant differences in survival could be detected (p=0.58, p=0.68 and p=0.05). Multivariate Cox regression analysis showed that the only independent prognostic factors were radicality after primary surgery (RH=5.34; 95% CI, 2.34-12.20; p<0.0001) and preoperative serum TN (RH=0.69, 95% CI, 0.57-0.82; p<0.0001), whereas plasma suPAR (4.18 ng/ml), age, histological type of tumour and serum CA 125 had no independent prognostic value. In conclusion, preoperative plasma suPAR level was of no prognostic value in this cohort of Danish stage III OC patients. PMID:15274388

Begum, Farah Diba; Høgdall, Claus K; Kjaer, Susanne K; Christensen, Lise; Blaakaer, Jan; Bock, Johannes E; Glud, Eva; Høyer-Hansen, Gunilla; Ring-Larsen, Helmer; Høgdall, Estrid V S

2004-01-01

305

The human urokinase-plasminogen activator gene (PLAU) is located on chromosome 10q24 centromeric to the HOX11 gene  

Energy Technology Data Exchange (ETDEWEB)

Urokinase-plasminogen activator is one of two soluble serine proteases that are produced by humans and that convert plasminogen, an inactive proenzyme present in plasma and other extracellular fluids, to plasmin, a protease with broad substrate specificities. Its activity is involved in processes requiring localized extracellular proteolysis such as fibrinolysis, tissue remodeling, and cell migration. Increased production of urokinase has been associated with cancer metastases. The gene for urokinase-plasminogen activator, PLAU, was mapped to chromosome 10q24-qter. By employing somatic cell genetics, polymerase chain reaction (PCR), and Southern blot analysis, the authors assign PLAU to chromosome 10q24. Human chromosome segment 10q23-q25 contains the genes for terminal deoxynucleotidyltransferase, cytochrome P450IIC, glutamic-oxaloacetic transaminase, and plasma retinol binding protein, which form a syntenic group on murine chromosome 19. It is therfore of interest that PLAU and glutamate dehydrogenase, which are on murine chromosome 14, also map in or close to this region of human chromosome 10.

Stein, P.M.; Stass, S.A.; Kagan, J. (Univ. of Texas, Houston (United States))

1993-04-01

306

Interaction of surface molecules on Cryptococcus neoformans with plasminogen.  

Science.gov (United States)

Microbial pathogens are known to express molecules that interact with host proteins, leading to invasion and colonization. For example, some pathogenic microorganisms express proteins that bind to and enhance the activity of plasminogen. In this way, pathogens utilize the host fibrinolytic system to promote invasion. We found that triosephosphate isomerase (TPI), a glycolytic enzyme produced by Staphylococcus aureus, bound to mannooligosaccharides from the pathogenic capsulated fungus Cryptococcus neoformans and human plasminogen, suggesting that TPI is a moonlighting protein. Several C. neoformans surface proteins are thought to be plasminogen-binding proteins. Here, we examined the ability of surface polymers (including polysaccharides) to bind plasminogen. Heat-killed C. neoformans cells transformed plasminogen into plasmin in a dose-dependent manner in the presence of tissue plasminogen activator. Soluble polysaccharides were found to bind plasminogen based on surface plasmon resonance (SPR) analysis. Neutral polysaccharides fractionated using DEAE column chromatography bound and activated plasminogen. However, the fraction containing glucuronoxylomannan (the primary component of the capsule) did not activate plasminogen. In addition, binding between glucuronoxylomannan and plasminogen was weak. Components of the neutral polysaccharides were identified as mannose, galactose, glucose and xylose. In conclusion, neutral polysaccharides that may affect fibrinolysis were detected on the surface of C. neoformans. PMID:24373348

Ikeda, Reiko; Ichikawa, Tomoe

2014-05-01

307

Tc-99m labeled tissue-type plasminogen activator; Preparation, stability and preliminary imaging of thrombus-bearing rats  

Energy Technology Data Exchange (ETDEWEB)

Tissue-type plasminogen activator (t-PA) is a thrombolytic agent that directly binds to fibrin formed in clots. In terms of radiolabeling and nuclear imaging, t-PA has several advantages in Tc-99m labeling: it is stable in acidic solution at pH 3, which is suitable for labeling Tc-99m by a method of stannous reduction and blood disappearance after administration is rapid, which is desirable for imaging targets using short-lived radionuclides. Recombinant t-PA was labeled with Tc-99m by a method of stannous reduction without significant degradation of biochemical activity, over 95% of which was retained after the labeling procedure. Labeling efficiency in paper chromatography was over 98%. The moiety of hydrolyzed Tc-99m that was not eluted through the Sephadex column was estimated to be less than 10%. Tc-99m labeled t-PA, however, appeared to become unstable when diluted with normal saline. Nevertheless, in in vitro fibrin binding, Tc-99m labeled t-PA showed high affinity with fibrin: 80% of 100 ng/ml of Tc-99m t-PA bound to 10{sup -5} mol of the fibrinogen. Preliminary animal studies also showed a concentration of Tc-99m labeled t-PA at fresh thrombi formed in the inferior vena cava. Tc-99m labeled t-PA appears to have potential for thrombus imaging and the preparation of an instant kit. (author).

Itoh, Kazuo; Tsukamoto, Eriko; Nishibe, Toshiya; Sakurama, Shoki; Ieko, Masahiro; Tanabe, Tatsuzo; Furudate, Masayori (Hokkaido Univ., Sapporo (Japan). School of Medicine)

1991-07-01

308

Construction of thrombus-targeted microbubbles carrying tissue plasminogen activator and their in vitro thrombolysis efficacy: a primary research.  

Science.gov (United States)

Thrombosis is the common mechanism of various diseases of heart and vasculature and their major morbility and mortality. An efficient, safe and easy thrombolysis method is needed. We tried to develop a new type of ultrasound microbubbles carrying thrombolytics and simultaneously targeting to thrombus, which could bind with thrombus specifically and release the encapsulated drug locally under the ultrasound exposure. Microbubbles carrying tissue plasminogen activator (tPA) and Arg-Gly-Asp-Ser tetrapeptide (RGDS) were prepared by lyopyilization. Their properties were detected, including morphology, particle size, surface potential and pH. The results showed that the microbubbles were suitable for intravenous injection. The envelope rate of tPA, detected by ELISA, was (81.12 +/- 2.44%), and the conjugate rate of RGDS, detected by flow cytometer, was (94.49 +/- 6.19%). The tPA encapsulated in microbubbles kept fibrinolysis activity under the conditions of both natural releasing and ultrasound exposure, checked by agarose fibrin plate process. The contrast-enhanced ultrasonography (CEU) in rabbit liver showed that they were good for enhanced ultrasound imaging. The in vitro thrombolysis of the microbubbles to the blood clots from healthy human was detected with a mimical flowing model propelled by peristaltic pump. The drug-loaded microbubbles plus ultrasound irradiation got higher thrombolysis with the lowest dosage. The tPA-loaded microbubbles targeting to thrombus can be prepared by lyopyilization, which will bring out a novel way for the targeting drug-released thrombolysis therapy. PMID:20155435

Hua, Xing; Liu, Ping; Gao, Yun-Hua; Tan, Kai-Bin; Zhou, Li-Na; Liu, Zheng; Li, Xin; Zhou, Shi-Wen; Gao, Yue-Juan

2010-07-01

309

Tissue-type plasminogen activator-induced fibrinolysis is enhanced in patients with breast, lung, pancreas and colon cancer.  

Science.gov (United States)

Although cancer-mediated changes in hemostatic proteins unquestionably promote hypercoagulation, the effects of neoplasia on fibrinolysis in the circulation are less well defined. The goals of the present investigation were to determine if plasma obtained from patients with breast, lung, pancreas and colon cancer was less or more susceptible to lysis by tissue-type plasminogen activator (tPA) compared to plasma obtained from normal individuals. Archived plasma obtained from patients with breast (n?=?18), colon/pancreas (n?=?27) or lung (n?=?19) was compared to normal individual plasma (n?=?30) using a thrombelastographic assay that assessed fibrinolytic vulnerability to exogenously added tPA. Plasma samples were activated with tissue factor/celite, had tPA added, and had data collected until clot lysis occurred. Additional, similar samples had potato carboxypeptidase inhibitor added to assess the role played by thrombin-activatable fibrinolysis inhibitor in cancer-modulated fibrinolysis. Rather than inflicting a hypofibrinolytic state, the three groups of cancers demonstrated increased vulnerability to tPA (e.g. decreased time to lysis, increased speed of lysis, decreased clot lysis time). However, hypercoagulation manifested as increased speed of clot formation and strength compensated for enhanced fibrinolytic vulnerability, resulting in a clot residence time that was not different from normal individual thrombi. In sum, enhanced hypercoagulability associated with cancer was in part diminished by enhanced fibrinolytic vulnerability to tPA. PMID:24674880

Nielsen, Vance G; Matika, Ryan W; Ley, Michele L B; Waer, Amy L; Gharagozloo, Farid; Kim, Samuel; Nfonsam, Valentine N; Ong, Evan S; Jie, Tun; Warneke, James A; Steinbrenner, Evangelina B

2014-04-01

310

Respiratory burst function of ovine neutrophils  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Respiratory burst function resulting in the release of reactive oxygen species such as superoxide anion (O2- from neutrophils is one of the key mechanisms of the innate immune system, and maladaptive control of this mechanism is thought to play a pivotal role in the development of pathologies such as acute lung injury and sepsis. Ovine models of these pathologies are limited by the poor understanding of ovine neutrophil respiratory burst function. Results Aspects of ovine neutrophil respiratory burst function to be characterised were: i the maximum rate of O2- generated (Vmax; ii the time taken to reach Vmax; iii the total amount of O2- generated during the reaction; and iv the duration of the reaction. As well as for unstimulated neutrophils, these aspects were also characterised after incubation with a priming agonist (platelet activating factor [PAF], tumour necrosis factor alpha [TNF-?] and lipopolysaccharides [LPS] activating agonists (N-formylmethionyl-leucyl-phenylalanine [fMLP] and phorbol 12-myristate 13-acetate [PMA] or a combination of a priming and an activating agonist. In the absence of priming or activating agonists, ovine neutrophils displayed a low level of respiratory burst function which was not enhanced by either PAF, TNF-?, LPS or fMLP, but was significantly enhanced by PMA. The PMA-induced respiratory burst function was further enhanced by pre-incubation with PAF, but not with TNF-? or LPS. By varying the length of pre-incubation with PAF it was demonstrated that this effect decreased as the duration of pre-incubation with PAF increased, and that PAF was enhancing PMA's effects rather than PMA enhancing PAF's effects. Conclusion This study successfully adapted a commonly used method of measuring human neutrophil respiratory burst function to characterise different aspects of ovine neutrophil respiratory burst function. This improved understanding of ovine neutrophils will facilitate the validitation of ovine biomedical models of human pathologies in which neutrophils have been implicated.

Fraser John F

2009-05-01

311

Elevated plasma plasminogen activator inhibitor type-1 is an independent predictor of coronary microvascular dysfunction in hypertension  

International Nuclear Information System (INIS)

Elevated plasma plasminogen activator inhibitor-1 (PAI-1) is related to cardiovascular events, but its role in subclinical coronary microvascular dysfunction remains unknown. Thus, in the present study it was investigated whether elevated plasma PAI-1 activity is associated with coronary microvascular dysfunction in hypertensive patients. Thirty patients with untreated essential hypertension and 10 age-matched healthy controls were studied prospectively. Myocardial blood flow (MBF) was measured by using 15O-water positron emission tomography. Clinical variables associated with atherosclerosis (low-density lipoprotein-cholesterol, high-density lipoprotein (HDL)-cholesterol, triglyceride, homeostasis model assessment (HOMA-IR), and PAI-1 activity) were assessed to determine their involvement in coronary microvascular dysfunction. Adenosine triphosphate (ATP)-induced hyperemic MBF and coronary flow reserve (CFR) were significantly lower in hypertensive patients than in healthy controls (ATP-induced MBF: 2.77±0.82 vs 3.49±0.71 ml·g-1·min-1; p<0.02 and CFR: 2.95±1.06 vs 4.25±0.69; p<0.001). By univariate analysis, CFR was positively correlated with HDL-cholesterol (r=0.46, p<0.02), and inversely with HOMA-IR (r=-0.39, p<0.05) and PAI-1 activity (r=-0.61, p<0.001). By multivariate analysis, elevated PAI-1 activity remained a significant independent determinant of diminished CFR. Elevated plasma PAI-1 activity was independently associated with coronary microvascular dysfunction, which suggests that plasma PAI-1 activity is an important clue linking hypofibrinolysis to the development of atherosclerosis. (author)

2007-03-01

312

sup 1 H NMR structural characterization of a recombinant kringle 2 domain from human tissue-type plasminogen activator  

Energy Technology Data Exchange (ETDEWEB)

The kringle 2 domain of human tissue-type plasminogen activator (t-PA) has been characterized via {sup 1}H NMR spectroscopy at 300 and 620 MHz. The experiments were performed on the isolated domain obtained by expression of the 174-263 portion of t-PA in Escherichia coli. The spectrum of t-Pa kringle 2 is characteristic of a globular structure and shows overall similarity to that of the plasminogen (PGN) kringle 4. Spectral comparison with human and bovine PGN kringle 4 identified side-chain resonances from Leu{sup 46}, which afford a fingerprint of kringle folding, and from most of the aromatic ring spin systems. Ligand-binding studies confirm that t-PA kringle 2 binds L-lysine with an association constant K{sub a} {approximately} 11.9 mM{sup {minus}1}. The data indicate that homologous or conserved residues relative to those that compose the lysine-binding sites of PGN kringles 1 and 4 are involved in the binding of L-lysine to t-PA kringle 2. These include Tyr{sup 36} and, within the kringle inner loop, Trp{sup 62}, His{sup 64}, Trp{sup 72}, and Tyr{sup 74}. Several labile NH protons of t-PA kringle 2 exhibit retarded H-exchange kinetics, requiring more than a week in {sup 2}H{sub 2}O for full deuteration in the presence of L-lysine at 37{degree}C. This reveals that kringle 2 is endowed with a compact, dynamically stable conformation. Proton Overhauser experiments in {sup 1}H{sub 2}O, centered on well-resolved NH resonances between 9.8 and 12 ppm, identify signals arising from the His{sup 48a} imidazole NH3 proton and the three Trp indole NH1 protons. Overall, the data indicate a highly structured conformation for the recombinant t-PA kringle 2 that is closely related to that of the previously investigated PGN kringles 1, 4, and 5.

Byeon, I.J.L.; Llinas, M. (Carnegie Mellon Univ., Pittsburgh, PA (USA)); Kelley, R.F. (Genentech, Inc., South San Francisco, CA (USA))

1989-11-28

313

Elevated plasma levels of vascular endothelial growth factor and plasminogen activator inhibitor-1 decrease during improvement of psoriasis.  

DEFF Research Database (Denmark)

OBJECTIVE AND DESIGN: An evaluation of angiogenesis related molecules during open treatment of psoriasis. MATERIALS AND SUBJECTS: Plasma samples and skin biopsies from 16 patients with psoriasis and plasma samples from 13 healthy controls. TREATMENT: Ranitidine 300 mg orally twice daily for 6 months. METHODS: Vascular endothelial growth factor (VEGF) and plasminogen activator inhibitor-1 (PAI-1) were determined by ELISA methods in plasma collected from the patients before treatment and after 1, 3 and 6 months. Vessel counts were performed in biopsies from affected skin areas taken before treatment and after 3 and 6 months. The results were compared to simultaneous PASI scores. RESULTS: Pre-treatment plasma levels of VEGF and PAI-1 were significantly elevated in patients compared with levels in healthy persons (p = 0.02 and p = 0.04, respectively). The plasma levels decreased significantly during treatment (p = 0.03 and p = 0.01, respectively), and the decrease in combined levels correlated with the decrease in PASI score. However, the vessel density in affected skin did not change during treatment. CONCLUSIONS: Increased pre-treatment levels of VEGF and PAI-1 and decrease during improvement of the disease suggest that the two molecules may play a role in pathogenesis of psoriasis.

Nielsen, Hans Jørgen; Christensen, Ib Jarle

2002-01-01

314

The liberated domain I of urokinase plasminogen activator receptor--a new tumour marker in small cell lung cancer  

DEFF Research Database (Denmark)

The prognosis of small cell lung cancer (SCLC) remains poor with a 5-year survival rate of 4-6%. In non-small cell lung cancer (NSCLC), high levels of intact and cleaved forms of the receptor for urokinase plasminogen activator (uPAR) are significantly associated with short overall survival. Our aim was therefore to determine the prognostic value of the different uPAR forms in blood from SCLC patients. Serum samples from 92 treatment naive SCLC patients were analysed. Intact uPAR, uPAR(I-III), intact and cleaved uPAR, uPAR(I-III) + uPAR(II-III) and the liberated domain I, uPAR(I) were measured using time-resolved fluorescence immunoassays (TR-FIA 1-3). Assessment of association of the uPAR forms to overall survival (OS) was done using Cox regression analysis adjusted for clinical covariates [age, gender, stage, lactate dehydrogenase (LDH), WHO performance status (PS)]. Multivariate survival analysis demonstrated that high levels of uPAR(I) were significantly (p = 0.009) associated with short overall survival (OS). Patients with uPAR(I) levels above the second tertile had a hazard ratio (HR) of 1.9 (95% confidence interval (CI): 1.1-3.3), compared to patients with levels below the first tertile. High serum uPAR(I) levels are associated with short OS in SCLC patient, independent of LDH and PS.

Almasi, Charlotte E; Drivsholm, Lars

2013-01-01

315

Current perspectives on the use of intravenous recombinant tissue plasminogen activator (tPA) for treatment of acute ischemic stroke.  

Science.gov (United States)

In 1995, the NINDS (National Institute of Neurological Disorders and Stroke) tPA (tissue plasminogen activator) Stroke Study Group published the results of a large multicenter clinical trial demonstrating efficacy of intravenous tPA by revealing a 30% relative risk reduction (absolute risk reduction 11%-15%) compared with placebo at 90 days in the likelihood of having minimal or no disability. Since approval in 1996, tPA remains the only drug treatment for acute ischemic stroke approved by the US Food and Drug Administration. Over the years, an abundance of research and clinical data has supported the safe and efficacious use of intravenous tPA in all eligible patients. Despite such supporting data, it remains substantially underutilized. Challenges to the utilization of tPA include narrow eligibility and treatment windows, risk of symptomatic intracerebral hemorrhage, perceived lack of efficacy in certain high-risk subgroups, and a limited pool of neurological and stroke expertise in the community. With recent US census data suggesting annual stroke incidence will more than double by 2050, better education and consensus among both the medical and lay public are necessary to optimize the use of tPA for all eligible stroke patients. Ongoing and future research should continue to improve upon the efficacy of tPA through more rapid stroke diagnosis and treatment, refinement of advanced neuroimaging and stroke biomarkers, and successful demonstration of alternative means of reperfusion. PMID:24591838

Chapman, Sherita N; Mehndiratta, Prachi; Johansen, Michelle C; McMurry, Timothy L; Johnston, Karen C; Southerland, Andrew M

2014-01-01

316

A novel mRNA binding protein complex promotes localized plasminogen activator inhibitor-1 accumulation at the myoendothelial junction  

Science.gov (United States)

Objective Plasminogen activator inhibitor-1 (PAI-1) has previously been shown to be key to the formation of myoendothelial junctions (MEJs) in normal and pathological states (e.g., obesity). We there for sought to identify the mechanism whereby PAI-1 could be selectively accumulated at the MEJ. Methods and Results We identified PAI-1 protein enrichment at the MEJ in obese mice and in response to TNF-? with a vascular cell co-culture. However, PAI-1 mRNA was also found at the MEJ and transfection with a PAI-1-GFP with TNF-? did not demonstrate trafficking of the protein to the MEJ. We there for hypothesized the PAI-1 mRNA was being locally translated and identified serpine binding protein-1, which stabilizes PAI-1 mRNA, as being enriched in obese mice and after treatment with TNF-?, while staufen, which degrades PAI-1 mRNA, was absent in obese mice and after TNF- ? application. We identified nicotinamide phosphoribosyl transferase as a serpine binding protein-1 binding partner with a functional tau-like microtubule binding domain. Application of peptides against the microtubule binding domain significantly decreased the number of MEJs and the amount of PAI-1 at the MEJ. Conclusions We conclude that PAI-1 can be locally translated at the MEJ due to a unique mRNA binding protein complex.

Heberlein, Katherine R.; Han, Jenny; Straub, Adam C.; Best, Angela K.; Kaun, Christoph; Wojta, Johann; Isakson, Brant E.

2012-01-01

317

Multifunctional roles of urokinase plasminogen activator (uPA) in cancer stemness and chemoresistance of pancreatic cancer.  

Science.gov (United States)

Pancreatic ductal adenocarcinoma (PDAC) is almost always lethal. One of the underlying reasons for this lethality is believed to be the presence of cancer stem cells (CSC), which impart chemoresistance and promote recurrence, but the mechanisms responsible are unclear. Recently the poor prognosis of PDAC has been correlated with increased expression of urokinase plasminogen activator (uPA). In the present study we examine the role of uPA in the generation of PDAC CSC. We observe a subset of cells identifiable as a side population (SP) when sorted by flow cytometry of MIA PaCa-2 and PANC-1 pancreatic cancer cells that possess the properties of CSC. A large fraction of these SP cells are CD44 and CD24 positive, are gemcitabine resistant, possess sphere-forming ability, and exhibit increased tumorigenicity, known characteristics of cancer stemness. Increased tumorigenicity and gemcitabine resistance decrease after suppression of uPA. We observe that uPA interacts directly with transcription factors LIM homeobox-2 (Lhx2), homeobox transcription factor A5 (HOXA5), and Hey to possibly promote cancer stemness. uPA regulates Lhx2 expression by suppressing expression of miR-124 and p53 expression by repressing its promoter by inactivating HOXA5. These results demonstrate that regulation of gene transcription by uPA contributes to cancer stemness and clinical lethality. PMID:23864708

Asuthkar, Swapna; Stepanova, Victoria; Lebedeva, Tatiana; Holterman, Aixuan L; Estes, Norman; Cines, Douglas B; Rao, Jasti S; Gondi, Christopher S

2013-09-01

318

Characterization of a helicase-like transcription factor involved in the expression of the human plasminogen activator inhibitor-1 gene.  

Science.gov (United States)

A 5.4-kb cDNA encoding the protein that binds to the B Box of the plasminogen activator inhibitor-1 (PAI-1) gene was isolated and sequenced. The protein, named helicase-like transcription factor (HLTF), contains a DNA-binding domain, a RING finger domain, and seven helicase domains and is homologous to SWI/SNF proteins. Two HLTF mRNAs of 5.5 and 4.5 kb were detected in most human tissues, a single gene was located on chromosome 3q24-25, and the protein was located in the nucleoplasm. Two HLTF proteins differing in translation start site (Met-1 or Met-123) were obtained by in vitro translation in reticulocyte lysate or by immunoprecipitation from HeLa cell nuclear extracts. In vitro transcription from the PAI-1 promoter in HeLa cell extracts was inhibited by HLTF antibodies and by the HLTF DNA binding domain. Over-expression of HLTF or HLTFMet123 produced a three-fold induction of PAI-1-LUC transient expression in HeLa cells. Mutation of the PAI-1 B Box led to an eight-fold reduction of basal PAI-1-LUC expression in these cell lines, but did not affect the four- to six-fold induction by phorbol esters. PMID:8672239

Ding, H; Descheemaeker, K; Marynen, P; Nelles, L; Carvalho, T; Carmo-Fonseca, M; Collen, D; Belayew, A

1996-06-01

319

Intracisternal administration of tissue plasminogen activator improves cerebrospinal fluid flow and cortical perfusion after subarachnoid hemorrhage in mice.  

Science.gov (United States)

Early brain injury (EBI) during the first 72 h after subarachnoid hemorrhage (SAH) is an important determinant of clinical outcome. A hallmark of EBI, global cerebral ischemia, occurs within seconds of SAH and is thought to be related to increased intracranial pressure (ICP). We tested the hypothesis that ICP elevation and cortical hypoperfusion are the result of physical blockade of cerebrospinal fluid (CSF) flow pathways by cisternal microthrombi. In mice subjected to SAH, we measured cortical blood volume (CBV) using optical imaging, ICP using pressure transducers, and patency of CSF flow pathways using intracisternally injected tracer dye. We then assessed the effects of intracisternal injection of recombinant tissue plasminogen activator (tPA). ICP rose immediately after SAH and remained elevated for 24 h. This was accompanied by a decrease in CBV and impaired dye movement. Intracisternal administration of tPA immediately after SAH lowered ICP, increased CBV, and partially restored CSF flow at 24 h after SAH. Lowering ICP without tPA, by draining CSF, improved CBV at 1 h, but not 24 h after SAH. These findings suggest that blockade of CSF flow by microthrombi contributes to the early decline in cortical perfusion in an ICP-dependent and ICP-independent manner and that intracisternal tPA may reduce EBI and improve outcome after SAH. PMID:24526376

Siler, Dominic A; Gonzalez, Jorge A; Wang, Ruikang K; Cetas, Justin S; Alkayed, Nabil J

2014-04-01

320

The alteration of plasminogen activator inhibitor-1 expression by linoleic acid and fenofibrate in HepG2 cells.  

Science.gov (United States)

The present study investigated the influence of linoleic acid and fenofibrate on plasminogen activator inhibitor-1 (PAI-1) expression in HepG2 cells and the mechanism possibly involved. Using gene recombination techniques, chloromycetin acetyltransferase (CAT) reporter gene plasmids containing nuclear factor-kappaB response element deletion (del1-PAI-pCAT) or very-low-density lipoprotein/fatty acid response element deletion (del2-PAI-pCAT) in the PAI-1 promoter were constructed and transiently transfected into HepG2 cells, respectively. Linoleic acid and fenofibrate were added to induce the transfected cells. The PAI-1 expression in mRNA and protein level was significantly induced by linoleic acid, but suppressed by fenofibrate. In the HepG2 cells transfected with PAI-pCAT plasmid, the PAI-1 transcription activity was significantly induced by linoleic acid, but suppressed by fenofibrate. Under transfection with del1-PAI-pCAT, both linoleic acid and fenofibrate increased the PAI-1 transcriptional activity; whereas in those cells transfected with del2-PAI-pCAT, fenofibrate significantly reduced PAI-1 transcriptional activity but no change was found with linoleic acid stimulation. Peroxisome proliferator-activated receptor alpha may be one of transcription factors playing a role in the upregulation of PAI-1 gene expression by linoleic acid in HepG2 cells. The inhibition of the nuclear factor-kappaB signaling pathway may be involved in the downregulation of PAI-1 gene expression by fenofibrate. PMID:17179821

Ye, Ping; He, Yan-Li; Wang, Qiong; Liu, Yong-Xue

2007-01-01

 
 
 
 
321

Plasma carboxypeptidases as regulators of the plasminogen system.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Carboxy-terminal lysine residues on the surface of cells and fibrin bind plasminogen and control its activation. Since plasma contains basic carboxypeptidases, which remove carboxy-terminal lysines from protein substrates, we investigated if these enzymes are involved in the regulation of plasminogen binding sites. Plasma reduced plasminogen binding to cells, and this effect could be ascribed to the activity of the plasma carboxypeptidases. Purified carboxypeptidase N, which is constitutively...

Redlitz, A.; Tan, A. K.; Eaton, D. L.; Plow, E. F.

1995-01-01

322

Bacterial endotoxin enhances colorectal cancer cell adhesion and invasion through TLR-4 and NF-kappaB-dependent activation of the urokinase plasminogen activator system.  

LENUS (Irish Health Repository)

Perioperative exposure to lipopolysaccharide (LPS) is associated with accelerated metastatic colorectal tumour growth. LPS directly affects cells through Toll-like receptor 4 (TLR-4) and the transcription factor NF-kappaB. The urokinase plasminogen activator (u-PA) system is intimately implicated in tumour cell extracellular matrix (ECM) interactions fundamental to tumour progression. Thus we sought to determine if LPS directly induces accelerated tumour cell ECM adhesion and invasion through activation of the u-PA system and to elucidate the cellular pathways involved. Human colorectal tumour cell lines were stimulated with LPS. u-PA concentration, u-PA activity, active u-PA, surface urokinase plasminogen activator receptor (u-PAR) and TLR-4 expression were assessed by ELISA, colorimetric assay, western blot analysis and flow cytometry respectively. In vitro tumour cell vitronectin adhesion and ECM invasion were analysed by vitronectin adhesion assay and ECM invasion chambers. u-PA and u-PAR function was inhibited with anti u-PA antibodies or the selective u-PA inhibitors amiloride or WXC-340, TLR-4 by TLR-4-blocking antibodies and NF-kappaB by the selective NF-kappaB inhibitor SN-50. LPS upregulates u-PA and u-PAR in a dose-dependent manner, enhancing in vitro tumour cell vitronectin adhesion and ECM invasion by >40% (P<0.01). These effects were ameliorated by u-PA and u-PAR inhibition. LPS activates NF-kappaB through TLR-4. TLR-4 and NF-kappaB inhibition ameliorated LPS-enhanced u-PA and u-PAR expression, tumour cell vitronectin adhesion and ECM invasion. LPS promotes tumour cell ECM adhesion and invasion through activation of the u-PA system in a TLR-4- and NF-kappaB-dependent manner.

Killeen, S D

2009-05-19

323

Expression of protease nexin-1 and plasminogen activators during follicular growth and the periovulatory period in cattle.  

Science.gov (United States)

Extracellular matrix remodeling occurs during ovarian follicular development, mediated by plasminogen activators (PAs) and PA inhibitors including protease nexin-1 (PN-1). In the present study we measured expression/activity of the PA system in bovine follicles at different stages of development by timed collection of ovaries during the first follicular wave and during the periovulatory period, and in follicles collected from an abattoir. The abundance of mRNA encoding PN-1, tissue-type PA (tPA), urokinase (uPA) and PA inhibitor-1 (PAI-1) were initially upregulated by human chorionic gonadotropin (hCG) in bovine preovulatory follicular wall homogenates. PN-1, PAI-1 and tPA mRNA expression then decreased near the expected time of ovulation, whereas uPA mRNA levels remained high. PN-1 concentration in follicular fluid (FF) decreased and reached the lowest level at the time of ovulation, whereas plasmin activity in FF increased significantly after hCG. Follicles collected from the abattoir were classified as non-atretic, early-atretic or atretic based on FF estradiol and progesterone content: PN-1 protein levels in FF were significantly higher in non-atretic than in atretic follicles, and plasmin activity was correspondingly higher in the atretic follicles. No changes in PN-1 levels in FF were observed during the growth of pre-deviation follicles early in a follicular wave. These results indicate that PN-1 may be involved in the process of atresia in non-ovulatory dominant follicles and the prevention of precocious proteolysis in periovulatory follicles. PMID:16388016

Cao, Mingju; Buratini, José; Lussier, Jacques G; Carrière, Paul D; Price, Christopher A

2006-01-01

324

The angiotensin-converting enzyme inhibitor captopril inhibits poly(ADP-ribose)polymerase activation and exerts beneficial effects in an ovine model of burn and smoke injury  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We investigated the effect of the angiotensin converting enzyme (ACE) inhibitor captopril in a clinically relevant ovine model of smoke and burn injury, with special reference to oxidative stress, activation of poly(ADP-ribose) polymerase in the lung and in circulating leukocytes. Female, adult sheep (28–40 kg) were divided into 3 groups. After tracheostomy and under deep anesthesia both vehicle-control (n=5) and captopril (20 mg/kg/d, iv., starting 0.5 hour before the injury) treated (n=5)...

Asmussen, Sven; Bartha, Eva; Olah, Gabor; Sbrana, Elena; Rehberg, Sebastian W.; Yamamoto, Yusuke; Enkhbaatar, Perenlei; Hawkins, Hal K.; Ito, Hiroshi; Cox, Robert A.; Traber, Lillian D.; Traber, Daniel L.; Szabo, Csaba

2011-01-01

325

Plasminogen activator inhibitor-1 deficiency ameliorates insulin resistance and hyperlipidemia but not bone loss in obese female mice.  

Science.gov (United States)

We previously demonstrated that plasminogen activator inhibitor-1 (PAI-1), an inhibitor of fibrinolysis, is involved in type 1 diabetic bone loss in female mice. PAI-1 is well known as an adipogenic factor induced by obesity. We therefore examined the effects of PAI-1 deficiency on bone and glucose and lipid metabolism in high-fat and high-sucrose diet (HF/HSD)-induced obese female mice. Female wild-type (WT) and PAI-1-deficient mice were fed with HF/HSD or normal diet for 20 weeks from 10 weeks of age. HF/HSD increased the levels of plasma PAI-1 in WT mice. PAI-1 deficiency suppressed the levels of blood glucose, plasma insulin, and total cholesterol elevated by obesity. Moreover, PAI-1 deficiency improved glucose intolerance and insulin resistance induced by obesity. Bone mineral density (BMD) at trabecular bone as well as the levels of osterix, alkaline phosphatase, and receptor activator of nuclear factor ?B ligand mRNA in tibia were decreased by HF/HSD in WT mice, and those changes by HF/HSD were not affected by PAI-1 deficiency. HF/HSD increased the levels of plasma TNF-? in both WT and PAI-1-deficient mice, and the levels of plasma TNF-? were negatively correlated with trabecular BMD in tibia of female mice. In conclusion, we revealed that PAI-1 deficiency does not affect the trabecular bone loss induced by obesity despite the amelioration of insulin resistance and hyperlipidemia in female mice. Our data suggest that the changes of BMD and bone metabolism by obesity might be independent of PAI-1 as well as glucose and lipid metabolism. PMID:24605827

Tamura, Yukinori; Kawao, Naoyuki; Yano, Masato; Okada, Kiyotaka; Matsuo, Osamu; Kaji, Hiroshi

2014-05-01

326

Role of LFB3 in cell-specific cAMP induction of the urokinase-type plasminogen activator gene.  

Science.gov (United States)

In previous work we suggested that a kidney-specific transcription factor LFB3 cooperates with cAMP-response element (CRE)-binding proteins within a cAMP regulatory unit comprised of three protein-binding domains and located 3.4 kilobase pairs upstream of the urokinase-type plasminogen activator (uPA) gene in LLC-PK1 cells (Menoud, P.-A., Matthies, R., Hofsteenge, J., and Nagamine, Y. (1993) Nucleic Acids Res. 21, 1845-1852). The two domains contain a CRE-like sequence, and the third domain is recognized by LFB3. The absolute requirement of LFB3 as well as the cooperation among the three domains for cAMP regulation were confirmed by transient transfection assays in F9 teratocarcinoma cells, in which the level of LFB3 was negligible. Suspecting a possible feedback regulation of LFB3 mRNA expression during cAMP-dependent uPA gene induction in LLC-PK1 cells, we measured LFB3 mRNA levels after cAMP treatment and found a strong reduction. This reduction was not due to a change in template activity of the LFB3 gene because run-on transcription showed no significant change in LFB3 gene transcription. RNA synthesis inhibitor-chase experiments indicated that the down-regulation was post-transcriptional. Interestingly, when the inhibitor was added at the same time as cAMP, the cAMP-induced decrease in LFB3 mRNA levels was abrogated, suggesting that ongoing RNA synthesis is required for the decrease. Similar effects on LFB3 mRNA metabolism were observed with all agents that induce uPA mRNA in LLC-PK1 cells, including 12-O-tetradecanoylphorbol-13-acetate, okadaic acid, colchicine, and cytochalasin. We discuss the significance of this regulation in uPA gene expression. PMID:7665606

Marksitzer, R; Stief, A; Menoud, P A; Nagamine, Y

1995-09-15

327

Hypoxia dysregulates the production of adiponectin and plasminogen activator inhibitor-1 independent of reactive oxygen species in adipocytes  

International Nuclear Information System (INIS)

Low plasma levels of adiponectin (hypoadiponectinemia) and elevated circulating concentrations of plasminogen activator inhibitor (PAI)-1 are causally associated with obesity-related insulin resistance and cardiovascular disease. However, the mechanism that mediates the aberrant production of these two adipokines in obesity remains poorly understood. In this study, we investigated the effects of hypoxia and reactive oxygen species (ROS) on production of adiponectin and PAI-1 in 3T3-L1 adipocytes. Quantitative PCR and immunoassays showed that ambient hypoxia markedly suppressed adiponectin mRNA expression and its protein secretion, and increased PAI-1 production in mature adipocytes. Dimethyloxallyl glycine, a stabilizer of hypoxia-inducible factor 1? (HIF-1?), mimicked the hypoxia-mediated modulations of these two adipokines. Hypoxia caused a modest elevation of ROS in adipocytes. However, ablation of intracellular ROS by antioxidants failed to alleviate hypoxia-induced aberrant production of adiponectin and PAI-1. On the other hand, the antioxidants could reverse hydrogen peroxide (H2O2)-induced dysregulation of adiponectin and PAI-1 production. H2O2 treatment decreased the expression levels of peroxisome proliferator-activated receptor gamma (PPAR?) and CCAAT/enhancer binding protein (C/EBP?), but had no effect on HIF-1?, whereas hypoxia stabilized HIF-1? and decreased expression of C/EBP?, but not PPAR?. Taken together, these data suggest that hypoxia and ROS decrease adiponectin production and augment PAI-1 expression in adipocytes via distinct signaling pathways. These effects may contribute to hypoadiponectinemia and elevated PAI-1 levels in obesity, type 2 diabetes, and cardiovascular diseases

2006-03-10

328

Production of Desmodus rotundus salivary plasminogen activator alpha1 (DSPAalpha1) in tobacco is hampered by proteolysis.  

Science.gov (United States)

The high fibrin specificity of Desmodus rotundus salivary plasminogen activator alpha1 (DSPAalpha1 or desmoteplase (INN)) makes it a promising candidate for the treatment of acute ischemic stroke. In the current study we explored the use of transgenic tobacco plants and BY-2 suspension cells as alternative production platforms for this drug. Four different N-terminal signal peptides, from plants and animals, were used to translocate the recombinant DSPAalpha1 protein to the endomembrane system. Intact recombinant DSPAalpha1 was produced in transgenic plants and BY-2 cells, although a certain degree of degradation was observed in immunoblotted extracts. The choice of signal peptide had no major influence on the degradation pattern or recombinant protein accumulation, which reached a maximum level of 38 microg/g leaf material. N-terminal sequencing of purified, His6-tagged DSPAalpha1 revealed only minor changes in the position of signal peptide cleavage compared to the same protein expressed in Chinese hamster ovary cells. However, correctly processed recombinant DSPAalpha1 was also detected. The enzymatic activity of the recombinant protein was confirmed using an in vitro assay with unpurified and purified samples, demonstrating that plants are suitable for the production of functional DSPAalpha1. In contrast to whole plant cell extracts, no recombinant DSPAalpha1 was detected in the culture supernatant of transgenic BY-2 cells. Further analysis showed that recombinant DSPAalpha1 is subject to proteolysis and that endogenous secreted BY-2 proteases are responsible for DSPAalpha1 degradation in the culture medium. The addition of a highly concentrated protease inhibitor mixture or 5 mM EDTA reduced DSPAalpha1 proteolysis, improving the accumulation of intact product in the culture medium. Strategies to improve the plant cell suspension system for the production of secreted recombinant proteins are discussed. PMID:15685597

Schiermeyer, Andreas; Schinkel, Helga; Apel, Stefanie; Fischer, Rainer; Schillberg, Stefan

2005-03-30

329

SERPINE2, an inhibitor of plasminogen activators, is highly expressed in the human endometrium during the secretory phase  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background SERPINE2, also known as protease nexin-1, belongs to the serine protease inhibitor (SERPIN superfamily. It is one of the potent SERPINs that modulates the activity of plasminogen activators (PAs. PAs and their SERPIN inhibitors, such as SERPINB2 and SERPINE1, were expressed in the human endometrium and were implicated in implantation. However, expression data about SERPINE2 in the human endometrium is still unknown. Thus, we conducted an investigation to reveal the spatiotemporal and cellular expression of SERPINE2 in the human uterus during the menstrual cycle. Methods Seven patients who underwent a hysterectomy and samples of 120 archived patients' endometrial curettage or parts of the uterus that were formalin-fixed and embedded in paraffin. Western blotting was performed to evaluate the specificity and sensitivity of the antibody. Immunohistochemistry was conducted to localize the SERPINE2 expression site. Quantitative analysis was conducted to evaluate expression levels of SERPINE2 in various sub-phases of the menstrual cycle. Results The SERPINE2 protein was primarily detected in the uterine fluid during the mid- and late-secretory phases of the menstrual cycle. It was predominantly expressed in the luminal and glandular epithelium, less in the myometrium, and only dispersedly in certain stromal cells throughout the menstrual cycle. A quantitative analysis of expression levels of SERPINE2 in the glandular epithelium revealed that it was highly expressed in the endometrium during the secretory phase compared to the proliferative phase. Conclusions The SERPINE2 protein is highly expressed in the endometrium during the secretory phase, indicating that it may participate in tissue remodeling involved in implantation.

Hwu Yuh-Ming

2011-03-01

330

Angiotensin-converting enzyme inhibition augments coronary release of tissue plasminogen activator in women but not in men.  

Science.gov (United States)

The renin-angiotensin system regulates the vascular fibrinolytic balance. In the human forearm vasculature, angiotensin-converting enzyme (ACE) inhibitors (ACE-Is) increase the release of t-PA through endogenous bradykinin. We tested the hypothesis that ACE inhibition and sex modulate the endogenous coronary release of tissue plasminogen activator (t-PA) in hypertensive patients. Seventy-three patients underwent diagnostic coronary angiography and had normal coronary angiograms. Thirty-three patients (21 men and 12 women) were treated with imidapril (5 mg/day) for 4 weeks (ACE-I group), and 40 (23 men and 17 women) were not treated with ACE-I (non-ACE-I group). All of the women were postmenopausal. Coronary blood flow in the left anterior descending artery was evaluated by measuring Doppler flow velocity. Net coronary t-PA release was determined as (coronary sinus-aorta gradient of t-PA)x(coronary blood flow)x[(100-hematocrit)/100]. Age, arterial pressure, heart rate, lipid levels, coronary flow, and the plasma level of t-PA at either aorta or coronary sinus were comparable among the 4 groups. In women, net t-PA release in the ACE-I group was significantly higher than that in the other groups (P<0.05; man non-ACE-I group: 1.4+/-2.6 ng/mL; woman non-ACE-I group: 1.4+/-3.1 ng/mL; man ACE-I group: -1.8+/-2.8 ng/mL; woman ACE-I group: 14.8+/-3.6 ng/mL). Correction for smoking status gave similar results. There was a significant negative correlation between serum ACE activity and coronary t-PA release in women (r=-0.38; P<0.05) but not in men. ACE inhibition increases coronary release of t-PA in women but not in men. PMID:20606106

Matsumoto, Tetsuya; Takashima, Hiroyuki; Nakae, Ichiro; Yamane, Tetsunobu; Hayashi, Hideki; Horie, Minoru

2010-09-01

331

Impact of the 4G/5G polymorphism in the plasminogen activator inhibitor-1 gene on primary nephrotic syndrome.  

Science.gov (United States)

The aim of the present study was to investigate whether the four guanosines (4G)/five guanosines (5G) polymorphism in the gene coding for plasminogen activator inhibitor-1 (PAI-1) affects the clinical features of primary nephrotic syndrome (PNS). A cohort of 200 biopsy-diagnosed PNS patients was studied, with 40 healthy subjects as controls. The PAI-1 gene polymorphism was detected by polymerase chain reaction and DNA sequencing. Associations between the PAI-1 4G/5G polymorphism and clinical features and pathological types of PNS were analyzed. The results indicated that the PAI-1 genotype distribution is significantly different between patients with PNS and healthy controls, with significantly higher numbers of the 4G/4G genotype and lower numbers of the 5G5G genotype detected in PNS patients compared to controls (both PIgAN) and membranous nephropathy (MN) were associated with significantly increased frequencies of the 4G/4G and 4G/5G genotypes, as well as of the 4G allele. The increased 4G frequency was also detected in patients with minimal change disease (MCD). Significantly increased international normalized ratio (INR) and prolonged activated partial thromboplastin time (APTT) were observed in 4G/4G compared to 5G/5G PNS subjects. The response to steroids was not significantly different among the three genotypes. In conclusion, the 4G allele of the PAI-1 gene appears to be associated with PNS, especially in MN and IgAN patients. These findings suggest that specific targeting may be required for the treatment of PNS patients with the 4G/4G genotype. PMID:24435552

Luo, Yuezhong; Wang, Chao; Tu, Haitao

2014-03-01

332

Cytosolic Proteins Contribute to Surface Plasminogen Recruitment of Neisseria meningitidis?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Plasminogen recruitment is a common strategy of pathogenic bacteria and results in a broad-spectrum surface-associated protease activity. Neisseria meningitidis has previously been shown to bind plasminogen. In this study, we show by several complementary approaches that endolase, DnaK, and peroxiredoxin, which are usually intracellular proteins, can also be located in the outer membrane and act as plasminogen receptors. Internal binding motifs, rather than C-terminal lysine residues, are res...

2007-01-01

333

Ovine toxoplasmosis.  

Science.gov (United States)

Congenital infection with Toxoplasma gondii is an important cause of abortion in sheep worldwide. The cat is the definitive host of the parasite, and infected cats may shed millions of oocysts in their faeces resulting in extensive environmental contamination and an important source of infection for grazing herbivorous animals. Studies looking at development of specific antibodies in sheep, as an indicator of exposure to T. gondii, have shown that there is an increase in seroprevalence associated with age indicating that most infections in sheep occur following birth. The stage of gestation when transplacental transmission of T. gondii to the developing foetus occurs is critical in determining the clinical outcome. The importance of endogenous transplacental transmission in persistently infected ewes and its clinical importance is a subject of current debate. Ewes infected prior to mating develop immune responses that help protect against disease in a subsequent pregnancy and also against experimental challenge administered during pregnancy. Both innate and adaptive immune responses are activated following T. gondii infection and experiments involving the chronic cannulation of peripheral lymph nodes in sheep have allowed the dynamics of the immune responses to be analysed in real time. A live vaccine, Toxovax is the only commercially available vaccine worldwide to protect against congenital toxoplasmosis. PMID:19995468

Innes, Elisabeth A; Bartley, Paul M; Buxton, David; Katzer, Frank

2009-12-01

334

Increase of cellular fibrinolysis in human lung cancer cell line by radiation. Relationship between urokinase-type plasminogen activator (uPA) and metastasis and invasion  

Energy Technology Data Exchange (ETDEWEB)

It is well known that urokinase-type plasminogen activator (uPA) activates fibrinolysis of tumor cells and accelerates their metastasis and invasion. Human adenosquamous cell line, AOI cells, were stimulated to produce and accumulate of uPA by radiation. In AOI cells, there was relationship between uPA production and accumulation and the radiation doses. It was suggested that radiation had the possibility to accelerate the metastasis and invasion by increasing the production and accumulation of uPA from cancer cells. (author)

Itoh, Y.H.; Oguri, Takashi; Miyata, Nobuki [Aichi Medical Univ., Nagakute (Japan); Nakatsugawa, Shigekazu

1996-09-01

335

Npas4 Is Activated by Melatonin, and Drives the Clock Gene Cry1 in the Ovine Pars Tuberalis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Seasonal mammals integrate changes in the duration of nocturnal melatonin secretion to drive annual physiologic cycles. Melatonin receptors within the proximal pituitary region, the pars tuberalis (PT), are essential in regulating seasonal neuroendocrine responses. In the ovine PT, melatonin is known to influence acute changes in transcriptional dynamics coupled to the onset (dusk) and offset (dawn) of melatonin secretion, leading to a potential interval-timing mechanism capable of decoding c...

West, A.; Dupre?, S. M.; Yu, L.; Paton, I. R.; Miedzinska, K.; Mcneilly, A. S.; Davis, J. R. E.; Burt, D. W.; Loudon, A. S. I.

2013-01-01

336

Course and prerequisites of Lys-plasminogen formation during fibrinolysis  

International Nuclear Information System (INIS)

Plasmin-catalyzed modification of the native plasma zymogen Glu1-plasminogen to its more reactive LYS78 form has been shown to be enhanced in the presence of fibrin. The aim of the present work has been to characterize the influence of fibrinopeptide release, fibrin polymerization, and plasmin cleavage of fibrin on the rate of Lys78-plasminogen formation. 125I-Labeled Glu1- to Lys78-plasminogen conversion was catalyzed by preformed Lys78-plasmin, or by plasmin generated during plasminogen activation with tissue plasminogen activator or urokinase. The two forms of plasminogen were quantitated following separation by polyacrylamide gel electrophoresis in acetic acid/urea. Plasmin generated by plasminogen activator was monitored by a fixed-time amidolytic assay. The rate of Lys78-plasminogen formation was correlated, in separate experiments, to the simultaneous, plasmin-catalyzed cleavage of 125I-labeled fibrinogen or fibrin to fragments X, Y, and D. The radiolabeled components were quantitated after separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The results show that the formation of both bathroxobin-catalyzed des-A-fibrin and thrombin-catalyzed des-AB-fibrin leads to marked stimulation of Lys78-plasminogen formation, whereas inhibition of fibrin polymerization, with Gly-Pro-Arg-Pro, abolishes the stimulatory effect. The rate of Lys78-plasminogen formation varies markedly in the course of fibrinolysis. It is concluded that efficient conversion of Glu1 to Lys78-plasminogen requires formation of des-A or des-AB fragment X polymer, in the absence of significant quantities of fragments Y and D. Under these conditions, Lys78-plasminogen formation is a potent stimulatory mechanism in fibrinolysis

1988-04-05

337

Coagulation factors, fibrinogen and plasminogen activator inhibitor-1, are differentially regulated by yellow fever virus infection of hepatocytes.  

Science.gov (United States)

Yellow fever virus (YFV) infection poses a great risk to un-vaccinated individuals living or traveling in the endemic regions of Africa and South America. It is estimated that approximately 30,000 people die each year of this disease. The liver is the main target of YFV, where as many as 80% of the hepatocytes may become involved in the infection. The overwhelming infection of the liver is associated with the observed hemorrhagic disease manifestations such as petechiae, ecchymoses, and hematemesis which are all thought to be linked with the observed coagulation abnormalities that include prolonged clotting times, reduction in clotting factors, fibrin-split products (D-dimers) and elevated prothrombin times. Many factors involved in the coagulation pathway are produced by hepatocytes, such as fibrinogen (FBG) and plasminogen activator inhibitor-1 (PAI-1). Both of these proteins have been indicated in another flavivirus related disease, dengue, as having roles related to the bleeding abnormalities observed and overall outcome of infection. In this study we wanted to determine if FBG and PAI-1 expression levels by human hepatocytes was disrupted or altered by infection with either wild-type Asibi or vaccine strain17-D YFVs. Our findings indicate that YFV infection does affect the transcriptional and translational expression of FBG and PAI-1 in human hepatocytes and that these results are further affected by IL-6 during early stages of infection. These results may lead to further understanding of the molecular mechanism associated with bleeding abnormalities observed during late stage YFV infection. PMID:23639427

Woodson, Sara E; Freiberg, Alexander N; Holbrook, Michael R

2013-08-01

338

Low vs standard dose of recombinant tissue plasminogen activator in treating East Asian patients with acute ischemic stroke  

Directory of Open Access Journals (Sweden)

Full Text Available Background : Intravenous recombinant tissue plasminogen activator (rtPA has been approved to treat eligible patients with acute ischemic stroke within 4.5 hours of onset. The rationale for using a lower dose in Asian patients came from concerns about intracerebral hemorrhage because of the racial differences in blood coagulation-fibrinolysis factors. Aim : The aim of this systemic review was to compare the data from previous studies to address the efficacy and safety of using low-dose vs standard-dose rtPA in treating patients with acute ischemic stroke. Material and Methods : Previous studies were searched and analyzed. The confidence interval was calculated at 95%. Baseline characteristics and outcomes of the patients were compared between two doses of rtPA (0.6 vs 0.9 mg/kg, using Z test for two independent proportions. Results : Patients who received standard-dose rtPA had significantly higher favorable outcome at 3 months (33.1 vs 47.2%, P<0.0001, without significant difference in the rates of symptomatic intracerebral hemorrhage (3.5 vs 4.3%, P = 0.42 and mortality (13.1 vs 11.7%, P = 0.56. However, patients in the low-dose group were older and had more severe stroke. Conclusions : Patients receiving standard-dose rtPA seem to have higher rates of favorable outcome. However, there were significant differences in baseline characteristics between the two groups. A further, well-designed, randomized study in the same population is still needed to clarify the suspected benefit of the standard dose for East Asian patients.

Dharmasaroja Pornpatr

2011-01-01

339

Thrombolysis with Intravenous Tissue Plasminogen Activator (rt-PA) Predicts Favorable Discharge Disposition in Patients with Acute Ischemic Stroke  

Science.gov (United States)

Background and Purpose Acute ischemic stroke patients receiving IV tissue plasminogen activator (rt-PA) within 3 hours of symptom onset are 30% more likely to have minimal disability at three months. During hospitalization, short-term disability is subjectively measured by discharge disposition, whether to home, Inpatient Rehabilitation (IR), Skilled Nursing Facility (SNF) or Subacute Care (Sub). There are no studies assessing the role of rt-PA use as a predictor of post-stroke disposition. Methods Retrospective analysis of all ischemic stroke patients admitted to the University of Texas Houston Medical School (UTHMS) Stroke Service between Jan 2004 and Oct 2009. Baseline demographics and National Institute of Health Stroke Scale (NIHSS) score were collected. Cerebrovascular disease risk factors were used for risk stratification. Results Home vs. IR, SNF, Sub Of 2225 acute ischemic stroke patients, 1019 were discharged home, 1206 to another level of care. Patients who received rt-PA therapy were 1.9 times more likely to be discharged home (P = <0.0001; OR 1.945, 95% CI 1.538 to 2.459). IR vs. SNF, Sub / SNF vs. Sub Of 1206 acute ischemic stroke patients, 719 patients were discharged to acute IR, 371 were discharged to SNF, 116 to Sub. There were no differences in disposition between patients who received rt-PA therapy. Conclusions Stroke patients who receive IV rt-PA for acute ischemic stroke are more 1.9 times more likely to be discharged directly home after hospitalization. This study is limited by its retrospective nature and the undetermined role of psychosocial factors related to discharge.

Ifejika-Jones, Nneka L.; Harun, Nusrat; Mohammed-Rajput, Nareesa A.; Noser, Elizabeth A.; Grotta, James C.

2011-01-01

340

Interactions of plasminogen activator inhibitor-1 with vitronectin involve an extensive binding surface and induce mutual conformational rearrangements  

DEFF Research Database (Denmark)

In order to explore early events during the association of plasminogen activator inhibitor-1 (PAI-1) with its cofactor vitronectin, we have applied a robust strategy that combines protein engineering, fluorescence spectroscopy, and rapid reaction kinetics. Fluorescence stopped-flow experiments designed to monitor the rapid association of PAI-1 with vitronectin indicate a fast, concentration-dependent, biphasic binding of PAI-1 to native vitronectin but only a monophasic association with the somatomedin B (SMB) domain, suggesting that multiple phases of the binding interaction occur only when full-length vitronectin is present. Nonetheless, in all cases, the initial fast interaction is followed by slower fluorescence changes attributed to a conformational change in PAI-1. Complementary experiments using an engineered, fluorescently silent PAI-1 with non-natural amino acids showed that concomitant structural changes occur as well in native vitronectin. Furthermore, we have measured the effect of vitronectin on the rate of insertion of the reactive center loop into beta-sheet A of PAI-1 during reaction with target proteases. With a variety of PAI-1 variants, we observe that both full-length vitronectin and the SMB domain have protease-specific effects on the rate of loop insertion but that the two exhibit clearly different effects. These results support a model for PAI-1 binding to vitronectin in which the interaction surface extends beyond the region of PAI-1 occupied by the SMB domain. In support of this model are recent results that define a PAI-1-binding site on vitronectin that lies outside the somatomedin B domain (Schar, C. R., Blouse, G. E., Minor, K. H., and Peterson, C. B. (2008) J. Biol. Chem. 283, 10297-10309) and the complementary site on PAI-1 (Schar, C. R., Jensen, J. K., Christensen, A., Blouse, G. E., Andreasen, P. A., and Peterson, C. B. (2008) J. Biol. Chem. 283, 28487-28496).

Blouse, Grant E; Dupont, Daniel Miotto

2009-01-01

 
 
 
 
341

The effect of plasminogen activator inhibitor-1 -675 4G/5G polymorphism on familial Mediterranean fever (FMF) disease.  

Science.gov (United States)

Familial Mediterranean fever (FMF) is an autosomal recessive disease that is the most common of a rare group of disorders collectively termed familial hereditary periodic fever syndromes, also known as autoinflammatory syndromes. FMF is predominantly affecting people of Mediterranean descent and clinically characterized by intermittent attacks of fever with peritonitis and abdominal pain, pleuritis, arthritis, or erysipelas-like rashes. Amyloidosis due to chronic inflammation progressing to renal failure is one of the most serious potential complications of this disease.Patients with inflammatory diseases, such as systemic lupus erythematosus and rheumatoid arthritis, and conditions with chronic subclinical inflammation, like obesity and diabetes mellitus, are now considered to have an increased risk of atherosclerotic cardiovascular complications. FMF is also an inflammatory disease, and it is accepted that even during attack-free periods significant inflammatory reaction continues. However, whether this inflammatory process causes premature atherosclerosis is not known due to a lack of data.Different studies have investigated the association between the fibrinolytic and inflammatory process parameters. PAI-1 is paracrine secretion of pro- and antiinflammatory cytokines, thereby playing a possible role in the adiposity-related inflammation and atherosclerosis. The patients with IRS have higher values of fibrinogen, factor VII, VIII, Von Willebrand factor and Plasminogen Activator Inhibitor (PAI) compared to control subjects. So that we aimed in this study to investigate whether FMF patients with/without amyloidosis and with M694V homozygote mutation, have increased risk for atherosclerotic cardiovascular complications and to determine the strength of association between MEFV gene-mutation types. To our knowledge, this is the first case control and cross-sectional study in the pediatric age groups. PMID:19033264

Ozel Demiralp, Duygu; Ekim, Mesiha; Akar, Nejat

2009-01-01

342

Transcatheter Thrombolysis with High-Dose Bolus Tissue Plasminogen Activator in Iatrogenic Arterial Occlusion after Femoral Arterial Catheterization  

International Nuclear Information System (INIS)

Purpose: To assess the efficacy of percutaneous local thrombolysis with high-dose bolus recombinant tissue plasminogen activator (rt-PA) in patients with acute limb ischemia due to arterial thrombosis after cardiac catheterization.Methods: We treated eight patients (7 men; mean age 56 years) with thrombotic occlusion of both the common femoral artery (CFA) and external iliac artery (EIA) in six patients and of the CFA only in two patients. Two 5 mg boluses of rt-PA were injected into the proximal clot through a 5 Fr end-hole catheter and subsequently two additional boluses of 5 mg rt-PA were given through a catheter with multiple side-holes. In case of a significant amount of residual thrombus, a continuous infusion of 2.5 mg/hr of rt-PA was started.Results: Successful lysis was achieved in all patients. The mean duration of lysis was 2 hr 41 min. The mean total amount of rt-PA delivered was 23.16 mg. In four patients unmasked flow-limited dissections confined to the CFA were managed by prolonged balloon dilatation, while in the remaining four patients with extension of the dissection to the external iliac artery one or two Easy Wallstents were implanted. There was prompt relief of lower limb ischemic symptoms and signs in all patients. Two groin hematomas were conservatively treated.Clinical and color Doppler flow imaging follow-up with a mean duration of 15 months, showed no reappearance of ischemic symptoms or development of restenosis in any of the patients. One patient died 6 months after thrombolysis.Conclusions: Transcatheter thrombolysis with high-dose bolus rt-PA is a safe and effective treatment inpatients with iatrogenic arterial occlusion after femoral catheterization. Underlying dissections should be treated by prolonged balloon dilatation but stent implantation is often required

2002-01-01

343

Influence of bone marrow fat embolism on coagulation activation in an ovine model of vertebroplasty  

Digital Repository Infrastructure Vision for European Research (DRIVER)

BACKGROUND: Intraoperative cardiovascular deterioration as a result of pulmonary embolization of bone marrow fat is a potentially serious complication during vertebroplasty. The release of fatty material and thromboplastin from the bone marrow cavity during vertebroplasty may activate the coagulation cascade resulting in thrombogenesis, and pharmacological prophylaxis may therefore prevent cardiovascular complications. Thus, the effects of bone marrow fat embolism on coagulation activation du...

Krebs, J.; Ferguson, S. J.; Hoerstrup, S. P.; Goss, B. G.; Haeberli, A.; Aebli, N.

2008-01-01

344

Plasminogen activator activity in lung and alveolar macrophages of rats exposed to graded single doses of. gamma. rays to the right hemithorax  

Energy Technology Data Exchange (ETDEWEB)

Male rats were sacrificed 2 or 6 months after a single dose of 0-30 Gy of /sup 60/Co ..gamma.. rays to the right hemithorax. At autopsy, macrophages were lavaged from the right lung, counted, and frozen. The right (irradiated) and the left (shielded) lungs were frozen, then assayed for plasminogen activator (PLA) activity by the fibrin plate lysis method. Freeze-thawed macrophages were assayed for both PLA activity (/sup 125/I-fibrin clot lysis method) and fibrinolytic inhibitor activity (inhibition of urokinase-induced fibrin lysis). There was a linear, dose-dependent decrease in right lung PLA activity over the dose range of 10-30 Gy at 2 and 6 months postirradiation, reductions of 3.1 and 2.6% per Gy, respectively. PLA activity at all radiation doses was 10-15% higher at 6 months than at 2 months indicative of a partial recovery of this endothelial function in the irradiated lung. PLA activity per 10/sup 6/ macrophages decreased with increasing radiation dose at both autopsy times, closely paralleling lung PLA activity. This radiation-induced decrease in macrophage PLA activity was not due to increased fibrinolytic inhibitor activity in the irradiated macrophages. These data quantitate the dose response and time course of radiation-induced fibrinolytic defects in rat lung and suggest that information obtained from a minimally invasive procedure such as bronchoalveolar lavage may serve as an index of the degree of pulmonary fibrinolytic dysfunction after irradiation.

Ts' ao, C.; Ward, W.F.

1985-09-01

345

Plasminogen activator activity in lung and alveolar macrophages of rats exposed to graded single doses of ? rays to the right hemithorax  

International Nuclear Information System (INIS)

Male rats were sacrificed 2 or 6 months after a single dose of 0-30 Gy of "6"0Co ? rays to the right hemithorax. At autopsy, macrophages were lavaged from the right lung, counted, and frozen. The right (irradiated) and the left (shielded) lungs were frozen, then assayed for plasminogen activator (PLA) activity by the fibrin plate lysis method. Freeze-thawed macrophages were assayed for both PLA activity ("1"2"5I-fibrin clot lysis method) and fibrinolytic inhibitor activity (inhibition of urokinase-induced fibrin lysis). There was a linear, dose-dependent decrease in right lung PLA activity over the dose range of 10-30 Gy at 2 and 6 months postirradiation, reductions of 3.1 and 2.6% per Gy, respectively. PLA activity at all radiation doses was 10-15% higher at 6 months than at 2 months indicative of a partial recovery of this endothelial function in the irradiated lung. PLA activity per 10"6 macrophages decreased with increasing radiation dose at both autopsy times, closely paralleling lung PLA activity. This radiation-induced decrease in macrophage PLA activity was not due to increased fibrinolytic inhibitor activity in the irradiated macrophages. These data quantitate the dose response and time course of radiation-induced fibrinolytic defects in rat lung and suggest that information obtained from a minimally invasive procedure such as bronchoalveolar lavage may serve as an index of the degree of pulmonary fibrinolytic dysfunction after irradiation

1985-01-01

346

Glyceraldehyde-3-Phosphate Dehydrogenase of Streptococcus pneumoniae Is a Surface-Displayed Plasminogen-Binding Protein  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The recruitment of plasminogen endows the bacterial cell surface of Streptococcus pneumoniae with proteolytic activity. In this study we demonstrate specific plasmin- and plasminogen-binding activity for the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which is located in the cytoplasm as well as on the surface of pneumococci. GAPDH exhibits a high affinity for plasmin and a significantly lower affinity for plasminogen.

Bergmann, Simone; Rohde, Manfred; Hammerschmidt, Sven

2004-01-01

347

Characterization of the plasminogen activator of herpesvirus-transformed cells and examination of its correlation with the tumorigenic and metastatic ability of in vivo-derived sublines  

International Nuclear Information System (INIS)

Herpes simplex virus type 2-transformed hamster embryo fibroblasts (333-8-9 cells) produce increased amounts of plasminogen activator (PA) compared with normal hamster cells. The 333-8-9 PA activity was quantitated in comparison to a PA standard, urokinase (UK). Using a direct PA assay in which 125I-labeled plasminogen is cleaved, a linear dose-response was seen over a 1000-fold range in UK concentration when plotted on a semi-logarithmic scale. Extracellular PA activity secreted by the HSV-2-transformed cell line, 333-8-9, followed a similar dose-response slop. The optimum pH and osmolarity for detection of the 333-8-9 extracellular PA activity were pH 8.9 and approximately 150 mOsmol, respectively. Secretion of PA by the 333-8-9 cells did not vary significantly on a per cell basis over cell densities ranging from 0.1 to 8.0 x 107 cells/T-75 cm2 flask. This assay was accurate, reproducible, and demonstrated that the 333-8-9 cells produced at least a 20-fold greater amount of PA activity than their normal cell counterparts. Based on the molecular weight (50-58 Kd) of the secreted 333-8-9 cell PA and lack of fibrin stimulation of the PA activity, it is concluded to be a urokinase-type PA

1986-01-01

348

Quantitative MRI reveals the elderly ischemic brain is susceptible to increased early blood–brain barrier permeability following tissue plasminogen activator related to claudin 5 and occludin disassembly  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Great uncertainty exists as to whether aging enhances the detrimental effects of tissue plasminogen activator (tPA) on vascular integrity of the ischemic brain. We hypothesized that tPA treatment would augment ischemic injury by causing increased blood–brain barrier (BBB) breakdown as determined by quantitative serial T1 and T2 magnetic resonance imaging (MRI), and the transfer constant for gadolinium-diethylenetriamine penta-acetic acid (Gd-DTPA) from blood to brain in aged (18 to 20 month...

Kaur, Jaspreet; Tuor, Ursula I.; Zhao, Zonghang; Barber, Philip A.

2011-01-01

349

Acute coronary thrombolysis with recombinant human tissue-type plasminogen activator: initial patency and influence of maintained infusion on reocclusion rate  

Digital Repository Infrastructure Vision for European Research (DRIVER)

An intravenous infusion of 40 mg of recombinant tissue-type plasminogen activator (rt-PA) was given intravenously over 90 minutes to 123 patients with acute myocardial infarction (AMI) of less than 4 hours' duration. A coronary angiogram was recorded at the end of the infusion in 119 patients. Central assessment of the angiograms revealed a patent infarct-related artery in 78 patients (patency rate 66%, 95% confidence limits 57 to 74%). Patients with a patent infarct-related artery at the fir...

Verstraete, M.; Brower, R. W.; Collen, D.; Bone, D. P.; Zwaan, C.; Erbel, R.; Hillis, W. S.; Lennane, R. J.; Lubsen, J.; Mathey, D.; Reid, D. S.; Rutsch, W. R.; Schartl, M.; Schofer, J.; Serruys, P. W. J. C.

1987-01-01

350

Induction of plasminogen activator inhibitor 1 gene expression in murine liver by lipopolysaccharide. Cellular localization and role of endogenous tumor necrosis factor-alpha.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We previously demonstrated that lipopolysaccharide (LPS) induces plasminogen activator inhibitor 1 (PAI-1) gene expression primarily in endothelial cells in most organs of the mouse, with maximal induction by 3 hours. Here we show that induction in the liver occurs in a distinctly different pattern. For example, the increase in PAI-1 mRNA in liver was biphasic with an initial peak at 1 to 2 hours and a second peak at 6 to 8 hours. Moreover, in situ hybridization experiments revealed that PAI-...

Fearns, C.; Loskutoff, D. J.

1997-01-01

351

Tissue Plasminogen Activator alters intracellular sequestration of zinc through interaction with the transporter ZIP4  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Glutamatergic neurons contain free zinc packaged into neurotransmitter-loaded synaptic vesicles. Upon neuronal activation, the vesicular contents are released into the synaptic space, whereby the zinc modulates activity of postsynaptic neurons though interactions with receptors, transporters and exchangers. However, high extracellular concentrations of zinc trigger seizures and are neurotoxic if substantial amounts of zinc re-enter the cells via ion channels and accumulate in the cytoplasm. T...

Emmetsberger, Jaime; Mirrione, Martine M.; Zhou, Chun; Fernandez-monreal, Monica; Siddiq, Mustafa M.; Ji, Kyungmin; Tsirka, Stella E.

2010-01-01

352

Role of prostaglandins in determining the increased cardiac sympathetic nerve activity in ovine sepsis.  

Science.gov (United States)

Effective treatment of sepsis remains a significant challenge in intensive care units. During sepsis, there is widespread activation of the sympathetic nervous system, which is thought to have both beneficial and detrimental effects. The sympathoexcitation is thought to be partly due to the developing hypotension, but may also be a response to the inflammatory mediators released. Thus, we investigated whether intracarotid infusion of prostaglandin E2 (PGE2) induced similar cardiovascular changes to those caused by intravenous infusion of Escherichia coli in sheep and whether inhibition of prostaglandin synthesis, with the nonselective cyclooxygenase inhibitor indomethacin, administered at 2 and 8 h after the onset of sepsis, reduced sympathetic nerve activity (SNA), and heart rate (HR). Studies were performed in conscious sheep instrumented to measure mean arterial pressure (MAP), HR, cardiac SNA (CSNA), and renal SNA (RSNA). Intracarotid infusion of PGE2 (50 ng·kg(-1)·min(-1)) increased temperature, CSNA, and HR, but not MAP or RSNA. Sepsis, induced by infusion of E. coli, increased CSNA, but caused an initial, transient inhibition of RSNA. At 2 h of sepsis, indomethacin (1.25 mg/kg bolus) increased MAP and caused reflex decreases in HR and CSNA. After 8 h of sepsis, indomethacin did not alter MAP, but reduced CSNA and HR, without altering baroreflex control. These findings indicate an important role for prostaglandins in mediating the increase in CSNA and HR during the development of hyperdynamic sepsis, whereas prostaglandins do not have a major role in determining the early changes in RSNA. PMID:24789991

Booth, Lindsea C; Ramchandra, Rohit; Calzavacca, Paolo; May, Clive N

2014-07-01

353

Functionally stable plasminogen activator inhibitor-1 in a family with cardiovascular disease and vitiligo.  

Science.gov (United States)

Vitiligo is a common skin condition with a complex pathophysiology characterized by the lack of pigmentation due to melanocyte degeneration. In this study, we investigated PAI-1 antigen (Ag) and activity levels in a 34 year old male with extensive vascular disease, alopecia areata and vitiligo. Fasting PAI-1 Ag and activity levels were measured at 9 a.m. in the subject and family members. Both PAI-1 Ag (67 ± 38 vs. 18.6 ± 6.5 ng/ml, P vitiligo is not known, it is likely due to post-translational modifications or increased binding affinity for a stabilizing cofactor. In conclusion, enhanced stability of PAI-1 may contribute to the pathophysiology of vascular disease and associated melanocyte degeneration. Systemic or local treatment with PAI-1 inhibitors may offer a potential treatment alternative to the near orphan status for vitiligo drug development. PMID:24197654

Agirbasli, Mehmet; Eren, Mesut; Yasar, Songul; Delil, Kenan; Goktay, Fatih; Oner, Ebru Toksoy; Vaughan, Douglas E

2014-07-01

354

Phorbol ester stimulates the synthesis and phosphorylation of a 48 kDA-intra-cellular protein in plasminogen activator secreting melanoma cells  

International Nuclear Information System (INIS)

Phorbol ester (12-O-tetradecanoyl-phorbol 13-acetate) stimulates the secretion of tissue-type plasminogen activator by the melanoma cell line, Bowes. This effect is associated with increased levels of mRNAs for both tissue-type plasminogen activator and a 48 KDa-protein. Labelling of melanoma cells with L-["3"2S]methionine allowed to identify an intracellular protein which was identical with the in vitro translation product of the 48 kDa-protein mRNA. As detectable by L-["3"5S]methionine labelling, the protein was mainly localized in the cytosol. In vitro phosphorylation reactions, carried out on subcellular fractions revealed a membrane-associated protein which also had the characteristics of the 48 kDa-protein. Phosphorylation did not require Ca"2"+-ions. Reconstitution experiments between membrane and cytosol fractions of phorbol ester-treated and untreated cells showed that the 48 kDa protein occurs in a cytosolic, unphosphorylated and a membrane-bound, phosphorylated form and that the former is converted to the latter by a phorbol ester activated, membrane-associated protein kinase

1986-05-29

355

A protease of Bacteroides gingivalis degrades cell surface and matrix glycoproteins of cultured gingival fibroblasts and induces secretion of collagenase and plasminogen activator.  

Science.gov (United States)

To assess the direct effects of Bacteroides gingivalis on periodontal cells, human gingival fibroblasts were cultured in the presence of B. gingivalis extracts or a trypsinlike enzyme partially purified from the bacteria by chromatography on benzamidine-Sepharose and Sephacryl S-200. Analysis of cell surface glycoproteins by the periodate-[3H]borohydride labeling technique combined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE)-fluorography demonstrated that fibronectin and some other high-molecular-weight cell surface glycoproteins were degraded by a 35,000-Mr(35K) B. gingivalis protease. Immunostaining of the fibroblast cultures showed degradation of intercellular matrix fibronectin by the 35K protease. The pattern of fibronectin degradation was monitored by examining the reaction products with the SDS-PAGE-immunoblotting technique. The protease degraded fibronectin rapidly and more extensively than did corresponding amounts of pancreatic trypsin. Collagenase secretion by the fibroblasts was assayed by incubating cell culture medium with soluble type I [3H]collagen at 25 degrees C followed by SDS-PAGE-fluorography analysis of the reaction products. The medium was also assayed for plasminogen activator activity by using a casein-agarose diffusion plate assay. The fibroblasts cultured with the 35K protease secreted increased amounts of collagenase and plasminogen activator into the medium. The results suggest that periodontal infection by B. gingivalis causes proteolytic damage of the host cell surface structures. Concomitantly, B. gingivalis may induce the cells to degrade their pericellular matrix. PMID:2535833

Uitto, V J; Larjava, H; Heino, J; Sorsa, T

1989-01-01

356

Characteristics of ovine cytotoxic lymphocytes  

International Nuclear Information System (INIS)

Experiments were conducted to examine characteristics of the effector cells responsible for cell-mediated cytotoxicity in the sheep. Conditions for the production and assay of ovine T cell growth factor (TCGF) activity were evaluated. Peripheral blood leukocytes (PBL) were stimulated with concanavalin A (Con A) in the presence of 2% autologous serum or serum-free media. A 28 h proliferation assay with 2.5 x 104 h Con A blasts per well was optimal for detection of TCGF. Peak TCGF activity occurred with a 30-37kD molecular weight fraction. Ovine PBL were used for in vitro generation of genetically-restricted cytotoxic T lymphocytes (CTL). Peripheral blood leukocytes from sheep that had been previously inoculated with live vaccinia virus were stimulated by being cultured in vitro on glutaraldehyde-fixed vaccinia-infected autologous skin fibroblasts. Cytotoxic T lymphocyte activity was assessed in a 6 h 51Cr-release assay on autologous and allogeneic fibroblasts targets. Killing was restricted to virus-infected autologous targets. In vitro generation of both anti-vaccinia and anti-TNP CTL activity could be enhanced by the addition of TCGF containing media from ConA-stimulated PBL

1987-01-01

357

Plasminogen activator inhibitor 1: Mechanisms of its synergistic regulation by growth factors  

Energy Technology Data Exchange (ETDEWEB)

My research is on the synergistic regulation of PAI-1 by EGF and TGF-?. The mechanism of synergistic regulation of PAI-1 by EGF and TGF-? are addressed. Methods are described for effective identification of RNA accessible sites for antisense oligodexoxynucleotides (ODNs) and siRNA. In this study effective AS-ODN sequences for both Lcn2 and Bcl2 were identified by in vitro tiled microarray studies. Our results suggest that hybridization of ODN arrays to a target mRNA under physiological conditions might be used as a rapid and reliable in vitro method to accurately identify targets on mRNA molecules for effective antisense and potential siRNA activity in vivo.

Song, Xiaoling

2011-12-01

358

Intraclot recombinant tissue-type plasminogen activator reduces perihematomal edema and mortality in patients with spontaneous intracerebral hemorrhage.  

Science.gov (United States)

The study aimed to investigate the impact of intraclot recombinant tissue-type plasminogen activator (rt-PA) on perihematomal edema (PHE) development in patients with intracerebral hemorrhage (ICH) treated with minimally invasive surgery (MIS) and the effects of intraclot rt-PA on the 30-day survival. We reviewed the medical records of ICH patients undergoing MIS between October 2011 and July 2013. A volumetric analysis was done to assess the change in PHE and ICH volumes at pre-MIS (T1), post-MIS (T2) and day 10-16 (T3) following diagnostic computed tomographic scans (T0). Forty-three patients aged 52.8±11.1 years with (n=30) or without rt-PA (n=13) were enrolled from our institutional ICH database. The median rt-PA dose was 1.5 (1) mg, with a maximum dose of 4.0 mg. The ratio of clot evacuation was significantly increased by intraclot rt-PA as compared with controls (77.9%±20.4% vs. 64%±15%; P=0.046). From T1 to T2, reduction in PHE volume was strongly associated with the percentage of clot evacuation (?=0.34; P=0.027). In addition, PHE volume was positively correlated with residual ICH volume at the same day (? ranging from 0.39-0.56, P<0.01). There was no correlation between the cumulative dose of rt-PA and early (T2) PHE volume (?=0.24; P=0.12) or delayed (T3) PHE volume (?=0.19; P=0.16). The 30-day mortality was zero in this cohort. In the selected cohort of ICH patients treated with MIS, intraclot rt-PA accelerated clot removal and had no effects on PHE formation. MIS aspiration and low dose of rt-PA seemed to be feasible to reduce the 30-day mortality in patients with severe ICH. A large, randomized study addressing dose titration and long-term outcome is needed. PMID:24710926

Lian, Li-Fei; Xu, Feng; Tang, Zhou-Ping; Xue, Zheng; Liang, Qi-Ming; Hu, Qi; Zhu, Wen-Hao; Kang, Hui-Cong; Liu, Xiao-Yan; Wang, Fu-Rong; Zhu, Sui-Qiang

2014-04-01

359

Para-aminobenzamidine linked regenerated cellulose membranes for plasminogen activator purification: effect of spacer arm length and ligand density.  

Science.gov (United States)

Despite membrane-based separations offering superior alternative to packed bed chromatographic processes, there has been a substantial lacuna in their actual application to separation processes. One of the major reasons behind this is the lack of availability of appropriately modified or end-group modifiable membranes. In this paper, an affinity membrane was developed using a commercially available serine protease inhibitor, para-aminobenzamidine (pABA). The membrane modification was optimized for protein binding capacity by varying: (i) the length of the spacer arm (SA; 5-atoms, 7-atoms, and 14-atoms) linking the ligand to membrane surface; (ii) the affinity ligand (pABA) density on membrane surface (5-25nmol/cm(2)). Resulting membranes were tested for their ability to bind plasminogen activators (PAs) from mono- and multi-component systems in batch mode. The membrane containing pABA linked through 7-atoms SA but similar ligand density as in the case of 5- or 14-atoms long SA was found to bind up to 1.6-times higher amounts of PA per nmoles of immobilized ligand from conditioned HeLa cell culture media. However, membranes with similar ligand densities but different lengths of SA, showed comparable binding capacities in mono-component system. In addition, the length of SA did not affect the selectivity of the ligand for PA. A clear inverse linear correlation was observed between ligand density and binding capacity until the point of PA binding optima was reached (11±1.0nmol/cm(2)) in mono- and multi-component systems for 7- as well as 14-atoms SA. Up to 200-fold purification was achieved in a single step separation of PA from HeLa conditioned media using these affinity membranes. The issues of ligand leaching and reuse of the membranes were also investigated. An extensive regeneration procedure allowed the preservation of approximately 95% of the PA binding capacity of the membranes even after five cycles of use. PMID:23703544

Fasoli, Ezio; Reyes, Yiaslin Ruiz; Guzman, Osiris Martinez; Rosado, Alexandra; Cruz, Vivian Rodriguez; Borges, Amaris; Martinez, Edmarie; Bansal, Vibha

2013-07-01

360

Prognostic value analysis of urokinase-type plasminogen activator receptor in oral squamous cell carcinoma: an immunohistochemical study  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Oral squamous cell carcinoma (OSCC represents the most common oral malignancy. Despite recent advances in therapy, up to 50% of the cases have relapse and/or metastasis. There is therefore a strong need for the identification of new biological markers able to predict the clinical behaviour of these lesions in order to improve quality of life and overall survival. Among tumour progression biomarkers, already known for their involvement in other neoplasia, a crucial role is ascribed to the urokinase-type plasminogen activator receptor (uPAR, which plays a multiple role in extracellular proteolysis, cell migration and tissue remodelling not only as a receptor for the zymogen pro-uPA but also as a component for cell adhesion and as a chemoattractant. The purpose of this study was to gain information on the expression of uPAR in OSCC and to verify whether this molecule can have a role as a prognostic/predictive marker for this neoplasia. Methods In a retrospective study, a cohort of 189 OSCC patients was investigated for uPAR expression and its cellular localization by immunohistochemistry. As standard controls, 8 normal oral mucosal tissues free of malignancy, obtained from patients with no evidence or history of oral cavity tumours, were similarly investigated. After grouping for uPAR expression, OSCCs were statistically analyzed for the variables age, gender, histological grading (G, tumour size, recurrence, TNM staging and overall survival rate. Results In our immunohistochemical study, 74 cases (39.1% of OSCC showed a mostly cytoplasmic positivity for uPAR, whereas 115 were negative. uPAR expression correlated with tumour differentiation grade and prognosis: percentage of positive cases was the greatest in G3 (70.4% and patients positives for uPAR expression had an expectation of life lower than those for uPAR negatives. Conclusion The results obtained in this study suggest a role of uPAR as a potential biomarker useful to identify higher risk subgroups of OSCC patients.

Rocchetti Romina

2008-08-01

 
 
 
 
361

Combined Approach to Lysis Utilizing Eptifibatide and Recombinant Tissue Plasminogen Activator in Acute Ischemic Stroke-Enhanced Regimen Stroke Trial  

Science.gov (United States)

Background and Purpose In a previous study, 0.3 and 0.45 mg/kg of intravenous recombinant tissue plasminogen activator (rt-PA) were safe when combined with eptifibatide 75 mcg/kg bolus and a 2-hour infusion (0.75 mcg/kg per minute). The Combined Approach to Lysis Utilizing Eptifibatide and rt-PA in Acute Ischemic Stroke–Enhanced Regimen (CLEAR-ER) trial sought to determine the safety of a higher-dose regimen and to establish evidence for a phase III trial. Methods CLEAR-ER was a multicenter, double-blind, randomized safety study. Ischemic stroke patients were randomized to 0.6 mg/kg rt-PA plus eptifibatide (135 mcg/kg bolus and a 2-hour infusion at 0.75 mcg/kg per minute) versus standard rt-PA (0.9 mg/kg). The primary safety end point was the incidence of symptomatic intracranial hemorrhage within 36 hours. The primary efficacy outcome measure was the modified Rankin Scale (mRS) score ?1 or return to baseline mRS at 90 days. Analysis of the safety and efficacy outcomes was done with multiple logistic regression. Results Of 126 subjects, 101 received combination therapy, and 25 received standard rt-PA. Two (2%) patients in the combination group and 3 (12%) in the standard group had symptomatic intracranial hemorrhage (odds ratio, 0.15; 95% confidence interval, 0.01–1.40; P=0.053). At 90 days, 49.5% of the combination group had mRS ?1 or return to baseline mRS versus 36.0% in the standard group (odds ratio, 1.74; 95% confidence interval, 0.70–4.31; P=0.23). After adjusting for age, baseline National Institutes of Health Stroke Scale, time to intravenous rt-PA, and baseline mRS, the odds ratio was 1.38 (95% confidence interval, 0.51–3.76; P=0.52). Conclusions The combined regimen of intravenous rt-PA and eptifibatide studied in this trial was safe and provides evidence that a phase III trial is warranted to determine efficacy of the regimen.

Pancioli, Arthur M.; Adeoye, Opeolu; Schmit, Pamela A.; Khoury, Jane; Levine, Steven R.; Tomsick, Thomas A.; Sucharew, Heidi; Brooks, Claudette E.; Crocco, Todd J.; Gutmann, Laurie; Hemmen, Thomas M.; Kasner, Scott E.; Kleindorfer, Dawn; Knight, William A.; Martini, Sharyl; McKinney, James S.; Meurer, William J.; Meyer, Brett C.; Schneider, Alexander; Scott, Phillip A.; Starkman, Sidney; Warach, Steven; Broderick, Joseph P.

2014-01-01

362

Collagenase and tissue plasminogen activator production in developing rat calvariae: normal progression despite fetal exposure to microgravity  

Science.gov (United States)

Exposure to zero gravity has been shown to cause a decrease in bone formation. This implicates osteoblasts as the gravity-sensing cell in bone. Osteoblasts also are known to produce neutral proteinases, including collagenase and tissue plasminogen activator (tPA), which are thought to be important in bone development and remodeling. The present study investigated the effects of zero gravity on development of calvariae and their expression of collagenase and tPA. After in utero exposure to zero gravity for 9 days on the NASA STS-70 space shuttle mission, the calvariae of rat pups were examined by immunohistochemistry for the presence and location of these two proteinases. The ages of the pups were from gestational day 20 (G20) to postnatal (PN) day 35. Both collagenase and tPA were found to be present at all ages examined, with the greatest amount of both proteinases present in the PN14 rats. At later ages, high amounts were maintained for tPA but collagenase decreased substantially between ages PN21 to PN35. The location of collagenase was found to be associated with bone-lining cells, osteoblasts, osteocytes, and in the matrix along cement lines. In contrast, tPA was associated with endothelial cells lining the blood vessels entering bone. The presence and developmental expression of these two proteinases appeared to be unaffected by the exposure to zero gravity. The calvarial thickness of the pups was also examined; again the exposure to zero gravity showed little to no effect on the growth of the calvariae. Notably, from G20 to PN14, calvarial thickness increased dramatically, reaching a plateau after this age. It was apparent that elevated collagenase expression correlated with rapid bone growth in the period from G20 to PN14. To conclude, collagenase and tPA are present during the development of rat calvariae. Despite being produced by the same cell in vitro, i.e., the osteoblast, they are located in distinctly different places in bone in vivo. Their presence, developmental expression, and quantity do not seem to be affected by a brief exposure to zero gravity in utero.

Davis, B. A.; Sipe, B.; Gershan, L. A.; Fiacco, G. J.; Lorenz, T. C.; Jeffrey, J. J.; Partridge, N. C.

1998-01-01

363

Urokinase plasminogen activator receptor affects bone homeostasis by regulating osteoblast and osteoclast function  

DEFF Research Database (Denmark)

The uPAR and its ligand uPA are expressed by both osteoblasts and osteoclasts. Their function in bone remodeling is unknown. We report that uPAR-lacking mice display increased BMD, increased osteogenic potential of osteoblasts, decreased osteoclasts formation, and altered cytoskeletal reorganization in mature osteoclasts. INTRODUCTION: Urokinase receptor (uPAR) is actively involved in the regulation of important cell functions, such as proliferation, adhesion, and migration. It was previously shown that the major players in bone remodeling, osteoblasts and osteoclasts, express uPAR and produce urokinase (uPA). The purpose of this study was to investigate the role of uPAR in bone remodeling. MATERIALS AND METHODS: In vivo studies were performed in uPAR knockout (KO) and wildtype (WT) mice on a C57Bl6/SV129 (75:25) background. Bone mass was analyzed by pQCT. Excised tibias were subjected to mechanical tests. UPAR KO calvaria osteoblasts were characterized by proliferation assays, RT-PCR for important proteins secreted during differentiation, and immunoblot for activator protein 1 (AP-1) family members. In vitro osteoclast formation was tested with uPAR KO bone marrow monocytes in the presence of macrophage-colony stimulating factor (M-CSF) and RANKL. Phalloidin staining in osteoclasts served to study actin ring and podosome formation. RESULTS: pQCT revealed increased bone mass in uPAR-null mice. Mechanical tests showed reduced load-sustaining capability in uPAR KO tibias. uPAR KO osteoblasts showed a proliferative advantage with no difference in apoptosis, higher matrix mineralization, and earlier appearance of alkaline phosphatase (ALP). Surface RANKL expression at different stages of differentiation was not altered. AP-1 components, such as JunB and Fra-1, were upregulated in uPAR KO osteoblasts, along with other osteoblasts markers. On the resorptive side, the number of osteoclasts formed in vitro from uPAR KO monocytes was decreased. Podosome imaging in uPAR KO osteoclasts revealed a defect in actin ring formation. CONCLUSIONS: The defective proliferation and differentiation of bone cells, coincident with both aberrant expression of transcription factors and cytoskeletal organization, are typical uPAR-dependent molecular phenotypes, and we have now shown their function in osteoblasts and osteoclasts function in vivo. Udgivelsesdato: 2007-Sep

Furlan, Federico; Galbiati, Clara

2007-01-01

364

Inhibitory effect of berberine on the invasion of human lung cancer cells via decreased productions of urokinase-plasminogen activator and matrix metalloproteinase-2  

International Nuclear Information System (INIS)

Berberine, a compound isolated from medicinal herbs, has been reported with many pharmacological effects related to anti-cancer and anti-inflammation capabilities. In this study, we observed that berberine exerted a dose- and time-dependent inhibitory effect on the motility and invasion ability of a h