WorldWideScience
1

The ovine urokinase plasminogen activator and its receptor cDNAs: molecular cloning, characterization and expression in various tissues.  

Science.gov (United States)

The activation of plasminogen plays a crucial role in a variety of extracellular proteolytic events such as, fibrinolysis, cell migration, ovulation, involution of the mammary gland and the activation of other protease classes and growth factors. In this paper we describe the isolation of the full-length cDNAs of ovine urokinase plasminogen activator (u-PA) and its receptor (u-PAR) using a polymerase chain reaction based strategy. The ovine u-PA cDNA comprised of 2350 bp and it is characterized by a coding region of 1302 bp, and 5'- and 3'-UTR regions of 129 and 919 bp, respectively. The deduced amino acid sequence consists of 433 amino acids. The ovine u-PAR cDNA is comprised of 1247 bp and it is characterized by a coding region of 957 bp and 5'- and 3'-UTR regions of 44 and 246 bp respectively. The deduced amino acid sequence consists of 318 amino acids. Three-dimensional models of the putative protein products of both cDNAs showed that the proteins bear a high similarity with their human counterparts. Real-time PCR revealed high levels of u-PA expression in the adipose tissue, followed by that in mammary gland and kidney. Lower levels of expression were detected in the adrenal glands, heart, ovaries, spleen, liver and cerebellum. A similar pattern was observed in u-PAR expression with noticeably lower levels of expression in heart, liver and cerebellum. To the best of our knowledge, this is the first paper reporting expression of u-PA and u-PAR in the adipose tissue. These data strengthen the suggestion that adipose tissue functions as an endocrine organ besides an energy storage organ. Furthermore, u-PA and u-PAR mRNA levels were 7 and 8.5 fold higher respectively in involuting mammary tissue obtained from non-lactating ewes compared to that detected in mammary tissue obtained from lactating ewes. These data are consistent with the notion that upregulation of u-PA and u-PAR expression may play a key role in the process of involution of the mammary gland. PMID:19389465

Theodorou, Giorgos; Bizelis, Iosif; Rogdakis, Emmanuel; Politis, Ioannis

2009-08-15

2

The ovine plasminogen activator inhibitors type 1 and type 2 cDNAs: molecular cloning, characterization and expression in various tissues.  

Science.gov (United States)

Two serine proteases, urokinase and tissue type, control the activation of plasminogen to plasmin. These proteases are in turn specifically inhibited by plasminogen activator inhibitors type 1 and 2 (PAI-1 and PAI-2), both of which belong to the serine protease inhibitor (serpin) superfamily. Very little information is available on the role of PAI-1 and PAI-2 in ruminants, in mammary gland involution and in the adipose tissue. In this paper we describe the isolation of the full-length cDNAs of ovine PAI-1 and PAI-2 using a polymerase chain reaction based strategy. The ovine PAI-1 cDNA comprised of 1460bp and it is characterized by a coding region of 1209bp, and 5'- and 3'-UTR regions of 147 and 104bp, respectively. The deduced amino acid sequence consists of 402 amino acids. The ovine PAI-2 cDNA is comprised of 2128bp and it is characterized by a coding region of 957bp and 5'- and 3'-UTR regions of 58 and 819bp respectively. The deduced amino acid sequence consists of 416 amino acids. Three-dimensional models of the putative protein products of both cDNAs showed that the proteins bear a high similarity with their human counterparts. Real-time PCR revealed that the two inhibitors were predominantly expressed in the ovine mammary gland and adipose tissue. Furthermore, PAI-1 and PAI-2 mRNA levels were higher in the involuting mammary tissue and the adipose tissue obtained from non-lactating ewes compared to the corresponding values in tissues obtained from lactating ewes. These data are consistent with the notion that the plasminogen activation cascade plays a key role in involution of the mammary gland. The upregulation of expression of both inhibitors in the adipose tissue during the non-lactating period is a rather enigmatic observation. However, the expression of both inhibitors (PAI-1 and PAI-2) together with that of urokinase type plasminogen activator and its receptor previously reported by our group, strengthen the suggestion that the adipose tissue functions as an endocrine besides an energy storage organ. PMID:20114071

Theodorou, Giorgos; Bizelis, Iosif; Rogdakis, Emmanuel; Politis, Ioannis

2010-04-01

3

Functional Stability of Plasminogen Activator Inhibitor-1  

OpenAIRE

Plasminogen activator inhibitor-1 (PAI-1) is the main inhibitor of plasminogen activators, such as tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA), and a major regulator of the fibrinolytic system. PAI-1 plays a pivotal role in acute thrombotic events such as deep vein thrombosis (DVT) and myocardial infarction (MI). The biological effects of PAI-1 extend far beyond thrombosis including its critical role in fibrotic disorders, atherosclerosis, renal an...

Songul Yasar Yildiz; Pinar Kuru; Ebru Toksoy Oner; Mehmet Agirbasli

2014-01-01

4

Does Plasminogen Activator Inhibitor-1 Drive Lymphangiogenesis?  

OpenAIRE

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...

Bruyere, Francoise; Melen-lamalle, Laurence; Blacher, Silvia; Detry, Benoi?t; Masset, Anne; Lecomte, Julie; Lambert, Vincent; Maillard, Catherine; Hoyer-hansen, Gunilla; Lund, Leif R.; Foidart, Jean-michel; Noe?l, Agne?s

2010-01-01

5

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

6

Antipsoriatic therapies inhibit epidermal plasminogen activator activity.  

Science.gov (United States)

Urokinase (UK, Mr 55,000) and tissue-type plasminogen activator (tPA, Mr 74,000) are serine proteinases involved in many biological processes, ie, cell migration, neoplastic transformation, and extracellular proteolysis. Cutaneous fibrinolytic activity (dependent on the activity of UK and tPA) was studied with the autohistographic fibrin film method in 40 patients affected by psoriasis vulgaris before and after topical (anthralin, betamethasone valerate, hydrocolloid occlusive dressing) or systemic psoralen-ultraviolet-light (PUVA) treatments. Autohistographic studies also were performed after apposition of monoclonal antibodies directed against the catalytic site of UK and tPA. Finally, UK and tPA were localized immunohistochemically in the psoriatic plaques and in controls using the immunoperoxidase procedure based on the biotin/avidin system. UK and tPA immunoreactivity was present in the cytoplasm and around the outlines of keratinocytes in the psoriatic patches before treatment and in the patches not cleared after treatment, while it was not detectable in normal epidermis, in the unaffected psoriatic epidermis, and in the cleared psoriatic skin. Cutaneous fibrinolytic activity was present in the cases in which UK and tPA were detected histochemically and, in the psoriatic epidermis, it was abolished by preincubation with anti-tPA but not with anti-UK antibodies. This study suggests that established topical and systemic treatments for psoriasis possess UK and tPA antagonist activity. PMID:2121655

Lotti, T; Bonan, P; Cannarozzo, G; Fedi, A M; Panconesi, E

1990-09-01

7

Plasminogen activator and its assay of the activity  

International Nuclear Information System (INIS)

Plasminogen activators (PA) are specific proteolytic enzymes. Which convert the inactive proenzyme to plasmin. The plasmin formed is a potent and nonspecific protease which cleaves blood fibrin clots into soluble polypeptide. The author describes some biochemical characteristic of the different components of the plasminogen activator system, current methods for assay of the activity of the PA. The potential application of PA as diagnostic tools in diseases of the thrombi

8

Ovulation efficiency is reduced in mice that lack plasminogen activator gene function: functional redundancy among physiological plasminogen activators.  

OpenAIRE

Several lines of indirect evidence suggest that plasminogen activation plays a crucial role in degradation of the follicular wall during ovulation. However, single-deficient mice lacking tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA), or PA inhibitor type 1(PAI-1) gene function were recently found to have normal reproduction, although mice with a combined deficiency of tPA and uPA were significantly less fertile. To investigate whether the reduced fertilit...

Leonardsson, G.; Peng, X. R.; Liu, K.; Nordstro?m, L.; Carmeliet, P.; Mulligan, R.; Collen, D.; Ny, T.

1995-01-01

9

Activation of immobilized plasminogen by tissue activator. Multimolecular complex formation  

International Nuclear Information System (INIS)

Ternary complex formation of tissue plasminogen activator (TPA) and plasminogen (Plg) with thrombospondin (TSP) or histidine-rich glycoprotein (HRGP) has been demonstrated using an enzyme-linked immunosorbent assay, an affinity bead assay, and a rocket immunoelectrophoresis assay. The formation of these complexes was specific, concentration dependent, saturable, lysine binding site-dependent, and inhibitable by fluid phase plasminogen. Apparent Kd values were approximately 12-36 nM for the interaction of TPA with TSP-Plg complexes and 15-31 nM with HRGP-Plg complexes. At saturation the relative molar stoichiometry of Plg:TPA was 3:1 within the TSP-containing complexes and 1:1 within HRGP-containing complexes. The activation of Plg to plasmin by TPA on TSP- and HRGP-coated surfaces was studied using a synthetic fluorometric plasmin substrate (D-Val-Leu-Lys-7-amino-4-trifluoromethyl coumarin). Kinetic analysis demonstrated a marked increase in the affinity of TPA for plasminogen in the presence of surface-associated TSP or HRGP. Complex formation of locally released tissue plasminogen activator with Plg immobilized on TSP or HRGP surfaces may thus play an important role in effecting proteolytic events in nonfibrin-containing microenvironments

10

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

OpenAIRE

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

11

Epithelial glycoprotein-330 mediates endocytosis of plasminogen activator-plasminogen activator inhibitor type-1 complexes  

DEFF Research Database (Denmark)

Epithelial glycoprotein 330 (gp330) is structurally similar to the multifunctional alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein (alpha 2MR/LRP), gp330 and alpha 2MR/LRP bind Ca2+ with high affinity, and both receptors bind and mediate endocytosis of alpha 2MR-associated protein (RAP). In the present report, we describe that affinity-purified gp330 from rabbit renal cortex binds plasminogen activator inhibitor type-1 (PAI-1) complexed with urokinase-type plasminogen activator (uPA). alpha 2M-methylamine, which binds with high affinity to alpha 2MR/LRP, did not bind to gp330. The apparent Kd for binding of uPA.PAI-1 complexes was about 0.8 nM at 4 degrees C. The binding was calcium-dependent and inhibited by recombinant RAP (rRAP) and tissue type plasminogen activator-PAI-1 complexes. Thin sections of rabbit renal proximal tubules bound 125I-labeled uPA.PAI-1 and rRAP in the apical part of proximal tubules corresponding to the localization of gp330. The binding of 125I-uPA.PAI-1 complexes in tubules was abolished by excess unlabeled rRAP, and a rRAP-inhibitable endocytosis and degradation of labeled uPA.PAI-1 complexes was demonstrated by perfusion of isolated rabbit proximal tubules. The results establish an endocytotic function of gp330 and suggest that gp330 is an important component of the fibrinolytic system in gp330-containing epithelial as found in, for example, kidney and lung.

Moestrup, S K; Nielsen, Susanne

1993-01-01

12

A role for tissue plasminogen activator in thrombotic thrombocytopenic purpura.  

Science.gov (United States)

Thrombotic thrombocytopenic purpura (TTP) is a life-threatening disease characterized by generalized microvascular occlusion. TTP has been related to severe deficiency of ADAMTS13, an enzyme that cleaves von Willebrand factor multimers into less adhesive molecules. However, ADAMTS13 deficiency correlates poorly with severity of thrombocytopenia or microangiopathic hemolysis, with the frequency of neurologic complications or the response to plasma exchange. Also, some patients with severe hereditary ADAMTS13 deficiency consistently relapse every few weeks, whereas others remain asymptomatic into their forties. Taken together, these findings suggest that an additional element is missing in the pathophysiology of TTP. We postulate that both low ADAMTS13 activity and low tissue-plasminogen activator activity are required to trigger TTP attacks. Tissue-plasminogen activator end product, plasmin, extensively degrades von Willebrand factor, breaking-down the bonds between platelets and the blood vessel wall, so that low tissue-plasminogen activator activity prevents a mechanism similar to that of ADAMTS13. The hypothesis that low tissue-plasminogen activator activity plays an important role in TTP pathogenesis is further substantiated by TTP comorbidity. Problems prevalent in patients with TTP attacks or with long-term TTP remission, including increased body mass index, major depression, cognitive abnormalities, hypertension, and premature death, are somehow associated with low tissue-plasminogen activator activity. PMID:25459148

Hoirisch-Clapauch, Silvia; Nardi, Antonio Egidio

2014-12-01

13

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; Pedersen, Katrine Egelund

14

Role of residue Y99 in tissue plasminogen activator.  

OpenAIRE

The crystal structure of the fibrinolytic enzyme tissue plasminogen activator (tPA) shows that the bulky side chain of Y99 hinders access to the active site by partially occluding the S2 site and may be responsible for the low catalytic activity of tPA toward plasminogen. We have tested the role of Y99 by replacing it with Leu, the residue found in more proficient proteases like trypsin and thrombin. The Y99L replacement results in an increase in the k(cat)/Km for chromogenic substrates due t...

Vindigni, A.; Winfield, M.; Ayala, Y. M.; Di Cera, E.

2000-01-01

15

4G/5G Plasminogen Activator Inhibitor-1 Polymorphisms and Haplotypes Are Associated with Pneumonia  

OpenAIRE

Rationale: Plasminogen activator inhibitor (PAI)-1 inhibits urokinase and tissue plasminogen activator, required for host response to infection. Whether variation within the PAI-1 gene is associated with increased susceptibility to infection is unknown.

Yende, Sachin; Angus, Derek C.; Ding, Jingzhong; Newman, Anne B.; Kellum, John A.; Li, Rongling; Ferrell, Robert E.; Zmuda, Joseph; Kritchevsky, Stephen B.; Harris, Tamara B.; Garcia, Melissa; Yaffe, Kristine; Wunderink, Richard G.

2007-01-01

16

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-Å 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 ?-sheet B, and the 147-loop directly contacts PAI-1 ?-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

17

Dehydroepiandrosterone reduces plasma plasminogen activator inhibitor type 1 and tissue plasminogen activator antigen in men.  

Science.gov (United States)

Dehydroepiandrosterone (DHEA) may help prevent heart disease in men. To test the hypothesis that DHEA might exert its effects by enhancing endogenous fibrinolytic potential, a double-blind, placebo-controlled study was conducted that assessed the effects of DHEA administration on plasma plasminogen activator inhibitor type 1 (PAI-1) and tissue plasminogen activator (tPA) antigen. Eighteen men received 50 mg DHEA orally and 16 men received a placebo capsule thrice daily for 12 days. Serum DHEA-sulfate and plasma PAI-1 and tPA antigen were measured before and after treatment. In the DHEA group, serum DHEA-sulfate (from 7.5 +/- 1.2 micromol/L to 20.2 +/- 1.5 micromol/L (P < 0.0001), androstenedione (from 2.6 +/- 0.2 nmol/L to 4.0 +/- 0.4 nmol/L; P < 0.005) and estrone (from 172 +/- 21 pmol/L to 352 +/- 28 pmol/L; P < 0.005) increased, whereas plasma PAI-1 (from 55.4 +/- 3.8 ng/mL to 38.6 +/- 3.3 ng/mL; P < 0.0001) and tPA antigen (from 8.1 +/- 1.9 ng/mL to 5.4 +/- 1.3 ng/mL; P < 0.0005) decreased. In the placebo group, serum DHEA-sulfate declined slightly from 8.0 +/- 3.3 micromol/L to 7.3 +/- 3.4 micromol/L (P < 0.05), but no other measured steroid changed. Plasma PAI-1 and tPA antigen did not change in the placebo group. These findings suggest that DHEA administration reduces plasma PAI-1 and tPA antigen concentrations in men. PMID:8615394

Beer, N A; Jakubowicz, D J; Matt, D W; Beer, R M; Nestler, J E

1996-05-01

18

Photonic Activation of Plasminogen Induced by Low Dose UVB.  

Science.gov (United States)

Activation of plasminogen to its active form plasmin is essential for several key mechanisms, including the dissolution of blood clots. Activation occurs naturally via enzymatic proteolysis. We report that activation can be achieved with 280 nm light. A 2.6 fold increase in proteolytic activity was observed after 10 min illumination of human plasminogen. Irradiance levels used are in the same order of magnitude of the UVB solar irradiance. Activation is correlated with light induced disruption of disulphide bridges upon UVB excitation of the aromatic residues and with the formation of photochemical products, e.g. dityrosine and N-formylkynurenine. Most of the protein fold is maintained after 10 min illumination since no major changes are observed in the near-UV CD spectrum. Far-UV CD shows loss of secondary structure after illumination (33.4% signal loss at 206 nm). Thermal unfolding CD studies show that plasminogen retains a native like cooperative transition at ~70 ºC after UV-illumination. We propose that UVB activation of plasminogen occurs upon photo-cleavage of a functional allosteric disulphide bond, Cys737-Cys765, located in the catalytic domain and in van der Waals contact with Trp761 (4.3 Å). Such proximity makes its disruption very likely, which may occur upon electron transfer from excited Trp761. Reduction of Cys737-Cys765 will result in likely conformational changes in the catalytic site. Molecular dynamics simulations reveal that reduction of Cys737-Cys765 in plasminogen leads to an increase of the fluctuations of loop 760-765, the S1-entrance frame located close to the active site. These fluctuations affect the range of solvent exposure of the catalytic triad, particularly of Asp646 and Ser74, which acquire an exposure profile similar to the values in plasmin. The presented photonic mechanism of plasminogen activation has the potential to be used in clinical applications, possibly together with other enzymatic treatments for the elimination of blood clots. PMID:25635856

Correia, Manuel; Snabe, Torben; Thiagarajan, Viruthachalam; Petersen, Steffen Bjørn; Campos, Sara R R; Baptista, António M; Neves-Petersen, Maria Teresa

2015-01-01

19

Nuclear translocation of urokinase-type plasminogen activator  

OpenAIRE

Urokinase-type plasminogen activator (uPA) participates in diverse (patho)physiological processes through intracellular signaling events that affect cell adhesion, migration, and proliferation, although the mechanisms by which these occur are only partially understood. Here we report that upon cell binding and internalization, single-chain uPA (scuPA) translocates to the nucleus within minutes. Nuclear translocation does not involve proteolytic activation or degradation of scuPA. Neither the ...

Stepanova, Victoria; Lebedeva, Tatiana; Kuo, Alice; Yarovoi, Serge; Tkachuk, Sergei; Zaitsev, Sergei; Bdeir, Khalil; Dumler, Inna; Marks, Michael S.; Parfyonova, Yelena; Tkachuk, Vsevolod A.; Higazi, Abd Al-roof; Cines, Douglas B.

2008-01-01

20

Thrombin-specific inactivation of endothelial cell derived plasminogen activator  

International Nuclear Information System (INIS)

Although thrombin (T) has diverse functions in the overall hemostatic mechanism, relatively little is known about its direct effect on components of the fibrinolytic enzyme system. The authors have investigated the interaction of T with plasminogen activators (PA) derived from bovine aortic endothelial cells (EC) in culture (2-5th passage, preconfluent monolayers). Varying concentrations of purified bovine or human thrombin were added to EC-conditioned media (CM). CM + T mixtures were assayed at various times for PA activity using purified plasminogen and a sensitive 125I-fibrinogenolytic or caseinolytic assay. T (5 nM), but not plasmin or trypsin at equivalent concentrations, resulted in a time-dependent inhibition of the PA activity in CM. T had no effect on the PA activity of urokinase, streptokinase or preformed plasmin. The ability of T to inactivate the EC-derived PA was abolished by prior treatment of T with active site-directed reagents. SDS-PAGE and zymography with copolymerized fibrinogen and plasminogen revealed further specificity in that only one of the multiple-molecular weight forms of PA present in EC-CM was inactivated by T. The authors conclude that in a highly specific fashion, T inactivates the predominant PA present in EC-CM by limited proteolysis. Thus, another potentially important function of T is suggested which may have particular significance in the temporal regulation of coagulation and fibrinolysis at the blood-endothelium interface

21

Immunohistochemical localisation of tissue plasminogen activator in human brain tumours.  

OpenAIRE

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

22

Intraventricular recombinant tissue plasminogen activator for lysis of intraventricular haemorrhage.  

OpenAIRE

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

23

Imaging active urokinase plasminogen activator in prostate cancer.  

Science.gov (United States)

The increased proteolytic activity of membrane-bound and secreted proteases on the surface of cancer cells and in the transformed stroma is a common characteristic of aggressive metastatic prostate cancer. We describe here the development of an active site-specific probe for detecting a secreted peritumoral protease expressed by cancer cells and the surrounding tumor microenvironment. Using a human fragment antigen-binding phage display library, we identified a human antibody termed U33 that selectively inhibited the active form of the protease urokinase plasminogen activator (uPA, PLAU). In the full-length immunoglobulin form, U33 IgG labeled with near-infrared fluorophores or radionuclides allowed us to noninvasively detect active uPA in prostate cancer xenograft models using optical and single-photon emission computed tomography imaging modalities. U33 IgG labeled with (111)In had a remarkable tumor uptake of 43.2% injected dose per gram (%ID/g) 72 hours after tail vein injection of the radiolabeled probe in subcutaneous xenografts. In addition, U33 was able to image active uPA in small soft-tissue and osseous metastatic lesions using a cardiac dissemination prostate cancer model that recapitulated metastatic human cancer. The favorable imaging properties were the direct result of U33 IgG internalization through an uPA receptor-mediated mechanism in which U33 mimicked the function of the endogenous inhibitor of uPA to gain entry into the cancer cell. Overall, our imaging probe targets a prostate cancer-associated protease, through a unique mechanism, allowing for the noninvasive preclinical imaging of prostate cancer lesions. Cancer Res; 75(7); 1225-35. ©2015 AACR. PMID:25672980

LeBeau, Aaron M; Sevillano, Natalia; Markham, Kate; Winter, Michael B; Murphy, Stephanie T; Hostetter, Daniel R; West, James; Lowman, Henry; Craik, Charles S; VanBrocklin, Henry F

2015-04-01

24

Blood urokinase plasminogen activator system in chronic urticaria.  

Science.gov (United States)

The evidence gathered has pointed to the fibrinolytic system, which apart from its major role in hemostasis may also be involved in inflammatory and immune processes. To understand better the role of fibrinolysis in urticaria, we measured plasma levels of the urokinase system associated molecules such as urokinase-type plasminogen activator (uPA), its soluble receptor (suPAR; CD87) and an inhibitor, plasminogen activator inhibitor type 1 (PAI-1) activity in chronic urticaria (CU) patients. Plasma was obtained from symptomatic sixteen CU patients (12 females and 4 males) showing positive response to autologous serum skin test (ASST), 28 CU patients with negative ASST (20 females and 8 males) as well as from healthy subjects matched by sex and age. The plasma level of uPA and suPAR antigens, PAI-1 activity did not differ significantly among the three subjects groups. The data obtained suggest that CU patients showing positive response to ASST have plasma profile of the urokinase system-associated proteins, which is not markedly different as compared with CU patients with negative ASST as well as healthy subjects. Our findings have also confirmed the earlier studies, suggesting that systemic fibrinolysis may not be involved in chronic urticaria. PMID:17102952

Kasperska-Zajac, Alicja; Brzoza, Zenon; Rogala, Barbara

2007-01-01

25

Downregulation of urokinase-type plasminogen activator and plasminogen activator inhibitor-1 by grape seed proanthocyanidin extract.  

Science.gov (United States)

Urokinase plasminogen activator (uPA) system, comprising of uPA, its receptor uPAR and inhibitor, type 1 plasminogen activator inhibitor (PAI-1), plays a vital role in various biological processes involving extracellular proteolysis, fibrinolysis, cell migration and proliferation. The timely occurence of these processes are essential for normal wound healing. This study examines the regulation of uPA and PAI-1 by a natural polyphenol-rich compound, grape seed extract (GSE). GSE is reported to have beneficial effects in promoting wound healing. Fibroblast cells exposed to different doses of GSE for 18hours were processed for further studies such as ELISA, RT-PCR, western blotting, fibrinolytic assay, cell surface plasmin activity assay and in vitro wound healing assay. GSE treatment caused a significant downregulation of uPA and PAI-1 expression, both at the RNA and protein levels. ELISA also revealed a dose-dependent decrease in uPA and PAI-1 activities. Functional significance of the downregulation was evident in decreased fibrinolytic activity, concomittant with decreased cell-surface plasmin activity. In vitro wound healing studies showed that GSE also retarded the migration of cells towards the wounded region. PMID:19640694

Sandra, Dhungana; Radha, Madhyastha; Harishkumar, Madhyastha; Yuichi, Nakajima; Sayuri, Omura; Masugi, Maruyama

2010-01-01

26

[Polymorphism of the plasminogen activator inhibitor type 1 gene, plasminogen level and thrombosis in patients with antiphospholipid syndrome].  

Science.gov (United States)

The frequency of venous and arterial thromboses and plasminogen level were investigated in 78 patients with antiphospholipid syndrome (APS), 35 of whom with systemic lupus erythematosus (SLE+APS) and 43 - with primary APS (PAPS). The levels and genotype of plasminogen activator inhibitor type 1 (PAI-1) were determined in 45 patients with APS, of whom 21 patients with SLE + APS and 24 patients with PAPS. A control group included 10 healthy individuals without autoimmune disease signs and thromboses on period of investigation and in past history. It was shown for the first time that for one third of 67 patients with APS and thromboses high positive levels of antiphospholipid antibodies (aPL) are associated with low plasminogen levels. The levels of PAI-1 antigen measured by ELIZA method, which detects active, latent and bound with plasminogen activator PAI-1, were opposed with frequency of thromboses in APS patients. Correlation between the high and increased levels of PAI-1 and high positive aPL levels was found for one third of 43 patients with APS and thrombosis. One of the possible mechanisms of this interconnection was considered. It was shown that arterial and, to a more extent, venous thromboses are associated with the 4G/5G polymorphism of PAI-1 gene and high plasma level of the inhibitor in 79% of APS patients. At the presence of the 4G allele patients with SLE+APS had higher PAI-1 levels than patients with PAPS. The obtained results show that measuring the levels of plasminogen and PAI-1 as well as the 4G/5G polymorphism of PAI-1 gene which is associated with thromboses may have the practical significance for identification of high risk of thrombosis in APS patients. PMID:24749249

A?sina, R B; Mukhametova, L I; Ostriakova, E V; Seredavkina, N V; Patrushev, L I; Patrusheva, N L; Reshetniak, T M; Gulin, D A; Gershkovich, K B; Nasonov, E L; Varfolomeev, S D

2014-01-01

27

Comparative molecular analysis of ovine and bovine Streptococcus uberis isolates.  

Science.gov (United States)

Streptococcus uberis causes clinical and subclinical mastitis in cattle and sheep, but it is unknown whether the composition of Strep. uberis populations differs between host species. To address this, we characterized a collection of bovine and ovine Strep. uberis isolates with shared geographical and temporal origins by means of an expanded multilocus sequence typing scheme. Among 14 ovine and 35 bovine isolates, 35 allelic profiles were detected. Each allelic profile was associated with a single host species and all but one were new to the multilocus sequence typing database. The median number of new alleles per isolate was higher for ovine isolates than for bovine isolates. None of the ovine isolates belonged to the global clonal complexes 5 or 143, which are commonly associated with bovine mastitis and which have a wide geographical distribution. Ovine isolates also differed from bovine isolates in carriage of plasminogen activator genes, with significantly higher prevalence of pauB in ovine isolates. Isolates that were negative for yqiL, one of the targets of multilocus sequence typing, were found among ovine and bovine isolates and were not associated with a specific sequence type or global clonal complex. One bovine isolate carried a gapC allele that was probably acquired through lateral gene transfer, most likely from Streptococcus salivarius. We conclude that ovine isolates are distinct from bovine isolates of Strep. uberis, and that recombination between isolates from different host species or bacterial species could contribute to changes in virulence gene profiles with relevance for vaccine development. PMID:23200465

Gilchrist, T L; Smith, D G E; Fitzpatrick, J L; Zadoks, R N; Fontaine, M C

2013-02-01

28

Arrhenius temperature dependence of in vitro tissue plasminogen activator thrombolysis  

International Nuclear Information System (INIS)

Stroke is a devastating disease and a leading cause of death and disability. Currently, the only FDA approved therapy for acute ischemic stroke is the intravenous administration of the thrombolytic medication, recombinant tissue plasminogen activator (tPA). However, this treatment has many contraindications and can have dangerous side effects such as intra-cerebral hemorrhage. These treatment limitations have led to much interest in potential adjunctive therapies, such as therapeutic hypothermia (T ? 35 deg. C) and ultrasound enhanced thrombolysis. Such interest may lead to combining these therapies with tPA to treat stroke, however little is known about the effects of temperature on the thrombolytic efficacy of tPA. In this work, we measure the temperature dependence of the fractional clot mass loss ?m(T) resulting from tPA exposure in an in vitro human clot model. We find that the temperature dependence is well described by an Arrhenius temperature dependence with an effective activation energy Eeff of 42.0 ± 0.9 kJ mole-1. Eeff approximates the activation energy of the plasminogen-to-plasmin reaction of 48.9 kJ mole-1. A model to explain this temperature dependence is proposed. These results will be useful in predicting the effects of temperature in future lytic therapies

29

Arrhenius temperature dependence of in vitro tissue plasminogen activator thrombolysis  

Energy Technology Data Exchange (ETDEWEB)

Stroke is a devastating disease and a leading cause of death and disability. Currently, the only FDA approved therapy for acute ischemic stroke is the intravenous administration of the thrombolytic medication, recombinant tissue plasminogen activator (tPA). However, this treatment has many contraindications and can have dangerous side effects such as intra-cerebral hemorrhage. These treatment limitations have led to much interest in potential adjunctive therapies, such as therapeutic hypothermia (T {<=} 35 deg. C) and ultrasound enhanced thrombolysis. Such interest may lead to combining these therapies with tPA to treat stroke, however little is known about the effects of temperature on the thrombolytic efficacy of tPA. In this work, we measure the temperature dependence of the fractional clot mass loss {delta}m(T) resulting from tPA exposure in an in vitro human clot model. We find that the temperature dependence is well described by an Arrhenius temperature dependence with an effective activation energy E{sub eff} of 42.0 {+-} 0.9 kJ mole{sup -1}. E{sub eff} approximates the activation energy of the plasminogen-to-plasmin reaction of 48.9 kJ mole{sup -1}. A model to explain this temperature dependence is proposed. These results will be useful in predicting the effects of temperature in future lytic therapies.

Shaw, George J [Department of Emergency Medicine, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0769 (United States); Dhamija, Ashima [Department of Emergency Medicine, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0769 (United States); Bavani, Nazli [Department of Emergency Medicine, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0769 (United States); Wagner, Kenneth R [Department of Neurology, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0769 (United States); Holland, Christy K [Department of Biomedical Engineering, University of Cincinnati College of Medicine, Cincinnati, OH 45267-0769 (United States)

2007-06-07

30

Keeping the blood flowing—plasminogen activator genes and feeding behavior in vampire bats  

Science.gov (United States)

The blood feeding vampire bats emerged from New World leaf-nosed bats that fed on fruit and insects. Plasminogen activator, a serine protease that regulates blood coagulation, is known to be expressed in the saliva of Desmodus rotundus (common vampire bat) and is thought to be a key enzyme for the emergence of blood feeding in vampire bats. To better understand the evolution of this biological function, we studied the plasminogen activator (PA) genes from all vampire bat species in light of their feeding transition to bird and subsequently mammalian blood. We include the rare species Diphylla ecaudata and Diaemus youngi, where plasminogen activator had not previously been studied and demonstrate that PA gene duplication observed in Desmodus is not essential to the vampire phenotype, but relates to the emergence of predominant mammalian blood feeding in this species. Plasminogen activator has evolved through gene duplication, domain loss, and sequence evolution leading to change in fibrin-specificity and susceptibility to plasminogen activator inhibitor-1. Before undertaking this study, only the four plasminogen activator isoforms from Desmodus were known. The evolution of vampire bat plasminogen activators can now be linked phylogenetically to the transition in feeding behavior among vampire bat species from bird to mammalian blood.

Tellgren-Roth, Åsa; Dittmar, Katharina; Massey, Steven E.; Kemi, Cecilia; Tellgren-Roth, Christian; Savolainen, Peter; Lyons, Leslie A.; Liberles, David A.

2009-01-01

31

Keeping the blood flowing-plasminogen activator genes and feeding behavior in vampire bats.  

Science.gov (United States)

The blood feeding vampire bats emerged from New World leaf-nosed bats that fed on fruit and insects. Plasminogen activator, a serine protease that regulates blood coagulation, is known to be expressed in the saliva of Desmodus rotundus (common vampire bat) and is thought to be a key enzyme for the emergence of blood feeding in vampire bats. To better understand the evolution of this biological function, we studied the plasminogen activator (PA) genes from all vampire bat species in light of their feeding transition to bird and subsequently mammalian blood. We include the rare species Diphylla ecaudata and Diaemus youngi, where plasminogen activator had not previously been studied and demonstrate that PA gene duplication observed in Desmodus is not essential to the vampire phenotype, but relates to the emergence of predominant mammalian blood feeding in this species. Plasminogen activator has evolved through gene duplication, domain loss, and sequence evolution leading to change in fibrin-specificity and susceptibility to plasminogen activator inhibitor-1. Before undertaking this study, only the four plasminogen activator isoforms from Desmodus were known. The evolution of vampire bat plasminogen activators can now be linked phylogenetically to the transition in feeding behavior among vampire bat species from bird to mammalian blood. PMID:18791694

Tellgren-Roth, Asa; Dittmar, Katharina; Massey, Steven E; Kemi, Cecilia; Tellgren-Roth, Christian; Savolainen, Peter; Lyons, Leslie A; Liberles, David A

2009-01-01

32

Transgenic chickens expressing human urokinase-type plasminogen activator.  

Science.gov (United States)

Urokinase-type plasminogen activator is a serine protease that is clinically used in humans for the treatment of thrombolytic disorders and vascular diseases such as acute ischemic stroke and acute peripheral arterial occlusion. This study explored the feasibility of using chickens as a bioreactor for producing human urokinase-type plasminogen activator (huPA). Recombinant huPA gene, under the control of a ubiquitous Rous sarcoma virus promoter, was injected into the subgerminal cavity of freshly laid chicken eggs at stage X using the replication-defective Moloney murine leukemia virus (MoMLV)-based retrovirus vectors encapsidated with VSV-G (vesicular stomatitis virus G) glycoprotein. A total of 38 chicks, out of 573 virus-injected eggs, hatched and contained the huPA gene in their various body parts. The mRNA transcript of the huPA gene was present in various organs, including blood and egg, and was germ-line transmitted to the next generation. The level of active huPA protein was 16-fold higher in the blood of the transgenic chicken than in the nontransgenic chicken (P volume of semen ejaculate, sperm concentration, and sperm viability. Taken together, our data suggest that huPA transgenic chickens could be successfully produced by the retroviral vector system. Transgenic chickens, expressing the huPA under the control of a ubiquitous promoter, may not only be used as a bioreactor for pharming of the huPA drug but also be useful for studying huPA-induced bleeding and other disorders. PMID:23960123

Lee, Sung Ho; Gupta, Mukesh Kumar; Ho, Young Tae; Kim, Teoan; Lee, Hoon Taek

2013-09-01

33

Novel protein interactors of urokinase-type plasminogen activator receptor.  

Science.gov (United States)

The urokinase-type plasminogen activator receptor (uPAR) has been implicated in tumor growth and metastasis. The crystal structure of uPAR revealed that the external surface is largely free to interact with a number of proteins. Additionally, due to absence of an intracellular cytoplasmic protein domain, many of the biological functions of uPAR necessitate interactions with other proteins. Here, we used yeast two-hybrid screening of breast cancer cDNA library to identify hSpry1 and HAX1 proteins as putative candidate proteins that interact with uPAR bait constructs. Interaction between these two candidates and uPAR was confirmed by GST-pull down, co-immunoprecipitation assays and confocal microscopy. These novel interactions that have been identified may also provide further evidence that uPAR can interact with a number of other proteins which may influence a range of biological functions. PMID:20696135

Mekkawy, Ahmed H; De Bock, Charles E; Lin, Zhen; Morris, David L; Wang, Yao; Pourgholami, Mohammad H

2010-09-01

34

Mutant and chimeric recombinant plasminogen activators. Production in eukaryotic cells and preliminary characterization.  

OpenAIRE

Mutant urokinase-type plasminogen activator (u-PA) genes and hybrid genes between tissue-type plasminogen activator (t-PA) and u-PA have been designed to direct the synthesis of new plasminogen activators and to investigate the structure-function relationship in these molecules. The following classes of constructs were made starting from cDNA encoding human t-PA or u-PA: 1) u-PA mutants in which the Arg156 and Lys158 were substituted with threonine, thus preventing cleavage by thrombin and pl...

Pierard, Luc; Jacobs, P.; Gheysen, D.; Hoylaerts, M.; Andre, B.; Topisirovic, L.; Cravador, A.; Foresta, F.; Herzog, A.; Collen, D.

1987-01-01

35

Mice deficient in plasminogen activator inhibitor-1 have improved skeletal muscle regeneration  

Science.gov (United States)

journal article, "Mice deficient in plasminogen activator inhibitor-1 have improved skeletal muscle regeneration," by Timothy Koh, Scott Bryer, Augustina Pucci, and Thomas Sisson, found in the American Journal of Physiology-Cell Physiology.

PhD Timothy J. Koh (University of Illinois at Chicago Department of Movement Sciences)

2005-07-01

36

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

37

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

OpenAIRE

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. This 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 f...

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

2012-01-01

38

Functional properties of the recombinant kringle-2 domain of tissue plasminogen activator produced in Escherichia coli  

International Nuclear Information System (INIS)

The kringle-2 domain (residues 176-262) of tissue-type plasminogen activator (t-PA) was cloned and expressed in Escherichia coli. The recombinant peptide, which concentrated in cytoplasmic inclusion bodies, was isolated, solubilized, chemically refolded, and purified by affinity chromatography on lysine-Sepharose to apparent homogeneity. [35S]Cysteine-methionine-labeled polypeptide was used to study the interactions of kringle-2 with lysine, fibrin, and plasminogen activator inhibitor-1. The kringle-2 domain bound to lysine-Sepharose and to preformed fibrin with a Kd = 104 +/- 6.2 microM (0.86 +/- 0.012 binding site) and a Kd = 4.2 +/- 1.05 microM (0.80 +/- 0.081 binding site), respectively. Competition experiments and direct binding studies showed that the kringle-2 domain is required for the formation of the ternary t-PA-plasminogen-intact fibrin complex and that the association between the t-PA kringle-2 domain and fibrin does not require plasmin degradation of fibrin and exposure of new COOH-terminal lysine residues. We also observed that kringle-2 forms a complex with highly purified guanidine-activated plasminogen activator inhibitor-1, dissociable by 0.2 M epsilon-aminocaproic acid. The kringle-2 polypeptide significantly inhibited tissue plasminogen activator/plasminogen activator inhibitor-1 interaction. The kringle-2 domain bound to plasminogen activator inhibitor-1 in a specific and saturable manner with a Kd = 0.51 +/- 0.055 microM (0.35 +/- 0.026 binding +/- 0.055 microM (0.35 +/- 0.026 binding site). Therefore, the t-PA kringle-2 domain is important for the interaction of t-PA not only with fibrin, but also with plasminogen activator inhibitor-1 and thus represents a key structure in the regulation of fibrinolysis

39

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

40

Characterization of a recombinant fusion protein of the finger domain of tissue-type plasminogen activator with a truncated single chain urokinase-type plasminogen activator.  

OpenAIRE

Human recombinant single chain urokinase-type plasminogen activator (recombinant scu-PA) and a hybrid between human tissue-type plasminogen activator (t-PA) and scu-PA, obtained by ligation of cDNA fragments encoding the NH2-terminal region (amino acids 1-67) of t-PA and the COOH-terminal region (amino acids 136-411) of scu-PA, were expressed in a mammalian cell system. The proteins were purified from conditioned culture media containing 2% fetal calf serum by chromatography on zinc chelate-S...

Gheysen, D.; Lijnen, H. R.; Pierard, Luc; Foresta, F.; Demarsin, E.; Jacobs, P.; Wilde, Marc; Bollen, A.; Collen, D.

1987-01-01

41

Effects of Korean Red Ginseng extract on tissue plasminogen activator and plasminogen activator inhibitor-1 expression in cultured rat primary astrocytes  

OpenAIRE

Korean Red Ginseng (KRG) is an oriental herbal preparation obtained from Panax ginseng Meyer (Araliaceae). To expand our understanding of the action of KRG on central nervous system (CNS) function, we examined the effects of KRG on tissue plasminogen activator (tPA)/plasminogen activator inhibitor-1 (PAI-1) expression in rat primary astrocytes. KRG extract was treated in cultured rat primary astrocytes and neuron in a concentration range of 0.1 to 1.0 mg/mL and the expression of functional tP...

Ko, Hyun Myung; Joo, So Hyun; Kim, Pitna; Park, Jin Hee; Kim, Hee Jin; Bahn, Geon Ho; Kim, Hahn Young; Lee, Jongmin; Han, Seol-heui; Shin, Chan Young; Park, Seung Hwa

2013-01-01

42

Rapid purification of a high-affinity plasminogen activator from human blood plasma by specific adsorption on fibrin/Celite.  

OpenAIRE

A preparation of fibrin precipitated over a solid Celite (diatomaceous earth) matrix that selectively binds 50-70% of the plasminogen activator present in human blood plasma is described. Affinity chromatography of plasma on fibrin/Celite followed by gel filtration led to a 29,000-fold purification of the plasminogen activator. The activator, referred to as the high-affinity plasminogen activator, is characterized by its ability to be strongly adsorbed by fibrin. Smaller amounts of other plas...

Husain, S. S.; Lipinski, B.; Greuwich, V.

1981-01-01

43

Regulation of epithelial sodium channels in urokinase plasminogen activator deficiency.  

Science.gov (United States)

Epithelial sodium channels (ENaC) govern transepithelial salt and fluid homeostasis. ENaC contributes to polarization, apoptosis, epithelial-mesenchymal transformation, etc. Fibrinolytic proteases play a crucial role in virtually all of these processes and are elaborated by the airway epithelium. We hypothesized that urokinase-like plasminogen activator (uPA) regulates ENaC function in airway epithelial cells and tested that possibility in primary murine tracheal epithelial cells (MTE). Both basal and cAMP-activated Na(+) flow through ENaC were significantly reduced in monolayers of uPA-deficient cells. The reduction in ENaC activity was further confirmed in basolateral membrane-permeabilized cells. A decrease in the Na(+)-K(+)-ATPase activity in the basolateral membrane could contribute to the attenuation of ENaC function in intact monolayer cells. Dysfunctional fluid resolution was seen in uPA-disrupted cells. Administration of uPA and plasmin partially restores ENaC activity and fluid reabsorption by MTEs. ERK1/2, but not Akt, phosphorylation was observed in the cells and lungs of uPA-deficient mice. On the other hand, cleavage of ? ENaC is significantly depressed in the lungs of uPA knockout mice vs. those of wild-type controls. Expression of caspase 8, however, did not differ between wild-type and uPA(-/-) mice. In addition, uPA deficiency did not alter transepithelial resistance. Taken together, the mechanisms for the regulation of ENaC by uPA in MTEs include augmentation of Na(+)-K(+)-ATPase, proteolysis, and restriction of ERK1/2 phosphorylation. We demonstrate for the first time that ENaC may serve as a downstream signaling target by which uPA controls the biophysical profiles of airway fluid and epithelial function. PMID:25172911

Chen, Zaixing; Zhao, Runzhen; Zhao, Meimi; Liang, Xinrong; Bhattarai, Deepa; Dhiman, Rohan; Shetty, Sreerama; Idell, Steven; Ji, Hong-Long

2014-10-15

44

Soluble urokinase plasminogen activator receptor during allogeneic stem cell transplantation  

DEFF Research Database (Denmark)

Previous studies have found that soluble urokinase plasminogen activation receptor (suPAR) increases during inflammatory and malignant illness and elevated suPAR levels may be associated with poor clinical outcome. The purpose of this study was to investigate plasma levels of suPAR during the course of allogeneic stem cell transplantation (SCT). Twenty SCT patients were included in the study. suPAR was measured by ELISA in daily taken plasma samples during the pretransplant conditioning with chemotherapy and weekly for 1 month after infusion of the graft. suPAR levels before the start of the conditioning were significantly elevated when compared to those of healthy controls. During the conditioning in particular treatment with antithymocyte globulin was associated with significantly increased suPAR levels (P = 0.012). At day +7 after infusion of the graft, suPAR levels had decreased to pretreatment levels. High suPAR levels at day 0 were associated with increased mortality (P = 0.011). The present study foundincreased suPAR levels during the conditioning in SCT patients. Further, the data indicated that increased suPAR levels may be associated with increased mortality, suggesting suPAR as a candidate for further studies as an outcome predictor in SCT.

Haastrup, E; Andersen, J

2011-01-01

45

Soluble urokinase plasminogen activator receptor during allogeneic stem cell transplantation.  

Science.gov (United States)

Previous studies have found that soluble urokinase plasminogen activation receptor (suPAR) increases during inflammatory and malignant illness and elevated suPAR levels may be associated with poor clinical outcome. The purpose of this study was to investigate plasma levels of suPAR during the course of allogeneic stem cell transplantation (SCT). Twenty SCT patients were included in the study. suPAR was measured by ELISA in daily taken plasma samples during the pretransplant conditioning with chemotherapy and weekly for 1 month after infusion of the graft. suPAR levels before the start of the conditioning were significantly elevated when compared to those of healthy controls. During the conditioning in particular treatment with antithymocyte globulin was associated with significantly increased suPAR levels (P = 0.012). At day +7 after infusion of the graft, suPAR levels had decreased to pretreatment levels. High suPAR levels at day 0 were associated with increased mortality (P = 0.011). The present study found increased suPAR levels during the conditioning in SCT patients. Further, the data indicated that increased suPAR levels may be associated with increased mortality, suggesting suPAR as a candidate for further studies as an outcome predictor in SCT. PMID:21223347

Haastrup, E; Andersen, J; Ostrowski, S R; Høyer-Hansen, G; Jacobsen, N; Heilmann, C; Ullum, H; Müller, K

2011-04-01

46

Thrombolysis with recombinant tissue plasminogen activator in 7 children.  

Science.gov (United States)

The information about the thromboembolic events, the optimal treatment choice, the dose, and duration of antithrombotic therapy in children are limited. More clinical data are required. Recombinant tissue plasminogen activator (r-tPA) is increasingly used in pediatric thrombosis. We retrospectively analyzed the clinical course of 7 children (9.3 ± 2.1 years; 34 days to 16 years) with arterial thrombosis (n = 1) and intracardiac thrombosis (n = 6). The children were treated with r-tPA. The dose ranged between 0.2 and 0.4 mg/kg per h infused for 3 to 4 hours. This dose was repeated between 2 to 7 times till the thrombolysis was achieved. Treatment side effects were closely monitored. Complete clot lysis was achieved in all cases. None of them had severe bleeding except mild recurrent epistaxis occurring in 2 cases. In conclusion, r-tPA is an effective and safe therapy under close hemostatic control in children. PMID:22496087

Evim, Melike Sezgin; Bostan, Özlem; Baytan, Birol; Semizel, Evren; Günes, Adalet Meral

2013-09-01

47

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

48

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

2007-05-01

49

An Active Site Water Network in the Plasminogen Activator Pla from Yersinia pestis  

Energy Technology Data Exchange (ETDEWEB)

The plasminogen activator Pla from Yersinia pestis is an outer membrane protease (omptin) that is important for the virulence of plague. Here, we present the high-resolution crystal structure of wild-type, enzymatically active Pla at 1.9 {angstrom}. The structure shows a water molecule located between active site residues D84 and H208, which likely corresponds to the nucleophilic water. A number of other water molecules are present in the active site, linking residues important for enzymatic activity. The R211 sidechain in loop L4 is close to the nucleophilic water and possibly involved in the stabilization of the oxyanion intermediate. Subtle conformational changes of H208 result from the binding of lipopolysaccharide to the outside of the barrel, explaining the unusual dependence of omptins on lipopolysaccharide for activity. The Pla structure suggests a model for the interaction with plasminogen substrate and provides a more detailed understanding of the catalytic mechanism of omptin proteases.

Eren, Elif; Murphy, Megan; Goguen, Jon; van den Berg, Bert (UMASS, MED)

2010-08-13

50

Expression of Active Human Tissue-Type Plasminogen Activator in Escherichia coli  

OpenAIRE

The formation of native disulfide bonds in complex eukaryotic proteins expressed in Escherichia coli is extremely inefficient. Tissue plasminogen activator (tPA) is a very important thrombolytic agent with 17 disulfides, and despite numerous attempts, its expression in an active form in bacteria has not been reported. To achieve the production of active tPA in E. coli, we have investigated the effect of cooverexpressing native (DsbA and DsbC) or heterologous (rat and yeast protein disulfide i...

Qiu, Ji; Swartz, James R.; Georgiou, George

1998-01-01

51

Tissue plasminogen activator modulates emotion in a social context.  

Science.gov (United States)

Tissue plasminogen activator (tPA) is known to play physiologically and pathologically crucial roles in the central nervous system. However, it is still obscure whether it affects social behavior. We investigated social behavior and neuronal activation after social stimulation in tPA knockout (KO) mice. In a resident-intruder test, the latency to the first contact was significantly delayed in tPA KO mice compared with that in tPA heterogenic (Het) mice. However, tPA KO mice spent significantly more time undertaking active behavior than tPA Het mice did. In a sociability test, tPA KO mice significantly spent more time and walked a greater distance in the chamber containing an empty cage than tPA Het mice. Furthermore, tPA KO mice approached an empty cage more frequently than tPA Het mice did. In a social novelty test, there was no difference in the duration spent sniffing a stimulator mouse between genotypes. However, tPA KO mice approached even a familiar mouse more frequently than tPA Het mice did. tPA KO mice spent similar durations in a chamber containing a familiar mouse and that containing an unfamiliar mouse, and tPA KO mice walked a significantly greater distance in the former chamber than tPA Het mice did. Furthermore, at the cingulate and prelimbic cortices, the number of cFos-positive cells was significantly increased in tPA KO mice compared with that in tPA Het mice after social stimulation. Our results suggest that tPA modulates emotion in a social context through the function of the prefrontal cortex. PMID:25499620

Nakamura, Kazuki; Takabe, Ayumi; Shimizu, Fuki; Takahashi, Maiko; Matsuo, Osamu; Mitsui, Shinichi

2015-03-15

52

Prognostic significance of urokinase-type plasminogen activator and plasminogen activator inhibitor-1 in primary breast cancer  

DEFF Research Database (Denmark)

The uPA-mediated pathway of plasminogen activation is central to cancer metastasis. Whether uPA and PAI-1 are related to local recurrence, metastatic spread or both is not clear. We present a retrospective study of 429 primary breast cancer patients with a median follow-up of 5.1 years, in which the levels of uPA and PAI-1 in tumour extracts were analysed by means of an enzyme-linked immunosorbent assay. The median values of uPA and PAI-1, which were used as cut-off points, were 4.5 and 11.1 ng mg(-1) protein respectively. The levels of uPA and PAI-1 were correlated with tumour size, degree of anaplasia, steroid receptor status and number of positive nodes. Patients with high content of either uPA or PAI-1 had increased risk of relapse and death. We demonstrated an independent ability of PAI-1 to predict distant metastasis (relative risk 1.7, confidence limits 1.22 and 2.46) and that neither uPA nor PAI-1 provided any information regarding local recurrence.

Knoop, Ann; Andreasen, Peter A

1998-01-01

53

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 es 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

54

Soluble urokinase plasminogen activator receptor in blood transfusion components.  

Science.gov (United States)

Post-transfusion infectious complications associated with allogeneic blood components may depend on storage time and may be related to extracellular accumulation of bioactive substances during storage. The glycoprotein, soluble urokinase plasminogen activator receptor (suPAR), which is located in specific granules of neutrophils, plays a role in inflammation and remodelling of the extracellular matrix. Using enzyme-linked immunosorbent assay, suPAR was determined in serum, plasma and blood cell lysates. In addition, suPAR was measured in whole blood (WB), buffy-coat-depleted saline-adenine-glucose-mannitol (SAGM) blood, platelet-rich plasma (PRP) and buffy-coat-derived platelet (BCP) pools with and without pre-storage leucofiltration, and in non-filtered WB, SAGM blood and platelet concentrates prepared using apheresis (APC) at different time points during storage. Mean suPAR concentration was significantly higher in cell lysates, compared to that in a corresponding serum (P = 0.007) and in plasma samples (P = 0.004). Mean suPAR levels in WB, BCP and SAGM were significantly reduced using leucofiltration (WB: 3.4 versus 2.0 ng mL(-1); BCP: 1.6 versus 1.1 ng mL(-1); SAGM: 2.8 versus 0.19 ng mL(-1)), whereas no difference was observed in PRP. In non-filtered WB, SAGM and APC, extracellular suPAR accumulated significantly in a storage-time-dependent manner (WB: P suPAR, compared to both serum and plasma. This can be explained by the release of suPAR from intracellular granules during cell lysis. The amount of suPAR is significantly increased during storage of blood transfusion components, an accumulation that is reduced using pre-storage leucofiltration. PMID:15285727

Riisbro, R; Brünner, N; Christensen, I J; Nielsen, H J

2004-08-01

55

NMR studies of urokinase-type plasminogen activator  

International Nuclear Information System (INIS)

U-PA (urokinase-type plasminogen activator or urokinase) has been studied under a variety of solution conditions by 1-D and 2-D NMR spectroscopy. Very high quality spectra could be obtained despite the high molecular weight (46 kDa) by appropriate choice of solution conditions. Comparison of spectra of u-PA with spectra of the isolated kringle and protease domains, of the EGF-kringle pair, and of a 23 residue peptide, corresponding to part of the linker between the kringle and the protease domain, enabled sequential assignments in the u-PA spectrum to be made for resonances of the EGF-like and the kringle domain, and domain specific assignments for many others. Simulations of lineshapes in 2-D spectra enabled effective correlation times for the different domains, both isolated and in the intact protein, to be determined. These have permitted a model of the u-PA dynamics to be put forward involving extensive, but not unrestricted, motion between all three domains. In detailed studies of thermal and chemical denaturation the stability of the domains has been proved not to be affected by their isolation. Furthermore, each of the domains has been shown to unfold independently in the intact protein. Three unfolding transitions have been found in the non-inactivated protease domain, corresponding to two transitions of the C-terminal subdomain and a third for the N-terminal subdomain. The extremely high stability of the N-terminal subdomain has been used to isolate this regibdomain has been used to isolate this region by limited proteolysis with thermolysin. The isolated fragment is again highly stable. Finally spectra of different forms of u-PA, namely the protein of human urinary origin, the zymogen single chain u-PA and the non-inactivated protein, have been analysed to address questions which arose in the course of these studies. (author)

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Enhanced expression of urokinase plasminogen activator and its receptor in pancreatic carcinoma.  

OpenAIRE

Urokinase plasminogen activator (uPA) is a serine proteinase that has been suggested to play an important role in cancer invasion and metastasis. It binds to a specific membrane receptor denominated uPA receptor (uPAR). uPA activates plasminogen to form plasmin, which participates in tissue degradation and proteolysis. Binding of uPA to its receptor accelerates UPA's own activation from pro-uPA, enhancing the activity of the uPA/uPAR cascade. Using immunohistochemistry and Northern blot analy...

Cantero, D.; Friess, H.; Deflorin, J.; Zimmermann, A.; Bru?ndler, M. A.; Riesle, E.; Korc, M.; Bu?chler, M. W.

1997-01-01

57

Receptor-mediated endocytosis of plasminogen activators and activator/inhibitor complexes  

DEFF Research Database (Denmark)

Recent findings have elucidated the mechanism for clearance from the extracellular space of the two types of plasminogen activators, urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA), and their type-1 inhibitor (PAI-1). Activator/PAI-1 complexes and uncomplexed t-PA bind to the multi-ligand receptors alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein (alpha 2MR) and epithelial glycoprotein 330 (gp330). These receptors mediate endocytosis and degradation of u-PA/PAI-1 complex bound to the glycosyl phosphatidyl inositol-anchored urokinase receptor (u-PAR) on cell surfaces, and participate, in cooperation with other receptors, in hepatic clearance of activator/PAI-1 complexes and uncomplexed t-PA from blood plasma. The alpha 2MR- and gp330-mediated endocytosis of a ligand (u-PA/PAI-1 complex) initially bound to another receptor (u-PAR) is a novel kind of interaction between membrane receptors. Binding to alpha 2MR and gp330 is a novel kind of molecular recognition of serine proteinases and serpins.

Andreasen, P A; Sottrup-Jensen, L

1994-01-01

58

Plasminogen activator inhibitor-1 (PAI-1 and urokinase plasminogen activator (uPA in sputum of allergic asthma patients.  

Directory of Open Access Journals (Sweden)

Full Text Available Urokinase plasminogen activator (uPA and its inhibitor (PAI-1 have been associated with asthma. The aim of this study was to evaluate concentration of uPA and PAI-1 in induced sputum of house dust mite allergic asthmatics (HDM-AAs. The study was performed on 19 HDM-AAs and 8 healthy nonatopic controls (HCs. Concentration of uPA and PAI-1 was evaluated in induced sputum supernatants using ELISA method. In HDM-AAs the median sputum concentration of uPA (128 pg/ml; 95% CI 99 to 183 pg/ml and PAI-1 (4063 pg/ml; 95%CI 3319 to 4784 pg/ml were significantly greater than in HCs (17 pg/ml; 95%CI 12 to 32 pg/ml; p<0.001 and 626 pg/ml; 95%CI 357 to 961 pg/ml; p<0.001 for uPA and PAI-1 respectively. The sputum concentration of uPA correlated with sputum total cell count (r=0.781; p=0.0001 and with logarithmically transformed exhaled nitric oxide concentration (eNO (r=0.486; p=0.035 but not with FEV1 or bronchial reactivity to histamine. On the contrary, the sputum PAI-1 concentration correlated with FEV1 (r=-0,718; p=0.0005 and bronchial reactivity to histamine expressed as log(PC20 (r=-0.824; p<0.0001 but did not correlate with sputum total cell count or eNO. The results of this study support previous observations linking PAI-1 with airway remodeling and uPA with cellular inflammation. Moreover, the observed effect of uPA seems to be independent of its fibrynolytic activity.

Sebastian Zukowski

2008-06-01

59

Amiloride lowers blood pressure and attenuates urine plasminogen activation in patients with treatment-resistant hypertension.  

Science.gov (United States)

In conditions with albuminuria, plasminogen is aberrantly filtered across the glomerular barrier and activated along the tubular system to plasmin. In the collecting duct, plasmin activates epithelial sodium channels (ENaC) proteolytically. Hyperactivity of ENaC could link microalbuminuria/proteinuria to resistant hypertension. Amiloride, an ENaC inhibitor, inhibits urokinase-type plasminogen activator. We hypothesized that amiloride (1) reduces blood pressure (BP); (2) attenuates plasminogen-to-plasmin activation; and (3) inhibits urine urokinase-type plasminogen activator in patients with resistant hypertension and type 2 diabetes mellitus (T2DM).In an open-label, non-randomized, 8-week intervention study, a cohort (n = 80) of patients with resistant hypertension and T2DM were included. Amiloride (5 mg/d) was added to previous triple antihypertensive treatment (including a diuretic and an inhibitor of the renin-angiotensin-aldosterone system) and increased to 10 mg if BP control was not achieved at 4 weeks. Complete dataset for urine analysis was available in 60 patients. Systolic and diastolic BP measured by ambulatory BP monitoring and office monitoring were significantly reduced. Average daytime BP was reduced by 6.3/3.0 mm Hg. Seven of 80 cases (9%) discontinued amiloride due to hyperkalemia >5.5 mol/L, the most frequent adverse event. Urinary plasmin(ogen) and albumin excretions were significantly reduced after amiloride treatment (P < .0001). Urokinase activity was detectable in macroalbuminuric urine, with a tendency toward reduction in activity after amiloride treatment. Amiloride lowers BP, urine plasminogen excretion and activation, and albumin/creatinine ratio, and is a relevant add-on medication for the treatment of resistant hypertension in patients with T2DM and microalbuminuria. PMID:25492830

Oxlund, Christina S; Buhl, Kristian B; Jacobsen, Ib A; Hansen, Mie R; Gram, Jeppe; Henriksen, Jan Erik; Schousboe, Karoline; Tarnow, Lise; Jensen, Boye L

2014-12-01

60

Amiloride lowers blood pressure and attenuates urine plasminogen activation in patients with treatment-resistant hypertension  

DEFF Research Database (Denmark)

In conditions with albuminuria, plasminogen is aberrantly filtered across the glomerular barrier and activated along the tubular system to plasmin. In the collecting duct, plasmin activates epithelial sodium channels (ENaC) proteolytically. Hyperactivity of ENaC could link microalbuminuria/proteinuria to resistant hypertension. Amiloride, an ENaC inhibitor, inhibits urokinase-type plasminogen activator. We hypothesized that amiloride (1) reduces blood pressure (BP); (2) attenuates plasminogen-to-plasmin activation; and (3) inhibits urine urokinase-type plasminogen activator in patients with resistant hypertension and type 2 diabetes mellitus (T2DM).In an open-label, non-randomized, 8-week intervention study, a cohort (n = 80) of patients with resistant hypertension and T2DM were included. Amiloride (5 mg/d) was added to previous triple antihypertensive treatment (including a diuretic and an inhibitor of the renin-angiotensin-aldosterone system) and increased to 10 mg if BP control was not achieved at 4 weeks.Complete dataset for urine analysis was available in 60 patients. Systolic and diastolic BP measured by ambulatory BP monitoring and office monitoring were significantly reduced. Average daytime BP was reduced by 6.3/3.0 mm Hg. Seven of 80 cases (9%) discontinued amiloride due to hyperkalemia >5.5 mol/L, the most frequent adverse event. Urinary plasmin(ogen) and albumin excretions were significantly reduced after amiloride treatment (P < .0001). Urokinase activity was detectable in macroalbuminuric urine, with a tendency toward reduction in activity after amiloride treatment. Amiloride lowers BP, urine plasminogen excretion and activation, and albumin/creatinine ratio, and is a relevant add-on medication for the treatment of resistant hypertension in patients with T2DM and microalbuminuria.

Oxlund, Christina S; Buhl, Kristian B

2014-01-01

61

Human breast cancer cell-mediated bone collagen degradation requires plasminogen activation and matrix metalloproteinase activity  

OpenAIRE

Abstract Background Breast cancer cells frequently metastasize to the skeleton and induce extensive bone destruction. Cancer cells produce proteinases, including matrix metalloproteinases (MMPs) and the plasminogen activator system (PAS) which promote invasion of extracellular matrices, but whether these proteinases degrade bone matrix is unclear. To characterize the role that breast cancer cell proteinases play in bone degradation we compared the effects of three human breast cancer cell lin...

Hill Peter A; Morgan Hayley

2005-01-01

62

A comparative study of amyloid-beta (1-42) as a cofactor for plasminogen activation by vampire bat plasminogen activator and recombinant human tissue-type plasminogen activator  

OpenAIRE

The activity of both human tissue-type plasminogen activator (t-PA) and the PA from the saliva of the vampire bat, Desmodus rotundus, (DSPA) is critically dependent on the presence of a cofactor. The most efficient cofactor for both PAs is fibrin, but fibrinogen and amyloid beta peptides also have cofactor activities for human t-PA. Compared to t-PA, DSPA has a more stringent requirement for fibrin as a cofactor. The present study was undertaken to compare cofactor activities of amyloid beta ...

Kruithof, Egbert; Schleuning, Wolf-dieter

2004-01-01

63

Rapid and highly sensitive solid-phase radioassay for plasminogen activators  

International Nuclear Information System (INIS)

A rapid and highly sensitive solid-phase radioassay for the measurement of plasminogen activators is presented. The method employs a convenient and stable 125I-fibrinogen-latex bead product and can reproducibly detect 0.25 milli Ploug units/ml of urokinase. This represents a 100-fold increase in sensitivity over previously published radioisotopic solid-phase technique and a 120-fold increase over the sensitivity of the fibrin plate method. Since the assay can readily detect plasminogen activator levels in euglobulin solutions prepared from pre- and post-venous occlusion plasma, it may be useful for rapidly detecting and monitoring the fibrinolytic potential of patients predisposed to thromboembolic disease

64

Endotoxin induction of an inhibitor of plasminogen activator in bovine pulmonary artery endothelial cells  

International Nuclear Information System (INIS)

The effects of bacterial lipopolysaccharide (endotoxin) on the fibrinolytic activity of bovine pulmonary artery endothelial cells were examined. Endotoxin suppressed the net fibrinolytic activity of cell extracts and conditioned media in a dose-dependent manner. The effects of endotoxin required at least 6 h for expression. Cell extracts and conditioned media contained a 44-kDa urokinase-like plasminogen activator. Media also contained multiple plasminogen activators with molecular masses of 65-75 and 80-100 kDa. Plasminogen activators in extracts and media were unchanged by treatment of cells with endotoxin. Diisopropyl fluorophosphate (DFP)-abolished fibrinolytic activity of extracts and conditioned media. DFP-treated samples from endotoxin-treated but not untreated cells inhibited urokinase and tissue plasminogen activator, but not plasmin. Inhibitory activity was lost by incubation at pH 3 or heating to 560C for 10 min. These treatments did not affect inhibitory activity of fetal bovine serum. Incubation of 125I-urokinase with DFP-treated medium from endotoxin-treated cells produced an inactive complex with an apparent molecular mass of 80-85 kDa

65

Relationships between activators and inhibitors of plasminogen, and the progression of small abdominal aortic aneurysms  

DEFF Research Database (Denmark)

plasmin is a common activator of the known proteolytic systems involved in the aneurysmal degradation, and is reported to be associated with the expansion of abdominal aortic aneurysms (AAA). The aim of this study was to study the activating pathways of plasminogen as predictors of the progression of AAA.

Lindholt, Jes Sanddal; JØrgensen, B

2003-01-01

66

Levels of plasminogen activator inhibitor type 1 and urokinase plasminogen activator receptor in non-small cell lung cancer as measured by quantitative ELISA and semiquantitative immunohistochemistry  

DEFF Research Database (Denmark)

The components of the plasminogen activation system have been reported to have prognostic impact in several cancer types, e.g. breast-, colon-, gastric- and lung cancer. Most of these studies have used quantification by enzyme-linked immunosorbent assay (ELISA) on tumour tissue extracts. However, results in non-small cell lung cancer (NSCLC) studies obtained by quantitative ELISA and semiquantitative immunohistochemistry differ. If the prognostic value of the components of the plasminogen activation system is to be exploited clinically in the future, it is important to choose an easy and valid methodology. In the present study we investigated levels of plasminogen activator inhibitor type 1 (PAI-I) and urokinase plasminogen activator receptor (uPAR), as quantitated by ELISA in tumour extracts from 64 NSCLC patients (38 squamous cell carcinomas, 26 adenocarcinomas), and compared them to staining intensity as semiquantitated by immunohistochemistry for PAI-1 and uPAR on corresponding cryostat sections. A significant association (r = 0.49), P <0.0001) was found between the PAI-1 levels measured by ELISA and semiquantitated by immunohistochemistry. No association was found for uPAR. When correlating levels of PAI-1 and uPAR determined by ELISA and immunohistochemistry, respectively, to survival status, no significant correlation was found for any of the subgroups. At present neither of the methods examined in the present study can be recommended as superior for quantitating PAI-1 and uPAR with the aim of predicting prognosis. In conclusion, a larger comparative study is needed to clarify the relationship between ELISA and immunohistochemical results, before a methodology for clinical use can be chosen in non-small cell lung cancer.

Pappot, Helle; Skov, Birgit Guldhammer

1997-01-01

67

Seahorse-derived peptide suppresses invasive migration of HT1080 fibrosarcoma cells by competing with intracellular ?-enolase for plasminogen binding and inhibiting uPA-mediated activation of plasminogen.  

Science.gov (United States)

?-Enolase is a glycolytic enzyme and a surface receptor for plasminogen. ?-Enolase-bound plasminogen promotes tumor cell invasion and cancer metastasis by activating plasmin and consequently degrading the extracellular matrix degradation. Therefore, ?-enolase and plasminogen are novel targets for cancer therapy. We found that the amino acid sequence of a peptide purified from enzymatic hydrolysates of seahorse has striking similarities to that of ?-enolase. In this study, we report that this peptide competes with cellular ?-enolase for plasminogen binding and suppresses urokinase plasminogen activator (uPA)-mediated activation of plasminogen, which results in decreased invasive migration of HT1080 fibrosarcoma cells. In addition, the peptide treatment decreased the expression levels of uPA compared to that of untreated controls. These results provide new insight into the mechanism by which the seahorse-derived peptide suppresses invasive properties of human cancer cells. Our findings suggest that this peptide could emerge as a potential therapeutic agent for cancer. PMID:24602611

Kim, Yong-Tae; Kim, Se-kwon; Jeon, You-Jin; Park, Sun Joo

2014-12-01

68

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

69

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

70

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

Directory of Open Access Journals (Sweden)

Full Text Available 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. This 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

2012-11-01

71

Paradoxical embolus illustrating speed of action of recombinant tissue plasminogen activator in massive pulmonary embolism  

OpenAIRE

A case of a patient who presented with massive pulmonary embolism (PE) requiring thrombolysis with alteplase is reported. The subsequent presence of a patent foramen ovale and paradoxical embolism clinically demonstrated the speed of action of the recombinant tissue plasminogen activator. The advantage of this class of medication when considering the treatment options for a PE in an acute setting is highlighted.

Madani, H.; Ransom, P. A.

2007-01-01

72

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.

1299-13-01

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Production of Plasminogen Activator in Cultures of Superior Cervical Ganglia and Isolated Schwann Cells  

Science.gov (United States)

Plasminogen activator has been implicated in tissue remodeling and cell migration during embryogenesis. In the developing nervous system, these processes are evident in the migration of neurons, axonal extension, Schwann cell migration, and the ensheathment and myelination of nerves. We have studied the production of plasminogen activator in cultures of superior cervical ganglia under conditions in which both neurons and glia are present. We have found that a principal source of the enzyme in these cultures is the glial cells and that the enzyme could not be detected at the growing tips of neurites. Plasminogen activator is also produced by Schwann cells isolated from neonatal rat sciatic nerve. The production of the enzyme by these cells is stimulated 6- to 10-fold by cholera toxin. Isolated Schwann cells and glial cells in the ganglion explant cultures produce the tissue form of plasminogen activator, a form of the enzyme not often found in nonmalignant cells. Preliminary experiments suggest that neuronal-glial interactions may regulate enzyme production by Schwann cells.

Alvarez-Buylla, Arturo; Valinsky, Jay E.

1985-05-01

74

Identification of a pituitary factor responsible for enhancement of plasminogen activator activity in breast tumor cells.  

OpenAIRE

Sheep pituitary glands contain a protein that stimulates plasminogen activator (PA) activity 3- to 20-fold in serum-free cultures of T47D, MTW9/PL, and SC115 breast tumor cells. This protein was found to be similar to basic fibroblast growth factor (bFGF) in size, cationic nature, and affinity for heparin. Purified human placental bFGF, a homologue of human and bovine pituitary bFGF, was effective in stimulating mammary tumor cell PA at a concentration of 1 ng/ml. Antibodies to placental bFGF...

Mira-y-lopez, R.; Joseph-silverstein, J.; Rifkin, D. B.; Ossowski, L.

1986-01-01

75

In vitro degradation of radiolabelled, intact basement membrane mediated by cellular plasminogen activator  

International Nuclear Information System (INIS)

A simple and unique procedure for the isolation of intact basement membrane from Syrian hamster lung was developed. The abilities of malignant fibrosarcoma cell lines to degrade the [3H]basement membrane was examined based on the solubilization of the insoluble material after degradation. When added to growing tumor cells in the presence of growth medium and serum, the [3H]basement membrane was solubilized extensively. The reaction was linear for 24 h at which time up to 90% of the labelled material had been released. A preneoplastic cell line was also capable of degrading the [3H]basement membrane. The solubilization of the [3H]basement membrane was primarily due to degradation of the glycoproteins of the basement membrane. The abilities of the tumor cells to degrade the [3H]basement membrane correlated with their fibrinolytic activity and inhibitors of plasmin inhibited the reaction. Furthermore, the activity of the cells in this assay was dependent of [3H]basement membrane was observed if plasminogen depleted serum was employed, but complete degradation was accomplished if purified plasminogen was added to the medium with plasminogen-depleted serum. These results indicate a role for plasminogen activator in the pathogenesis of invasive tumor cells. (author)

76

Structure-activity relationships of 11 new congeners of the SMTP plasminogen modulator.  

Science.gov (United States)

The fungal metabolite Stachybotrys microspora triprenyl phenols (SMTPs) are small-molecule plasminogen modulators that enhance plasminogen activation. The SMTP molecule consists of a tricyclic ?-lactam moiety, an isoprene side-chain and an N-linked side-chain. Previous investigations have demonstrated that the N-linked side-chain is crucial for its activity. In this study, we have isolated 11 new SMTP congeners with a variety of N-linked side-chain structures, to investigate structure-activity relationships. Active compounds included congeners with a carboxyl or a sulfonic acid group in the N-linked side-chain, whereas not all the congeners with a carboxyl group were active. Of these congeners, that with methionine or tyrosine as the N-linked side-chain moiety was more active than that with an aliphatic amino acid. Congeners without ionizable group in the N-linked side-chain were essentially inactive. PMID:20842143

Hasegawa, Keiko; Koide, Haruki; Hu, Weimin; Nishimura, Naoko; Narasaki, Ritsuko; Kitano, Yoshikazu; Hasumi, Keiji

2010-10-01

77

Pocket Protein-Independent Repression of Urokinase-Type Plasminogen Activator and Plasminogen Activator Inhibitor 1 Gene Expression by E2F1  

OpenAIRE

Expression of genes of the plasminogen activator (PA) system declines at the G0/G1-S-phase boundary of the cell cycle. We found that overexpression of E2F1-3, which acts mainly in late G1, inhibits promoter activity and endogenous expression of the urokinase-type PA (uPA) and PA inhibitor 1 (PAI-1) genes. This effect is dose dependent and conserved in evolution. Mutation analysis indicated that both the DNA-binding and transactivation domains of E2F1 are necessary for this regulation. Interes...

Koziczak, Magdalena; Krek, Wilhelm; Nagamine, Yoshikuni

2000-01-01

78

In vitro stimulation of plasminogen activator release from vein walls by adrenaline.  

OpenAIRE

The effect of adrenaline on plasminogen activator release was studied in vitro in human vein biopsy specimens, in which the fibrinolytic activity was determined according to the fibrin slide technique. The tissue slides were covered with a thin fibrin film containing 10(-9) and 10(-7) M adrenaline and exposed for 30 to 60 minutes. In both concentrations highly significant (p less than 0.001) enhancement of fibrinolytic activity was shown, and the enhancement of fibrinolysis was most pronounce...

Kjaeldgaard, A.; Kjaeldgaard, M.

1986-01-01

79

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

80

Excitotoxin-induced neuronal degeneration and seizure are mediated by tissue plasminogen activator  

Science.gov (United States)

NEURONAL degeneration in the hippocampus, a region of the brain important for acquisition of memory in humans, occurs in various pathological conditions, including Alzheimer's disease, brain ischaemia and epilepsy. When neuronal activity is stimulated in the adult rat and mouse hippocampus, tissue plasminogen activator (tPA), a serine protease that converts inactive plasminogen to the active protease plasmin, is transcriptionally induced1,2. The activity of tPA in neural tissue is correlated with neurite outgrowth3, regeneration4 and migration5, suggesting that it might be involved in neuronal plasticity. Here we show that tPA is produced primarily by microglia in the hippocampus. Using excitotoxins to induce neuronal cell loss, we demonstrate that tPA-deficient mice are resistant to neuronal degeneration. These mice are also less susceptible to pharmacologically induced seizures than wild-type mice. These findings identify a role for tPA in neuronal degeneration and seizure.

Tsirka, Stella E.; Gualandris, Anna; Amaral, David G.; Strickland, Sidney

1995-09-01

81

Plasminogen N-terminal activation peptide modulates the activity of angiostatin-related peptides on endothelial cell proliferation and migration  

International Nuclear Information System (INIS)

Angiostatin, a potent inhibitor of angiogenesis, is derived from the fibrinolytic proenzyme, plasminogen, by enzymatic processing. Plasminogen N-terminal activation peptide (PAP) is one of the products concomitantly released aside from angiostatin (kringles 1-4) and mini-plasminogen (kringle 5 plus the catalytic domain) when plasminogen is processed. To determine whether PAP alone or together with the angiostatin-related peptides derived from the processing of plasminogen modulate the proliferation and motility of endothelial cells, we have generated a recombinant PAP and used it to study its effects on endothelial cells in the presence and absence of the angiostatin-related peptides. Our results showed that PAP alone slightly increased the migration but not the proliferation of endothelial cells. However, in the presence of the angiostatin-related peptides, PAP attenuated the inhibitory activity of the angiostatin-related peptides on the proliferation and migration of endothelial cells. The inhibitory effect of PAP on the angiostatin-related peptides could be due to its binding to the kringle domains of the latter peptides

82

Simultaneous Combined Intravenous Recombinant Tissue Plasminogen Activator and Endovascular Therapy for Hyperacute Middle Cerebral Artery M1 occlusion  

OpenAIRE

We investigated the efficacy and safety of combined intravenous (IV) recombinant tissue plasminogen activator (rtPA) and simultaneous endovascular therapy (ET) for hyperacute middle cerebral artery (MCA) M1 occlusion.

Toyota, S.; Sugiura, S.; Iwaisako, K.

2011-01-01

83

A kringle-containing protease with plasminogen-like activity in the basal chordate Branchiostoma belcheri.  

Science.gov (United States)

Plg (plasminogen), a member of the serine protease superfamily, is a key component constituting the fibrinolytic system, and its evolutionary origin remains unknown during the course of animal evolution. In the present study, we isolated a cDNA, designated BbPlgl, encoding a kringle-containing protease with plasminogen-like activity from the basal chordate Branchiostoma belcheri. The deduced protein, BbPlgl, consisted of 430 amino acids, which is structurally characterized by the presence of an N-terminal signal peptide of 16 amino acids, 2 kringle domains with a Lys-binding site structure, a serine protease domain with the putative tPA (tissue plasminogen activator)-cleavage site (between Arg297 and Val298), the catalytic triad His237-Asp288-Ser379 expected for protease function, and a potential N-linked glycosylation site, all characteristic of Plgs. Besides, the recombinant refolded BbPlgl was readily activated by human uPA (urokinase plasminogen activator), and exhibited Plg-like activity. BbPlgl was also able to auto-activate at neutral and alkaline pH at 4 degrees C without the addition of uPA, and the activation was accelerated by addition of human uPA. These results demonstrate that BbPlgl is a novel member of the Plg family, with a domain structure of K-K-SP (kringle-kringle-serine protease) lacking the PAN domain, pushing the evolutionary origin of Plg to the protochordate. In addition, BbPlgl displays a tissue-specific expression pattern in B. belcheri, with the most abundant expression in the hepatic caecum and hind-gut, agreeing with the notion that the hepatic caecum of amphioxus is the precursor of the vertebrate liver. PMID:19193194

Liu, Mingying; Zhang, Shicui

2009-12-01

84

Prostate Cancer Cell-Derived Urokinase-Type Plasminogen Activator Contributes to Intraosseous Tumor Growth and Bone Turnover1  

OpenAIRE

A variety of proteases have been implicated in prostate cancer (PC) bone metastasis, but the individual contributions of these enzymes remain unclear. Urokinase-type plasminogen activator (uPA), a serine protease, can activate plasminogen and stimulate signaling events on binding its receptor uPAR. In the present study, we investigated the functional role of PC cell-associated uPA in intraosseous tumor growth and bone matrix degradation. Using a severe combined immunodeficient-human mouse mod...

Dong, Zhong; Saliganan, Allen D.; Meng, Hong; Nabha, Sanaa M.; Sabbota, Aaron L.; Sheng, Shijie; Bonfil, R. Daniel; Cher, Michael L.

2008-01-01

85

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

86

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

87

C-reactive protein increases plasminogen activator inhibitor–1 expression in human endothelial cells  

OpenAIRE

C-reactive protein (CRP) is an inflammatory marker which predicts cardiovascular disease. However, it is not fully understood whether CRP has direct effects on endothelial functions and gene expression. The purpose of current study was to determine the effects and molecular mechanisms of CRP on the expression of plasminogen activator inhibitor-1 (PAI-1) in human endothelial cells. Human coronary artery endothelial cells (HCAEC) were treated with CRP at clinically relevant concentrations for d...

Chen, Changyi; Nan, Bicheng; Lin, Peter; Yao, Qizhi

2007-01-01

88

Urokinase Plasminogen Activator Receptor-Deficient Mice Demonstrate Reduced Hyperoxia-Induced Lung Injury  

OpenAIRE

Patients with respiratory failure often require supplemental oxygen therapy and mechanical ventilation. Although both supportive measures are necessary to guarantee adequate oxygen uptake, they can also cause or worsen lung inflammation and injury. Hyperoxia-induced lung injury is characterized by neutrophil infiltration into the lungs. The urokinase plasminogen activator receptor (uPAR) has been deemed important for leukocyte trafficking. To determine the expression and function of neutrophi...

Zoelen, Marieke A. D.; Florquin, Sandrine; Beer, Regina; Pater, Jennie M.; Verstege, Marleen I.; Meijers, Joost C. M.; Poll, Tom

2009-01-01

89

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

OpenAIRE

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...

Colaneri, Massimo; Quarti, Andrea; Pozzi, Marco; Gasparini, Stefano; Carloni, Ines; Benedictis, Fernando Maria

2014-01-01

90

Increased tissue plasminogen activator levels in patients with nonvalvular atrial fibrillation  

OpenAIRE

OBJECTIVE: To determine whether plasma tissue plasminogen activator (tPA) levels (a) are higher in patients with novalvular atrial fibrillation (NVAF) than in control subjects in sinus rhythm; (b) differ between NVAF patients with and without a history of an embolic event (transient ischemic attack or embolic stroke); and (c) differ in control subjects with and without a history of thrombotic stroke. DESIGN: Cross-sectional study. SETTING: Internal medicine outpatient group practice and antic...

Kahn, S. R.; Solymoss, S.; Flegel, K. M.

1997-01-01

91

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

OpenAIRE

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

92

Prognostic value of urokinase plasminogen activator in primary breast carcinoma: comparison of two immunoassay methods.  

OpenAIRE

Urokinase-type plasminogen activator (uPA) is a potentially important prognostic factor in breast cancer for identifying patients at high risk of recurrence. This retrospective study assessed two enzyme-linked immunosorbent assay (ELISA) methods measuring uPA antigen levels in 499 primary breast cancer cytosols. Both uPA methods were applied to cytosols used routinely for oestrogen (ER) and progesterone (PgR) receptor assays. uPA was determined using a classical ELISA method (Imubind; America...

Bouchet, C.; Spyratos, F.; Haca?¨ne, K.; Durcos, L.; Ba?©cette, V.; Oglobine, J.

1998-01-01

93

Prognostic role of urokinase-type plasminogen activator in human gliomas.  

OpenAIRE

Urokinase-type plasminogen activator (u-PA) is a 54-kd enzyme shown to participate in tissue degradation under certain normal and pathological conditions, including cancer invasion and metastasis. Increased u-PA expression has been found in cancers of the breast, lung, colon, and prostate, and correlated with worse outcome in patients with lung and breast cancer. We examined the correlation between u-PA expression in gliomas and patient survival. Seventy-seven gliomas from 41 men and 36 women...

Hsu, D. W.; Efird, J. T.; Hedley-whyte, E. T.

1995-01-01

94

Tissue plasminogen activator via cross-collateralization for tandem internal carotid and middle cerebral artery occlusion  

OpenAIRE

Tandem internal carotid and middle cerebral artery occlusion after carotid dissection predicts poor outcome after systemic thrombolysis. Current treatments include the use of endovascular carotid stenting, which carries with it a high risk of propagating further embolic events and worsening the dissection. New strategies for avoiding the aforementioned side-effects include recanalization using cross-collaterals for delivery of intra-lesional tissue plasminogen activator (tPA). We present two ...

Bulsara, Ketan R.; Ediriwickrema, Asiri; Pepper, Joshua; Robertson, Fergus; Aruny, John; Schindler, Joseph

2013-01-01

95

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

OpenAIRE

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...

Smirnov Vladimir N; Trushkina Polina M; Romanova Nina V; Agaphonov Michael O; Ter-Avanesyan Michael D

2002-01-01

96

Inhibition of plasminogen activation by apo(a): role of carboxyl-terminal lysines and identification of inhibitory domains in apo(a)[S  

Science.gov (United States)

Apo(a), the distinguishing protein component of lipoprotein(a) [Lp(a)], exhibits sequence similarity to plasminogen and can inhibit binding of plasminogen to cell surfaces. Plasmin generated on the surface of vascular cells plays a role in cell migration and proliferation, two of the fibroproliferative inflammatory events that underlie atherosclerosis. The ability of apo(a) to inhibit pericellular plasminogen activation on vascular cells was therefore evaluated. Two isoforms of apo(a), 12K and 17K, were found to significantly decrease tissue-type plasminogen activator-mediated plasminogen activation on human umbilical vein endothelial cells (HUVECs) and THP-1 monocytes and macrophages. Lp(a) purified from human plasma decreased plasminogen activation on THP-1 monocytes and HUVECs but not on THP-1 macrophages. Removal of kringle V or the strong lysine binding site in kringle IV10 completely abolished the inhibitory effect of apo(a). Treatment with carboxypeptidase B to assess the roles of carboxyl-terminal lysines in cellular receptors leads in most cases to decreases in plasminogen activation as well as plasminogen and apo(a) binding; however, inhibition of plasminogen activation by apo(a) was unaffected. Our findings directly demonstrate that apo(a) inhibits pericellular plasminogen activation in all three cell types, although binding of apo(a) to cell-surface receptors containing carboxyl-terminal lysines does not appear to play a major role in the inhibition mechanism. PMID:24478033

Romagnuolo, Rocco; Marcovina, Santica M.; Boffa, Michael B.; Koschinsky, Marlys L.

2014-01-01

97

Characterization and purification of tissue plasminogen activator and its binding to the surface of cerebellar neutrons  

International Nuclear Information System (INIS)

Early studies directed at understanding the possible role of the fibrinolytic system in the brain had demonstrated the production of plasminogen activator (PA) by developing mouse cerebellum. This thesis describes neuronal PA as being primarily tissue plasminogen activator (tPA). This tPA was isolated from a mouse neuronal cell line, and a highly specific antibody was developed in rabbits. The antibody isolated from rabbit antiserum effectively and specifically inhibits murine tPA and blocks the catalytic activity of both the intact molecule and its B-chain, while exhibiting little or nor reactivity with mouse or human urokinase (uPA), human plasmin or plasminogen. This antibody was used as an immunoaffinity ligand to obtain highly purified murine tPA to investigate further the interactions of the protein with cerebellar neurons. Using 125I-tPA, the high-affinity binding to cerebellar granule neurons is rapid, time-dependent, saturable, reversible and specific for tPA but not for other related serine proteases. Neither the catalytic site nor the carbohydrate moiety of tPA appear to be involved in the binding. Autoradiography shows the specific tPA binding is to granule neurons in these cultures

98

Proteolytic Regulation of Epithelial Sodium Channels by Urokinase Plasminogen Activator: CUTTING EDGE AND CLEAVAGE SITES.  

Science.gov (United States)

Plasminogen activator inhibitor 1 (PAI-1) level is extremely elevated in the edematous fluid of acutely injured lungs and pleurae. Elevated PAI-1 specifically inactivates pulmonary urokinase-type (uPA) and tissue-type plasminogen activators (tPA). We hypothesized that plasminogen activation and fibrinolysis may alter epithelial sodium channel (ENaC) activity, a key player in clearing edematous fluid. Two-chain urokinase (tcuPA) has been found to strongly stimulate heterologous human ??? ENaC activity in a dose- and time-dependent manner. This activity of tcuPA was completely ablated by PAI-1. Furthermore, a mutation (S195A) of the active site of the enzyme also prevented ENaC activation. By comparison, three truncation mutants of the amino-terminal fragment of tcuPA still activated ENaC. uPA enzymatic activity was positively correlated with ENaC current amplitude prior to reaching the maximal level. In sharp contrast to uPA, neither single-chain tPA nor derivatives, including two-chain tPA and tenecteplase, affected ENaC activity. Furthermore, ? but not ? subunit of ENaC was proteolytically cleaved at ((177)GR?KR(180)) by tcuPA. In summary, the underlying mechanisms of urokinase-mediated activation of ENaC include release of self-inhibition, proteolysis of ? ENaC, incremental increase in opening rate, and activation of closed (electrically "silent") channels. This study for the first time demonstrates multifaceted mechanisms for uPA-mediated up-regulation of ENaC, which form the cellular and molecular rationale for the beneficial effects of urokinase in mitigating mortal pulmonary edema and pleural effusions. PMID:25555911

Ji, Hong-Long; Zhao, Runzhen; Komissarov, Andrey A; Chang, Yongchang; Liu, Yongfeng; Matthay, Michael A

2015-02-27

99

Host Plasminogen Activator Inhibitor-1 Promotes Human Skin Carcinoma Progression in a Stage-Dependent Manner  

Directory of Open Access Journals (Sweden)

Full Text Available Angiogenesis and tumor expansion are associated with extracellular matrix remodeling and involve various proteases such as the plasminogen (Pig/plasminogen activator (PA system. Recently, several experimental data have implicated the plasminogen activator inhibitor-1 (PAI-1 in tumor angiogenesis in murine systems. However, little is known about PAI-1 functions in human skin carcinoma progression. By generating immunodeficient mice (in Rag-1-/- or nude background deleted for PAI-1 gene (PAI-1-/- , we have evaluated the impact of host PAI-1 deficiency on the tumorigenicity of two malignant human skin keratinocyte cell lines HaCaT II-4 and HaCaT A5-RT3 forming low-grade and high-grade carcinomas, respectively. When using the surface transplantation model, angiogenesis and tumor invasion of these two cell lines are strongly reduced in PAI-1-deficient mice as compared to the wild-type control animals. After subcutaneous injection in PAI-1-/- mice, the tumor incidence is reduced for HaCaT II-4 cells, but not for those formed by HaCaT A5-RT3 cells. These data indicate that PAI-1 produced by host cells is an important contributor to earlier stages of human skin carcinoma progression. It exerts its tumor-promoting effect in a tumor stage-dependent manner, but PAI-1 deficiency is not sufficient to prevent neoplastic growth of aggressive tumors of the human skin.

Catherine Maillard

2005-01-01

100

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

101

Inhalation of urokinase-type plasminogen activator reduces airway remodeling in a murine asthma model.  

Science.gov (United States)

The airway remodeling that occurs in asthma is characterized by an excess of extracellular matrix deposition in the submucosa, hyperplasia/hypertrophy of smooth muscle, goblet cell metaplasia, and accumulation of fibroblasts/myofibroblasts. The urokinase-type plasminogen activator (uPA)/plasmin system participates in pericellular proteolysis and is capable of directly degrading matrix components, activating latent proteinases, and activating growth factors. In a mouse ovalbumin (OVA) asthma model, we increased plasminogen activator activity in the lung by administering exogenous uPA or by using mice genetically deficient in the uPA inhibitor plasminogen activator inhibitor-1 (PAI-1) to assess the role of this system in asthma pathogenesis. After intraperitoneal OVA sensitization, mice inhaled OVA plus uPA (500 IU/mouse) or saline by ultrasonic nebulization for 3 wk. When studied 24 h after the final exposure, the groups with upregulated plasmin activity had significantly reduced subepithelial fibrosis within the airway walls and had decreased airway hyperresponsiveness (AHR) to methacholine. Morphometric analysis showed that subepithelial wall thickening of the bronchi (subepithelial area ratio) was also reduced, as were collagen and alpha-smooth muscle actin. Upregulation of plasmin activity also increased the level of hepatocyte growth factor activity in bronchoalveolar lavage fluid, whereas the release of transforming growth factor-beta was decreased. The administration of uPA 1 wk after the last OVA inhalation also significantly reduced lung hydroxyproline content and AHR. These results show that enhancing uPA/plasmin activity lessens the airway remodeling in a murine asthma model. PMID:19098125

Kuramoto, Emi; Nishiuma, Teruaki; Kobayashi, Kazuyuki; Yamamoto, Masatsugu; Kono, Yuko; Funada, Yasuhiro; Kotani, Yoshikazu; Sisson, Thomas H; Simon, Richard H; Nishimura, Yoshihiro

2009-03-01

102

Construction and expression of a recombinant antibody-targeted plasminogen activator  

International Nuclear Information System (INIS)

Covalent linkage of tissue-type plasminogen activator (t-PA) to a monoclonal antibody specific for the fibrin ? 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 ?2b constant region and the catalytic ? 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

103

Tissue-type plasminogen activator promotes murine myofibroblast activation through LDL receptor–related protein 1–mediated integrin signaling  

OpenAIRE

The activation of interstitial fibroblasts to become ?-SMA–positive myofibroblasts is an essential step in the evolution of chronic kidney fibrosis, as myofibroblasts are responsible for the production and deposition of the ECM components that are a hallmark of the disease. Here we describe a signaling pathway that leads to this activation. Tissue-type plasminogen activator (tPA) promoted TGF-?1–mediated ?-SMA and type I collagen expression in rat kidney interstitial fibroblasts. This ...

Hu, Kebin; Wu, Chuanyue; Mars, Wendy M.; Liu, Youhua

2007-01-01

104

Changes in coagulation and tissue plasminogen activator after the treatment of cerebral infarction with lumbrokinase.  

Science.gov (United States)

This paper aimed to investigate the effect of lumbrokinase on the anticoagulation and fibrinolysis in treating cerebral infarction. Lumbrokinase was used in patients with cerebral infarction. Patients were randomly divided into treatment group (n = 31) and control group (n = 20). Single blind method was used in this investigation. The Chinese stroke score was used to evaluate the results of treatment before and after administration of lumbrokinase. Kaolin partial thromboplastin time (KPTT), prothrombin time (PT), fibrinogen content, vWF content were analyzed, and tissue plasminogen activator (t-PA) activity, plasminogen activator inhibitor (PAI) activity, D-dimer level were assayed. In both groups, the stroke score decreased after administration, but in the treatment group, it was more obvious. In the treatment group, KPTT was prolonged, t-PA activity and D-dimer level increased, while the content of fibrinogen decreased significantly. There were no significant changes of PT and PAI activity in both groups. It is concluded that lumbrokinase is beneficial to the treatment of cerebral infarction. The effect of lumbrokinase is related to the inhibition of intrinsic coagulation pathway and the activation of fibrinolysis via an increase of t-PA activity. PMID:11321442

Jin, L; Jin, H; Zhang, G; Xu, G

2000-01-01

105

Fractionation of heparin by chromatography on a tissue plasminogen activator-Sepharose column.  

OpenAIRE

Heparin stimulates the activity of tissue plasminogen activator (t-PA) and binds to t-PA. To study this interaction, a complex between t-PA and N-acetylated heparin was formed and then linked to Sepharose. This procedure selectively links the t-PA to the column because the acetylated heparin has no free amino groups. The procedure also protects the heparin-binding site(s) on the enzyme during coupling to the matrix. The t-PA column separates heparin into two fractions, one with low affinity f...

Andrade-gordon, P.; Strickland, S.

1990-01-01

106

The Urokinase Receptor Promotes Cancer Metastasis Independently of Urokinase-Type Plasminogen Activator in Mice  

OpenAIRE

The urokinase receptor (uPAR) promotes metastasis of human malignancies; however, its mechanism of action remains incompletely understood. Established models focus on the ability of uPAR to bind urokinase-type plasminogen activator (uPA) and promote protease activation in the tumor cell microenvironment; however, uPAR also regulates cell signaling and migration by both uPA-dependent and -independent mechanisms in vitro. The significance of uPAR as a cell-signaling receptor in vivo remains unc...

Jo, Minji; Takimoto, Shinako; Montel, Valerie; Gonias, Steven L.

2009-01-01

107

Urokinase-type plasminogen activator and its receptor synergize to promote pathogenic proteolysis  

OpenAIRE

Urokinase-type plasminogen activator (uPA) is a potent catalyst of extracellular proteolysis, which also binds to a high-affinity plasma membrane receptor (uPAR). Binding of uPA may influence pericellular proteolysis and/or activate intracellular signal transduction. Transgenic mice overexpressing either uPA or uPAR in basal epidermis and hair follicles had no detectable cutaneous alterations. In contrast, bi-transgenic mice overexpressing both uPA and uPAR, obtained by crossing the two trans...

Zhou, Hong-ming; Nichols, Anthony; Meda, Paolo; Vassalli, Jean-dominique

2000-01-01

108

Seasonal variation of plasminogen activator activity in spermatozoa and seminal plasma of boar, buck, bull and stallion.  

Science.gov (United States)

Plasminogen activators (PA) are proteolytic enzymes present in the spermatozoa and seminal plasma of various species. They play a role in the binding of the spermatozoon and its penetration through the layers surrounding the oocyte. Plasminogen activator activity (PAA) is modulated by hormones that have a seasonal variation, such as testosterone and melatonin. The present study investigates the seasonal variation of PA activity in sperm extracts and seminal plasma of four farm animal species: boar, buck, bull and stallion. Semen samples were collected every second week during a 12-month period and PAA was determined. With respect to sperm enzyme activity, the boar showed a peak from late January until the beginning of April, whereas the activity in the bull was at the highest levels from April until October and gradually declined during autumn and winter period. Plasminogen activator activity of stallion spermatozoa peaked during March and April, and remained low throughout the rest of the year, whereas in the buck sperm, PAA increased from late October until the end of January. No biologically significant variation was detected regarding the seminal PAA activity in any of the species studied. While seasonality of reproduction is typically studied from the female perspective, the present data provide compelling information about a factor that may affect the reproductive ability of the male. PMID:20412514

Zervos, I A; Lavrentiadou, S N; Tsantarliotou, M P; Georgiadis, M P; Kokolis, N A; Taitzoglou, I A

2010-12-01

109

Accessibility of receptor-bound urokinase to type-1 plasminogen activator inhibitor  

International Nuclear Information System (INIS)

Urokinase plasminogen activator (uPA) interacts with a surface receptor and with specific inhibitors, such as plasminogen activator inhibitor type 1 (PAI-1). These interactions are mediated by two functionally independent domains of the molecule: the catalytic domain (at the carboxyl terminus) and the growth factor domain (at the amino terminus). The authors have now investigated whether PAI-1 can bind and inhibit receptor-bound uPA. Binding of 125I-labeled ATF (amino-terminal fragment of uPA) to human U937 monocyte-like cells can be competed for by uPA-PAI-1 complexes, but not by PAI-1 alone. Preformed 125I-labeled uPA-PAI-1 complexes can bind to uPA receptor with the same binding specificity as uPA. PAI-1 also binds to, and inhibits the activity of, receptor-bound uPA in U937 cells, as shown in U937 cells by a caseinolytic plaque assay. Plasminogen activator activity of these cells is dependent on exogenous uPA, is competed for by receptor-binding diisopropyl fluorophosphate-treated uPA, and is inhibited by the addition of PAI-1. In conclusion, in U937 cells the binding to the receptor does not shield uPA from the action of PAI-1. The possibility that in adherent cells a different localization of PAI-1 and uPA leads to protection of uPA from PAI-1 is to be considered

110

Tissue plasminogen activator (tPA) and matrix metalloproteinases in the pathogenesis of stroke: therapeutic strategies.  

Science.gov (United States)

Today there exists only one FDA-approved treatment for ischemic stroke; i.e., the serine protease tissue-type plasminogen activator (tPA). In the aftermath of the failed stroke clinical trials with the nitrone spin trap/radical scavenger, NXY-059, a number of articles raised the question: are we doing the right thing? Is the animal research truly translational in identifying new agents for stroke treatment? This review summarizes the current state of affairs with plasminogen activators in thrombolytic therapy. In addition to therapeutic value, potential side effects of tPA also exist that aggravate stroke injury and offset the benefits provided by reperfusion of the occluded artery. Thus, combinational options (ultrasound alone or with microspheres/nanobubbles, mechanical dissociation of clot, activated protein C (APC), plasminogen activator inhibitor-1 (PAI-1), neuroserpin and CDP-choline) that could offset tPA toxic side effects and improve efficacy are also discussed here. Desmoteplase, a plasminogen activator derived from the saliva of Desmodus rotundus vampire bat, antagonizes vascular tPA-induced neurotoxicity by competitively binding to low-density lipoprotein related-receptors (LPR) at the blood-brain barrier (BBB) interface, minimizing the tPA uptake into brain parenchyma. tPA can also activate matrix metalloproteinases (MMPs), a family of endopeptidases comprised of 24 mammalian enzymes that primarily catalyze the turnover and degradation of the extracellular matrix (ECM). MMPs have been implicated in BBB breakdown and neuronal injury in the early times after stroke, but also contribute to vascular remodeling, angiogenesis, neurogenesis and axonal regeneration during the later repair phase after stroke. tPA, directly or by activation of MMP-9, could have beneficial effects on recovery after stroke by promoting neurovascular repair through vascular endothelial growth factor (VEGF). However, any treatment regimen directed at MMPs must consider their pleiotropic nature and the likelihood of either beneficial or detrimental effects that might depend on the timing of the treatment in relation to the stage of brain injury. PMID:18673209

Adibhatla, Rao Muralikrishna; Hatcher, James F

2008-06-01

111

Distal hinge of plasminogen activator inhibitor-1 involves its latency transition and specificities toward serine proteases  

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Full Text Available Abstract Background The plasminogen activator inhibitor-1 (PAI-1 spontaneously converts from an inhibitory into a latent form. Specificity of PAI-1 is mainly determined by its reactive site (Arg346-Met347, which interacts with serine residue of tissue-type plasminogen activator (tPA with concomitant formation of SDS-stable complex. Other sites may also play roles in determining the specificity of PAI-1 toward serine proteases. Results To understand more about the role of distal hinge for PAI-1 specificities towards serine proteases and for its conformational transition, wild type PAI-1 and its mutants were expressed in baculovirus system. WtPAI-1 was found to be about 12 fold more active than the fibrosarcoma PAI-1. Single site mutants within the Asp355-Arg356-Pro357 segment of PAI-1 yield guanidine activatable inhibitors (a that can still form SDS stable complexes with tPA and urokinase plasminogen activator (uPA, and (b that have inhibition rate constants towards plasminogen activators which resemble those of the fibrosarcoma inhibitor. More importantly, latency conversion rate of these mutants was found to be ~3–4 fold faster than that of wtPAI-1. We also tested if Glu351 is important for serine protease specificity. The functional stability of wtPAI-1, Glu351Ala, Glu351Arg was about 18 ± 5, 90 ± 8 and 14 ± 3 minutes, respectively, which correlated well with both their corresponding specific activities (84 ± 15 U/ug, 112 ± 18 U/ug and 68 ± 9 U/ug, respectively and amount of SDS-stable complex formed with tPA after denatured by Guanidine-HCl and dialyzed against 50 mM sodium acetate at 4°C. The second-order rate constants of inhibition for uPA, plasmin and thrombin by Glu351Ala and Glu351Arg were increased about 2–10 folds compared to wtPAI-1, but there was no change for tPA. Conclusion The Asp355-Pro357 segment and Glu351 in distal hinge are involved in maintaining the inhibitory conformation of PAI-1. Glu351 is a specificity determinant of PAI-1 toward uPA, plasmin and thrombin, but not for tPA.

Shaltiel Shmuel

2003-07-01

112

The role of plasminogen activator inhibitor-1 in gastric mucosal protection.  

Science.gov (United States)

Gastric mucosal health is maintained in response to potentially damaging luminal factors. Aspirin and nonsteroidal anti-inflammatory drugs (NSAIDs) disrupt protective mechanisms leading to bleeding and ulceration. The plasminogen activator system has been implicated in fibrinolysis following gastric ulceration, and an inhibitor of this system, plasminogen activator inhibitor (PAI)-1, is expressed in gastric epithelial cells. In Helicobacter pylori-negative patients with normal gastric histology taking aspirin or NSAIDs, we found elevated gastric PAI-1 mRNA abundance compared with controls; the increase in patients on aspirin was independent of whether they were also taking proton pump inhibitors. In the same patients, aspirin tended to lower urokinase plasminogen activator mRNA. Immunohistochemistry indicated PAI-1 localization to epithelial cells. In a model system using MKN45 or AGS-GR cells transfected with a PAI-1 promoter-luciferase reporter construct, we found no evidence for upregulation of PAI-1 expression by indomethacin, and, in fact, cyclooxygenase products such as PGE2 and PGI2 weakly stimulated expression. Increased gastric PAI-1 mRNA was also found in mice following gavage with ethanol or indomethacin, but plasma PAI-1 was unaffected. In PAI-1(-/-) mice, gastric hemorrhagic lesions in response to ethanol or indomethacin were increased compared with C57BL/6 mice. In contrast, in PAI-1-H/K? mice in which PAI-1 is overexpressed in parietal cells, there were decreased lesions in response to ethanol and indomethacin. Thus, PAI-1 expression is increased in gastric epithelial cells in response to mucosal irritants such as aspirin and NSAIDs probably via an indirect mechanism, and PAI-1 acts as a local autoregulator to minimize mucosal damage. PMID:23494120

Kenny, Susan; Steele, Islay; Lyons, Suzanne; Moore, Andrew R; Murugesan, Senthil V; Tiszlavicz, Laszlo; Dimaline, Rod; Pritchard, D Mark; Varro, Andrea; Dockray, Graham J

2013-05-01

113

Hormonal regulation of plasminogen activator production in porcine kidney cells (LLC-PK1)  

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-PK1, 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-PK1 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-PK1 cells has been examined using two-dimensional gel electrophoresis of 35S 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 an 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

114

Cellular localization of type 1 plasminogen activator inhibitor messenger RNA and protein in murine renal tissue.  

OpenAIRE

Type 1 plasminogen activator inhibitor (PAI-1) may be markedly increased in the plasma of patients with endotoxemia and/or renal disease. To investigate renal PAI-1 production during acute endotoxemia, a murine model system was used. Mice were injected with either saline alone or saline containing 50 micrograms endotoxin, and sacrificed 3 hours later and their tissues analyzed for PAI-1 messenger RNA (mRNA) and antigen. Northern blot analysis confirmed that the level of renal PAI-1 mRNA was g...

Keeton, M.; Eguchi, Y.; Sawdey, M.; Ahn, C.; Loskutoff, D. J.

1993-01-01

115

Successful salvage of thrombosed arterio-venous fistula with thrombolytic therapy using tissue plasminogen activator.  

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A functioning vascular access is crucial to the wellbeing of patients on hemodialysis. Thrombosis is the most common complication of arteriovenous fistula (AVF) resulting in late fistula failure; Its treatment is difficult, and results are often suboptimal. Interventional treatment of AVF thrombosis may not be available all the time, and timely application of an available noninterventional treatment may salvage the fistula. We report the successful treatment of AVF thrombosis using local thrombolytic therapy using tissue plasminogen activator in a patient, for the first time in India. PMID:25838651

Parameswaran, S; Satheesh, S; Morkhandikar, S; Shankar, V; Jayasurya, R; Padhi, R K; Priyamvada, P S; Swaminathan, R P

2015-01-01

116

Plasminogen activator inhibitor-1 gene-deficient mice. I. Generation by homologous recombination and characterization.  

OpenAIRE

Homozygous plasminogen activator inhibitor-1 (PAI-1)-deficient (PAI-1-/-) mice were generated by homologous recombination in D3 embryonic stem cells. Deletion of the genomic sequences encompassing the transcription initiation site and the entire coding regions of murine PAI-1 was demonstrated by Southern blot analysis. A 3.0-kb PAI-1-specific mRNA was identified by Northern blot analysis in liver from PAI-1 wild type (PAI-1+/+) but not from PAI-1-/- mice. Plasma PAI-1 levels, measured 2-4 h a...

Carmeliet, P.; Kieckens, L.; Schoonjans, L.; Ream, B.; Nuffelen, A.; Prendergast, G.; Cole, M.; Bronson, R.; Collen, D.; Mulligan, R. C.

1993-01-01

117

Urokinase-type plasminogen activator in endothelial cells during acute inflammation of the appendix.  

OpenAIRE

Urokinase and tissue-type plasminogen activators (u-PA and t-PA) were identified immunohistochemically in normal and inflamed human appendices by means of polyclonal and monoclonal antibodies. In addition, extracts of the tissues were analyzed for u-PA and t-PA by ELISA. Twelve appendices (five normal and seven with acute inflammation) were analyzed. In the normal appendices, there was a strong staining of the endothelial cells for t-PA, whereas there was negative staining for u-PA. In contra...

Grøndahl-hansen, J.; Kirkeby, L. T.; Ralfkiaer, E.; Kristensen, P.; Lund, L. R.; Danø, K.

1989-01-01

118

The nature of interactions between tissue-type plasminogen activator and platelets  

International Nuclear Information System (INIS)

To elucidate interactions responsible for inhibition of aggregation of platelets in platelet-rich plasma (PRP) harvested from whole blood preincubated with t-PA, experiments were performed with PRP and washed platelets under diverse conditions of preincubation. Both ADP and collagen induced aggregation were inhibited in PRP unless aprotinin had been added to the preincubated whole blood concomitantly with t-PA. However, in washed platelets prepared after the same exposure aggregation was intact. When washed platelets were supplemented with fibrinogen degradation products (FDPs) in concentrations simulating those in whole blood preincubated with t-PA, aggregation induced with either ADP or collagen was inhibited. Thus, the inhibition in PRP depended on generation of FDPs by activated plasminogen. The functional integrity of surface glycoprotein (GP) IIb/IIIa receptors in washed platelets was documented by autoradiography after SDS-PAGE of surface labeled GPs and by fibrinogen binding despite preincubation of the whole blood or washed platelets themselves with t-PA and plasminogen as long as exogenous calcium (greater than or equal to 0.1 microM) was present. In contrast, when calcium was absent, the platelet GP IIb/IIIa receptor was rendered susceptible to degradation by plasmin, and aggregation was inhibited by preincubation at 37 degrees C even if aprotinin was present when aggregation was being assayed. These observations reconcile disparate results in the literaturconcile disparate results in the literature from studies in vivo and in vitro by demonstrating that inhibition of aggregation of platelets in PRP and in whole blood reflects indirect effects of plasminogen activation rather than direct effects of t-PA or plasmin on the platelets themselves

119

The effect of recombinant tissue plasminogen activator on MMP-2 and MMP-9 activities in vitro.  

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One of the most significant side effects during recombinant tissue plasminogen activator (rtPA) for acute stroke treatment is intracranial bleeding. Gelatinases [matrix metalloproteinase (MMP)-2 and MMP-9] are one of the agents involved in the blood-brain barrier destruction resulting in secondary bleeding into the ischemic area during stroke. Previous papers revealed that patients with high baseline MMP-9 serum level have higher risk of intracranial bleeding after thrombolytic therapy. Our objective was to evaluate rtPA influence on serum MMP-2 and MMP-9 activities in vitro. Nine sera obtained from healthy donors were applied for experiment. The commercially available rtPA (Actylise) were diluted with included solvent and additionally with phosphate-buffered saline (PBS) to get concentrations: 2, 4, 8, and 16 ?g/ml. Next, 100 ?l of serum was mixed with equal proportion with different concentrations of rtPA to obtain final rtPA concentrations: 1, 2, 4, and 8 ?g/ml. The sera together with rtPA were incubated for 1 or 2 hours at 37 °C. The activity of gelatinases was estimated with zymography. The activities of MMP-9 (92 kDa) and MMP-2 (72 kDa) were increased by incubation with rtPA in a dose-dependent manner. Simultaneously, the activity of band at 200 kDa (MMP-9/MMP-9 homodimer) was decreased. The activity of gelatinases incubated for 2 hours was elevated in comparison with 1-hour incubation; however, the increase was observed even for sample without rtPA. In conclusion, this study showed that rtPA can increase the biological activity of MMP-2 and MMP-9 on posttranslational level. PMID:24963695

Golab, Piotr; Kielbus, Michal; Bielewicz, Joanna; Kurzepa, Jacek

2015-01-01

120

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

121

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

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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

122

Targeting the autolysis loop of urokinase-type plasminogen activator with conformation-specific monoclonal antibodies  

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Tight regulation of serine proteases is essential for their physiological function, and unbalanced states of protease activity have been implicated in a variety of human diseases. One key example is the presence of uPA (urokinase-type plasminogen activator) in different human cancer types, with high levels correlating with a poor prognosis. This observation has stimulated efforts into finding new principles for intervening with uPA's activity. In the present study we characterize the so-called autolysis loop in the catalytic domain of uPA as a potential inhibitory target. This loop was found to harbour the epitopes for three conformation-specific monoclonal antibodies, two with a preference for the zymogen form pro-uPA, and one with a preference for active uPA. All three antibodies were shown to have overlapping epitopes, with three common residues being crucial for all three antibodies, demonstrating a direct link between conformational changes of the autolysis loop and the creation of a catalytically matureactive site. All three antibodies are potent inhibitors of uPA activity, the two pro-uPA-specific ones by inhibiting conversion of pro-uPA to active uPA and the active uPA-specific antibody by shielding the access of plasminogen to the active site. Furthermore, using immunofluorescence, the conformation-specific antibodies mAb-112 and mAb-12E6B10 enabled us to selectively stain pro-uPA or active uPA on the surface of cultured cells. Moreover, in various independent model systems, the antibodies inhibited tumour cell invasion and dissemination, providing evidence for the feasibility of pharmaceutical intervention with serine protease activity by targeting surface loops that undergo conformational changes during zymogen activation.

BØtkjær, Kenneth AlrØ; Fogh, Sarah

2011-01-01

123

Detection of deep venous thrombosis with indium 111-labelled monoclonal antibody against tissue plasminogen activator  

International Nuclear Information System (INIS)

The administration of a radiolabelled monoclonal antibody against tissue plasminogen activator allows detection of areas with increased fibrinolytic activity, i.e. those with an active thrombotic lesion. Eight patients with phlebographically verified deep venous thrombosis were examined. At the time of immunoscintigraphy study they were examined receiving anticoagulant therapy. Some 75-85 MBq 111In-labelled antibody were injected, and scintigrams were obtained after 30 min and after 24 h. The precise site of the thrombus could not be visualized after 30 min due to high background activity, whereas after 24 h it was detectable in all patients. The thrombus/background ratios achieved are twice as high as those observed in a human antifibrin antibody study. These preliminary data suggest a high sensitivity of our PA-specific antibody for the detection of active deep venous thrombosis in man, and our antibody seems to offer theoretical advantages over both platelet and fibrin-specific antibodies. (orig.)

124

Effects of gemfibrozil and ciprofibrate on plasma levels of tissue-type plasminogen activator, plasminogen activator inhibitor-1 and fibrinogen in hyperlipidaemic patients.  

Science.gov (United States)

Evaluation of fibrate treatment in humans has focused primarily on its anti-lipidaemic effects. A potentially favourable haemostasis-modulating activity of fibrates has also been recognized but the data are not consistent. We sought to learn more about this variability by examining the effects of gemfibrozil and ciprofibrate on plasma levels of tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1) and fibrinogen in primary hyperlipidaemic patients after six and twelve weeks of treatment using different assay systems for PAI-1 and fibrinogen. Although both fibrates effectively lowered triglyceride and cholesterol levels, no effect on the elevated baseline antigen levels of t-PA and PAI-1 was observed after fibrate treatment. However, both fibrates influenced plasma fibrinogen levels, albeit in a different way. Fibrinogen antigen levels were elevated by 17.6% (p <0.05) and 24.3% (p <0.001) with gemfibrozil after six and twelve weeks, respectively, whereas with ciprofibrate there was no effect. Using a Clauss functional assay with either a mechanical end point or a turbidity-based end point, no significant change in fibrinogen levels was seen after six weeks of gemfibrozil treatment. However, after twelve weeks, gemfibrozil enhanced functional fibrinogen levels by 7.2% (p <0.05) as assessed by the Clauss mechanical assay, but decreased functional fibrinogen levels by 12.5% (p <0.0001) when a Clauss assay based on turbidity was used. After six or twelve weeks of ciprofibrate treatment, functional fibrinogen levels were decreased by 10.1% (p <0.001) and 10.5% (p <0.0001), respectively on the basis of Clauss mechanical and by 14.2% (p <0.001) and 28.2% (p <0.0001), respectively with the Clauss turbidimetric assay. A remarkable and consistent finding with both fibrates was the decrease in functionality of fibrinogen as assessed by the ratio of functional fibrinogen (determined by either of the two Clauss assays) to fibrinogen antigen. Taken together, our results indicate that at least part of the variability in the effects of fibrates on haemostatic parameters can be explained by intrinsic differences between various fibrates, by differences in treatment period and/or by the different outcomes of various assay systems. Interestingly, the two fibrates tested both reduced the functionality of fibrinogen. PMID:9364979

Kockx, M; de Maat, M P; Knipscheer, H C; Kastelein, J J; Kluft, C; Princen, H M; Kooistra, T

1997-10-01

125

Angiostatic activity of human plasminogen fragments is highly dependent on glycosylation  

International Nuclear Information System (INIS)

To assess the importance of carbohydrate moieties to the anti-angiogenic activity of plasminogen fragments, we cloned the fragment corresponding to amino acids Val79 to Thr346 (Kint3-4) that presents the three glycosylation sites. The activity of glycosylated and unglycosylated Kint3-4 was tested in murine sponge implant model. We observed a significant decrease in the neovascularization on the sponge after treatment with Kint3-4 by histological examination and determination of the hemoglobin levels. The effects were more intense with the glycosylated than the unglycosylated protein. 99mTechnecium-labeled red blood cells confirmed the inhibition of cell infiltration in the implanted sponge. Studies using melanoma B16F1 implanted in a mouse demonstrated that treatment with glycosylated Kint3-4 (0.15 nmol/48 h) during 14 days suppresses tumor growth by 80%. The vascular endothelial growth factor mRNA levels on the tumor were reduced after treatment. Kint3-4 is a potent plasminogen fragment that has been found to inhibit tumor growth. (author)

126

Rapid localization of indium-111-labeled inhibited recombinant tissue plasminogen activator in a rabbit thrombosis model  

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The thrombus localizing properties of indium-111-recombinant tissue plasminogen activator ({sup 111}In-rt-PA) have been investigated in an effort to achieve prompt and accurate detection of thrombi. Unlike previous studies with rt-PA, the active plasminogen catalytic site was permanently inhibited with peptides of chloromethyl ketone so that the radiotracer binds to fibrin without causing fibrinolysis. Thrombi were created in the external jugular vein of 14 male New Zealand white rabbits followed by injection of {sup 111}In-rt-PA. The agent cleared rapidly in vivo with a half-time of 4.6 min. The thrombus: blood ratio in nonheparinized rabbits (n = 7) was 6.39 +/- 0.86. The ratio in heparinized rabbits (n = 4) was 3.11 +/- 0.23. Thrombi were clearly visible in the planar images of both groups 1 hr postinjection. The combination of rapid thrombus localization and positive images, especially in the presence of anticoagulation, suggests that further work is warranted with rt-PA thrombus imaging.

Butler, S.P.; Kader, K.L.; Owen, J.; Wang, T.S.; Fawwaz, R.A.; Alderson, P.O. (Columbia-Presbyterian Medical Center, New York, NY (USA))

1991-03-01

127

Rapid localization of indium-111-labeled inhibited recombinant tissue plasminogen activator in a rabbit thrombosis model  

International Nuclear Information System (INIS)

The thrombus localizing properties of indium-111-recombinant tissue plasminogen activator (111In-rt-PA) have been investigated in an effort to achieve prompt and accurate detection of thrombi. Unlike previous studies with rt-PA, the active plasminogen catalytic site was permanently inhibited with peptides of chloromethyl ketone so that the radiotracer binds to fibrin without causing fibrinolysis. Thrombi were created in the external jugular vein of 14 male New Zealand white rabbits followed by injection of 111In-rt-PA. The agent cleared rapidly in vivo with a half-time of 4.6 min. The thrombus: blood ratio in nonheparinized rabbits (n = 7) was 6.39 +/- 0.86. The ratio in heparinized rabbits (n = 4) was 3.11 +/- 0.23. Thrombi were clearly visible in the planar images of both groups 1 hr postinjection. The combination of rapid thrombus localization and positive images, especially in the presence of anticoagulation, suggests that further work is warranted with rt-PA thrombus imaging

128

Urokinase plasminogen activator receptor on invasive cancer cells: A prognostic factor in distal gastric adenocarcinoma  

DEFF Research Database (Denmark)

Gastric cancer is the second cancer causing death worldwide. The five-year survival for this malignancy is below 25% and few parameters have shown an impact on the prognosis of the disease. The receptor for urokinase plasminogen activator (uPAR) is involved in extracellular matrix degradation by mediating cell surface associated plasminogen activation, and its presence on gastric cancer cells is linked to micrometastasis and poor prognosis. Using immunohistochemistry, the prognostic significance of uPAR was evaluated in tissue samples from a retrospective series of 95 gastric cancer patients. uPAR was expressed by neoplastic cells, macrophages, myofibroblasts and neutrophils in both intestinal and diffuse subtypes. No association was demonstrated between the expression of uPAR on cancer cells and histological subtype (p = 0.64) or TNM stage (p = 0.75). Univariate analysis revealed a significant association between the expression of uPAR on tumor cells in the peripheral invasion zone and overall survival of gastric cancer patients (HR = 2.16; 95% CI: 1.13-4.14; p = 0.02). Multivariate analysis showed that uPAR immunoreactivity in cancer cells at the invasive front is an independent prognostic factor for overall survival in gastric cancer (HR = 2.39; 95% CI: 1.22-4.69; p = 0.011). In consequence, scoring of uPAR-positive cancer cells may be a direct measure for the invasive potential of gastric adenocarcinomas.

Alpizar, Warner Enrique Alpizar; Christensen, Ib Jarle

2012-01-01

129

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

130

The Role of Plasminogen Activator Inhibitor-1 in the Metabolic Syndrome and Its Regulation  

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Full Text Available Obesity is a major risk factor for cardiovascular disease (CVD. Lipid abnormalities, hypertension, impaired glucose tolerance or diabetes, are cardiovascular risk factors that are frequently present in patients with obesity. Haemostatic and fibrinolytic disturbances are also considered to be important risk factors for CVD hence, a potential link between CVD, obesity and the metabolic syndrome arises. Regulation of the fibrinolytic system can occur at the level of plasminogen activators and plasminogen activator inhibitor-1 (PAI-1. PAI-1, a glycoprotein, is one of the most important inhibitors of fibrinolysis. Regulation of this serine protease inhibitor may have a beneficial effect on other conditions associated with the metabolic syndrome. Human adipose tissue is a source of PAI-1. PAI-1 production may in turn be controlled by a number of hormones and cytokines which are secreted by adipose tissue in addition to dietary factors. In this review we summarise the current knowledge regarding the role of altered fibrinolytic function in obesity, CVD and hence the metabolic syndrome. Regulatory factors including different dietary components, weight loss and dietary intervention will also be discussed.

Martha Phelan

2014-07-01

131

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

132

Role of urokinase plasminogen activator and plasminogen activator inhibitor mRNA expression as prognostic factors in molecular subtypes of breast cancer  

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Full Text Available Isabell Witzel,1 Karin Milde-Langosch,1 Marcus Schmidt,2 Thomas Karn,3 Sven Becker,3 Ralph Wirtz,4 Achim Rody,5 Elena Laakmann,1 Dina Schütze,1 Fritz Jänicke,1 Volkmar Müller1 1Department of Gynecology, University Medical Center, Hamburg, 2Department of Obstetrics and Gynecology, University Hospital, Mainz, 3Department of Obstetrics and Gynecology, University Hospital, Frankfurt, 4STRATIFYER Molecular Pathology GmbH, Cologne, 5Department of Obstetrics and Gynecology, University Medical Center Schleswig-Holstein, Luebeck, Germany Background: Protein levels of urokinase plasminogen activator (uPA and its inhibitor (PAI-1 determined by enzyme-linked immunosorbent assay from fresh-frozen tumor tissue have been evaluated as prognostic factors in prospectively randomized trials in breast cancer. However, the role of uPA and PAI-1 in the context of breast cancer subtypes and for mRNA expression of these factors is less clear. Methods: We evaluated uPA and PAI-1 mRNA expression using the Affymetrix HG-U 133A array within molecular subgroups of breast cancer in cohorts of patients with systemic treatment (cohort A, n=362 and without systemic treatment (cohort B, n=200. We validated mRNA expression in a cohort of HER2-positive breast cancer patients (cohort C, n=290. Luminal, triple-negative, and HER2-positive subcohorts were defined by ESR1 and ERBB2 mRNA expression using predefined cutoffs. Results: In the entire cohort A, elevated PAI-1 but not uPA mRNA expression was associated with shorter disease-free survival (P=0.007 for PAI and 0.069 for uPA. Regarding different molecular subgroups, 67% (n=244 of tumors were luminal, 14% (n=49 were HER2-positive, and 19% (n=69 were triple-negative. Elevated PAI-1 mRNA expression was associated with shorter disease-free survival only in the HER2-positive subgroup (P=0.031. The same disease-free survival results were found for uPA in HER2-positive patients (P=0.011. In contrast, no association between either marker and survival was observed in the luminal or triple-negative subgroups. In the HER2-positive validation cohort C, elevated uPA and PAI-1 mRNA expression also showed strong associations with shorter disease-free survival (P=0.014 for PAI-1, P<0.001 for uPA. Conclusion: In this study, the prognostic impact of uPA and PAI-1 expression was mainly observed in patients with HER2-positive tumors. Keywords: urokinase plasminogen activator, urokinase plasminogen activator inhibitor-1, HER2, breast cancer, prognosis

Witzel I

2014-11-01

133

Long-term stability of recombinant tissue plasminogen activator at -80 C  

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Full Text Available Abstract Background Recombinant tissue plasminogen activator (tPA is a thrombolytic widely used clinically in the treatment of acute thrombotic disease such as ischemic stroke, myocardial infarction, and deep venous thrombosis. This has led to much interest in tPA based lytic therapies leading to laboratory based in-vitro and in-vivo investigations using this drug. However, tPA reconstituted in solution exhibits full activity for only 6–8 hours, according to the manufacturer. Therefore, methods to store reconstituted tPA for long durations while maintaining activity would be of assistance to laboratories using this enzyme. Findings In this work, the enzymatic activity of tPA stored at -80 C over time was measured, using an ELISA technique that measured the amount of active tPA bound to plasminogen activator inhibitor 1 (PAI-1 in a given sample. Sample of tPA solution mixed to a concentration of 1 (mg/ml were stored in cryogenic vials at -80 C for up to 7 years. For a given sample, aliquots were assayed for tPA activity, and compared with a tPA standard to determine relative enzymatic activity. Results are reported as means with standard errors, and 12 measurements were performed for each sample age. Conclusion There was no decrease in tPA activity for samples stored up to 7 years. Such cryogenic storage is a viable method for the preservation of tPA solution for laboratory investigations of tPA-based lytic therapies.

Sperling Matthew

2009-06-01

134

Anthelmintic activity of extracts of Artemisia absinthium against ovine nematodes.  

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The increasing prevalence of anthelmintic resistant strains of helminths, drug residues in animal products and high cost of conventional anthelmintics has created an interest in studying medicinal plants as an alternative source of anthelmintics. Artemisia absinthium Linn. (Tethwen) is used traditionally by people as a vermifuge in addition to its other livestock uses. The objective of this study was to evaluate the anthelmintic efficacy of crude aqueous extracts (CAE) and crude ethanolic extracts (CEE) of the aerial parts of A. absinthium in comparison to albendazole against the gastrointestinal (GI) nematodes of sheep. To fulfill the objectives, the worm motility inhibition assay was utilized in order to investigate the direct effects of plant extracts on the survival of the adult nematodes under in vitro conditions and faecal egg count reduction assay to investigate the effects on faecal egg output of GI nematodes under in vivo conditions. Significant anthelmintic effects of CAE and CEE on live adult Haemonchus contortus worms (P absinthium. The better activity of CEE can be attributed to the greater concentration of alcohol soluble active anthelmintic principle/s and a more rapid transcuticular absorption of the CEE into the body of the worms when compared with the CAE. The results of the present study suggest that A. absinthium extracts are a promising alternative to the commercially available anthelmintics for the treatment of GI nematodes of sheep. PMID:19070963

Tariq, K A; Chishti, M Z; Ahmad, F; Shawl, A S

2009-03-01

135

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

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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

136

Probiotic in rennet paste can affect lipase activity of rennet and lipolysis in ovine cheese  

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Full Text Available Lambs were subjected to three different feeding regimes (mother suckling MS, artificial rearing AR, and artificial rearing with 7log10 cfu/ml Lactobacillus acidophilus supplementation to the milk substitute ARLb and slaughtered at 20d and 40d of age for each feeding treatment. Lambs abomasa were processed to rennet paste and lipases activity was evaluated. Rennet paste was used for Pecorino cheese production. Free fatty acids (FFAs and conjugated linoleic acids (CLAs were detected in cheese at 60d of ripening. Lipase activity was found higher in ARLb than in MS and AR rennet from lambs slaughtered at an older age. A reduction of all FFAs was observed in all cheeses when passing from 20 d to 40d of slaughtering. CLAs were more abundant in ARLb cheeses at both 20 and 40d. Milk substitute with Lb. acidophilus improves enzymatic features of rennet, and health and nutritional characteristics of ovine cheese.

Marzia Albenzio

2010-01-01

137

Reduced plasminogen binding and delayed activation render ?'-fibrin more resistant to lysis than ?A-fibrin.  

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Fibrin (Fn) clots formed from ?'-fibrinogen (?'-Fg), a variant with an elongated ?-chain, are resistant to lysis when compared with clots formed from the predominant ?A-Fg, a finding previously attributed to differences in clot structure due to delayed thrombin-mediated fibrinopeptide (FP) B release or impaired cross-linking by factor XIIIa. We investigated whether slower lysis of ?'-Fn reflects delayed plasminogen (Pg) binding and/or activation by tissue plasminogen activator (tPA), reduced plasmin-mediated proteolysis of ?'-Fn, and/or altered cross-linking. Clots formed from ?'-Fg lysed more slowly than those formed from ?A-Fg when lysis was initiated with tPA/Pg when FPA and FPB were both released, but not when lysis was initiated with plasmin, or when only FPA was released. Pg bound to ?'-Fn with an association rate constant 22% lower than that to ?A-Fn, and the lag time for initiation of Pg activation by tPA was longer with ?'-Fn than with ?A-Fn. Once initiated, however, Pg activation kinetics were similar. Factor XIIIa had similar effects on clots formed from both Fg isoforms. Therefore, slower lysis of ?'-Fn clots reflects delayed FPB release, which results in delayed binding and activation of Pg. When clots were formed from Fg mixtures containing more than 20% ?'-Fg, the upper limit of the normal level, the delay in lysis was magnified. These data suggest that circulating levels of ?'-Fg modulate the susceptibility of clots to lysis by slowing Pg activation by tPA and provide another example of the intimate connections between coagulation and fibrinolysis. PMID:25128532

Kim, Paul Y; Vu, Trang T; Leslie, Beverly A; Stafford, Alan R; Fredenburgh, James C; Weitz, Jeffrey I

2014-10-01

138

Crystal structure of plasminogen activator inhibitor-1 in an active conformation with normal thermodynamic stability  

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The serpin plasminogen activator inhibitor-1 (PAI-1) is a crucial regulator in fibrinolysis and tissue remodeling. PAI-1 has been associated with several pathological conditions and is a validated prognostic marker in human cancers. However, structural information about the native inhibitory form of PAI-1 has been elusive because of its inherent conformational instability and rapid conversion to a latent, inactive structure. Here we report the crystal structure of PAI-1 W175F at 2.3 ? resolution as the first model of the metastable native molecule. Structural comparison with a quadruple mutant (14-1B) previously used as representative of the active state uncovered key differences. The most striking differences occur near the region that houses three of the four mutations in the 14-1B PAI-1 structure. Prominent changes are localized within a loop connecting ?-strand 3A with the F helix, in which a previously observed 3(10)-helix is absent in the new structure. Notably these structural changes are found near the binding site for the cofactor vitronectin. Because vitronectin is the only known physiological regulator of PAI-1 that slows down the latency conversion, the structure of this region is important. Furthermore, the previously identified chloride-binding site close to the F-helix is absent from the present structure and likely to be artifactual, because of its dependence on the 14-1B mutations. Instead we found a different chlorine-binding site that is likely to be present in wild type PAI-1 and that more satisfactorily accounts for the chlorine stabilizing effect on PAI-1.

Jensen, Jan K; Thompson, Lawrence C

2011-01-01

139

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

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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.

S?atirog?lu, O?mer; Durakog?lugil, Murtaza Emre; Ug?urlu, Yavuz; S?ahin, I?smail; Dog?an, Sitki; Ergu?l, Elif; Karadag?, Zakir; Bostan, Mehmet

2014-01-01

140

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

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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

141

Effect of retinoic acid on plasminogen activator expression in human melanoma cells.  

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The plasminogen activation system comprises various proteases that contribute to the invasive potential and metastatic spread of the tumour cell. Two such proteases are tissue-type (tPA) and urokinase-type (uPA) plasminogen activators. Both these enzymes convert plasminogen into the active zymogen plasmin, which has a broad substrate specificity and is capable of degrading a wide range of extracellular matrix molecules. In this study, we examined the effect of retinoic acid (RA) on uPA and tPA secretion in the highly metastatic C8161 and the poorly metastatic Hs294T human melanoma cell lines using a specific enzyme-linked immunosorbent assay (ELISA) detection system, and correlated this production with RA receptor (RAR) expression. Over a range of dilutions, we were able to show that the highly metastatic C8161 cells secreted 0.95 ng of uPA/cell compared with 4.41 fg/cell for the Hs294T cells, whereas the Hs294T cells secreted 24.5 fg of tPA/cell compared with 4.35 fg/cell for the C8161 cells. On exposure of the cells to RA (10(-10)-10(-5) M) for 4 days, uPA secretion was increased 3.4-fold in the C8161 cell line and 1.6-fold in the Hs294T cell line using 10(-8) M RA. In addition, tPA expression was increased in both cell lines by 3.7-fold in the C8161 cells and 3.8-fold in the Hs294T cells with 10(-6) M RA treatment. Increases in PA expression by RA have been reported to involve RAR alpha and RAR beta expression. We were able to detect RAR beta and gamma expression in both cell lines, with and without RA treatment, but were unable to detect expression of RAR alpha. This suggests that another mechanism must exist to regulate the RA modulation of tPA and uPA secretion in these cell lines that does not require RAR alpha expression. PMID:10504054

Alexander, C L; Edward, M; MacKie, R M

1999-08-01

142

Hyperoxia enhanced the production of plasminogen activator by cultured pulmonary artery endothelial cells  

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Pulmonary artery endothelial cell (EC) monolayers exposed to hyperoxia show increased permeability to albumin and marked alterations in actin filament distribution. The mechanism for these cytoskeletal changes is unknown. The authors now report that cells exposed to hyperoxia (95% O{sub 2}) produce significant quantities of urokinase-type plasminogen activators (u-PA). Zymographic analysis revealed that plasminogen-dependent caseinolytic activity in conditioned media from O{sub 2}-exposed cells was several fold higher than controls, especially at 48 hr. Cell-associated lytic activity was markedly increased at 48 and 72 hr when oxidative effects on the cytoskeleton were most apparent. Amidolytic assays confirmed these findings. (1) Conditioned media, CTA U/ml (mean {+-} SEM): Control 0.73{+-}0.05 vs 48 hr O{sub 2} 2.09{+-}0.50, (2) Cell-associated activity (preparations enriched for adhesion plaques), CTA mU/10{sup 6} cells: Control 0.17{+-}0.06; O{sub 2}: 24 hr 0.18{+-}0.07; 48 hr 0.46{+-}0.08; 72 hr 0.48{+-}0.08; O{sub 2} 48 hr or 72 hr vs Control. Immunocytochemical analysis demonstrated breakdown and restructuring of fibronectin and collagen components of the extracellular matrix at 48 and 72 hours. They conclude that hyperoxia enhanced both PA production and dismantling of matrix by EC. These changes may in part be responsible for some of the actin filament alterations which occur in conjunction with O{sub 2}-induced permeability increases.

Phillips, P.G.; Ryan, T.J.; Birnby, L.; Tsan, M.F. (Albany Medical College, NY (United States) New York State Health Lab., Albany (United States))

1990-02-26

143

Plasminogen activator inhibitor-1 suppresses profibrotic responses in fibroblasts from fibrotic lungs.  

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Idiopathic pulmonary fibrosis (IPF) is a fatal lung disease characterized by progressive interstitial scarification. A hallmark morphological lesion is the accumulation of myofibroblasts or fibrotic lung fibroblasts (FL-fibroblasts) in areas called fibroblastic foci. We previously demonstrated that the expression of both urokinase-type plasminogen activator (uPA) and the uPA receptor are elevated in FL-fibroblasts from the lungs of patients with IPF. FL-fibroblasts isolated from human IPF lungs and from mice with bleomycin-induced pulmonary fibrosis showed an increased rate of proliferation compared with normal lung fibroblasts (NL-fibroblasts) derived from histologically "normal" lung. Basal expression of plasminogen activator inhibitor-1 (PAI-1) in human and murine FL-fibroblasts was reduced, whereas collagen-I and ?-smooth muscle actin were markedly elevated. Conversely, alveolar type II epithelial cells surrounding the fibrotic foci in situ, as well as those isolated from IPF lungs, showed increased activation of caspase-3 and PAI-1 with a parallel reduction in uPA expression. Transduction of an adenovirus PAI-1 cDNA construct (Ad-PAI-1) suppressed expression of uPA and collagen-I and attenuated proliferation in FL-fibroblasts. On the contrary, inhibition of basal PAI-1 in NL-fibroblasts increased collagen-I and ?-smooth muscle actin. Fibroblasts isolated from PAI-1-deficient mice without lung injury also showed increased collagen-I and uPA. These changes were associated with increased Akt/phosphatase and tensin homolog proliferation/survival signals in FL-fibroblasts, which were reversed by transduction with Ad-PAI-1. This study defines a new role of PAI-1 in the control of fibroblast activation and expansion and its role in the pathogenesis of fibrosing lung disease and, in particular, IPF. PMID:25648892

Marudamuthu, Amarnath S; Shetty, Shwetha K; Bhandary, Yashodhar P; Karandashova, Sophia; Thompson, Michael; Sathish, Venkatachalem; Florova, Galina; Hogan, Taryn B; Pabelick, Christina M; Prakash, Y S; Tsukasaki, Yoshikazu; Fu, Jian; Ikebe, Mitsuo; Idell, Steven; Shetty, Sreerama

2015-04-10

144

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

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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

145

Selection and characterization of camelid nanobodies towards urokinase-type plasminogen activator.  

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Urokinase-type plasminogen activator (uPA) is a trypsin-like serine protease that plays a vital role in extracellular conversion of inactive plasminogen into catalytically active plasmin. Activated plasmin facilitates the release of several proteolytic enzymes, which control processes like pericellular proteolysis and remodeling of ECM. uPA and the receptor uPAR, are overexpressed in a number of malignant tumours and uPA/uPAR play major roles in adhesion, migration, invasion and metastasis of cancer cells. Elevated levels of uPA have been reported as a risk biomarker for disease relapse, increased cancer malignancy and poor survival prognosis. For these reasons uPA is considered an important target for anticancer drug therapy. In this study we isolated two camel single domain antibodies (nanobodies) from a naïve library by phage display. The nanobody sequences were sequence-optimized for Escherichia coli expression, cloned into the pET22-B(+) vector system, expressed in BL-21 cells and purified from the periplasmic fraction by IMAC. ELISA tests demonstrated that the purified nanobodies were specific for uPA when tested towards other trypsin-like serine proteases. The apparent affinities of the nanobodies were determined by competitive ELISA to 80nM and 522nM, respectively. The best binder did not inhibit uPA (nAb-C3), however the lowest affinity binder (nAb-C8) was able to inhibit the uPA-mediated cleavage of the substrate S-2444. The results validate the naïve library as a resource for retrieval of relevant lead molecules and the novel uPA-nanobodies can be useful pharmacological tools to study uPA structure-function relationships. PMID:25749705

Kaczmarek, Jakub Zbigniew; Skottrup, Peter Durand

2015-06-01

146

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

147

Concentrations of plasminogen activator inhibitor-1 (PAI-1 and urokinase plasminogen activator (uPA in induced sputum of asthma patients after allergen challenge  

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Full Text Available Urokinase plasminogen activator (uPA and its inhibitor (PAI-1 are involved in tiisue remodeling and repairprocesses associated with acute and chronic inflammation. The aim of the study was to evaluate the effect of allergen challengeon concentration of uPA and PAI-1 in induced sputum of house dust mite allergic asthmatics (HDM-AAs. ThirtyHDM-AAs and ten healthy persons (HCswere recruited for the study. In 24 HDM-AAs bronchial challenge with Dermatophagoidespteronyssinus (Dp and in 6 HDM-AAs sham challenege with saline were performed. In HDM-AAs sputumwas induced 24 hours before (T0 and 24 hours (T24 after the challenge. Concentration of uPA and PAI-1 in induced sputumwere determined using immunoenzymatic assays. At T0 in HDM-AAs mean sputum uPA (151±96 pg/ml and PAI-1(4341±1262 pg/ml concentrations were higher than in HC (18.8±6.7 pg/ml; p=0.0002 and 596±180 pg/ml; p<0.0001; foruPA and PAI-1 respectively. After allergen challenge further increase in sputum uPA (187±144 pg/ml; p=0.03 and PAI-1(6252±2323 pg/ml; p<0.0001 concentrations were observed. Moreover, in Dp challenged, but not in saline challengedHDM-AAs the mean uPA/PAI-1 ratio decreased significantly at T24. No significant increase in the studied parameters werefound in sham challenged patients. In HDM-AAs allergen exposure leads to activation of the plasmin system in the airways.Greater increase of the PAI-1 concentration than uPA concentration after allergen challenge may promote airway remodelingand play an important role in the development of bronchial hyperreactivity.

Krzysztof Kowal,

2010-04-01

148

Concentrations of plasminogen activator inhibitor-1 (PAI-1 and urokinase plasminogen activator (uPA in induced sputum of asthma patients after allergen challenge.  

Directory of Open Access Journals (Sweden)

Full Text Available Urokinase plasminogen activator (uPA and its inhibitor (PAI-1 are involved in tiisue remodeling and repair processes associated with acute and chronic inflammation. The aim of the study was to evaluate the effect of allergen challenge on concentration of uPA and PAI-1 in induced sputum of house dust mite allergic asthmatics (HDM-AAs. Thirty HDM-AAs and ten healthy persons (HCswere recruited for the study. In 24 HDM-AAs bronchial challenge with Dermatophagoides pteronyssinus (Dp and in 6 HDM-AAs sham challenege with saline were performed. In HDM-AAs sputum was induced 24 hours before (T0 and 24 hours (T24 after the challenge. Concentration of uPA and PAI-1 in induced sputum were determined using immunoenzymatic assays. At T0 in HDM-AAs mean sputum uPA (151 Â? 96 pg/ml and PAI-1 (4341 Â? 1262 pg/ml concentrations were higher than in HC (18.8 Â? 6.7 pg/ml; p=0.0002 and 596 Â? 180 pg/ml; p<0.0001; for uPA and PAI-1 respectively. After allergen challenge further increase in sputum uPA (187 Â? 144 pg/ml; p=0.03 and PAI-1 (6252 Â? 2323 pg/ml; p<0.0001 concentrations were observed. Moreover, in Dp challenged, but not in saline challenged HDM-AAs the mean uPA/PAI-1 ratio decreased significantly at T24. No significant increase in the studied parameters were found in sham challenged patients. In HDM-AAs allergen exposure leads to activation of the plasmin system in the airways. Greater increase of the PAI-1 concentration than uPA concentration after allergen challenge may promote airway remodeling and play an important role in the development of bronchial hyperreactivity.

Marcin Moniuszko

2011-04-01

149

Elevated soluble urokinase plasminogen activator receptor in plasma from patients with idiopathic myelofibrosis or polycythaemia vera.  

Science.gov (United States)

Extracellular proteolytic enzymes of the urokinase-type plasminogen activator (uPA) system and the family of metalloproteases play a crucial role in the matrix degradation and tissue remodelling processes characteristic of malignant disorders. The receptor for urokinase plasminogen activator (uPAR) serves to localise and intensify the action of uPA and is expressed on the surface of malignant as well as tumour stromal cells including fibroblasts. A soluble form of uPAR (suPAR) cleaved from its glycosyl-phosphatidylinositol anchor is detected in plasma from healthy individuals and increased levels of suPAR have been found in advanced malignancy, suggesting that suPAR may be a marker of extensive tissue remodelling. In an attempt to clarify whether suPAR might be a marker for bone marrow tissue remodelling we measured plasma suPAR levels in a patient cohort comprising 17 with myelofibrosis (MF), 17 with polycythaemia vera (PV), 15 with essential thrombocythaemia (ET), one with a transitional myeloproliferative disorder evolving from PV and 30 controls. Compared with controls suPAR levels were significantly higher in the patients (P suPAR levels differed significantly with the highest levels found in patients with MF and PV (MF vs. PV vs. ET; P = 0.0003). When comparing suPAR levels of the individual patient subgroups with controls, only suPAR levels of PV and MF patients were significantly increased (P suPAR plasma values that were above the mean +2 standard deviations (SD) of controls. The concentration of suPAR was significantly correlated to plasma lactate dehydrogenase, thrombomodulin, and complex of tPA:PAI-1 in the patients. There was no difference between patients and controls when comparing plasma uPA levels. Increased plasma suPAR levels in patients with chronic myeloproliferative disorders may reflect increased uPAR production in the bone marrow, leading to enhanced bone marrow remodelling. PMID:12270061

Jensen, Morten Krogh; Riisbro, Rikke; de Nully Brown, Peter; Brünner, Nils; Hasselbalch, Hans Carl

2002-07-01

150

Histone Deacetylase Inhibition Enhances Tissue Plasminogen Activator Release Capacity in Atherosclerotic Man  

Science.gov (United States)

The expression of the tissue plasminogen activator (t-PA) gene appears to be under epigenetic control and can be affected by histone deacetylation inhibition. The study aimed to test if histone deacetalyase inhibitor treatment lead to increased t-PA release or reduced exhaustion in t-PA release in response to stimulation, as well as change in plasminogen activator inhibitor-1 (PAI-1) in subjects with coronary disease. In this clinical study, 16 post-myocardial infarction subjects, the perfused forearm model was used with isoprenaline provocation during 20 minutes, to stimulate local t-PA release. Each subject was measured twice on the same day (repeated stimuli sequences) as well as on two different occasions, without treatment and after four weeks of treatment with valproic acid (500 mg, twice daily). Net forearm release for t-PA in response to isoprenaline at minutes 1.5, 3, 6, 9, 12, 15 and 18 was measured, allowing assessment of cumulative t-PA release. There was a reduction in the exhaustion of cumulative t-PA release during repeated and prolonged stimulation with valproic acid treatment compared to non-treatment. Plasma PAI-1 antigen was decreased following treatment compared to non-treatment (18.4 ± 10.0 vs. 11.0 ± 7.1 nanograms/ml respectively, mean with 95% confidence interval). These findings demonstrate that histone deacetylation inhibition increases the capacity for endogenous t-PA release in subjects with vascular disease. Furthermore, the fibrinolytic balance is favored with suppressed PAI-1 levels. More studies are needed to establish the clinical relevance of these findings. Trial registration EU Clinical Trials Register 2012-004950-27 PMID:25807501

Svennerholm, Kristina; Haney, Michael; Biber, Björn; Ulfhammer, Erik; Saluveer, Ott; Larsson, Pia; Omerovic, Elmir; Jern, Sverker; Bergh, Niklas

2015-01-01

151

Structure of the human plasminogen activator inhibitor 1 gene: nonrandom distribution of introns  

International Nuclear Information System (INIS)

Plasminogen activator inhibitor 1 (PAI-1) is the primary inhibitor of tissue-type plasminogen activator and thus performs an essential role in the regulation of the fibrinolytic process. It is a member of a large family of serine protease inhibitors (serpins). The authors determined the structure of the PAI-1 gene in order to more completely investigate the relationship of PAI-1 to other serpins and, at the same time, to begin to delineate structure-function relations in PAI-1 itself. A human genomic cosmid DNA library was screened and found to contain two independent clones, each harboring the entire PAI-1 gene. Restriction site mapping, electron microscopic inspection of heteroduplexes, and nucleotide sequence analysis all demonstrate that the PAI-1 gene is approximately 12.2 kilobase pairs in length and consists of nine exons and eight introns. All intron-exon boundaries are in accord with the GT-AG rule, including a cryptic acceptor splice site found in intron 7. The intron-exon pattern of the PAI-1 gene is distinct from that of most other serpins except that intron 3 of PAI-1 occupies an identical position as intron E of ovalbumin. Comparison of the authors data with the proposed subdomain structure of serpins suggests that seven of the eight introns may occupy a nonrandom position in the gene. These introns either delineate boundaries of individual structural subdomains or are located in random coil regions of the protein. Transcription of the PAI-1 gene in cultn. Transcription of the PAI-1 gene in cultured vascular endothelial cells results in two distinct mRNA species. The data suggest that these two transcripts arise by alternative polyadenylation

152

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

153

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

OpenAIRE

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

154

Intrapleural Adenoviral Delivery of Human Plasminogen Activator Inhibitor–1 Exacerbates Tetracycline-Induced Pleural Injury in Rabbits  

Science.gov (United States)

Elevated concentrations of plasminogen activator inhibitor–1 (PAI-1) are associated with pleural injury, but its effects on pleural organization remain unclear. A method of adenovirus-mediated delivery of genes of interest (expressed under a cytomegalovirus promoter) to rabbit pleura was developed and used with lacZ and human (h) PAI-1. Histology, ?-galactosidase staining, Western blotting, enzymatic and immunohistochemical analyses of pleural fluids (PFs), lavages, and pleural mesothelial cells were used to evaluate the efficiency and effects of transduction. Transduction was selective and limited to the pleural mesothelial monolayer. The intrapleural expression of both genes was transient, with their peak expression at 4 to 5 days. On Day 5, hPAI-1 (40–80 and 200–400 nM of active and total hPAI-1 in lavages, respectively) caused no overt pleural injury, effusions, or fibrosis. The adenovirus-mediated delivery of hPAI-1 with subsequent tetracycline-induced pleural injury resulted in a significant exacerbation of the pleural fibrosis observed on Day 5 (P = 0.029 and P = 0.021 versus vehicle and adenoviral control samples, respectively). Intrapleural fibrinolytic therapy (IPFT) with plasminogen activators was effective in both animals overexpressing hPAI-1 and control animals with tetracycline injury alone. An increase in intrapleural active PAI-1 (from 10–15 nM in control animals to 20–40 nM in hPAI-1–overexpressing animals) resulted in the increased formation of PAI-1/plasminogen activator complexes in vivo. The decrease in intrapleural plasminogen-activating activity observed at 10 to 40 minutes after IPFT correlates linearly with the initial concentration of active PAI-1. Therefore, active PAI-1 in PFs affects the outcome of IPFT, and may be both a biomarker of pleural injury and a molecular target for its treatment. PMID:23002099

Karandashova, Sophia; Florova, Galina; Azghani, Ali O.; Komissarov, Andrey A.; Koenig, Kathy; Tucker, Torry A.; Allen, Timothy C.; Stewart, Kris; Tvinnereim, Amy

2013-01-01

155

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

OpenAIRE

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

156

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

Directory of Open Access Journals (Sweden)

Full Text Available 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 activator system in the regulation of fibronectin matrix assembly in the osteoblast-like cell line, MG-63. Results Treatment of MG-63 cells with P25, a peptide ligand for uPAR, resulted in an increase in assembly of fibronectin matrix which was associated with an increase in the number of activated ?1 integrins on the cell surface. Overexpression of uPAR in MG-63 cells increased the effect of P25 on fibronectin matrix assembly and ?1 integrin activation. P25 had no effect on uPAR null fibroblasts, confirming a role for uPAR in this process. The addition of plasminogen activator inhibitor Type I (PAI-1 to cells increased the P25-induced fibronectin polymerization, as well as the number of activated integrins. This positive regulation of PAI-1 on fibronectin assembly was independent of PAI-1's anti-proteinase activity, but acted through PAI-1 binding to the somatomedin B domain of vitronectin. Conclusion These results indicate that vitronectin modulates fibronectin matrix assembly in osteosarcoma cells through a novel mechanism involving cross-talk through the plasminogen activator system.

McKeown-Longo Paula J

2006-03-01

157

Testicular peritubular cells in culture secrete specific inhibitor(s) of plasminogen activators  

International Nuclear Information System (INIS)

Rat peritubular myoid cells in culture secrete component(s) which inhibit the activity of plasminogen activators (PA) produced by Sextoli cells, but do not inhibit the activity of plasmin. The PA inhibitor(s) (PA-I) block both urokinase-type PA (u-PA) secreted by sertoli cells under basal conditions, and tissue-type PA (t-PA) synthesized by Sertoli cells stimulated by follicle stimulating hormone or by cAMP derivatives. The approximate molecular mass of the PA-I is 55 kDa, as determined by gel exclusion chromatography; by polyacrylamide gel electrophoresis; and by reverse autography. Formation of high molecular mass complexes between PA-I and 125I-uPA is prevented by PMSF, indicating that the inhibitor-protease interaction occurs via the active center of the serine protease. Most of the peritubular cell PA-I activity can be immunoprecipitated by antibodies directed against a vascular endothelial cell PA-I. The authors conclude that peritubular myoid cells secrete one or more specific inhibitors of t-PA and u-PA. Possible functions of the PA-I in the modulation of seminiferous tubule restructuring during spermatogenesis will be discussed in relation to their previous observations that testicular PA production and secretion are greatest at discrete stages of the cycle

158

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 condit functional under physiologic conditions and has a increased fibrin binding compared to the two-chain form

159

Targeting of plasminogen activator inhibitor 1 improves fibrinolytic therapy for tetracycline-induced pleural injury in rabbits.  

Science.gov (United States)

Endogenous active plasminogen activator inhibitor 1 (PAI-1) was targeted in vivo with monoclonal antibodies (mAbs) that redirect its reaction with proteinases to the substrate branch. mAbs were used as an adjunct to prourokinase (single-chain [sc] urokinase [uPA]) intrapleural fibrinolytic therapy (IPFT) of tetracycline-induced pleural injury in rabbits. Outcomes of scuPA IPFT (0.25 or 0.0625 mg/kg) with 0.5 mg/kg of mouse IgG or mAbs (MA-33H1F7 and MA-8H9D4) were assessed at 24 hours. Pleural fluid (PF) was collected at 0, 10, 20, and 40 minutes and 24 hours after IPFT and analyzed for plasminogen activating (PA), uPA, fibrinolytic activities, levels of total plasmin/plasminogen, ?-macroglobulin (?M), mAbs/IgG antigens, free active uPA, and ?M/uPA complexes. Anti-PAI-1 mAbs, but not mouse IgG, delivered with an eightfold reduction in the minimal effective dose of scuPA (from 0.5 to 0.0625 mg/kg), improved the outcome of IPFT (P tetracycline-induced pleural injury. PAI-1-neutralizing mAbs improved IPFT by increasing the durability of intrapleural PA activity. These results suggest a novel, well-tolerated IPFT strategy that is tractable for clinical development. PMID:25140386

Florova, Galina; Azghani, Ali; Karandashova, Sophia; Schaefer, Chris; Koenig, Kathleen; Stewart-Evans, Kris; Declerck, Paul J; Idell, Steven; Komissarov, Andrey A

2015-04-01

160

Validation of an ELISA for the quantitation of lanoteplase, a novel plasminogen activator.  

Science.gov (United States)

An ELISA was developed and validated for the quantitation of lanoteplase in human citrated plasma. The ELISA employed a monoclonal anti-lanoteplase antibody absorbed onto 96-well microtiter plates to capture lanoteplase in citrated human plasma samples containing PPACK, a protease inhibitor. The captured lanoteplase was detected using a biotinylated rabbit anti-lanoteplase polyclonal antibody. The standard curve range in human plasma for the ELISA was 7-100 ng/ml. Assessment of individual standard curve variability indicated reproducible responses with r2 values of > or = 0.985. The accuracy (% DEV) and precision (%RSD) estimates for the ELISA based on the predicted values from quality control (QC) samples were within 7.3% and 11%, respectively. Cross-reactivity with t-PA was determined to be less than 11% by ELISA. The stability of lanoteplase was established in human citrated PPACK plasma for 24 hours at 4 degrees C, for 2 months at -20 degrees C, for 22 months at -70 degrees C, three weeks at room temperature, and through four freeze/thaw cycles. To quantify lanoteplase plasminogen activator (PA) activity, a commercially available chromogenic activity assay was also validated. This method and its application is described briefly here. The lanoteplase ELISA as well as the commercial activity method were successfully employed to evaluate the pharmacokinetic parameters of lanoteplase in support of clinical Phase II/III studies. PMID:10595857

Stouffer, B; Habte, S; Vachharajani, N; Tay, L

1999-11-01

161

Inhibition of plasminogen activator inhibitor-1 binding to endocytosis receptors of the low-density-lipoprotein receptor family by a peptide isolated from a phage display library  

OpenAIRE

Abstract The functions of the serpin plasminogen activator inhibitor-1 (PAI-1) are based on molecular interactions with its target proteases urokinase-type and tissue-type plasminogen activator (uPA and tPA), with vitronectin, and with endocytosis receptors of the low density lipoprotein family. Understanding the significance of these interactions would be facilitated by the ability to block them individually. Using phage display, we have identified the disulphide constrained pepti...

Jensen, Jan K.; Malmendal, Anders; Schiøtt, Birgit; Skeldal, Sune; Pedersen, Katrine E.; Celik, Leyla; Nielsen, Niels C.; Andreasen, Peter A.; Wind, Troels

2006-01-01

162

Regulation of proteinases during mouse peri-implantation development: urokinase-type plasminogen activator expression and cross talk with matrix metalloproteinase 9  

OpenAIRE

Trophoblast cells express urokinase-type plasminogen activator (PLAU) and may depend on its activity for endometrial invasion and tissue remodeling during peri-implantation development. However, the developmental regulation, tissue distribution, and function of PLAU are not completely understood. In this study, the expression of PLAU and its regulation by extracellular matrix proteins was examined by RT-PCR, immunocytochemistry, and plasminogen–casein zymography in cultured mouse embryos. T...

Marti?nez-herna?ndez, M. G.; Baiza-gutman, L. A.; Castillo-tra?pala, A.; Armant, D. Randall

2010-01-01

163

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 1013 kPa for 21 h. 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 1.0 mm. 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

164

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.

2011-01-01

165

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 propoy adds additional information to the proposed role of plasminogen activators in the pathogenic pathways of radiation damage in the CNS. 38 refs., 6 figs

166

Effects of addition of tissue-type plasminogen activator in in vitro fertilization medium on bovine embryo development and quality.  

Science.gov (United States)

Plasminogen activators/Plasmin system plays pivotal role in regulating reproductive functions of mammals. Here, we examined the effects of modification of in vitro fertilization medium (IVF medium) with the addition of tissue-type plasminogen activator (t-PA), on bovine embryo development and quality, assessed by quantification of expression of various genes related to metabolism, oxidation, implantation and apoptosis. In addition, plasminogen activator activity (PAA) and plasminogen activator inhibition (PAI) were measured in the spent media. After conventional IVM, 2016 cumulus-oocyte complexes (COCs) were divided into four groups with modified composition of the IVF medium containing t-PA and/or its inhibitor epsilon-aminocaproic acid (control, t-PA, t-PA+?-ACA, ?-ACA). Presumptive zygotes were cultured for 8 days in synthetic oviductal fluid (SOF) medium; gene expression studies were carried out on morulae and blastocysts. t-PA alone significantly suppressed cleavage and blastocyst formation rates, but this effect was neutralized by the addition of ?-ACA. PAA in the treated group was significantly reduced by ?-ACA, but without total elimination. Significant differences were detected in the expression of genes related to apoptosis and/or cell cycle arrest (BAX, BCL2L1, KAT2B) between embryos produced in t-PA-modified media and controls, giving an overall notion that the inferior developmental competence of treated embryos may be attributed to apoptotic phenomena induced by t-PA. In conclusion, it appears that excessive t-PA content in the IVF media, suppresses blastocyst formation rate, possibly due to induction of apoptotic phenomena. PMID:25405906

Krania, F; Dovolou, E; Rekkas, C A; Theodosiadou, E K; Pappas, I; Amiridis, G S

2015-02-01

167

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

168

Enhanced levels of urokinase plasminogen activator and its soluble receptor in common variable immunodeficiency.  

Science.gov (United States)

Common variable immunodeficiency (CVID) is a heterogeneous syndrome characterized by defective immunoglobulin production and high frequency of bacterial infections, autoimmunity and manifestations of chronic inflammation. The urokinase plasminogen activator (uPA), its cell bound and soluble receptor (uPAR, suPAR) have complex biological functions involving innate immune defense mechanisms and regulation of inflammation. Based on this dual role, we hypothesized that the uPA system could be affected in CVID, and examined expression of components of the uPA system in subgroups of CVID. All CVID-patients had increased plasma levels of suPAR with particularly high levels in those with splenomegaly and thrombocytopenia. Plasma uPA levels were also raised in these patients, and both suPAR and uPA levels correlated with the monocyte activation marker neopterin. Monocytes from CVID patients had increased expression of uPAR. We show an increased activation of the uPA system possibly contributing to the inflammatory phenotype seen in subgroups of CVID patients. PMID:19232508

Fevang, Børre; Eugen-Olsen, Jesper; Yndestad, Arne; Brosstad, Frank; Beiske, Klaus; Aukrust, Pål; Frøland, Stig S

2009-06-01

169

Enhanced levels of urokinase plasminogen activator and its soluble receptor in common variable immunodeficiency  

DEFF Research Database (Denmark)

Common variable immunodeficiency (CVID) is a heterogeneous syndrome characterized by defective immunoglobulin production and high frequency of bacterial infections, autoimmunity and manifestations of chronic inflammation. The urokinase plasminogen activator (uPA), its cell bound and soluble receptor (uPAR, suPAR) have complex biological functions involving innate immune defense mechanisms and regulation of inflammation. Based on this dual role, we hypothesized that the uPA system could be affected in CVID, and examined expression of components of the uPA system in subgroups of CVID. All CVID-patients had increased plasma levels of suPAR with particularly high levels in those with splenomegaly and thrombocytopenia. Plasma uPA levels were also raised in these patients, and both suPAR and uPA levels correlated with the monocyte activation marker neopterin. Monocytes from CVID patients had increased expression of uPAR. We show an increased activation of the uPA system possibly contributing to the inflammatory phenotype seen in subgroups of CVID patients.

Fevang, BØrre; Eugen-Olsen, Jesper

2009-01-01

170

Tissue Plasminogen Activator Alters Intracellular Sequestration of Zinc through Interaction with the Transporter ZIP4  

Energy Technology Data Exchange (ETDEWEB)

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.

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

2010-09-17

171

PGE2 reduces MMP-14 and increases plasminogen activator inhibitor-1 in cardiac fibroblasts.  

Science.gov (United States)

Prostaglandin E2 (PGE2) is elevated during cardiac injury and we have previously shown that mice lacking the PGE2 EP4 receptor display dilated cardiomyopathy (DCM) with increased expression of the membrane type matrix metalloproteinase, MMP-14. We thus hypothesized that PGE2 regulates expression of MMP-14 and also affects fibroblast migration. Primary cultures of neonatal rat ventricular fibroblasts (NVFs) were used to test the effects of PGE2. Gene and protein expression was assessed by real time RT-PCR and Western blot, MMP activity was determined by zymography and migration of NVF was assessed by motility in a transwell system. PGE2 reduced expression of MMP-14 and these effects were antagonized by an EP4 antagonist. An EP4 agonist mimicked the effect of PGE2. PGE2 also increased mRNA and protein levels of plasminogen activator inhibitor-1 (PAI-1), an inhibitor of MMP activation. However, PGE2-stimulation of PAI-1 was mediated by the EP1/EP3 receptor and not EP4. Migration of NVF was assessed by motility in a transwell system. Treatment of NVFs with PGE2 reduced the number of cells migrating toward 10% FCS. Treatment with the EP2 agonist also reduced migration but did not affect MMP-14 expression or PAI-1. Our results suggest that PGE2 utilizes different receptors and mechanisms to ultimately decrease MMP expression and NVF migration. PMID:25263346

Kassem, Kamal M; Clevenger, Margarette H; Szandzik, David L; Peterson, Edward; Harding, Pamela

2014-10-01

172

Plasminogen activator inhibitor type-1 : structure-function studies and its use as a reference for intramolecular distance measurements  

OpenAIRE

Inhibitors belonging to the serpin (serine protease inhibitor) family control proteases involved in various physiological processes. All serpins have a common tertiary structure based on the dominant b-sheet A, but they have different inhibitory specificity. The specificity of a serpin is determined by the Pl-Pl’ peptide bond acting as a bait for the target protease which is made up of an exposed reactive centre loop (RCL). The serpin plasminogen activator inhibitor type-1 (PAI-1) is the ma...

Ha?gglo?f, Peter

2003-01-01

173

Intrapleural Adenoviral Delivery of Human Plasminogen Activator Inhibitor–1 Exacerbates Tetracycline-Induced Pleural Injury in Rabbits  

OpenAIRE

Elevated concentrations of plasminogen activator inhibitor–1 (PAI-1) are associated with pleural injury, but its effects on pleural organization remain unclear. A method of adenovirus-mediated delivery of genes of interest (expressed under a cytomegalovirus promoter) to rabbit pleura was developed and used with lacZ and human (h) PAI-1. Histology, ?-galactosidase staining, Western blotting, enzymatic and immunohistochemical analyses of pleural fluids (PFs), lavages, and pleural mesothelial...

Karandashova, Sophia; Florova, Galina; Azghani, Ali O.; Komissarov, Andrey A.; Koenig, Kathy; Tucker, Torry A.; Allen, Timothy C.; Stewart, Kris; Tvinnereim, Amy; Idell, Steven

2013-01-01

174

Fibrinolysis, inflammation and cardiovascular disease; Epidemiological studies on Plasminogen activator inhibitor-type 1 and C-reactive protein  

OpenAIRE

Plasminogen activator inhibitor-type 1 (PAI-1) is the main inhibitor of fibrinolysis and a potential risk factor for cardiovascular disease. In addition to its regulating role in fibrinolysis, PAI-1 may have detrimental effects in the cardiovascular system also through other processes, e.g. inflammation. Although PAI-1 is generally elevated in the presence of cardiovascular disease, it is not yet clear whether it is a causal risk factor. A polymorphism in the promoter region of the PAI-1 gene...

Hoekstra, T.

2003-01-01

175

The Role of Proteasome Beta Subunits in Gastrin-Mediated Transcription of Plasminogen Activator Inhibitor-2 and Regenerating Protein1  

OpenAIRE

The hormone gastrin physiologically regulates gastric acid secretion and also contributes to maintaining gastric epithelial architecture by regulating expression of genes such as plasminogen activator inhibitor 2 (PAI-2) and regenerating protein 1(Reg1). Here we examine the role of proteasome subunit PSMB1 in the transcriptional regulation of PAI-2 and Reg1 by gastrin, and its subcellular distribution during gastrin stimulation. We used the gastric cancer cell line AGS, permanently transfecte...

O’hara, Adrian; Howarth, Alice; Varro, Andrea; Dimaline, Rod

2013-01-01

176

Involvement of tissue plasminogen activator “tPA” in ethanol-induced locomotor sensitization and conditioned-place preference  

OpenAIRE

Ethanol is one of the most abused drugs in the western societies. It is well established that mesolimbic dopaminergic neurons mediate the rewarding properties of ethanol. In our previous studies we have shown that the serine protease tissue plasminogen activator (tPA) is involved in the rewarding properties of morphine and amphetamine. In the current study, we investigated the role of tPA in ethanol-induced behavioral sensitization and conditioned-place preference (CPP). Ethanol treatment dos...

Bahi, Amine; Dreyer, Jean-luc

2012-01-01

177

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

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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

178

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

OpenAIRE

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 ...

Serce Nuran Bektas; Boesl Andreas; Klaman Irina; von Serényi Sonja; Noetzel Erik; Press Michael F; Dimmler Arno; Hartmann Arndt; Sehouli Jalid; Knuechel Ruth; Beckmann Matthias W; Fasching Peter A; Dahl Edgar

2012-01-01

179

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

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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

180

Increased expression of recombinant human tissue plasminogen activator in Leishmania tarentolae.  

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Recombinant tissue plasminogen activator (rt-PA) is one of the most important thrombolytic agents for treating cardiovascular obstructions such as stroke. Glycoprotein rt-PA is a serine protease, consisting of 527 amino acids of which 35 are cysteine residues. A variety of recombinant protein expression systems have been developed for heterologous gene expression in prokaryotic and eukaryotic hosts. In recent years, Leishmania tarentolae has been considered because of its safety aspects and special attributes in expression of complex proteins. In this study, two expression cassettes, each one including two copies of t-PA cDNA, were used for integration into the L. tarentolae genome by electroporation. Transformed clones were selected in the presence of appropriate antibiotics. Expression of active rt-PA was confirmed by Western blot and Zymography tests. Real-time PCR analysis was applied to investigate the presence of multiple t-PA gene copies in the parasite genome. Correlation of t-PA gene dosage and production rate was confirmed with real-time PCR. It was shown that the expression level of rt-PA in L. tarentolae is at least 480 IU/mL of culture media. This concentration of rt-PA is seven times higher than what was reported in previous studies in L. tarentolae and some other eukaryotic systems. PMID:21058320

Hemayatkar, Mahdi; Mahboudi, Fereidoun; Majidzadeh-A, Keivan; Davami, Fatemeh; Vaziri, Behrouz; Barkhordari, Farzaneh; Adeli, Ahmad; Mahdian, Reza; Davoudi, Noushin

2010-11-01

181

Down-regulation of survivin suppresses uro-plasminogen activator through transcription factor JunB.  

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Survivin, a member of the inhibitors of apoptosis protein family, is expressed during development and in various human cancers. However, the clinical relevance of survivin in cancer is still a matter of debate. Genes induced by hepatocyte growth factor (HGF) were screened using cDNA microarray technology in the stomach cancer cell lines, NUGC3 and MKN28. The levels of JunB, survivin, and uro-plasminogen activator (uPA) were up-regulated in cells treated with HGF in a dose-dependent manner. HGF-induced up regulation of JunB, survivin, and uPA was inhibited by pre-treatment with a MEK inhibitor (PD 98059). HGF-induced up-regulation of uPA was repressed by survivin knockdown. HGF enhanced the binding activity of JunB to the survivin promoter in control cells, but not in the JunB-shRNA cells. Transfection with survivin- shRNA resulted in a decrement of cell proliferation, as determined with MTT assays. In an in vitro invasion assay, significantly fewer cells transfected with survivin shRNA than control cells were able to invade across a Matrigel membrane barrier. In conclusion, survivin appeared to play an important role in the up-regulation of uPA induced by HGF via JunB and might contribute to HGF-mediated tumor invasion and metastasis, which may serve as a promising target for gastric cancer therapy. PMID:21734448

Lee, Kyung Hee; Choi, Eun Young; Koh, Sung Ae; Kim, Min Kyoung; Kim, Kyeong Ok; Lee, Si Hyung; Jang, Byung Ik; Kim, Se Won; Kim, Sang Woon; Song, Sun Kyo; Choi, Joon Hyuk; Kim, Jae-Ryong

2011-09-30

182

Resistin regulates the expression of plasminogen activator inhibitor-1 in 3T3-L1 adipocytes.  

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Resistin and plasminogen activator inhibitor-1 (PAI-1) are adipokines, which are secreted from adipocytes. Increased plasma resistin and PAI-1 levels aggravate metabolic syndrome through exacerbation of insulin resistance and induction of chronic inflammation. However, the relationship between resistin and PAI-1 gene expression remains unclear. Previously, we found that resistin regulates lipid metabolism via carbohydrate responsive element-binding protein (ChREBP) during adipocyte maturation (Ikeda et al., 2013) [6]. In this study, to clarify the relationship between expression of resistin and PAI-1, PAI-1 expression in differentiated 3T3-L1 adipocytes was measured after transfection with anti-resistin siRNA. We found that PAI-1 gene expression and secreted PAI-1 protein were significantly decreased by resistin knockdown. Furthermore, phosphorylation of Akt, which can inhibit PAI-1 expression, was accelerated and the activity of protein phosphatase 2A (PP2A) was suppressed in resistin knockdown 3T3-L1 adipocytes. In addition, the expression of glucose transporter type 4, a ChREBP target gene, was reduced and was associated with inhibition of PP2A. The addition of culture medium collected from COS7 cells transfected with a resistin expression plasmid rescued the suppression of PAI-1 expression in resistin knockdown 3T3-L1 adipocytes. Our findings suggest that resistin regulates PAI-1 expression in 3T3-L1 adipocytes via Akt phosphorylation. PMID:24667608

Ikeda, Yoshito; Tsuchiya, Hiroyuki; Hama, Susumu; Kajimoto, Kazuaki; Kogure, Kentaro

2014-05-30

183

SIRT1-mediated epigenetic downregulation of plasminogen activator inhibitor-1 prevents vascular endothelial replicative senescence.  

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The inactivation of plasminogen activator inhibitor-1 (PAI-1) has been shown to exert beneficial effects in age-related vascular diseases. Limited information is available on the molecular mechanisms regarding the negatively regulated expression of PAI-1 in the vascular system. In this study, we observed an inverse correlation between SIRT1, a class III histone deacetylase, and PAI-1 expression in human atherosclerotic plaques and the aortas of old mice, suggesting that internal negative regulation exists between SIRT1 and PAI-1. SIRT1 overexpression reversed the increased PAI-1 expression in senescent human umbilical vein endothelial cells (HUVECs) and aortas of old mice, accompanied by decreased SA-?-gal activity in vitro and improved endothelial function and reduced arterial stiffness in vivo. Moreover, the SIRT1-mediated inhibition of PAI-1 expression exerted an antisenescence effect in HUVECs. Furthermore, we demonstrated that SIRT1 is able to bind to the PAI-1 promoter, resulting in a decrease in the acetylation of histone H4 lysine 16 (H4K16) on the PAI-1 promoter region. Thus, our findings suggest that the SIRT1-mediated epigenetic inhibition of PAI-1 expression exerts a protective effect in vascular endothelial senescence. PMID:25040736

Wan, Yan-Zhen; Gao, Peng; Zhou, Shuang; Zhang, Zhu-Qin; Hao, De-Long; Lian, Li-Shan; Li, Yong-Jun; Chen, Hou-Zao; Liu, De-Pei

2014-10-01

184

Genetic association of urokinase-type plasminogen activator gene rs2227564 site polymorphism with sporadic Alzheimer's disease in the Han Chinese population?  

OpenAIRE

A missense C/T polymorphism in exon 6 (the NCBI rsID is rs2227564) of the urokinase-type plasminogen activator gene has been identified as a possible hot spot for Alzheimer's disease risk. The present study analyzed urokinase-type plasminogen gene polymorphisms of rs2227564 with sporadic Alzheimer's disease by PCR-restriction fragment length polymorphism. Results showed that CC, CT and TT genotype distribution frequencies had significant differences between sporadic Alzheimer's disease patien...

Ji, Xuelian; Jia, Longfei; Jia, Jianping; Qi, Li

2012-01-01

185

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 absocture of ABUSV-PA, as proved by UV absorption and CD spectroscopy

186

Soluble urokinase plasminogen activator receptor levels in tuberculosis patients at high risk for multidrug resistance.  

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The soluble urokinase plasminogen activator receptor (suPAR) has been shown to be a strong prognostic biomarker for tuberculosis (TB). In the present study, the profiles of plasma suPAR levels in pulmonary TB patients at high risk for multidrug resistance were analyzed and compared with those in multidrug resistant (MDR)-TB patients. Forty patients were prospectively included, consisting of 10 MDR-TB patients and 30 TB patients at high risk for MDR, underwent clinical assesment. Plasma suPAR levels were measured using ELISA (SUPARnostic, Denmark) and bacterial cultures were performed in addition to drug susceptibility tests. All patients of suspected MDR-TB group demonstrated significantly higher suPAR levels compared with the healthy TB-negative group (1.79?ng/mL). Among the three groups at high risk for MDR-TB, only the relapse group (7.87?ng/mL) demonstrated suPAR levels comparable with those of MDR-TB patients (7.67?ng/mL). suPAR levels in the two-month negative acid-fast bacilli conversion group (9.29 ng/mL) were higher than positive control, whereas levels in the group consisting of therapy failure patients (5.32?ng/mL) were lower. Our results strongly suggest that suPAR levels enable rapid screening of suspected MDR-TB patients, but cannot differentiate between groups. PMID:23304490

Mardining Raras, Tri Yudani; Astuti, Triwahju; Noor Chozin, Iin

2012-01-01

187

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

188

Urokinase-type plasminogen activator: a new target for male contraception?  

Science.gov (United States)

Urokinase-type plasminogen activator (uPA) is closely related to male reproduction. With the aim of investigating the possibility for uPA as a potential contraceptive target, in the present work, Kunming male mice were immunized by human uPA subcutaneous injection at three separate doses for 3 times. Then the potency of the anti-human uPA antibody in serum was analyzed, and mouse fertility was evaluated. Serum antibody titers for human uPA in immunized groups all reached 1:10,240 or higher levels by enzyme linked immunosorbent assay, and mating experiments revealed that pregnancy rates and the mean number of embryos implanted after mating declined obviously (P groups. However, the mating capacity and reproductive organ weights had no obvious change, and histological analysis of the testes and epididymides also showed normal morphology for immunized male mice. Sperm function tests suggested that the sperm concentration, sperm viability, sperm motility, and in vitro fertilization rate for the cauda epididymis sperm in uPA-immunized groups were lower than those in the controls (P < 0.05). Together, these observations indicated that subcutaneous injection human uPA to the male mice could effectively reduce their fertility, and uPA could become a new target for immunocontraception in male contraceptive development. PMID:25578931

Qin, Ying; Han, Yan; Xiong, Cheng-Liang; Li, Hong-Gang; Hu, Lian; Zhang, Ling

2015-01-01

189

Pneumatic Displacement of a Dense Submacular Hemorrhage with or without Tissue Plasminogen Activator  

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Full Text Available Background: To evaluate the efficacy of treating a dense submacular hemorrhage withpneumatic displacement with or without tissue plasminogen activator (tPA.Methods: Twenty-four patients with a dense submacular hemorrhage were treated withintravitreal expansile gas, with or without an intravitreal injection of tPA, inorder to displace the submacular blood. The main outcome measurementsinclude preoperative and postoperative visual acuity, postoperative fluoresceinangiography (FAG results and additional postoperative treatments.Results: Total or subtotal subfoveal blood displacement was achieved in all 24 eyes.After a mean follow-up of 15.5 months (range 6-50 months, final visualacuity had improved two or more lines in 11 (45.8% of the 24 eyes, andmeasured 20/100 or better in 10 (4l.7% of the 11 eyes. Based on the FAGresults for 14 cases, nine eyes (64.3% received additional postoperativelaser treatment. Final visual acuity of 20/100 or better was achieved in four(40% of the 10 eyes, with a choroidal neovascular membrane (CNVMdetected on FAG, and dye leakage not detected in three (75% of the foureyes.Conclusions: Pneumatic displacement, with or without intravitreal injection of tPA, seemsuseful in displacing dense submacular hemorrhage and facilitating visualimprovement, although the visual result is often limited by the progression ofthe underlying macular disease. In patients with age-related macular degeneration,more treatable CNVM may be detected on postoperative FAG.

Po-Min Yang

2005-12-01

190

A human monoclonal antibody scFv to urokinase plasminogen activator.  

Science.gov (United States)

A human monoclonal antibody can be a good method for tumor diagnosis and treatment. This study is aimed at the generation of human antibody fragments against urokinase plasminogen activator (uPA) known to be related to tumor metastasis using the naive human antibody phage display library. Three clones--A2, A8, and E4--were selected from 1 x 10(10) sized human naïve antibody phage library using BIAcore rescue and screen. Clone A8 was finally selected by flow cytometry against Hep3 and HT1080, uPA overexpressing tumor cell lines. A8 clone consisted of 324 bp lambda and 402 bp heavy chains. The affinity (K(D)) of purified A8 antibody fragments was 1.44 x 10(-8) M(-1). The antibody fragment was reacted with HT1080 in a dose-dependent manner but not reacted with LS513 normal fibroblast. In this study, uPA specific human monoclonal antibody fragment A8 was made with BIAcore selection. Selected A8 was bound specifically to uPA expressed on the tumor cell surface. Further study for the application of A8 antibody clones will be needed. PMID:20443707

Bae, Ki-Beom; Jeong, Young-Joo; Won, Hae Jeong; Hong, Kwan-Hee; Choi, Il-Whan; Seo, Su-Kil; Park, Sae-Gwang

2010-04-01

191

Plasma Soluble Urokinase Plasminogen Activator Receptor in Children with Urinary Tract Infection  

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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. PMID:21918598

Wittenhagen, Per; Andersen, Jesper Brandt; Hansen, Anita; Lindholm, Lone; Rønne, Frederik; Theil, Jørn; Tvede, Michael; Eugen-Olsen, Jesper

2011-01-01

192

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)

193

ADAM9 promotes lung cancer metastases to brain by a plasminogen activator-based pathway.  

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The transmembrane cell adhesion protein ADAM9 has been implicated in cancer cell migration and lung cancer metastasis to the brain, but the underpinning mechanisms are unclear and clinical support has been lacking. Here, we demonstrate that ADAM9 enhances the ability of tissue plasminogen activator (tPA) to cleave and stimulate the function of the promigratory protein CDCP1 to promote lung metastasis. Blocking this mechanism of cancer cell migration prolonged survival in tumor-bearing mice and cooperated with dexamethasone and dasatinib (a dual Src/Abl kinase inhibitor) treatment to enhance cytotoxic treatment. In clinical specimens, high levels of ADAM9 and CDCP1 correlated with poor prognosis and high risk of mortality in patients with lung cancer. Moreover, ADAM9 levels in brain metastases derived from lung tumors were relatively higher than the levels observed in primary lung tumors. Our results show how ADAM9 regulates lung cancer metastasis to the brain by facilitating the tPA-mediated cleavage of CDCP1, with potential implications to target this network as a strategy to prevent or treat brain metastatic disease. PMID:25060522

Lin, Chen-Yuan; Chen, Hung-Jen; Huang, Cheng-Chung; Lai, Liang-Chuan; Lu, Tzu-Pin; Tseng, Guan-Chin; Kuo, Ting-Ting; Kuok, Qian-Yu; Hsu, Jennifer L; Sung, Shian-Ying; Hung, Mien-Chie; Sher, Yuh-Pyng

2014-09-15

194

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

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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.

Hansen, Line; Larsen, Esben Kjær Unmack

2013-01-01

195

Extracellular proteolysis of reelin by tissue plasminogen activator following synaptic potentiation.  

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The secreted glycoprotein reelin plays an indispensable role in neuronal migration during development and in regulating adult synaptic functions. The upstream mechanisms responsible for initiating and regulating the duration and magnitude of reelin signaling are largely unknown. Here we report that reelin is cleaved between EGF-like repeats 6-7 (R6-7) by tissue plasminogen activator (tPA) under cell-free conditions. No changes were detected in the level of reelin and its fragments in the brains of tPA knockouts, implying that other unknown proteases are responsible for generating reelin fragments found constitutively in the adult brain. Induction of NMDAR-independent long-term potentiation with the potassium channel blocker tetraethylammonium chloride (TEA-Cl) led to a specific up-regulation of reelin processing at R6-7 in wild-type mice. In contrast, no changes in reelin expression and processing were observed in tPA knockouts following TEA-Cl treatment. These results demonstrate that synaptic potentiation results in tPA-dependent reelin processing and suggest that extracellular proteolysis of reelin may regulate reelin signaling in the adult brain. PMID:24892761

Trotter, J H; Lussier, A L; Psilos, K E; Mahoney, H L; Sponaugle, A E; Hoe, H-S; Rebeck, G W; Weeber, E J

2014-08-22

196

Endogenous tissue type plasminogen activator facilitates NMDA-induced retinal damage  

International Nuclear Information System (INIS)

To investigate the role of tissue plasminogen activator (tPA) in retinal damage, tPA-deficient and wild-type mice were employed. Two different retinal neuron insult models were used in the present study. One is an excitotoxin-treated retinal model, created by direct intravitreal injection of glutamate analogs, NMDA or kainic acid (KA), and the other is an ischemia-reperfusion model induced by transient elevation of intraocular pressure. TdT-dUTP terminal nick-end labeling (TUNEL) method was used to examine the retinal cell nuclear damage. The number of TUNEL-positive cells in ganglion cell layer (GCL) and inner nuclear layer (INL) in tPA-deficient mice after low-, but not high-dose NMDA was significantly less compared to wild type. In contrast, neither intravitreal KA or transient ischemia produced significant difference in retinal damage in tPA vs. wild-type mice. These data show that tPA-deficient mice are resistant to retinal damage by intravitreal injection of NMDA, and indicate that tPA plays a role in the retinal cell damage induced by excitotoxins, especially NMDA

197

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

198

Urokinase-type plasminogen activator in endothelial cells during acute inflammation of the appendix.  

Science.gov (United States)

Urokinase and tissue-type plasminogen activators (u-PA and t-PA) were identified immunohistochemically in normal and inflamed human appendices by means of polyclonal and monoclonal antibodies. In addition, extracts of the tissues were analyzed for u-PA and t-PA by ELISA. Twelve appendices (five normal and seven with acute inflammation) were analyzed. In the normal appendices, there was a strong staining of the endothelial cells for t-PA, whereas there was negative staining for u-PA. In contrast, the endothelial cells in the inflamed appendices showed u-PA immunoreactivity, and negative or very weak reactions for t-PA. In the inflamed appendix, there was also a labeling of u-PA in fibroblast-like cells and in interstitial areas. The specificity of the staining was supported by a variety of staining controls and also by analysis of tissue extracts with ELISA, showing that on the average the inflamed appendices contained more than twice as much mu-PA per mg of protein as the normal appendices and less than one third of the amount of t-PA. PMID:2508479

Grøndahl-Hansen, J; Kirkeby, L T; Ralfkiaer, E; Kristensen, P; Lund, L R; Danø, K

1989-10-01

199

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

200

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)

201

Medicolegal considerations with intravenous tissue plasminogen activator in stroke: a systematic review.  

Science.gov (United States)

Background. Intravenous tPA (tissue plasminogen activator) therapy remains underutilized in patients with Acute Ischemic Stroke (AIS). Anecdotal data indicates that physicians are increasingly liable for administering and for failure to administer tPA. Methods. An extensive search of Medline, Embase, Westlaw, LexisNexis Legal, and Google Scholar databases was performed. Case studies that involved malpractice litigation in ischemic stroke and thrombolytic therapy were analyzed systematically. Results. We identified 789 ischemic stroke litigation cases, of which 46 cases were related to intravenous tPA and stroke litigation. Case descriptions of 40 cases were available. Data for verdicts were available for 38 patients. The most frequent plaintiff claim was related to failure to administer intravenous tPA (38, 95%). Only 2 (5.0%) claim involved complications of treatment with tPA. Hospitals were defendants in majority of the 36 cases. Physicians were involved in 33 cases. While ED physicians were involved in 25 (60.52%) cases, neurologists were involved in 8 (20.0%) cases. There were 26 (65%) defendant-favored and 12 (30%) plaintiff-favored verdicts. Conclusion. Physicians and hospitals are at an increased risk of litigation in patients with AIS when in IV-tPA is being considered for treatment. While majority of the cases litigated were cases where tPA was not administered, only about 1 in 20 cases was litigated when complications occurred. PMID:24083048

Bhatt, Archit; Safdar, Adnan; Chaudhari, Dhara; Clark, Diane; Pollak, Amber; Majid, Arshad; Kassab, Mounzer

2013-01-01

202

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

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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 pHOMA-IR. SFA intake was predictor of PAI-1 independent of body fat, HOMA-IR and total-cholesterol. In addition, PAI-1 was an independent predictor of blood pressure. HOMA-IR and QUICKI presented significantly higher and lower, respectively, in High-SFA compared to Moderate-SFA intake. High-SFA influenced 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

203

Plasminogen activator inhibitor-1 regulates infiltration of macrophages into melanoma via phosphorylation of FAK-Tyr?²?.  

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Tumor-infiltrating macrophages are potential candidates for cancer immunotherapy. However, the detailed molecular mechanism underlying macrophage infiltration into tumors is poorly understood. Based on our previous finding that plasminogen activator inhibitor-1 (PAI-1) enhances vitronectin-dependent migration of macrophages, we investigated the potential role of PAI-1 in macrophage invasion into melanoma. Experimental evidence obtained from spheroid confrontation assay clearly showed that PAI-1 overexpression significantly enhanced the invasion of RAW 264.7 cells into B16F10 melanoma. We further demonstrated that PAI-1 induces phosphorylation of focal adhesion kinase (FAK) at Tyr(925), which, in turn, mediated the invasion of macrophages into the melanoma. This work further illustrates that low-density lipoprotein receptor related-protein 1 (LRP1) is essential for PAI-1-mediated FAK phosphorylation and macrophage invasion into melanoma. In conclusion, our study demonstrates a novel role of PAI-1 in macrophage invasion into melanoma and provides insights into the underlying molecular mechanism. PMID:25063025

Thapa, Bikash; Koo, Bon-Hun; Kim, Yeon Hyang; Kwon, Hyung-Joo; Kim, Doo-Sik

2014-08-01

204

Plasma soluble urokinase plasminogen activator receptor (suPAR) levels in ankylosing spondylitis.  

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Recent studies demonstrated that soluble urokinase plasminogen activator receptor (suPAR) is a valuable marker in the recognition of an inflammatory response. Ongoing inflammation leads to elevated plasma suPAR levels. We aimed to characterize plasma suPAR levels in ankylosing spondylitis (AS) patients compared to healthy individuals in order to reveal if suPAR could be used as a clinical marker of inflammation in AS. We measured plasma suPAR and C-reactive protein (CRP) levels as well as erythrocyte sedimentation rate (ESR) in 33 AS patients at various stages of disease duration and activity and 29 healthy controls. CRP and ESR values were higher in AS patients than in healthy individuals, while suPAR values were comparable (median [interquartile range]: 2.97 [2.57-3.80] ng/mL vs. 2.80 [2.06-3.42] ng/mL, P>0.05). In AS patients, a correlation was detected between BASDAI scores and CRP as well as ESR values but not suPAR levels (P=0.0005, r=0.57 and P=0.01, r=0.43, respectively). Unlike in many other inflammatory conditions, plasma suPAR levels do not reflect inflammation in AS. To assess the inflammatory status in AS, ESR and particularly CRP values are still more appropriate clinical markers. In line with earlier findings, our results indicate that, unlike suPAR, both of these markers are positively correlated with disease activity in AS. PMID:22999906

Toldi, Gergely; Szalay, Balázs; Bek?, Gabriella; Kovács, László; Vásárhelyi, Barna; Balog, Attila

2013-01-01

205

Plasminogen activators from the saliva of Desmodus rotundus (common vampire bat): unique fibrin specificity.  

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The saliva of D. rotundus contains at least four plasminogen activators (PAs) which all require fibrin as a cofactor. D. rotundus salivary PAs (DSPAs) exhibit a sequential array of structural motifs such as "Finger" (F), "EGF" (E), "Kringle" (K) and "Protease" (P) which was elucidated by cDNA cloning and sequencing. The respective domain organizations are: FEKP (DSPA alpha 1 and DSPA alpha 2), EKP (DSPA beta) and KP (DSPA gamma). In all four forms the plasmin-sensitive site of tPA is obliterated, indicating that they function as single-chain enzymes. DSPA alpha 1 differs from alpha 2 by amino acid substitutions found mainly in the F, E and K domain, 11% of the total sequence. DSPA beta and gamma, while being closely related to alpha 2, still exhibit 2 and 13 amino acid exchanges, respectively. These sequence heterogeneities, together with results of Southern blot hybridization experiments, strongly suggest that the four DSPA mRNA species originate from different genes. All four forms of DSPA have been expressed in animal cell culture and DSPA alpha 1 was chosen for a detailed pharmacological characterization. In vitro DSPA alpha 1 activity is enhanced 50,000-fold in the presence of fibrin, whereas the activity of single chain tPA is only enhanced 100-fold. At equally effective thrombolytic doses DSPA causes lower bleeding incidence in a rat mesenteric vein model and exhibits high potency, clot selectivity, and speed in the dissolution of fibrin embolized into the lung of anesthetized rats. In the copper coil-induced dog coronary heart infarction model, at doses that achieve patency at equal rates, reocclusion is significantly less frequent than with tPA. These results indicate that DSPA alpha 1 may be a safer and more efficacious thrombolytic agent than the PAs currently in clinical use. PMID:1309059

Schleuning, W D; Alagon, A; Boidol, W; Bringmann, P; Petri, T; Krätzschmar, J; Haendler, B; Langer, G; Baldus, B; Witt, W

1992-12-01

206

Oviduct-specific expression of tissue plasminogen activator in laying hens  

Scientific Electronic Library Online (English)

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; Liu, Xiang; Jueken, Aniwashi; Shiyong, Xu.

207

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

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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

208

Optimizing production of recombinant tissue plasminogen activator in non-pathogenic Leishmania by two genetic constructs  

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Full Text Available "nBackground: Recombinant tissue plasminogen activator (rt-PA is one of the most important thrombolytic agents used in patients with vascular occlusions such as acute ischemic stroke or myocardial infarction. A variety of recombinant protein expression systems have been developed for heterologous gene expression in prokaryotic and eukaryotic hosts. In recent years, Leishmania tarentolae (L. tarentolae, a non-pathogenic trypanosomatid protozoa, has come under consideration because of its safety and immunogenicity as a vaccine vector and special attributes in the expression of complex proteins. This study was done to improve rt-PA expression in this protozoon and create the opportunity for the replacement of rt-PA gene with other genes for the production of other complex proteins."n "n Methods: Two expression cassettes were used for the integration of two copies of t-PA cDNA, one copy in each cassette, into the parasite genome by electroporation. The transformed clones were selected by antibiotic resistancy. The expression of active secreted rt-PA was confirmed by Western blot analysis and Chromolize assay."n "n Results: Appearance of a 64 kD band in nitrocellulose membrane in the Western blot analysis confirmed the presence of full-length rt-PA in the culture media. Chromolize assay showed the expression levels of active recombinant t-PA in single and double transfected L. tarentolae clones- 375 IU/ml and 480 IU/ml of the culture media, respectively."n "n Conclusion: The produced rt-PA in the culture media containing the transfected cells was at least seven times higher than what has been reported in previous studies on L. tarentolae or on some other eukaryotic systems.

Hemayatkar M

2011-02-01

209

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

210

Antibody-mediated targeting of the urokinase-type plasminogen activator proteolytic function neutralizes fibrinolysis in vivo  

DEFF Research Database (Denmark)

Urokinase-type plasminogen activator (uPA) plays a central role in tissue remodeling processes. Most of our understanding of the role of uPA in vivo is derived from studies using gene-targeted uPA-deficient mice. To enable in vivo studies on the specific interference with uPA functionality in mouse models, we have now developed murine monoclonal antibodies (mAbs) directed against murine uPA by immunization of uPA-deficient mice with the recombinant protein. Guided by enzyme-linked immunosorbent assay, Western blotting, surface plasmon resonance, and enzyme kinetic analyses, we have selected two highly potent and inhibitory anti-uPA mAbs (mU1 and mU3). Both mAbs recognize epitopes located on the B-chain of uPA that encompasses the catalytic site. In enzyme activity assays in vitro, mU1 blocked uPA-catalyzed plasminogen activation as well as plasmin-mediated pro-uPA activation, whereas mU3 only was directed against the first of these reactions. We additionally provide evidence that mU1, but not mU3, successfully targets uPA-dependent processes in vivo. Hence, systemic administration of mU1 (i) rescued mice treated with a uPA-activable anthrax protoxin and (ii) impaired uPA-mediated hepatic fibrinolysis in tissue-type plasminogen activator (tPA)-deficient mice, resulting in a phenotype mimicking that of uPA;tPA double deficient mice. Importantly, this is the first report demonstrating specific antagonist-directed targeting of mouse uPA at the enzyme activity level in a normal physiological process in vivo.

Lund, Ida K; Jögi, Annika

2008-01-01

211

Increased plasminogen activator inhibitor-1 levels in patients with isolated coronary artery ectasia.  

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Isolated coronary artery ectasia (ICAE) is defined as the ectasia of the coronary arteries without concomitant coronary artery stenosis. The etiology and the clinical course of ICAE are still not clear. Increased levels of plasminogen activator inhibitor-1 (PAI-1) inhibit vasa vasorum, leading to diminished vessel wall supply and thus contributes to aortic aneurysm expansion. Whether the same process has role in coronary artery ectasia is not known. The aim of this study is to investigate the association between PAI-1 and coronary artery ectasia in patients without concomitant obstructive coronary artery disease. Among 2830 patients who underwent coronary angiography between March 2010 and 2011, 55 patients (40 male, 15 female, mean age 60 ± 8 years) with ICAE, formed our study group. 27 patients with similar patient characteristics, with angiographically proven normal coronary arteries, were enrolled as the control group. The basal characteristics were similar between two groups. PAI-1 levels were statistically higher in the ICAE group compared to the control group (104.13 ± 56.65 and 63.39 ± 35.01 ng/dl, respectively) (P = 0.008). A significant positive correlation between CAE and PAI-1 (r = 0.358, P = 0.007) was also demonstrated. Serum high sensitive C reactive protein (hsCRP) levels did not differ between two groups (P > 0.05). The plasma PAI-1 levels were significantly higher in ICAE patients compared to normal coronary artery group. Increased PAI-1 levels may diminish vasa vasorum by antiangiogenic activity leading to coronary ectasia. PMID:21850503

Cicek, Yuksel; Durakoglugil, Murtaza Emre; Erdogan, Turan; Yilmaz, Adnan; Uydu, Huseyin Avni; Saglam, Hayrettin; Cetin, Mustafa; Satiroglu, Omer; Bostan, Mehmet; Canga, Aytun; Temiz, Ahmet

2012-01-01

212

Inhibition of urokinase-type plasminogen activator expression by dihydroartemisinin in breast cancer cells.  

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The aim of the present study was to investigate the inhibitory effects of dihydroartemisinin (DHA) on the primary tumor growth and metastasis of the human breast cancer cell line, MDA-MB-231, in vitro. The expression levels of urokinase-type plasminogen activator (uPA) were detected by immunocytochemistry in two cell lines (MCF-7 and MDA-MB-231). The MDA-MB-231 cell activity was inhibited by various concentration gradients of DHA. The inhibitory rate, cell growth curve and apoptotic morphological observations were obtained using the MTT assay at 0, 24, 48 and 72 h. Cell scratch migration was performed at various time-points to test the cell proliferation and migration capacity. Reverse transcription-polymerase chain reaction was used to analyze the effect of DHA on uPA mRNA expression in breast cancer cells. The human breast cancer cell line, MDA-MB-231, possesses higher metastatic potential and relatively higher expression of uPA when compared with the MCF-7 cell line. DHA was found to inhibit the proliferation and migration capacity of the cell line, MDA-MB-231, in vitro. The growth inhibition occurred in a time- and dose-dependent manner, with IC50 values of 117.76±0.04, 60.26±0.12 and 52.96±0.07 ?mol/l following 24, 48 and 72 h, respectively. The inhibition of uPA was observed to decrease breast cancer cell growth and migration. Thus, results of the present study indicate that DHA may be used for further studies with regard to breast cancer therapy. PMID:24765140

Zhang, Shuqun; Ma, Yinan; Jiang, Jiantao; Dai, Zhijun; Gao, Xiaoyan; Yin, Xiaoran; Xi, Wentao; Min, Weili

2014-05-01

213

Soluble urokinase plasminogen activator receptor: an indicator of pneumonia severity in children.  

Science.gov (United States)

Enhanced level of soluble urokinase plasminogen activator receptor (suPAR) level has been associated with activation of the immune system. It may be a novel biomarker for pneumonia severity, yet data on this subject are limited. In the present study we seek to determine the suPAR level in hospitalized children with community-acquired pneumonia (CAP), its correlation with pneumonia severity, and to compare the suPAR level between pneumonia and healthy conditions. The study encompassed a total of 596 children: 447 with pneumonia and 119 healthy. suPAR was measured in 227 out of the 447 pneumonia patients and in all healthy subjects. We used clinical indicators (fever, time for defeverscence, heart and breath rate, saturation, and length of antibiotic treatment and of hospitalization) and laboratory indicators (CRP, procalcitonin, white blood cell count, and sodium) to assess the CAP severity. The finding were that the suPAR concentration in children with pneumonia was significantly higher (median 7.11 ng/mL) than in healthy individuals (4.68 ng/mL). We found a positive correlation between the suPAR and the following factors: fever, time for defeverscence, length of hospital stay, and elevated CRP and procalcitonin levels. There was a reverse correlation with sodium concentration and capillary blood saturation. Moreover, the suPAR level was significantly higher in children with a severe course of pneumonia compared with those having non-severe pneumonia (7.79 vs. 6.87 ng/mL; p?=?0.006). In conclusion, suPAR elevation is observed in pneumonia and may reflect its severity. PMID:25315615

Wrotek, A; Jackowska, T; Pawlik, K

2015-01-01

214

Soluble Urokinase-Type Plasminogen Activator Receptor Levels in Patients With Schizophrenia  

DEFF Research Database (Denmark)

BACKGROUND: The etiology of schizophrenia remains largely unknown but alterations in the immune system may be involved. In addition to the psychiatric symptoms, schizophrenia is also associated with up to 20 years reduction in life span. Soluble urokinase-type plasminogen activator receptor (suPAR) is a protein that can be measured in blood samples and reflects the levels of inflammatory activity. It has been associated with mortality and the development of type 2 diabetes and cardiovascular disease. METHODS: suPAR levels in patients with schizophrenia were compared to healthy controls from the Danish Blood Donor Study. SuPAR levels were dichotomized at >4.0 ng/ml, which is considered the threshold for low grade inflammation. A multiple logistic regression model was used and adjusted for age, sex, and current smoking. RESULTS: In total we included 1009 subjects, 105 cases with schizophrenia (10.4%) and 904 controls (89.6%). The mean suPAR values were 4.01 ng/ml (SD = 1.43) for the cases vs 1.91 ng/ml (SD = 1.35) for the controls (P 4.0 ng/ml yielded: schizophrenia, OR: 46.15 95% CI 22.69-93.87, P < .001; age, OR: 1.02 95% CI 0.99-1.02, P = .15; male sex, OR: 0.70 95% CI 0.35-1.36, P = .29; and current smoking, OR: 3.51 95% CI 1.78-6.94, P < .001. CONCLUSIONS: Patients with schizophrenia had significantly higher suPAR levels than healthy controls. Further studies are warranted to clarify if elevated suPAR levels are involved in the pathophysiology of schizophrenia and/or the increased mortality found in patients with schizophrenia.

Nielsen, Jimmi; RØge, Rasmus

2014-01-01

215

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

216

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

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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

217

Localization of human malignant tumors with radioiodinated recombinant tissue plasminogen activator  

International Nuclear Information System (INIS)

Human recombinant tissue plasminogen activator (tPA), labeled with 131I(1.1 to 6.2 mCi) by the iodogen method, was administered intravenously to 15 patients with various soft-tissue malignant tumors after blocking of thyroidal radioiodine uptake. Gamma camera imaging was performed 4 and 24 hr after injection; three patients were also imaged 5 days following injection. We observed accumulation of radioactivity in primary and secondary lesions in 11 patients. In this preliminary study we did not detect any definite association between the magnitude of uptake and type of tumor. Tumors were usually visualized already after 4 hr but the uptake was more intense at 24 hr. The target-to-nontarget ratios at 24 hr, determined by computer analysis of stored images, varied from less than 1.2 to 2.1. This is the first demonstration of accumulation of radiolabeled tPA in malignant tissue. We do not know the mechanism of the uptake but because tPA is known to be avidly bound to fibrin, a component of the stroma of many malignant tumors, it is possible that [131I]tPA is bound to fibrin rather than taken up by the malignant cell; various possible cell uptake mechanisms are discussed. Due to the relatively early maximal uptake of this radiopharmaceutical it will be possible to substitute 123I for 131I, a possibility suggesting a potential clinical use of radioiodinated tPA for the detection of malignant tumors of various originrigin

218

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

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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

219

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 bcrose 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)

220

Tissue plasminogen activator in trabecular meshwork attenuates steroid induced outflow resistance in mice.  

Science.gov (United States)

Tissue plasminogen activator, a serine protease encoded by the PLAT gene is present in the trabecular meshwork (TM) and other ocular tissues and has been reported to be downregulated by treatment with steroids in vitro. Steroids are known to cause changes in outflow facility of aqueous humor in many species. In the present study, we tested whether overexpression of PLAT can prevent and/or reverse the outflow facility of mouse eyes treated with steroids. Animals received bilateral injection with 20 µl of triamcinolone acetonide (TA) (40 mg/ml) suspension subconjunctivally to induce outflow facility changes. Some animals received unilateral intracameral injection with 2 µl of adenoviral suspension [3-4 x 10(12) virus genomes per milliliter (vg/ml)] carrying sheep PLAT cDNA (AdPLAT) either concurrently with TA injection or one week after TA injection, whereas others received bilateral intracameral injection with 2 µl of adenoviral suspension (9 x 10(12) vg/ml) carrying no transgene (AdNull) concurrently with TA injection. Animals were sacrificed one week after AdPLAT or AdNull treatment. Endogenous mRNA expression levels of mouse PAI-1 and MMP-2, -9 and -13 were also measured using qRT-PCR. Outflow facility one week after AdPLAT administration was increased by 60% and 63% respectively for animals that had not or had been pretreated with steroids. Overexpression of PLAT significantly upregulated expression of PAI-1, MMP-2, -9 and -13 compared to the levels found in TA only treated eyes. These findings suggest that overexpression of PLAT in TM of mouse eyes can both prevent and reverse the decrease in outflow facility caused by steroid treatment and is associated with upregulation of MMPs. PMID:23977299

Kumar, Sandeep; Shah, Shaily; Tang, Hai Michael; Smith, Matthew; Borrás, Teresa; Danias, John

2013-01-01

221

Increased tissue plasminogen activator levels in patients with nonvalvular atrial fibrillation  

Science.gov (United States)

OBJECTIVE: To determine whether plasma tissue plasminogen activator (tPA) levels (a) are higher in patients with novalvular atrial fibrillation (NVAF) than in control subjects in sinus rhythm; (b) differ between NVAF patients with and without a history of an embolic event (transient ischemic attack or embolic stroke); and (c) differ in control subjects with and without a history of thrombotic stroke. DESIGN: Cross-sectional study. SETTING: Internal medicine outpatient group practice and anticoagulation clinic in 2 teaching hospitals. PATIENTS: Seventy-four NVAF patients (24 with and 50 without a history of an embolic event), separated into 3 groups: no prior embolic event and no warfarin use (group 1), no prior embolic event and warfarin use (group 2), and prior embolic event and warfarin use (group 3). Forty control subjects in sinus rhythm (29 without and 11 with prior thrombotic stroke). OUTCOME MEASURES: Plasma tPA levels. RESULTS: The age-adjusted mean tPA levels exceeded the upper limit of normal in all 3 NVAF groups but not in the control groups. The NVAF patients had significantly higher mean tPA levels than the control subjects (p = 0.015). The levels did not differ significantly between the NVAF patients with a history of an embolic event and those without such a history. The control subjects with a history of thrombotic stroke had significantly higher mean tPA levels than the other control subjects (p = 0.03). CONCLUSIONS: NVAF patients, regardless of their history of embolic events, and control patients with a history of thrombotic stroke have higher tPA levels than subjects in sinus rhythm without a history of stroke. A prospective, longitudinal study involving NVAF patients is required to determine whether high baseline tPA levels are associated with, and perhaps causally related to, an increased risk of stroke. PMID:9307554

Kahn, S R; Solymoss, S; Flegel, K M

1997-01-01

222

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

223

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

224

Granulosa-thecal cell interactions in the regulation of plasminogen activator activity during ovarian follicular development in the hen.  

Science.gov (United States)

In the present studies, hen granulosa and thecal cells from the first (F1) and fourth (F4) largest and developing large white follicles (LWF) were cultured alone or cocultured, and plasminogen activator (PA) activity was determined. The PA of the cells (PAc) and the medium (PAm) was measured through use of the chromogenic substrate Val-Leu-Lys-p-nitroanilide, and the total PA (PAt) was calculated. The PA activity of cultured granulosa cells from all stages of follicular development increased with time of culture. Granulosa cell PAc and PAm activity differed with follicular development: the LWF granulosa cells had the highest levels of activity, the F4 had intermediate levels, and the F1 cells had the lowest. Thecal cell PA activity increased during culture but was unaffected by the stage of follicular development. Cocultures of granulosa and thecal cells from F1 follicles exhibited PA activity 2- to 3-fold higher than the sum of the activities of granulosa and thecal cells cultured alone. The PA activity of granulosa cells from LWF was not affected by coculture with thecal cells. Conditioned medium from thecal cells (TCCM) of all stages of follicular development stimulated PAc activity of granulosa cells from F1 and F4 follicles. Conditioned medium from thecal cells of F4 and LWF caused small inhibitory effects on the PAc activity of granulosa cells from LWF. Zymographic analysis of the PA activity of F1 granulosa cell cultures indicates that the enzyme activity is associated with a molecular mass of about 33 kDa, which is consistent with that of urokinase type PA. Thecal cell PA activity was unaffected by granulosa cell-conditioned medium.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8286588

Wang, L; Croze, F; Morley, P; Tsang, B K

1993-11-01

225

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

226

Synergistic and multidimensional regulation of plasminogen activator inhibitor type 1 expression by transforming growth factor type ? and epidermal growth factor  

Energy Technology Data Exchange (ETDEWEB)

The major physiological inhibitor of plasminogen activator, type I plasminogen activator inhibitor (PAI-1), controls blood clotting and tissue remodeling events that involve cell migration. Transforming growth factor type ? (TGF?) and epidermal growth factor (EGF) interact synergistically to increase PAI-1 mRNA and protein levels in human HepG2 and mink Mv1Lu cells. Other growth factors that activate tyrosine kinase receptors can substitute for EGF. EGF and TGF? regulate PAI-1 by synergistically activating transcription, which is further amplified by a decrease in the rate of mRNA degradation, the latter being regulated only by EGF. The combined effect of transcriptional activation and mRNA stabilization results in a rapid 2-order of magnitude increase in the level of PAI-1. TGF? also increases the sensitivity of the cells to EGF, thereby recruiting the cooperation of EGF at lower than normally effective concentrations. The contribution of EGF to the regulation of PAI-1 involves the MAPK pathway, and the synergistic interface with the TGF? pathway is downstream of MEK1/2 and involves phosphorylation of neither ERK1/2 nor Smad2/3. Synergism requires the presence of both Smad and AP-1 recognition sites in the promoter. This work demonstrates the existence of a multidimensional cellular mechanism by which EGF and TGF? are able to promote large and rapid changes in PAI-1 expression.

Song, Xiaoling; Thalacker, F.W.; Nilsen-Hamilton, Marit

2012-04-06

227

Study of the sites of plasminogen molecule which are responsible for inhibitory effect of Lys-plasminogen on platelet aggregation.  

Science.gov (United States)

Plasminogen/plasmin system is involved in such important processes as thrombosis, inflammation and cancer. Plasmin and plasminogen mediate their action through plasminogen-binding proteins on the cell surface. Lys-plasminogen, but not Glu-plasminogen, shows inhibitory effect on platelet aggregation induced by ADP, collagen and thrombin in preparations of both: platelet-rich plasma and washed platelets. We have shown that the kringle domains of Lys-plasminogen mediate interaction of this proenzyme with platelet- surface proteins. The aim of the work is to study the role of certain kringle domains in the inhibitory effect of Lys-plasminogen and to determine possible plasminogen-binding proteins on the platelet surface. All studied plasminogen fragments (K1-3, K4 and K5) abolished the inhibitory effect of Lys-plasminogen on platelet aggregation. We observed that K5 was more effective than K1-3 and K4. Biotin-labeled Lys-plasminogen, Glu-plasminogen and plasminogen fragment K1-3 possessed the highest affinity for actin, whereas the binding of biotin-labeled mini-plasminogen and K4 to actin was negligible. We have suggested that inhibitory effect of Lys-plasminogen is due to the interaction of kringle domains of this proenzyme with membrane-bound proteins which are exposed on the platelet surface during activation and are involved in thrombus formation. PMID:25816591

Roka-Moya, Y M; Zhernossekov, D D; Yusova, E I; Kapustianenko, L G; Grinenko, T V

2014-01-01

228

Cell Signaling by Urokinase-type Plasminogen Activator Receptor Induces Stem Cell–like Properties in Breast Cancer Cells  

OpenAIRE

Signaling by urokinase-type plasminogen activator receptor (uPAR) can cause epithelial-mesenchymal transition (EMT) in cultured breast cancer cells. In this report, we show that uPAR signaling can also induce cancer stem cell (CSC)-like properties. Ectopic overexpression of uPAR in human MDA-MB-468 breast cancer cells promoted emergence of a CD24-/CD44+ phenotype, characteristic of CSCs, while increasing the cell surface abundance of integrin subunits ?1/CD29 and ?6/CD49f that represent ...

Jo, Minji; Eastman, Boryana M.; Webb, Drue L.; Stoletov, Konstantin; Klemke, Richard; Gonias, Steven L.

2010-01-01

229

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

DEFF Research Database (Denmark)

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-methylpentanoyl]-L-tryptophan methylamide) eventually heal, whereas skin wounds in galardin-treated Plg-deficient mice do not heal. Furthermore, plasmin is biochemically detectable in wound extracts from uPA;tPA double-deficient mice. In vivo administration of a plasma kallikrein (pKal)-selective form of the serine protease inhibitor ecotin exacerbates the healing impairment of uPA;tPA double-deficient wounds to a degree indistinguishable from that observed in Plg-deficient mice, and completely blocks the activity of pKal, but not uPA and tPA in wound extracts. These findings demonstrate that an additional plasminogen activator provides sufficient plasmin activity to sustain the healing process albeit at decreased speed in the absence of uPA, tPA and galardin-sensitive MMPs and suggest that pKal plays a role in plasmin generation.

Lund, Leif R; Green, Kirsty A

2006-01-01

230

Radiation-Induced Hypomethylation Triggers Urokinase Plasminogen Activator Transcription in Meningioma Cells1  

Science.gov (United States)

Our previous studies have shown the role of radiation-induced urokinase plasminogen activator (uPA) expression in the progression of meningioma. In the present study, we investigated whether modulation of DNA methylation profiles could regulate uPA expression. Initially, radiation treatment was found to induce hypomethylation in meningioma cells with a decrease in DNA (cytosine-5)-methyltransferase 1 (DNMT1) and methyl-CpG binding domain protein (MBD) expression. However, oxidative damage by H2O2 or pretreatment of irradiated cells with N-acetyl cysteine (NAC) did not show any influence on these proteins, thereby indicating a radiation-specific change in the methylation patterns among meningioma cells. Further, we identified that hypomethylation is coupled to an increase in uPA expression in these cells. Azacytidine treatment induced a dose-dependent surge of uPA expression, whereas pre-treatment with sodium butyrate inhibited radiation-induced uPA expression, which complemented our prior results. Methylation-specific polymerase chain reaction on bisulfite-treated genomic DNA revealed a diminished methylation of uPA promoter in irradiated cells. Transfection with small hairpin RNA (shRNA)-expressing plasmids targeting CpG islands of the uPA promoter showed a marked decline in uPA expression with subsequent decrease in invasion and proliferation of meningioma cells. Further, radiation treatment was found to recruit SP1 transcription factor, which was abrogated by shRNA treatment. Analysis on signaling events demonstrated the activation of MAP kinase kinase (MEK)-extracellular signal-regulated kinase (ERK) in radiation-treated cells, while U0126 (MEK/ERK inhibitor) blocked hypomethylation, recruitment of SP1, and uPA expression. In agreement with our in vitro data, low DNMT1 levels and high uPA were found in intracranial tumors treated with radiation compared to untreated tumors. In conclusion, our data suggest that radiation-mediated hypomethylation triggers uPA expression in meningioma cells. PMID:23441133

Velpula, Kiran Kumar; Gogineni, Venkateswara Rao; Nalla, Arun Kumar; Dinh, Dzung H; Rao, Jasti S

2013-01-01

231

Radiation-induced hypomethylation triggers urokinase plasminogen activator transcription in meningioma cells.  

Science.gov (United States)

Our previous studies have shown the role of radiation-induced urokinase plasminogen activator (uPA) expression in the progression of meningioma. In the present study, we investigated whether modulation of DNA methylation profiles could regulate uPA expression. Initially, radiation treatment was found to induce hypomethylation in meningioma cells with a decrease in DNA (cytosine-5)-methyltransferase 1 (DNMT1) and methyl-CpG binding domain protein (MBD) expression. However, oxidative damage by H(2)O(2) or pretreatment of irradiated cells with N-acetyl cysteine (NAC) did not show any influence on these proteins, thereby indicating a radiation-specific change in the methylation patterns among meningioma cells. Further, we identified that hypomethylation is coupled to an increase in uPA expression in these cells. Azacytidine treatment induced a dose-dependent surge of uPA expression, whereas pre-treatment with sodium butyrate inhibited radiation-induced uPA expression, which complemented our prior results. Methylation-specific polymerase chain reaction on bisulfite-treated genomic DNA revealed a diminished methylation of uPA promoter in irradiated cells. Transfection with small hairpin RNA (shRNA)-expressing plasmids targeting CpG islands of the uPA promoter showed a marked decline in uPA expression with subsequent decrease in invasion and proliferation of meningioma cells. Further, radiation treatment was found to recruit SP1 transcription factor, which was abrogated by shRNA treatment. Analysis on signaling events demonstrated the activation of MAP kinase kinase (MEK)-extracellular signal-regulated kinase (ERK) in radiation-treated cells, while U0126 (MEK/ERK inhibitor) blocked hypomethylation, recruitment of SP1, and uPA expression. In agreement with our in vitro data, low DNMT1 levels and high uPA were found in intracranial tumors treated with radiation compared to untreated tumors. In conclusion, our data suggest that radiation-mediated hypomethylation triggers uPA expression in meningioma cells. PMID:23441133

Velpula, Kiran Kumar; Gogineni, Venkateswara Rao; Nalla, Arun Kumar; Dinh, Dzung H; Rao, Jasti S

2013-02-01

232

Radiation-Induced Hypomethylation Triggers Urokinase Plasminogen Activator Transcription in Meningioma Cells  

Directory of Open Access Journals (Sweden)

Full Text Available Our previous studies have shown the role of radiation-induced urokinase plasminogen activator (uPA expression in the progression of meningioma. In the present study, we investigated whether modulation of DNA methylation profiles could regulate uPA expression. Initially, radiation treatment was found to induce hypomethylation in meningioma cells with a decrease in DNA (cytosine-5-methyltransferase 1 (DNMT1 and methyl-CpG binding domain protein (MBD expression. However, oxidative damage by H2O2 or pretreatment of irradiated cells with N-acetyl cysteine (NAC did not show any influence on these proteins, thereby indicating a radiation-specific change in the methylation patterns among meningioma cells. Further, we identified that hypomethylation is coupled to an increase in uPA expression in these cells. Azacytidine treatment induced a dose-dependent surge of uPA expression, whereas pre-treatment with sodium butyrate inhibited radiation-induced uPA expression, which complemented our prior results. Methylation-specific polymerase chain reaction on bisulfite-treated genomic DNA revealed a diminished methylation of uPA promoter in irradiated cells. Transfection with small hairpin RNA (shRNA-expressing plasmids targeting CpG islands of the uPA promoter showed a marked decline in uPA expression with subsequent decrease in invasion and proliferation of meningioma cells. Further, radiation treatment was found to recruit SP1 transcription factor, which was abrogated by shRNA treatment. Analysis on signaling events demonstrated the activation of MAP kinase kinase (MEK-extracellular signal-regulated kinase (ERK in radiation-treated cells, while U0126 (MEK/ERK inhibitor blocked hypomethylation, recruitment of SP1, and uPA expression. In agreement with our in vitro data, low DNMT1 levels and high uPA were found in intracranial tumors treated with radiation compared to untreated tumors. In conclusion, our data suggest that radiation-mediated hypomethylation triggers uPA expression in meningioma cells.

Kiran Kumar Velpula

2013-02-01

233

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

OpenAIRE

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...

Vieira, Monica L.; Vasconcellos, Silvio A.; Gonc?ales, Amane P.; Morais, Zenaide M.; Nascimento, Ana L. T. O.

2009-01-01

234

Prostate Cancer Cell-Derived Urokinase-Type Plasminogen Activator Contributes to Intraosseous Tumor Growth and Bone Turnover  

Directory of Open Access Journals (Sweden)

Full Text Available A variety of proteases have been implicated in prostate cancer (PC bone metastasis, but the individual contributions of these enzymes remain unclear. Urokinase-type plasminogen activator (uPA, a serine protease, can activate plasminogen and stimulate signaling events on binding its receptor uPAR. In the present study, we investigated the functional role of PC cell-associated uPA in intraosseous tumor growth and bone matrix degradation. Using a severe combined immunodeficient-human mouse model, we found that PC3 cells were the major source of uPA in the experimental bone tumor. Injection of uPA-silenced PC3 cells in bone xenografts resulted in significant reduction of bone tumor burdens and protection of trabecular bones from destruction. The suppressed tumor growth was associated with the level of uPA expression but not with its activity. An increase in the expression of PAI-1, the endogenous uPA inhibitor, was found during in vitro tumor-stromal interactions. Up-regulation of PAI-1 in bone stromal cells and preosteoclasts/osteoblasts was due to soluble factor(s released by PC cells, and the enhanced PAI-1 expression in turn stimulated PC cell migration. Our results indicate that both tumor-derived uPA and tumor-stroma-induced PAI-1 play important roles in intraosseous metastatic PC growth through regulation of a uPA-uPAR-PAI-1 axis by autocrine/paracrine mechanisms.

Zhong Dong

2008-05-01

235

Expression analysis and purification of human recombinant tissue type plasminogen activator (rt-PA) from transgenic tobacco plants.  

Science.gov (United States)

Recombinant tissue-type plasminogen activator (rt-PA) has been produced in different hosts. In this research, transgenic tobacco was selected for production of human rt-PA. Transgenic plants were analyzed by polymerase chain reaction (PCR) and reverse-transcription (RT)-PCR. The protein was extracted by Lysine Sepharose chromatography column and was further purified by HiTrap desalting column. The function of eluted protein was analyzed on zymography gel. The results showed that the 1.7-kb cDNA of tissue-type plasminogen activator (t-PA) (as well as a shortened 650-bp transcript of t-PA) has been expressed in transgenic plants. The anticipated 63-kD protein band and an additional 53-kD protein were observed in transgenic plants. Finally, zymography assay revealed that the purified rt-PA has anticipated appropriate activity comparable to a positive control drug (Alteplase). On the whole, we can say that transgenic tobacco is a good alternative host for production of t-PA. PMID:21442553

Nabiabad, Haidar Saify; Yaghoobi, Mohammad Mehdi; Javaran, Mokhtar Jalali; Hosseinkhani, Saman

2011-01-01

236

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

237

Metastatic behavior of human melanoma cell lines in nude mice correlates with urokinase-type plasminogen activator, its type-1 inhibitor, and urokinase-mediated matrix degradation  

OpenAIRE

Five out of six human melanoma cell lines tested were able to degrade in vitro a smooth muscle cell extracellular matrix in a plasmin- dependent way. In three of these five cell lines, this process was mediated by tissue-type plasminogen activator (t-PA) and in the other two cell lines by urokinase-type plasminogen activator (u-PA). All melanoma cell lines produced t-PA mRNA and protein, whereas only the two cell lines showing u-PA-mediated matrix degradation produced u-PA mRNA and protein. T...

1991-01-01

238

Plasminogen and plasmin in Alzheimer's disease.  

Science.gov (United States)

In Alzheimer's disease, abnormal accumulation of A? leads to neuronal death and impaired A? degradation may play an important role in this accumulation. Plasmin is the key active protease in the plasminogen system and is capable of cleaving A?. Here we investigate plasminogen mRNA levels, plasminogen and plasmin protein levels and plasmin activity levels in post-mortem AD and control brain tissue. Plasminogen and plasmin distribution in the human brain was demonstrated by immunoperoxidase staining. Plasminogen mRNA levels were measured in 20 AD, 20 control and 15 Vascular dementia (VaD) brains by real-time PCR (RT-PCR). In an expanded cohort of 38 AD and 38 control brains plasminogen and plasmin protein levels were measured by dot blot and Western blot, respectively, while plasmin activity levels were measured by fluorogenic assay. Plasminogen and plasmin were present mainly in the neurons. Plasminogen mRNA levels were unaltered in the AD or VaD groups compared to the controls. Plasminogen and plasmin protein levels were not significantly altered in AD compared to controls. Plasmin activity was reduced in AD but this did not reach statistical significance. In contrast to some studies these data do not support the involvement of plasmin in the abnormal accumulation of A? in AD. PMID:20709033

Barker, Rachel; Love, Seth; Kehoe, Patrick G

2010-10-01

239

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

240

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.

2000-01-01

241

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 symecting 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)

242

Soluble urokinase-type plasminogen activator receptor is a novel biomarker predicting acute exacerbation in COPD  

Directory of Open Access Journals (Sweden)

Full Text Available Aziz Gumus,1 Nejat Altintas,2 Halit Cinarka,1 Aynur Kirbas,3 Muge Haziroglu,1 Mevlut Karatas,1 Unal Sahin1 1Department of Pulmonary Medicine, School of Medicine, Recep Tayyip Erdogan University, Rize, Turkey; 2Department of Pulmonary Medicine, School of Medicine, Namik Kemal University, Tekirdag, Turkey; 3Department of Clinical Biochemistry, School of Medicine, Recep Tayyip Erdogan University, Rize, Turkey Background: Chronic obstructive pulmonary disease (COPD is a chronic inflammatory condition, and progresses with acute exacerbations. (AE. During AE, levels of acute phase reactants such as C-reactive protein (CRP and inflammatory cells in the circulation increase. Soluble urokinase-type plasminogen activator receptor (suPAR levels increase in acute viral and bacterial infections and in diseases involving chronic inflammation. The purpose of this study was to investigate the effectiveness of suPAR in predicting diagnosis of AE of COPD (AE-COPD and response to treatment. Methods: The study population consisted of 43 patients diagnosed with AE-COPD and 30 healthy controls. suPAR, CRP, and fibrinogen levels were measured on the first day of hospitalization and on the seventh day of treatment. Results: We found that fibrinogen (P<0.001, CRP (P<0.001, and suPAR (P<0.001 were significantly higher in patients with AE-COPD than in healthy controls. Fibrinogen (P<0.001, CRP (P=0.001, and suPAR (P<0.001 were significantly decreased by the seventh day of treatment. However, the area under receiver operator characteristic curve showed that suPAR is superior to CRP and fibrinogen in distinguishing AE-COPD. There was a correlation between fibrinogen, CRP, and suPAR. However, only fibrinogen was a powerful predictor of suPAR in multiple linear regression. In multiple logistic regression, only suPAR and fibrinogen were strong predictors of AE-COPD (P=0.002 and P=0.014, respectively. Serum suPAR was negatively correlated with forced expiratory volume in 1 second (r=-478, P=0.001. Conclusion: suPAR is a marker of acute inflammation. It is well correlated with such inflammation markers as CRP and fibrinogen. suPAR can be used as a predictor of AE-COPD and in monitoring response to treatment. Keywords: biomarker, chronic obstructive pulmonary disease, inflammation, suPAR

Gumus A

2015-02-01

243

Characterisation of urokinase plasminogen activator receptor variants in human airway and peripheral cells  

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Full Text Available Abstract Background Expression of the urokinase plasminogen activator receptor (UPAR has been shown to have clinical relevance in various cancers. We have recently identified UPAR as an asthma susceptibility gene and there is evidence to suggest that uPAR may be upregulated in lung diseases such as COPD and asthma. uPAR is a key receptor involved in the formation of the serine protease plasmin by interacting with uPA and has been implicated in many physiological processes including proliferation and migration. The current aim was to determine key regulatory regions and splice variants of UPAR and quantify its expression in primary human tissues and cells (including lung, bronchial epithelium (HBEC, airway smooth muscle (HASM and peripheral cells. Results Using Rapid Amplification of cDNA Ends (RACE a conserved transcription start site (-42 to -77 relative to ATG was identified and multiple transcription factor binding sites predicted. Seven major splice variants were identified (>5% total expression, including multiple exon deletions and an alternative exon 7b (encoding a truncated, soluble, 229aa protein. Variants were differentially expressed, with a high proportion of E7b usage in lung tissue and structural cells (55–87% of transcripts, whereas classical exon 7 (encoding the GPI-linked protein was preferentially expressed in peripheral cells (~80% of transcripts, often with exon 6 or 5+6 deletions. Real-time PCR confirmed expression of uPAR mRNA in lung, as well as airway and peripheral cell types with ~50–100 fold greater expression in peripheral cells versus airway cells and confirmed RACE data. Protein analysis confirmed expression of multiple different forms of uPAR in the same cells as well as expression of soluble uPAR in cell supernatants. The pattern of expression did not directly reflect that seen at the mRNA level, indicating that post-translational mechanisms of regulation may also play an important role. Conclusion We have identified multiple uPAR isoforms in the lung and immune cells and shown that expression is cell specific. These data provide a novel mechanism for uPAR regulation, as different exon splicing may determine uPAR function e.g. alternative E7b results in a soluble isoform due to the loss of the GPI anchor and exon deletions may affect uPA (ligand and/or integrin binding and therefore influence downstream pathways. Expression of different isoforms within the lung should be taken into consideration in studies of uPAR in respiratory disease.

Sayers Ian

2009-07-01

244

Assessment of plasminogen synthesis in vitro by mouse tumor cells using a competition radioimmunoassay for mouse plasminogen  

International Nuclear Information System (INIS)

A sensitive, specific competition radioimmunoassay for mouse plasmin(ogen) has been developed in order to determine whether mouse tumor cells can synthesize plasminogen in vitro. The rabbit anti-BALB/c mouse plasminogen antibodies used in the assay react with the plasminogen present in serum from BALB/c, C3H, AKR and C57BL/6 mice, and also recognized mouse plasmin. The competition radiommunoassay can detect as little as 50 ng of mouse plasminogen. No competition was observed with preparations of fetal calf, human and rabbit plasminogens. A variety of virus-transformed and mouse tumor cell lines were all found to contain less than 100 ng mouse plasminogen/mg of cell extract protein. Thus, if the plasminogen activator/plasmin system is important in the growth or movement of this group of tumor cells, the cells will be dependent upon the circulatory system of the host for their plasminogen supply. (Auth.)

245

Inhibition of histone deacetylase activity down-regulates urokinase plasminogen activator and matrix metalloproteinase-9 expression in gastric cancer.  

Science.gov (United States)

Histone acetylation and deacetylaion play important roles in chromatin remodeling and gene expression. An imbalance of these reactions leads to aberrant behavior of the cells in the cell cycle, which in turn contributes to carcinogenesis. Histone deacetylase (HDAC) inhibitors have been shown to have anti-tumor effects in clinical trials. However, the exact mechanisms by which HDAC inhibitors exert anti-tumor effects and modulate gene expression are not completely understood, and remain a subject of intense investigation. In the current study, we determined whether HDACs regulate urokinase plasminogen activator (uPA), matrix metalloproteinase-9 (MMP-9), and tumor invasion. Using cDNA microarray analysis, we found that hepatocyte growth factor (HGF) induced HDAC5 expression in gastric cancer cell lines, NUGC-3 and MKN-28. TSA, a HDAC inhibitor, decreased HGF-induced HADC-5 expression and also repressed uPA and MMP-9 expression. TSA inhibited cell proliferation in both cell lines. In vitro Matrigel invasion assays showed that the HDAC inhibitor decreased cancer cell invasion. Furthermore, GO6976, a PKC inhibitor, significantly inhibited not only HGF-induced HDAC5 expression but also cell invasion. These results demonstrated that HDACs regulate HGF-induced uPA and MMP-9 expression through a PKC-dependent signal pathway in gastric cancer cells. Our data probably suggest that such activities serve as anti-tumor mechanisms of the HDAC inhibitor. PMID:20559690

Lee, Kyung Hee; Choi, Eun Young; Kim, Min Kyoung; Kim, Kyeong Ok; Jang, Byung Ik; Kim, Se Won; Kim, Sang Woon; Song, Sun Kyo; Kim, Jae-Ryong

2010-10-01

246

Lack of association between plasminogen activator inhibitor type-1 (PAI-1) gene 4G/5G polymorphism and osteoarthritis.  

Science.gov (United States)

This study was conducted in Turkish osteoarthritis patients to determine the frequency of 4G/5G polymorphism genotypes of plasminogen activator inhibitor type-1 gene and to examine the role of this polymorphism in osteoarthritis development. Genomic DNA obtained from 200 persons (140 patients with osteoarthritis and 60 healthy controls) was used in the study. DNA was amplified by polymerase chain reaction using 4G allele- and 5G allele-specific primers. Polymerase chain reaction products were assessed with CCD camera by being exposed to 2% agarose gel electrophoresis. No statistically significant difference between the groups with respect to genotype distribution was found (P > 0.05) in the study. The 4G allele frequency was indicated as 44% and 5G allele was as 56% in patients, whereas this was 45-55% in the control group. This study has established that 4G/5G polymorphism genotypes of plasminogen activator inhibitor type-1 gene do not play a role in the development of osteoarthritis in the Turkish population. PMID:21240498

Bayram, Banu; Sayin, Emrah; Erkasap, Nilüfer; Onlü, Harun; Ozkurt, Mete; Sahin, Fezan; Türko?lu, Züleyha

2012-01-01

247

Mammalian protein secretion without signal peptide removal. Biosynthesis of plasminogen activator inhibitor-2 in U-937 cells  

International Nuclear Information System (INIS)

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

248

Mammalian protein secretion without signal peptide removal. Biosynthesis of plasminogen activator inhibitor-2 in U-937 cells  

Energy Technology Data Exchange (ETDEWEB)

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

Ye, R.D.; Wun, T.C.; Sadler, J.E.

1988-04-05

249

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

250

Collagen metabolism and enzymes of the urokinase plasminogen activator system in chronic myeloproliferative disorders: correlation between plasma-soluble urokinase plasminogen activator receptor and serum markers for collagen metabolism.  

Science.gov (United States)

Extracellular proteolytic enzymes of the urokinase-type plasminogen activator (uPA) system and the family of metalloproteinases (MMPs) catalyse the matrix degradation and remodelling processes characteristic of invasive malignant disorders. In a cohort of 50 patients with chronic myeloproliferative disorders (MPD) serum markers for collagen metabolism were compared to plasma levels of enzymes of the uPA and MMP system. Serum aminoterminal propeptide of type III procollagen (S-PIIINP) (P levels differed significantly with the highest values found in patients with MF (MF vs. PV vs. ET; P = 0.0027). Serum concentration of carboxyterminal telopeptide of type I collagen (S-ICTP) (P = 0.0006), reflecting type I collagen degradation, was significantly higher in patients compared with controls (median 4.0 micro g/L vs. 2.7 micro g/L). When comparing S-ICTP measurements between patient subgroups and controls there were only significantly higher values among MF and PV patients (MF vs. controls; P plasma-soluble urokinase plasminogen receptor (suPAR) disclosed a significant relationship between suPAR and S-PIIINP (r = 0.48; P = 0.0009), S-hyaluronan (r = 0.56; P Plasma levels of MMP-2 and -9 were not correlated to serum markers for collagen metabolism. These findings suggest that enzymes of the uPA system might participate in the bone marrow remodelling processes characteristic of MPD. PMID:12950237

Jensen, Morten Krogh; Riisbro, Rikke; Holten-Andersen, Mads N; Brown, Peter de Nully; Junker, Peter; Brünner, Nils; Hasselbalch, Hans C

2003-10-01

251

Cytokeratin 8 ectoplasmic domain binds urokinase-type plasminogen activator to breast tumor cells and modulates their adhesion, growth and invasiveness  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Generation of plasmin is a characteristic of tumor cells, promoting the degradation of extracellular matrix, tumor progression and metastasis. The process is accelerated if plasminogen and plasminogen activator are bound to their cell surface receptors. Results In this study we show that the monoclonal antibody that recognizes an epitope on the cytokeratin 8 (CK8 ectoplasmic domain (anti-CK MAb inhibits plasminogen activation mediated by urokinase-type plasminogen activator (uPA in MCF-7 and MCF-10A neoT cells. The ectoplasmic domain of CK8 acts as a binding site for plasminogen, however, by using confocal microscopy, we demonstrated that it is also co-localized with uPA. CK8, therefore, function also as a receptor for uPA on the cell surface, and the presence of anti-CK MAb may prevent the binding of uPA to a designated CK8 motif. The consequent inhibition of plasmin generation resulted in changed cell morphology, enhanced cell adhesion to fibronectin, reduced invasion potential, and an enhanced G1/S transition. Moreover, surface plasmon resonance analysis showed that the synthetic dodecapeptide corresponding to the epitope sequence (VKIALEVEIATY, binds uPA in the nanomolar range. Conclusion These novel findings suggest a model in which CK8, together with uPA, plasminogen and fibronectin, constitutes a signaling platform capable of modulating cell adhesion/growth-dependent signal transduction in breast tumor cells. Anti-CK MAb, which competes for the binding site for uPA, could be used as an agent to reduce the invasive potential of breast tumor cells.

Doljak Bojan

2009-10-01

252

Reactive oxygen species regulate urokinase plasminogen activator expression and cell invasion via mitogen-activated protein kinase pathways after treatment with hepatocyte growth factor in stomach cancer cells  

OpenAIRE

Abstract Background Reactive oxygen species (ROS) are closely associated with the intracellular signal cascade, thus strongly implicating involvement in tumor progression. However, the mechanism by which ROS are generated and how ROS target downstream molecules to trigger tumor metastasis is unclear. In this study, we investigated the underlying signal pathways in ROS-induced urokinase plasminogen activator (uPA) expression in the human gastric cancer cells, NUGC-3 and MKN-28. Methods and Res...

Kim Jae-Ryong; Kim Sang; Lee Kyung

2009-01-01

253

Fibrinolytic activity in a human fibrosarcoma cell line and evidence for the induction of plasminogen activator secretion during tumor formation.  

Science.gov (United States)

Seven clones were isolated from the HT1080 human fibrosarcoma cell line using a fibrinagarose overlay technique. Three of these clones induced lysis of the fibrin overlay, whereas four did not. The extracellular and intracellular levels of protease were then measured using 125I-fibrin plates incubated with acid-treated human serum. The extracellular protease can be directly assayed in the medium from cells incubated with 10% fetal calf serum. Although there were large differences in the amounts of protease secreted by these two sets of clones, the intracellular levels of protease were similar. No significant differences were found between the abilities of the cells to grow in soft agar or as tumors in immunosuppressed hamsters. However, cells grown from tumors derived from all the low secretors of protease showed an increase in the amount of protease secreted. It appeared, therefore, that the secretion of protease might be selected for or induced during tumor growth. Further detailed studies with one of the low secreting clones (clone E) suggested an inductive rather than a selective mechanism for this increase in extracellular plasminogen activator. PMID:1182804

Jones, P A; Laug, W E; Benedict, W F

1975-10-01

254

Regulation of proteinases during mouse peri-implantation development: urokinase-type plasminogen activator expression and cross talk with matrix metalloproteinase 9.  

Science.gov (United States)

Trophoblast cells express urokinase-type plasminogen activator (PLAU) and may depend on its activity for endometrial invasion and tissue remodeling during peri-implantation development. However, the developmental regulation, tissue distribution, and function of PLAU are not completely understood. In this study, the expression of PLAU and its regulation by extracellular matrix proteins was examined by RT-PCR, immunocytochemistry, and plasminogen-casein zymography in cultured mouse embryos. There was a progressive increase in Plau mRNA expression in blastocysts cultured on gestation days 4-8. Tissue-type plasminogen activator (55?kDa) and PLAU (a triplet of 40, 37, and 31?kDa) were present in conditioned medium and embryo lysates, and were adsorbed to the culture plate surface. The temporal expression pattern of PLAU, according to semi-quantitative gel zymography, was similar in non-adhering embryos and embryos cultured on fibronectin, laminin, or type IV collagen, although type IV collagen and laminin upregulated Plau mRNA expression. Immunofluorescence revealed PLAU on the surface of the mural trophectoderm and in non-spreading giant trophoblast cells. Exogenous human plasminogen was transformed to plasmin by cultured embryos and activated endogenous matrix metalloproteinase 9 (MMP9). Indeed, the developmental expression profile of MMP9 was similar to that of PLAU. Our data suggest that the intrinsic developmental program predominantly regulates PLAU expression during implantation, and that PLAU could be responsible for activation of MMP9, leading to localized matrix proteolysis as trophoblast invasion commences. PMID:21075828

Martínez-Hernández, M G; Baiza-Gutman, L A; Castillo-Trápala, A; Armant, D Randall

2011-02-01

255

Effects of a High-Fat Diet on Spontaneous Metastasis of Lewis Lung Carcinoma in Plasminogen Activator Inhibitor-1 Deficient and Wild-Type Mice  

OpenAIRE

This study investigated the effects of a high-fat diet on spontaneous metastasis of Lewis lung carcinoma (LLC) in plasminogen activator inhibitor-1 deficient (PAI-1?/?) and wild-type mice. The high-fat diet increased the number of pulmonary metastases by 60% (p

Yan, Lin; Demars, Lana C.

2014-01-01

256

Effects of a high-fat diet on spontaneous metastasis of Lewis lung carcinoma in plasminogen activator inhibitor-1 deficient and wild-type mice  

Science.gov (United States)

We investigated the effects of plasminogen activator inhibitor-1 (PAI-1) deficiency on spontaneous metastasis of Lewis lung carcinoma (LLC) in PAI-1 deficient (PAI-1-/-) and wildtype mice (C57BL/6J background) fed the AIN93G diet or that diet modified with 45% calories from fat. The high-fat diet i...

257

Influence of macrophage products on the release of plasminogen activator, collagenase, beta-glucuronidase and prostaglandin E2 by articular chondrocytes.  

Science.gov (United States)

We describe the effects of products of mononuclear phagocytes on the secretory activity of chondrocytes. The primary confluent cultures of rabbit articular chondrocytes were exposed to standard medium alone or enriched with conditioned medium obtained from cultures of rabbit peritoneal macrophages, the mouse macrophage cell line P388D1 or human blood mononuclear cells. Four markers of release were assessed, the neutral proteinases plasminogen activator and collagenase, the acid hydrolase beta-glucuronidase and prostaglandin E2, and the kinetics of their changes were monitored. Chondrocytes that were cultured in standard medium secreted large amounts of plasminogen activator, some beta-glucuronidase, but no collagenase, and released only minor amounts of prostaglandin E2. The addition of conditioned medium from rabbit macrophages induced a rapid release of large quantities of prostaglandin E2 and an abundant secretion of collagenase, while abolishing or strongly decreasing plasminogen activator secretion. In addition, beta-glucuronidase secretion was markedly enhanced. The decrease in secretion of plasminogen activator appeared to reflect a diminished production, since no evidence was found for the generation of inhibitors or for an accelerated extracellular breakdown of the enzyme. Conditioned media of the mouse and human mononuclear cells influenced the secretory activities of rabbit articular chondrocytes in a similar way, suggesting that the factor (or factors) acting on chondrocytes is produced by a variety of macrophages, and that its action is not species-restricted. The time course and concentration-dependence of the effects observed indicate that the secretion of plasminogen activator and collagenase are influenced in a strictly reciprocal fashion by the macrophage products. The release of prostaglandin E2 paralleled that of collagenase. PMID:6331394

Evêquoz, V; Schnyder, J; Trechsel, U; Baggiolini, M; Fleisch, H

1984-04-15

258

Expression of the receptor for urokinase-type plasminogen activator in normal and neoplastic blood cells and hematopoietic tissue  

DEFF Research Database (Denmark)

Expression of the receptor for the urokinase type plasminogen activator (uPAR) has been studied by flow cytometry and immunohistology in normal blood and bone marrow cells, in vitro activated lymphoid cells, and tissue samples from reactive lymph nodes (n = 6), thymus (n = 2) and malignant lymphomas (n = 82), or leukemias (n = 32). HL-60 myeloid precursor cells and CD34-positive normal stem cells also were analyzed. In the normal cells, staining was confined to monocytes, macrophages, neutrophils, and myeloid precursors. No labelling was seen of normal or activated lymphoid cells. Purified CD34-positive hematopoietic progenitors were uPAR negative, but expressed uPAR during differentiation in short-term liquid culture stimulated in vitro by recombinant interleukin (IL)-1, IL-3, IL-6, granulocyte-macrophage colony stimulating factor (CSF), granulocyte-CSF, and stem cell factor. Enhanced uPAR expression was also seen in HL-60 cells after induction of differentiation with dimethyl sulfoxide or 1 alpha,25-dihydroxyvitamin D3. In lymphomas and leukemias, the staining pattern was similar to that seen in the normal cells with labelling of monocytic and myeloid that seen in the normal cells with labelling of monocytic and myeloid malignancies, but not of the neoplastic cells in B-cell or T-cell lymphomas or Hodgkin's disease. In conclusion, uPAR is a differentiation marker for myeloid and monocytic cells, and may act to facilitate migration of these cells in normal and pathologic conditions by cell-associated plasminogen activation. Whether expression of uPAR in myeloid and monocytic malignancies relates to their growth and behavior will be an important topic for investigations in the future.

Plesner, T; Ralfkiaer, E

1994-01-01

259

Signal transduction in the platelet activation induced by IgG anti-streptokinase and anisoylated plasminogen-streptokinase activator complex.  

Science.gov (United States)

Streptokinase (SK) is one of the plasminogen activators currently used in therapeutics. SK antibodies may appear in the blood after thrombolytic therapy with SK or after-hemolytic streptococci infection. Such antibodies may both activate platelets and neutralize the ability of SK to convert plasminogen into plasmin. We previously demonstrated that platelet activation induced by the combination of IgG anti-SK and anisoylated plasminogen-SK activator complex (APSAC) is mediated by Fc gamma RIIa1 receptor. However, the mechanism by which IgG anti-SK and APSAC (or SK) transduce an activating signal across the platelet plasma membrane remains unknown. We have demonstrated in the present study that the platelet aggregation induced by the combination of IgG anti-SK and APSAC is accompanied by an increase in inositol phosphate, Ca2+ mobilization and thromboxane (Tx) A2 generation. Neomycin, erbstatin and GF 109203X, which inhibit phospholipase C (PLC), protein tyrosine kinase (PTK) and protein kinase C (PKC) activities, respectively, abolished platelet aggregation induced by IgG anti-SK plus APSAC, indicating the pivotal roles of the PLC, PTK and PKC pathways in this immunological activation. In addition, TxA2 generation is also important since aspirin, a cyclooxygenase inhibitor and SQ 29548, a TxA2 receptor antagonist, showed significant inhibition of the platelet response. The contribution of released ADP was confirmed using apyrase, which significantly inhibited IgG anti-SK plus APSAC-induced platelet aggregation. Finally, WEB 2086, a platelet-activating factor (PAF) receptor antagonist, was not effective, indicating that PAF is not involved in this process. APSAC- or SKinduced platelet activation may limit the therapeutic effectiveness of the drug and may contribute to the pathogenesis of early reocclusion. The study of the mechanism leading to APSAC-induced platelet activation could be relevant for a better understanding of the physiopathology of immune complex disorder diseases and thrombolytic treatment failure. PMID:16793641

1997-04-01

260

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

261

Biochemical actions of glucocorticoids on macrophages in culture. Specific inhibition of elastase, collagenase, and plasminogen activator secretion and effects on other metabolic functions  

Energy Technology Data Exchange (ETDEWEB)

The effects of glucocorticoids on biochemical functions of macrophages from man, mouse, rabbit, and guinea pig were examined. Secretion of plasminogen activator by human peripheral blood monocytes was decreased 50% with 1 nM dexamethasone. Differentiation of murine monocytic and granulocytic colonies in agar from bone marrow precursors was decreased 50% at 7 days with 20 nM dexamethasone. Secretion of elastase, collagenase, and plasminogen activator by resident and thioglycollate-elicited mouse peritoneal macrophages was decreased by dexamethasone, cortisol, and triamcinolone acetonide (1 to 1,000 nM), but not by progesterone, estradiol, and dihydrotestosterone (1,000 nM); in contast, secretion of lysozyme was not affected by glucocorticoids. The inhibition of macrophage secretion by dexamethasone was both time and dose dependent. Inhibition of macrophage secretion increased with increasing glucocorticoid concentration. Half-maximum inhibition of secretion of elastase, collagenase, and plasminogen activator was seen at dexamethasone concentrations (1 to 10 nM) similar to those that half-saturated the specific glucocorticoid receptors. At high concentrations of dexamethasone (100 to 1,000 nM) the secretion of plasminogen activator was inhibited to a greater extent (>95%) than the secretion of elastase (60 to 80%).Progesterone alone had no effect on secretion, but blocked the inhibitory effects of dexamethasone and cortisol. Secretion of collagenase, neutral proteinases, and plasminogen activator by elicited rabbit alveolar macrophages was inhibited with glucocorticoids (0.1 to 100 nM) but not with progesterone or sex steroids. Secretion of a neutral elastinolytic proteinase by guinea pig alveolar macrophages was also inhibited by dexamethasone.

Werb, Z.

1978-01-01

262

IL-1?-stimulated urokinase plasminogen activator expression through NF-?B in gastric cancer after HGF treatment.  

Science.gov (United States)

The potential of hepatocyte growth factor (HGF) to regulate the expression of urokinase plasminogen activator (uPA) in a gastric cancer cell is not widely acknowledged. To identify the genes associated with the plasminogen activator proteolytic axis by HGF, we used cDNA microarray technology and selected genes upregulated or downregulated in two gastric cell lines (NUGC-3 and MKN-28). First, IL-1? RNA and protein were confirmed to be upregulated. Then, we investigated the effect of IL-1? induced by HGF on the uPA system, facilitating the migration and invasion of cancer cells in the metastatic process. The role for IL-1? in HGF-induced upregulation of uPA was determined by knockdown of IL-1? with IL-1? shRNA and a chromatin immune precipitation assay. The levels of IL-1? and uPA were upregulated in cells treated with HGF in a dose-dependent manner. HGF-induced upregulation of uPA was suppressed by IL-1? knockdown. HGF enhanced the binding activity of NF-?B to the uPA promoter in control cells, but not in the IL-1? shRNA cells. We confirmed the functional role of HGF inactivation of the uPA promoter by a reporter gene assay. Downregulation of IL-1? using IL-1? shRNA also decreased cell proliferation and in vitro cell invasion. IL-1? stimulated uPA expression through ERK and NF-?B in gastric cancer, which may therefore be promising targets for gastric cancer therapy. PMID:24626561

Lee, Kyung Hee; Choi, Eun Young; Koh, Sung Ae; Kim, Min Kyoung; Jang, Byung Ik; Kim, Sang Woon; Kim, Jae-Ryong

2014-05-01

263

Serase-1B, a new splice variant of polyserase-1/TMPRSS9, activates urokinase-type plasminogen activator and the proteolytic activation is negatively regulated by glycosaminoglycans.  

Science.gov (United States)

Polyserase-1 (polyserine protease-1)/TMPRSS9 (transmembrane serine protease 9) is a type II transmembrane serine protease (TTSP) that possesses unique three tandem serine protease domains. However, the physiological function of each protease domain remains poorly understood. We discovered a new splice variant of polyserase-1, termed Serase-1B, which contains 34 extra amino acids consisting a SEA module (a domain found in sea urchin sperm protein, enterokinase and agrin) adjacent to the transmembrane domain and the first protease domain with a mucin-like box at the C-terminus. The tissue distribution of this enzyme by RT (reverse transcription)-PCR analysis revealed high expression in the liver, small intestine, pancreas, testis and peripheral blood CD14+ and CD8+ cells. To investigate the role of Serase-1B, a full-length form recombinant protein was produced. Interestingly, recombinant Serase-1B was partly secreted as a soluble inactive precursor and it was also activated by trypsin. This activated enzyme selectively cleaved synthetic peptides for trypsin and activated protein C, and it was inhibited by several natural serine protease inhibitors, such as aprotinin, alpha2-antiplasmin and plasminogen activator inhibitor 1. In addition, Serase-1B efficiently converted pro-uPA (urokinase-type plasminogen activator) into active uPA and this activation was strongly inhibited by these natural inhibitors. Furthermore, this activation was also negatively regulated by glycosaminoglycans. Our results indicate that Serase-1B is a novel member of TTSPs that might be involved in uPA/plasmin-mediated proteolysis and possibly implicated in biological events such as fibrinolysis and tumour progression. PMID:16872279

Okumura, Yuushi; Hayama, Masaki; Takahashi, Etsuhisa; Fujiuchi, Mieko; Shimabukuro, Aki; Yano, Mihiro; Kido, Hiroshi

2006-12-15

264

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)

265

Relationship between plasminogen activator inhibitor-1 gene polymorphism and early local hemostatic activation in patients with percutaneous coronary intervention procedure  

International Nuclear Information System (INIS)

Objective: To investigate the relationship between plasminogen activatorinhibitor-1 (PAI-1) 4 G/5 G gene polymorphism and local homeostatic activation of PAI-1, D-dimers (DD), activated factor VII (F VII Ia) and P-Selectin (CD62P), on patients under percutaneous coronary intervention (PCI)procedures, and to evaluate its prognostic value on acute stent thrombosis by gene polymorphism analysis. Methods: 20 stable angina patients with a 70% diameter stenosis by visual estimation during angiography and a clinical indication for revascularization were selected. Lesions were treated with the use of standard interventional techniques, both stents implantation underwent with adjunctive balloon angioplasty. Simultaneous blood samples were drawn in sequence from the ostium of the coronary artery before balloon angioplasty through guiding catheter, from the distal coronary artery just beyond the dilated segment after balloon angioplasty and after stent implantation, through aspiration catheter. Markers of PAI-1 and CD62P were measured by ELISA. Markers of F VII and DD were measured by technique chronometrique and ELISA VIDAS respectively. Prevalence of the 4 G/5 G polymorphism was investigated using DNA analysis. Results: The distribution of PAI-1 genotypes in French people was as follows: 4 G/4 G in 30.0%, 4 G/5 G in 60.0% and 5 G/5 G in 10.0%. Among the patients, the frequency of the 4 G and 5 G allele were 0.60 and 0.40 respectively. In patients with the 4 G/5 G polymorphism In patients with the 4 G/5 G polymorphism of PAI-1 gene, the activities of the PAI-1, DD and F VIIa in the coronary circulation were significantly increased after balloon angioplasty in comparing with those before balloon angioplasty (P=0.01, respectively). However, there were no significant differences between the levels of hemostatic activation at ostium before balloon angioplasty and distal to lesion after stent implantation in patients with the 4 G/5 G genotype. Conclusions: Balloon angioplasty more easily induces vessel shrinkage and arterial wall injury and transient local haemostatic activation in comparing with stent implantation. This reponse would be more obvious in patients with 4 G/5 G polymorphism of the PAI-1 gene. Pretreatment with double chain antiplatelets might effectively control the early local activation of platelet in patients undergoing PCI procedure. (authors)

266

Prognostic importance of the soluble plasminogen activator receptor, suPAR, in plasma from rectal cancer patients.  

Science.gov (United States)

Colorectal cancer is one of the most common tumour types with approximately one third of the tumours located within the rectum. Rectal cancer differs somewhat from colon cancer, e.g. regarding the method of operation and the use of preoperative radiotherapy due to a tendency for local tumour recurrence. Proteolytic enzymes have been identified as key molecules in tumour invasion and metastasis, and factors within the urokinase-plasminogen activation (uPA) system have been associated with prognosis in several tumour types, including colorectal cancer. Recently, methods have been developed to analyse the soluble fraction of the plasminogen activator receptor (suPAR) in blood samples. An association between elevated suPAR levels and poor prognosis has recently been demonstrated in colorectal cancer. We have measured suPAR levels in pretreatment plasma samples from 173 rectal cancer patients in order to confirm its prognostic strength in this clinical entity. suPAR levels were determined in ethylenediamine tetraacetic acid (EDTA) plasma by a kinetic enzyme-linked immunosorbent assay (ELISA) and analysed with respect to sex, age, Dukes' stage, tumour differentiation grade and survival. In a univariate analysis, continuous suPAR plasma levels were associated with survival (PsuPAR values. Patients with suPAR values within the upper quartile had significantly shorter survival (hazard ratio (HR) 2.2, 95% confidence interval (CI) 1.3-43.7, P=0.002). In a multivariate Cox analysis, increasing suPAR values predicted shorter survival independent from Dukes' stage and tumour differentiation grade with an adjusted HR of 2.2 per ng/ml suPAR (95% CI 1.2-4.0, P=0.01). This study thus confirms that measurement of suPAR in preoperative plasma samples gives independent prognostic information in rectal cancer patients, higher values being associated with shorter survival. PMID:11267858

Fernebro, E; Madsen, R R; Fernö, M; Brünner, N; Bendahl, P; Christensen, I J; Johnson, A; Nilbert, M

2001-03-01

267

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

268

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

269

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

270

Tissue plasminogen activator contributes to morphine tolerance and induces mechanical allodynia via astrocytic IL-1? and ERK signaling in the spinal cord of mice  

OpenAIRE

Accumulating evidence indicates that activation of spinal cord astrocytes contributes importantly to nerve injury and inflammation-induced persistent pain and chronic opioid-induced antinociceptive tolerance. Phosphorylation of extracellular signal-regulated kinase (pERK) and induction of interleukin-1 beta (IL-1?) in spinal astrocytes have been implicated in astrocytes-mediated pain. Tissue plasminogen activator (tPA) is a serine protease that has been extensively used to treat stroke. We e...

Berta, Temugin; Liu, Yen-chin; Xu, Zhen-zhong; Ji, Ru-rong

2013-01-01

271

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 aepatocytes, 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

272

Caveolin-1 mediates tissue plasminogen activator-induced MMP-9 up-regulation in cultured brain microvascular endothelial cells.  

Science.gov (United States)

Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates blood-brain barrier injury and increases the risk of symptomatic cerebral hemorrhage. The mechanism through which tPA enhances MMP-9 activity is not well understood. Here we report an important role of caveolin-1 in mediating tPA-induced MMP-9 synthesis. Brain microvascular endothelial cell line bEnd3 cells were incubated with 5 or 20 ?g/ml tPA for 24 hrs before analyzing MMP-9 levels in the conditioned media and cellular extracts by gelatin zymography. tPA at a dose of 20 ?g/mL tPA, but not 5 ?g/mL, significantly increased MMP-9 level in cultured media while decreasing it in cellular extracts. Concurrently, tPA treatment induced a 2.3-fold increase of caveolin-1 protein levels in endothelial cells. Interestingly, knockdown of Cav-1 with siRNA inhibited tPA-induced MMP-9 mRNA up-regulation and MMP-9 increase in the conditioned media, but did not affect MMP-9 decrease in cellular extracts. These results suggest that caveolin-1 critically contributes to tPA-mediated MMP-9 up-regulation, but may not facilitate MMP-9 secretion in endothelial cells. Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates ischemic blood brain barrier (BBB) injury and increases the risk of symptomatic cerebral hemorrhage. Our results suggest a novel mechanism underlying this tPA-MMP 9 axis. In response to tPA treatment, caveolin-1 protein levels increased in endothelial cells, which mediate MMP-9 mRNA up-regulation and its secretion into extracellular space. Caveolin-1 may, however, not facilitate MMP-9 secretion in endothelial cells. Our data suggest caveolin-1 as a novel therapeutic target for protecting the BBB against ischemic damage. The schematic outlines tPA-induced MMP-9 upreguation. PMID:25683686

Jin, Xinchun; Sun, Yanyun; Xu, Ji; Liu, Wenlan

2015-03-01

273

Inhibition of urokinase plasminogen activator “uPA” activity alters ethanol consumption and conditioned place preference in mice  

Directory of Open Access Journals (Sweden)

Full Text Available Elyazia Al Maamari,* Mouza Al Ameri, Shamma Al Mansouri, Amine Bahi*Department of Anatomy, College of Medicine and Health Sciences, United Arab Emirates University, Al Ain, United Arab Emirates*These authors contributed equally to this workAbstract: Urokinase plasminogen activator, uPA, is a serine protease implicated in addiction to drugs of abuse. Using its specific inhibitor, B428, we and others have characterized the role of uPA in the rewarding properties of psychostimulants, including cocaine and amphetamine, but none have examined the role of uPA in ethanol use disorders. Therefore, in the current study, we extended our observations to the role of uPA in ethanol consumption and ethanol-induced conditioned place preference. The general aim of the present series of experiments was to investigate the effects of the administration of the B428 on voluntary alcohol intake and ethanol conditioned reward. A two-bottle choice, unlimited-access paradigm was used to compare ethanol intake between vehicle- and 3, 10, and 30 mg/kg B428-administered mice. For this purpose, the mice were presented with an ethanol solution (2.5%–20% and water, at each concentration for 4 days, and their consumption was measured daily. Consumption of saccharin and quinine solutions was also measured. Systemic administration of B428 dose-dependently decreased ethanol intake and preference. Additionally, B428 mice did not differ from vehicle mice in their intake of graded solutions of tastants, suggesting that the uPA inhibition did not alter taste function. Also, ethanol metabolism was not affected following B428 injection. More importantly, 1.5 g/kg ethanol-induced conditioned place preference acquisition was blocked following B428 administration. Taken together, our results are the first to implicate uPA inhibition in the regulation of ethanol consumption and preference, and suggest that uPA may be considered as a possible therapeutic drug target for alcoholism and abstinence.Keywords: B428, CPP, two-bottle choice

Al Maamari E

2014-09-01

274

The association between the 4G/5G polymorphism in the promoter of the plasminogen activator inhibitor-1 gene and extension of postsurgical calf vein thrombosis.  

Science.gov (United States)

The objective of this study was to evaluate whether the presence of a plasminogen activator inhibitor type 1 (PAI-1) promoter polymorphism 4G/5G could significantly influence the proximal extension of vein thrombosis in spite of anticoagulant treatment in patients with calf vein thrombosis (CVT) following orthopaedic, urological and abdominal surgery. We studied 168 patients with CVT, who had undergone orthopaedic, urological and abdominal surgery, subdivided as follows: first, 50 patients with thrombosis progression; second, 118 patients without thrombosis progression. The 4G/5G polymorphism of the plasminogen activator inhibitor 1 was evaluated in all patients and in 70 healthy matched controls. We also studied PAI-1 activity in plasma. The presence of 4G/5G genotype was significantly increased in the group of patients with the extension of thrombotic lesions and was associated with an increase in CVT extension risk (odds ratio adjusted for sex 2.692; 95% confidence interval 1.302-4.702). Moreover, we observed a significant increase of PAI-1 plasma activity in patients with extension of thrombotic lesion vs. patients without extension (P=0.0001). Patients with 4G/5G genotype in the promoter of the plasminogen activator inhibitor - 1 gene present a higher risk of extension of thrombotic lesions. PMID:23222167

Ferrara, Filippo; Meli, Francesco; Raimondi, Francesco; Montalto, Salvatore; Cospite, Valentina; Novo, Giuseppina; Novo, Salvatore

2013-04-01

275

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

Science.gov (United States)

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/inflammatory responses. PAI-1, a key regulator for fibrinolysis and coagulation, acts as an APP during acute phase response (APR) such as acute lung injury (ALI), inflammation, and sepsis. However, the role of PAI-1 in the pathogenesis of these diseases still remains unclear, especially in bacterial pneumonia. In this study, we showed that PAI-1 expression is upregulated following nontypeable Haemophilus influenzae (NTHi) infection. PAI-1 knockout (KO) mice failed to generate early immune responses against NTHi. Failure of generating early immune responses in PAI-1 KO mice resulted in reduced bacterial clearance and prolonged disease process, which in turn led to enhanced inflammation at late stage of infection. Moreover, we also found that NTHi induces PAI-1 via activation of TLR2-MyD88-MKK3-p38 MAPK signaling pathway. These data suggest that PAI-1 plays critical role in earl host defense response against NTHi infection. Our study thus reveals a novel role of PAI-1 in infection caused by NTHi, one of the most common gram-negative bacterial pathogens in respiratory systems. PMID:21945446

Lim, Jae Hyang; Woo, Chang-Hoon; Li, Jian-Dong

2011-10-14

276

Interrelations between blood-brain barrier permeability and matrix metalloproteinases are differently affected by tissue plasminogen activator and hyperoxia in a rat model of embolic stroke  

OpenAIRE

Abstract Background In ischemic stroke, blood-brain barrier (BBB) regulations, typically involving matrix metalloproteinases (MMPs) and inhibitors (TIMPs) as mediators, became interesting since tissue plasminogen activator (tPA)-related BBB breakdown with risk of secondary hemorrhage was considered to involve these mediators too. Despite high clinical relevance, detailed interactions are purely understood. After a pilot study addressing hyperoxia as potential neuroprotective co-treatment to t...

Michalski Dominik; Hobohm Carsten; Weise Christopher; Pelz Johann; Heindl Marita; Kamprad Manja; Kacza Johannes; Härtig Wolfgang

2012-01-01

277

Expression of Urokinase Plasminogen Activator Receptor on Monocytes from Patients with Relapsing-Remitting Multiple Sclerosis: Effect of Glatiramer Acetate (Copolymer 1)  

OpenAIRE

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system in which peripheral blood monocytes play an important role. We have previously reported that patients with chronic progressive MS (CPMS) have significantly increased numbers of circulating monocytes which express the urokinase plasminogen activator receptor (uPAR). In the present study, we examined the expression of uPAR on monocytes in patients with relapsing-remitting multiple sclerosis (RRMS) not curren...

Balabanov, Roumen; Lisak, Deena; Beaumont, Thomas; Lisak, Robert P.; Dore-duffy, Paula

2001-01-01

278

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

OpenAIRE

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

279

Detection of mRNAs for urokinase-type plasminogen activator, its receptor, and type 1 inhibitor in giant cell tumors of bone with in situ hybridization.  

OpenAIRE

Although giant cell tumor of bone (GCT) is generally considered to be an uncommon benign neoplasm, it can pursue an aggressive course with local recurrence and metastasis. Attempts to predict the biological behavior of GCT with histopathological parameters, however, have not been successful. The urokinase-type plasminogen activation system has been implicated in tumor invasion and metastasis and abnormalities of the components of this system have been found in several malignancies. In this st...

Zheng, M. H.; Fan, Y.; Panicker, A.; Smith, A.; Robertson, T.; Wysocki, S.; Robbins, P.; Papadimitriou, J. M.; Wood, D. J.

1995-01-01

280

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

281

Reactive protein, plasminogen activator inhibitor type-1 (PAI-1) levels, PAI-1 promoter 4G/5G polymorphism and acute myocardial infarction  

OpenAIRE

Objective To investigate the relationship between CRP, plasminogen activator inhibitor type 1 (PAI-1) levels, PAI-1 gene promoter 4G/5G polymorphism and the type of acute myocardial infarction (ST elevation myocardial infarction, STEMI vs. the non-ST elevation Myocardial infarction, NSTEMI). Methods One hundred seventy-six consecutive patients with AMI were included for the study, of whom 60 had STEMI and 56 had NSTEMI, and 60 adults without cardiovascular and cerebrovascular disease were s...

Xue-Lei Cao; Chang-Yong Zhou; Lei Yin; Shao-Chun Wang; Xiu-Ling Jia; Huan Huang; Xiao-Hong Sun

2010-01-01

282

Neuroproteome Changes after Ischemia/Reperfusion Injury and Tissue Plasminogen Activator Administration in Rats: A Quantitative iTRAQ Proteomics Study  

OpenAIRE

The thrombolytic, recombinant tissue plasminogen activator (rt-PA) is the only approved therapy for acute ischemic stroke (AIS). When administered after AIS, rt-PA has many adverse pleiotropic actions, which are currently poorly understood. The identification of proteins showing differential expression after rt-PA administration may provide insight into these pleiotropic actions. In this study we used a 2D-LC MS/MS iTRAQ proteomic analysis, western blotting, and pathway analysis to analyze ch...

Merali, Zamir; Gao, Meah Mingyang; Bowes, Tim; Chen, Jian; Evans, Kenneth; Kassner, Andrea

2014-01-01

283

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

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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 (pplasma 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; pplasma 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

284

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

285

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

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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

286

Effects of antiinflammatory agents on mouse skin tumor promotion, epidermal DNA synthesis, phorbol ester-induced cellular proliferation, and production of plasminogen activator. [Fluocinolone acetonide, fluocinonide, fluclorolone acetonide  

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The antiinflammatory steroids fluocinolone acetonide, fluocinonide, and fluclorolone acetonide were found to be very effective inhibitory agents of mouse skin tumor promotion. These steroids also drastically inhibited epidermal DNA synthesis and epidermal cellular proliferation induced by a phorbal ester tumor promoter. In addition, these compounds were potent inhibitors of plasminogen activator production in tumor cell cultures. The clinically used nonsteroidal antiinflammatory agents oxyphenbutazone, indomethacin, and Seclazone also inhibited tumor promotion but were much less effective. Although these agents are useful against inflammatory disorders in general when given p.o., in our studies they had little effect on inflammation and epidermal cellular proliferation induced by a phorbol ester tumor promoter when given topically. The aforementioned nonsteroidal antiinflammatory agents also had little effect on epidermal DNA synthesis. Oxyphenbutazone and indomethacin were less potent inhibitors of plasminogen activator production in tumor cells than were the antiinflammatory steroids, and Seclazone produced a negligible inhibition. There is, therefore, a general correlation in the potencies of a series of steroidal antiinflammatory agents for inhibition of tumor promotion and their ability to inhibit plasminogen activator production by tumor cell cultures and epidermal DNA synthesis.

Viaje, A.; Slaga, T.J.; Wigler, M.; Weinstein, I.B.

1977-05-01

287

The Soluble Plasminogen Activator Receptor as a Biomarker on Monitoring the Therapy Progress of Pulmonary TB-AFB(+) Patients.  

Science.gov (United States)

The role of soluble soluble urokinase-type plasminogen activator receptor (suPAR) as a biological marker for TB treatment efficacy on active pulmonary TB-AFB(+) patients was investigated. Twenty pulmonary TB-AFB(+) patients participated in a cohort study for six months. The plasma suPAR level was measured using ELISA method before treatment, two months, four months and six months after treatment. At the same time clinical parameters were also measured. Results indicated that all patients (n = 20) showed highest plasma suPAR levels before treatment (median 12.775?ng/mL) and significantly decreased ( P = .0001level (median 4.772?ng/mL). Interestingly, the significant reduced of suPAR level was parallel to treatment efficacy and correlated with other clinical and laboratory parameters, that is, decreasing of patients' complaints, increasing of BMI (r = -0.281), thoracic imaging improvement, sputum conversion, decreasing of ESR (r = 0.577) and monocytes count (r = 0.536) with exception the width of lesion in thoracic imaging. In conclusion, the suPAR level in could reflect the progress of TB therapy. PMID:22567258

Mardining Raras, Tri Yudani; Noor Chozin, Iin

2010-01-01

288

Risk factors associated with serum levels of the inflammatory biomarker soluble urokinase plasminogen activator receptor in a general population.  

Science.gov (United States)

The soluble urokinase plasminogen activator receptor (suPAR) is a biomarker of mortality risk in various patient populations. However, little is known about the implications of lifestyle for suPAR levels in the general population. Lifestyle, demographic, and cardiovascular disease (CVD) risk factor data were collected from 5,538 participants in the Danish population-based Inter99 study. Their suPAR levels were measured using a sandwich enzyme-linked immunosorbent assay. In the final adjusted model, smoking and morbid obesity were strongly associated with higher suPAR levels (P unhealthy diet and alcohol abstinence in men were also associated with higher suPAR levels. Physical activity in leisure time had a modest impact on suPAR levels in univariate analysis, but not in the final adjusted model. In conclusion, smoking and morbid obesity were strongly associated with higher serum suPAR levels in this general population. Diet and alcohol consumption also seemed to impact suPAR levels. Lifestyle changes are likely to affect suPAR since ex-smokers had suPAR levels comparable to those of never-smokers. PMID:25574132

Haupt, Thomas H; Kallemose, Thomas; Ladelund, Steen; Rasmussen, Line Jh; Thorball, Christian W; Andersen, Ove; Pisinger, Charlotta; Eugen-Olsen, Jesper

2014-01-01

289

Genetic association of five plasminogen activator inhibitor-1 (PAI-1) polymorphisms and idiopathic recurrent pregnancy loss in Korean women.  

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Plasminogen activator inhibitor-1 (PAI-1) is important for maintaining pregnancy. Aberrantly increased PAI-1 levels may contribute to thrombosis and inflammation, leading to pregnancy loss. This study investigated the association of PAI-1 polymorphisms (PAI-1 rs2227631 [-844G>A], rs1799889 [-675 4G/5G], rs6092 [43G>A], rs2227694 [9785G>A], and rs7242 [11053T>G]) with idiopathic recurrent pregnancy loss (RPL) in Korean women. We screened 308 RPL patients and 227 control participants for five PAI-1 polymorphisms. Genotyping of PAI-1 was performed by polymerase chain reaction-restriction fragment length polymorphism assay. PAI-1 4G4G and -844AA/4G4G/11053GG genotypes were associated with RPL. PAI-1 -844A/4G/43G/9785G/11053G haplotype was connected to hypofibrinolytic status (i.e. increased levels of plasma PAI-1, increased numbers of platelets, reduced prothrombin time, and reduced activated partial thromboplastin time). Moreover, PAI-1 11053TG+GG frequency was positively related to plasma homocysteine and urate levels, whereas -844AA frequency was associated with plasma folate concentrations according to ordinal logistic regression analysis. Based on these results, we propose that PAI-1 -844G>A, 4G/5G, and 11053T>G polymorphisms are markers of RPL. PMID:23903286

Jeon, Young Joo; Kim, Young Ran; Lee, Bo Eun; Choi, Yi Seul; Kim, Ji Hyang; Shin, Ji Eun; Rah, HyungChul; Cha, Sun Hee; Lee, Woo Sik; Kim, Nam Keun

2013-10-01

290

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

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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

291

Association between Plasminogen Activator Inhibitor-1 Gene Polymorphisms and Recurrent Pregnancy Loss: A Systematic Review and Meta-Analysis.  

Science.gov (United States)

Human plasminogen activator inhibitor-1 (PAI-1) is closely related to embryonic development and pregnancy success. The association between PAI-1 gene polymorphisms (PAI-1-844G/A and PAI-1-675G/A) and the risk of recurrent pregnancy loss (RPL) is controversial. Therefore, we perform this review to clarify the association between PAI-1 gene polymorphisms and RPL risk. We performed a systematic search for studies that described the effect of PAI-1 polymorphisms on RPL risk. The odds ratios (ORs) with corresponding 95% confidence intervals (CIs) were considered under recessive genetic models. Furthermore, we conducted a subgroup analysis based on the studies' geographic regions of origin. Data were analyzed using Stata 11.2 software. Eighteen studies were included, and a high degree of statistical heterogeneity existed among the studies. In this study, we found a significant association between the PAI-1-675G/A polymorphism and the risk of RPL under the recessive model (OR = 1.70, 95% CI = 1.21-2.38). However, no significant association between the PAI-1-844G/A polymorphism and RPL was noted. PAI-1-675G/A (4G/5G) polymorphisms play a potential role in RPL. The screening of PAI-1 (4G/5G) gene mutations should be included during an RPL diagnostic workup, and patients should be treated using anticoagulant therapy during pregnancy if necessary. PMID:25250948

Chen, Hui; Nie, Shuping; Lu, Ming

2015-04-01

292

Association between the plasminogen activator inhibitor-1 4G/5G polymorphism and venous thrombosis. A meta-analysis.  

Science.gov (United States)

The effect of the 675 insertion/deletion (4G/5G) polymorphism of plasminogen activator inhibitor-1 (PAI-1) gene on the risk of venous thromboembolism (VTE) remains controversial. In this study, we performed a meta-analysis of published data regarding this issue. A comprehensive electronic search was carried out up until September 2006. A total of 22 articles were included in the analysis that was performed using random effects models. Eighteen papers, concerning patients without another known risk factor, comprised 2,644 cases and 3,739 controls. The alleles contrast (4G vs. 5G allele) yielded a statistically significant odds ratio (OR) of 1.153 (95% confidence interval [CI]: 1.068-1.246). In a sub-analysis of five studies that included 256 cases with another genetic risk factor and 147 controls, the combined per-allele OR was still significant (OR: 1.833,95% CI: 1.325-2.536). On the contrary, the analysis of five studies regarding cases with a non-genetic risk factor for VTE (antiphospholipid antibody syndrome, Behcet disease) provided insignificant results in all aspects. There was no evidence for heterogeneity and publication bias in all analyses. Based on our findings, the 4G allele appears to increase the risk of venous thrombosis, particularly in subjects with other genetic thrombophilic defects. Recommendation for detection of this polymorphism in evaluating thrombophilia in such patients might be considered. PMID:17549286

Tsantes, Argirios E; Nikolopoulos, Georgios K; Bagos, Pantelis G; Rapti, Evdoxia; Mantzios, Georgios; Kapsimali, Violeta; Travlou, Anthi

2007-06-01

293

A flexible multidomain structure drives the function of the urokinase-type plasminogen activator receptor (uPAR)  

DEFF Research Database (Denmark)

The urokinase-type plasminogen activator receptor (uPAR) provides a rendezvous between proteolytic degradation of the extracellular matrix and integrin mediated adhesion to vitronectin. These processes are however tightly linked as the high-affinity binding of urokinase regulates the binding of uPAR to matrix-embedded vitronectin. Although crystal structures exist to define the corresponding static bi- and trimolecular receptor complexes it is evident that the dynamic property of uPAR plays a decisive role for its function. In the present study, we combine small angle X-ray scattering, hydrogen-deuterium exchange, and surface plasmon resonance to develop a structural model describing the allosteric regulation of uPAR. We show that the flexibility of its N-terminal domain provides the key for understanding this allosteric mechanism. Importantly, our model has direct implications for understanding uPAR-assisted cell adhesion and migration as well as for translational research including targeted intervention therapy and non-invasive tumor imaging in vivo.

Mertens, Haydyn D.T.; Kjærgaard, Magnus

2012-01-01

294

Effect of dipeptidyl peptidase-4 inhibitor, vildagliptin on plasminogen activator inhibitor-1 in patients with diabetes mellitus.  

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Dipeptidyl peptidase-4 (DPP-4) inhibitors may affect the serum levels of plasminogen activator inhibitor-1 (PAI-1) associated with triglyceride (TG) metabolism, which is a prognostic factor for cardiovascular disease, in diabetic patients. We conducted an 8-week, prospective, randomized study in which we assigned type 2 diabetic patients who were inadequately controlled with antidiabetic therapy to the vildagliptin group (50 mg bid, n = 49) or the control group (n = 49). The primary efficacy parameter was the change in the serum level of PAI-1, and the secondary end point was the change in the serum levels of TG-rich lipoproteins. In the vildagliptin group, significant decrease of the serum PAI-1 level by 16.3% (p <0.0001) and significant decreases of the serum TG, remnant-like particle cholesterol, and apolipoprotein B levels by 12.1% (p = 0.002), 13.9% (p = 0.003), and 9.5% (p <0.0001), respectively, were observed. No such changes were observed in the control group. Multivariate regression analyses identified the absolute change from the baseline (?) of the PAI-1, but not that of the fasting blood glucose or hemoglobin A1c, as independent predictors of the ?TG, ? remnant-like particle cholesterol, and ? apolipoprotein B. In conclusion, treatment of type 2 diabetes with vildagliptin might prevent the progression of atherosclerotic cardiovascular disease in diabetic patients by decreasing the serum PAI-1 levels and improving TG metabolism. PMID:25637323

Tani, Shigemasa; Takahashi, Atsuhiko; Nagao, Ken; Hirayama, Atsushi

2015-02-15

295

Meta-Analysis of the Association between Plasminogen Activator Inhibitor-1 4G/5G Polymorphism and Recurrent Pregnancy Loss.  

Science.gov (United States)

Background The association between plasminogen activator inhibitor-1 (PAI-1) 4G/5G polymorphism and recurrent pregnancy loss (RPL) risk is still contradictory. We thus performed a meta-analysis. Material and Methods Relevant studies were searched for in PubMed, Web of Science, Embase, and Cochrane Library. An odds ratio (OR) with a 95% confidence interval (CI) was used to assess the association between PAI-1 4G/5G polymorphism and RPL risk. Results A total of 22 studies with 4306 cases and 3076 controls were included in this meta-analysis. We found that PAI-1 4G/5G polymorphism was significantly associated with an increased RPL risk (OR=1.89; 95% CI 1.34-2.67; P=0.0003). In the subgroup analysis by race, PAI-1 4G/5G polymorphism was significantly associated with an increased RPL risk in Caucasians (OR=2.23; 95% CI 1.44-3.46; P=0.0003). However, no significant association was observed in Asians (OR=1.47; 95% CI 0.84-2.59; P=0.18). Conclusions In conclusion, this meta-analysis suggests that PAI-1 4G/5G polymorphism might be associated with RPL development in Caucasians. PMID:25862335

Li, Xuejiao; Liu, Yukun; Zhang, Rui; Tan, Jianping; Chen, Libin; Liu, Yinglin

2015-01-01

296

D-dimer, plasminogen activator inhibitor-1, prothrombin fragments and protein C - role in prothrombotic state of colorectal cancer.  

Science.gov (United States)

The relationship between malignant tumors and blood coagulation disorders is generally well known. The authors studied blood coagulation in patients with colorectal cancer and evaluated some prothrombotic markers. The authors analyzed by latex-aglutination method, ProC Global test, Asserachrom PAI-1 test and Enzygnost F 1+2 test the group of 137 patients with malignant tumor of colon and rectum, drew attention to the relationship between level of D-dimer, PAI-1, F 1+2, Protein C and the progress of malignant tumor, its localization, clinical stage, histopathology type, method of surgery considering the stapling use. Very aggressive and advanced tumors have significantly higher level of D-dimer, plasminogen activator inhibitor I (PAI-1). Prothrombotic fragments 1+2 were significantly higher by anastomosis dehiscence. Protein C level was lower in the age from sixty to seventy and in advanced clinical stage. Pre-operative surveys of D-dimer, PAI-1, prothrombotic fragments and Protein C give informations about risk of thrombosis of malignant diseases, their clinical stage and histological type. D-dimer and PAI-1 have a real clinical value and can be reliable prothrombotic marker. PMID:21391740

Mytnik, M; Stasko, J

2011-01-01

297

Recombinant T cell receptor ligand treatment improves neurological outcome in the presence of tissue plasminogen activator in experimental ischemic stroke.  

Science.gov (United States)

RTL1000 is a partial human MHC molecule coupled to a human myelin peptide. We previously demonstrated that RTL1000 was protective against experimental ischemic stroke in HLA-DR2 transgenic (DR2-Tg) mice. Since thrombolysis with recombinant tissue plasminogen activator (t-PA) is a standard therapy for stroke, we determined if RTL1000 efficacy is altered when combined with t-PA in experimental stroke. Male DR2-Tg mice underwent 60 min of intraluminal middle cerebral artery occlusion (MCAO). t-PA or vehicle was infused intravenously followed by either a single or four daily subcutaneous injections of RTL1000 or vehicle. Infarct size was measured by 2, 3, 5-triphenyltetrazolium chloride staining at 24 or 96 h of reperfusion. Our data showed that t-PA alone reduced infarct size when measured at 24 h but not at 96 h after MCAO. RTL1000 alone reduced infarct size both at 24 and 96 h after MCAO. Combining RTL1000 with t-PA did not alter its ability to reduce infarct size at either 24 or 96 h after MCAO and provides additional protection in t-PA treated mice at 24 h after ischemic stroke. Taken together, RTL1000 treatment alone improves outcome and provides additional protection in t-PA-treated mice in experimental ischemic stroke. PMID:24953050

Zhu, Wenbin; Libal, Nicole L; Casper, Amanda; Bodhankar, Sheetal; Offner, Halina; Alkayed, Nabil J

2014-10-01

298

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

299

Estriol-induced fibrinolysis due to the activation of plasminogen to plasmin by nitric oxide synthesis in platelets.  

Science.gov (United States)

Estriol, an oestrogen, at 0.6?nmol/l was reported to inhibit ADP-induced platelet aggregation through nitric oxide synthesis. As nitric oxide has been reported to cause fibrinolysis due to the activation of plasminogen to plasmin, the role of estriol as a fibrinolytic agent was investigated. Also, the mechanism of estriol-induced nitric oxide synthesis in anucleated platelets was investigated. The estriol-induced lysis of platelet-rich plasma (PRP) clot was determined by photography of the clot lysis and by the assay of fibrin degradation products in the lysate and was obtained by SDS-PAGE. Nitric oxide was determined by methemoglobin method. The platelet membrane protein was isolated from the platelets by using Triton X-100 (0.05% v/v). The binding of estriol to the protein was determined by Scatchard plot by using an ELISA for estriol. Estriol at 0.6?nmol/l was found to lyse the clotted PRP due to fibrinolysis that produced fibrin degradation products in the lysate. The amino acid analysis of the platelet membrane protein, which resembles with nitric oxide synthase (NOS) activity, was activated nearly 10-fold over the control in the presence of estriol and was identified to be a human serum albumin precursor (Mr. 69?kDa) that binds to estriol with Kd1 of 6.0?×?10?mol/l and 39?±?2 molecules of estriol bound the NOS molecule. The estriol-induced nitric oxide is capable of inducing fibrinolysis of the clotted PRP. The binding of estriol to platelet membrane NOS activated the enzyme in the absence of DNA in the platelet. PMID:24695088

Jana, Pradipta; Maiti, Smarajit; Kahn, Nighat N; Sinha, Asru K

2015-04-01

300

Reducing hemorrhagic complication by dabigatran via neurovascular protection after recanalization with tissue plasminogen activator in ischemic stroke of rat.  

Science.gov (United States)

This study assesses the risks and benefits of tissue plasminogen activator (tPA) treatment under oral anticoagulation with dabigatran compared with warfarin or vehicle control in transient middle cerebral artery occlusion (tMCAO). After pretreatment with warfarin (0.2 mg/kg/day), dabigatran (20 mg/kg/day), or vehicle (0.5% carboxymethyl cellulose sodium salt) for 7 days, tMCAO was induced for 120 min, followed by reperfusion and tPA (10 mg/kg/10 ml). Clinical parameters, including cerebral infarction volume, hemorrhagic volume, and blood coagulation, were examined. At 24 hr after reperfusion, markers for the neurovascular unit at the peri-ischemic lesion were immunohistochemically examined in brain sections, and MMP-9 activity was measured by zymography. Paraparesis and intracerebral hemorrhage volume were significantly improved in the dabigatran-pretreated group compared with the warfarin-pretreated group. A marked dissociation between astrocyte foot processes and the basal lamina or pericyte was observed in the warfarin-pretreated group, which was greatly improved in the dabigatran-pretreated group. Furthermore, a remarkable activation of MMP-9 in the ipsilateral warfarin-pretreated rat brain was greatly reduced in dabigatran-pretreated rats. The present study reveals that the mechanism of intracerebral hemorrhage with warfarin-pretreatment plus tPA in ischemic stroke rats is the dissociation of the neurovascular unit, including the pericyte. Neurovascular protection by dabigatran, which was first shown in this study, could partially explain the reduction in hemorrhagic complication by dabigatran reported from clinical study. PMID:24265137

Kono, Syoichiro; Deguchi, Kentaro; Omote, Yoshio; Yunoki, Taijun; Yamashita, Toru; Kurata, Tomoko; Ikeda, Yoshio; Abe, Koji

2014-01-01

301

A plasminogen activator inhibitor type 1 mutant retards diabetic nephropathy in db/db mice by protecting podocytes.  

Science.gov (United States)

A mutant non-inhibiting plasminogen activator inhibitor type 1 (PAI-1), termed PAI-1R, which reduces endogenous PAI-1 activity, has been shown to inhibit albuminuria and reduce glomerulosclerosis in experimental diabetes. The mechanism of the reduction of albuminuria is unclear. This study sought to determine whether the administration of PAI-1R protected podocytes from injury directly, thereby reducing albuminuria in the db/db mouse, a model of type 2 diabetes. Untreated uninephrectomized db/db mice developed significant mesangial matrix expansion and albuminuria at week 22 of age, associated with segmental podocyte foot-process effacement, reduction of renal nephrin, podocin and zonula occludin-1 production and induction of renal desmin and B7-1 generation. In contrast, treatment with PAI-1R at 0.5 mg (kg body weight)(-1) i.p., twice daily from week 20 to 22, reduced glomerular matrix accumulation, fibronectin and collagen production and albuminuria by 36, 62, 65 and 31%, respectively (P < 0.05), without affecting blood glucose level or body weight. Podocyte morphology and protein markers were also significantly attenuated by PAI-1R administration. Importantly, recombinant PAI-1 downregulated nephrin and zonula occludin-1 but increased desmin and B7-1 mRNA expression and protein production by podocytes in vitro, similar to the effects of transforming growth factor-?1. These observations provide evidence that PAI-1, in a manner similar to transforming growth factor-?1, directly induces podocyte injury, particularly in the setting of diabetes, where elevated PAI-1 may contribute to the progression of albuminuria. Reducing the increased PAI-1 activity by administration of PAI-1R, in fact, reduces podocyte injury, thereby reducing albuminuria. Therefore, PAI-1R provides an additional therapeutic effect in slowing the progression of diabetic nephropathy via the protection of podocytes. PMID:24443353

Zhang, Jiandong; Gu, Chunyan; Lawrence, Daniel A; Cheung, Alfred K; Huang, Yufeng

2014-05-01

302

Urinary-type plasminogen activator receptor/alpha 3 beta 1 integrin signaling, altered gene expression, and oral tumor progression.  

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Oral squamous cell carcinoma (OSCC) has 50% 5-year survival rate, highlighting our limited understanding of the molecular events that contribute to disease progression. Microarray analyses of primary oral tumors have identified urinary-type plasminogen activator (uPA) and its receptor (uPAR) as key genes associated with human OSCC progression. The uPAR functions as both a proteinase receptor and an integrin ligand, modifying proteolysis, migration, integrin signaling, and cellular transcription. In the current study, uPAR expression levels were modified in OSCC cells followed by analysis of tumor growth in an in vivo orthotopic xenograft model and by transcriptional profiling. Overexpression of uPAR resulted in more infiltrative and less differentiated tumors, with ill-defined borders, cytologic atypia, and enhanced vascularity. Analysis of serial sections of both murine experimental tumors and microarrayed human OSCC showed a statistically significant association between uPAR and alpha(3) integrin colocalization in areas exhibiting extracellular signal-regulated kinase phosphorylation, suggesting that uPAR/alpha(3) integrin interaction potentiates extracellular signal-regulated kinase signaling in vivo. This is supported by cDNA microarray analysis, which showed differential expression of 148 genes (113 upregulated and 35 downregulated). Validation of gene expression changes in human OSCC using immunohistochemistry and quantitative real-time PCR showed increased growth factors, proteinases/inhibitors, and matrix components in uPAR-overexpressing tumors. Together, these results support a model wherein increased uPAR expression promotes alpha(3)beta(1) integrin association, resulting in increased mitogen-activated protein kinase signaling and transcriptional activation, leading to the formation of more aggressive tongue tumors. This combined approach has efficacy to identify additional biomarkers and/or prognostic indicators associated with aggressive human OSCC. PMID:20145038

Ghosh, Supurna; Koblinski, Jennifer; Johnson, Jeffrey; Liu, Yueying; Ericsson, Aaron; Davis, J Wade; Shi, Zonggao; Ravosa, Matthew J; Crawford, Susan; Frazier, Shellaine; Stack, M Sharon

2010-02-01

303

Relationship between Circulating Tumor Cells, Blood Coagulation, and Urokinase-Plasminogen-Activator System in Early Breast Cancer Patients.  

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Cancer is a risk factor for venous thromboembolism (VTE) and plasma d-dimer (DD) and tissue factor (TF) are established VTE associated markers. Circulating tumor cells (CTCs) are associated with the risk of VTE in metastatic breast cancer. This study aimed to correlate CTCs, blood coagulation and the urokinase plasminogen activator (uPA) system in primary breast cancer (PBC) patients. This prospective study included 116 PBC patients treated by primary surgery. CTCs were detected by quantitative RT-PCR assay for expression of epithelial (CK19) or epithelial-mesenchymal transition (EMT) genes (TWIST1, SNAIL1, SLUG, ZEB1, FOXC2). Plasma DD, TF, uPA system proteins were detected by enzyme-linked immunosorbent assays, while expressions of uPA system in surgical specimens were evaluated by immunohistochemistry. CTCs were detected in 27.6% patients. Patients with CTCs had a significantly higher mean plasma DD (ng/mL) than those of patients without CTCs (632.4 versus 365.4, p = 0.000004). There was no association between plasma TF and CTCs. Epithelial CTCs exhibit higher expression of uPA system genes compared to EMT_CTCs. Patients with CTCs had higher plasma uPA proteins than those of patients without CTCs; there was no correlation between tissue expression of uPA system, CTCs, DD or TF levels. In multivariate analysis CTCs and patients age were independent factors associated with plasma DD. We found association between plasma DD and CTCs indicating a potential role for activation of the coagulation cascade in the early metastatic process. CTCs could be directly involved in coagulation activation or increased CTCs could be marker of aggressive disease and increased VTE risk. PMID:25623304

Mego, Michal; Karaba, Marian; Minarik, Gabriel; Benca, Juraj; Sedlácková, Tatiana; Tothova, Lubomira; Vlkova, Barbora; Cierna, Zuzana; Janega, Pavol; Luha, Jan; Gronesova, Paulina; Pindak, Daniel; Fridrichova, Ivana; Celec, Peter; Reuben, James M; Cristofanilli, Massimo; Mardiak, Jozef

2015-03-01

304

Effects of Pharmacological Inhibition and Genetic Deficiency of Plasminogen Activator Inhibitor-1 in Radiation-Induced Intestinal Injury  

International Nuclear Information System (INIS)

Purpose: To investigate effects of plasminogen activator inhibitor 1 (PAI-1) genetic deficiency and pharmacological PAI-1 inhibition with PAI-039 in a mouse model of radiation-induced enteropathy. Methods and Materials: Wild-type (Wt) and PAI-1-/- knockout mice received a single dose of 19 Gy to an exteriorized localized intestinal segment. Sham and irradiated Wt mice were treated orally with 1 mg/g of PAI-039. Histological modifications were quantified using a radiation injury score. Moreover, intestinal gene expression was monitored by real-time PCR. Results: At 3 days after irradiation, PAI-039 abolished the radiation-induced increase in the plasma active form of PAI-1 and limited the radiation-induced gene expression of transforming growth factor ?1 (TGF-?1), CTGF, PAI-1, and COL1A2. Moreover, PAI-039 conferred temporary protection against early lethality. PAI-039 treatment limited the radiation-induced increase of CTGF and PAI-1 at 2 weeks after irradiation but had no effect at 6 weeks. Radiation injuries were less severe in PAI-1-/- mice than in Wt mice, and despite the beneficial effect, 3 days after irradiation, PAI-039 had no effects on microscopic radiation injuries compared to untreated Wt mice. Conclusions: A genetic deficiency of PAI-1 is associated with amelioration of late radiation enteropathy. Pharmacological inhibition of PAI-1 by PAI-039 positively impacts the early, acute phase increase in plasma PAI-1 and the associated rade in plasma PAI-1 and the associated radiation-induced gene expression of inflammatory/extracellular matrix proteins. Since PAI-039 has been shown to inhibit the active form of PAI-1, as opposed to the complete loss of PAI-1 in the knockout animals, these data suggest that a PAI-1 inhibitor could be beneficial in treating radiation-induced tissue injury in acute settings where PAI-1 is elevated.

305

Tissue plasminogen activator induces microglial inflammation via a noncatalytic molecular mechanism involving activation of mitogen-activated protein kinases and Akt signaling pathways and AnnexinA2 and Galectin-1 receptors  

OpenAIRE

Inflammatory responses mediated by glial cells play a critical role in many pathological situations related to neurodegeneration such as Alzheimer's disease. Tissue plasminogen activator (tPA) is a serine protease which best-known function is fibrinolysis, but it is also involved in many other physiological and pathological events as microglial activation. Here, we found that tPA is required for Aβ-mediated microglial inflammatory response and tumor necrosis factor-α release. We further...

Serratosa, Joan; Tusell, Josep Maria; Saura, Josep; Planas, Anna M.; Navarro, Pilar

2012-01-01

306

Antithrombotic activity of a monoclonal antibody inducing the substrate form of plasminogen activator inhibitor type 1 in rat models of venous and arterial thrombosis.  

Science.gov (United States)

1. Elevated plasminogen activator inhibitor 1 (PAI-1) is a risk factor for thrombosis, and inhibitors of the interaction between PAI-1 and tissue plasminogen activator (t-PA) have antithrombotic and prothrombolytic activity in animals. We describe the antithrombotic effects in the rat of a monoclonal antibody (MA33H1) which converts PAI-1 to a non-inhibitory substrate. 2. The activity of MA33H1 against rat PAI-1 was confirmed using two-chain t-PA and a chromogenic substrate. MA33H1 was evaluated in rat venous (thromboplastin + stasis in the abdominal vena cava) and arterial (electric current applied to a carotid artery) thrombosis models. The effects on tail-transection bleeding time were studied. 3. MA33H1 at 100 ng ml(-1) inhibited both human (44.1%) and rat PAI-1 (49.7%). This effect was concentration-dependent. Its effect on human PAI-1 was not significantly inhibited by 1 microg ml(-1) fibrin or a approximately 7 fold molar excess of vitronectin (1 nM). Inhibition of rat PAI-1 was unchanged by fibrin, but vitronectin reduced inhibition from 0.5 nM. 4. In the venous thrombosis model, MA33H1 significantly reduced thrombus weights by 38 and 58.6% at 50 and 100 microg kg(-1) min(-1) i.v. respectively. This effect was inhibited by tranexamic acid. In the arterial model, MA33H1 significantly increased the delay to occlusive thrombus formation by 58 and 142% at 50 and 100 microg kg(-1) min(-1) i.v., and did not affect bleeding time at 300 microg kg(-1) min(-1) i.v. 5. Thus, a monoclonal antibody which transforms PAI-1 to a t-PA substrate prevents thrombus formation in the rat with no effect on bleeding time at a higher dose. PMID:9776340

Berry, C N; Lunven, C; Lechaire, I; Girardot, C; O'Connor, S E

1998-09-01

307

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 P4G allele was also significantly higher in PNS patients compared to healthy controls (P4G/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

308

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

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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

309

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 togf 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

310

Interferon-tau activates multiple signal transducer and activator of transcription proteins and has complex effects on interferon-responsive gene transcription in ovine endometrial epithelial cells.  

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Interferon-tau (IFNtau), a type I IFN produced by sheep conceptus trophectoderm, is the signal for maternal recognition of pregnancy. Although it is clear that IFNtau suppresses transcription of the estrogen receptor alpha and oxytocin receptor genes and induces expression of various IFN-stimulated genes within the endometrial epithelium, little is known of the signal transduction pathway activated by the hormone. This study determined the effects of IFNtau on signal transducer and activator of transcription (STAT) activation, expression, DNA binding, and transcriptional activation using an ovine endometrial epithelial cell line. IFNtau induced persistent tyrosine phosphorylation and nuclear translocation of STAT1 and -2 (10 min to 48 h), but transient phosphorylation and nuclear translocation of STAT3, -5a/b, and -6 (10 to gene factor-3 and STAT1 homodimers formed and bound an IFN-stimulated response element (ISRE) and gamma-activated sequence (GAS) element, respectively. IFNtau increased transcription of GAS-driven promoters at 3 h, but suppressed their activity at 24 h. In contrast, the activity of an ISRE-driven promoter was increased at 3 and 24 h. These results indicate that IFNtau activates multiple STATs and has differential effects on ISRE- and GAS-driven gene transcription. PMID:11145571

Stewart, M D; Stewart, D M; Johnson, G A; Vyhlidal, C A; Burghardt, R C; Safe, S H; Yu-Lee, L Y; Bazer, F W; Spencer, T E

2001-01-01

311

Fibronectin, Plasminogen Activator Inhibitor Type 1 (PAI-1) And Uterine Artery Doppler Velocimetry as Markers of Preeclampsia  

OpenAIRE

Objective: The purpose of this study was to examine plasma levels of fibronectin and plasminogen inhibitor type 1 (PAI-1), and alterations in uterine artery (UtA) waveforms throughout normotensive and preeclamptic pregnancies and to analyze its predictive value for the detection of preeclampsia within the second trimester of pregnancy.

Kristina Biskupska Bodova; Kamil Biringer; Karol Dokus; Jela Ivankova; Jan Stasko; Jan Danko

2011-01-01

312

Transcriptional Regulation of Urokinase-type Plasminogen Activator Receptor by Hypoxia-Inducible Factor 1 Is Crucial for Invasion of Pancreatic and Liver Cancer  

Directory of Open Access Journals (Sweden)

Full Text Available Angioinvasion is critical for metastasis with urokinase-type plasminogen activator receptor (uPAR and tumor hypoxia-activated hypoxia-inducible factor 1 (HIF-1 as key players. Transcriptional control of uPAR expression by HIF has never been reported. The aim of the present study, therefore, was to test whether tumor hypoxia-induced HIF expression may be linked to transcriptional activation of uPAR and dependent angioinvasion. We used human pancreatic cancer cells and a model of parental and derived HIF-1?-deficient mouse liver cancer cell lines and performed Northern blot analysis, nuclear runoff assays, electrophoretic mobility shift assay, polymerase chain reaction-generated deletion mutants, luciferase assays, Matrigel invasion assays, and in vivo angioinvasion assays in the chorioallantoic membrane of fertilized chicken eggs. Urokinase-type plasminogen activator receptor promoter analysis resulted in four putative HIF binding sites. Hypoxia strongly induced de novo transcription of uPAR mRNA. With sequential deletion mutants of the uPAR promoter, it was possible to identify one HIF binding site causing a nearly 200-fold increase in luciferase activity. Hypoxia enhanced the number of invading tumor cells in vitro and in vivo. In contrast, HIF-1?-deficient cells failed to upregulate uPAR expression, to activate luciferase activity, and to invade on hypoxia. Taken together, we show for the first time that uPAR is under transcriptional control of HIF and that this is important for hypoxia-induced metastasis.

Peter Büchler

2009-02-01

313

Serum levels of soluble urokinase plasminogen activator receptor is associated with parasitemia in children with acute Plasmodium falciparum malaria infection.  

Science.gov (United States)

Serum levels of soluble urokinase plasminogen activator receptor (suPAR) are significantly elevated and of prognostic value in patients suffering from serious infectious diseases such as HIV and tuberculosis. Our objective was to investigate suPAR levels during symptomatic malaria infection and 7 days after treatment. Children younger than 6 years who presented with fever or other symptoms compatible with malaria were enrolled. Blood films and samples were collected on day 0 and day 7. Twenty-five children were allocated to each of three groups according to the amount of Plasmodium falciparum detected in their initial blood film. Children in group 1 had parasite densities in excess of 20 parasites per 200 leucocytes. The median plasma suPAR level was 6.49 ng/mL (interquartile range [IQR]: 4.90-7.61) and correlated to parasitemia (Spearman 0.43, P suPAR level to median 3.48 ng/mL (IQR: 3.08-3.91) (P suPAR level was median 2.91 ng/mL (IQR: 2.27-4.40) and decreased with median 0.5 ng/mL following treatment (P = 0.0002). Group 3 showed to be negative in their blood slides and most received antibiotic treatment. suPAR decreased from median 3.26 ng/mL (IQR: 2.77-4.46) to median 2.47 ng/mL (IQR: 2.01-3.75), on day 7 (P = 0.006). This study demonstrates an important association between suPAR and acute malaria infection in humans. PMID:15491469

Perch, M; Kofoed, Pe; Fischer, T K; Có, F; Rombo, L; Aaby, P; Eugen-Olsen, J

2004-05-01

314

Coagulation factors, fibrinogen and plasminogen activator inhibitor-1, are differentially regulated by yellow fever virus infection of hepatocytes.  

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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

315

Characterization of the interaction between heterodimeric ?v?6 integrin and urokinase plasminogen activator receptor (uPAR) using functional proteomics.  

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Urokinase plasminogen activator receptor (uPAR) and the epithelial integrin ?v?6 are thought to individually play critical roles in cancer metastasis. These observations have been highlighted by the recent discovery (by proteomics) of an interaction between these two molecules, which are also both implicated in the epithelial-mesenchymal transition (EMT) that facilitates escape of cells from tissue barriers and is a common signature of cancer metastases. In this study, orthogonal in cellulo and in vitro functional proteomic approaches were used to better characterize the uPAR·?v?6 interaction. Proximity ligation assays (PLA) confirmed the uPAR·?v?6 interaction on OVCA429 (ovarian cancer line) and four different colon cancer cell lines including positive controls in cells with de novo ?6 subunit expression. PLA studies were then validated using peptide arrays, which also identified potential physical sites of uPAR interaction with ?v?6, as well as verifying interactions with other known uPAR ligands (e.g., uPA, vitronectin) and individual integrin subunits (i.e., ?v, ?1, ?3, and ?6 alone). Our data suggest that interaction with uPAR requires expression of the complete ?? heterodimer (e.g., ?v?6), not individual subunits (i.e., ?v, ?1, ?3, or ?6). Finally, using in silico structural analyses in concert with these functional proteomics studies, we propose and demonstrate that the most likely unique sites of interaction between ?v?6 and uPAR are located in uPAR domains II and III. PMID:25318615

Ahn, Seong Beom; Mohamedali, Abidali; Anand, Samyuktha; Cheruku, Harish R; Birch, Debra; Sowmya, Gopichandran; Cantor, David; Ranganathan, Shoba; Inglis, David W; Frank, Ronald; Agrez, Michael; Nice, Edouard C; Baker, Mark S

2014-12-01

316

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 pf 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

317

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

318

Upper extremity acute compartment syndrome during tissue plasminogen activator therapy for pulmonary embolism in a morbidly obese patient  

Science.gov (United States)

Introduction Deep vein thrombosis (DVT) and pulmonary embolism (PE) are more frequently observed in morbidly obese patients. Tissue plasminogen activator (tPA) is a thrombolytic agent which dissolves the thrombus more rapidly than conventional heparin therapy and reduces the mortality and morbidity rates associated with PE. Compartment syndrome is a well-known and documented complication of thrombolytic treatment. In awake, oriented and cooperative patients, the diagnosis of compartment syndrome is made based on clinical findings including swelling, tautness, irrational and continuous pain, altered sensation, and severe pain due to passive stretching. These clinical findings may not be able to be adequately assessed in unconscious patients. Presentation of case In this case report, we present compartment syndrome observed, for which fasciotomy was performed on the upper right extremity of a 46-year old morbidly obese, conscious female patient who was receiving tPA due to a massive pulmonary embolism. Discussion Compartment syndrome had occurred due to the damage caused by the repeated unsuccessful catheterisation attempts to the brachial artery and the accompanying tPA treatment. Thus, the bleeding that occurred in the volar compartment of the forearm and the anterior compartment of the arm led to acute compartment syndrome (ACS). After relaxation was brought about in the volar compartment of the forearm and the anterior compartment of the arm, the circulation in the limb was restored. Conclusion As soon as the diagnosis of compartment syndrome is made, an emergency fasciotomy should be performed. Close follow-up is required to avoid wound healing problems after the fasciotomy. PMID:25618841

Tuna, Serkan; Duymus, Tahir Mutlu; Mutlu, Serhat; Ketenci, Ismail Emre; Ulusoy, Ayhan

2015-01-01

319

Recombinant production of a hybrid plasminogen activator composed of surfactant protein B and low-molecular-weight urokinase.  

Science.gov (United States)

Intraalveolar fibrin deposition is commonly observed during acute inflammatory and chronic interstitial lung diseases and may contribute to impairment of surfactant function and gas exchange. We recently described a chemically cross-linked chimeric protein consisting of surfactant protein (SP)-B and urokinase (uPA) for targeting alveolar fibrin under conditions such as acute respiratory distress syndrome (ARDS) or lung fibrosis. We now investigated the feasibility of a recombinant production of a fusion protein encoding mature SP-B and uPA, termed SPUC. Four different SPUC proteins (N-term SP-B/C-term uPA, N-term uPA/C-term SP-B, each +/- His-tag) were prepared by cloning the cDNA encoding mature SP-B and low-molecular-weight uPA into the expression vector pcDNA3.1. CHO-cells were transfected with the constructs and the supernatant and cell lysates were analyzed for expression of SPUC. Using a chromogenic substrate assay uPA activity was found in supernatants and lysates of transfected cells with highest activities related to the N-term uPA/C-term SP-B (+/- His-tag) construct in supernatants 48h after transfection. Casein enzymography showed an enzymatically active fusion proteins with a molecular weight of approximately 42 kDa in the supernatant of cells transfected with the N-term uPA/C-term SP-B (+/- His-tag) construct, but only a minor activity with the N-term SP-B/C-term uPA construct. The N-term uPA/C-term SP-B construct was also shown to possess higher resistance towards inhibition by plasminogen activator inhibitor-1. We conclude that recombinant production of a fusion protein consisting of mature SP-B and uPA is feasible, when the SP-B moiety is fused to the C-terminus of urokinase. PMID:19132247

Ruppert, Clemens; Mahavadi, Poornima; Wygrecka, Malgorzata; Weaver, Timothy E; Magdolen, Viktor; Idell, Steven; Preissner, Klaus T; Seeger, Werner; Günther, Andreas; Markart, Philipp

2008-12-01

320

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

321

Urokinase-type Plasminogen Activator (uPA) is Inhibited with QLT0267 a Small Molecule Targeting Integrin-linked Kinase (ILK)  

OpenAIRE

Urokinase-type plasminogen activator (uPA) is associated with cancer recurrence where the most evidence comes from studies in breast cancer. According to the European Organization for Research and Treatment of Cancer, uPA is considered one of the most prominent biomarkers for cancer recurrence and therefore new agents are needed to inhibit it. Whether uPA is also expressed in pediatric cancers is yet unknown. If it is then uPA inhibitors might also help children with recurrent cancers. In thi...

Santos, Nancy Dos; Habibi, Golareh; Wang, Michelle; Law, Jennifer H.; Andrews, Heather N.; Wei, Daniel; Triche, Timothy; Dedhar, Shoukat; Dunn, Sandra E.

2007-01-01

322

Transcriptional regulation of the rat tissue type plasminogen activator gene: localization of DNA elements and nuclear factors mediating constitutive and cyclic AMP-induced expression.  

OpenAIRE

We have characterized tissue type plasminogen activator (tPA) promoter elements and nuclear factors required for follicle-stimulating hormone (FSH)-induced transcription of the rat tPA gene in granulosa cells and constitutive expression of the gene in the rat neuroblastoma cell line B103. Run-on transcription analysis of isolated nuclei revealed that B103 cells transcribe the tPA gene at a high and constitutive level, while FSH was found to induce tPA gene transcription in a rapid and transie...

Ohlsson, M.; Leonardsson, G.; Jia, X. C.; Feng, P.; Ny, T.

1993-01-01

323

Preparation of ultrasound microbubbles crosslinked to albumin nanoparticles packaged with tissue-type plasminogen activator gene plasmid and method of in vivo transfection  

OpenAIRE

Ji Jun, Ji Shang-Yi, He Xia, Ling Wen-PingDepartment of Pathology, ShenZhen Sun Yat-Sen Cardiovascular Hospital, ShenZhen, GuangDong Province, People's Republic of ChinaAims: To observe the effect of constructed ultrasound microbubble crosslinked to albium nanoparticles packaged with tissue-type plasminogen activator (tPA) gene plasmid on the in vivo transfection.Methods: The rabbits were chosen for all experiments. A highly expressive gene plasmid for tPA was constructed and packaged...

Jun J; Shang-Yi J; Xia H; Wen-Ping L

2011-01-01

324

Npas4 is activated by melatonin, and drives the clock gene Cry1 in the ovine pars tuberalis.  

Science.gov (United States)

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 changes in day length (photoperiod). Melatonin offset at dawn is linked to cAMP accumulation, which directly induces transcription of the clock gene Per1. The rise of melatonin at dusk induces a separate and distinct cohort, including the clock-regulated genes Cry1 and Nampt, but little is known of the up-stream mechanisms involved. Here, we used next-generation sequencing of the ovine PT transcriptome at melatonin onset and identified Npas4 as a rapidly induced basic helix-loop-helix Per-Arnt-Sim domain transcription factor. In vivo we show nuclear localization of NPAS4 protein in presumptive melatonin target cells of the PT (?-glycoprotein hormone-expressing cells), whereas in situ hybridization studies identified acute and transient expression in the PT of Npas4 in response to melatonin. In vitro, NPAS4 forms functional dimers with basic helix loop helix-PAS domain cofactors aryl hydrocarbon receptor nuclear translocator (ARNT), ARNT2, and ARNTL, transactivating both Cry1 and Nampt ovine promoter reporters. Using a combination of 5'-deletions and site-directed mutagenesis, we show NPAS4-ARNT transactivation to be codependent upon two conserved central midline elements within the Cry1 promoter. Our data thus reveal NPAS4 as a candidate immediate early-response gene in the ovine PT, driving molecular responses to melatonin. PMID:23598442

West, A; Dupré, S M; Yu, L; Paton, I R; Miedzinska, K; McNeilly, A S; Davis, J R E; Burt, D W; Loudon, A S I

2013-06-01

325

Npas4 Is Activated by Melatonin, and Drives the Clock Gene Cry1 in the Ovine Pars Tuberalis  

OpenAIRE

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

326

Heparin-binding epidermal growth factor-like growth factor: p91 activation induction of plasminogen activator/inhibitor, and tubular morphogenesis in human microvascular endothelial cells.  

Science.gov (United States)

Epidermal growth factor (EGF) or transforming growth factor-alpha (TGF-alpha) stimulates cell migration, proliferation and the formation of tube-like structures of human microvascular endothelial cells in culture. Heparin-binding EGF-like growth factor(HB-EGF), which shows 35% homology with EGF/TGF-alpha, is a member of the EGF family, and it is ubiquitous in many tissues and organs. We examined whether or not HB-EGF induced angiogenic responses in human microvascular endothelial cells. HB-EGF inhibited the binding of (125) I-EGF to the EGF receptor and induced autophosphorylation of the receptor on endothelial cells. Exogenous HB-EGF induced the loss of more than 70% of the EGF receptor from the cell surface within 30 min, with similar kinetics to that of EGF. The level of c-fos mRNA markedly increased at 30 min in response to HB-EGF as well as EGF. A gel shift assay demonstrated the activation of the transcription factor p91 by HB-EGF and EGF. This factor directly interacts with EGF receptor and mediates the activation of c-fos gene promoter. HB-EGF enhanced the mRNA expression of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) mRNA. However, the enhancement of t-PA and PAI-1 by HB-EGF was less than that by EGF. Heparitinase/chlorate, which digests the heparan sulfate proteoglycan of the endothelial cell surface, restored both t-PA and PAI-1 mRNA levels in response to HB-EGF in a manner similar to that by EGF. HB-EGF at 10 ng/ml developed tube-like structures in type I collagen gel at similar levels to that of EGF at 10 ng/ml, suggesting that HB-EGF is also a potent angiogenic factor in the model system for angiogenesis. The tubulogenesis activity of HB-EGF is discussed in relation to the expression of the t-PA and PAI-1 genes. PMID:8609052

Ushiro, S; Ono, M; Izumi, H; Kohno, K; Taniguchi, N; Higashiyama, S; Kuwano, M

1996-01-01

327

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

328

Relationship of circulating soluble urokinase plasminogen activator receptor (suPAR) levels to disease control in asthma and asthmatic pregnancy.  

Science.gov (United States)

Asthma has a high burden of morbidity if not controlled and may frequently complicate pregnancy, posing a risk for pregnancy outcomes. Elevated plasma level of the inflammatory biomarker soluble urokinase plasminogen activator receptor (suPAR) is related to a worse prognosis in many conditions such as infectious, autoimmune, or pregnancy-related diseases; however the value of suPAR in asthma and asthmatic pregnancy is unknown. The present study aimed to investigate the suPAR, CRP and IL-6 levels in asthma (asthmatic non-pregnant, ANP; N?=?38; female N?=?27) and asthmatic pregnancy (AP; N?=?15), compared to healthy non-pregnant controls (HNP; N?=?29; female N?=?19) and to healthy pregnant women (HP; N?=?58). The relationship between suPAR levels and asthma control was also evaluated. The diagnostic efficacy of suPAR in asthma control was analyzed using ROC analysis. IL-6 and CRP levels were comparable in all study groups. Circulating suPAR levels were lower in HP and AP than in HNP and ANP subjects, respectively (2.01 [1.81-2.38] and 2.39 [2.07-2.69] vs. 2.60 [1.82-3.49] and 2.84 [2.33-3.72] ng/mL, respectively, p?=?0.0001). suPAR and airway resistance correlated in ANP (r?=?0.47, p?=?0.004). ROC analysis of suPAR values in ANP patients with PEF above and below 80% yielded an AUC of 0.75 (95% CI: 0.57-0.92, p?=?0.023) and with ACT total score above and below 20 an AUC of 0.80 (95% CI: 0.64-0.95, p?=?0.006). The cut-off value of suPAR to discriminate between controlled and not controlled AP and ANP was 4.04 ng/mL. In conclusion, suPAR may help the objective assessment of asthma control, since it correlates with airway resistance and has good sensitivity in the detection of impaired asthma control. Decrease in circulating suPAR levels detected both in healthy and asthmatic pregnant women presumably represents pregnancy induced immune tolerance. PMID:23565268

Ivancsó, István; Toldi, Gergely; Bohács, Anikó; Eszes, Noémi; Müller, Veronika; Rigó, János; Vásárhelyi, Barna; Losonczy, György; Tamási, Lilla

2013-01-01

329

Intravenous Thrombolysis with Recombinant Tissue Plasminogen Activator for Ischemic Stroke Patients over 80 Years Old: The Fukuoka Stroke Registry  

Science.gov (United States)

Objectives The benefit of intravenous recombinant tissue plasminogen activator (rt-PA) therapy for very old patients with acute ischemic stroke remains unclear. The aim of this study was to elucidate the efficacy and safety of intravenous rt-PA therapy for patients over 80 years old. Methods Of 13,521 stroke patients registered in the Fukuoka Stroke Registry in Japan from June 1999 to February 2013, 953 ischemic stroke patients who were over 80 years old, hospitalized within 3 h of onset, and not treated with endovascular therapy were included in this study. Among them, 153 patients were treated with intravenous rt-PA (0.6 mg/kg). For propensity score (PS)-matched case-control analysis, 148 patients treated with rt-PA and 148 PS-matched patients without rt-PA therapy were selected by 1?1 matching with propensity for using rt-PA. Clinical outcomes were neurological improvement, good functional outcome at discharge, in-hospital mortality, and hemorrhagic complications (any intracranial hemorrhage [ICH], symptomatic ICH, and gastrointestinal bleeding). Results In the full cohort of 953 patients, rt-PA use was associated positively with neurological improvement and good functional outcome, and negatively with in-hospital mortality after adjustment for multiple confounding factors. In PS-matched case-control analysis, patients treated with rt-PA were still at lower risk for unfavorable clinical outcomes than non-treated patients (neurological improvement, odds ratio 2.67, 95% confidence interval 1.61–4.40; good functional outcome, odds ratio 2.23, 95% confidence interval 1.16–4.29; in-hospital mortality, odds ratio 0.30, 95% confidence interval 0.13–0.65). There was no significant association between rt-PA use and risk of hemorrhagic complications in the full and PS-matched cohorts. Conclusions Intravenous rt-PA therapy was associated with improved clinical outcomes without significant increase in risk of hemorrhagic complications in very old patients (aged>80 years) with acute ischemic stroke. PMID:25329379

Matsuo, Ryu; Kamouchi, Masahiro; Fukuda, Haruhisa; Hata, Jun; Wakisaka, Yoshinobu; Kuroda, Junya; Ago, Tetsuro; Kitazono, Takanari

2014-01-01

330

Prognostic value analysis of urokinase-type plasminogen activator receptor in oral squamous cell carcinoma: an immunohistochemical study  

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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

331

Probiotic in rennet paste can affect lipase activity of rennet and lipolysis in ovine cheese  

OpenAIRE

Lambs were subjected to three different feeding regimes (mother suckling MS, artificial rearing AR, and artificial rearing with 7log10 cfu/ml Lactobacillus acidophilus supplementation to the milk substitute ARLb) and slaughtered at 20d and 40d of age for each feeding treatment. Lambs abomasa were processed to rennet paste and lipases activity was evaluated. Rennet paste was used for Pecorino cheese production. Free fatty acids (FFAs) and conjugated linoleic acids (CLAs) were detected in chees...

Marzia Albenzio; Rosaria Marino; Mariangela Caroprese; Antonella Santillo

2010-01-01

332

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.

Furlan, Federico; Galbiati, Clara

2007-01-01

333

Antagonistic anti-urokinase plasminogen activator receptor (uPAR) antibodies significantly inhibit uPAR-mediated cellular signaling and migration.  

Science.gov (United States)

Interactions between urokinase plasminogen activator receptor (uPAR) and its various ligands regulate tumor growth, invasion, and metastasis. Antibodies that bind specific uPAR epitopes may disrupt these interactions, thereby inhibiting these processes. Using a highly diverse and naïve human fragment of the antigen binding (Fab) phage display library, we identified 12 unique human Fabs that bind uPAR. Two of these antibodies compete against urokinase plasminogen activator (uPA) for uPAR binding, whereas a third competes with beta1 integrins for uPAR binding. These competitive antibodies inhibit uPAR-dependent cell signaling and invasion in the non-small cell lung cancer cell line, H1299. Additionally, the integrin-blocking antibody abrogates uPAR/beta1 integrin-mediated H1299 cell adhesion to fibronectin and vitronectin. This antibody and one of the uPAR/uPA antagonist antibodies shows a significant combined effect in inhibiting cell invasion through Matrigel/Collagen I or Collagen I matrices. Our results indicate that these antagonistic antibodies have potential for the detection and treatment of uPAR-expressing tumors. PMID:20501655

Duriseti, Sai; Goetz, David H; Hostetter, Daniel R; LeBeau, Aaron M; Wei, Ying; Craik, Charles S

2010-08-27

334

Plasminogen activator inhibitor type 1 gene is located at region q21. 3-q22 of chromosome 7 and genetically linked with cystic fibrosis  

Energy Technology Data Exchange (ETDEWEB)

The regional chromosomal location of the human gene for plasminogen activator inhibitor type 1 (PAI1) was determined by three independent methods of gene mapping. PAI1 was localized first to 7cen-q32 and then to 7q21.3-q22 by Southern blot hybridization analysis of a panel of human and mouse somatic cell hybrids with a PAI1 cDNA probe and in situ hybridization, respectively. The authors frequent HindIII restriction fragment length polymorphism (RFLP) of the PAI1 gene with an information content of 0.369. In family studies using this polymorphism, genetic linkage was found between PAI1 and the loci for erythropoietin (EPO), paraoxonase (PON), the met protooncogene (MET), and cystic fibrosis (CF), all previously assigned to the middle part of the long arm of chromosome 7. The linkage with EPO was closest with an estimated genetic distance of 3 centimorgans, whereas that to CF was 20 centimorgans. A three-point genetic linkage analysis and data from previous studies showed that the most likely order of these loci is EPO, PAI1, PON, (MET, CF), with PAI1 being located centromeric to CF. The PAI1 RFLP may prove to be valuable in ordering genetic markers in the CF-linkage group and may also be valuable in genetic analysis of plasminogen activation-related diseases, such as certain thromboembolic disorders and cancer.

Klinger, K.W.; Winqvist, R.; Riccio, A.; Andreasen, P.A.; Sartorio, R.; Nielsen, L.S.; Stuart, N.; Stanislovitis, P.; Watkins, P.; Douglas, R.

1987-12-01

335

Inhibition of plasminogen activator inhibitor-1 binding to endocytosis receptors of the low density lipoprotein receptor family by a peptide isolated from a phage displayed library  

DEFF Research Database (Denmark)

The functions of the serpin plasminogen activator inhibitor-1 (PAI-1) are based on molecular interactions with its target proteases urokinase-type and tissue-type plasminogen activator (uPA and tPA), with vitronectin, and with endocytosis receptors of the low density lipoprotein family. Understanding the significance of these interactions would be facilitated by the ability to block them individually. Using phage display, we have identified the disulphide constrained peptide motif CFGWC with affinity for natural human PAI-1. The three-dimensional structure of a peptide containing this motif (DVPCFGWCQDA) was determined by NMR. A binding site in the so-called flexible joint region of PAI-1 was suggested by molecular modelling and validated through binding studies with various competitors and site-directed mutagenesis of PAI-1. The peptide with an N-terminal biotin inhibited the binding of the uPA-PAI-1 complex to the endocytosis receptors low density lipoprotein receptor-related protein (LRP-1A) and very low density lipoprotein receptor (VLDLR) in vitro and inhibited endocytosis of the uPA-PAI-1 complex in U937 cells. We conclude that the isolated peptide represents a novel approach to pharmacological interference with the functions of PAI-1 based on inhibition of one specific molecular interaction.

Jensen, Jan K.; Malmendal, Anders

2006-01-01

336

Inhibition of plasminogen activator inhibitor-1 binding to endocytosis receptors of the low-density-lipoprotein receptor family by a peptide isolated from a phage display library  

DEFF Research Database (Denmark)

The functions of the serpin PAI-1 (plasminogen activator inhibitor-1) are based on molecular interactions with its target proteases uPA and tPA (urokinase-type and tissue-type plasminogen activator respectively), with vitronectin and with endocytosis receptors of the low-density-lipoprotein family. Understanding the significance of these interactions would be facilitated by the ability to block them individually. Using phage display, we have identified the disulfide-constrained peptide motif CFGWC with affinity for natural human PAI-1. The three-dimensional structure of a peptide containing this motif (DVPCFGWCQDA) was determined by liquid-state NMR spectroscopy. A binding site in the so-called flexible joint region of PAI-1 was suggested by molecular modelling and validated through binding studies with various competitors and site-directed mutagenesis of PAI-1. The peptide with an N-terminal biotin inhibited the binding of the uPA-PAI-1 complex to the endocytosis receptors low-density-lipoprotein-receptor-related protein 1A (LRP-1A) and very-low-density-lipoprotein receptor (VLDLR) in vitro and inhibited endocytosis of the uPA-PAI-1 complex in U937 cells. We conclude that the isolated peptide represents a novel approach to pharmacological interference with the functions of PAI-1 based on inhibition of one specific molecular interaction.

Jensen, Jan Kristian; Malmendal, Anders

2006-01-01

337

Plasminogen activator inhibitor type 1 gene is located at region q21.3-q22 of chromosome 7 and genetically linked with cystic fibrosis  

International Nuclear Information System (INIS)

The regional chromosomal location of the human gene for plasminogen activator inhibitor type 1 (PAI1) was determined by three independent methods of gene mapping. PAI1 was localized first to 7cen-q32 and then to 7q21.3-q22 by Southern blot hybridization analysis of a panel of human and mouse somatic cell hybrids with a PAI1 cDNA probe and in situ hybridization, respectively. The authors frequent HindIII restriction fragment length polymorphism (RFLP) of the PAI1 gene with an information content of 0.369. In family studies using this polymorphism, genetic linkage was found between PAI1 and the loci for erythropoietin (EPO), paraoxonase (PON), the met protooncogene (MET), and cystic fibrosis (CF), all previously assigned to the middle part of the long arm of chromosome 7. The linkage with EPO was closest with an estimated genetic distance of 3 centimorgans, whereas that to CF was 20 centimorgans. A three-point genetic linkage analysis and data from previous studies showed that the most likely order of these loci is EPO, PAI1, PON, (MET, CF), with PAI1 being located centromeric to CF. The PAI1 RFLP may prove to be valuable in ordering genetic markers in the CF-linkage group and may also be valuable in genetic analysis of plasminogen activation-related diseases, such as certain thromboembolic disorders and cancer

338

High levels of tissue plasminogen activator (tPA antigen precede the development of type 2 diabetes in a longitudinal population study. The Northern Sweden MONICA Study  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Impaired fibrinolysis is found in impaired glucose tolerance and type 2 diabetes, associated with components of the metabolic syndrome. There are no data concerning fibrinolysis in subjects with normal glucose tolerance that convert to diabetes. Methods We studied the activities of tissue plasminogen activator (tPA and plasminogen activator inhibitor-1 (PAI-1 and the levels of tPA antigen (a marker of endothelial dysfunction in 551 subjects with normal glucose tolerance in 1990 in relation to incident diabetes during nine years of follow-up. Results Subjects with diabetes at follow-up (n = 15 had significantly lower baseline tPA activity and higher PAI-1 activity and tPA antigen than non-converters. The risk of diabetes increased linearly across quartiles of PAI-activity (p = 0.007 and tPA antigen (p p = 0.026. The risk of diabetes with low tPA activity or high PAI-1 activity persisted after adjustment for age and sex but diminished to a non-significant level after further adjustments. The odds ratio of diabetes for high tPA antigen was 10.4 (95% confidence interval 2.7–40 adjusted for age and sex. After further adjustment for diastolic blood pressure, waist circumference, insulin, triglycerides, fasting and post load glucose the odds ratio was 6.5 (1.3–33, p = 0.024. Conclusions Impaired fibrinolysis and endothelial dysfunction are evident in subjects with normal glucose tolerance who later develop diabetes. High tPA antigen is predictive of future diabetes independent from the metabolic syndrome.

Jansson Jan-Håkan

2003-12-01

339

Targeting Tumor Cell Invasion and Dissemination In Vivo by an Aptamer That Inhibits Urokinase-type Plasminogen Activator through a Novel Multifunctional Mechanism  

DEFF Research Database (Denmark)

Data accumulated over the latest two decades have established that the serine protease urokinase-type plasminogen activator (uPA) is a potential therapeutic target in cancer. When designing inhibitors of the proteolytic activity of serine proteases, obtaining sufficient specificity is problematic, because the topology of the proteases' active sites are highly similar. In an effort to generate highly specific uPA inhibitors with new inhibitory modalities, we isolated uPA-binding RNA aptamers by screening a library of 35 nucleotides long 2'-fluoro-pyrimidine RNA molecules using a version of human pro-uPA lacking the epidermal growth factor-like and kringle domains as bait. One pro-uPA-binding aptamer sequence, referred to as upanap-126, proved to be highly specific for human uPA. Upanap-126 delayed the proteolytic conversion of human pro-uPA to active uPA, but did not inhibit plasminogen activation catalyzed by two-chain uPA. The aptamer also inhibited the binding of pro-uPA to uPAR and the binding of vitronectin to the preformed pro-uPA/uPAR complex, both in cell-free systems and on cell surfaces. Furthermore, upanap-126 inhibited human tumor cell invasion in vitro in the Matrigel assay and in vivo in the chick embryo assay of cell escape from microtumors. Finally, upanap-126 significantly reduced the levels of tumor cell intravasation and dissemination in the chick embryo model of spontaneous metastasis. Together, our findings show that usage of upanap-126 represents a novel multifunctional mechanistic modality for inhibition of uPA-dependent processes involved in tumor cell spread.

Botkjaer, Kenneth A; Deryugina, Elena I

2012-01-01

340

Targeting tumor cell invasion and dissemination in vivo by an aptamer that inhibits urokinase-type plasminogen activator through a novel multifunctional mechanism  

DEFF Research Database (Denmark)

Data accumulated over the latest two decades have established that the serine protease urokinase-type plasminogen activator (uPA) is a potential therapeutic target in cancer. When designing inhibitors of the proteolytic activity of serine proteases, obtaining sufficient specificity is problematic, because the topology of the proteases' active sites are highly similar. In an effort to generate highly specific uPA inhibitors with new inhibitory modalities, we isolated uPA-binding RNA aptamers by screening a library of 35 nucleotides long 2'-fluoro-pyrimidine RNA molecules using a version of human pro-uPA lacking the epidermal growth factor-like and kringle domains as bait. One pro-uPA-binding aptamer sequence, referred to as upanap-126, proved to be highly specific for human uPA. Upanap-126 delayed the proteolytic conversion of human pro-uPA to active uPA, but did not inhibit plasminogen activation catalyzed by two-chain uPA. The aptamer also inhibited the binding of pro-uPA to uPAR and the binding of vitronectin to the preformed pro-uPA/uPAR complex, both in cell-free systems and on cell surfaces. Furthermore, upanap-126 inhibited human tumor cell invasion in vitro in the Matrigel assay and in vivo in the chick embryo assay of cell escape from microtumors. Finally, upanap-126 significantly reduced the levels of tumor cell intravasation and dissemination in the chick embryo model of spontaneous metastasis. Together, our findings show that usage of upanap-126 represents a novel multifunctional mechanistic modality for inhibition of uPA-dependent processes involved in tumor cell spread.

Botkjaer, Kenneth A; Deryugina, Elena I

2012-01-01

341

Effects of cytokine-suppressive anti-inflammatory drugs on inflammatory activation in ex vivo human and ovine fetal membranes.  

Science.gov (United States)

Intrauterine infection and inflammation are responsible for the majority of early (Ureaplasma parvum or saline control and subjected to explant culture. The effects of treatment with CSAIDs or vehicle (1% DMSO) on accumulation of PGE2 and cytokines (human interleukin 6 (IL6), IL10 and TNF?; ovine IL8 (oIL8)) were assessed in conditioned media at various time points (3-20 ?h). In human membranes, the IKK? inhibitor TPCA-1 (7 ??M) and p38 MAPK inhibitor SB239063 (20 ??M) administered to the amniotic compartment were the most effective in inhibiting accumulation of cytokines and PGE2 in the fetal compartment. NAC (10 ?mM) inhibited accumulation of PGE2 and IL10 only; NBDI (10 ??M) had no significant effect. In addition to the fetal compartment, SB239063 also exerted consistent and significant inhibitory effects in the maternal compartment. TPCA-1 and SB239063 suppressed oIL8 production, while all CSAIDs tested suppressed ovine PGE2 production. These results support the further investigation of intra-amniotically delivered CSAIDs for the prevention of inflammation-mediated PTB. PMID:24493151

Stinson, Lisa F; Ireland, Demelza J; Kemp, Matthew W; Payne, Matthew S; Stock, Sarah J; Newnham, John P; Keelan, Jeffrey A

2014-03-01

342

Induction of alpha2-antiplasmin inhibits E-cadherin processing mediated by the plasminogen activator/plasmin system, leading to suppression of progression of oral squamous cell carcinoma via upregulation of cell-cell adhesion.  

Science.gov (United States)

The plasminogen activator/plasmin system is one of the main protease systems involved in tumor cell invasion and metastasis. Our previous study has shown that plasmin degrades E-cadherin and promotes cell dissemination by downregulation of E-cadherin-mediated cell-cell adhesion in oral squamous cell carcinoma (SCC) cells. To examine the effect of downregulation of the plasminogen activator/plasmin system by alpha2-antiplasmin (alpha2-AP) on cell-cell adhesion mediated by E-cadherin in oral SCC cells, the oral SCC cell line SCCKN was stably transfected with alpha2-AP cDNA. Induction of alpha2-AP expression led to the inhibition of the proteolysis of E-cadherin by plasminogen activator/plasmin in SCC cells, resulting in the enhancement of the cell aggregation and the suppression of the cell motility. Moreover, alpha2-AP also reduced the ability of SCC cells to invade type I collagen gel, and suppressed tumorigenicity in vivo. These results suggested that downregulation of the plasminogen activator/ plasmin system by alpha2-AP might be a potent therapeutic approach to prevent the progression of oral SCC. PMID:17203182

Hayashido, Yasutaka; Hamana, Tomoaki; Ishida, Yasutaka; Shintani, Tomoaki; Koizumi, Koh-Ichi; Okamoto, Tetsuji

2007-02-01

343

In vitro and in vivo antiangiogenic activity of a novel deca-peptide derived from human tissue-type plasminogen activator kringle 2  

International Nuclear Information System (INIS)

A synthetic deca-peptide corresponding to the amino acid sequence Arg54-Trp63 of human tissue-type plasminogen activator (t-PA) kringle 2 domain, named TKII-10, is produced and tested for its ability to inhibit endothelial cell proliferation, migration, tube formation in vitro, and angiogenesis in vivo. At the same time, another peptide TKII-10S composed of the same 10 amino acids as TKII-10, but in a different sequence, is also produced and tested. The results show that TKII-10 potently inhibits VEGF-stimulated endothelial cell migration and tube formation in a dose-dependent, as well as sequence-dependent, manner in vitro while it is inactive in inhibiting endothelial cell proliferation. Furthermore, TKII-10 potently inhibits angiogenesis in chick chorioallantoic membrane and mouse cornea. The middle four amino acids DGDA in their sequence play an important role in TKII-10 angiogenesis inhibition. These results suggest that TKII-10 is a novel angiogenesis inhibitor that may serve as a prototype for antiangiogenic drug development.

344

Influence of growth factors on the plasminogen activator activity of avian granulosa cells from follicles at different maturational stages of preovulatory development.  

Science.gov (United States)

Granulosa cells from the first (F1), third (F3) and fifth and sixth (F5-6) preovulatory follicles and the small yellow follicles (SYFs; diameter 6-8 mm) were cultured for 21 h in the absence and presence of murine and human epidermal growth factors, fibroblast growth factor, transforming growth factors alpha and beta-I (TGF alpha, TGF beta), platelet-derived growth factor and insulin-like growth factor-I at concentrations of 0.1-100 ng/ml. Plasminogen activator (PA) activities in the cell (PAc) and in the medium (PAm) were measured by fibrinolysis and fibrin overlay methods. Basal PAc and PAm activities were highest in cell cultures from the less mature follicles (F5-6 and SYF) and decreased as the follicles matured (F3 > F1). PAc activity was greater than PAm activity, irrespective of the stage of follicular development. All growth factors examined at the 100 ng/ml level were effective in increasing PAc and PAm activities in cultures of granulosa cells from F1 follicles. However, only TGF alpha was able to increase PA activities at lower concentrations. The stimulation of the PA activities of granulosa cells from F3 follicles was inconsistent. None of the growth factors significantly increased PA activities in granulosa cells from F5-6 follicles and SYFs, as determined by fibrinolysis. The major PAc and PAm species (characterized by fibrin overlay) had a molecular mass of about 35 kDa, which is characteristic of the urokinase type. Both assay methods detected a stimulatory effect of the growth factors on PA activities in the granulosa cells from F1 follicles. However, an increase in PA activities in cells from F3 and F5-6 follicles and SYFs was indicated only after fibrin overlay analysis. Tritiated thymidine was incorporated into the DNA of granulosa cells at all stages of follicular development and was enhanced by all growth factors, although TGF alpha and TGF beta were the most effective and had a ranked order of activity: F3, F5-6 > F1, SYF. The present findings show that, of the growth factors examined, TGF alpha may be an effective regulator of PA activity in avian granulosa cells during follicular development, in addition to its observed mitogenic action. PMID:8148037

Lafrance, M; Croze, F; Tsang, B K

1993-12-01

345

Staurosporine induces ganglion cell differentiation in part by stimulating urokinase-type plasminogen activator expression and activation in the developing chick retina  

International Nuclear Information System (INIS)

Highlights: ? Staurosporine mediates stimulation of RGC differentiation in vitro cultured retinal neuroblasts. ? Staurosporine mediates uPA activation during RGC differentiation in vitro. ? Inhibition of uPA blocks the staurosporine mediated RGC differentiation both in vitro and in ovo. ? Thus, uPA may play a role in the staurosporine-mediated stimulation of RGC differentiation. -- Abstract: Here, we investigated whether staurosporine-mediated urokinase-type plasminogen activator (uPA) activation is involved in retinal ganglion cell (RGC) differentiation. Retinal cells were isolated from developing chick retinas at embryonic day 6 (E6). Relatively few control cells grown in serum-free medium started to form processes by 12 h. In contrast, staurosporine-treated cells had processes within 3 h, and processes were evident at 8 h. Immunofluorescence staining showed that Tuj-1-positive cells with shorter neurites could be detected in control cultures at 18 h, whereas numerous Tuj-1 positive ganglion cells with longer neuritic extensions were seen in staurosporine-treated cultures. BrdU-positive proliferating cells were more numerous in control cultures than in staurosporine-treated cultures, and the BrdU staining was not detected in post-mitotic Tuj-1 positive ganglion cells. Western blotting of cell lysates showed that staurosporine induced high levels of the active form of uPA. The staurosporine-induced uPA signal was localized predominantly in the soma, neurites and axons of Tuj-1-positive ganglion cells. Amiloride, an inhibitor of uPA, markedly reduced staurosporine-induced Tuj-1 staining, neurite length, neurite number, and uPA staining versus controls. In developing retinas in ovo, amiloride administration remarkably reduced the staurosporine-induced uPA staining and RGC differentiation. Taken together, our in vitro and in vivo data collectively indicate that uPA plays a role in the staurosporine-mediated stimulation of RGC differentiation.

346

PACAP Interacts with PAC1 Receptors to Induce Tissue Plasminogen Activator (tPA) Expression and Activity in Schwann Cell-Like Cultures.  

Science.gov (United States)

Regeneration of peripheral nerves depends on the abilities of rejuvenating axons to migrate at the injury site through cellular debris and altered extracellular matrix, and then grow along the residual distal nerve sheath conduit and reinnervate synaptic targets. Considerable evidence suggest that glial cells participate in this process, although the mechanisms remain to be clarified. In cell culture, regenerating neurites secrete PACAP, a peptide shown to induce the expression of the protease tissue plasminogen activator (tPA) in neural cell types. In the present studies, we tested the hypothesis that PACAP can stimulate peripheral glial cells to produce tPA. More specifically, we addressed whether or not PACAP promoted the expression and activity of tPA in the Schwann cell line RT4-D6P2T, which shares biochemical and physical properties with Schwann cells. We found that PACAP dose- and time-dependently stimulated tPA expression both at the mRNA and protein level. Such effect was mimicked by maxadilan, a potent PAC1 receptor agonist, but not by the PACAP-related homolog VIP, suggesting a PAC1-mediated function. These actions appeared to be mediated at least in part by the Akt/CREB signaling cascade because wortmannin, a PI3K inhibitor, prevented peptide-driven CREB phosphorylation and tPA increase. Interestingly, treatment with BDNF mimicked PACAP actions on tPA, but acted through both the Akt and MAPK signaling pathways, while causing a robust increase in PACAP and PAC1 expression. PACAP6-38 totally blocked PACAP-driven tPA expression and in part hampered BDNF-mediated effects. We conclude that PACAP, acting through PAC1 receptors, stimulates tPA expression and activity in a Akt/CREB-dependent manner to promote proteolytic activity in Schwann-cell like cultures. PMID:25658447

Castorina, Alessandro; Waschek, James A; Marzagalli, Rubina; Cardile, Venera; Drago, Filippo

2015-01-01

347

Promotor polymorphisms of plasminogen activator inhibitor-1 and other thrombophilic genotypes in cerebral venous thrombosis: a case-control study in adults.  

Science.gov (United States)

Plasminogen activator inhibitor 1 (PAI-1) is the main inhibitor of tissue-type and urokinase-type plasminogen activator. A 4G/5G polymorphism in the promoter region of the PAI-1 gene has been reported to enhance the plasma levels of PAI-1. In particular, the 4G allele (guanosine deletion) has been linked with increased plasma PAI-1 levels, which may lead to impaired activity of the fibrinolytic system, thus increasing the incidence of thrombotic events. The aim of this case-control study was to analyze whether variants of the PAI-1 promotor genotype 4G/4G, 4G/5G and 5G/5G, in particular the 4G/5G-variant, constitute an independent risk factor of cerebral venous thrombosis (CVT). A total of 136 consecutive patients with proven CVT were compared to 1,054 DNA specimens of healthy controls from a population-based cohort. PAI-1 promotor polymorphisms were evaluated using polymerase chain reaction. No significant association of CVT with PAI-1 4G/5G was found in either the additive (OR 1.04; 95 % CI 0.78-1.38) or in the dominant model (OR 1.24; 95 % CI 0.72-2.13). Also, the prevalence of the other genotypes (4G/4G and 5G/5G) in patients was not significantly different from controls. When considering the variants of the PAI-1 promoter genotype in combination with known genetical thrombophilias, no differences were found either. As was expected, the prothrombin (G20210A) genotype was confirmed as an independent risk factor for CVT. We conclude that the 4G allele of the PAI-1 polymorphism does not increase the risk of CVT in adults. PMID:22527222

Ringelstein, Marius; Jung, Alexander; Berger, Klaus; Stoll, Monika; Madlener, Katharina; Klötzsch, Christof; Schlachetzki, Felix; Stolz, Erwin

2012-11-01

348

Correlation between plasminogen activator inhibitor-1 (PAI-1) promoter 4G/5G polymorphism and metabolic/proinflammatory factors in polycystic ovary syndrome.  

Science.gov (United States)

Polycystic Ovary Syndrome (PCOS) is the most common cause of subfertility associated to metabolic disorders. The aim of this study was to correlate metabolic and proinflammatory factors in women with PCOS. The frequency of Plasminogen Activator Inhibitor-1 (PAI-1) promoter 4?G/5?G polymorphism was also compared to healthy controls. We evaluated 79 PCOS and 79 healthy women. PAI-1 levels are positively correlated with proinflammatory factors in PCOS group. 4?G allele in PAI-1 gene was more frequent in PCOS and the 4G/4?G genotype was associated with increased PAI-1 levels. A correlation between insulin resistance and proinflammatory and overweight was also observed. C-reactive protein, serum levels of vascular cell adhesion molecule-1 (sVCAM-1), Lipid Accumulation Product (LAP) and vitamin D are good tools to evaluated factors associated to cardiovascular risk in women with PCOS. PMID:23898913

Sales, M F; Sóter, M O; Candido, A L; Fernandes, A P; Oliveira, F R; Ferreira, A C S; Sousa, M O; Ferreira, C N; Gomes, K B

2013-10-01

349

Plasminogen Activator Inhibitor-1-675 4G/5G and Methylenetetrahydrofolate Reductase Gene Variants in Young Acute Myocardial Infarction and Juvenile Ischemic Stroke  

Directory of Open Access Journals (Sweden)

Full Text Available Cardiovascular diseases often recognize a hereditary-familial genetic risk patterns and a substantial proportion of variability in clinical features of atherosclerosis could probably be explained by genetic factors. Aim of this study is to compare prevalence of factor V Leiden, factor II 20210A, PAI-1-675 4G/5G and MTHFR C677T gene variants in 2 cardiovascular disease clinical features: young acute myocardial infarction (<50 years and ischemic stroke. Plasminogen Activator Inhibitor-1-675 4G/5G didn`t show significant difference between patients and controls while, between patient groups, the polymorphism was most present in myocardial infarction group than in juvenile stroke. Polymorphisms in Homocysteine metabolism: MTHFR C677T variants was significantly higher in patients with myocardial infarction compared with those affected by ischemic Stroke.

2008-01-01

350

Elevated plasma tissue plasminogen activator and anti-THP-1 antibodies are independently associated with decreased graft survival in cardiac transplant recipients.  

Science.gov (United States)

Hemostatic and immunologic factors have been implicated in future cardiac events in patients with coronary artery disease. The role of these factors and their interaction is less established in cardiac transplant recipients. We sought to characterize the role of these factors in these patients. Cardiac transplant patients who presented for surveillance coronary angiography and/or endomyocardial biopsy were eligible for enrollment. Ninety-nine consecutive patients were enrolled. Plasma levels of tissue-type plasminogen activator (t-PA), plasminogen activator inhibitor-1, von Willebrand factor, fibrin D-dimer, and anti-t-PA antibody were determined by enzyme-linked immunosorbent assays. Anti-THP-1 cell antibodies directed against a monocytic leukemia cell line were detected by incubating patient plasma with THP-1 cells. Bound antibody was detected using goat peroxidase-labeled immunoglobulin G directed against human immunoglobulins. Lipids were measured by enzymatic methods. Multivariate analysis identified the presence of anti-THP-1 cell antibodies (risk ratio 4.41, p = 0.002), t-PA antigen (risk ratio 1.10, p = 0.033), donor age 20 to 26 years (risk ratio 8.83, p = 0.042), and donor age >36 years (risk ratio 15.53, p = 0.009) as predictors of allograft failure. Altered hemostatic function, as demonstrated by elevated plasma t-PA antigen levels, is predictive of subsequent allograft failure in cardiac transplant recipients. In addition, the presence of anti-THP-1 cell antibodies in these patients is predictive of allograft failure. PMID:11423054

Warshofsky, M K; Dominguez, M; Eisenberg, M S; Wasserman, H S; Sciacca, R; Wang, W; Simon, A D; Morse, J H; Schwartz, A; Anglés-Cano, E; Rabbani, L E

2001-07-01

351

Lysosomal degradation of receptor-bound urokinase-type plasminogen activator is enhanced by its inhibitors in human trophoblastic choriocarcinoma cells  

DEFF Research Database (Denmark)

We have studied the effect of plasminogen activator inhibitors PAI-1 and PAI-2 on the binding of urokinase-type plasminogen activator (u-PA) to its receptor in the human choriocarcinoma cell line JAR. With 125I-labeled ligands in whole-cell binding assays, both uncomplexed u-PA and u-PA-inhibitor complexes bound to the receptor with a Kd of approximately 100 pM at 4 degrees C. Transferring the cells to 37 degrees C led to degradation to amino acids of up to 50% of the cell-bound u-PA-inhibitor complexes, whereas the degradation of uncomplexed u-PA was 15%; the remaining ligand was recovered in an apparently intact form in the medium or was still cell associated. The degradation could be inhibited by inhibitors of vesicle transport and lysosomal hydrolases. By electron microscopic autoradiography, both 125I-u-PA and 125I-u-PA-inhibitor complexes were located over the cell membrane at 4 degrees C, with the highest density of grains over the membrane at cell-cell interphases, but, after incubation at 37 degrees C, 17 and 27% of the grains for u-PA and u-PA-PAI-1 complexes, respectively, appeared over lysosomal-like bodies. These findings suggest that the u-PA receptor possesses a clearance function for the removal of u-PA after its complex formation with a specific inhibitor. The data suggest a novel mechanism by which receptor-mediated endocytosis is initiated by the binding of a secondary ligand. Full text

Jensen, Poul Henning; Christensen, Erik IlsØ

1990-01-01

352

Epidermal growth factor receptor-mediated regulation of urokinase plasminogen activator expression and glioblastoma invasion via C-SRC/MAPK/AP-1 signaling pathways.  

Science.gov (United States)

One of the major pathophysiological features of malignant astrocytomas is their ability to infiltrate surrounding brain tissue. The epidermal growth factor receptor (EGFR) and proteases are known to be overexpressed in glioblastomas (GBMs), but the interaction between the activation of the EGFR and urokinase plasminogen activator (uPA) in promoting astrocytic tumor invasion has not been fully elucidated. Here, we characterized the signal transduction pathway(s) by which EGF regulates uPA expression and promotes astrocytoma invasion. We show that EGFR activation and constitutively active EGFR vIII in GBM cell lines upregulate uPA expression. Small-molecule inhibitors of mitogen-activated protein kinase, tyrosine kinase, and small interfering RNA targeting c-Src blocked uPA upregulation. Similarly, mutations in the activator protein 1 binding site of the uPA promoter reduced EGF-induced increases in uPA promoter activity. Treatment of GBM cells with EGF increased in vitro cell invasion, and the invasive phenotype was attenuated by gene silencing of uPA using small interfering RNA and short hairpin RNA. In addition, uPA knockdown clones formed smaller well-circumscribed tumors than nontarget U1242 control cells in a xenograft GBM mouse model in vivo. In summary, these results suggest that c-Src, mitogen-activated protein kinase, and a composite activator protein 1 on the uPA promoter are responsible for EGF-induced uPA expression and GBM invasion. PMID:20467333

Amos, Samson; Redpath, Gerard T; Dipierro, Charles G; Carpenter, Joan E; Hussaini, Isa M

2010-06-01

353

Bacterial plasminogen receptors: in vitro evidence for a role in degradation of the mammalian extracellular matrix.  

OpenAIRE

The potential of bacterium-bound plasmin to degrade mammalian extracellular matrix and to enhance bacterial penetration through basement membrane was assessed with the adherent strain SH401-1 of Salmonella enterica serovar Typhimurium. Typhimurium SH401-1 was able to bind plasminogen and to enhance the tissue-type plasminogen activator-mediated activation of the single-chain plasminogen to the two-chain plasmin. The end product, the enzymatically active, bacterium-bound plasmin activity, was ...

La?hteenma?ki, K.; Virkola, R.; Pouttu, R.; Kuusela, P.; Kukkonen, M.; Korhonen, T. K.

1995-01-01

354

Nicotine stimulates urokinase-type plasminogen activator receptor expression and cell invasiveness through mitogen-activated protein kinase and reactive oxygen species signaling in ECV304 endothelial cells  

Energy Technology Data Exchange (ETDEWEB)

Urokinase-type plasminogen activator receptor (uPAR) expression is elevated during inflammation, tissue remodeling and in many human cancers. This study investigated the effect of nicotine, a major alkaloid in tobacco, on uPAR expression and cell invasiveness in ECV304 endothelial cells. Nicotine stimulated uPAR expression in a dose-dependent manner and activated extracellular signal-regulated kinases-1/2 (Erk-1/2), c-Jun amino-terminal kinase (JNK) and p38 mitogen activated protein kinase (MAPK). Specific inhibitors of MEK-1 (PD98059) and JNK (SP600125) inhibited the nicotine-induced uPAR expression, while the p38 MAPK inhibitor SB203580 did not. Expression vectors encoding dominant negative MEK-1 (pMCL-K97M) and JNK (TAM67) also prevented nicotine-induced uPAR promoter activity. The intracellular hydrogen peroxide (H{sub 2}O{sub 2}) content was increased by nicotine treatment. The antioxidant N-acetylcysteine prevented nicotine-activated production of reactive oxygen species (ROS) and uPAR expression. Furthermore, exogenous H{sub 2}O{sub 2} increased uPAR mRNA expression. Deleted and site-directed mutagenesis demonstrated the involvement of the binding sites of transcription factor nuclear factor-kappaB (NF-?B) and activator protein (AP)-1 in the nicotine-induced uPAR expression. Studies with expression vectors encoding mutated NF-?B signaling molecules and AP-1 decoy confirmed that NF-?B and AP-1 were essential for the nicotine-stimulated uPAR expression. MAPK (Erk-1/2 and JNK) and ROS functioned as upstream signaling molecules in the activation of AP-1 and NF-?B, respectively. In addition, ECV304 endothelial cells treated with nicotine displayed markedly enhanced invasiveness, which was partially abrogated by uPAR neutralizing antibodies. The data indicate that nicotine induces uPAR expression via the MAPK/AP-1 and ROS/NF-?B signaling pathways and, in turn, stimulates invasiveness in human ECV304 endothelial cells. -- Highlights: ? Endothelial cells treated with nicotine displayed enhanced invasiveness. ? Nicotine induces uPAR expression and, in turn, stimulates invasiveness. ? MAPK/AP-1 and ROS/NF-?B signals are involved in nicotine-induced uPAR.

Khoi, Pham Ngoc; Park, Jung Sun; Kim, Nam Ho; Jung, Young Do, E-mail: ydjung@chonnam.ac.kr

2012-03-01

355

Alpha-Enolase Is Upregulated on the Cell Surface and Responds to Plasminogen Activation in Mice Expressing a ?133p53? Mimic  

Science.gov (United States)

The p53 protein is a master regulator of the stress response. It acts as a tumor suppressor by inducing transcriptional activation of p53 target genes, with roles in apoptosis, cell cycle arrest and metabolism. The discovery of at least 12 isoforms of p53, some of which have tumor-promoting properties, has opened new avenues of research. Our previous work studied tumor phenotypes in four mouse models with different p53 backgrounds: wild-type p53, p53 null, mutant p53 lacking the proline domain (m?pro), and a mimic for the human ?133p53? p53 isoform (?122p53). To identify the major proteins affected by p53 function early in the response to DNA damage, the current study investigated the entire proteome of bone marrow, thymus, and lung in the four p53 models. Protein extracts from untreated controls and those treated with amsacrine were analyzed using two-dimensional fluorescence difference gel electrophoresis. In the bone marrow, reactive proteins were universally decreased by wild-type p53, including ?-enolase. Further analysis of ?-enolase in the p53 models revealed that it was instead increased in ?122p53 hematopoietic and tumor cell cytosol and on the cell surface. Alpha-enolase on the surface of ?122p53 cells acted as a plasminogen receptor, with tumor necrosis factor alpha induced upon plasminogen stimulation. Taken together, these data identified new proteins associated with p53 function. One of these proteins, ?-enolase, is regulated differently by wild-type p53 and ?122p53 cells, with reduced abundance as part of a wild-type p53 response and increased abundance with ?122p53 function. Increased cell surface ?-enolase on ?122p53 cells provides a possible explanation for the model’s pro-inflammatory features and suggests that p53 isoforms may direct an inflammatory response by increasing the amount of ?-enolase on the cell surface. PMID:25643152

Sawhney, Sonal; Hood, Kylie; Shaw, Alisha; Braithwaite, Antony W.; Stubbs, Richard; Hung, Noelyn A.; Royds, Janice A.; Slatter, Tania L.

2015-01-01

356

Staurosporine induces ganglion cell differentiation in part by stimulating urokinase-type plasminogen activator expression and activation in the developing chick retina  

Energy Technology Data Exchange (ETDEWEB)

Highlights: Black-Right-Pointing-Pointer Staurosporine mediates stimulation of RGC differentiation in vitro cultured retinal neuroblasts. Black-Right-Pointing-Pointer Staurosporine mediates uPA activation during RGC differentiation in vitro. Black-Right-Pointing-Pointer Inhibition of uPA blocks the staurosporine mediated RGC differentiation both in vitro and in ovo. Black-Right-Pointing-Pointer Thus, uPA may play a role in the staurosporine-mediated stimulation of RGC differentiation. -- Abstract: Here, we investigated whether staurosporine-mediated urokinase-type plasminogen activator (uPA) activation is involved in retinal ganglion cell (RGC) differentiation. Retinal cells were isolated from developing chick retinas at embryonic day 6 (E6). Relatively few control cells grown in serum-free medium started to form processes by 12 h. In contrast, staurosporine-treated cells had processes within 3 h, and processes were evident at 8 h. Immunofluorescence staining showed that Tuj-1-positive cells with shorter neurites could be detected in control cultures at 18 h, whereas numerous Tuj-1 positive ganglion cells with longer neuritic extensions were seen in staurosporine-treated cultures. BrdU-positive proliferating cells were more numerous in control cultures than in staurosporine-treated cultures, and the BrdU staining was not detected in post-mitotic Tuj-1 positive ganglion cells. Western blotting of cell lysates showed that staurosporine induced high levels of the active form of uPA. The staurosporine-induced uPA signal was localized predominantly in the soma, neurites and axons of Tuj-1-positive ganglion cells. Amiloride, an inhibitor of uPA, markedly reduced staurosporine-induced Tuj-1 staining, neurite length, neurite number, and uPA staining versus controls. In developing retinas in ovo, amiloride administration remarkably reduced the staurosporine-induced uPA staining and RGC differentiation. Taken together, our in vitro and in vivo data collectively indicate that uPA plays a role in the staurosporine-mediated stimulation of RGC differentiation.

Kim, Yeoun-Hee [Department of Biology, College of Natural Sciences, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Chang, Yongmin [Department of Molecular Medicine, Kyungpook National University College of Medicine, Kyungpook National University, 200 Dongduk-Ro Jung-Gu, Daegu 700-714 (Korea, Republic of); Jung, Jae-Chang, E-mail: jcjung@knu.ac.kr [Department of Biology, College of Natural Sciences, Kyungpook National University, Daegu 702-701 (Korea, Republic of)

2012-06-22

357

Plasminogen-stimulated inflammatory cytokine production by airway smooth muscle cells is regulated by annexin A2.  

Science.gov (United States)

Plasminogen has a role in airway inflammation. Airway smooth muscle (ASM) cells cleave plasminogen into plasmin, a protease with proinflammatory activity. In this study, the effect of plasminogen on cytokine production by human ASM cells was investigated in vitro. Levels of IL-6 and IL-8 in the medium of ASM cells were increased by incubation with plasminogen (5-50 ?g/ml) for 24 hours (P reduced (P asthma. PMID:23721211

Schuliga, Michael; Langenbach, Shenna; Xia, Yuxiu C; Qin, Chengxue; Mok, Josephine S L; Harris, Trudi; Mackay, Graham A; Medcalf, Robert L; Stewart, Alastair G

2013-11-01

358

Glycosylated ovine prolactin.  

OpenAIRE

A modification of prolactin, in which the asparagine at position 31 carries a carbohydrate unit, was isolated from ovine pituitary glands. Sequence and amino acid analyses identified the point of linkage of the carbohydrate. Glucosamine was found in acid hydrolysates, an indication that the carbohydrate is attached through N-acetylglucosamine. The glycosylated prolactin binds to concanavalin A and lentil lectin and is eluted with methyl alpha-D-mannopyranoside. During gel electrophoresis in s...

Lewis, U. J.; Singh, R. N.; Lewis, L. J.; Seavey, B. K.; Sinha, Y. N.

1984-01-01

359

Tumor necrosis factor induction of endothelial cell urokinase-type plasminogen activator mediated proteolysis of extracellular matrix and its antagonism by gamma-interferon.  

Science.gov (United States)

Tumor necrosis factor (TNF) has a profound capacity to alter the endothelial cell phenotype that includes morphologic and functional changes that may be central for proinflammatory processes. Recent observations have indicated that TNF can promote the synthesis and secretion of urokinase plasminogen activator (uPA) in low passage human endothelial cells that normally release little uPA. In this report we have confirmed and expanded upon these initial observations in human endothelial cells and describe the ability of gamma-interferon (gamma-IFN) to inhibit TNF-induced uPA synthesis and secretion in a dose-dependent manner (0.025 to 25 ng/mL). Analysis of cell-free conditioned medium derived from gamma-IFN-treated cultures by micro-enzyme-linked immunosorbent assay (ELISA) methodologies using uPA- and plasminogen activator inhibitor type 1 (PAI-1)-specific monoclonal antibodies (MoAbs) indicate that the decrease in uPA activity observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) zymography is a direct result of a decrease in extracellular uPA antigen and is not a consequence of increased PAI-1 antigen. These findings are supported by Northern blot analyses that indicate that gamma-IFN treatment of endothelial cells resulted in a decreased steady state level of uPA messenger RNA (mRNA) with no measurable change in PAI-1 mRNA. This inhibitory response is specific for gamma-IFN because alpha-IFN fails to elicit a similar inhibitory response. In addition, TNF augmented extracellular proteolysis of radiolabeled subendothelial extracellular matrix (ECM) in a dose-dependent manner. The observed increase in ECM degradation mediated by TNF treatment of endothelial cells was dependent on the presence of plasminogen and could be inhibited by an anticatalytic uPA MoAb implying the requirement of proteolytically active uPA in this process. gamma-IFN (25 ng/mL) treatment of endothelial cells antagonized TNF-promoted degradation of radiolabeled ECM at a concentration that completely inhibited TNF-mediated uPA expression and activity. In addition, endothelial cells treated with TNF (18 hours) displayed the ability to invade ECM and reorganize individual cells into tube-like structures that were not evident in untreated control cultures when grown on Matrigel-coated culture dishes. gamma-IFN treatment of endothelial cells propagated on Matrigel was observed to inhibit TNF-mediated ECM invasion and tube formation at concentrations that were analogous to those required for the inhibition of uPA expression and activity. In summary, these observations suggest a novel homeostatic control mechanism for endothelial cell regulation of subendothelial ECM degradation promoted by TNF and inhibited by gamma-IFN.(ABSTRACT TRUNCATED AT 400 WORDS) PMID:1732009

Niedbala, M J; Picarella, M S

1992-02-01

360

Ochratoxin A inhibits the production of tissue factor and plasminogen activator inhibitor-2 by human blood mononuclear cells: Another potential mechanism of immune-suppression  

International Nuclear Information System (INIS)

The mycotoxin ochratoxin A (OTA), an ubiquitous contaminant of food products endowed with a wide spectrum of toxicity, affects several functions of mononuclear leukocytes. Monocytes/macrophages play a major role in fibrin accumulation associated with immune-inflammatory processes through the production of tissue factor (TF) and plasminogen activator inhibitor 2 (PAI-2). We studied the effect of OTA on TF and PAI-2 production by human blood mononuclear cells (MNC). The cells were incubated for 3 or 18 h at 37 deg. C with non toxic OTA concentrations in the absence and in the presence of lipopolysaccharide (LPS) or other inflammatory agents. TF activity was measured by a one-stage clotting test. Antigen assays were performed by specific ELISAs in cell extracts or conditioned media and specific mRNAs were assessed by RT-PCR. OTA had no direct effect on TF and PAI-2 production by MNC. However, OTA caused a dose-dependent reduction in LPS-induced TF (activity, antigen and mRNA) and PAI-2 (antigen and mRNA) production with > 85% inhibition at 1 ?g/ml. Similar results were obtained when monocyte-enriched preparations were used instead of MNC. TF production was also impaired by OTA (1 ?g/ml) when MNC were stimulated with phorbol myristate acetate (98% inhibition), IL-1? (83%) or TNF-? (62%). The inhibition of TF and PAI-2 induction might represent a hitherto unrecognized mechanism whereby OTA exerts immunosuppressant activity

361

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

Science.gov (United States)

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 expressed on neutrophils, mediates endogenous thrombolysis through a urokinase-dependent mechanism. Here we show that membrane GPI-anchored uPAR is decreased or absent on granulocytes and platelets of patients with PNH, while soluble uPAR (suPAR) levels are increased in patients' plasma. Serum suPAR concentrations correlated with the number of GPI-negative neutrophils and were highest in patients who later develop thrombosis. In vitro, suPAR is released from PNH hematopoietic cells and from platelets upon activation, suggesting that these cells are the probable source of plasma suPAR in the absence of GPI anchor synthesis and trafficking of uPAR to the cell membrane. In vitro, the addition of recombinant suPAR results in a dose-dependent decrease in the activity of single-chain urokinase. We hypothesized that suPAR, prevents the interaction of urokinase with membrane-anchored uPAR on residual normal cells. PMID:18954937

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

2008-12-01

362

Inhibitory effects of C-type natriuretic peptide on the differentiation of cardiac fibroblasts, and secretion of monocyte chemoattractant protein-1 and plasminogen activator inhibitor-1.  

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The present study aimed to investigate the effect of C-type natriuretic peptide (CNP) on the function of cardiac fibroblasts (CFs). Western blotting was used to investigate the expression of myofibroblast marker proteins: ?-smooth muscle actin (?-SMA), extra domain-A fibronectin, collagen I and collagen III, and the activity of extracellular signal-regulated kinase 1/2 (ERK1/2). Immunofluorescence was used to examine the morphological changes; a transwell assay was used to analyze migration, and reverse transcription-quantitative polymerase chain reaction and ELISA were employed to determine the mRNA expression and protein secretion of monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1). The results demonstrated that CNP significantly reduced the protein expression of ?-SMA, fibronectin, collagen I and collagen III, and suppressed the migratory ability of CFs. Additionally, the mRNA and protein expression of MCP-1 and PAI-1 was inhibited under the CNP treatment; and this effect was mediated by the inhibition of the ERK1/2 activity. In conclusion, CNP inhibited cardiac fibroblast differentiation and migration, and reduced the secretion of MCP-1 and PAI-1, which demonstrates novel mechanisms to explain the antifibrotic effect of CNP. PMID:25352084

Li, Zhi-Qiang; Liu, Ying-Long; Li, Gang; Li, Bin; Liu, Yang; Li, Xiao-Feng; Liu, Ai-Jun

2015-01-01

363

Modulation of ovine SBD-1 expression by 17beta-estradiol in ovine oviduct epithelial cells  

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Full Text Available Abstract Background Mucosal epithelia, including those of the oviduct, secrete antimicrobial innate immune molecules (AIIMS. These have bactericidal/bacteriostatic functions against a variety of pathogens. Among the AIIMs, sheep ?-defensin-1 (SBD-1 is one of the most potent. Even though the SBD-1 is an important AIIM and it is regulated closely by estrogenic hormone, the regulation mechanism of 17?-estradiol has not been clearly established. We investigated the effects of E2 and agonist or inhibitor on ovine oviduct epithelial cells in regard to SBD-1 expression using reverse transcription quantitative PCR (RT-qPCR. In addition, three different pathways were inhibited separately or simultaneously to confirm the effect of different inhibitors in the regulation mechanism. Results 17beta-estradiol (E2 induced release of SBD-1 in ovine oviduct epithelial cells. SBD-1 expression was mediated through G-protein-coupled receptor 30 (GPR30 and Estrogen Receptors (ERs activation in ovine oviduct epithelial cell. Inhibition of gene expression of protein kinase A (PKA, protein kinase C (PKC, and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-?B led to a decreased SBD-1 expression. Conclusions Taken together, E2-induced up-regulation of SBD-1 expressions were GPR30-dependent during prophase and ERs-dependent during later-stage in ovine oviduct epithelial cells, and we assume that the effect was completed by the PKA, PKC, and NF-?B pathways simultaneous.

Wen Shiyong

2012-08-01

364

Differences between binding of one-chain and two-chain tissue plasminogen activators to non-cross-linked and cross-linked fibrin clots  

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Interaction of tissue plasminogen activator (t-PA) with fibrin plays a key role in regulation of plasminogen activation and clot dissolution. Previous investigations of t-PA-fibrin interaction, using incorporation of t-PA into polymerizing fibrin clots, have suggested that no significant differences exist in the binding of one-chain or two-chain t-PA to non-cross-linked or cross-linked fibrin. In the present study, binding of 125I-labeled and affinity-purified one-chain and two-chain forms of t-PA to preformed non-cross-linked or cross-linked, sonicated suspension of fibrin was investigated. Interaction of one-chain t-PA with cross-linked fibrin involved a single type of binding site with dissociation constant (kd) of 0.58 mumol/L and a stoichiometry (n) of 1.5. Interaction of one-chain t-PA with non-cross-linked fibrin, however, involved two classes of binding sites with dissociation constants of 0.32 and 1.5 mumol/L and corresponding number of binding sites equal to 0.57 and 2.0, respectively. In contrast to the binding of one-chain t-PA to cross-linked fibrin by a limited number of sites, two-chain t-PA appeared to involve a considerably greater number of sites (minimum six) whose dissociation constant was 3.2 mumol/L. Interaction of two-chain t-PA with non-cross-linked fibrin also showed the presence of many binding sites (minimum seven) with approximate dissociation constant of 6.4 mumol/L, as well as a few (n = 0.012) high-affinity sites with a kd of 0.011 mumolgh-affinity sites with a kd of 0.011 mumol/L epsilon-Aminocaproic acid did not completely reverse the binding of either one-chain t-PA or two-chain t-PA to fibrin. The present findings suggest that the fibrin-binding properties of t-PA undergo considerable changes on proteolytic conversion from one-chain to two-chain t-PA, catalyzed under physiologic conditions by plasmin

365

Human circadian system causes a morning peak in prothrombotic plasminogen activator inhibitor-1 (PAI-1) independent of the sleep/wake cycle.  

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Serious adverse cardiovascular events peak in the morning, possibly related to increased thrombosis in critical vessels. Plasminogen activator inhibitor-1 (PAI-1), which inhibits fibrinolysis, is a key circulating prothrombotic factor that rises in the morning in humans. We tested whether this morning peak in PAI-1 is caused by the internal circadian system or by behaviors that typically occur in the morning, such as altered posture and physical activity. Twelve healthy adults underwent a 2-week protocol that enabled the distinction of endogenous circadian effects from behavioral and environmental effects. The results demonstrated a robust circadian rhythm in circulating PAI-1 with a peak corresponding to ?6:30 am. This rhythm in PAI-1 was 8-times larger than changes in PAI-1 induced by standardized behavioral stressors, including head-up tilt and 15-minute cycle exercise. If this large endogenous morning peak in PAI-1 persists in vulnerable individuals, it could help explain the morning peak in adverse cardiovascular events. PMID:24200683

Scheer, Frank A J L; Shea, Steven A

2014-01-23

366

Dissecting the effect of RNA aptamer binding on the dynamics of plasminogen activator inhibitor 1 using hydrogen/deuterium exchange mass spectrometry  

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RNA aptamers, selected from large synthetic libraries, are attracting increasing interest as protein ligands, with potential uses as prototype pharmaceuticals, conformational probes, and reagents for specific quantification of protein levels in biological samples. Very little is known, however, about their effects on protein conformation and dynamics. We have employed hydrogen/deuterium exchange (HDX) mass spectrometry to study the effect of RNA aptamers on the structural flexibility of the serpin plasminogen activator inhibitor-1 (PAI-1). The aptamers have characteristic effects on the biochemical properties of PAI-1. In particular, they are potent inhibitors of the structural transition of PAI-1 from the active state to the inactive, so-called latent state. This transition is one of the largest conformational changes of a folded protein domain without covalent modification. Binding of the aptamers to PAI-1 is associated with substantial and widespread protection against deuterium uptake in PAI-1. The aptamers induce protection against exchange with the solvent both in the protein-aptamer interface as well as in other specific areas. Interestingly, the aptamers induce substantial protection against exchange in ?-helices B, C and I. This observation substantiates the relevance of structural instability in this region for transition to the latent state and argues for involvement of flexibility in regions not commonly associated with regulation of latency transition in serpins.

Trelle, Morten B; Dupont, Daniel Miotto

2014-01-01

367

Tissue-type plasminogen activator acts as a cytokine that triggers intracellular signal transduction and induces matrix metalloproteinase-9 gene expression.  

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Tissue-type plasminogen activator (tPA), a serine protease well known for generating plasmin, has been demonstrated to induce matrix metalloproteinase-9 (MMP-9) gene expression and protein secretion in renal interstitial fibroblasts. However, exactly how tPA transduces its signal into the nucleus to control gene expression is unknown. This study investigated the mechanism by which tPA induces MMP-9 gene expression. Both wild-type and non-enzymatic mutant tPA were found to induce MMP-9 expression in rat kidney interstitial fibroblasts (NRK-49F), indicating that the actions of tPA are independent of its proteolytic activity. tPA bound to the low density lipoprotein receptor-related protein-1 (LRP-1) in NRK-49F cells, and this binding was competitively abrogated by the LRP-1 antagonist, the receptor-associated protein. In mouse embryonic fibroblasts (PEA-13) lacking LRP-1, tPA failed to induce MMP-9 expression. Furthermore, tPA induced rapid tyrosine phosphorylation on the beta subunit of LRP-1, which was followed by the activation of Mek1 and its downstream Erk-1 and -2. Blockade of Erk-1/2 activation by the Mek1 inhibitor abolished MMP-9 induction by tPA in NRK-49F cells. Conversely, overexpression of constitutively activated Mek1 induced Erk-1/2 phosphorylation and MMP-9 expression. In mouse obstructed kidney, tPA, LRP-1, and MMP-9 were concomitantly induced in the renal interstitium. Collectively, these results suggest that besides its classical proteolytic activity, tPA acts as a cytokine that binds to the cell membrane receptor LRP-1, induces its tyrosine phosphorylation, and triggers intracellular signal transduction, thereby inducing specific gene expression in renal interstitial fibroblasts. PMID:16303771

Hu, Kebin; Yang, Junwei; Tanaka, Sakae; Gonias, Steven L; Mars, Wendy M; Liu, Youhua

2006-01-27

368

Clonal variation of expression of the genes coding for plasminogen activators, their inhibitors and the urokinase receptor in HT1080 sarcoma cells.  

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The human sarcoma cell line HT1080 was found, by in situ hybridization, to consist of cells expressing various levels of urokinase (uPA) and tissue type (tPA) plasminogen activator (PA) suggesting clonal variation of expression of these genes. Colonies originating from single HT1080 cells were, therefore, established and screened for PA activity using a fibrin agarose overlay. Colonies inducing lysis (clone C+ and H+) or no lysis (clones B- and M-) were isolated and tested for mRNA levels of uPA, tPA, uPA receptor (uPAR) and the 3 PA inhibitors (PAI), PAI-1, PAI-2 and protease-nexin I. The different clones revealed considerable variation of expression of the different PA and PAI genes, with lysis-inducing clones expressing mainly the PA genes, whereas non-lysing clones demonstrated higher expression of the PAI genes. Amplification or loss of specific genes was excluded by Southern blotting. The protein levels of cellular and secreted PA and PAI determined by ELISA and Western blots demonstrated a pattern similar to that observed for PA and PAI mRNA concentrations, suggesting clonal differences either on the level of transcription or in RNA processing and/or stability. Due to complex interactions between PA and PAI, neither mRNA nor protein levels of the different genes were predictive for the amount of functional PA activity present in the supernatant or on the cell surface of the different clones. Receptor-bound uPA activity was found to be considerably higher in lysis-inducing than in non-lysing clones and the activity was dependent on neutralization by PAI-1 rather than on the level of uPAR mRNA. PMID:1325952

Laug, W E; Wang, K; Mundi, R; Rideout, W; Kruithof, E K; Bogenmann, E

1992-09-01

369

Evidence for a pre-latent form of the serpin plasminogen activator inhibitor-1 with a detached beta-strand 1C  

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Latency transition of plasminogen activator inhibitor-1 (PAI-1) occurs spontaneously in the absence of proteases and results in stabilization of the molecule through insertion of its reactive center loop (RCL) as a strand in beta-sheet A and detachment of beta-strand 1C (s1C) at the C-terminal hinge of the RCL. This is one of the largest structural rearrangements known for a folded protein domain without a concomitant change in covalent structure. Yet, the sequence of conformational changes during latency transition remains largely unknown. We have now mapped the epitope for the monoclonal antibody H4B3 to the cleft revealed upon s1C detachment and shown that H4B3 inactivates recombinant PAI-1 in a time-dependent manner. With fluorescence spectroscopy, we show that insertion of the RCL is accelerated in the presence of H4B3, demonstrating that the loss of activity is the result of latency transition. Considering that the epitope for H4B3 appears to be occluded by s1C in active PAI-1, this finding suggests theexistence of a pre-latent conformation on the path from active to latent PAI-1 characterized by at least partial detachment of s1C. Functional characterization of mutated PAI-1 variants suggests that a salt-bridge between Arg273 and Asp224 may stabilize the pre-latent conformation. The binding of H4B3 and of a peptide targeting the cleft revealed upon s1C detachment was hindered by the glycans attached to Asn267. Conclusively, we have provided evidence for the existence of an equilibrium between active PAI-1 and a pre-latent form, characterized by reversible detachment of s1C and formation of a glycan-shielded cleft in the molecule.

Dupont, Daniel M; Blouse, Grant E

2006-01-01

370

Up-regulation of urokinase plasminogen activator messenger ribonucleic acid and protein in hen granulosa cells by transforming growth factor alpha in vitro during follicular development.  

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The aim of the present study was to examine the regulatory role of transforming growth factor alpha (TGF alpha) on urokinase plasminogen activator (uPA) gene expression and protein levels in hen granulosa cells from different stages of ovarian follicular development in vitro. Granulosa cells from the first (F1), the second and third (F2-3), and the fourth, fifth, and sixth (F4-6) largest preovulatory follicles were cultured for 21 h in the absence and presence of TGF alpha (10 ng/ml). The uPA mRNA abundance and protein content were determined by Northern and Western blot analysis, respectively. Cell-associated and secreted PA activity was measured by a fibrinolysis assay and characterized by zymography. Hen granulosa cells produce a uPA with a molecular mass of about 35 kDa and a transcript size of approximately 2.5 kb. Basal uPA mRNA abundance, protein content, and activity were highest in granulosa cells from F4-6 follicles and decreased with follicular maturation. Granulosa cell uPA mRNA levels, protein content, and activity were increased in the presence of TGF alpha, reaching maximal levels in granulosa cells from less mature follicles, although the percentage of stimulation was higher in cells from late stages of follicular development. These findings clearly demonstrate specific expression of uPA in proliferatively active granulosa cells and responsiveness of uPA to TGF alpha at both transcriptional and translational levels. They support the concept that PA of the urokinase type plays an important role in extracellular matrix remodeling during TGF alpha-induced granulosa cell proliferation and ovarian follicular growth. PMID:9160733

Li, J; Croze, F; Yan, W; Haché, R J; Tsang, B K

1997-05-01

371

Camptothecin induces urokinase-type plasminogen activator gene-expression in human RC-K8 malignant lymphoma and H69 small cell lung cancer cells.  

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Full Text Available We previously reported that anthracyclines, which could generate reactive oxygen species (ROS, could induce the urokinase-type plasminogen activator (uPA gene expression in human RC-K8 malignant lymphoma cells and in H69 small cell lung cancer (SCLC cells. In screening other uPA-inducible anti-cancer agents, we found that camptothecin (CPT and its derivative, SN38, could induce uPA in RC-K8 and H69 cells. CPT and SN38, which are also used for the treatment of lymphoma and SCLC, significantly increased the uPA accumulation in the conditioned media of both cells in a dose-dependent manner. The maximum induction of uPA mRNA levels was observed 24 h after stimulation. Pretreatment with pyrrolidine dithiocarbamate (PDTC, an anti-oxidant, inhibited the CPT-induced uPA mRNA expression. Thus, CPT induces uPA through gene expression, and, therefore, CPT may influence the tumor-cell biology by up-regulating the uPA/plasmin system.

Shibakura M

2002-10-01

372

Prognostic value of plasma soluble urokinase plasminogen activator receptor (suPAR) in Danish patients with recurrent epithelial ovarian cancer (REOC).  

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The level of the soluble urokinase plasminogen activator receptor (suPAR) is elevated in tumour tissue from several types of cancer. This is the first study aiming to predict the prognosis for survival by the use of a pre-chemotherapeutic plasma suPAR value in 71 patients with recurrent epithelial ovarian cancer (REOC). For determination of suPAR, pre-chemotherapeutic blood samples from the patients with REOC were processed into plasma (EDTA) within one working day from venipuncture. The plasma suPAR level is not correlated with performance status (p=0.41), FIGO stage (p=0.09), treatment-free interval (TFI) of 12 months (p=0.26), site of recurrence (peritoneum, p=0.50 or pelvis, p=0.44), age (p=0.43), or serum CA125 (p=0.09). Univariate as well as multivariate analyses cannot demonstrate that high pre-chemotherapeutic levels of plasma suPAR (p=0.22, p=0.80) are associated with shorter survival of REOC patients. Multivariate analysis showed that only TFI of 12 months (p=0.001) and performance score status of 2 (p=0.02) were independent prognostic factors. Our study indicates that pre-chemotherapeutic measurement of plasma suPAR level in REOC patients may not be useful to identify a subgroup of patients with poor prognosis. PMID:17004970

Begum, Farah Diba; Høgdall, Estrid V S; Riisbro, Rikke; Christensen, Ib Jarle; Engelholm, Svend A; Jørgensen, Morten; Pedersen, Bent Nørgaard; Høgdall, Claus K

2006-10-01

373

Usefulness of soluble urokinase plasminogen activator receptor to predict repeat myocardial infarction and mortality in patients with ST-segment elevation myocardial infarction undergoing primary percutaneous intervention.  

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The plasma level of the inflammatory biomarker soluble urokinase plasminogen activator receptor (suPAR) is an independent predictor of cardiovascular disease and all-cause mortality in healthy subjects. The prognostic capability of suPAR, its temporal course, and its relation to plasma C-reactive protein (CRP) in patients with ST-segment