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1

Safety of topical thrombins: the ongoing debate  

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Full Text Available Abstract Until recently, only bovine-derived thrombin was available for use as a stand-alone topical hemostat or as a component of other hemostatic devices. Concerns over a number of case reports of immune-mediated coagulopathies associated with the use of bovine-derived thrombin resulted in a United States Food and Drug Administration warning letter being issued in 1996 and the later addition of a boxed warning ("Black Box Warning" to all bovine-derived thrombin products. Since 2007, both a human-pooled plasma thrombin product and a recombinant thrombin have entered the market. With the addition of these two products to the topical thrombin class, a unique situation has developed in which only a single member (bovine-derived thrombin within the class carries the Food and Drug Administration's strongest cautionary language about possible adverse events related to an agent's use. Neither the human-pooled plasma thrombin nor the recombinant thrombin products have a boxed warning; although, the human-pooled plasma product does include a precaution/warning about infectious agent transmission - a warning common to products derived from human sources. This report will address this unique situation and the impact, clinical and non-clinical, that thrombin choice may have. Since alternatives are now available, institutions may need to revisit their formulary choice of thrombin preparation, taking into consideration the potential risks associated with bovine-derived products.

Lomax Christopher

2009-09-01

2

Topical recombinant thrombin at a concentration of 1000 IU/mL reliably shortens in vivo TTH and delivers durable hemostasis in the presence of heparin anticoagulation and clopidogrel platelet inhibition in a rabbit model of vascular bleeding  

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Full Text Available Abstract Background This study was designed to evaluate the effect of recombinant human thrombin (rThrombin concentration on time to hemostasis (TTH, clot durability, and clot strength in settings that replicate the heparinization and platelet inhibition often found in surgical populations. Methods A modified, anticoagulated rabbit arteriovenous shunt preparation was selected to model vascular anastomotic bleeding. Rabbits were treated with heparin or heparin + clopidogrel and TTH was measured after applying a range of topical rThrombin concentrations or placebo, in combination with absorbable gelatin sponge, USP. Treatments (placebo, rThrombin were randomly assigned and the investigator was blinded to treatment. TTH was evaluated with the Kaplan-Meier method. After hemostasis was achieved, clot burst assessment was performed for heparin + clopidogrel treated animals. Clot viscoelastic strength and kinetics were measured in ex-vivo samples using thromboelastography (TEG methods. Results TTH decreased with increasing concentrations of rThrombin in heparin-treated animals and was shorter after treatment with 1000 IU/mL rThrombin (73 seconds than with 125 IU/mL rThrombin (78 seconds; p = 0.007. TTH also decreased with increasing concentrations of rThrombin in heparin + clopidogrel treated animals; again it was significantly shorter after treatment with 1000 IU/mL rThrombin (71 seconds than with 125 IU/mL rThrombin (177 seconds; p  Conclusion In an animal model designed to replicate the anti-coagulation regimens encountered in clinical settings, topical rThrombin at 1000 IU/mL more reliably controlled the pharmacological effects of heparin or heparin + clopidogrel on hemostasis than rThrombin at 125 IU/mL. Results from in vitro assessments confirmed a positive relationship between the amount of rThrombin activity and both the rate of clot formation and clot strength.

Garcia Richard

2009-11-01

3

Chemical modifications and amino acid substitutions in recombinant hirudin that increase hirudin-thrombin affinity.  

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Recombinant hirudin (r-hirudin), unlike the naturally occurring leech protein, lacks a sulfate ester on Tyr-63 which reduces its binding affinity to thrombin by 3-10-fold. We demonstrate that nitration or iodination of Tyr-63 restores hirudin-thrombin affinity to levels similar to or exceeding that of the natural inhibitor. In contrast, nitration of Tyr-3 reduces the affinity of hirudin for thrombin. These chemical modifications results in multiple reaction products that are readily separated by reverse-phase HPLC. The mechanism of the observed changes in thrombin affinity may involve a reduction in the pK of the hydroxyl group of tyrosine due to substitution of the electrophilic iodo or nitro group on the phenyl ring, resulting in an increased negative charge at neutral pH. For Tyr-63, this effect mimics the sulfatotyrosine of natural hirudin, leading to an increased thrombin affinity at the anion-binding exosite. For Tyr-3, the increased polarity may destabilize its interaction within the apolar-binding site of thrombin. Substitution of the highly conserved Tyr-3 residue with Phe or Trp not only enables specific and quantitative chemical modification at Tyr-63 but also independently increases hirudin-thrombin affinity. Kinetic analysis of thrombin inhibition showed that enhanced binding by r-hirudin(nitro-Tyr-63) is due to an increase in the association rate between hirudin and thrombin whereas the reduced binding of r-hirudin(nitro-Tyr-3) results from a large increase in the dissociation rate. These observations indicate that specific segments within both the amino- and carboxy-terminal regions of hirudin interact with thrombin. PMID:1991108

Winant, R C; Lazar, J B; Johnson, P H

1991-02-01

4

Functional characterization of recombinant batroxobin, a snake venom thrombin-like enzyme, expressed from Pichia pastoris.  

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A thrombin-like enzyme of Bothrops atrox moojeni venom, batroxobin, specifically cleaves fibrinogen alpha chain, resulting in the formation of non-crosslinked fibrin clots. The cDNA encoding batroxobin was cloned, expressed in Pichia pastoris and the molecular function of purified recombinant protein was also characterized. The recombinant batroxobin had an apparent molecular weight of 33 kDa by SDS-PAGE analysis and biochemical activities similar to those of native batroxobin. The purified recombinant protein strongly converted fibrinogen into fibrin clot in vitro, and shortened bleeding time and whole blood coagulation time in vivo. However, it did not make any considerable alterations on other blood coagulation factors. Several lines of experimental evidence in this study suggest that the recombinant batroxobin is a potent pro-coagulant agent. PMID:15280019

You, Weon-Kyoo; Choi, Won-Seok; Koh, You-Seok; Shin, Hang-Cheol; Jang, Yangsoo; Chung, Kwang-Hoe

2004-07-30

5

Optimization of expression of untagged and histidine-tagged human recombinant thrombin precursors in Escherichia coli.  

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The present study is focused on preparation of proper Escherichia coli expression system to ensure high yields of various modified precursors of human recombinant thrombin, a potential biopharmaceutical reagent. Two thrombin precursors, the smallest single-chain ?-thrombin precursor prethrombin-2 and its shortened form prethrombin-2?13, and their His-tagged forms were used. In order to determine the effect of the different lengths and amino acid compositions of affinity His-tag on the target protein expression level, a variety of the His-tag sequences were used. We found out that the protein expression efficiency was closely related to the codons used for encoding of amino acids of fusion histidine tag. Optimization of culture medium composition is another way to increase yield of the target protein. Suitable medium composition can ensure cell growth to high densities which is related to total yield of expressed protein. In this study, a new optimized complex medium for batch fermentation was developed. Addition of nutrients like a yeast extract and enzymatic casein hydrolysate to the defined medium components had a positive impact on protein expression, where relatively high expression level of the target protein from total amount of cellular proteins was achieved. Further, we have focused on trace element solution composition, and the optimized nickel and selenium concentrations were determined. Our results show that the composition of essential trace metal solution has a major impact not only on expression level, but it can also affect cell growth rate. PMID:24878753

Osadská, Michaela; Bo?ková, Hana; Krahulec, Ján; Stuchlík, Stanislav; Tur?a, Ján

2014-11-01

6

Optimization of the production of triabin, a novel thrombin inhibitor, in High Five™ insect cells infected with a recombinant baculovirus.  

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The isolation of a new type of thrombin inhibitor, called triabin, from the saliva of the hematophagous bug Triatoma pallidipennis, has recently been described. In the in vitro platelet aggregation inhibition assay triabin has a similar potency as the thrombin inhibitor hirudin now in phase III clinical trials. However, in another in vitro assay using a low molecular weight substrate for thrombin, triabin does not inhibit thrombin completely even at 6 fold higher molar doses in comparison with hirudin. This means that triabin has a novel mode of action towards thrombin making triabin into an interesting candidate as a therapeutic agent. Recently it has been shown that a recombinant baculovirus can be efficiently used for the triabin production in insect cells and that the yields in adherent cultures of High Five™ cells (approx. 20 mg l-1) were about 7 fold higher than in adherent cultures of Sf9 cells (approx. 3 mg l- 1). To optimize the triabin yield from the baculovirus/insect cell expression system, experiments were performed with suspension adapted cultures of High Five™ cells to investigate the effects of the state of the host cell, of the multiplicity of infection, of the cell density at the time of infection and of supplementation of the medium with nutrients and oxygen. Triabin yields of up to 200 mg l-1, as determined by an activity assay, could finally be obtained here. PMID:22359057

Vallazza, M; Petri, T

1999-03-01

7

Activation-induced exposure of the thrombin anion-binding exosite. Interactions of recombinant mutant prothrombins with thrombomodulin and a thrombin exosite-specific antibody.  

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The activation of serine protease zymogens involves conformational changes that increase the affinity of substrate binding and the activity of the catalytic center. The activation of prothrombin is particularly complex and requires several cleavages in the proenzyme region in addition to the conserved activation cleavage after Arg320. To understand how these cleavages lead to the exposure of the thrombin anion-binding exosite, a major macromolecular recognition site, interactions of recombinant human prothrombin derivatives with thrombomodulin, and an exosite-specific antibody were studied by competition binding and immunoprecipitation. By either method, the anion-binding exosite is not functional on prethrombin 2, which is cleaved after Arg271 and lacks fragment 1.2, nor on meizothrombin, which is cleaved only after Arg320. In contrast, the exosite is fully exposed on meizothrombin des-F1, which is cleaved after both Arg320 and Arg155 and therefore lacks amino-terminal fragment 1 (F1). Thus, two events are required to create the exosite. First, cleavage after Arg320 causes conformational changes that are much more extensive than those accompanying the activation of trypsinogen. Second, removal of amino-terminal F1 is necessary, perhaps to relieve steric hindrance. These results indicate that the F1 fragment regulates access to the thrombin exosite. The properties of meizothrombin des-F1 suggest that this prothrombin derivative could have a biological function. PMID:8106418

Wu, Q; Picard, V; Aiach, M; Sadler, J E

1994-02-01

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Purification of A Recombinant Thrombin-like Enzyme, Gloshedobin by Egg Yolk Antibody-Coupled Adsorbents  

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Full Text Available The gloshedobin, a snake venom thrombin-like enzyme was biosynthesized in the soluble form and purified by egg yolk antibody-coupled adsorbents from E. coli. As His-tag is not favored from the point of view of high-level and soluble expression, we herein constructed a recombinant gloshedobin without His-tag and developed a novel egg yolk antibody (IgY-immunoaffinity chromatography for its purification in a higher activity yield. The purification process involving Octyl Sepharose FF, IgY-immunoaffinity chromatography and Source Q, yielded 454.7U mg1 protein of interest and 34.8% activity recovery. The anti-gloshedobin IgY was obtained from eggs by injecting diluted gloshedobin into the breast muscle of laying hens and then purified by several steps including 3.5%, 8.5% and 12% polyethylene glycol-6000 precipitation and affinity chromatography using gloshedobin-coupled agarose gel. The obtained IgY was covalently linked to CNBr pre-activated agarose gel, Sepharose 4B FF, to yield immunoaffinity adsorbents. Both immunological and enzymatic activities of the purified enzyme were determined by western-blotting analysis and fibrinogen clotting assay, respectively.

Qing Yang

2005-01-01

9

Recombinant DNA Technology. A Topics Course for Undergraduates.  

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Describes the development of a topics course offered jointly by the chemistry and biology departments at Macalester College (Minnesota). Outlines the syllabus for the course. Discusses teaching and laboratory methods used. (CW)

Parson, Kathleen A.

1988-01-01

10

Topics in cosmology: Structure formation, dark energy and recombination  

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The field of theoretical cosmology consists of numerous, inter-related branches, whose ambitious goal is to uncover the history of the universe from its beginning to its future. Achieving this, no doubt, requires a deep understanding of many areas of physics. In this thesis I touch upon a few of these areas in which I worked during my PhD studies. Chapter (2) describes our work in finding the accretion and merger history of dark matter halos. Dark matter halos are the collapsed dark matter structures in the late time evolution of the universe, whose existence is vital for the formation of galaxies in the Universe as they act as the potential wells where normal matter (collectively called Baryons) can accumulate, cool, and form stars. It is then no surprise that the properties of galaxies depends on the properties of the dark matter halo in which it resides, including its merger history, i.e. the number of times it merged with other halos. Even though these merger rates can be calculated theoretically for infinitesimal time steps, in order to find the merger history over an extended period of time one had to use either Monte-Carlo simulations to build up the total rates of merging and accreting from the infinitesimal rates or use N-body simulations. In chapter (2) we show how we used random walk formalism to write down an analytical (integral) equation for the merger history of halos. We have solved this equation numerically and find very good agreement with Monte-Carlo simulations. This work can be used in theories of galaxy formation and evolution. We then switch from the overdense regions of the Universe, halos, to the underdense ones, voids. These structures have not attracted as much attention from cosmologists as their overdense counterparts in probing the cosmological models. We show here that the shapes of voids as a probe can be of use for future surveys to pin down the equation of state of the dark energy, i.e. the ratio of its pressure to its energy density. As first approximation, voids can be considered to be ellipsoids whose axis ratio evolution depends on the cosmological parameters. This, together with the fact that the initial distribution of the axis ratios is known (because the intial density field is Gaussian) can be used to infer the equation of state of the dark energy statistically from the observation of voids at different redshifts and with different sizes. The standard method of Fisher matrices is then used to forecast how well a future survey can measure the equation of state. We find promising results with constraints coming from void ellipticity measurements comparable to those of other standard methods. Chapter (4) goes farther back in the history of the Universe. During the recombination era, when the Universe was around a thousandth of its present size, it became cool enough that free electrons got captured by free protons to make hydrogen atoms. Consequently, the Thompson scattering of photons off of free electrons dropped dramatically and the Universe became transparent to photon propagation. The Cosmic Microwave Background (CMB) is a remnant from this epoch, consisting of photons last scattered off of a free electron. A wealth of information is contained in the statistical properties of the CMB field. However, in order to take full advantage of this probe one needs to know the recombination history, i.e. the evolution of the number density of free electrons as a function of time, to sub-percent level accuracy during this era. There are a plethora of phenomena, from radiative transfer effects to atomic and molecular ones, that have the potential to change the recombination history to this level. Our work was to calculate the effect that the formation of hydrogen molecules will have on the recombination history. Even though the abundance of hydrogen molecules is very small, they still have the potential to change the recombination history by reshuffling photons from the blue side of the Ly-alpha line to its red side and vise-versa. To find the magnitude of the effect, we solve the appropri

Alizadeh, Esfandiar

11

BLA 125248 Thrombin (Recombinant) Recothrom  

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... (~et Z(~II~Y~~·Cil:I.~::; M~\\lI~ PTA 11l1pld~i GLP IV. SC (~et ZGI i~~~~·h:;:1 :\\'cti: Oa' '.~~e of th~ ttllll ';rcp:c;!f' i:n.::udi~ dli 3pplic3tk.u ci rb. ...

12

Triabin, a highly potent exosite inhibitor of thrombin.  

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Triabin, a new thrombin inhibitor, has been purified from the saliva of Triatoma pallidipennis, a blood-sucking triatomine bug. It forms a noncovalent complex with thrombin at a molar ratio of 1:1, inhibits thrombin-induced platelet aggregation, and prolongs thrombin clotting time and activated partial thromboplastin time. However, it only minimally suppresses the amidolytic activity of thrombin, as measured by a chromogenic peptide substrate assay. It completely blocks trypsin-catalyzed cleavage of thrombin, probably via protection of the anion-binding exosite and inhibits the effect of thrombomodulin on thrombin in a dose-dependent fashion. These results indicate that the inhibitor is directed toward the anion-binding exosite of thrombin. The protein was partially sequenced and the information used to isolate cDNA clones from a T. pallidipennis salivary gland library. Four slightly polymorphic variants coding for mature proteins of 142 amino acids preceded by a putative leader sequence were obtained. The recombinant protein expressed in the periplasmic space of Escherichia coli has a biological activity similar to that of salivary triabin, as tested in a thrombin-induced platelet aggregation assay. In addition, recombinant triabin inhibits thrombin-catalyzed hydrolysis of fibrinogen with a Ki of about 3 pM. PMID:7499380

Noeske-Jungblut, C; Haendler, B; Donner, P; Alagon, A; Possani, L; Schleuning, W D

1995-12-01

13

Direct thrombin inhibitors.  

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Direct thrombin inhibitors interact with thrombin and block its catalytic activity on a wide range of substrates. Their action is in contrast to heparin and its derivatives, which inhibit thrombin and other coagulation serine proteases via antithrombin, and to the warfarin-type drugs that interfere with synthesis of the precursors of the coagulation serine proteases. There are three direct thrombin inhibitors approved for clinical use at present (lepirudin, bivalirudin, argatroban) and another in advanced clinical testing (melagatran/ximelagatran). The chemical structure, kinetics of thrombin inhibition, pharmacokinetics, and clinical use of each of these agents are discussed. PMID:12124681

Kaplan, Karen L; Francis, Charles W

2002-07-01

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Inhibition of thrombin-mediated cellular effects by triabin, a highly potent anion-binding exosite thrombin inhibitor.  

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Triabin, a 17 kDa protein from the saliva of the assassin bug Triatoma pallidipennis is a potent thrombin inhibitor interfering with the anion-binding exosite of the enzyme. The recombinant protein, produced by the baculovirus/insect cell system, was used to study the inhibitory effect on thrombin-mediated cellular responses. The thrombin (1 nM)-stimulated aggregation of washed human platelets and the rise in cytoplasmic calcium in platelets were inhibited by triabin at nanomolar concentrations. In contrast, the rise in calcium induced by the thrombin receptor-activating peptide (10 microM) was not suppressed by triabin. In isolated porcine pulmonary arteries, preconstricted with PGF 2 alpha thrombin (2 nM) elicited an endothelium-dependent relaxation which was inhibited by triabin in the same concentration range as found for the inhibition of platelet aggregation. Higher concentrations of triabin were required to diminish the contractile response of endotheliumdenuded pulmonary vessels to thrombin (10 nM). In cultured bovine coronary smooth muscle cells, the mitogenic activity of thrombin (3 nM), measured by [3H]thymidine incorporation, was also suppressed by triabin. In all these assays, the inhibitory effect of triabin was dependent on the thrombin concentration used. These studies suggest that the new anion-binding exosite thrombin inhibitor triabin is one of the most potent inhibitors of thrombin-mediated cellular effects. PMID:9241757

Glusa, E; Bretschneider, E; Daum, J; Noeske-Jungblut, C

1997-06-01

15

Whole blood thrombin: development of a process for intra-operative production of human thrombin.  

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Thrombin-based clotting agents currently used for topical hemostasis with absorbable sponges, fibrin sealants, and platelet gels are primarily derived from bovine or pooled human plasma sources. Autologous thrombin has important safety advantages in that it does not carry the same safety concerns as pooled plasma-derived products and it avoids exposure to risks associated with bovine-derived proteins. The goal of our research was to develop a rapid, reliable, and simple to perform process to generate autologous human thrombin in the intraoperative setting, from patient whole blood as the starting source material. Using whole blood instead of plasma as the starting material, it is possible to avoid the inherent delay in thrombin availability associated with a primary step of plasma isolation. In this study, we varied several key processing parameters to maximize thrombin production, reproducibility and stability. Autologous thrombin production was generated using a dedicated, single use disposable with a sterile reagent. The disposable consists of a tubular reaction chamber containing glass microsphere beads to activate the alternative pathway of the coagulation cascade. At the end of the process, thrombin-activated serum was harvested from the reaction chamber. The average activity of the thrombin produced at room temperature by this system was 82.8 +/- 15.9 IU/mL. The total processing time was room, has been developed. The device is simple to use, requires 30 minutes, and can consistently produce thrombin solutions that achieve rapid clotting of platelet concentrates, plasma, and fibrinogen concentrates even when mixed at thrombin to blood product ratios of 1:11. PMID:17486869

Kumar, Vijay; Chapman, John R

2007-03-01

16

Recombiner  

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Air containing hydrogen can be oxidized by heating in a container called a recombiner, in order to avoid the collection of hydrogen. The container is long and a large number of straight heating bars are arranged in parallel in it and they are flanged to a lid. The heating bars are surrounded by tubes, in order to obtain good heat transfer by a narrow annular gap. (orig.)

17

DABIGATRAN ETEXILATE: NEW DIRECT THROMBIN INHIBITORS ANTICOAGULANTS  

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Thrombin plays a key role in thrombotic events, and therefore thrombin inhibition represents a therapeutic target for numerous thromboembolic diseases. Thrombin is responsible for the conversion of soluble fibrinogen to fibrin; clot stabilization through activation of factor XIII and the formation of cross-linkage among fibrin molecules; and the generation of additional thrombin through activation of factors V, VIII, and XI. Direct thrombin inhibitors are an innovative class of anticoagulant...

Patel Kinjal B; Galani Varsha; Patel Paresh B; Mehta Hiren R

2011-01-01

18

Human thrombins. Production, evaluation, and properties of alpha-thrombin.  

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Human alpha-thrombin, the thromboplastin activation product of prothrombin with high clotting and esterase activity, was produced from Cohn Fraction III paste. The procedure started with 0.4 to 3.2 kg of frozen paste and was completed in 2 or 3 days. Some 23 g of thrombin were recorded for 65 quantitated preparations made from 11 lots of Fraction III paste. These preparations were obtained at protein concentrations of 3.9 +/- 1.3 mg/ml with a yield of 340 +/- 110 mg/kg of paste, which represented 48 +/- 14% of the clotting potential extracted as prothrombin. They had specific clotting activities of 2.8 +/- 0.4 U.S. (NIH) units/microng of protein and titrated to 88 +/- 8% active with p-nitrophenyl-p'-guanidinobenzoate (NPGB). Those (N - 29) examined by labeling with [14C]diisopropyl phosphorofluoridate (iPr2P-F) and electrophoresing in sodium dodecyl sulfate (SDS)-polyacrylamide gels were found to contain only (N = 4) or predominantly alpha-thrombin (97 +/- 3%) and corresponding amounts of ists degradation product, beta-thrombin (2.6 +/- 3.1%). No plasmin(ogen), prothrombin complex factors (II, VII, IX, IXalpha, X, Xalpha), or prothrombin fragments were detected in representative preparations. As produced in 0.75 M NaCl, pH approximately 6, thrombin was stable for approximately 1 week at 4 degrees and for greater than 1 year at less than or equal to 50 degrees; freeze-dried thrombin stored at 4 degrees for greater than 1 year displayed stable clotting activity and no vial to vial variation, permitting its use for reference purposes. Human thrombin generated by Taipan snake venom activation was compared with that produced by rapid thromboplastin activation: after treatment with [14C]iPr2P-F, greater than 95% of the label in both thrombins migrated at the same rate during electrophoresis in SDS; identical pairs of NH2-terminal residues were released in three consecutive Edman degradation cycles. PMID:16908

Fenton, J W; Fasco, M J; Stackrow, A B

1977-06-10

19

Effect of Electronic Polarization to Human ?-Thrombin  

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The polarized protein-specific charges (PPC) of human ?-thrombin (thrombin) and its inhibitor (L86) are made possible by employing the recently developed molecular fractionation with conjugate caps approach incorporated the Poisson—Boltzmann model. Molecular dynamics (MD) simulations of thrombin have been carried out to investigate the dynamics and stability of the thrombin-inhibitor using PPC and AMBER charges respectively. Detailed analysis and comparison of MD results show that the PPC can correctly describe the polarized state of the thrombin and L86. Especially, the root-mean-square deviation of backbone atoms and the hydrogen bonds using PPC are more stable than the AMBER charge. The present results indicate that protein polarization plays critical roles in maintaining the compact structure of thrombin.

Duan, Li-Li; Li, Zong-Chao; He, Xiang; Zhang, Qing-Gang

2014-04-01

20

Effect of non-heparin thrombin antagonists on thrombin generation, platelet function, and clot structure in whole blood.  

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Platelet contractile force (PCF), which is absent in blood obtained during cardiopulmonary bypass, significantly recovers after protamine sulfate administration. In vitro studies reveal this effect to be primarily caused by heparin. Because many of heparin's effects are mediated by suppression of thrombin generation and activity, this study assessed the influence of thrombin inhibition on PCF. The effects of natural and synthetic antithrombins were measured. Clots were formed by the addition of batroxobin (0.21 microg/mL) to whole blood (platelet count 200,000/microL). Force development was measured from the moment of batroxobin addition. After 1200 s of clotting, purified antithrombin III (22 microM) reduced PCF by 74%. Thrombomodulin (0.014 microM) reduced PCF by 60%. At 0.040 microM, PCF was reduced by 82% (6.5-1.2 Kdynes). Hirudin decreased PCF in a dose-dependent fashion, with complete suppression at concentrations > or = 0.30 microM. At concentrations between 0.04 and 0.29 microM, Lepirudin (Refludan, a recombinant therapeutic hirudin) produced dose-dependent delay and suppression of PCF. Above 0.29 microM Lepirudin, PCF was totally suppressed. At 1.60 microM, bivalirudin (a synthetic, 20 amino acid peptide) delayed and reduced PCF by 50%. At 6.40 micro;M, PCF was completely suppressed. Although 20 microM of P-PACK II (d-Phenylalanyl-L-Phenylalanylarginine- chloro-methyl ketone 2 HCl) had little effect, 40 microM delayed onset of force development from 300 to 600 s and reduced PCF at 1200 s from 5.2 to 3.3 Kdynes. At 120 microM, force development was totally suppressed. Four micromol Thromstop (BNas-Gly-(pAM)Phe-Pip) delayed force development by greater than 800 s and PCF at 1200 s was reduced by 70%. At 0.20 microM, Argatroban (a synthetic polypeptide direct thrombin antagonist) delayed onset of PCF from 300 to 540 s and decreased PCF by 40%. At a concentration of 0.40 microM and above, Argatroban totally suppressed PCF. These results indicate that some of the antiplatelet effects of heparin are the result of thrombin inhibition and that low-level thrombin generation is essential for clot retraction. The sensitivity of PCF to the presence of thrombin may permit monitoring of antithrombin agents via this assay. PMID:14515016

Carr, Marcus E; Angchaisuksiri, Pantep; Carr, Sheryl L; Martin, Erika J

2003-01-01

 
 
 
 
21

DABIGATRAN ETEXILATE: NEW DIRECT THROMBIN INHIBITORS ANTICOAGULANTS  

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Full Text Available Thrombin plays a key role in thrombotic events, and therefore thrombin inhibition represents a therapeutic target for numerous thromboembolic diseases. Thrombin is responsible for the conversion of soluble fibrinogen to fibrin; clot stabilization through activation of factor XIII and the formation of cross-linkage among fibrin molecules; and the generation of additional thrombin through activation of factors V, VIII, and XI. Direct thrombin inhibitors are an innovative class of anticoagulants that bind directly to thrombin to inhibit its actions and impede the clotting process. Dabigatran is the first direct thrombin inhibitor, orally available first approval by US Food and Drugs Administration in 2010. Specifically and reversibly inhibits thrombin, so the duration of action is predictable. The anticoagulant effect correlates well with plasma drug concentrations, which implies an effective anticoagulation with low bleeding risk without major problems of interactions with other drugs. The predictable pharmacokinetics and pharmacodynamics characteristics of dabigatran may facilitate dental management of patients who until now have been in treatment with traditional anticoagulants, given that it doesn’t require routine laboratory monitoring in the vast majority of patients treated. They also present a profile of drug interactions very favorable.

Patel Kinjal B

2011-04-01

22

Aptamer Based Microsphere Biosensor for Thrombin Detection  

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Full Text Available We have developed an optical microsphere resonator biosensor using aptamer asreceptor for the measurement of the important biomolecule thrombin. The sphere surface ismodified with anti-thrombin aptamer, which has excellent binding affinity and selectivityfor thrombin. Binding of the thrombin at the sphere surface is monitored by the spectralposition of the microsphere’s whispering gallery mode resonances. A detection limit on theorder of 1 NIH Unit/mL is demonstrated. Control experiments with non-aptameroligonucleotide and BSA are also carried out to confirm the specific binding betweenaptamer and thrombin. We expect that this demonstration will lead to the development ofhighly sensitive biomarker sensors based on aptamer with lower cost and higher throughputthan current technology.

Xudong Fan

2006-08-01

23

Thrombin generation test in microfluidic systems  

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The thrombin generation test is one of the diagnostic tests currently in use as a universal method for measuring hemostatic disorders. We envisioned that conventional monitoring of thrombin generation could be miniaturized resulting in a time-saving, accurate, easy-to-operate, and cost-efficient test. For the translation of the conventional thrombin generation test to microfluidic devices, our focus was directed to parameters such as the detection limit, temperature, protein-surface interactions (i.e., hydrophilicity of microchannels), and mixing behavior. Scaling down to microchannels (e.g., capillaries) resulted in volume reduction and allowed us to study the effect of a microchannel surface (either hydrophilic or hydrophobic) on the thrombin activity. Finally, the use of a micromixer enabled us to perform efficient on-chip mixing, resulting in the successful measurement of a thrombin generation in a microfluidic device.

Koch, Kaspar; van Berkel, Sander S.; van de Wal, Marloes M. E. B.; Nieuwland, Pieter J.; van Hest, Jan C. M.; Rutjes, Floris P. J. T.

2009-05-01

24

Systemic thrombin generation by glucose  

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Full Text Available Background: Systemic thrombin activity (F2a, i.e. thrombin protected and transported by a2- macroglobulin, is a new biomarker for the activation state of coagulation in vivo. F2a > 120% of normal diagnoses a pathologic disseminated intravascular coagulation (PIC in humans, either acute or chronic. Since glucose triggers intrinsic coagulation, the present work aimed to quantify systemic thrombin generation induced by glucose in vivo in mice. Material and Methods: Balb/c mice were i.p. injected with different concentrations of glucose (0 - 0.3 mmoles. After 0 - 3 h EDTA-blood was withdrawn, centrifuged, and the plasma was stabilized 1 + 1 with 2.5 M arginine, pH 8.6, and analyzed for systemically circulating F2a (that is F2a.?2M. The F2a.?2M activity in mice without glucose injection was defined as 100% of murine norm. Results: 1 h after i.p. injection 0.1 - 0.3 mmoles glucose resulted in about 1.4 fold increase of plasmatic glucose and in about 2.5 fold increase of systemic F2a activity. At the 45 min time interval between i.p. injection of 0.038 mmoles glucose and blood withdrawing an approximately 1.5fold increase of plasma glucose caused a 4fold increase in systemic F2a. Discussion: When systemic F2a reaches 120% of the normal, the normal human intravascular coagulation (NIC turns to the pre-phase of pathologic plasmatic intravascular coagulation (PIC-0 also defined as pre-PIC. At 150% systemic F2a, the PIC-0 changes to PIC-1 which is the common pathologic plasmatic intravascular coagulation (typical PIC. At 200% systemic F2a, PIC-1 changes to PIC-2 (consumption PIC. The present assay technique seems to be suitable in judging the coagulation activation state of any mammalian blood. Diabetic patients should be monitored for the new biomarker systemic F2a similarly as for the old biomarker glycated hemoglobin (HbA1c. The target systemic F2a range should be NIC, preferably around 100% of normal.

Hani Harb

2012-02-01

25

The unresolved safety concerns of bovine thrombin  

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Full Text Available Abstract A recent review has suggested that bovine thrombin is not associated with an increased risk of bleeding in surgical populations. In spite of extremely limited evidence available, many valuable resources (e.g. safety surveillance and post-marketing programs, case reports were excluded in reaching this conclusion. While waiting for the adequately powered, controlled clinical trials to address the effects of bovine thrombin on bleeding and thrombotic events, the potential risk cannot be simply ignored. Rather, continued vigilance in the post-surgical setting for bleeding events that may be associated with the development of acquired coagulation factor inhibitors following bovine thrombin administration is warranted.

Javidroozi Mazyar

2008-09-01

26

PROTON BRIDGING IN THE INTERACTIONS OF THROMBIN WITH SMALL INHIBITORS†  

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Thrombin is the pivotal serine protease enzyme in the blood cascade system. Phe-Pro-Arg-chloromethylketone (PPACK), phosphate and phosphonate ester inhibitors form a covalent bond with the active-site Ser of thrombin. PPACK, a mechanism-based inhibitor, and the phosphate/phosphonate esters form adducts that mimic intermediates formed in reactions catalyzed by thrombin. Therefore, the dependence of the inhibition of human ?-thrombin on the concentration of these inhibitors, pH, and temperatur...

Kovach, Ildiko M.; Kelley, Paul; Eddy, Carol; Jordan, Frank; Baykal, Ahmet

2009-01-01

27

On the mechanism(s) of thrombin induced angiogenesis.  

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Promotion of tumour progression by thrombin is suggested by several clinical and laboratory observations. A plausible explanation for this effect of thrombin may be related to our previous findings that thrombin is a potent promoter of angiogenesis in the chick chorioallantoic membrane system (CAM) and in the Matrigel system in vivo. In this report we summarise the cellular and molecular actions of thrombin that could be contributing to the activation of angiogenic cascade. Treatment of endothelial cells with thrombin leads to activation of gelatinase A, which may allow for local dissolution of basement membrane, an essential first step of angiogenesis. Similarly thrombin-treated endothelial cells have diminished ability to adhere to collagen type IV and laminin. This new phenotype of endothelial cells can migrate and survive without attachment to extracellular matrix. Thrombin-treatment of endothelial cells increases the vectorial secretion of extracellular matrix proteins, a process essential at the final steps of angiogenesis. In addition, thrombin potentiates the VEGF-induced mitogenesis of endothelial cells. This can be explained by the upregulation of the VEGF receptors (KDR & flt-1) by thrombin treatment. All the aforementioned effects of thrombin are receptor mediated, dose-dependent and require only brief exposure of endothelial cells to thrombin for these actions of thrombin. The transduction mechanisms involved are via protein kinase C (PKC) and MAP-kinase pathways. PMID:10949654

Maragoudakis, M E; Tsopanoglou, N E

2000-01-01

28

Vesicular aptasensor for the detection of thrombin.  

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Self-assembled phospholipid vesicles are functionalized with thrombin-binding aptamers using a thiol-click reaction. The resulting aptasensors signal the binding of the analyte to the vesicle surface by changes of the emission properties of membrane co-embedded reporter dyes. PMID:25205174

Müller, Andreas; König, Burkhard

2014-10-28

29

Thrombin in myocardial ischemia-reperfusion during cardiac surgery.  

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Thrombin is a multifunctional protease with procoagulant, pro-inflammatory, and pro-apoptotic effects. Thrombin has direct potentially adverse effects on the endothelium and on cardiomyocytes, which are independent of its procoagulant effects, and it has emerged as a possible mediator of ischemia-reperfusion injury. Several lines of experimental evidence specifically implicate thrombin to be involved in myocardial ischemia-reperfusion injury. Cardiopulmonary bypass increases thrombin generation progressively, but reperfusion after myocardial ischemia induces an additional distinct and rapid increase in thrombin generation. Clinical studies have shown that thrombin formation during cardiac surgery, especially during myocardial reperfusion, is involved with myocardial damage and impaired hemodynamic recovery. Therefore, strategies to improve thrombin control during cardiopulmonary bypass might be beneficial. PMID:19559265

Raivio, Peter; Lassila, Riitta; Petäjä, Jari

2009-07-01

30

Thrombin regulates the function of human blood dendritic cells  

International Nuclear Information System (INIS)

Thrombin is the key enzyme in the coagulation cascade and activates endothelial cells, neutrophils and monocytes via protease-activated receptors (PARs). At the inflammatory site, immune cells have an opportunity to encounter thrombin. However little is known about the effect of thrombin for dendritic cells (DC), which are efficient antigen-presenting cells and play important roles in initiating and regulating immune responses. The present study revealed that thrombin has the ability to stimulate blood DC. Plasmacytoid DC (PDC) and myeloid DC (MDC) isolated from PBMC expressed PAR-1 and released MCP-1, IL-10, and IL-12 after thrombin stimulation. Unlike blood DC, monocyte-derived DC (MoDC), differentiated in vitro did not express PAR-1 and were unresponsive to thrombin. Effects of thrombin on blood DC were significantly diminished by the addition of anti-PAR-1 Ab or hirudin, serine protease inhibitor. Moreover, thrombin induced HLA-DR and CD86 expression on DC and the thrombin-treated DC induced allogenic T cell proliferation. These findings indicate that thrombin plays a role in the regulation of blood DC functions

31

Calibrated automated thrombin generation in frozen-thawed platelet-rich plasma to detect hypercoagulability.  

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To enhance the practical applicability of the calibrated automated thrombogram (CAT) we investigated whether frozen-thawed platelet-rich plasma (ft-PRP) can be used to assess the function of the protein C inhibitory pathway, while preserving the natural phospholipid composition. Recalcified ft-PRP triggered with 0.5 pM recombinant human tissue factor shows a median thrombin potential of 1,779 nM x min, against 1,576 nM x min for fresh PRP. To obtain approximately 70% inhibition, 6.7 nM activated protein C (APC) has to be added, instead of 25 nM in fresh PRP; so the relative APC resistance of PRP appears to depend upon the presence of intact platelets. Factor VIII, added to normal ft-PRP to obtain a concentration of 3.3 U/ml, increases the thrombin potential in the presence of APC 1.5-fold, from 524 to 808 nM x min, in keeping with previously published increases in thrombotic risk in patients with high factor VIII levels. We conclude that thrombography in ft-PRP, with and without added APC, can be used to assess known risk factors for thrombosis, which allows the design of large clinical studies aimed at proving the relationship between thrombin potential and clinical outcome. PMID:12853709

Regnault, Véronique; Béguin, Suzette; Lecompte, Thomas

2003-01-01

32

Thrombin inhibits osteoclast differentiation through a non-proteolytic mechanism.  

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Thrombin stimulates expression of interleukin 6 and cyclooxygenase 2 by osteoblasts, both of which enhance osteoblast-mediated osteoclast differentiation by increasing the ratio of receptor activator of nuclear factor ?B ligand (RANKL) expression to that of osteoprotegerin (OPG) in osteoblasts. We hypothesised that thrombin would also increase this ratio and thereby stimulate osteoclast differentiation in mixed cultures of osteoblastic cells and osteoclast precursors. In primary mouse osteoblasts, but not in bone marrow stromal cells, thrombin increased the ratio of RANKL to OPG expression. Thrombin inhibited differentiation of osteoclasts, defined as tartrate-resistant acid phosphatase (TRAP)-positive cells with three or more nuclei, in mouse bone marrow cultures treated with osteoclastogenic hormones; this effect was not mediated by the major thrombin receptor, protease-activated receptor 1, nor did it require thrombin's proteolytic activity. Thrombin also caused a decrease in the number of TRAP-positive cells with fewer than three nuclei. Thrombin (active or inactive) also inhibited osteoclast differentiation and bone resorption, respectively, in cultures of mouse spleen cells and human peripheral blood mononuclear cells induced to undergo osteoclastogenesis by treatment with RANKL and macrophage colony-stimulating factor. Osteoclast differentiation in spleen cells was inhibited when they were exposed to thrombin from days 0 to 3 or 3 to 5 of culture but not days 5 to 7 when most fusion occurred. Thrombin inhibited expression of RANK by spleen cells. These observations indicate that, although thrombin stimulates production of osteoclastogenic factors by osteoblastic cells, it inhibits the early stages of RANKL-induced osteoclast differentiation through a direct effect on osteoclast precursors that does not require thrombin's proteolytic activity. PMID:23419317

Sivagurunathan, S; Pagel, C N; Loh, L H; Wijeyewickrema, L C; Pike, R N; Mackie, E J

2013-06-01

33

Quantitative detection of thrombin activity in an ischemic stroke model.  

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Thrombin, a central factor in thrombogenesis, affects cells in the brain through protease activated receptors. Low levels of thrombin activity are neuroprotective while higher levels are deleterious, and we have therefore developed a new method for its direct quantitative measurement in brain slices following stroke. Thrombin activity was measured by a fluorescent substrate on fresh coronal slices taken from the ipsilateral and contralateral hemispheres 24-72 h following permanent right middle cerebral artery occlusion. Prolyl endopeptidase and aminopeptidases were inhibited as a critical step to insure the specificity of the assay for thrombin detection. Infarct volume was assessed using TTC staining. Thrombin activity in the right ischemic hemisphere was significantly higher compared to the contralateral hemisphere (32?±?6 and 27?±?10 mU/ml, mean?±?SE in the two most affected slices from the ischemic hemisphere vs. 21?±?6 and 8?±?2 mU/ml in corresponding contralateral slices; p?Thrombin levels in the ischemic and contralateral hemispheres were significantly higher compared to healthy control mice and were above the range known to be protective to brain cells. A significant correlation was found between thrombin activity in the ischemic hemisphere and the infarct volume. Results of studies based on this method may translate into potential thrombin based therapies. PMID:23900720

Bushi, Doron; Chapman, Joab; Katzav, Aviva; Shavit-Stein, Efrat; Molshatzki, Noa; Maggio, Nicola; Tanne, David

2013-11-01

34

Hypersensitivity to thrombin of platelets from hypercholesterolemic rats  

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Hypersensitivity of platelets to thrombin has been associated with hypercholesterolemia. The authors have examined the mechanisms involved in this hypersensitivity. Rats were given diets rich in milk fat and containing added cholesterol and taurocholate to produce hypercholesterolemia (HC) (262 +/- 25 mg%) or added sitosterol as a normocholesterolemic control (NC) (89 +/- 6 mg%). Washed platelets were prelabelled with 14C-serotonin. In the presence of acetylsalicyclic acid (ASA) (to inhibit thromboxane A2 (TXA2) formation) and creatine phosphate/creatine phosphokinase (CP/CPK) (to remove released ADP), HC platelets aggregated more (26 +/- 1%) and released more 14C (9.1 +/- 2.0%) than NC platelets (aggregation: 0%, p 14C release: 1.5 +/- 0.5%, p 2 formation is involved in the hypersensitivity of HC platelets to thrombin. Total binding of 125I-thrombin to HC platelets was less than that to NC platelets but HC platelets were smaller and had less protein than NC platelets; the thrombin binding per mg platelet protein was the same for HC and NC platelets, indicating that hypersensitivity to thrombin of HC platelets does not result from increased thrombin binding. Thus, hypersensitivity of HC platelets to thrombin is not due to TXA2 formation, the action of released ADP or increased thrombin binding

35

Bexarotene Topical  

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Targretin® Topical Gel ... Topical bexarotene is used to treat cutaneous T-cell lymphoma (CTCL, a type of skin cancer) that ... Topical bexarotene comes as a gel to apply to the skin. It is usually applied once every ...

36

Thrombin domains: structure, function and interaction with platelet receptors.  

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Thrombin plays a pivotal role in different biological phenomena, such as hemostasis, thrombosis, and cell differentiation. Indeed this protease catalyzes the conversion of fibrinogen into fibrin, the activation of coagulation factors V, VIII, XI, and XIII, but is also involved in the activation of many cell types and platelets. Thrombin bears some recognition domains and insertion loops, not found among other serine proteases of the coagulation system. In this review the properties of these thrombin domains, which regulate the specificity of the enzyme's interaction with substrates and inhibitors, are particularly emphasized. The example of thrombin interaction with the platelet membrane receptors, namely GpIb and PAR1, shows how the concerted action of the insertion loops and recognition domains is the key to solve the apparent enigma as to how thrombin can be at the same time a very efficient and specific enzyme for different substrates and inhibitors. PMID:14739624

De Cristofaro, Raimondo; De Candia, Erica

2003-06-01

37

Mechanisms of platelet activation by thrombin: a short history.  

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Platelet activation by thrombin is relevant to arterial thrombosis, therefore it is an attractive target for the development of new antithrombotic drugs. In the 1970s the platelet membrane complex glycoprotein (GP) Ib-V-IX was shown to have a high affinity binding site for thrombin on GPIb? and a substrate cleaved by thrombin, GPV. For several years it was considered to be involved in platelet activation by thrombin. The discovery of the protease activated receptors (PARs) in 1991 was a major breakthrough in the field. The first member of this family of receptors to be discovered was PAR1, a seven transmembrane G-protein coupled receptor which, upon cleavage by thrombin, unmasks a new amino-terminus able to bind intramolecularly to PAR1 itself thus inducing signaling. On human platelets PAR1 and, later PAR4, were demonstrated to mediate most of the platelet responses to thrombin. However, after the discovery of PARs, different groups demonstrated that GPIb? is required to stimulate a full platelet activation by thrombin. A model where thrombin binds to the GPIb receptor prior to proteolysis of the PAR receptors was supported by several lines of evidence. A role for GPV as inhibitor of GPIb? signaling has been shown by using GPV knock-out mice. Crystallographic data suggested that thrombin bound to GPIb? might be able to interact with other GPIb? molecules on the same or other platelets, shedding light on a new role for thrombin binding to GPIb?. Finally, anti-PAR1 molecules were developed which are now in phase II and III clinical studies as antithrombotic drugs. PMID:22137742

De Candia, Erica

2012-03-01

38

Planar Hall magnetoresistive aptasensor for thrombin detection.  

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The use of aptamer-based assays is an emerging and attractive approach in disease research and clinical diagnostics. A sensitive aptamer-based sandwich-type sensor is presented to detect human thrombin using a planar Hall magnetoresistive (PHR) sensor in cooperation with superparamagnetic labels. A PHR sensor has the great advantages of a high signal-to-noise ratio, a small offset voltage and linear response in the low-field region, allowing it to act as a high-resolution biosensor. In the system presented here, the sensor has an active area of 50 µm × 50 µm with a 10-nm gold layer deposited onto the sensor surface prior to the binding of thiolated DNA primary aptamer. A polydimethylsiloxane well of 600-µm radius and 1-mm height was prepared around the sensor surface to maintain the same specific area and volume for each sensor. The sensor response was traced in real time upon the addition of streptavidin-functionalized magnetic labels on the sensor. A linear response to the thrombin concentration in the range of 86 pM-8.6 µM and a lower detection limit down to 86 pM was achieved by the proposed present method with a sample volume consumption of 2 µl. The proposed aptasensor has a strong potential for application in clinical diagnosis. PMID:24727201

Sinha, B; Ramulu, T S; Kim, K W; Venu, R; Lee, J J; Kim, C G

2014-09-15

39

APTAMER-BASED SERRS SENSOR FOR THROMBIN DETECTION  

Energy Technology Data Exchange (ETDEWEB)

We describe an aptamer-based Surface Enhanced Resonance Raman Scattering (SERRS) sensor with high sensitivity, specificity, and stability for the detection of a coagulation protein, human a-thrombin. The sensor achieves high sensitivity and a limit of detection of 100 pM by monitoring the SERRS signal change upon the single step of thrombin binding to immobilized thrombin binding aptamer. The selectivity of the sensor is demonstrated by the specific discrimination of thrombin from other protein analytes. The specific recognition and binding of thrombin by the thrombin binding aptamer is essential to the mechanism of the aptamer-based sensor, as shown through measurements using negative control oligonucleotides. In addition, the sensor can detect 1 nM thrombin in the presence of complex biofluids, such as 10% fetal calf serum, demonstrating that the immobilized, 5{prime}-capped, 3{prime}-capped aptamer is sufficiently robust for clinical diagnostic applications. Furthermore, the proposed sensor may be implemented for multiplexed detection using different aptamer-Raman probe complexes.

Cho, H; Baker, B R; Wachsmann-Hogiu, S; Pagba, C V; Laurence, T A; Lane, S M; Lee, L P; Tok, J B

2008-07-02

40

Label-free impedimetric biosensor for thrombin using the thrombin-binding aptamer as receptor  

International Nuclear Information System (INIS)

This study presents the further establishment of impedimetric biosensors with aptamers as receptors. Aptamers are short single-stranded oligonucleotides which bind analytes with a specific region of their 3D structure. Electrical impedance spectroscopy is a sensitive method for analyzing changes on the electrode surface, e.g. caused by receptor-ligand-interactions. Fast and inexpensive prototyping of electrodes on the basis of commercially available compact discs having a 24 carat gold reflective layer was investigated. Electrode structures (CDtrodes [1]) in the range from few millimetres down to 100 microns were realized. The well-studied thrombin-binding aptamer (TBA) was used as receptor for characterizing these micro- and macro-electrodes. The impedance signal showed a linear correlation for concentrations of thrombin between 1.0 nM to 100 nM. This range corresponds well with most of the references and may be useful for the point-of-care testing (POCT).

 
 
 
 
41

Dynamic changes in thrombin generation in abdominal sepsis in mice.  

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Systemic inflammatory response syndrome and severe infections are associated with major derangements in the coagulation system. The purpose of this study was to examine the dynamic alterations in thrombin generation in abdominal sepsis. Abdominal sepsis was induced by cecal ligation and puncture (CLP) in C57/Bl6 mice. Cecal ligation and puncture caused a systemic inflammatory response, with neutrophil recruitment and tissue damage in the lung as well as thrombocytopenia and leukocytopenia. Thrombin generation, coagulation factors, lung histology, and myeloperoxidase activity was determined 1, 3, 6, and 24 h after induction of CLP. It was found that thrombin generation was increased 1 h after CLP and that thrombin generation started to decrease at 3 h and was markedly reduced 6 and 24 h after CLP induction. Platelet-poor plasma from healthy mice could completely reverse the inhibitory effect of CLP on thrombin generation, suggesting that sepsis caused a decrease in the levels of plasma factors regulating thrombin generation in septic animals. Indeed, it was found that CLP markedly decreased plasma levels of prothrombin, factor V, and factor X at 6 and 24 h. Moreover, we observed that CLP increased plasma levels of activated protein C at 6 h, which returned to baseline levels 24 h after CLP induction. Finally, pretreatment with imipenem/cilastatin attenuated the CLP-evoked decrease in thrombin generation and consumption of prothrombin 24 h after CLP induction. Our novel findings suggest that thrombin generation is initially increased and later decreased in abdominal sepsis. Sepsis-induced reduction in thrombin generation is correlated to changes in the plasma levels of coagulation factors and activated protein C. These findings help explain the dynamic changes in global hemostasis in abdominal sepsis. PMID:24978891

Wang, Yongzhi; O Braun, Oscar; Zhang, Su; Luo, Lingtao; Norström, Eva; Thorlacius, Henrik

2014-10-01

42

Thrombin inhibitors from the freshwater cyanobacterium Anabaena compacta.  

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Bioassay-guided investigation of the cyanobacterium Anabaena compacta extracts afforded spumigin J (1) and the known thrombin inhibitor spumigin A (2). The absolute configuration of 1 was analyzed by advanced Marfey's methodology. Compounds 1 and 2 inhibited thrombin with EC(50) values of 4.9 and 2.1 ?M, and 0.7 and 0.2 ?M in the cathepsin B inhibitory assay, respectively. The MM-GBSA methodology predicted spumigin A with 2S-4-methylproline as the better thrombin inhibitor. PMID:22950366

Anas, Andrea Roxanne J; Kisugi, Takaya; Umezawa, Taiki; Matsuda, Fuyuhiko; Campitelli, Marc R; Quinn, Ronald J; Okino, Tatsufumi

2012-09-28

43

Three Decades of Recombinant DNA.  

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Discusses highlights in the development of genetic engineering, examining techniques with recombinant DNA, legal and ethical issues, GenBank (a national database of nucleic acid sequences), and other topics. (JN)

Palmer, Jackie

1985-01-01

44

A thrombin receptor function for platelet glycoprotein Ib–IX unmasked by cleavage of glycoprotein V  

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Glycoprotein (GP) V is a major substrate cleaved by the protease thrombin during thrombin-induced platelet activation. Previous analysis of platelets from GP V-null mice suggested a role for GP V as a negative modulator of platelet activation by thrombin. We now report the mechanism by which thrombin activates GP V ?/? platelets. We show that proteolytically inactive forms of thrombin induce robust stimulatory responses in GP V null mouse platelets, via the platele...

Ramakrishnan, Vanitha; Deguzman, Francis; Bao, Ming; Hall, Scott W.; Leung, Lawrence L.; Phillips, David R.

2001-01-01

45

The effect of thrombin on a 6-hydroxydopamine model of Parkinson's disease depends on timing  

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Recent results in animal models suggest that thrombin may modulate brain injury in Parkinson's disease (PD). High doses of thrombin (?20 U) can damage dopaminergic neurons, while we have found that low dose thrombin (1 U), given several days before a brain insult (thrombin preconditioning), is protective in models of PD and stroke. However, the effects of such low levels of thrombin at the time of, or after, exposure to the dopamine neurotoxin 6-hydroxydopamine (6-OHDA) have not been examin...

Cannon, Jason R.; Hua, Ya; Richardson, Rudy J.; Xi, Guohua; Keep, Richard F.; Schallert, Timothy

2007-01-01

46

Fibrinogen and thrombin concentrations are critical for fibrin glue adherence in rat high-risk colon anastomoses  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english OBJECTIVE: Fibrin glues have not been consistently successful in preventing the dehiscence of high-risk colonic anastomoses. Fibrinogen and thrombin concentrations in glues determine their ability to function as sealants, healers, and/or adhesives. The objective of the current study was to compare [...] the effects of different concentrations of fibrinogen and thrombin on bursting pressure, leaks, dehiscence, and morphology of high-risk ischemic colonic anastomoses using fibrin glue in rats. METHODS: Colonic anastomoses in adult female Sprague-Dawley rats (weight, 250-350 g) treated with fibrin glue containing different concentrations of fibrinogen and thrombin were evaluated at post-operative day 5. The interventions were low-risk (normal) or high-risk (ischemic) end-to-end colonic anastomoses using polypropylene sutures and topical application of fibrinogen at high (120 mg/mL) or low (40 mg/mL) concentrations and thrombin at high (1000 IU/mL) or low (500 IU/mL) concentrations. RESULTS: Ischemia alone, anastomosis alone, or both together reduced the bursting pressure. Glues containing a low fibrinogen concentration improved this parameter in all cases. High thrombin in combination with low fibrinogen also improved adherence exclusively in low-risk anastomoses. No differences were detected with respect to macroscopic parameters, histopathology, or hydroxyproline content at 5 days post-anastomosis. CONCLUSIONS: Fibrin glue with a low fibrinogen content normalizes the bursting pressure of high-risk ischemic left-colon anastomoses in rats at day 5 after surgery.

Eliseo Portilla-de, Buen; Abel, Orozco-Mosqueda; Caridad, Leal-Cortés; Gonzalo, Vázquez-Camacho; Clotilde, Fuentes-Orozco; Andrea Socorro, Alvarez-Villaseñor; Michel Dassaejv, Macías-Amezcua; Alejandro, González-Ojeda.

2014-04-01

47

Oxybutynin Topical  

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Oxybutynin topical gel is used to treat overactive bladder (a condition in which the bladder muscles contract uncontrollably and ... Topical oxybutynin comes as a gel to apply to the skin. It is usually applied once a ...

48

Tretinoin Topical  

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Tretinoin comes in topical liquid, cream, and gel. Tretinoin usually is used daily at bedtime or once every 2 or 3 days. Follow ... nonmedicated cosmetics on cleansed skin. Do not use topical preparations with a lot of alcohol, menthol, spices, ...

49

Fluorouracil Topical  

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Fluorouracil cream and topical solution are used to treat actinic or solar keratoses (scaly or crusted lesions [skin areas] caused by years of too much exposure to sunlight). Fluorouracil cream and topical solution are also used to treat a type ...

50

Bimatoprost Topical  

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Topical bimatoprost is used to treat hypotrichosis (less than the normal amount of hair) of the eyelashes ... the growth of longer, thicker, and darker lashes. Topical bimatoprost is in a class of medications called ...

51

Topical anesthesia.  

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OBJECTIVE: To consider topical anesthetic options available to primary care physicians, indications for their use, and efficacy and safety of these agents as supported by the literature. QUALITY OF EVIDENCE: Five randomized controlled trials were retrieved that compared various topical anesthetics as well as topical anesthetics versus infiltrative anesthesia. MAIN FINDINGS: A combination of lidocaine, epinephrine, and tetracaine (LET) is currently the topical anesthetic of choice for repair o...

Keyes, P. D.; Tallon, J. M.; Rizos, J.

1998-01-01

52

Cellular signaling events involved in thrombin activation of vascular endothelium  

International Nuclear Information System (INIS)

Although cytosolic calcium ([Ca])/sub I/), inositol trisphosphate (IP3) and diacylglycerol (DAG) are important second messengers involved in stimulus-response coupling to certain hormones, little information is available regarding their role in the activation of vascular endothelial cells (EC) during coagulation. The authors have used cultured human umbilical vein EC to examine thrombin effects on [Ca]/sub I/ and on IP3 and DAG formation. Thrombin (1 U/ml) induces a rapid (peak 2+ indicator)-loaded EC. In addition thrombin stimulates concentration-dependent (.001-1 U/ml) increases in calcium efflux and IP3 formation in EC prelabeled with 45Ca or 3H-myoinositol. Peak IP3 (182+/-14% control) is rapid (3H-arachidonic acid-labeled EC, thrombin increases DAG (protein kinase C activator) at 15 sec (133+/-8% control), as well as 5 min (148+/-12% control). EC preincubation with 4?-phorbol 12-myristate 13-acetate (10-7M, 5 min), a potent activator of protein kinase C, blocks thrombin (1 U/ml)-induced increases in [Ca]/sub I/ and IP3. Thus, thrombin may trigger at least two distinct signaling pathways (IP3/calcium; DAG/protein kinase C) in EC. In addition, DAG stimulation of protein kinase C may influence this mediator's action in vascular EC

53

Calibrated automated thrombin generation in normal uncomplicated pregnancy.  

Science.gov (United States)

Pregnancy is associated with substantial changes in the haemostatic system and a six-fold higher incidence of venous thromboembolism. Conventional global tests, such as prothrombin time and activated partial thromboplastin time, do not definitely detect this hypercoagulable condition. We investigated whether the changes in haemostatic system during pregnancy are reflected in the calibrated automated thrombography (CAT). Thrombin generation was measured in platelet-poor plasma (PPP) of 150 healthy pregnant women without any pregnancy associated diseases by means of CAT. In addition, prothrombin (FII), antithrombin (AT), protein S, protein C, tissue factor pathway inhibitor (TFPI), plasminogen activator inhibitor-1 (PAI-1), thrombin-antithrombin complex (TAT), and prothrombin fragments 1+2 (F1+2) were measured. Endogenous thrombin potential (ETP) and peak of thrombin generation increased significantly with gestational weeks, while lag time and time to peak remained unchanged. A significant increase of PAI-1, TFPI, F1+2 and TAT as well as a significant decrease of free protein S, protein S antigen, and protein S activity was observed. Levels of AT and protein C remained stable during pregnancy. Division of population in trimester of pregnancy and analysis of differences between the trimesters showed rather similar results. Our study shows that endogenous thrombin potential does increase with duration of normal uncomplicated pregnancy. Whether parameters of continuous thrombin generation will correlate with thrombembolic disease remains to be shown. PMID:18278182

Rosenkranz, Andrea; Hiden, Michael; Leschnik, Bettina; Weiss, Eva-Christine; Schlembach, Dietmar; Lang, Uwe; Gallistl, Siegfried; Muntean, Wolfgang

2008-02-01

54

Activated protein C attenuates coagulation-associated over-expression of fibrinolytic activity by suppressing the thrombin-dependent inactivation of PAI-1.  

Science.gov (United States)

Several activated coagulation factors have been reported to enhance fibrinolysis by neutralizing plasminogen activator inhibitor type 1 (PAI-1) activity. We evaluated the physiological relevance of this mechanism using the euglobulin clot lysis time (ECLT) assay in the presence and absence of Ca2+, which is controlled by PAI-1 and mimics physiological thrombolysis. We found that the ECLT (18.5 +/- 0.6 h) was shortened by Ca2+ (5 mm) (6.6 +/- 0.1 h). A significant difference was observed in thrombin generation by the presence of Ca2+ in the euglobulin fraction. Prothrombin was almost fully converted to thrombin within 15 min in the presence of Ca2+, whereas essentially no conversion was observed without Ca2+. The presence of activated protein C (aPC) suppressed thrombin generation, and attenuated the shortening of ECLT in a dose-dependent manner, an effect enhanced by phospholipid and protein S. In the absence of Ca2+, aPC did not prolong the ECLT. After addition of biotin-labeled recombinant PAI-1 to the euglobulin fraction, PAI-1 was cleaved to lower molecular weight forms only in the presence of Ca2+. This cleavage did not occur in the presence of aPC, suggesting that thrombin was the catalyst for PAI-1 cleavage. The cleavage and inactivation of PAI-1 by generated thrombin is proposed to be responsible for the shortening of ECLT by Ca2+ and for coagulation-associated over-expression of fibrinolysis. Under such conditions, aPC appeared to suppress thrombin generation and to normalize highly activated fibrinolysis. PMID:14675098

Urano, T; Castellino, F J; Ihara, H; Suzuki, Y; Ohta, M; Suzuki, K; Mogami, H

2003-12-01

55

Sensitive and selective detection of thrombin by using a cyclic peptide as affinity ligand.  

Science.gov (United States)

Here we describe a sensitive assay for thrombin by using a high binding-affinity cyclic peptide against thrombin as affinity ligand. The cyclic peptide is immobilized on the magnetic beads or microplates to selectively capture thrombin. The enriched thrombin then catalyzes the cleavage of a substrate of thrombin to a detectable product. The detection of thrombin is finally achieved by measuring the generated product. This assay enables the detection of thrombin at tens fM in 100 µL of sample solution when fluorogenic substrate was applied, while detection limits reached pM level when chromogenic substrate was used. Thrombin in plasma sample can be detected with this assay. This cyclic peptide affinity ligand shows potentials for thrombin analysis in other detection formats. PMID:25048449

Zhao, Qiang; Gao, Jie

2015-01-15

56

Biochemical characterization of bovine plasma thrombin-activatable fibrinolysis inhibitor (TAFI)  

DEFF Research Database (Denmark)

BACKGROUND: TAFI is a plasma protein assumed to be an important link between coagulation and fibrinolysis. The three-dimensional crystal structures of authentic mature bovine TAFI (TAFIa) in complex with tick carboxypeptidase inhibitor, authentic full lenght bovine plasma thrombin-activatable fibrinolysis inhibitor (TAFI), and recombinant human TAFI have recently been solved. In light of these recent advances, we have characterized authentic bovine TAFI biochemically and compared it to human TAFI. RESULTS: The four N-linked glycosylation sequons within the activation peptide were all occupied in bovine TAFI, similar to human TAFI, while the sequon located within the enzyme moiety of the bovine protein was non-glycosylated. The enzymatic stability and the kinetic constants of TAFIa differed somewhat between the two proteins, as did the isoelectric point of TAFI, but not TAFIa. Equivalent to human TAFI, bovine TAFI was a substrate for transglutaminases and could be proteolytically cleaved by trypsin or thrombin/solulin complex, although small differences in the fragmentation patterns were observed. Furthermore, bovine TAFI exhibited intrinsic activity and TAFIa attenuated tPA-mediated fibrinolysis similar to the human protein. CONCLUSION: The findings presented here suggest that the properties of these two orthologous proteins are similar and that conclusions reached using the bovine TAFI may be extrapolated to the human protein.

Thaysen-Andersen, Morten; HØjrup, Peter

2009-01-01

57

Thrombin, a mediator of cerebrovascular inflammation in AD and hypoxia  

Directory of Open Access Journals (Sweden)

Full Text Available Considerable evidence implicates hypoxia and vascular inflammation in Alzheimer’s disease (AD. Thrombin, a multifunctional inflammatory mediator, is demonstrable in the brains of AD patients both in the vessel walls and senile plaques. Hypoxia-inducible factor 1? (HIF-1?, a key regulator of the cellular response to hypoxia, is also upregulated in the vasculature of human AD brains. The objective of this study is to investigate inflammatory protein expression in the cerebrovasculature of transgenic AD mice and to explore the role of thrombin as a mediator of cerebrovascular inflammation and oxidative stress in AD and in hypoxia-induced changes in brain endothelial cells. Immunofluorescent analysis of the cerebrovasculature in AD mice demonstrates significant (p<0.01-0.001 increases in thrombin, HIF-1?, interleukin-6 (IL-6, monocyte chemoattractant protein-1 (MCP-1, matrix metalloproteinases (MMPs, and reactive oxygen species (ROS compared to controls. Administration of the thrombin inhibitor dabigatran (100 mg/kg to AD mice for 34 wks significantly decreases expression of inflammatory proteins and ROS. Exposure of cultured brain endothelial cells to hypoxia for 6 h causes an upregulation of thrombin, HIF-1?, MCP-1, IL-6 and MMP2 and ROS. Treatment of endothelial cells with the dabigatran (1 nM reduces ROS generation and inflammatory protein expression (p<0.01-0.001. The data demonstrate that inhibition of thrombin in culture blocks the increase in inflammatory protein expression and ROS generation evoked by hypoxia. Also, administration of dabigatran to transgenic AD mice diminishes expression of inflammatory proteins and ROS in the cerebromicrovasculature. Taken together, these results suggest that inhibiting thrombin generation could have therapeutic value in AD and other disorders where hypoxia, inflammation and oxidative stress are involved.

PaulaGrammas

2013-05-01

58

Percutaneous ultrasound-guided thrombin injection for the treatment of iatrogenic femoral artery pseudoaneurysms  

International Nuclear Information System (INIS)

usion: Percutaneous ultrasound-guided thrombin injection of is a quick, effective and safe treatment for iatrogenic femoral pseudoaneurysms. For larger pseudoaneurysms, although it is worth attempting more than one thrombin injection, endovascular repair may eventually be required.

59

Scanning electrochemical microscopy for study of aptamer-thrombin interfacial interactions on gold disk microelectrodes.  

Science.gov (United States)

A feasibility for the determination of thrombin on gold disk microelectrodes (GDMs) using scanning electrochemical microscopy (SECM) is reported. The assembly process step-by-step of thrombin aptasensor on GDMs is monitored by SECM. SECM analysis reveals the immobilization of thrombin aptamers on GDMs. The interaction between thrombin aptamers and thrombin on GDMs is imaged by SECM with feedback mode using ferrocenemethanol as an electrochemical mediator. The formation of thrombin/thrombin aptamer complex on GDMs results in a decrease in the tip peak current on spatial SECM images. This method is able to linearly and selectively detect thrombin over a linear range from 10(-12) to 10(-5)M with a detection limit of 6.07 fM. PMID:24407695

Bai, Huei-Yu; del Campo, F Javier; Tsai, Yu-Chen

2014-03-01

60

[Thrombin--a regulator of reparative processes in wound healing].  

Science.gov (United States)

Thrombin, binding to receptors of the protease activated receptor (PAR) family, is involved in wound healing by inducing the reparation processes and regulating the activity of mast cells, which secrete mediators of inflammation. Using thrombin receptor agonist peptide (TRAP-6) for the activation of rat mast cells, effect of several receptors, including PAR-1, on mast cells was demonstrated. It was shown that TRAP increases the concentration of Ca2+ in the cytoplasm of mast cells and regulates cell degranulation, while releasing nitrogen oxide. Thrombin encapsulated in poly(N-vinyl caprolactam)-calcium alginate (PVCL-Ca-Alg) hydrogel films promotes wound healing in rats as demonstrated by the acceleration of fibroblast proliferation and neovascularization. PMID:9612571

Strukova, S M; Dugina, T N; Chistov, I V; Markvicheva, E A; Kuptsova, S V; Kolokol'chikova, E G; Rumsh, L D; Zubov, V P; Gluza, E

1998-04-01

 
 
 
 
61

Cellular signaling events involved in thrombin activation of vascular endothelium  

Energy Technology Data Exchange (ETDEWEB)

Although cytosolic calcium ((Ca))/sub I/), inositol trisphosphate (IP/sub 3/) and diacylglycerol (DAG) are important second messengers involved in stimulus-response coupling to certain hormones, little information is available regarding their role in the activation of vascular endothelial cells (EC) during coagulation. The authors have used cultured human umbilical vein EC to examine thrombin effects on (Ca)/sub I/ and on IP/sub 3/ and DAG formation. Thrombin (1 U/ml) induces a rapid (peak < 15 sec) increase in (Ca)/sub I/ (300% basal levels) in suspensions of fura-2 (fluorescent Ca/sup 2 +/ indicator)-loaded EC. In addition thrombin stimulates concentration-dependent (.001-1 U/ml) increases in calcium efflux and IP/sub 3/ formation in EC prelabeled with /sup 45/Ca or /sup 3/H-myoinositol. Peak IP/sub 3/ (182+/-14% control) is rapid (<15 sec) and temporally similar to the rise in (Ca)/sub I/. In /sup 3/H-arachidonic acid-labeled EC, thrombin increases DAG (protein kinase C activator) at 15 sec (133+/-8% control), as well as 5 min (148+/-12% control). EC preincubation with 4..beta..-phorbol 12-myristate 13-acetate (10/sup -7/M, 5 min), a potent activator of protein kinase C, blocks thrombin (1 U/ml)-induced increases in (Ca)/sub I/ and IP/sub 3/. Thus, thrombin may trigger at least two distinct signaling pathways (IP/sub 3//calcium; DAG/protein kinase C) in EC. In addition, DAG stimulation of protein kinase C may influence this mediator's action in vascular EC.

Brock, T.A.; Capasso, E.L.; Gimbrone, M.A. Jr.

1986-03-05

62

Clindamycin Topical  

Science.gov (United States)

Topical clindamycin is used to treat acne. Clindamycin is in a class of medications called lincomycin antibiotics. It works by slowing or stopping the growth of bacteria that cause acne and by decreasing ...

63

Estradiol Topical  

Science.gov (United States)

... used to treat and prevent hot flushes (hot flashes; sudden strong feelings of heat and sweating) in ... and moisture (not in the bathroom). Do not freeze topical estradiol. Keep estradiol gel away from open ...

64

Tacrolimus Topical  

Science.gov (United States)

Tacrolimus ointment is used to treat the symptoms of eczema (atopic dermatitis; a skin disease that causes ... whose eczema has not responded to another medication. Tacrolimus is in a class of medications called topical ...

65

Intravitreal thrombin activity is elevated in retinal vein occlusion.  

Science.gov (United States)

To evaluate whether intravitreal thrombin activity is elevated in eyes with branch retinal vein occlusion (BRVO) and central retinal vein occlusion (CRVO) in comparison to healthy controls. Prospective clinical case series of 19 patients with BRVO, 13 patients suffering from CRVO and nine participants serving as controls. Vitreous taps were extracted from the central vitreous body, 200??l frozen/thawed sample was immediately stabilized with 200??l 5% human albumin, and 200??l mixture thereof was stabilized with 200??l 2.5?mol/l arginine, pH 8.6. Thrombin activity was determined chromogenically. Intravitreal levels of vascular endothelial growth factor (VEGF) as a marker for blood-retina barrier (BRB) breakdown were measured by a commercial chemiluminescent enzyme immuno assay (R&D). Intravitreal thrombin activity and VEGF levels were 1.6?±?1.2?mIU/ml (mean value?±?SD; range: 0.2-4.2?mIU/ml) and 554?±?568?pg/ml (range: 20-2005?pg/ml) in BRVO-affected eyes, 2.6?±?1.2?mIU/ml (range: 0.8-5.2?mIU/ml) and 1332?±?1350?pg/ml (range: 58-3943?pg/ml) in eyes suffering from CRVO as well as 0.8?±?0.8?mIU/ml (range: 0.2-2.7?mIU/ml) and 115?±?120?pg/ml (range: 32-431?pg/ml) in controls. There are significant differences of intravitreal thrombin activity and intravitreal VEGF levels between eyes with BRVO, CRVO, and controls (P?=?0.007 and P?=?0.003, Kruskal-Wallis test). Intravitreal thrombin activity is significantly correlated with intravitreal VEGF levels (r?=?0656; P?thrombin activity might serve as a new marker for BRB breakdown or macular fibrin deposition in ophthalmology. Significant differences of intravitreal thrombin activity between eyes with BRVO, CRVO, and healthy controls might offer new therapeutic strategies for RVO-affected eyes. The effect of oral and intravitrealy injected direct thrombin inhibitors needs to be evaluated in further investigations. PMID:24625582

Bertelmann, Thomas; Stief, Thomas; Sekundo, Walter; Mennel, Stefan; Nguyen, Nauke; Koss, Michael J

2014-10-01

66

Nonradiative recombination in semiconductors  

CERN Document Server

In recent years, great progress has been made in the understandingof recombination processes controlling the number of excessfree carriers in semiconductors under nonequilibrium conditions. As a result, it is now possible to give a comprehensivetheoretical description of these processes. The authors haveselected a number of experimental results which elucidate theunderlying physical problems and enable a test of theoreticalmodels. The following topics are dealt with: phenomenological theory ofrecombination, theoretical models of shallow and deep localizedstates, cascade model of carrier captu

Abakumov, VN; Yassievich, IN

1991-01-01

67

Thrombin regulates components of the fibrinolytic system in human mesangial cells  

International Nuclear Information System (INIS)

Besides its procoagulant activity, thrombin has been shown to stimulate cell proliferation and to regulate the fibrinolytic pathway. We report here the effect of purified human alpha thrombin on the synthesis of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) by cultured human mesangial cells. Thrombin (0 to 2.5 U/ml) increased in a time- and dose-dependent manner the production of t-PA and PAI-1 (2- to 3-fold increase of secreted t-PA and PAI-1 release during a 24 hour incubation). This effect was associated with a twofold increase in DNA synthesis measured by 3H-thymidine incorporation. Zymographic analysis and reverse fibrin autography showed that thrombin also increased the level of the 110 Kd t-PA-PAI-1 complex, whereas PAI-1 was present as a free 50 Kd form in the culture medium conditioned by unstimulated and thrombin-stimulated cells. Free t-PA was never observed. Both membrane binding and catalytic activity of thrombin were required since the effects of 1 U/ml thrombin were inhibited by addition 2 U/ml hirudin, which inhibits the membrane binding and catalytic activity of thrombin, and since DFP-inactivated thrombin, which has the ability to bind but which has no enzymatic activity, did not induce t-PA or PAI-1. Gamma thrombin, which does not bind to thrombin receptor, did not increase t-PA and PAI-1 releases. The effects of thrombin were probably mediated by protein kinase C activation since H7, an inhibitor of protein kinases, inhibited significantly thrombin effects on t-PA and PAI-1 production, and since addition of an activator of protein kinase A, 8-bromocyclic AMP (100 microM), induced a significant inhibition of the thrombin effect. The effects of thrombin were also suppressed by 1.25 micrograms/ml alpha amanitin, suggesting a requirement of de novo RNA synthesis

68

Thrombin regulates components of the fibrinolytic system in human mesangial cells  

Energy Technology Data Exchange (ETDEWEB)

Besides its procoagulant activity, thrombin has been shown to stimulate cell proliferation and to regulate the fibrinolytic pathway. We report here the effect of purified human alpha thrombin on the synthesis of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) by cultured human mesangial cells. Thrombin (0 to 2.5 U/ml) increased in a time- and dose-dependent manner the production of t-PA and PAI-1 (2- to 3-fold increase of secreted t-PA and PAI-1 release during a 24 hour incubation). This effect was associated with a twofold increase in DNA synthesis measured by 3H-thymidine incorporation. Zymographic analysis and reverse fibrin autography showed that thrombin also increased the level of the 110 Kd t-PA-PAI-1 complex, whereas PAI-1 was present as a free 50 Kd form in the culture medium conditioned by unstimulated and thrombin-stimulated cells. Free t-PA was never observed. Both membrane binding and catalytic activity of thrombin were required since the effects of 1 U/ml thrombin were inhibited by addition 2 U/ml hirudin, which inhibits the membrane binding and catalytic activity of thrombin, and since DFP-inactivated thrombin, which has the ability to bind but which has no enzymatic activity, did not induce t-PA or PAI-1. Gamma thrombin, which does not bind to thrombin receptor, did not increase t-PA and PAI-1 releases. The effects of thrombin were probably mediated by protein kinase C activation since H7, an inhibitor of protein kinases, inhibited significantly thrombin effects on t-PA and PAI-1 production, and since addition of an activator of protein kinase A, 8-bromocyclic AMP (100 microM), induced a significant inhibition of the thrombin effect. The effects of thrombin were also suppressed by 1.25 micrograms/ml alpha amanitin, suggesting a requirement of de novo RNA synthesis.

Villamediana, L.M.; Rondeau, E.; He, C.J.; Medcalf, R.L.; Peraldi, M.N.; Lacave, R.; Delarue, F.; Sraer, J.D. (INSERM Unite 64, Paris (France))

1990-11-01

69

Immobilized thrombin receptor agonist peptide accelerates wound healing in mice.  

Science.gov (United States)

To accelerate the healing processes in wound repair, attempts have been repeatedly made to use growth factors including thrombin and its peptide fragments. Unfortunately, the employment of thrombin is limited because of its high liability and pro-inflammatory actions at high concentrations. Some cellular effects of thrombin in wound healing are mediated by the activation of protease activated receptor-1 (PAR-1). The thrombin receptor agonist peptide (TRAP:SFLLRN) activates this receptor and mimics the effects of thrombin, but TRAP is a relatively weak agonist. We speculated that the encapsulated peptide may be more effective for PAR-1 activation than nonimmobilized peptide and developed a novel method for TRAP encapsulation in hydrogel films based on natural and synthetic polymers. The effects of an encapsulated TRAP in composite poly(N-vinyl caprolactam)-calcium alginate (PVCL) hydrogel films were investigated in a mouse model of wound healing. On day 7 the wound sizes decreased by about 60% under TRAP-chitosan-containing PVCL films, as compared with control films without TRAP. In the case of TRAP-polylysine-containing films no significant decrease in wound sizes was found. The fibroblast/macrophage ratio increased under TRAP-containing films on day 3 and on day 7. The number of proliferating fibroblasts increased to 150% under TRAP-chitosan films on day 7 as compared with control films. The number of [3H]-thymidine labeled endothelial and epithelial cells in granulation tissues was also enhanced. Thus, the immobilized TRAP to PVCL-chitosan hydrogel films were found to promote wound healing following the stimulation of fibroblast and epithelial cell proliferation and neovascularization. Furthermore, TRAP was shown to inhibit the secretion of the inflammatory mediator PAF from stimulated rat peritoneal mast cells due to augmentation of NO release from the mast cells. The encapsulated TRAP is suggested to accelerate wound healing due to the anti-inflammatory effects and earlier development of the proliferative phase of wound healing. PMID:11697718

Strukova, S M; Dugina, T N; Chistov, I V; Lange, M; Markvicheva, E A; Kuptsova, S; Zubov, V P; Glusa, E

2001-10-01

70

Prevention of venous thromboembolism following orthopaedic surgery: clinical potential of direct thrombin inhibitors.  

Science.gov (United States)

Patients undergoing total hip or total knee replacement are at high risk of venous thromboembolism (VTE), and are therefore considered to be populations well suited for the evaluation and dose optimisation of new anticoagulants. Deep vein thrombosis may lead to life-threatening pulmonary embolism, disabling morbidity in the form of the post-thrombotic syndrome, and risk of recurrent thrombotic events. There is increasing evidence that anticoagulant treatment for the prevention of VTE should be extended from 1 to at least 4 weeks after surgery. Anticoagulation with vitamin K antagonists (such as warfarin), low molecular weight heparin or unfractionated heparin effectively lowers the risk of VTE, but these anticoagulants have limitations such as the need for coagulation monitoring and subsequent dose adjustment (vitamin K antagonists), difficulty of continuing prophylaxis out of hospital because of the requirement for parenteral administration, and risk of heparin-induced thrombocytopenia. The development of new anticoagulants has been pursued with the aim of finding more effective, safer and/or more convenient therapies. Thrombin is a central regulator in the coagulation and inflammation process and several direct thrombin inhibitors (DTIs) with distinct pharmacological profiles, as well as pharmacological differences from the conventional anticoagulants, are currently in clinical use for certain indications or are under development. Clinical experience with parenterally administered DTIs has accumulated since the mid 1990s, although only desirudin (a recombinant hirudin) is currently approved for use in patients undergoing orthopaedic surgery. Two oral DTIs, ximelagatran and dabigatran etexilate, are in clinical development. Dabigatran etexilate has recently been evaluated in phase II clinical trials in patients undergoing total hip replacement. Several large phase III trials have now demonstrated the efficacy and safety of ximelagatran in the prevention of VTE following total hip or knee replacement. Ximelagatran can be used with an oral fixed dose without the need for coagulation monitoring or dose adjustment. Hence, it offers significant potential to facilitate the management of anticoagulation in or out of hospital. PMID:15018589

Eriksson, Bengt I; Dahl, Ola E

2004-01-01

71

Evaluation of DNA aptamers directed to thrombin as potential thrombus imaging agents  

International Nuclear Information System (INIS)

Two DNA aptamers directed against two separate exosites on human ?-thrombin were evaluated for thrombus-imaging potential. Aptamer ODN 1 is directed to the thrombin substrate binding site (exosite 1). Our finding that ODN 1 competes with fibrin for binding to exosite 1 on thrombin suggests that ODN 1 will not be useful for thrombus imaging. Aptamer ODN 2 is directed against the thrombin heparin binding site (exosite 2). ODN 2 bound to model thrombi that were formed either by clotting purified fibrinogen with thrombin, or by recalcifying citrated plasma. As the thrombin content of thrombi was increased the rate of ODN 2 uptake into preformed thrombi increased, whereas the rate of release of ODN 2 out of preformed thrombi decreased. This in vitro data suggested that ODN 2 might be useful for thrombus imaging because it can bind to exosite 2 on fibrin-bound thrombin. However, in a rabbit jugular vein model using thrombus supplemented with human thrombin, ODN 2 uptake was equal to the ovalbumin control, and did not reflect thrombin content. While the in vitro results with ODN 2 were consistent with thrombus imaging, the rapid clearance of ODN 2 from circulation, combined with slow mass transfer in the clot, seem to work against in vivo thrombin-dependent imaging or washout analysis

72

Thrombin increases inflammatory cytokine and angiogenic growth factor secretion in human adipose cells in vitro  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Abdominal obesity is associated with pro-thrombotic and inflammatory states. Therefore, the purpose of this study was to examine the expression of thrombin receptors (PAR1 and PAR4 human adipose tissue and whether thrombin stimulates an inflammatory cytokine and growth factor profile in human adipose tissue. Methods Human adipose tissue, isolated preadipocytes and differentiated adipocytes were used in this study. PAR1 and PAR4 mRNA and protein were detected by RT-PCR and immunoblot analysis in both adipose tissue and adipose microvessels. In separate studies, IL-1?, IL-6, MCP-1, TNF-?, IL-10, FGF-2, VEGF, and PDGF production were measured from adipose tissue (n = 5, adipocytes (n = 5, and preadipocytes (n = 3 supernatants with and without thrombin (1 or 10 U/ml; 24 hrs treatment. Results Thrombin increased cytokine secretion of IL-1?, IL-6, MCP-1 and TNF-? and growth factor secretion of VEGF from adipocytes along with MCP-1 and VEGF from preadipocytes. The direct thrombin inhibitor lepirudin given in conjunction with thrombin prevented the thrombin-mediated increase in cytokine and growth factor secretion. Conclusion Here we show that thrombin PAR1 and PAR4 receptors are present and that thrombin stimulates inflammatory cytokine generation and growth factor release in human adipose tissue and cells in vitro. These data suggest that thrombin may represent a molecular link between obesity and associated inflammation.

Phillips Shane A

2009-03-01

73

Thrombin aptasensing with inherently electroactive graphene oxide nanoplatelets as labels  

Science.gov (United States)

Graphene and its associated materials are commonly used as the transducing platform in biosensing. We propose a different approach for the application of graphene in biosensing. Here, we utilized graphene oxide nanoplatelets as the inherently electroactive labels for the aptasensing of thrombin. The basis of detection lies in the ability of graphene oxide to be electrochemically reduced, thereby providing a well-defined reduction wave; one graphene oxide nanoplatelet of dimension 50 × 50 nm can provide a reduction signal by accepting ~22 000 electrons. We demonstrate that by using graphene oxide nanoplatelets as an inherently electroactive label, we can detect thrombin in the concentration range of 3 pM-0.3 ?M, with good selectivity of the aptamer towards interferences by bovine serum albumin, immunoglobulin G and avidin. Therefore, the inherently electroactive graphene oxide nanoplatelets are a material which can serve as an electroactive label, in a manner similar to metallic nanoparticles.

Loo, Adeline Huiling; Bonanni, Alessandra; Pumera, Martin

2013-05-01

74

Thrombin generation by activated factor VII on platelet activated by different agonists. Extending the cell-based model of hemostasis  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Platelet activation is crucial in normal hemostasis. Using a clotting system free of external tissue factor, we investigated whether activated Factor VII in combination with platelet agonists increased thrombin generation (TG in vitro. Methods and results TG was quantified by time parameters: lag time (LT and time to peak (TTP, and by amount of TG: peak of TG (PTG and area under thrombin formation curve after 35 minutes (AUC?35min in plasma from 29 healthy volunteers using the calibrated automated thrombography (CAT technique. TG parameters were measured at basal conditions and after platelet stimulation by sodium arachidonate (AA, ADP, and collagen (Col. In addition, the effects of recombinant activated FVII (rFVIIa alone or combined with the other platelet agonists on TG parameters were investigated. We found that LT and TTP were significantly decreased (p 35min were significantly increased (p 35min (but not PTG when compared to platelet rich plasma activated with agonists in the absence of rFVIIa. Conclusion Platelets activated by AA, ADP, Col or rFVIIa triggered TG. This effect was increased by combining rFVIIa with other agonists. Our intrinsic coagulation system produced a burst in TG independent of external tissue factor activity an apparent hemostatic effect with little thrombotic capacity. Thus we suggest a modification in the cell-based model of hemostasis.

Herrera Maria

2006-04-01

75

Microplate assay for aptamer-based thrombin detection using a DNA-enzyme conjugate based on histidine-tag chemistry.  

Science.gov (United States)

We report a method to prepare a DNA-enzyme conjugate using histidine-tag (His-tag) chemistry. A DNA oligonucleotide was modified with nitrilotriacetate (NTA), whose K(d) was approximately 10?? (M?¹) toward a His-tag present on a recombinant protein via the complexation of Ni²?. His-tagged alkaline phosphatase (His-AP) was used as the model enzyme. Enzyme immobilization on the microplate revealed the conjugation of His-AP and the NTA-modified DNA via an Ni²? complex. SPR measurements also proved the conjugation of His-AP with the NTA-modified DNA via an Ni²? complex. The DNA-enzyme conjugate was then used for the detection of thrombin using a DNA aptamer. The DNA-AP conjugate successfully amplified the binding signal between the DNA aptamer and the thrombin, and the signal was measured as the fluorescent intensity derived from the AP-catalyzed reaction. The detection limit was 11 nM. Finally, we studied the effect of the release of the immobilized His-AP from the microplate on the AP activity, because the present strategy used a cleavable linker for the conjugation and the enzyme immobilization. The DNase-catalyzed release of the immobilized His-AP resulted in a 1.7-fold higher AP activity than observed when the His-AP was surface-immobilized. PMID:22178916

Shimada, Josui; Maruyama, Tatsuo; Kitaoka, Momoko; Kamiya, Noriho; Goto, Masahiro

2012-02-15

76

A critical role for thrombin in vertebrate lens regeneration.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Lens regeneration in urodele amphibians such as the newt proceeds from the dorsal margin of the iris where pigment epithelial cells (PEC) re-enter the cell cycle and transdifferentiate into lens. A general problem in regeneration research is to understand how the events of tissue injury or removal are coupled to the activation of plasticity in residual differentiated cells or stem cells. Thrombin, a pivotal regulator of the injury response, has been implicated as a regulator of cell cycle re-...

Imokawa, Yutaka; Simon, Andra?s; Brockes, Jeremy P.

2004-01-01

77

Cosmological Recombination  

Science.gov (United States)

In this thesis we focus on studying the physics of cosmological recombination and how the details of recombination affect the Cosmic Microwave Background (CMB) anisotropies. We present a detailed calculation of the spectral line distortions on the CMB spectrum arising from the Lyman-alpha and the lowest two-photon transitions in the recombination of hydrogen (H), and the corresponding lines from helium (He). The peak of these distortions mainly comes from the Lyman-alpha transition and occurs at about 170 microns, which is the Wien part of the CMB. The major theoretical limitation for extracting cosmological parameters from the CMB sky lies in the precision with which we can calculate the cosmological recombination process. With this motivation, we perform a multi-level calculation of the recombination of H and He with the addition of the spin-forbidden transition for neutral helium (He I), plus the higher order two-photon transitions for H and among singlet states of He I. We find that the inclusion of the spin-forbidden transition results in more than a percent change in the ionization fraction, while the other transitions give much smaller effects. Last we modify RECFAST by introducing one more parameter to reproduce recent numerical results for the speed-up of helium recombination. Together with the existing hydrogen `fudge factor', we vary these two parameters to account for the remaining dominant uncertainties in cosmological recombination. By using a Markov Chain Monte Carlo method with Planck forecast data, we find that we need to determine the parameters to better than 10% for He I and 1% for H, in order to obtain negligible effects on the cosmological parameters.

Wong, Wan Yan

2008-11-01

78

Combined administration of aspirin and a specific thrombin inhibitor in man.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

BACKGROUND: Heparin is of limited value as an antithrombotic drug in the presence of platelet activation and residual thrombus. Greater anticoagulant activity can be achieved in vivo with more specific thrombin inhibitors. Heparin may also increase the risk of bleeding by an effect on platelets that is independent of its thrombin inhibitory activity. METHODS AND RESULTS: The pharmacodynamic and pharmacokinetic effects of a novel thrombin inhibitor, argatroban, were examined alone and in combi...

Clarke, Rj; Mayo, G.; Fitzgerald, Ga; Fitzgerald, Dj

1991-01-01

79

Cosmological Recombination  

CERN Document Server

In this thesis we focus on studying the physics of cosmological recombination and how the details of recombination affect the Cosmic Microwave Background (CMB) anisotropies. We present a detailed calculation of the spectral line distortions on the CMB spectrum arising from the Lyman-alpha and the lowest two-photon transitions in the recombination of hydrogen (H), and the corresponding lines from helium (He). The peak of these distortions mainly comes from the Lyman-alpha transition and occurs at about 170 microns, which is the Wien part of the CMB. The major theoretical limitation for extracting cosmological parameters from the CMB sky lies in the precision with which we can calculate the cosmological recombination process. With this motivation, we perform a multi-level calculation of the recombination of H and He with the addition of the spin-forbidden transition for neutral helium (He I), plus the higher order two-photon transitions for H and among singlet states of He I. We find that the inclusion of the s...

Wong, Wan Yan

2008-01-01

80

A straightforward modification in the thrombin binding aptamer improving the stability, affinity to thrombin and nuclease resistance.  

Science.gov (United States)

Degradation of nucleic acids in biological environments is the major drawback of the therapeutic use of aptamers. Among the approaches used to circumvent this negative aspect, the introduction of 3'-3' inversion of polarity sites at the sequence 3'-end has successfully been proposed. However, the introduction of inversion of polarity at the ends of the sequence has never been exploited for G-quadruplex forming aptamers. In this communication we describe CD, UV, electrophoretic and biochemical investigations concerning thrombin binding aptamer analogues containing one or two inversions of polarity sites at the oligonucleotide ends. Data indicate that, in some cases, this straightforward chemical modification is able to improve, at the same time, the thermal stability, affinity to thrombin and nuclease resistance in biological environments, thus suggesting its general application as a post-SELEX modification also for other therapeutically promising aptamers adopting G-quadruplex structures. PMID:25296283

Esposito, Veronica; Scuotto, Maria; Capuozzo, Antonella; Santamaria, Rita; Varra, Michela; Mayol, Luciano; Virgilio, Antonella; Galeone, Aldo

2014-11-28

 
 
 
 
81

Percutaneous Ultrasound-Guided Thrombin Injection in Iatrogenic Arterial Pseudoaneurysms: Effectiveness and Complications  

Energy Technology Data Exchange (ETDEWEB)

To evaluate and describe the efficacy and side effects of a percutaneous thrombin injection under ultrasonography guidance for the treatment of iatrogenic pseudo aneurysms Eighteen consecutive iatrogenic pseudo aneurysm cases were treated with a thrombin injection. The thrombin was injected into the pseudo aneurysm cavity using a 22-gauge needle under ultrasonographic guidance. The causes of the pseudo aneurysms are as follows: post coronary angiography (9 cases), percutaneous coronary balloon angioplasty (5 cases), cerebral angiography (1 case), transhepatic chemo embolization (1 case), percutaneous trans femoral arterial stent insertion (1 case) and bone marrow aspiration for a marrow transplant (1 case). Only one case required a secondary thrombin injection due to recurrent flow in the pseudo aneurysm lumen, which was detected at the follow up Doppler ultrasound. Other seventeen cases were successfully treated on the first trial. There were no technical failures or complication related to the procedure. The average amount of thrombin injected was 733 IU. Nine out of 18 treated patients (50%) showed mild reactions to the thrombin including mild fever (4 cases), chilling sensation (3 cases), a chilling sensation with mild dyspnea (1 case), mild chest discomfort (1 case) after the thrombin injection. All these side effects were transient and improved several hours later. All the iatrogenic pseudo aneurysms were treated successfully with an ultrasound-guided percutaneous thrombin injection. There was a high rate of hypersensitivity to the bovine thrombin, which precaution should be taken to prevent more serious side effects

Koh, Young Hwan [Boramae Hospital, Seoul (Korea, Republic of); Kim, Hak Soo; Kim, Hyung Sik; Min, Seung Kee [Gachon Medical School, Incheon (Korea, Republic of)

2005-09-15

82

Pulmonary epithelial clearance of 99mTc-DTPA after thrombin-induced pulmonary microembolism  

International Nuclear Information System (INIS)

We investigated the effect of thrombin-induced pulmonary microembolism on the pulmonary clearance rate of aerosolized 99mTc diethylenetriamine pentaacetic acid (99mTc-DTPA) in awake, chronically prepared sheep. Chest activity was recorded after administration of a 0.44 micron aerosol of 99mTc-DTPA. Decay-corrected data were fit to an exponential and expressed as percent decrease per min (%/min). Sheep were given alpha-thrombin intravenously (80 U/kg for 10 min) 60 min after the aerosol administration. The clearance rate prior to alpha-thrombin was 0.35 +/- 0.05 %/min (mean +/- SEM). During alpha-thrombin administration, the clearance rate increased to 5.84 +/- 0.70 %/min (p less than 0.001 from baseline), but returned to 0.41 +/- 0.06 %/min within 30 min after the end of the thrombin infusion. The increased clearance rate during alpha-thrombin administration was not due to increased lung volume since alpha-thrombin did not change functional residual capacity. Moreover, the clearance rate was unchanged during gamma-thrombin administration, which does not induce coagulation, or during alpha-thrombin challenge in defibrinogenated animals. alpha-thrombin administration in neutrophil-depleted sheep caused a transient increase in DTPA clearance similar to that in control sheep, suggesting that the increase occurred independently of neutrophils. The results indicate that alpha-thrombin causes a large, transient increase in 99mTc-DTPA clearance, which may be the result of increased epithelial permeability. This response is dependent on the activation of intravascular coagulation

83

A novel hemostatic sealant composed of gelatin, transglutaminase and thrombin effectively controls liver trauma-induced bleeding in dogs  

Science.gov (United States)

Aim: A novel hemostatic sealant based on the in situ gel formation of gelatin catalyzed by thrombin and crosslinked by transglutaminase was designed. The aim of this study was to investigate the efficacy of the hemostatic sealant in control of bleeding caused by liver trauma in dogs. Methods: Hepatic trauma that mimicked the grade III–IV rupture of liver was made in 20 dogs. The traumatic lesion was topically administered the hemostatic sealant (treatment group, n=10), or a thrombin solution (control group, n=10). The time to achieve hemostasis and the blood loss were measured. Contrast-enhanced ultrasound (CEUS) examination was performed in each animal on d 3, d 7, and d 10 d postoperatively to study the healing of the lesions. Results: The mean time to achieve hemostasis in the treatment group was significantly shorter than that in the control group (1.20±0.33 vs 6.70±0.64 min, Psealant shows much better efficacy in hemostasis and may promote wound healing in dog liver trauma. PMID:23645012

Xie, Xia; Tian, Jiang-ke; Lv, Fa-qin; Wu, Rong; Tang, Wen-bo; Luo, Yu-kun; Huang, Ya-qin; Tang, Jie

2013-01-01

84

Synthetic alpha-thrombin receptor peptides activate G protein-coupled signaling pathways but are unable to induce mitogenesis.  

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alpha-Thrombin (thrombin) stimulates phospholipase C and modulates the activity of adenylate cyclase in a number of cell types via G protein-coupled receptors. It is also a potent growth factor, notably for a line of hamster fibroblasts (CCL39 cells). Recently, predicted amino acid sequences for human and hamster thrombin receptors have been reported that display a putative thrombin cleavage site in the N-terminal extracellular domain. Synthetic peptides corresponding to 14 residues carboxyl ...

Vouret-craviari, V.; Obberghen-schilling, E.; Rasmussen, U. B.; Pavirani, A.; Lecocq, J. P.; Pouysse?gur, J.

1992-01-01

85

Topical haemostatic agents for skin wounds: a systematic review  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Various agents and techniques have been introduced to limit intra-operative blood loss from skin lesions. No uniformity regarding the type of haemostasis exists and this is generally based on the surgeon's preference. To study the effectiveness of haemostatic agents, standardized wounds like donor site wounds after split skin grafting (SSG appear particularly suitable. Thus, we performed a systematic review to assess the effectiveness of haemostatic agents in donor site wounds. Methods We searched all randomized clinical trials (RCTs on haemostasis after SSG in Medline, Embase and the Cochrane Library until January 2011. Two reviewers independently assessed trial relevance and quality and performed data analysis. Primary endpoint was effectiveness regarding haemostasis. Secondary endpoints were wound healing, adverse effects, and costs. Results Nine relevant RCTs with a fair methodological quality were found, comparing epinephrine, thrombin, fibrin sealant, alginate dressings, saline, and mineral oil. Epinephrine achieved haemostasis significantly faster than thrombin (difference up to 2.5 minutes, saline or mineral oil (up to 6.5 minutes. Fibrin sealant also resulted in an up to 1 minute quicker haemostasis than thrombin and up to 3 minutes quicker than placebo, but was not directly challenged against epinephrine. Adverse effects appeared negligible. Due to lack of clinical homogeneity, meta-analysis was impossible. Conclusion According to best available evidence, epinephrine and fibrin sealant appear superior to achieve haemostasis when substantial topical blood loss is anticipated, particularly in case of (larger SSGs and burn debridement.

Ubbink Dirk T

2011-07-01

86

Batroxobin binds fibrin with higher affinity and promotes clot expansion to a greater extent than thrombin.  

Science.gov (United States)

Batroxobin is a thrombin-like serine protease from the venom of Bothrops atrox moojeni that clots fibrinogen. In contrast to thrombin, which releases fibrinopeptide A and B from the NH2-terminal domains of the A?- and B?-chains of fibrinogen, respectively, batroxobin only releases fibrinopeptide A. Because the mechanism responsible for these differences is unknown, we compared the interactions of batroxobin and thrombin with the predominant ?A/?A isoform of fibrin(ogen) and the ?A/?' variant with an extended ?-chain. Thrombin binds to the ?'-chain and forms a higher affinity interaction with ?A/?'-fibrin(ogen) than ?A/?A-fibrin(ogen). In contrast, batroxobin binds both fibrin(ogen) isoforms with similar high affinity (Kd values of about 0.5 ?M) even though it does not interact with the ?'-chain. The batroxobin-binding sites on fibrin(ogen) only partially overlap with those of thrombin because thrombin attenuates, but does not abrogate, the interaction of ?A/?A-fibrinogen with batroxobin. Furthermore, although both thrombin and batroxobin bind to the central E-region of fibrinogen with a Kd value of 2-5 ?M, the ?(17-51) and B?(1-42) regions bind thrombin but not batroxobin. Once bound to fibrin, the capacity of batroxobin to promote fibrin accretion is 18-fold greater than that of thrombin, a finding that may explain the microvascular thrombosis that complicates envenomation by B. atrox moojeni. Therefore, batroxobin binds fibrin(ogen) in a manner distinct from thrombin, which may contribute to its higher affinity interaction, selective fibrinopeptide A release, and prothrombotic properties. PMID:23612970

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

2013-06-01

87

Batroxobin Binds Fibrin with Higher Affinity and Promotes Clot Expansion to a Greater Extent than Thrombin*  

Science.gov (United States)

Batroxobin is a thrombin-like serine protease from the venom of Bothrops atrox moojeni that clots fibrinogen. In contrast to thrombin, which releases fibrinopeptide A and B from the NH2-terminal domains of the A?- and B?-chains of fibrinogen, respectively, batroxobin only releases fibrinopeptide A. Because the mechanism responsible for these differences is unknown, we compared the interactions of batroxobin and thrombin with the predominant ?A/?A isoform of fibrin(ogen) and the ?A/?? variant with an extended ?-chain. Thrombin binds to the ??-chain and forms a higher affinity interaction with ?A/??-fibrin(ogen) than ?A/?A-fibrin(ogen). In contrast, batroxobin binds both fibrin(ogen) isoforms with similar high affinity (Kd values of about 0.5 ?m) even though it does not interact with the ??-chain. The batroxobin-binding sites on fibrin(ogen) only partially overlap with those of thrombin because thrombin attenuates, but does not abrogate, the interaction of ?A/?A-fibrinogen with batroxobin. Furthermore, although both thrombin and batroxobin bind to the central E-region of fibrinogen with a Kd value of 2–5 ?m, the ?(17–51) and B?(1–42) regions bind thrombin but not batroxobin. Once bound to fibrin, the capacity of batroxobin to promote fibrin accretion is 18-fold greater than that of thrombin, a finding that may explain the microvascular thrombosis that complicates envenomation by B. atrox moojeni. Therefore, batroxobin binds fibrin(ogen) in a manner distinct from thrombin, which may contribute to its higher affinity interaction, selective fibrinopeptide A release, and prothrombotic properties. PMID:23612970

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

2013-01-01

88

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

89

Thrombin Receptor and Ventricular Arrhythmias after Acute Myocardial Infarction  

Science.gov (United States)

The mechanism mediating the development of ventricular arrhythmia (VA) after acute myocardial infarction (AMI) is still uncertain. Thrombin receptor (TR) activation has been proven to be arrhythmogenic in many other situations, and we hypothesize that it may participate in the genesis of post-AMI VA. Using a left coronary artery ligation rat model of AMI, we found that a local injection of hirudin into the left ventricle (LV) significantly reduced the ratio of VA durations to infarction sizing, whereas injection of thrombin receptor–activating peptide (TRAP) increased the ratios of VA duration to infarction sizing. The effects of TR activation on whole-cell currents were investigated in isolated myocytes. TRAP increased a glibenclamide-sensitive outward current. Pretreatment of rats with glibenclamide (4 mg/kg intraperitoneally) eliminated the effects of a local injection of TRAP on the ratios of VA durations to infarction sizing. TR mRNA and protein expression in the ischemic left ventricle had reached its peak by 20 min postligation in the rat AMI model (P < 0.05). TR-immunoreactive myocytes were observed in infarcted LV but were seldom seen in the right ventricle or in the normal heart. By 60 min, TR transcript levels had returned to control levels. We conclude that increased TR activation and expression in the infarcted LV after AMI may contribute to VA through a mechanism involving glibenclamide-sensitive potassium channels. PMID:18224254

Tang, Lilong; Deng, Chunyu; Long, Ming; Tang, Anli; Wu, Shulin; Dong, Yugang; Saravolatz, Louis D; Gardin, Julius M

2008-01-01

90

Thrombin binds to murine bone marrow-derived macrophages and enhances colony-stimulating factor-1-driven mitogenesis  

International Nuclear Information System (INIS)

The binding and mitogenic properties of thrombin have been established in various transformed cell lines. In such systems, thrombin induces cell division in the absence of exogenous growth factors, and the enzyme is considered to act directly as a mitogen. This study explores thrombin's interaction with nontransformed, growth factor-dependent cells. Binding of 125I-alpha-thrombin to colony-stimulating factor (CSF)-1-dependent bone marrow-derived macrophages is saturable, time-dependent, and displaceable by both unlabeled alpha-thrombin, and esterolytically inactive thrombin. Both dissociation studies of pre-bound radio-labeled thrombin and Scatchard analysis assisted by the program Ligand suggest adherence of thrombin-binding data to a multi-site model. There are an estimated 2 x 10(4) high affinity sites (Kd = 7 x 10(-9)M) and 2 x 10(6) low affinity sites (Kd = 9 x 10(-7)M) per cell. Quiescent bone marrow-derived macrophages were cultured with either 10(-8)M thrombin, 1000 units of CSF-1/ml, or both and [3H]thymidine incorporation was determined. Thrombin alone did not induce mitogenesis. CSF-1 induced mitogenesis with peak [3H] thymidine incorporation occurring 24 h after addition of the mitogen. This CSF-1-dependent mitogenic influence was enhanced greater than 2-fold by treatment with thrombin

91

Discovery of amino acid motifs for thrombin cleavage and validation using a model substrate.  

Science.gov (United States)

Understanding the active site preferences of an enzyme is critical to the design of effective inhibitors and to gaining insights into its mechanisms of action on substrates. While the subsite specificity of thrombin is understood, it is not clear whether the enzyme prefers individual amino acids at each subsite in isolation or prefers to cleave combinations of amino acids as a motif. To investigate whether preferred peptide motifs for cleavage could be identified for thrombin, we exposed a phage-displayed peptide library to thrombin. The resulting preferentially cleaved substrates were analyzed using the technique of association rule discovery. The results revealed that thrombin selected for amino acid motifs in cleavage sites. The contribution of these hypothetical motifs to substrate cleavage efficiency was further investigated using the B1 IgG-binding domain of streptococcal protein G as a model substrate. Introduction of a P(2)-P(1)' LRS thrombin cleavage sequence within a major loop of the protein led to cleavage of the protein by thrombin, with the cleavage efficiency increasing with the length of the loop. Introduction of further P(3)-P(1) and P(1)-P(1)'-P(3)' amino acid motifs into the loop region yielded greater cleavage efficiencies, suggesting that the susceptibility of a protein substrate to cleavage by thrombin is influenced by these motifs, perhaps because of cooperative effects between subsites closest to the scissile peptide bond. PMID:22050556

Ng, Natasha M; Pierce, James D; Webb, Geoffrey I; Ratnikov, Boris I; Wijeyewickrema, Lakshmi C; Duncan, Renee C; Robertson, Amy L; Bottomley, Stephen P; Boyd, Sarah E; Pike, Robert N

2011-12-01

92

Platelet activation and aggregation : the importance of thrombin activity--a laboratory model  

DEFF Research Database (Denmark)

This study introduces a new laboratory model of whole blood platelet aggregation stimulated by endogenously generated thrombin, and explores this aspect in haemophilia A in which impaired thrombin generation is a major hallmark. The method was established to measure platelet aggregation initiated by tissue factor evaluated by means of impedance aggregometry. Citrated whole blood from healthy volunteers and haemophilia A patients with the addition of inhibitors of the contact pathway and fibrin polymerization was evaluated. In healthy persons, a second wave of platelet aggregation was found to coincide with the thrombin burst and to be abolished by thrombin inhibitors. In this system, platelet aggregation in severe haemophilia A (n = 10) was found to be significantly decreased as compared with healthy individuals (912 ± 294 vs. 1917 ± 793 AU × min, P = 0.003), most probably due to the weak level of thrombin generation. For the first time, analysis of platelet aggregation as induced by endogenously generated thrombin was demonstrated. The new method makes it possible to explore the influence of the coagulation system on platelet function. In contrast to the general understanding, the data suggest that the impaired thrombin generation in haemophilia may affect platelet activation. Future studies will address whether our results may contribute to understanding differences in bleeding phenotypes and response to haemostatic substitution observed among patients.

Jensen, Maria Sander; Larsen, O H

2013-01-01

93

Thrombin selectively induces transcription of genes in human monocytes involved in inflammation and wound healing.  

Science.gov (United States)

Thrombin is essential for blood coagulation but functions also as a mediator of cellular signalling. Gene expression microarray experiments in human monocytes revealed thrombin-induced upregulation of a limited subset of genes, which are almost exclusively involved in inflammation and wound healing. Among these, the expression of F3 gene encoding for tissue factor (TF) was enhanced indicating that this physiological initiator of coagulation cascade may create a feed-forward loop to enhance blood coagulation. Activation of protease-activated receptor type 1 (PAR1) was shown to play a main role in promoting TF expression. Moreover, thrombin induced phosphorylation of ERK1/2, an event that is required for expression of thrombin-regulated genes. Thrombin also increased the expression of TF at the protein level in monocytes as evidenced by Western blot and immunostaining. Furthermore, FXa generation induced by thrombin-stimulated monocytes was abolished by a TF blocking antibody and therefore it is entirely attributable to the expression of tissue factor. This cellular activity of thrombin provides a new molecular link between coagulation, inflammation and wound healing. PMID:25057055

López, M L; Bruges, G; Crespo, G; Salazar, V; Deglesne, P-A; Schneider, H; Cabrera-Fuentes, H; Schmitz, M L; Preissner, K T

2014-11-01

94

Coiled-coil peptide based sensor for ultra-sensitive thrombin detection.  

Science.gov (United States)

Comb structured gold microelectrode array (CSGMA) functionalized with self-assembled monolayer of thiol terminated coiled-coil peptide (CCP) linked together by the thrombin specific cleavage site (Leu-Val-Pro-Arg-Gly-Ser) has been used to fabricate an ultrasensitive, disposable, electrochemical thrombin biosensor. CCP with thiol at one end provides the ease of CSGMA functionalization and the presence of thrombin specific peptide in the middle of coiled-coil peptide provides site for thrombin capture and detection. CCP/CSGMA electrodes were characterized using label-free electrochemical impedance (EIS) technique and exposed to solutions with different thrombin concentrations for its estimation. Results reveal that CCP/CSGMA electrodes have a limit of detection (LOD) of 10 fg/ml (28 fM) and are able to detect catalytic activity of thrombin within 30 min time frame. CCP/CSGMA electrodes were found to be selective against other IgG anti-bodies such as DO1 and HA. Thus, CCP/CSGMA electrodes provide high specificity toward thrombin detection and mechanistic details of binding and cleavage process. PMID:24355462

Kongsuphol, Patthara; Arya, Sunil K; Chung Wong, Chee; Polla, Lee Joseph; Park, Mi Kyoung

2014-05-15

95

Heparin coating of tantalum coronary stents reduces surface thrombin generation but not factor IXa generation.  

Science.gov (United States)

In the present study we used an in-vitro technique to examine initiation and propagation of blood coagulation at the surface of tantalum coronary stents exposed to flowing platelet-rich and platelet-free plasma. The time course of factor IXa production at the surface of the stent was not influenced by platelets. In spite of a significant factor IXa production, no thrombin activity was detected when the tantalum stent was exposed to platelet-free plasma; only when the stent was exposed to platelet-rich plasma was extensive thrombin production observed. These findings indicate that tantalum triggers blood coagulation, but that (adherent) platelets are essential for thrombin generation. Heparin-coated tantalum stents exposed to flowing platelet-rich plasma showed that factor IXa generation was slightly reduced compared with the bare stent. However, the heparin coating drastically delayed the onset of thrombin generation and largely reduced the steady-state production of thrombin. We found a clear relationship between the antithrombin binding capacity and the antithrombogenic potential of the heparin-coated stents. The mode of action of immobilized heparin is thought to abrogate thrombin generation by inhibiting thrombin-dependent positive feedback reactions at the surface of the coronary stent. PMID:9712292

Blezer, R; Cahalan, L; Cahalan, P T; Lindhout, T

1998-07-01

96

Increased thrombin generation and fibrinogen level after therapeutic plasma transfusion: relation to bleeding.  

Science.gov (United States)

In a clinical setting, fresh frozen plasma (FFP) is transfused to diluted patients with complicated surgery or trauma, as guided by prolonged conventional coagulation times or low fibrinogen levels. However, the limited sensitivity of these coagulation tests may restrict their use in measuring the effect of transfusion and hence predicting the risk of perioperative bleeding. We used the more sensitive, calibrated automated thrombogram (CAT) method to evaluate the result of therapeutic FFP transfusion to 51 patients with dilutional coagulopathy. Thrombin generation was measured in pre- and post-transfusion plasma samples in the presence of either platelets or phospholipids. For all patients, the transfusion led to higher plasma coagulation factor levels, a shortened activated partial thromboplastin time, and a significant increase in thrombin generation (peak height and endogenous thrombin potential). Interestingly, thrombin generation parameters and fibrinogen levels were higher in post-transfusion plasmas from patients who stopped bleeding (n = 32) than for patients with ongoing bleeding (n = 19). Plasmas from 15 of the 19 patients with ongoing bleeding were markedly low in either thrombin generation or fibrinogen level. We conclude that the thrombin generation method detects improved haemostatic activity after plasma transfusion. Furthermore, the data suggest that thrombin generation and fibrinogen are independent determinants of the risk of perioperative bleeding in this patient group. PMID:18217136

Schols, Saskia E M; van der Meijden, Paola E J; van Oerle, René; Curvers, Joyce; Heemskerk, Johan W M; van Pampus, Elisabeth C M

2008-01-01

97

Protamine sulfate down-regulates thrombin generation by inhibiting factor V activation.  

LENUS (Irish Health Repository)

Protamine sulfate is a positively charged polypeptide widely used to reverse heparin-induced anticoagulation. Paradoxically, prospective randomized trials have shown that protamine administration for heparin neutralization is associated with increased bleeding, particularly after cardiothoracic surgery with cardiopulmonary bypass. The molecular mechanism(s) through which protamine mediates this anticoagulant effect has not been defined. In vivo administration of pharmacologic doses of protamine to BALB\\/c mice significantly reduced plasma thrombin generation and prolonged tail-bleeding time (from 120 to 199 seconds). Similarly, in pooled normal human plasma, protamine caused significant dose-dependent prolongations of both prothrombin time and activated partial thromboplastin time. Protamine also markedly attenuated tissue factor-initiated thrombin generation in human plasma, causing a significant decrease in endogenous thrombin potential (41% +\\/- 7%). As expected, low-dose protamine effectively reversed the anticoagulant activity of unfractionated heparin in plasma. However, elevated protamine concentrations were associated with progressive dose-dependent reduction in thrombin generation. To assess the mechanism by which protamine mediates down-regulation of thrombin generation, the effect of protamine on factor V activation was assessed. Protamine was found to significantly reduce the rate of factor V activation by both thrombin and factor Xa. Protamine mediates its anticoagulant activity in plasma by down-regulation of thrombin generation via a novel mechanism, specifically inhibition of factor V activation.

Ni Ainle, Fionnuala

2009-08-20

98

Differentiation between proteolytic activation and autocatalytic conversion of human prothrombin. Activation of recombinant human prothrombin and recombinant D419N-prothrombin by snake venoms from Echis carinatus and Oxyuranus scutellatus.  

Science.gov (United States)

Recombinant human prothrombin (r-prothrombin) and recombinant mutant prothrombin with active site Asp419 substituted by Asn (D419N-prothrombin) were expressed in recombinant CHO cells, isolated and purified from the fermentation supernatant. The r-Prothrombin and D419N-prothrombin were digested by both Echis carinatus venom and Oxyuranus scutellatus venom. Prior to, during and after activation, generation of thrombin activity and the proteolytic degradation of the prothrombin polypeptide chain were analysed. Owing to the recombinant preparation and inactivity of D419N-prothrombin and its activation products, the proteolytic action of E.carinatus and O.scutellatus venoms could be studied without addition of thrombin inhibitor, without interference from autocatalytic digestion of prothrombin and in the absence of any other blood coagulation protease. The comparison between the activation of r-prothrombin and D419N-prothrombin by snake venoms permitted differentiation between proteolytic activation and autocatalytic conversion of prothrombin. Incubation of D419N-prothrombin with E.carinatus venom resulted in the generation of stable D419N-meizothrombin by hydrolysis of the peptide bond Arg320-Ile321. By contrast, O.scutellatus venom exhibited activity towards peptide bonds Arg320-Ile321 and Arg271-Thr272 and lower activity towards peptide bond Arg155-Ser156, thus converting D419-prothrombin into D419N-thrombin and also liberating Fragment-1, Fragment-2 and Fragment-1/2 activation peptide. Activation of r-prothrombin by E.carinatus and O.scutellatus venoms demonstrated the autocatalytic potential of prothrombin-derived molecules and indicated that meizothrombin hydrolysed the cleavage between Fragment-2 and thrombin A-chain in the meizothrombin molecule, but not in prothrombin, preferentially at position Arg284-Thr285. By contrast, both meizothrombin and thrombin exhibited no detectable activity towards peptide bond Arg320-Ile321 between thrombin A- and B-chain, although this site exhibits the optimum sequence for thrombin cleavage. PMID:8931132

Fischer, B E; Schlokat, U; Mitterer, A; Grillberger, L; Reiter, M; Mundt, W; Dorner, F; Eibl, J

1996-10-01

99

Thrombin-inhibiting nanoparticles rapidly constitute versatile and detectable anticlotting surfaces  

Science.gov (United States)

Restoring an antithrombotic surface to suppress ongoing thrombosis is an appealing strategy for treatment of acute cardiovascular disorders such as erosion of atherosclerotic plaque. An antithrombotic surface would present an alternative to systemic anticoagulation with attendant risks of bleeding. We have designed thrombin-targeted nanoparticles (NPs) that bind to sites of active clotting to extinguish local thrombin activity and inhibit platelet deposition while exhibiting only transient systemic anticoagulant effects. Perfluorocarbon nanoparticles (PFC NP) were functionalized with thrombin inhibitors (either D-phenylalanyl-L-prolyl-L-arginyl-chloromethyl ketone or bivalirudin) by covalent attachment of more than 15 000 inhibitors to each PFC NP. Fibrinopeptide A (FPA) ELISA demonstrated that thrombin-inhibiting NPs prevented cleavage of fibrinogen by both free and clot-bound thrombin. Magnetic resonance imaging (MRI) confirmed that a layer of thrombin-inhibiting NPs prevented growth of clots in vitro. Thrombin-inhibiting NPs were administered in vivo to C57BL6 mice subjected to laser injury of the carotid artery. NPs significantly delayed thrombotic occlusion of the artery, whereas an equivalent bolus of free inhibitor was ineffective. For thrombin-inhibiting NPs, only a short-lived (?10 min) systemic effect on bleeding time was observed, despite prolonged clot inhibition. Imaging and quantification of in vivo antithrombotic NP layers was demonstrated by MRI of the PFC NP. 19F MRI confirmed colocalization of particles with arterial thrombi, and quantitative 19F spectroscopy demonstrated specific binding and retention of thrombin-inhibiting NPs in injured arteries. The ability to rapidly form and image a new antithrombotic surface in acute vascular syndromes while minimizing risks of bleeding would permit a safer method of passivating active lesions than current systemic anticoagulant regimes.

Wheatley Myerson, Jacob; He, Li; Allen, John Stacy; Williams, Todd; Lanza, Gregory; Tollefsen, Douglas; Caruthers, Shelton; Wickline, Samuel

2014-09-01

100

Thrombin-inhibiting nanoparticles rapidly constitute versatile and detectable anticlotting surfaces.  

Science.gov (United States)

Restoring an antithrombotic surface to suppress ongoing thrombosis is an appealing strategy for treatment of acute cardiovascular disorders such as erosion of atherosclerotic plaque. An antithrombotic surface would present an alternative to systemic anticoagulation with attendant risks of bleeding. We have designed thrombin-targeted nanoparticles (NPs) that bind to sites of active clotting to extinguish local thrombin activity and inhibit platelet deposition while exhibiting only transient systemic anticoagulant effects. Perfluorocarbon nanoparticles (PFC NP) were functionalized with thrombin inhibitors (either D-phenylalanyl-L-prolyl-L-arginyl-chloromethyl ketone or bivalirudin) by covalent attachment of more than 15 000 inhibitors to each PFC NP. Fibrinopeptide A (FPA) ELISA demonstrated that thrombin-inhibiting NPs prevented cleavage of fibrinogen by both free and clot-bound thrombin. Magnetic resonance imaging (MRI) confirmed that a layer of thrombin-inhibiting NPs prevented growth of clots in vitro. Thrombin-inhibiting NPs were administered in vivo to C57BL6 mice subjected to laser injury of the carotid artery. NPs significantly delayed thrombotic occlusion of the artery, whereas an equivalent bolus of free inhibitor was ineffective. For thrombin-inhibiting NPs, only a short-lived (?10 min) systemic effect on bleeding time was observed, despite prolonged clot inhibition. Imaging and quantification of in vivo antithrombotic NP layers was demonstrated by MRI of the PFC NP. (19)F MRI confirmed colocalization of particles with arterial thrombi, and quantitative (19)F spectroscopy demonstrated specific binding and retention of thrombin-inhibiting NPs in injured arteries. The ability to rapidly form and image a new antithrombotic surface in acute vascular syndromes while minimizing risks of bleeding would permit a safer method of passivating active lesions than current systemic anticoagulant regimes. PMID:25200815

Myerson, Jacob Wheatley; He, Li; Allen, John Stacy; Williams, Todd; Lanza, Gregory; Tollefsen, Douglas; Caruthers, Shelton; Wickline, Samuel

2014-10-01

 
 
 
 
101

Measurement of dabigatran in standardly used clinical assays, whole blood viscoelastic coagulation, and thrombin generation assays.  

Science.gov (United States)

Dabigatran, a direct thrombin inhibitor, is increasingly used clinically as one of the new oral anticoagulants. This review summarizes the assays available to measure its activity and includes the relative sensitivity of the different assays for this agent. In addition to plasma-based clotting tests, assays commonly used in surgical/emergency settings, such as activated clotting time and thromboelastometry/thromboelastography, are reviewed. In addition, the thrombin generation assay is discussed as an important method to determine the potential risk of thrombosis or bleeding and its relevance to the measurement of direct thrombin inhibitors. PMID:25168938

van Ryn, Joanne; Grottke, Oliver; Spronk, Henri

2014-09-01

102

[Polymer coatings with immobilized thrombin and peptides: preparation and use for wound healing].  

Science.gov (United States)

Polymer dressings with encapsulated thrombin or synthetic peptides which can mimic thrombin action are employed for wound healing. Paper describes the method for preparation of these hydrogel composites of PVCL-CaAlg [poly(N-vinyl caprolactam-calcium alginate). The effect of encapsulated thrombin/peptides on tissue repair process have beet investigatat in vivo experiments using a mouse model of wound healing. The developed dressings accelerated wound healing: thascan be used as a basis for creation of novel formulations with controlled drug release for wound therapy. PMID:12698556

Markvicheva, E A; Kuptsova, S V; Rumsh, L D; Dugina, T N; Lange, M A; Chistov, I V; Strukova, S M; Zubov, V P

2002-01-01

103

[Topical corticosteroids versus topical inhibitors of calcineurin].  

Science.gov (United States)

Topical corticosteroids (TCC) have significantly shaped dermatological therapy for five decades. A few months ago the TCC were joined by competition, the topical inhibitors of calcineurin (TIC), wrongly termed topical immunomodulators. The present paper reviews the pharmacological effects and clinical efficacy of TIC, compares the risks, benefits and costs of those two groups of topical drugs and develops a position on the use of TIC. While TIC have ushered in a new era of topical anti-inflammatory therapy, the age of TCC is far from over. PMID:12669205

Niedner, R

2003-04-01

104

THE EVALUATION OF CLOTTING TIME IN BOVINE THROMBIN, REPTILASE ® , AND THROMBIN-LIKE FRACTION OF Crotalus durissus terrificus VENOM USING BOVINE, EQUINE, OVINE, BUBALINE AND HUMAN CRYOPRECIPITATES  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english The objective of this study was to evaluate the effects of the thrombin-like fraction of Crotalus durissus terrificus venom, Reptilase , and bovine thrombin of fibrinogen pools on bovine, equine, ovine, bubaline and human cryoprecipitates. The authors [...] also made a comparative study between animal and human cryoprecipitates to see if any there was any possibility of future use in medicine. Fibrinogen levels in cryoprecipitate were studied using 48 blood samples obtained as follows:12 samples from humans, 9 from bovine, 10 from equine, 10 from ovine and 7 from bubaline. The results obtained showed average levels of 375.50 mg % for humans, 218.33 mg % for bovine, 240.80 mg % for equine, 267.70 mg % for ovine and 664.00 mg % for bubaline. Upon the formation of pools of human and animal fibrinogens, the following results were obtained: 435 mg % for humans, 444 mg % for bovine, 337 mg % for equine, 390 mg % for ovine and 530 mg % for bubaline. Statistical analysis (using the analysis of variance for entirely randomized experiment for the calculation of F statistics) demonstrated that the bubaline fibrinogen level was higher than that of human, and both were higher than those of ovine, equine, and bovine. Clotting times were determined using different dilutions of bovine thrombin, thrombin-like fraction of Crotalus durissus terrificus venom, and Reptilase . Comparing these clotting times, results for human and bovine were found to be very similar, whereas using equine, ovine, and bubaline the results above a dilution of 1:3 were markedly different. The results obtained permitted the following conclusions to be drawn show that: 1) bovine thrombin presented better interactivity with fibrinogen extracted both from human and bovine cryoprecipitates; 2) there was similar behavior when bovine thrombin was substituted for Reptilase and for the thrombin-like fraction of Crotalus durissus terrificus venom; 3) cryoprecipitate from bovine can, in special circumstances, substitute human cryoprecipitate in medical practice; 4) human and bovine cryoprecipitates can be used with both Reptilase and Crotalus durissus terrificus fractions using a dilution up to 1:5; 5) the use of bovine cryoprecipitate can be recommended using either bovine thrombin, Reptilase , or thrombin-like fraction of Crotalus durissus terrificus venom.

I. A., THOMAZINI-SANTOS; M. J. S. M., GIANNINI; E., TOSCANO; P.E.A., MACHADO; C. R. G., LIMA; B., BARRAVIERA.

105

Interaction of hirudin with thrombin: Identification of a minimal binding domain of hirudin that inhibits clotting activity  

Energy Technology Data Exchange (ETDEWEB)

Hirudin, isolated from the European leech Hirudo medicinalis, is a potent inhibitor of thrombin, forming an almost irreversible thrombin-hirudin complex. Previously, the authors have shown that the carboxyl terminus of hirudin (residues 45-65) inhibits clotting activity and without binding to the catalytic site of thrombin. In the present study, a series of peptides corresponding to this carboxyl-terminal region of hirudin have been synthesized, and their anticoagulant activity and binding properties to thrombin were examined. Binding was assessed by their ability to displace {sup 125}I-hirudin 45-65 from Sepharose-immobilized thrombin and by isolation of peptide-thrombin complexes. They show that the carboxyl-terminal 10 amino acid residues 56-65 (Phe-Glu-Glu-Ile-Pro-Glu-Glu-Tyr-Leu-Gln) are minimally required for binding to thrombin and inhibition of clotting. Phe-56 was critical for maintaining anticoagulant activity as demonstrated by the loss of activity when Phe-56 was substituted with D-Phe, Glu, or Leu. In addition, they found that the binding of the carboxyl-terminal peptide of hirudin with thrombin was associated with a significant conformational change of thrombin as judged by circular dichroism. This conformational change might be responsible for the loss of clotting activity of thrombin.

Mao, S.J.T.; Yates, M.T.; Owen, T.J.; Krstenansky, J.L. (Merrell Dow Research Institute, Cincinnati, OH (USA))

1988-10-18

106

Behaviour of homologous 125I fibrinogen after thrombin and ancrod infusion in rabbits  

International Nuclear Information System (INIS)

The behaviour of radioactively labelled fibrinogen after infusion of thrombin or ancrod is investigated. Common factors and differences in the behaviour of fibrinogen after infusion of these two enzymes, which act proteolytically on the fibrinogen, are dealt with. Rabbits received an i.v. injection of homologous 125I-fibrinogen 3 days before ancrod or thrombin infusion. On the day of the experiments, one group of animals received an ancrod infusion (1.5 U/kg body weight for 30 minutes), the other a thrombin infusion (600 U/kg body weight for 60 minutes). Intravenous ancrod and thrombin infusions lowered the fibrinogen level to 30% or 50% of the initial value due to intravascular coagulation. About 50% of the 125I fibrinogen was transformed after ancrod exposure into a non-coagulating fraction of fibrinogen derivatives which produces no fibrinolytic decomposition products. (orig./AJ)

107

Theoretical modeling and experimental validation of surface stress in thrombin aptasensor.  

Science.gov (United States)

Adsorption of target molecules on the immobilized microcantilever surface produced beam displacement due to the differential surface stress generated between the immobilized and non-immobilized surface. Surface stress is caused by the intermolecular forces between the molecules. Van der Waals, electrostatic forces, hydrogen bonding, hydrophobic effect and steric hindrance are some of the intermolecular forces involved. A theoretical framework describing the adsorption-induced microcantilever displacement is derived in this paper. Experimental displacement of thrombin aptamer-thrombin interactions was carried out. The relation between the electrostatic interactions involved between adsorbates (thrombin) as well as adsorbates and substrates (thrombin aptamer) and the microcantilever beam displacement utilizing the proposed mathematical model was quantified and compared to the experimental value. This exercise is important to aid the designers in microcantilever sensing performance optimization. PMID:25122838

Lim, Yang Choon; Kouzani, Abbas Z; Kaynak, Akif; Dai, Xiujuan J; Littlefair, Guy; Duan, Wei

2014-12-01

108

A fluorescent sandwich assay for thrombin using aptamer modified magnetic beads and quantum dots  

International Nuclear Information System (INIS)

We describe an aptamer-based sandwich assay for thrombin by using a pair of thrombin-binding aptamers, namely one 15-mer aptamer (denoted as Apt15) and one 29-mer aptamer (denoted as Apt29). Either Apt29 or Apt15 can be used as capture aptamers on magnetic beads or reporter aptamers on the quantum dots to form the sandwich complex. Detection of thrombin is achieved by the fluorescent measurement of quantum dots in the sandwich complex. The choice of capture aptamers and reporter aptamers, and the effect of the addition order of the aptamers modified magnetic beads and the aptamers modified quantum dots were investigated. Detection of 0.05 nM thrombin was accomplished. The proteins hemoglobin, lysozyme, and transferrin did not interfere in this assay. (author)

109

Human Thrombin Detection Through a Sandwich Aptamer Microarray: Interaction Analysis in Solution and in Solid Phase  

Directory of Open Access Journals (Sweden)

Full Text Available We have developed an aptamer-based microarray for human thrombin detection exploiting two non-overlapping DNA thrombin aptamers recognizing different exosites of the target protein. The 15-mer aptamer (TBA1 binds the fibrinogen-binding site, whereas the 29-mer aptamer (TBA2 binds the heparin binding domain. Extensive analysis on the complex formation between human thrombin and modified aptamers was performed by Electrophoresis Mobility Shift Assay (EMSA, in order to verify in solution whether the chemical modifications introduced would affect aptamers/protein recognition. The validated system was then applied to the aptamer microarray, using the solid phase system devised by the solution studies. Finally, the best procedure for Sandwich Aptamer Microarray (SAM and the specificity of the sandwich formation for the developed aptasensor for human thrombin were optimized.

Alice Sosic

2011-10-01

110

Direct detection of aptamer-thrombin binding via surface-enhanced Raman spectroscopy  

Science.gov (United States)

In this study, we exploit the sensitivity offered by surface-enhanced Raman scattering (SERS) for the direct detection of thrombin using the thrombin-binding aptamer (TBA) as molecular receptor. The technique utilizes immobilized silver nanoparticles that are functionalized with thiolated thrombin-specific binding aptamer, a 15-mer (5'-GGTTGGTGTGGTTGG-3') quadruplex forming oligonucleotide. In addition to the Raman vibrational bands corresponding to the aptamer and blocking agent, new peaks (mainly at 1140, 1540, and 1635 cm-1) that are characteristic of the protein are observed upon binding of thrombin. These spectral changes are not observed when the aptamer-nanoparticle assembly is exposed to a nonbinding protein such as bovine serum albumin (BSA). This methodology could be further used for the development of label-free biosensors for direct detection of proteins and other molecules of interest for which aptamers are available.

Pagba, Cynthia V.; Lane, Stephen M.; Cho, Hansang; Wachsmann-Hogiu, Sebastian

2010-07-01

111

Thrombin-induced CCN2 expression as a target for anti-fibrotic therapy in scleroderma  

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Scleroderma (systemic sclerosis, SSc) is a fibrotic disease for which there is no therapy. CCN2 (connective tissue growth factor, CTGF) is a marker and mediator of fibrosis. Previously, it has been shown that thrombin induces CCN2 expression in fibroblasts. In a recent fascinating report, Bogatkevich et al. (Arthritis Rheum 60:3455–3464, 2009) show that dabigatran, an inhibitor of thrombin action, blocks the overexpression of CCN2 by scleroderma fibroblasts and reverses the contractile phen...

Leask, Andrew

2010-01-01

112

PKC isoenzymes differentially modulate the effect of thrombin on MAPK-dependent RPE proliferation.  

Science.gov (United States)

Thrombin signalling through PAR (protease-activated receptor)-1 is involved in cellular processes, such as proliferation, differentiation and cell survival. Following traumatic injury to the eye, thrombin signalling may participate in disorders, such as PVR (proliferative vitreoretinopathy), a human eye disease characterized by the uncontrolled proliferation, transdifferentiation and migration of otherwise quiescent RPE (retinal pigment epithelium) cells. PARs activate the Ras/Raf/MEK/ERK MAPK pathway (where ERK is extracellular-signal-regulated kinase, MAPK is mitogen-activated protein kinase and MEK is MAPK/ERK kinase) through the activation of G(alpha) and G(betagamma) heterotrimeric G-proteins, and the downstream stimulation of the PLC (phospholipase C)-beta/PKC (protein kinase C) and PI3K (phosphoinositide 3-kinase) signalling axis. In the present study, we examined the molecular signalling involved in thrombin-induced RPE cell proliferation, using rat RPE cells in culture as a model system for PVR pathogenesis. Our results showed that thrombin activation of PAR-1 induces RPE cell proliferation through Ras-independent activation of the Raf/MEK/ERK1/2 MAPK signalling cascade. Pharmacological analysis revealed that the activation of 'conventional' PKC isoforms is essential for proliferation, although thrombin-induced phosphorylation of ERK1/2 requires the activation of atypical PKCzeta by PI3K. Consistently, thrombin-induced ERK1/2 activation and RPE cell proliferation were prevented completely by PI3K or PKCzeta inhibition. These results suggest that thrombin induces RPE cell proliferation by joint activation of PLC-dependent and atypical PKC isoforms and the Ras-independent downstream stimulation of the Raf/MEK/ERK1/2 MAPK cascade. The present study is the first report demonstrating directly thrombin-induced ERK phosphorylation in the RPE, and the involvement of atypical PKCzeta in this process. PMID:18636965

Palma-Nicolas, Jose P; López, Edith; López-Colomé, Ana María

2008-12-01

113

Efficacy of fibrinogen/thrombin-coated equine collagen patch in controlling lymphatic leaks.  

Science.gov (United States)

We report the use of fibrinogen/thrombin-coated equine collagen patch (Tachosil(®) ) as a sealant agent in six patients who underwent heart surgery for congenital heart disease (CHD) and developed an intraoperative lymphatic leakage detected at the time of surgery. The use of fibrinogen/thrombin-coated equine collagen patch proved to be safe and effective in preventing the development of postoperative chylothorax. PMID:22583120

Vida, Vladimiro L; Padalino, Massimo A; Barzon, Elisa; Stellin, Giovanni

2012-07-01

114

Rapid purification of high purity thrombin and preparation of a novel hemostat for clinical purposes  

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Thrombin was prepared from crude prothrombin enriched plasma by activation using Russell’s viper venom. Prothrombin was prepared by barium sulphate adsorption and elution of prothrombin enriched fraction using high concentrations of sodium citrate. This fraction was directly activated with venom and thrombin was purified by SP Sepharose ion-exchange chromatography and subsequently over Phenyl-sepharose column. This product exhibits a purity of >98% with an activity of at least 6000U/mg or h...

Turaga, Krishna Kumar; Chakradhara Rao, P.; Sripad, G.

2008-01-01

115

A label-free electrochemical aptasensor for sensitive thrombin detection in whole blood  

International Nuclear Information System (INIS)

In this paper, we reported a novel label-free electrochemical aptasensors for thrombin detection in whole blood using self-assembled multilayers with carboxymethyl-PEG-carboxymethyl (CM-PEG-CM) and thrombin-binding aptamer (TBA). In the sensing strategy, CM-PEG-CM and TBA were assembled on the electrode surface via covalent binding. In the presence of target, the TBA on the outermost layer of the self-assembled multilayer would catch the target on the electrode interface, which makes a barrier for electrons and inhibits the electro-transfer, resulting in the decreased DPV signals. Using this strategy, a wide detection range (1 pM–160 nM) for target thrombin was obtained, with a low detection limit of 1.56 × 10?14 M. The control experiments were also carried out by using bull serum albumin (BSA) and lysozyme in the absence of thrombin. The results showed that the aptasensors had good specificity, stability and reproducibility to thrombin. Moreover, the aptasensors could be used for detection of thrombin in whole blood which could provide a promising platform for fabrication of aptamer based biosensors in clinical application

116

Ultrasensitive electrochemical aptasensor for thrombin based on the amplification of aptamer-AuNPs-HRP conjugates.  

Science.gov (United States)

Successful development of an ultrasensitive and highly specific electrochemical aptasensor for thrombin based on amplification of aptamer-gold nanoparticles-horseradish peroxidase (aptamer-AuNPs-HRP) conjugates was reported. In this electrochemical protocol, aptamer1 (Apt1) was immobilized on core/shell Fe(3)O(4)/Au magnetic nanoparticles (AuMNPs) and served as capture probe. Aptamer2 (Apt2) was dual labeled with AuNPs and HRP and used as detection probe. In the presence of thrombin, the sandwich format of AuMNPs-Apt1/thrombin/Apt2-AuNPs-HRP was fabricated. Remarkable signal amplification was realized by taking the advantage of AuNPs and catalytic reactions of HRP. Other proteins, such as human serum albumin, lysozyme, fibrinogen, and IgG did not show significant interference with the assay for thrombin. Linear response to thrombin concentration in the range of 0.1-60 pM and lower detection limit down to 30 fM (S/N=3) was obtained with the proposed method. This electrochemical aptasensor is simple, rapid (the whole detection period for a thrombin sample is less than 35 min), sensitive and highly specific, it shows promising potential in protein detection and disease diagnosis. PMID:21030239

Zhao, Jie; Zhang, Youyu; Li, Haitao; Wen, Yanqing; Fan, Xiaoyu; Lin, Fanbo; Tan, Liang; Yao, Shouzhuo

2011-01-15

117

Identification of the site of binding of sulfated, low molecular weight lignins on thrombin.  

Science.gov (United States)

Sulfated, low molecular weight lignins (LMWLs), designed recently as macromolecular mimetics of the low molecular weight heparins (LMWHs), were found to exhibit a novel allosteric mechanism of inhibition of human thrombin, factor Xa and plasmin, which translates into potent human blood anticoagulation potential. To identify the site of binding of sulfated LMWLs, a panel of site-directed thrombin mutants was studied. Substitution of alanine for Arg(93) or Arg(175) induced a 7-8-fold decrease in inhibition potency, while Arg(165)Ala, Lys(169)Ala, Arg(173)Ala and Arg(233)Ala thrombin mutants displayed a 2-4-fold decrease. Other exosite 2 residues including those that play an important role in heparin binding, such as Arg(101), Lys(235), Lys(236) and Lys(240), did not induce any deficiency in sulfated LMWL activity. Thrombin mutants with multiple alanine substitution of basic residues showed a progressively greater defect in inhibition potency. Comparison of thrombin, factor Xa, factor IXa and factor VIIa primary sequences reiterated Arg(93) and Arg(175) as residues likely to be targeted by sulfated LMWLs. The identification of a novel site on thrombin with capability of allosteric modulation is expected to greatly assist the design of new regulators based on the sulfated LMWL scaffold. PMID:21893043

Abdel Aziz, May H; Mosier, Philip D; Desai, Umesh R

2011-09-23

118

Pulsatile equibiaxial stretch inhibits thrombin-induced RhoA and NF-?B activation  

International Nuclear Information System (INIS)

This study investigated interactions between the effects of mechanical stretch and thrombin on RhoA activation in rat aortic smooth muscle cells (RASMC). Equibiaxial, pulsatile stretch, or thrombin produced a significant increase in RhoA activation. Surprisingly, in combination, 30 min of stretch inhibited the ability of thrombin to activate RhoA. NO donors and 8-bromo-cGMP significantly inhibited thrombin-induced RhoA activation. Interestingly, the nitric oxide synthase (NOS) inhibitor L-NAME increased basal RhoA activity, suggesting that NOS activity exerts a tonic inhibition on RhoA. Stretching RASMC increases nitrite production, consistent with the idea that NO contributes to the inhibitory effects of stretch. Thrombin stimulates MAP kinase and NF-?B pathways through Rho and these responses were blocked by 8-bromo-cGMP or stretch and restored by L-NAME. These data suggest that stretch, acting through NO and cGMP, can prevent the ability of thrombin to stimulate Rho signaling pathways that contribute to pathophysiological proliferative and inflammatory responses

119

A colorimetric aptamer biosensor based on cationic polymer and gold nanoparticles for the ultrasensitive detection of thrombin.  

Science.gov (United States)

A colorimetric assay for the ultrasensitive determination of thrombin based on cationic polymer and gold nanoparticles was presented, in which unmodified gold nanoparticles (AuNPs) was used as probes and 21-mer thrombin-binding aptamer (TBA) as sensing elements. Upon the addition of thrombin, TBA interacted specifically with thrombin to form a G-quadruplex structure. As a result, the conformation change facilitated the cationic polymer, poly(diallyldimethylammonium chloride) (PDDA)-induced AuNP aggregation. Thus, the visible change in color from wine-red to blue-purple was readily seen by the naked eye. The colorimetric sensor could detect thrombin down to 1 pM with high selectivity in the presence of other interferring proteins. Furthermore, the assay was successfully employed to determine thrombin in human serum sample, which suggested its great potential for diagnostic purposes. PMID:24463195

Chen, Zhengbo; Tan, Yuan; Zhang, Chenmeng; Yin, Lu; Ma, He; Ye, Nengsheng; Qiang, Hong; Lin, Yuqing

2014-06-15

120

Topical report review status  

International Nuclear Information System (INIS)

This report provides industry with procedures for submitting topical reports, guidance on how the U.S. Nuclear Regulatory Commission (NRC) processes and responds to topical report submittals, and an accounting, with review schedules, of all topical reports currently accepted for review schedules, of all topical reports currently accepted for review by the NRC. This report will be published annually. Each sponsoring organization with one or more topical reports accepted for review copies

 
 
 
 
121

Brasiliensin: A novel intestinal thrombin inhibitor from Triatoma brasiliensis (Hemiptera: Reduviidae) with an important role in blood intake  

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Every hematophagous invertebrate studied to date produces at least one inhibitor of coagulation. Among these, thrombin inhibitors have most frequently been isolated. In order to study the thrombin inhibitor from Triatoma brasiliensis and its biological significance for the bug, we sequenced the corresponding gene and evaluated its biological function. The T. brasiliensis intestinal thrombin inhibitor, termed brasiliensin, was sequenced and primers were designed to synthesize double strand RNA...

Araujo, R. N.; Campos, I. T. N.; Tanaka, A. S.; Santos, A.; Gontijo, N. F.; Lehane, M. J.; Pereira, M. H.

2007-01-01

122

Recombinant Technology and Probiotics  

Directory of Open Access Journals (Sweden)

Full Text Available Recombinant technology has led the way to monumental advances in the development of useful molecules, including the development of safe probiotics. The development of novel approaches using recombinant technology and probiotics that allow accurate targeting of therapeutics to the mucosa is an interesting area of research. The creation and use of recombinant probiotics expressing recombinantovalbumin, recombinant ovalbumin mutants and yet-to-be-designed recombinant hypo/non-allergenic molecules offer the opportunity to further investigate their effects for food, nutrition, environment andhealth. This review highlights advances in native probiotics and recombinant probiotics expressing native and recombinant molecules for food, nutrition, environment and health.

Icy D’Silva

2011-09-01

123

External quality assessment for thrombin generation tests: an exploration.  

Science.gov (United States)

External Quality Control of Diagnostic Assays and Tests (ECAT) surveys on thrombin generation rests (TGTs) have identified that various tests show a more than 30-fold difference in time to peak (TTP). The survey included pooled normal plasmas, microparticle (MP)-depleted plasmas, and factor (F)XII-deficient patient plasma. Between 4 and 11 laboratories participated per test; analyzed were a time (TTP) and quantity variable (AUC). MP depletion of plasma showed a progressive increase in TTP up to 29% with a decreasing tissue factor. The same was found for the AUC with the largest decrease of 38%. It was observed that MP depletion showed large individual differences. The FXII-deficient plasma showed no effect on TTP for rapid tests, but for slow tests it increased from 248 to 331%. The AUC declined gradually the slower the test, reaching a decline of 85%. The effects of FXII deficiency were not mimicked by the addition of corn trypsin inhibitor but were confirmed by inhibiting activated FXI. Interlaboratory variability was between 11% and 57% for all methods, showing differences mainly related to the used normal pooled plasma. The different sensitivities of TGTs to MPs and contact activation predict that they will associate differently with clinical situations, according to which of these aspects are important. Future ECAT surveys should include samples with variation in MPs and contact activation to match with features of the TGT variants. PMID:20979000

Kluft, Cornelis; Meijer, Piet

2010-10-01

124

Thrombin generation as a predictor of radiotherapy induced skin erythema  

International Nuclear Information System (INIS)

Background and purpose: Biological mechanisms underlying radiation induced erythema remain largely unknown, with no simple way to accurately predict or prevent extreme cases. Based on the recent findings in patients suffering from chronic urticaria, we sought to determine if similar mechanisms of hypercoagulation contributed to comparable skin reactions during radiotherapy. Materials and methods: Plasma levels of prothrombin factor 1+2 (F1+2), D-dimers and plasminogen activator inhibitor-1 (Pai-1) were tested in 32 women undergoing irradiation following breast conserving surgery for early breast cancer. Reflectance spectrophotometry was used to objectively assess erythema throughout the treatment by measuring the amount of light reflected from the skin surface as a function of wavelength. Correlations between peak levels of erythema and plasma biomarkers were then assessed. Results: Individual peak reflectance readings generally occurred between day 29 of treatment and 2 weeks post radiotherapy, and represented a median increase of 66% (range: 11-146%; p < 0.001) from baseline. Peak reflectance correlated with F1+2 and Pai-1 levels measured both at baseline and day 29 of treatment, and multivariate analysis indicated that these two baseline measurements were the best predictors of peak reflectance, accounting for 59% of the variability in erythema (p = 0.000004). Conclusions: Patients with signs of intravascular thrombin generation are at higher risk of radiotherapy-induced skin reactions, providing a new therapeutic avenue for possibly predicting and preventing this side effect of cancer treatment

125

Purification and Characterization of Human Thrombin Activatable Fibrinolysis Inhibitor (TAFI)  

DEFF Research Database (Denmark)

Thrombin Activatable Fibrinolysis inhibitor (TAFI) is a basic carboxypeptidase, circulating in plasma as an enzymatic inactive precursor. TAFI shares ~40% overall sequence identity with pancreas Carboxypeptidase B (PCPB) with the activation peptide being less conserved. Following activation of TAFI we observed a change in the isoelectric point (IP) of TAFIa. The IP of TAFIa was significantly more basic than the IP of TAFI. This was not observed in PCPB. Due to the change in IP, we have investigated the structural basis for this observation by mapping the N- and O-linked glycans. TAFI has 5 potential N-linked glycosylation sites, four of them located on the activation peptide. Only one potential O-linked glycosylation site is seen. In addition, TAFIa is unstable and loose enzymatic activity quickly. This is in contrast to the homologous PCPB. The structural basis for this observation is not known but could be caused by differences in thermodynamic stability. Disulfides are a major contributor to the structuralintegrity of proteins and disulfide permutations could explain the difference in enzymatic stability. To test this hypothesis we have identified the disulfide pattern of the eight cysteines in TAFI. In this study we have purified and characterized TAFI from human plasma.

Skottrup, Peter

126

The variable detergent sensitivity of proteases that are utilized for recombinant protein affinity tag removal.  

Science.gov (United States)

Recombinant proteins typically include one or more affinity tags to facilitate purification and/or detection. Expression constructs with affinity tags often include an engineered protease site for tag removal. Like other enzymes, the activities of proteases can be affected by buffer conditions. The buffers used for integral membrane proteins contain detergents, which are required to maintain protein solubility. We examined the detergent sensitivity of six commonly-used proteases (enterokinase, factor Xa, human rhinovirus 3C protease, SUMOstar, tobacco etch virus protease, and thrombin) by use of a panel of 94 individual detergents. Thrombin activity was insensitive to the entire panel of detergents, thus suggesting it as the optimal choice for use with membrane proteins. Enterokinase and factor Xa were only affected by a small number of detergents, making them good choices as well. PMID:21539919

Vergis, James M; Wiener, Michael C

2011-08-01

127

Diclofenac Topical (actinic keratosis)  

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Diclofenac topical gel (Solaraze; generic) is used to treat actinic keratosis (flat, scaly growths on the skin caused by ... products for osteoarthritis, read the monograph entitled diclofenac topical (osteoarthritis pain).

128

Ciclopirox Topical Solution  

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Ciclopirox topical solution is used along with regular nail trimming to treat fungal infections of the fingernails and toenails ( ... using ciclopirox without talking to your doctor.Ciclopirox topical solution will work best if you trim your ...

129

Diclofenac Topical (osteoarthritis pain)  

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Diclofenac topical gel (Voltaren) is used to relieve pain from osteoarthritis (arthritis caused by a breakdown of the lining ... knees, ankles, feet, elbows, wrists, and hands. Diclofenac topical liquid (Pennsaid) is used to relieve osteoarthritis pain ...

130

cDNA cloning and high-level expression of a thrombin-like enzyme from Agkistrodon acutus venom.  

Science.gov (United States)

Agkistrodon acutus (Guenther), a poisonous snake species of the family of Crotalidae, is mainly found south of the Yellow River in China. The main symptom of this poison is massive hemorrhage, in which thrombin-like enzymes (TLEs) in the venom play an important role. TLEs are abundant, especially in the venom of A. acutus. We isolated the total RNA from the venom gland tissue of A. acutus and amplified the cDNAs of the TLEs using reverse transcription-polymerase chain reaction (RT-PCR). The cDNAs were cloned into vector pThioHis B and were expressed as fusion proteins in the form of inclusion bodies, which accounted for nearly 50% of the total cell proteins. The inclusion bodies were washed, dissolved, refolded and purified by affinity chromatography. The purity was higher than 97%, as indicated by capillary zone electrophoresis. The renatured recombinant enzyme exhibited arginine esterase activity, as tested by the BAEE method, and also showed a fibrinogen cleavage effect, as detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). This method provides a fast and convenient system for studying the structure-function relationships in TLE isoenzymes, and also a practical way for mass production of TLEs in the pharmaceutical industry. PMID:12808469

Zha, X D; Ren, B; Liu, J; Xu, K S

2003-05-01

131

Cancer Topic Searches  

Science.gov (United States)

Cancer Topic Searches are prepared literature searches of the National Library of Medicine's PubMed database. Literature citations on more than 100 different topics can be obtained. All citations on a specific topic can be retrieved, or the results can be limited by selecting one of three date ranges.

132

Freshman Health Topics  

Science.gov (United States)

This article examines a cluster of health topics that are frequently selected by students in lower division classes. Topics address issues relating to addictive substances, including alcohol and tobacco, eating disorders, obesity, and dieting. Analysis of the topics examines their interrelationships and organization in the reference literature.…

Hovde, Karen

2011-01-01

133

Protease-activated receptor-1 (thrombin receptor) is expressed in mesenchymal portions of human hair follicle.  

Science.gov (United States)

Protease nexin-1, a serine protease inhibitor, is expressed specifically in the dermal papilla (DP) of anagen hair follicles and is suggested to be one of the modulators of the cyclic growth of hair follicles. Accumulating evidence has shown that protease nexin-1 plays its biologic role by inhibiting thrombin action in various systems other than the hair follicle. Thrombin has various physiologic functions including blood coagulation cascade, mostly via activation of protease-activated receptors (PAR). In this study, we investigated the expression of PAR mRNA using RT-PCR in dissected human hair follicles. We showed that PAR-1 mRNA was expressed specifically in the mesenchymal portions, including DP and connective tissue sheath, of anagen hair follicles. Furthermore, immunoreactivity for PAR-1 was detected in the DP and lower portion of connective tissue sheath in the anagen and catagen phases and in the DP of telogen hair follicles. Because only a pharmacologic level (100 nM) of thrombin significantly stimulated cell proliferation and DNA synthesis of the cultured dermal papilla cells, thrombin does not seem to have a mitogenic effect on dermal papilla cells physiologically. These results raise the possibility that thrombin is involved in the cyclic hair growth through its receptor of PAR-1. PMID:14632180

Anan, T; Sonoda, T; Asada, Y; Kurata, S; Takayasu, S

2003-10-01

134

Purification and variability in thrombin-like activity of Bothrops atrox venom from different geographic regions.  

Science.gov (United States)

Bothrops atrox snake venoms from two different Amazon regions, i.e., Manaus, AM (3 degrees 0.6'40"S; 60 degrees 0.1'6.0"W) and Tucurui, PA (3 degrees 0.42'30"S; 49 degrees 0.41'45"W), were analyzed with respect to the thrombin-like activity component by elution profile on gel-filtration and reverse phase HPLC chromatography, electrophoretic mobility on SDS-PAGE, and enzymatic activity on fibrinogen. Despite some individual discrepancies among venom specimens, the thrombin-like activity present in the Manaus pool was eluted earlier compared with the Tucurui pool but its enzymatic specific activity on thrombin was lower (s.a. = 6.0) than that observed in the Tucurui pool (s.a. = 134.0). However, the electrophoretic mobilities of the pools were similar, with most protein bands being concentrated around three main regions, i.e., protein bands with an apparent mr of 100 kDa, of 38-37 kDa and 30 kDa. However, no significant differences were observed in amidolytic activity on the synthetic substrate Tos-Gly-Pro-Arg-pNa, and there was no correlation between thrombin-like and amidolytic activities. A 32 kDa protein endowed with thrombin-like activity and specific activity of 2444 recognized and neutralized by horse anti-B. atrox antivenom, was purified by the successive use of gel filtration, electrofocusing and reverse phase HPLC. PMID:9620574

Cavinato, R A; Remold, H; Kipnis, T L

1998-02-01

135

Ethanol interferes with thrombin mediated changes in the morphology and cytoskeleton of human vascular endothelial cells  

Energy Technology Data Exchange (ETDEWEB)

The effect of physiological concentrations of ethanol (EtOH) on the response of human vascular endothelial cells (EC) to thrombin was examined Treatment of EC with EtOH concentrations of 20-85mM for 2-10 min. produced no significant changes in the morphology of 3- and 4-day monolayers established on fibronectin coated polystyrene. When examined immunofluorescently no significantly changes in the microfilament or microtubule structures were seen. Exposure of EC monolayers to 0.5 and 1 U/ml of thrombin for 1-60 minutes causes a concentration and time dependent monolayer retraction, evidenced by a general decrease in cell size, increase in visible gaps in the monolayer and redistribution of the microtubule and microfilament networks. Pretreatment of EC monolayers with EtOH for 3-5 minutes prior to addition of thrombin prevents the changes seen with thrombin alone. Immunofluorescent examination of the microfilament and microtubule structures suggests than EtOH may act in part via the microtubule network, which appears to be disorganized/disrupted when the EC are exposed to EtOH and then thrombin. Colchicine studies show that EC which have been pretreated with EtOH respond to colchicine differently then cells which have not previously seen EtOH. These data suggest that EtOH may alter EC monolayer responsiveness either by indirect changes which are reflected in cytoskeletal disorganization or possibly by direct influence on the cytoskeleton.

Pratt, K.J.; Rubin, R.; Hoek, J.; Williams, S.K. (Thomas Jefferson Univ., Philadelphia, PA (United States))

1991-03-15

136

Interaction of viper venom serine peptidases with thrombin receptors on human platelets.  

Science.gov (United States)

The serine peptidases, thrombocytin and PA-BJ, isolated from the venom of Bothrops atrox and Bothrops jararaca, respectively, induce platelet aggregation and granule secretion without clotting fibrinogen. The specific platelet aggregation activity of each enzyme was about 15 times lower than that of thrombin. This activity was blocked by monoclonal antibodies recognizing protease activated receptor 1 (PAR1) and by heparin, but not by hirudin nor thrombomodulin. Both enzymes induced calcium mobilization in platelets and desensitized platelets to the action of thrombin and the SFLLRN peptide. We compared the effect of thrombin, PA-BJ, and thrombocytin on the degradation of the soluble N-terminal domain of the PAR1 receptor. The major cleavage site by thrombin and both viper enzymes was Arg41-Ser42. In addition, a rapid cleavage of the peptide bond at Arg46-Asn47 by the viper enzymes was observed, resulting in the inactivation of the tethered ligand. PA-BJ and thrombocytin both cleaved at 41-42 and 46-47 peptide bonds, and fragment 42-103 disappeared rapidly. Both viper enzymes caused calcium mobilization in fibroblasts transfected with PAR4 and desensitized these cells to the thrombin action. In conclusion, both PAR1 and PAR4 mediate the effect of viper venom serine peptidases on platelets. PMID:10908720

Santos, B F; Serrano, S M; Kuliopulos, A; Niewiarowski, S

2000-07-21

137

Thrombin-induced Migration and Matrix Metalloproteinase-9 Expression Are Regulated by MAPK and PI3K Pathways in C6 Glioma Cells  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Glioblastoma multiforme is one of the most common and aggressive tumors in central nervous system. It often possesses characteristic necrotic lesions with hemorrhages, which increase the chances of exposure to thrombin. Thrombin has been known as a regulator of MMP-9 expression and cancer cell migration. However, the effects of thrombin on glioma cells have not been clearly understood. In the present study, influences of thrombin on glioma cell migration were examined using Boyden chamber mig...

Kim, Jiyoung; Lee, Jae-won; Kim, Song-in; Choi, Yong-joon; Lee, Won-ki; Jeong, Myung-ja; Cha, Sang-hoon; Lee, Hee Jae; Chun, Wanjoo; Kim, Sung-soo

2011-01-01

138

Thrombin enhances tumor cell adhesive and metastatic properties via increased alpha IIb beta 3 expression on the cell surface.  

Science.gov (United States)

The association between blood coagulation and cancer growth and metastatic dissemination is not yet completely understood. In this study we demonstrate that thrombin is capable of enhancing tumor cell adhesive properties and thereby increases tumor cell metastatic potential. Following exposure to alpha-thrombin, Walker 256 carcinosarcoma cells and B16 amelanotic melanoma cells became more adherent to both endothelial cell monolayers and the subendothelial matrix component, fibronectin. Preincubation of W256 and B16a cells with doses of alpha-thrombin from 0.01 to 10.0 U/ml produced a bell shape dose-response curve with the maximal effect (a 2-5-fold increase in adhesion) observed at 0.1 U/ml (corresponding to 0.8 nM). Complexes of alpha-thrombin with its inhibitors, hirudin and antithrombin III-heparin, diminished its effect on tumor cell adhesion. The effect of thrombin on tumor cell adhesion may be mediated by the alpha IIb beta 3 integrin as thrombin increased cell surface expression of the alpha IIb beta 3 complex. The significance of the in vitro observations was further substantiated by results of in vivo studies. Pretreatment of B16a cells with alpha-thrombin resulted in a 2-fold increase in the number of metastatic lung colonies in an experimental metastasis model. The data indicate a new role for thrombin in the metastatic spread of cancer. PMID:1281930

Wojtukiewicz, M Z; Tang, D G; Nelson, K K; Walz, D A; Diglio, C A; Honn, K V

1992-11-01

139

Thrombin stimulates VSMC proliferation through an EGFR-dependent pathway: involvement of MMP-2.  

Science.gov (United States)

In this study, the role of epidermal growth factor receptor (EGFR), extracellular signal-regulated kinase (ERK1/2), heparin-binding EGF-like growth factor (HB-EGF), general metalloproteinases, matrix metalloproteinases-2 (MMP-2) in mediating the mitogenic action of thrombin in rat vascular smooth muscle cells (VSMC) was investigated. The incubation of rat VSMC with thrombin (1 U/ml) for 5 min resulted in significant (p DNA synthesis by 3.6 ± 0.4-fold (p DNA synthesis correlates with the level of ERK1/2 activation rather than EGFR activation. These results further suggest that thrombin acts through EGFR and ERK 1/2 signaling pathways involving MMP-2 to upregulate proliferation of VSMC. PMID:25047892

Smiljanic, Katarina; Obradovic, Milan; Jovanovic, Aleksandra; Djordjevic, Jelena; Dobutovic, Branislava; Jevremovic, Danimir; Marche, Pierre; Isenovic, Esma R

2014-11-01

140

Discovery and evaluation of potent P1 aryl heterocycle-based thrombin inhibitors.  

Science.gov (United States)

In an effort to discover potent, clinically useful thrombin inhibitors, a rapid analogue synthetic approach was used to explore the P(1) region. Various benzylamines were coupled to a pyridine/pyrazinone P(2)-P(3) template. One compound with an o-thiadiazole benzylic substitution was found to have a thrombin K(i) of 0.84 nM. A study of ortho-substituted five-membered-ring heterocycles was undertaken and subsequently demonstrated that the o-triazole and tetrazole rings were optimal. Combination of these potent P(1) aryl heterocycles with a variety of P(2)-P(3) groups produced a compound with an extraordinary thrombin inhibitory activity of 1.4 pM. It is hoped that this potency enhancement in P(1) will allow for more diversification in the P(2)-P(3) region to ultimately address additional pharmacological concerns. PMID:15163182

Young, Mary Beth; Barrow, James C; Glass, Kristen L; Lundell, George F; Newton, Christina L; Pellicore, Janetta M; Rittle, Kenneth E; Selnick, Harold G; Stauffer, Kenneth J; Vacca, Joseph P; Williams, Peter D; Bohn, Dennis; Clayton, Franklin C; Cook, Jacquelynn J; Krueger, Julie A; Kuo, Lawrence C; Lewis, S Dale; Lucas, Bobby J; McMasters, Daniel R; Miller-Stein, Cynthia; Pietrak, Beth L; Wallace, Audrey A; White, Rebecca B; Wong, Bradley; Yan, Youwei; Nantermet, Philippe G

2004-06-01

 
 
 
 
141

Improving baculovirus recombination  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Recombinant baculoviruses have established themselves as a favoured technology for the high-level expression of recombinant proteins. The construction of recombinant viruses, however, is a time consuming step that restricts consideration of the technology for high throughput developments. Here we use a targeted gene knockout technology to inactivate an essential viral gene that lies adjacent to the locus used for recombination. Viral DNA prepared from the knockout fails to initiate an infecti...

Zhao, Yuguang; Chapman, David A. G.; Jones, Ian M.

2003-01-01

142

Abrogation of thrombin-induced increase in pulmonary microvascular permeability in PAR-1 knockout mice.  

Science.gov (United States)

We investigated the function of proteinase-activated receptor-1 (PAR-1) in the regulation of pulmonary microvascular permeability in response to thrombin challenge using PAR-1 knockout mice (-/-). Lungs were isolated and perfused with albumin (5 g/100 ml)-Krebs solution at constant flow (2 ml/min). Lung wet weight and pulmonary artery pressure (P(pa)) were continuously monitored. We determined the capillary filtration coefficient (K(fc)) and (125)I-labeled albumin (BSA) permeability-surface area product (PS) to assess changes in pulmonary microvessel permeability to liquid and albumin, respectively. Normal and PAR-1-null lung preparations received in the perfusate: 1) thrombin or 2) selective PAR-1 agonist peptide (TFLLRNPNDK-NH(2)). In control PAR-1 (+/+) mouse lungs, (125)I-albumin PS and K(fc) were significantly increased over baseline (by approximately 7- and 1.5-fold, respectively) within 20 min of alpha-thrombin (100 nM) challenge. PAR-1 agonist peptide (5 microM) gave similar results, whereas control peptide (5 microM; FTLLRNPNDK-NH(2)) was ineffective. At relatively high concentrations, thrombin (500 nM) or PAR-1 agonist peptide (10 microM) also induced increases in P(pa) and lung wet weight. All effects of thrombin (100 or 500 nM) or PAR-1 agonist peptide (5 or 10 microM) were prevented in PAR-1-null lung preparations. Baseline measures of microvessel permeability and P(pa) in the PAR-1-null preparations were indistinguishable from those in normal lungs. Moreover, PAR-1-null preparations gave normal vasoconstrictor response to thromboxane analog, U-46619 (100 nM). The results indicate that the PAR-1 receptor is critical in mediating the permeability-increasing and vasoconstrictor effects of thrombin in pulmonary microvessels. PMID:11120874

Vogel, S M; Gao, X; Mehta, D; Ye, R D; John, T A; Andrade-Gordon, P; Tiruppathi, C; Malik, A B

2000-12-18

143

Evaluation of Antithrombotic Activity of Thrombin DNA Aptamers by a Murine Thrombosis Model  

Science.gov (United States)

Aptamers are nucleic acid based molecular recognition elements with a high potential for the theranostics. Some of the aptamers are under development for therapeutic applications as promising antithrombotic agents; and G-quadruplex DNA aptamers, which directly inhibit the thrombin activity, are among them. RA-36, the 31-meric DNA aptamer, consists of two thrombin binding pharmacophores joined with the thymine linker. It has been shown earlier that RA-36 directly inhibits thrombin in the reaction of fibrinogen hydrolysis, and also it inhibits plasma and blood coagulation. Studies of both inhibitory and anticoagulation effects had indicated rather high species specificity of the aptamer. Further R&D of RA-36 requires exploring its efficiency in vivo. Therefore the development of a robust and adequate animal model for effective physiological studies of aptamers is in high current demand. This work is devoted to in vivo study of the antithrombotic effect of RA-36 aptamer. A murine model of thrombosis has been applied to reveal a lag and even prevention of thrombus formation when RA-36 was intravenous bolus injected in high doses of 1.4–7.1 µmol/kg (14–70 mg/kg). A comparative study of RA-36 aptamer and bivalirudin reveals that both direct thrombin inhibitors have similar antithrombotic effects for the murine model of thrombosis; though in vitro bivalirudin has anticoagulation activity several times higher compared to RA-36. The results indicate that both RA-36 aptamer and bivalirudin are direct thrombin inhibitors of different potency, but possible interactions of the thrombin-inhibitor complex with other components of blood coagulation cascade level the physiological effects for both inhibitors. PMID:25192011

Zavyalova, Elena; Samoylenkova, Nadezhda; Revishchin, Alexander; Golovin, Andrey; Pavlova, Galina; Kopylov, Alexey

2014-01-01

144

The effect of the REG2 Anticoagulation System on thrombin generation kinetics: a pharmacodynamic and pharmacokinetic first-in-human study.  

Science.gov (United States)

The REG2 Anticoagulation System consists of pegnivacogin, a subcutaneously administered aptamer factor IXa inhibitor, and its intravenous active control agent, anivamersen. Its effect on thrombin generation is unknown. A prospectively designed thrombin generation study was conducted within the phase 1 ascending dose study of REG2 to assess the effect of REG2 on thrombin generation kinetics. A total of 32 healthy volunteers were recruited into four cohorts of ascending dose pegnivacogin for the phase 1 study. In this pre-specified substudy, blood samples were drawn in the presence or absence of corn trypsin inhibitor at specified times within each dosing cohort. Thrombin generation was initiated with tissue factor and thrombin generation kinetics were measured using the Calibrated Automated Thrombogram (CAT). REG2 attenuated thrombin generation in a dose-dependent manner. All parameters of the CAT assay, except for lag time, showed a dose and concentration-dependent response to pegnivacogin [time to peak thrombin generation (PTm), endogenous thrombin potential, peak thrombin generation, and velocity index (VIx)]. Reversal of the effect of pegnivacogin with anivamersen demonstrated restoration of thrombin generation without rebound effect. This first-in-human study of the effect of the REG2 Anticoagulation System on thrombin generation demonstrates concentration-dependent suppression of thrombin generation that is reversible without rebound effect, as measured by the CAT assay. PMID:24880800

Vavalle, J P; Rusconi, C P; Zelenkofske, S; Wargin, W A; Ortel, T L; Alexander, J H; Povsic, T J; Becker, R C

2014-10-01

145

Therapeutic Recombinant Monoclonal Antibodies  

Science.gov (United States)

During the last two decades, the rapid growth of biotechnology-derived techniques has led to a myriad of therapeutic recombinant monoclonal antibodies with significant clinical benefits. Recombinant monoclonal antibodies can be obtained from a number of natural sources such as animal cell cultures using recombinant DNA engineering. In contrast to…

Bakhtiar, Ray

2012-01-01

146

Application of thrombin powder after tooth extraction in patients receiving anticoagulant therapy  

Directory of Open Access Journals (Sweden)

Full Text Available Patients with extreme hypocoagulation, which occurs either as an effect of some diseases with coagulation deficiency or because of the anticoagulant therapy (ACT, are a risk group for oral surgery. In the last decades decision to change or interrupt ACT before and after the procedure was abandoned and more often local hemostasis was being achieved by combining chemical and biological substances. The success of the surgical hemostasis and thrombin powder combination was tested on the group of 20 patients with ACT. The results were satisfactory despite thrombin powder solubility in the moist oral environment.

Marjanovi? Marjan

2002-01-01

147

Recombineering Homologous Recombination Constructs in Drosophila  

Science.gov (United States)

The continued development of techniques for fast, large-scale manipulation of endogenous gene loci will broaden the use of Drosophila melanogaster as a genetic model organism for human-disease related research. Recent years have seen technical advancements like homologous recombination and recombineering. However, generating unequivocal null mutations or tagging endogenous proteins remains a substantial effort for most genes. Here, we describe and demonstrate techniques for using recombineering-based cloning methods to generate vectors that can be used to target and manipulate endogenous loci in vivo. Specifically, we have established a combination of three technologies: (1) BAC transgenesis/recombineering, (2) ends-out homologous recombination and (3) Gateway technology to provide a robust, efficient and flexible method for manipulating endogenous genomic loci. In this protocol, we provide step-by-step details about how to (1) design individual vectors, (2) how to clone large fragments of genomic DNA into the homologous recombination vector using gap repair, and (3) how to replace or tag genes of interest within these vectors using a second round of recombineering. Finally, we will also provide a protocol for how to mobilize these cassettes in vivo to generate a knockout, or a tagged gene via knock-in. These methods can easily be adopted for multiple targets in parallel and provide a means for manipulating the Drosophila genome in a timely and efficient manner. PMID:23893070

Carreira-Rosario, Arnaldo; Scoggin, Shane; Shalaby, Nevine A.; Williams, Nathan David; Hiesinger, P. Robin; Buszczak, Michael

2013-01-01

148

Topical report review status  

International Nuclear Information System (INIS)

A Topical Report Review Status is scheduled to be published semi-annually. The primary purpose of this document is to provide periodic progress reports of on-going topical report reviews, to identify those topical reports for which the Nuclear Regulatory Commission (NRC) staff review has been completed and, to the extent practicable, to provide NRC management with sufficient information regarding the conduct of the topical report program to permit taking whatever actions deemed necessary or appropriate. This document is also intended to be a source of information to NRC Licensing Project Managers and other NRC personnel regarding the status of topical reports which may be referenced in applications for which they have responsibility. This status report is published primarily for internal NRC use in managing the topical report program, but is also used by NRC to advise the industry of report review status

149

(-)-Epigallocatechin-3-gallate decreases thrombin/paclitaxel-induced endothelial tissue factor expression via the inhibition of c-Jun terminal NH2 kinase phosphorylation  

International Nuclear Information System (INIS)

Patients with paclitaxel-eluting stents are concerned with stent thrombosis caused by premature discontinuation of dual antiplatelet therapy or clopidogrel resistance. This study investigates the effect of (-)-epigallocatechin-3-gallate (EGCG) on the expression of thrombin/paclitaxel-induced endothelial tissue factor (TF) expressions in human aortic endothelial cells (HAECs). EGCG was nontoxic to HAECs at 6 h up to a concentration of 25 ?mol/L. At a concentration of 25 ?mol/L, EGCG pretreatment potently inhibited both thrombin-stimulated and thrombin/paclitaxel-stimulated endothelial TF protein expression. Thrombin and thrombin/paclitaxel-induced 2.6-fold and 2.9-fold increases in TF activity compared with the control. EGCG pretreatment caused a 29% and 38% decrease in TF activity on thrombin and thrombin/paclitaxel treatment, respectively. Real-time polymerase chain reaction (PCR) showed that thrombin and thrombin/paclitaxel-induced 3.0-fold and 4.6-fold TF mRNA expressions compared with the control. EGCG pretreatment caused an 82% and 72% decrease in TF mRNA expression on thrombin and thrombin/paclitaxel treatment, respectively. The c-Jun terminal NH2 kinase (JNK) inhibitor SP600125 reduced thrombin/paclitaxel-induced TF expression. Furthermore, EGCG significantly inhibited the phosphorylation of JNK to 49% of thrombin/paclitaxel-stimulated HAECs at 60 min. Immunofluorescence assay did not show an inhibitory effect of EGCG on P65 NF-?B nuclear translocation in the thrombin/paclitaxel-stimulated endothelial cells. In conclusion, EGCG can inhibit TF expression in thrombin/paclitaxel-stimulated endothelial cells via the inhibition of JNK phosphorylation. The unique property of EGCG may be used to develop a new drug-eluting stent by co-coating EGCG and paclitaxel.

150

(-)-Epigallocatechin-3-gallate decreases thrombin/paclitaxel-induced endothelial tissue factor expression via the inhibition of c-Jun terminal NH2 kinase phosphorylation  

Energy Technology Data Exchange (ETDEWEB)

Patients with paclitaxel-eluting stents are concerned with stent thrombosis caused by premature discontinuation of dual antiplatelet therapy or clopidogrel resistance. This study investigates the effect of (-)-epigallocatechin-3-gallate (EGCG) on the expression of thrombin/paclitaxel-induced endothelial tissue factor (TF) expressions in human aortic endothelial cells (HAECs). EGCG was nontoxic to HAECs at 6 h up to a concentration of 25 {mu}mol/L. At a concentration of 25 {mu}mol/L, EGCG pretreatment potently inhibited both thrombin-stimulated and thrombin/paclitaxel-stimulated endothelial TF protein expression. Thrombin and thrombin/paclitaxel-induced 2.6-fold and 2.9-fold increases in TF activity compared with the control. EGCG pretreatment caused a 29% and 38% decrease in TF activity on thrombin and thrombin/paclitaxel treatment, respectively. Real-time polymerase chain reaction (PCR) showed that thrombin and thrombin/paclitaxel-induced 3.0-fold and 4.6-fold TF mRNA expressions compared with the control. EGCG pretreatment caused an 82% and 72% decrease in TF mRNA expression on thrombin and thrombin/paclitaxel treatment, respectively. The c-Jun terminal NH2 kinase (JNK) inhibitor SP600125 reduced thrombin/paclitaxel-induced TF expression. Furthermore, EGCG significantly inhibited the phosphorylation of JNK to 49% of thrombin/paclitaxel-stimulated HAECs at 60 min. Immunofluorescence assay did not show an inhibitory effect of EGCG on P65 NF-{kappa}B nuclear translocation in the thrombin/paclitaxel-stimulated endothelial cells. In conclusion, EGCG can inhibit TF expression in thrombin/paclitaxel-stimulated endothelial cells via the inhibition of JNK phosphorylation. The unique property of EGCG may be used to develop a new drug-eluting stent by co-coating EGCG and paclitaxel.

Wang, Huang-Joe [Institute of Biotechnology, National Tsing Hua University, No. 101, Section 2, Kuang Fu Road, Hsinchu 30013, Taiwan (China); Division of Cardiology, Department of Medicine, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung 40447, Taiwan (China); Lo, Wan-Yu [Department of Medical Research, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung 40447, Taiwan (China); Graduate Integration of Chinese and Western Medicine, China Medical University, No. 91, Hsueh-Shih Road, Taichung 40402, Taiwan (China); Lu, Te-Ling [School of Pharmacy, China Medical University, No. 91, Hsueh-Shih Road, Taichung 40402, Taiwan (China); Huang, Haimei, E-mail: hmhuang@life.nthu.edu.tw [Institute of Biotechnology, National Tsing Hua University, No. 101, Section 2, Kuang Fu Road, Hsinchu 30013, Taiwan (China)

2010-01-01

151

(-)-Epigallocatechin-3-gallate decreases thrombin/paclitaxel-induced endothelial tissue factor expression via the inhibition of c-Jun terminal NH2 kinase phosphorylation.  

Science.gov (United States)

Patients with paclitaxel-eluting stents are concerned with stent thrombosis caused by premature discontinuation of dual antiplatelet therapy or clopidogrel resistance. This study investigates the effect of (-)-epigallocatechin-3-gallate (EGCG) on the expression of thrombin/paclitaxel-induced endothelial tissue factor (TF) expressions in human aortic endothelial cells (HAECs). EGCG was nontoxic to HAECs at 6h up to a concentration of 25 micromol/L. At a concentration of 25 micromol/L, EGCG pretreatment potently inhibited both thrombin-stimulated and thrombin/paclitaxel-stimulated endothelial TF protein expression. Thrombin and thrombin/paclitaxel-induced 2.6-fold and 2.9-fold increases in TF activity compared with the control. EGCG pretreatment caused a 29% and 38% decrease in TF activity on thrombin and thrombin/paclitaxel treatment, respectively. Real-time polymerase chain reaction (PCR) showed that thrombin and thrombin/paclitaxel-induced 3.0-fold and 4.6-fold TF mRNA expressions compared with the control. EGCG pretreatment caused an 82% and 72% decrease in TF mRNA expression on thrombin and thrombin/paclitaxel treatment, respectively. The c-Jun terminal NH2 kinase (JNK) inhibitor SP600125 reduced thrombin/paclitaxel-induced TF expression. Furthermore, EGCG significantly inhibited the phosphorylation of JNK to 49% of thrombin/paclitaxel-stimulated HAECs at 60min. Immunofluorescence assay did not show an inhibitory effect of EGCG on P65 NF-kappaB nuclear translocation in the thrombin/paclitaxel-stimulated endothelial cells. In conclusion, EGCG can inhibit TF expression in thrombin/paclitaxel-stimulated endothelial cells via the inhibition of JNK phosphorylation. The unique property of EGCG may be used to develop a new drug-eluting stent by co-coating EGCG and paclitaxel. PMID:19944065

Wang, Huang-Joe; Lo, Wan-Yu; Lu, Te-Ling; Huang, Haimei

2010-01-01

152

Recombinant dengue type 1 virus NS5 protein expressed in Escherichia coli exhibits RNA-dependent RNA polymerase activity.  

Science.gov (United States)

The complete nonstructural NS5 gene of dengue type 1 virus, Singapore strain S275/90 (D1-S275/90) was expressed in Escherichia coli as a glutathione S-transferase (GST) fusion protein (126 kDa). The GST-NS5 fusion protein was purified and the recombinant NS5 protein released from the fusion protein by thrombin cleavage. The recombinant NS5 had a predicted molecular weight of 100 kDa and reacted with antiserum against D1-S275/90 virus in Western blot analysis. The purified recombinant NS5 protein possessed RNA-dependent RNA polymerase activity which was inhibited (>99%) by antibodies against the recombinant NS5 protein. The polymerase product was shown to be a negative-stranded RNA molecule, of template size, which forms a double-stranded complex with the template RNA. PMID:8607261

Tan, B H; Fu, J; Sugrue, R J; Yap, E H; Chan, Y C; Tan, Y H

1996-02-15

153

Choosing a Research Topic  

Science.gov (United States)

This website from the Chronicle of Higher Education features tips for graduate students on choosing a research topic. The author explains a variety of ways that a topic should be matched to the individual. A good choice can help lead to long-term career success.

Reis, Richard M.; Education, The C.

154

Mapping Topics and Topic Bursts in PNAS  

CERN Document Server

Scientific research is highly dynamic. New areas of science continually evolve;others gain or lose importance, merge or split. Due to the steady increase in the number of scientific publications it is hard to keep an overview of the structure and dynamic development of one's own field of science, much less all scientific domains. However, knowledge of hot topics, emergent research frontiers, or change of focus in certain areas is a critical component of resource allocation decisions in research labs, governmental institutions, and corporations. This paper demonstrates the utilization of Kleinberg's burst detection algorithm, co-word occurrence analysis, and graph layout techniques to generate maps that support the identification of major research topics and trends. The approach was applied to analyze and map the complete set of papers published in the Proceedings of the National Academy of Sciences (PNAS) in the years 1982-2001. Six domain experts examined and commented on the resulting maps in an attempt to ...

Mane, K; Mane, Ketan; B\\"orner, Katy

2004-01-01

155

[Thrombin-like serine proteases in Cerasted venoms (Cerasted cerastes and Cerastes vipera)].  

Science.gov (United States)

Cerastes cerastes and Cerastes vipera snake venoms are rich of thrombin-like, serine-protease polypeptides. Many proteins have been isolated, purified and characterized from these vipers. These proteins act in various way on blood coagulation pathway and platelet function. PMID:14722941

Marrakchi, N; el Ayeb, M

1998-01-01

156

Electrogenerated chemiluminescence aptasensor for ultrasensitive detection of thrombin incorporating an auxiliary probe.  

Science.gov (United States)

A novel electrogenerated chemiluminescence (ECL) aptasensor for ultrasensitive detection of thrombin incorporating an auxiliary probe was designed by employing specific anti-thrombin aptamer as a capture probe and a ruthenium(II) complex-tagged reporter probe as an ECL probe and an auxiliary probe to assist the ECL probe close to the surface of the electrode. The ECL aptasensor was fabricated by self-assembling a thiolated capture probe on the surface of gold electrode and then hybridizing the ECL probe with the capture probe, and further self-assembling the auxiliary probe. When analyte thrombin was bound with the capture probe, the part of the dehybridized ECL probe was hybridized with the neighboring auxiliary probe, led to the tagged ruthenium(II) complex close to the electrode surface, resulted in great increase in the ECL intensity. The results showed that the increased ECL intensity was directly related to the logarithm of thrombin concentrations in the range from 5.0 × 10(-15)M to 5.0 × 10(-12)M with a detection limit of 2.0 × 10(-15)M. This work demonstrates that employing an auxiliary probe which exists nearby the capture probe can enhance the sensitivity of the ECL aptasensor. This promising strategy will be extended to the design of other biosensors for detection of other proteins and genes. PMID:25159423

Li, Zhejian; Sun, Lijuan; Zhao, Ying; Yang, Libin; Qi, Honglan; Gao, Qiang; Zhang, Chengxiao

2014-12-01

157

Oral direct thrombin inhibition: an effective and novel approach for venous thromboembolism.  

Science.gov (United States)

Venous thromboembolism is a serious illness that affects patient morbidity and mortality and presents a significant management challenge to healthcare providers world-wide. Despite major achievements in the significant reduction of thromboembolic complications, the most common therapies currently used for prevention and treatment of venous thromboembolism--heparins and vitamin K antagonists such as warfarin--have several limitations. In particular, unfractionated heparin and warfarin show significant inter-patient variability in pharmacokinetics and pharmacodynamics, which makes regular coagulation monitoring necessary. Furthermore, only warfarin is suitable for long-term use, as it is administered orally. A new class of anticoagulants has been developed that directly target thrombin, a key enzyme in the blood coagulation cascade. Unlike warfarin and heparin, these direct thrombin inhibitors are able to inhibit fibrin-bound thrombin and so produce more effective inhibition of coagulation. Importantly, some members of this class of drugs have been developed for oral administration. Ximelagatran, which is converted to its active form melagatran, has predictable pharmacokinetics and pharmacodynamics. Therefore, ximelagatran can be administered in fixed doses with no need for coagulation monitoring. Its efficacy and safety profile have been demonstrated in preclinical and clinical studies. As the first oral agent in the new class, direct thrombin inhibitors, ximelagatran has significant potential for improving the prevention and treatment of venous thromboembolism. PMID:15586623

Haas, Sylvia

2004-01-01

158

Reperfusion of Splanchnic Artery Aneurysm Following Transcatheter Embolization: Treatment with Percutaneous Thrombin Injection  

International Nuclear Information System (INIS)

We describe a case of reperfusion of an aneurysm of the pancreaticoduodenal artery following transcatheter coil embolotherapy. The lesion was successfully treated by direct puncture of the aneurysm under computed tomographic guidance, followed by injection of thrombin. This technique is useful when an endovascular approach is not feasible

159

Prolyl endopeptidase and thrombin inhibitory diterpenoids from the bark of Xylopia aethiopica.  

Science.gov (United States)

The inhibitory effects of seven diterpenes, belonging to three different structural classes and isolated from the bark of Xylopia aethiopica, were investigated against the enzymes prolyl endopeptidase (PEP) and alpha-thrombin. Five compounds exhibited inhibitory activity against them. PMID:16195597

Diderot, Noungoue Tchamo; Silvere, Ngouela; Yasin, Amsha; Zareen, Seema; Fabien, Zelefack; Etienne, Tsamo; Choudhary, M Iqbal; Atta-Ur-Rahman

2005-09-01

160

Thrombin conducts epithelial?mesenchymal transition via protease?activated receptor?1 in human gastric cancer.  

Science.gov (United States)

Epithelial?mesenchymal transition (EMT) is thought to be a key step for cancer metastasis. Using an immunohistochemical approach with gastric carcinoma tissue, we found the expression of protease?activated receptor?1 (PAR1), along with a metalloproteinase known to activate PAR1, were associated with poorer prognosis, compared with expression?negative tumors, and activated PAR1 promotes gastric cancer cell invasion and proliferation in vivo. In this study we observed EMT induction by the PAR1 agonist ??thrombin, in human gastric cell lines stably expressing PAR1. We investigated ??thrombin-induced changes in the cell forms of pcDNA3.1?MKN45 (MKN45/Mock), pcDNA3.1?PAR1 transfected MKN45 (MKN45/PAR1), and MKN74. Expression levels of epithelial and mesenchymal markers as well as the distribution of transcriptional factors of E?cadherin in the cytoplasm and nucleus were also noted in these cell lines. We observed ??thrombin-induced morphological changes in MKN45/PAR1 and MKN74 cells. Western blotting and immunohistochemistry of these cells indicated a fall in the expression level of E?cadherin and an increase in fibronectin expression after 48 h. PAR1 activation also induced significant increases in nuclear levels of the Snail which is a repressor of E?cadherin gene expression. We found EMT in gastric cancer cell lines that underwent ??thrombin-induced PAR1 activation. PMID:25231630

Otsuki, Tadayoshi; Fujimoto, Daisuke; Hirono, Yasuo; Goi, Takanori; Yamaguchi, Akio

2014-12-01

 
 
 
 
161

An off-on-off electrochemiluminescence approach for ultrasensitive detection of thrombin.  

Science.gov (United States)

This work demonstrates an aptasensor for ultrasensitive electrochemiluminescence (ECL) detection of thrombin based on an "off-on-off" approach. The system is composed of an Eu(3+)-doped CdS nanocrystals (CdS:Eu NCs) film on glassy carbon electrode (GCE) as ECL emitter. Then gold nanoparticles (AuNPs) labeled hairpin-DNA probe (ssDNA1) containing thrombin-binding aptamer (TBA) sequence was linked on the NCs film, which led to ECL quenching (off) as a result of Förster-resonance energy transfer (FRET) between the CdS:Eu NC film and the proximal AuNPs. Upon the occurrence of hybridization with its complementary DNA (ssDNA2), an ECL enhancement (on) occurred owing to the interactions of the excited CdS:Eu NCs with ECL-induced surface plasmon resonance (SPR) in AuNPs at large separation. Thrombin could induce ssDNA1 forming a G-quadruplex and cause the AuNPs to be close to CdS:Eu NCs film again, which resulted in an enhanced ECL quenching (off). This "off-on-off" system showed a maximum 7.4-fold change of ECL intensity due to the configuration transformation of ssDNA1 and provides great sensitivity for detection of thrombin in a wide detection range from 50 aM to 1 pM. PMID:24699694

Deng, Li; Du, Ying; Xu, Jing-Juan; Chen, Hong-Yuan

2014-09-15

162

Improved thrombin binding aptamer by incorporation of a single unlocked nucleic acid monomer  

DEFF Research Database (Denmark)

A 15-mer DNA aptamer (named TBA) adopts a G-quadruplex structure that strongly inhibits fibrin-clot formation by binding to thrombin. We have performed thermodynamic analysis, binding affinity and biological activity studies of TBA variants modified by unlocked nucleic acid (UNA) monomers. UNA-U placed in position U3, U7 or U12 increases the thermodynamic stability of TBA by 0.15-0.50?kcal/mol. In contrast, modification of any position within the two G-quartet structural elements is unfavorable for quadruplex formation. The intramolecular folding of the quadruplexes is confirmed by T(m) versus ln c analysis. Moreover, circular dichroism and thermal difference spectra of the modified TBAs displaying high thermodynamic stability show bands that are characteristic for antiparallel quadruplex formation. Surface plasmon resonance studies of the binding of the UNA-modified TBAs to thrombin show that a UNA monomer is allowed in many positions of the aptamer without significantly changing the thrombin-binding properties. The biological effect of a selection of the modified aptamers was tested by a thrombin time assay and showed that most of the UNA-modified TBAs possess anticoagulant properties, and that the construct with a UNA-U monomer in position 7 is a highly potent inhibitor of fibrin-clot formation.

Pasternak, Anna; Hernandez, Frank J

2011-01-01

163

Engineering Thrombin for Selective Specificity toward Protein C and PAR1*  

Science.gov (United States)

Thrombin elicits functional responses critical to blood homeostasis by interacting with diverse physiological substrates. Ala-scanning mutagenesis of 97 residues covering 53% of the solvent accessible surface area of the enzyme identifies Trp215 as the single most important determinant of thrombin specificity. Saturation mutagenesis of Trp215 produces constructs featuring kcat/Km values for the hydrolysis of fibrinogen, protease-activated receptor PAR1, and protein C that span five orders of magnitude. Importantly, the effect of Trp215 replacement is context dependent. Mutant W215E is 10-fold more specific for protein C than fibrinogen and PAR1, which represents a striking shift in specificity relative to wild-type that is 100-fold more specific for fibrinogen and PAR1 than protein C. However, when the W215E mutation is combined with deletion of nine residues in the autolysis loop, which by itself shifts the specificity of the enzyme from fibrinogen and PAR1 to protein C, the resulting construct features significant activity only toward PAR1. These findings demonstrate that thrombin can be re-engineered for selective specificity toward protein C and PAR1. Mutations of Trp215 provide important reagents for dissecting the multiple functional roles of thrombin in the blood and for clinical applications. PMID:20404340

Marino, Francesca; Pelc, Leslie A.; Vogt, Austin; Gandhi, Prafull S.; Di Cera, Enrico

2010-01-01

164

Rigidification of the autolysis loop enhances Na[superscript +] binding to thrombin  

Energy Technology Data Exchange (ETDEWEB)

Binding of Na{sup +} to thrombin ensures high activity toward physiological substrates and optimizes the procoagulant and prothrombotic roles of the enzyme in vivo. Under physiological conditions of pH and temperature, the binding affinity of Na{sup +} is weak due to large heat capacity and enthalpy changes associated with binding, and the K{sub d} = 80 mM ensures only 64% saturation of the site at the concentration of Na{sup +} in the blood (140 mM). Residues controlling Na{sup +} binding and activation have been identified. Yet, attempts to improve the interaction of Na{sup +} with thrombin and possibly increase catalytic activity under physiological conditions have so far been unsuccessful. Here we report how replacement of the flexible autolysis loop of human thrombin with the homologous rigid domain of the murine enzyme results in a drastic (up to 10-fold) increase in Na{sup +} affinity and a significant improvement in the catalytic activity of the enzyme. Rigidification of the autolysis loop abolishes the heat capacity change associated with Na{sup +} binding observed in the wild-type and also increases the stability of thrombin. These findings have general relevance to protein engineering studies of clotting proteases and trypsin-like enzymes.

Pozzi, Nicola; Chen, Raymond; Chen, Zhiwei; Bah, Alaji; Di Cera, Enrico (St. Louis-MED)

2011-09-20

165

Aptamer modified organic-inorganic hybrid silica monolithic capillary columns for highly selective recognition of thrombin.  

Science.gov (United States)

A novel kind of aptamer modified organic-inorganic hybrid silica monolithic capillary column has been developed, via the covalent bonding of 5'-NH(2)-modified aptamer for human ?-thrombin on hybrid silica monolith, prepared by sol-gel method, with tetraethoxysilane and 3-aminopropyltriethoxysilane as precursors. Due to the large specific surface area of the hybrid matrix, the average coverage density of aptamer reached 568 pmol/?L, and the thrombin binding capacity was 1.15 ?g/?L, 14 times higher than that of aptamer modified open tubular capillaries. By such an affinity capillary column, the limit of detection of thrombin was decreased to 3.4 nM with a UV detector. Furthermore, even when thrombin was mixed with 1000 times more concentrated human serum, it could be selectively enriched and detected with the signal-to-noise ratio as ca.10. These results indicate that the developed preparation strategy for aptamer based hybrid silica monolithic capillary column might provide an effective method to achieve highly selective recognition of trace targets. PMID:23137349

Deng, Nan; Liang, Zhen; Liang, Yu; Sui, Zhigang; Zhang, Liyuan; Wu, Qi; Yang, Kaiguang; Zhang, Lihua; Zhang, Yukui

2012-12-01

166

Inhibition of intimal hyperplasia by direct thrombin inhibitors in an animal vein bypass model.  

Science.gov (United States)

Many functions of the coagulation system have nonthrombotic effects. The indirect thrombin inhibitor heparin has been previously shown to be effective in limiting intimal hyperplasia (IH). We sought to study the effect of thrombin on IH by using two direct thrombin inhibitors (DTIs), argatroban and lepirudin. Sprague-Dawley rats underwent interposition vein grafting to the carotid artery. Vein grafts were treated with either saline (n = 6) or one of the two DTIs (n = 6 for both). At 30 days, the rats were sacrificed and vessels were perfusion fixed. Sections of the proximal carotid artery, graft, and both anastomoses were stained with both hematoxlyin/eosin and von Gieson's elastin stain. Sections were examined and compared for luminal area and intima-to-media (IM) ratio. The vessels treated with DTIs had less (p argatroban 0.182 +/- 0.118) and better lumen preservation than the control vessels (lumen area of proximal anastomosis: control 1.69 +/- 0.9, lepirudin 2.45 +/- 0.74, argatroban 2.81 +/- 0.78). There were no thromboses in the DTI-treated vessels. Dilatation of the graft segment was noted in the argatroban group. Thus, DTIs are effective at reducing IH in a small-animal model, suggesting that inhibition of thrombin has a protective role in IH. In addition, a difference of action between DTIs is suggested by the dilatation seen only in the argatroban-treated graft sections. PMID:15253248

Mureebe, Leila; Turnquist, Susan E; Silver, Donald

2004-03-01

167

Fundamental study of recombination and recombineering in Escherichia coli  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Recombination and recombineering systems have been used in Escherichia coli to recombinant DNA sequences. With endonuclease and DNA lipase the bacterial plasmid and target DNA fragment can bind together and recombinant for a new DNA sequences. Red Proteins have been used in recombineering system to perform the function as the enzymes in recombination system, and faster and easier than the other way of recombinant new DNA sequences in E.coli. In this report we get to know the pr...

Sun, Xiaohang; Huang, Yang

2008-01-01

168

Implicit stage topics  

Directory of Open Access Journals (Sweden)

Full Text Available Il a souvent été proposé que les éléments spatio-temporels en position initiale de phrase spécifient le cadre de l’événement dénoté par la proposition et ont une interprétation thématique ou topicale. Alors que les topiques spatio-temporels explicites ont souvent été étudiés, Erteschik-Schir (1997, 1999 propose l’idée que les topiques spatio-temporels, ou topiques scéniques (stage topics peuvent aussi être implicites.Dans cet article, nous offrons des arguments en faveur de la notion de topique scénique implicite. Nous montrons qu’un certain nombre de cas d’inversion nominale en français, une configuration syntaxique qui est favorisée par la présence d’un topique scénique explicite, s’expliquent par la présence d’un topique scénique implicite. Le fait que les topiques scéniques implicites interagissent avec la structure syntaxique de la même façon que les topiques scéniques explicites constitue un argument empirique en faveur de leur existence.It has often been proposed that sentence-initial spatio-temporal elements specify the frame in which the whole proposition takes place and are topical (i.e. thematic. Whereas considerable attention has been paid to explicit spatio-temporal topics, Erteschik-Shir (1997, 1999 argues that spatio-temporal topics, or stage topics, can also be implicit.In this article we provide evidence in favour of the notion of implicit stage topic. We show that a certain number of nominal inversion cases in French, a syntactic configuration which is triggered by the presence of an explicit stage topic, are explained by the presence of an implicit stage topic. The fact that implicit stage topics interact with syntactic structure the same way explicit stage topics do constitutes a strong empirical argument in favour of their existence.

Karen Lahousse

2008-04-01

169

Protease-activated receptor-1 mediates thrombin-induced persistent sodium current in human cardiomyocytes.  

Science.gov (United States)

After the thrombus formation in cardiac cavities or coronaries, the serine protease thrombin is produced and can therefore reach the myocardial tissue by the active process of extravasation and binds to the G protein-coupled protease-activated receptor-1 (PAR1) expressed in human myocardium. The role of PAR1 was investigated in the thrombin effect on sodium current (I(Na)). I(Na) was recorded in freshly isolated human atrial myocytes by the whole-cell patch-clamp method. Action potentials (AP) were recorded in guinea pig ventricular tissue by the conventional glass microelectrode technique. Thrombin-activated PAR1 induced a tetrodotoxin-blocked persistent sodium current, I(NaP), in a concentration-dependent manner with an apparent EC(50) of 28 U/ml. The PAR1 agonist peptide SFLLR-NH(2) (50 microM) was able to mimic PAR1-thrombin action, whereas PAR1 antagonists N(3)-cyclopropyl-7-((4-(1-methylethyl)-phenyl)methyl)-7H-pyrrolo(3,2-f)quinazoline-1,3-diamine (SCH 203099; 10 microM) and 1-(3,5-di-tert-butyl-4-hydroxy-phenyl)-2-[3-(3-ethyl-3-hydroxy-pentyl)-2-imino-2,3-dihydro-imidazol-1-yl]-ethanone (ER 112787) (1 microM), completely inhibited it. The activated PAR1 involves the calcium-independent phospholipase-A(2) signaling pathway because two inhibitors of this cascade, bromoenol lactone (50 microM) and haloenol lactone suicide substrate (50 microM), block PAR1-thrombin-induced I(NaP).Asa consequence of I(NaP) activation, in guinea pig right ventricle papillary muscle, action potential duration (APD) were significantly increased by 20% and 15% under the respective action of 32 U/ml thrombin and 50 microM SFLLR-NH(2), and these increases in APD were prevented by 1 microM tetrodotoxin or markedly reduced by application of 1 microM SCH 203099 or ER 112787. Thrombin, through PAR1 activation, increases persistent component of the Na(+) current resulting in an uncontrolled sodium influx into the cardiomyocyte, which can contribute to cellular injuries observed during cardiac ischemia. PMID:18326052

Pinet, Caroline; Algalarrondo, Vincent; Sablayrolles, Sylvie; Le Grand, Bruno; Pignier, Christophe; Cussac, Didier; Perez, Michel; Hatem, Stephane N; Coulombe, Alain

2008-06-01

170

Characterization of a modified gold platform for the development of a label-free anti-thrombin aptasensor.  

Science.gov (United States)

This work reports the characterization of a modified gold surface as a platform for the development of a label free aptasensor for thrombin detection. The biorecognition platform was obtained by the self-assembly of 4-mercaptobenzoic acid onto a gold surface, covalent attachment of streptavidin and further immobilization of the biotinylated anti-thrombin aptamer. The biosensing platform was characterized by cyclic voltammetry, electrochemical impedance spectroscopy, surface plasmon resonance (SPR) and quartz crystal microbalance with dissipation monitoring. The biorecognition event aptamer-thrombin was detected from changes in the SPR angle produced as a consequence of the molecular interaction between the aptasensor and the target protein. The biosensing platform demonstrated to be highly selective for human thrombin even in the presence of large excess of bovine thrombin, bovine serum albumin, cytochrome C, lysozyme and myoglobin. The relationship between the changes in the SPR angle and thrombin concentration was linear up to 0.19 ?mol L(-1) (R(2)=0.992) while the detection limit was of 12.0 nmol L(-1) (240 fmol in the sample). This new sensing approach represents an interesting and promising alternative for the SPR-based quantification of thrombin. PMID:23017682

Jalit, Yamile; Gutierrez, Fabiana A; Dubacheva, Galina; Goyer, Cedric; Coche-Guerente, Liliane; Defrancq, Eric; Labbé, Pierre; Rivas, Gustavo A; Rodríguez, Marcela C

2013-03-15

171

The thrombin inhibitor Argatroban reduces breast cancer malignancy and metastasis via osteopontin-dependent and osteopontin-independent mechanisms.  

Science.gov (United States)

Osteopontin (OPN) has been clinically and experimentally associated with breast cancer metastasis. Proteolytic cleavage of OPN by thrombin has been reported to increase its biologic activity. The purpose of this study was to determine if inhibition of thrombin could reduce the malignancy-promoting effects of OPN on breast cancer cell behavior in vitro and in vivo. MDA-MB-468 human breast cancer cells were stably transfected to overexpress OPN (468-OPN) or a control vector (468-CON) and compared for functional differences in malignant/metastatic behavior in response to treatment with the thrombin-specific inhibitor Argatroban. Western blot analysis revealed that both 468-CON and 468-OPN cells produce thrombin and the thrombin-related protein tissue factor, and express very low levels of thrombin receptor (PAR-1). In vitro assays demonstrated that Argatroban treatment (25 microg/ml) of 468-OPN cells resulted in decreased cell growth, colony-forming ability, adhesion, and migration relative to untreated controls (P MDA-MB-468 breast cancer cells using both OPN-dependent and OPN-independent mechanisms, and suggest that thrombin inhibitors such as Argatroban may hold potential as therapeutic agents to combat breast cancer progression. PMID:18097747

Schulze, Erika B; Hedley, Benjamin D; Goodale, David; Postenka, Carl O; Al-Katib, Waleed; Tuck, Alan B; Chambers, Ann F; Allan, Alison L

2008-11-01

172

Monitoring Thrombin Generation and Screening Anticoagulants through Pulse Laser-Induced Fragmentation of Biofunctional Nanogold on Cellulose Membranes.  

Science.gov (United States)

Thrombin generation (TG) has an important part in the blood coagulation system, and monitoring TG is useful for diagnosing various health issues related to hypo-coagulability and hyper-coagulability. In this study, we constructed probes by using mixed cellulose ester membranes (MCEMs) modified with gold nanoparticles (Au NPs) for monitoring thrombin activity using laser desorption/ionization mass spectrometry (LDI-MS). The LDI process produced Au cationic clusters ([Aun](+); n = 1-3) that we detected through MS. When thrombin reacted with fibrinogen on the Au NPs-MCEMs, insoluble fibrin was formed, hindering the formation of Au cationic clusters and, thereby, decreasing the intensity of their signals in the mass spectrum. Accordingly, we incorporated fibrinogen onto the Au NPs-MCEMs to form Fib-Au NPs-MCEM probes to monitor TG with good selectivity (>1000-fold toward thrombin with respect to other proteins or enzymes) and sensitivity (limit of detection for thrombin of ca. 2.5 pM in human plasma samples). Our probe exhibited remarkable performance in monitoring the inhibition of thrombin activity by direct thrombin inhibitors. Analyses of real samples using our new membrane-based probe suggested that it will be highly useful in practical applications for the effective management of hemostatic complications. PMID:25141032

Li, Yu-Jia; Chiu, Wei-Jane; Unnikrishnan, Binesh; Huang, Chih-Ching

2014-09-10

173

Thrombin Inhibition with Dabigatran Protects against High-Fat Diet-Induced Fatty Liver Disease in Mice.  

Science.gov (United States)

Nonalcoholic fatty liver disease (NAFLD) is the hepatic manifestation of obesity and metabolic syndrome. Robust coagulation cascade activation is common in obese patients with NAFLD. We identified a critical temporal relationship between thrombin generation and the manifestation of hepatic steatosis, inflammation, and injury in C57BL/6J mice fed a high-fat diet (HFD) for 1, 2, and 3 months. Mice fed a HFD exhibited dramatic increases in hepatocellular injury and inflammation over time. Hepatic fibrin deposition preceded an increase in serum alanine aminotransferase, and the most dramatic changes in liver histopathology occurred in conjunction with a detectable increase in plasma thrombin-antithrombin levels at 3 months. To directly determine whether thrombin activity promotes NAFLD pathogenesis, mice were fed a HFD and simultaneously treated with the direct thrombin inhibitor dabigatran etexilate for 3 months. Notably, dabigatran treatment significantly reduced hepatic fibrin deposition, hepatic inflammation, hepatocellular injury, and steatosis in mice fed a HFD. Of interest, dabigatran treatment also significantly attenuated HFD-induced body weight gain. Gene expression analysis suggested that thrombin potentially drives NAFLD pathogenesis by altering the expression of genes associated with lipid metabolism and bile acid synthesis. Collectively, the results suggest that thrombin activity is central to HFD-induced body weight gain, liver injury, and inflammation and provide the proof-of-principle evidence that pharmacological thrombin inhibition could be effective in limiting NAFLD and associated pathologies. PMID:25138021

Kopec, Anna K; Joshi, Nikita; Towery, Keara L; Kassel, Karen M; Sullivan, Bradley P; Flick, Matthew J; Luyendyk, James P

2014-11-01

174

Thrombin generation in plasma of healthy adults and children: chromogenic versus fluorogenic thrombogram analysis.  

Science.gov (United States)

Coagulation tests and coagulation factor assays have been complemented recently with experimental tests to measure the total amount of thrombin formed. We have presently analyzed thrombin generation of healthy adult and paediatric plasma samples via a fluorogenic and a chromogenic method. The chromogenic method was performed on the fully automated Behring Coagulation System (BCS) and fluorogenic assays via Calibrated Automated Thrombography (CAT), after coagulation induction by various tissue factor (TF) concentrations. Sample distribution and variability were analyzed for the four main coagulation parameters, derived via computerized curve analysis in each method. Results for both methods were correlated. At the recommended TF concentration (300 pM), thrombin generation via BCS was less variable than via CAT (1-6 pM), but at comparable TF concentrations (1-6 pM), the CAT sensitivity was higher than that of BCS. Inhibition of intrinsic coagulation with the anti-factor VIII antibody BO2C11 revealed that the BCS detected extrinsic coagulation exclusively, at all TF concentrations tested. In contrast, at low TF concentrations (1 and 2.5 pM), via CAT, intrinsic coagulation pathway amplification was measured. At standardized TF concentrations (300 pM in BCS vs. 2.5 pM in CAT), different reference values between adults and children were found, for all parameters, except Tmax. In adult samples, the best correlation between both methods was observed for ETP(CAT) versus ETP(BCS) and for Peak height(CAT) versus Cmax(BCS), when thrombin generation was exclusively extrinsic (300 pM in BCS vs. 6 pM in CAT). In conclusion, differential thrombin generation characteristics in BCS and CAT are relevant for their clinical applicability. PMID:17849049

Devreese, Katrien; Wijns, Walter; Combes, Isabelle; Van kerckhoven, Soetkin; Hoylaerts, Marc F

2007-09-01

175

Auger recombination in silicon  

International Nuclear Information System (INIS)

The Auger recombination in n-Si is studied at room temperature using the technique of probing infrared light absorption (lambda approximately 2 ?m) by nonequilibrium carriers. Experimental value of the Auger recombination coefficient is determined (? approximately 1.5x10-30 cm6/s). At room temperature the phonon mechanism of the Auger recombination is shown to be more important than the resonance one

176

Recombination and Genetic Diversity  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english In this paper we present a spatial stochastic model for genetic recombination, that answers if diversity is preserved in an infinite population of recombinat-ing individuals distributed spatially. We show that, for finite times, recombination may maintain all the various potential different types, b [...] ut when time grows infinitely, the diversity of individuals extinguishes off. So under the model premisses, recombination and spatial localization alone are not enough to explain diversity in a population. Further we discuss an application of the model to a controversy regarding the diversity of "Major Histocompatibility Complex" (MHC).

T. C., Coutinho; T.T.da, Silva; G.L., Toledo.

177

Topics in Nuclear Astrophysics  

International Nuclear Information System (INIS)

Some topics in nuclear astrophysics are discussed, e.g.: highly evolved stellar cores, stellar evolution (through the temperature analysis of stellar surface), nucleosynthesis and finally the solar neutrino problem. (L.C.)

178

Topical photodynamic therapy  

Directory of Open Access Journals (Sweden)

Full Text Available Topical photodynamic therapy is a therapeutic modality in development, thus arises grate interest among dermatologists worldwide. It is an effective therapy for actinic keratosis, superficial BCC and Bowenos disease. Treatment efficacy, good cosmetics, low risk of skin cancer, low invasiveness, low rate of adverse events, facility for treating multiple or large lesions, especially in poor healing sites and, for penile, digital and facial involvement, low general toxicity and possibility of repeating the treatments with the same efficiency, enable topical photodynamic therapy to become increasingly practiced treatment modality. Researching aimed topical photodynamic therapy to prove as a treatment modality for clinical use in other dermatoses, is in experimental phase. To answer the question when dermatologist should consider using topical photodynamic therapy treatment modatility, we are present available date.

Polja?ki Mirjana

2006-01-01

179

Salicylic Acid Topical  

Science.gov (United States)

... to the skin that you are treating with topical salicylic acid unless your doctor tells you that you should: abrasive soaps or cleansers; skin care products that contain alcohol; other medications that are ...

180

Hancock research topics  

Science.gov (United States)

Here's what Greg Hancock writes about choosing topics his students will research during his Surface Processes class. You may also jump ahead to some examples that fit these criteria, or see some of the ...

 
 
 
 
181

Related Topics and News  

Science.gov (United States)

Related Topics and News Harold Varmus Appointed to President's Council of Advisors on Science and Technology Harold Varmus Attends White House Signing Harold Varmus Awarded Henry G. Friesen International Prize in Health Research Harold Varmus Featured

182

Topical treatment of melasma  

Directory of Open Access Journals (Sweden)

Full Text Available Melasma is a common hypermelanotic disorder affecting the face that is associated with considerable psychological impacts. The management of melasma is challenging and requires a long-term treatment plan. In addition to avoidance of aggravating factors like oral pills and ultraviolet exposure, topical therapy has remained the mainstay of treatment. Multiple options for topical treatment are available, of which hydroquinone (HQ is the most commonly prescribed agent. Besides HQ, other topical agents for which varying degrees of evidence for clinical efficacy exist include azelaic acid, kojic acid, retinoids, topical steroids, glycolic acid, mequinol, and arbutin. Topical medications modify various stages of melanogenesis, the most common mode of action being inhibition of the enzyme, tyrosinase. Combination therapy is the preferred mode of treatment for the synergism and reduction of untoward effects. The most popular combination consists of HQ, a topical steroid, and retinoic acid. Prolonged HQ usage may lead to untoward effects like depigmentation and exogenous ochronosis. The search for safer alternatives has given rise to the development of many newer agents, several of them from natural sources. Well-designed controlled clinical trials are needed to clarify their role in the routine management of melasma.

Bandyopadhyay Debabrata

2009-01-01

183

Cardiovascular and biochemical studies on the effects of thrombin and dabigatran and the interaction with vasopressor molecules  

Directory of Open Access Journals (Sweden)

Conclusion: The thrombin inhibitor, dabigatran reduces vascular oxidative stress and inflammation, improves endothelial function, and decreases atherosclerosis in rodents. [Int J Basic Clin Pharmacol 2014; 3(5.000: 874-878

R. Anand

2014-10-01

184

Oxygen-hydrogen recombiner  

International Nuclear Information System (INIS)

Purpose: To improve the oxygen-hydrogen removing performance, as well as enable to maintain the high performance in a range of increasing the processing gas flow rate within a recombiner with the reduced pressure and reduced amount of charged catalyst. Constitution: A plate-like metal catalyst comprising alumina added as a binder to the surface of a sponge-like metal support made of nickel-chromium alloy and particles of platinum type novel metal such as platinum or palladium having a catalytic activity supported on alumina is contained in a cartridge and filled within an oxygen-hydrogen recombiner. The recombiner is adapted so that the exhaust gas flow rate therein is within a range from 1 nm/sec to 4 nm/sec. It is possible with such a constitution to improve the recombining performance, reduce the pressure loss, decrease the size of the recombiner and facilitate the maintenance and check thereof. (Kawakami, Y.)

185

The impact of elective knee/hip replacement surgery and thromboprophylaxis with Rivaroxaban or Dalteparin on Thrombin Generation  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract Total hip/knee replacement surgeries are associated with an increased risk of venous thromboembolism and post-operative thromboprophylaxis has become standard treatment. The aims of this study were to: (1) assess the impact of hip/knee replacement surgery on ex vivo thrombin generation (TG), prothrombin-fragments 1+2 (F1+2), thrombin-antithrombin complexes (TAT) and D-dimer; (2) compare the anticoagulant effects of dalteparin and rivaroxaban on TG 24 hours after surgery. H...

1988-01-01

186

Structure of the thrombin complex with triabin, a lipocalin-like exosite-binding inhibitor derived from a triatomine bug.  

Science.gov (United States)

Triabin, a 142-residue protein from the saliva of the blood-sucking triatomine bug Triatoma pallidipennis, is a potent and selective thrombin inhibitor. Its stoichiometric complex with bovine alpha-thrombin was crystallized, and its crystal structure was solved by Patterson search methods and refined at 2.6-A resolution to an R value of 0.184. The analysis revealed that triabin is a compact one-domain molecule essentially consisting of an eight-stranded beta-barrel. The eight strands A to H are arranged in the order A-C-B-D-E-F-G-H, with the first four strands exhibiting a hitherto unobserved up-up-down-down topology. Except for the B-C inversion, the triabin fold exhibits the regular up-and-down topology of lipocalins. In contrast to the typical ligand-binding lipocalins, however, the triabin barrel encloses a hydrophobic core intersected by a unique salt-bridge cluster. Triabin interacts with thrombin exclusively via its fibrinogen-recognition exosite. Surprisingly, most of the interface interactions are hydrophobic. A prominent exception represents thrombin's Arg-77A side chain, which extends into a hydrophobic triabin pocket forming partially buried salt bridges with Glu-128 and Asp-135 of the inhibitor. The fully accessible active site of thrombin in this complex is in agreement with its retained hydrolytic activity toward small chromogenic substrates. Impairment of thrombin's fibrinogen converting activity or of its thrombomodulin-mediated protein C activation capacity upon triabin binding is explained by usage of overlapping interaction sites of fibrinogen, thrombomodulin, and triabin on thrombin. These data demonstrate that triabin inhibits thrombin via a novel and unique mechanism that might be of interest in the context of potential therapeutic applications. PMID:9342325

Fuentes-Prior, P; Noeske-Jungblut, C; Donner, P; Schleuning, W D; Huber, R; Bode, W

1997-10-28

187

Stability and bioactivity of thrombin binding aptamers modified with d-/l-isothymidine in the loop regions.  

Science.gov (United States)

Thrombin binding aptamer (TBA) is a 15-mer single-strand DNA that was identified by SELEX screening technology. It adopts a chair-type antiparallel G-quadruplex and can specifically interact with thrombin, thus inhibiting blood coagulation. Isonucleoside (isoNA) is a type of nucleoside isomer in which the base is shifted to 2'-positions of the glycosyl group, endowed with the ability to modulate local conformation of nucleotides, and l-isoNA could alter the conformation more due to the inversion of glycosyl configuration. Incorporation of l-isothymidine (l-isoT) at T3, T9, T12 positions and d-isoT at the T7 position in TBA's loop regions promoted the formation of G-quadruplex, resulting in enhanced affinity with thrombin and an increased anticoagulant effect. Computer simulation indicated that TBA-12L showed the strongest binding with thrombin, which was consistent with experimental results. The bioactivity of double isoNA incorporated TBA with d-IsoT at T7 and l-IsoT at T12 was comparable to that of TBA-12L, suggesting that the T12 of TBA was very important in interaction with thrombin. Our study also suggested that TBA might interact with two thrombin molecules through the TT loops (T3T4, T12T13) and TGT loop, but the second bonding did not show additional biological effects. PMID:25264858

Cai, Baobin; Yang, Xiantao; Sun, Lidan; Fan, Xinmeng; Li, Liyu; Jin, Hongwei; Wu, Yun; Guan, Zhu; Zhang, Liangren; Zhang, Lihe; Yang, Zhenjun

2014-10-21

188

Effect of thrombin concentration on the adhesion strength and clinical application of fibrin glue-soaked sponge.  

Science.gov (United States)

Fibrin glue-soaked gelatin sponge (FGGS) has been used for tissue sealing in neurosurgical practice, but too rapid clotting of fibrin glue occasionally prevents good fixation of FGGS. Dilution of thrombin may provide adequate manipulation time between mixing fibrinogen and thrombin on gelatin sponge and application into the tissue defects. The present study characterized the effect of thrombin dilution on the adhesion strength of FGGS and retrospectively assessed the clinical usage of the dilution for filling dead space or sealing arachnoid defect in 255 cases who underwent transsphenoidal surgery for the last 66 months. FGGS was prepared using three different concentrations of thrombin: 250 (standard), 50 (1:5 dilution), and 25 (1:10 dilution) units/ml, and incubated for three different periods (5, 20, and 60 seconds). FGGSs were applied over two adjacently positioned porcine skins placed on two metallic plates. The adhesion strength was evaluated by measuring maximum tensile strength during pulling out the sliding plate at a constant rate of displacement. The maximum adhesion strength was greater for FGGS with 1:10 diluted thrombin solution than for FGGS prepared with higher concentrations (p < 0.05). Adhesion strength did not decay for 20 seconds after the mixture. Only four of 255 cases (1.6%) required second reconstruction of sella floor due to the cerebrospinal fluid leakage. FGGS prepared with diluted thrombin solution can provide adequate adhesion strength for clinical use. PMID:23358164

Campos, Francia; Fujio, Shingo; Sugata, Sei; Tokimura, Hiroshi; Hanaya, Ryosuke; Bohara, Manoj; Arita, Kazunori

2013-01-01

189

Thrombin effectuates therapeutic arteriogenesis in the rabbit hindlimb ischemia model: A quantitative analysis by computerized in vivo imaging  

Science.gov (United States)

We report on an experimental mammalian controlled study that documents arteriogenic capacity of thrombin and utilizes computerized algorithms to quantify the newly formed vessels. Hindlimb ischemia was surgically invoked in 10 New Zealand white rabbits. After quiescence of endogenous angiogenesis heterologous bovine thrombin was intramuscularly injected (1500 units) in one hindlimb per rabbit (Group T). Contralateral limbs were infused with normal saline (Group C). Digital subtraction angiography (DSA) of both limbs was performed after thrombin infusion by selective cannulation of the abdominal aorta and digital images were post-processed with computerized algorithms in order to enhance newly formed vessels. Total vessel area and total vessel length were quantified. In vivo functional evaluation included measurements of blood flow volume at the level of the external iliac artery by Doppler ultrasonography both at baseline and at 20 days after thrombin infusion. Total vessel area and length (in pixels) were 14,713±1023 and 5466±1327 in group T versus 12,015±2557 and 4598±1269 in group C ( p=0.0062 and 0.1526, respectively). Blood flow volumes (ml/min) at baseline and at 20 days after thrombin infusion were 25.87±11.09 and 38.06±11.72 in group T versus 26.57±11.19 and 20.35±7.20 in group C ( p=0.8898 and 0.0007, respectively). Intramuscular thrombin effectuates an arteriogenic response in the rabbit hindlimb ischemia model. Computerized algorithms may enable accurate quantification of the neovascularization outcome.

Kagadis, George C.; Karnabatidis, Dimitrios; Katsanos, Konstantinos; Diamantopoulos, Athanassios; Samaras, Nikolaos; Maroulis, John; Siablis, Dimitrios; Nikiforidis, George C.

2006-12-01

190

Liquid crystal-based detection of thrombin coupled to interactions between a polyelectrolyte and a phospholipid monolayer.  

Science.gov (United States)

Herein, we describe a real-time, label-free biosensing strategy for thrombin detection that uses the orientational properties of nematic liquid crystals (LCs) and the interactions between a polyelectrolyte and a phospholipid monolayer. The imaging principle is based on the disruption of the orientation of 4-cyano-4'-pentylbiphenyl by reorganized 1,2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (DOPG) at the aqueous/LC interface. Positively charged, multiple arginine peptides (poly-l-arginine hydrochloride) interacted with negatively charged DOPG at the aqueous/LC interface, which caused reorganization of the phospholipid layer and induced an orientational transition of LCs from a homeotropic to a planar state. As a result, a dark to bright shift in the optical response was observed. Thrombin cleaves poly-l-arginine hydrochloride into peptides. Thus, when thrombin was added, the optical signals generated by the LCs reverted from bright to dark because of the weakened ability of the fragments to induce electrostatic interactions. The limit of detection of the LC-based sensor was 0.25ng/mL (6.7pM) thrombin, and the sensor was fully reusable. The detection limit of our LC-based interface sensor is 600 times lower than that of a previously reported enzyme-linked aptamer assay for the detection of thrombin. Thus, we have established a new, simple thrombin biosensor with high sensitivity and low interference. PMID:24708935

Zhang, Minmin; Jang, Chang-Hyun

2014-06-15

191

Topical Acne Treatments and Pregnancy  

Science.gov (United States)

... or visit us online at: www.OTISpregnancy.org . Topical Acne Treatments and Pregnancy In every pregnancy, a ... advice from your health care professional. What are topical acne treatments? Topical acne treatments are medications applied ...

192

Topics in Physical Mathematics  

CERN Document Server

This title adopts the view that physics is the primary driving force behind a number of developments in mathematics. Previously, science and mathematics were part of natural philosophy and many mathematical theories arose as a result of trying to understand natural phenomena. This situation changed at the beginning of last century as science and mathematics diverged. These two fields are collaborating once again; 'Topics in Mathematical Physics' takes the reader through this journey. The author discusses topics where the interaction of physical and mathematical theories has led to new points o

Marathe, Kishore

2010-01-01

193

Persistent protease-activated receptor 4 signaling mediates thrombin-induced microglial activation.  

Science.gov (United States)

We have previously reported that thrombin, the ultimate serine protease in the coagulation cascades, is a proinflammatory agent that causes proliferation and activation of brain microglial cells. However, participation of its principal receptor, the protease-activated receptor 1 (PAR1) appears to be limited to promoting microglial proliferation and not induction of inflammatory mediators. In the present study, we now report that thrombin action in promoting inflammatory mediators from brain microglia is mediated through another thrombin receptor, PAR4. Here we show that the PAR4 agonist peptide (PAR4AP, GYPGKF), but not the PAR1AP (TRAP, SFLLRN), induced tumor necrosis factor-alpha (TNF-alpha) production not only in cultured murine microglial cells in vitro but also in rat cortex in vivo. Down-regulation of PAR4 expression in microglial cultures by a specific antisense, but not a sense, oligonucleotide reduced PAR4AP-induced TNF-alpha. Mechanistic studies indicated that, in comparison with PAR1 signaling, prolonged increase of [Ca2+]i and phosphorylation of p44/42 mitogen-activated protein kinases, as well as NFkappaB activation may be responsible for PAR4AP-induced TNF-alpha production in microglia. Taken together, these results demonstrate that PAR4 activation mediates the potentially detrimental effects of thrombin on microglia, implying that perspectives of exploiting PAR1 as a potential anti-inflammatory target should be shifted toward PAR4 as a much more specific therapeutic target in brain inflammatory conditions associated with neurotrauma and neurodegenerations. PMID:12775717

Suo, Zhiming; Wu, Min; Citron, Bruce A; Gao, Chenhua; Festoff, Barry W

2003-08-15

194

Effects of the oral, direct thrombin inhibitor dabigatran on five common coagulation assays  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Dabigatran is an oral, reversible thrombin inhibitor that has shown promising results in large clinical trials. Laboratory monitoring is not needed but the effects on common coagulation assays are incompletely known. Dabigatran was added to plasma from healthy subjects in the concentration range 0-1,000 mu g/l and analysed using several reagents for activated thromboplastin time (APTT), prothrombin time (PT), fibrinogen, antithrombin, and activated protein C resistance. Typical trough concent...

Lindahl, Tomas; Baghaei, Fariba; Fagerberg Blixter, Inger; Gustafsson, Kerstin; Stigendal, Lennart; Sten-linder, Margareta; Strandberg, Karin; Hillarp, Andreas

2011-01-01

195

[Dabigatran etixilate (Pradaxa): oral anticoagulant acting as direct selective inhibitor of thrombin].  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Le dabigatran étexilate (Pradaxa®), administrable par voie orale, inhibe directement et sélectivement la thrombine (facteur II a) et, à ce titre, exerce un effet anticoagulant. Pradaxa® ne nécessite pas de surveillance ou d’adaptation de la dose, excepté en cas d’insuffisance rénale modérée ou chez les sujets âgés de plus de 75 ans, ainsi qu’en cas de traitement concomitant par amiodarone ou vérapamil. Il est actuellement indiqué pour la prophylaxie de la thromboembolie ve...

Lancellotti, Patrizio; Scheen, Andre?

2010-01-01

196

Dabigatran Etexilate, A Novel Oral Direct Thrombin Inhibitor, for Preventing Thromboembolic Events After Knee Replacement Arthroplasty  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Background: Dabigatran etexilate is one of the few direct thrombin inhibitors with anti-coagulant activities and the following distinctive features: taken orally, no need to closely monitor for complications, and no need for regular dose adjustments. Relying on the above mentioned valuable advantages, dabigatran etexilate can be considered as a premier choice for the prevention of venous thromboembolism after knee replacement arthroplasty. Methods: Forty five patients undergoing 50 knee repla...

Yegane A; Farahini H; Shahhoseini Gh; Moghtadaee M; Rajabpour S

2012-01-01

197

Modifying the substrate specificity of Carcinoscorpius rotundicauda serine protease inhibitor domain 1 to target thrombin.  

Science.gov (United States)

Protease inhibitors play a decisive role in maintaining homeostasis and eliciting antimicrobial activities. Invertebrates like the horseshoe crab have developed unique modalities with serine protease inhibitors to detect and respond to microbial and host proteases. Two isoforms of an immunomodulatory two-domain Kazal-like serine protease inhibitor, CrSPI-1 and CrSPI-2, have been recently identified in the hepatopancreas of the horseshoe crab, Carcinoscorpius rotundicauda. Full length and domain 2 of CrSPI-1 display powerful inhibitory activities against subtilisin. However, the structure and function of CrSPI-1 domain-1 (D1) remain unknown. Here, we report the crystal structure of CrSPI-1-D1 refined up to 2.0 Å resolution. Despite the close structural homology of CrSPI-1-D1 to rhodniin-D1 (a known thrombin inhibitor), the CrSPI-1-D1 does not inhibit thrombin. This prompted us to modify the selectivity of CrSPI-1-D1 specifically towards thrombin. We illustrate the use of structural information of CrSPI-1-D1 to modify this domain into a potent thrombin inhibitor with IC(50) of 26.3 nM. In addition, these studies demonstrate that, besides the rigid conformation of the reactive site loop of the inhibitor, the sequence is the most important determinant of the specificity of the inhibitor. This study will lead to the significant application to modify a multi-domain inhibitor protein to target several proteases. PMID:21188150

Giri, Pankaj Kumar; Tang, Xuhua; Thangamani, Saravanan; Shenoy, Rajesh T; Ding, Jeak Ling; Swaminathan, Kunchithapadam; Sivaraman, J

2010-01-01

198

The Emerging Role of the Thrombin Receptor (PAR-1) in Melanoma Metastasis - a Possible Therapeutic Target  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Melanoma remains as the deadliest form of skin cancer with limited and inefficient treatment options available for patients with metastatic disease. Within the last decade, the thrombin receptor, Protease Activated Receptor-1, has been described as an essential gene involved in the progression of human melanoma. PAR-1 is known to activate adhesive, invasive and angiogenic factors to promote melanoma metastasis. It is overexpressed not only in metastatic melanoma cell lines but is also highly ...

Villares, Gabriel J.; Zigler, Maya; Bar-eli, Menashe

2011-01-01

199

Successful endovascular treatment of a hemodialysis graft pseudoaneurysm by covered stent and direct percutaneous thrombin injection.  

LENUS (Irish Health Repository)

Vascular access for hemodialysis remains a challenge for nephrologists, vascular surgeons, and interventional radiologists alike. Arteriovenous fistula and synthetic grafts remain the access of choice for long-term hemodialysis; however, they are subject to complications from infection and repeated needle cannulation. Pseudoaneurysms are an increasingly recognized adverse event. At present, there are many minimally invasive methods to repair these wall defects. We present a graft pseudoaneurysm, which required a combination of endovascular stent graft placement and percutaneous thrombin injection for successful occlusion.

Keeling, Aoife N

2011-07-25

200

Histamine and thrombin modulate endothelial focal adhesion through centripetal and centrifugal forces.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We examined the contribution of actin-myosin contraction to the modulation of human umbilical vein endothelial cell focal adhesion caused by histamine and thrombin. Focal adhesion was measured as the electrical resistance across a cultured monolayer grown on a microelectrode. Actin-myosin contraction was measured as isometric tension of cultured monolayers grown on a collagen gel. Histamine immediately decreased electrical resistance but returned to basal levels within 3-5 min. Histamine did ...

Moy, A. B.; Engelenhoven, J.; Bodmer, J.; Kamath, J.; Keese, C.; Giaever, I.; Shasby, S.; Shasby, D. M.

1996-01-01

 
 
 
 
201

Postpyelolithotomy Renal Artery Pseudoaneurysm Management with Percutaneous Thrombin Injection: A Case Report  

International Nuclear Information System (INIS)

Renal artery pseudoaneurysm leading to life-threatening hematuria can occur after a surgical procedure such as pyelolithotomy, albeit rarely. With recent advances in transarterial embolization techniques, this minimally invasive procedure has become the treatment of choice, replacing surgery. We present a case of massive hematuria due to renal artery pseudoaneurysm developing after pyelolithotomy that was managed with percutaneus thrombin injection directly into the pseudoaneurysm

202

An easy way for the rapid purification of recombinant proteins from Helicobacter pylori using a newly designed expression vector.  

Science.gov (United States)

We constructed a H. pylori expression vector which consisted of both a His-tag and a GST tag as purification tools for recombinant protein and a chloramphenicol resistant cat gene as a reporter. The backbone of the vector pBK contained an ColEI origin of replication and a kanamycin resistant gene. A set of oligos for the His-tag and the PCR product of gst (glutathione S-transferase) gene were inserted sequentially in frame in the multi-cloning site of pBK. The orf of cat was inserted downstream of the gst to generate pBKHGC. The 3' part of H. pylori clpB and flaA were cloned into the vector which was introduced into H. pylori. Recombinant proteins were purified by GSH affinity column, digested with thrombin and were analyzed by western blotting. The final recombinant proteins were successfully purified. PMID:24972810

Kang, Hyung-Lyun; Jo, Jin-Sung; Kwon, Soon-Uck; Song, Jae-Young; Seo, Ji-Hyun; Cho, Myung-Je; Baik, Seung-Chul; Youn, Hee-Shang; Rhee, Kwang-Ho; Lee, Woo-Kon

2014-07-01

203

Direct thrombin inhibitor-bivalirudin functionalized plasma polymerized allylamine coating for improved biocompatibility of vascular devices.  

Science.gov (United States)

The direct thrombin inhibitor of bivalirudin (BVLD), a short peptide derived from hirudin, has drawn an increasing attention in clinical application because it is safer and more effective than heparin for diabetic patients with moderate- or high-risk for acute coronary syndromes (ACS). In this study, BVLD was covalently conjugated on plasma polymerized allylamine (PPAam) coated 316L stainless steel (SS) to develop an anticoagulant surface. QCM-D real time monitoring result shows that 565±20 ng/cm2 of BVLD was bound to the PPAam surface. Infrared spectroscopy (IR) and X-ray photoelectron spectroscopy (XPS) confirmed the immobilization of BVLD. The conjugation of BVLD onto the PPAam coating led to enhanced binding of thrombin, and the activity of the thrombin adsorbed on its surface was effectively inhibited. As a result, the BVLD immobilized PPAam (BVLD-PPAam) substrate prolonged the clotting times, and exhibited inhibition in adhesion and activation of platelets and fibrinogen. We also found that the BVLD-PPAam coating significantly enhanced endothelial cell adhesion, proliferation, migration and release of nitric oxide (NO) and secretion of prostaglandin I2 (PGI2). In vivo results indicate that the BVLD-PPAam surface restrained thrombus formation by rapidly growing a homogeneous and intact endothelium on its surface. These data suggest the potential of this multifunctional BVLD-PPAam coating for the application not only in general vascular devices such as catheters, tubes, oxygenator, hemodialysis membranes but also vascular grafts and stents. PMID:22877639

Yang, Zhilu; Tu, Qiufen; Maitz, Manfred F; Zhou, Shuo; Wang, Jin; Huang, Nan

2012-11-01

204

Identification of a novel thrombin-like phospholipase A2 from Gloydius ussuriensis snake venom.  

Science.gov (United States)

The coagulant effects of phospholipase A2 with Gln at 49 sites (Gln49-PLA2), purified from Gloydius ussurensis snake venom, were investigated on human citrated plasma and fibrinogen. Gln49-PLA2 clotted human plasma dose-dependently from 180.67 +/- 1.86 s to 19.00 +/- 0.58 s, and reduced the re-calcification time from 7.46 +/- 1.17 to 0.75 +/- 0.33 min and the prothrombin time from 12.4 +/- 0.29 s to 6.95 +/- 0.20 s, but it could not activate factor XIII, and the procoagulant effects were inhibited by heparin. The specific clotting activities of Gln49-PLA2 were equivalent to 1100 NIH thrombin U/mg on human fibrinogen, and the specific arginine esterase activity on the substrate BAEE was 1747 U/mg. Gln49-PLA2 hydrolyzed fibrinopeptide A faster than fibrinopeptide B, and the fibrinongenolytic ability was inhibited by the serine protease inhibitor phenyl-methylsulphonyl fluoride, but not by the metalloprotease inhibitor ethylenediamine tetraacetic acid. This finding demonstrates that Gln49-PLA2 is consistent with thrombin-like properties, and therefore should be a new thrombin-like serine protease. PMID:17982312

Zhang, Qiuyan; Wang, Jingyun; Han, Ying; Xie, Qun; An, Lijia; Bao, Yongming

2007-12-01

205

[Heparin cofactor II, a thrombin inhibitor with a still not clarified physiologic role].  

Science.gov (United States)

Heparin Cofactor II (HCII) is a glycoprotein in human plasma which inactivates thrombin rapidly in the presence of dermatan sulfate. Inhibition occurs by formation of a stable equimolar complex between HCII and thrombin. HCII association with thrombotic events has not always been observed, thus decreased HCII does not appear to be a strong risk factor for thromboembolic events. Reduced HCII levels have been detected in different clinical conditions, such as hepatic failure, disseminated intravascular coagulation, thalasemina, sickle cell anemia. Increased physiological levels have been found in pregnant women and oral contraception. In our laboratory, we measured HCII plasmatic levels in the normal Buenos Aires city population and in patients under different clinical conditions, such as sepsis, diabetis, burns, oral anticoagulation and in patients treated with heparin, hyperhomcysteinemia in whom septic and diabetic patients showed decreased values. HCII thrombin inhibition possibly takes place in extravascular sites where dermatan sulfate is present. HCII activity would be important in the regulation of wound healing, inflammation, or neuronal development. PMID:10349131

Rossi, E B; Duboscq, C L; Kordich, L C

1999-01-01

206

Posttraumatic lingual artery pseudoaneurysm treated with ultrasound-guided percutaneous thrombin injection.  

Science.gov (United States)

Pseudoaneurysms of the lingual artery are extremely rare and are commonly iatrogenic in nature or less frequently a result of blunt or penetrating trauma. Traditionally, these vascular abnormalities have been repaired with open or endovascular techniques. Although ultrasound-guided percutaneous thrombin injection has become a standard treatment for superficial pseudoaneurysms, there are no reports of this being used in the treatment of lingual artery pseudoaneurysms. We report the case of a 26-year-old man who suffered a penetrating head and neck injury after an improvised explosive device blast in Iraq who presented with persistent oropharyngeal swelling. Color-flow Doppler ultrasonography revealed the classic yin/yang sign of a pseudoaneurysm, and a computed tomography scan was obtained that revealed a right lingual artery pseudoaneurysm. With the lack of endovascular capabilities and the excessive risk of open surgery, thrombin was injected directly into the pseudoaneurysm under ultrasound guidance. A computed tomography scan and Doppler ultrasonography revealed complete resolution of the aneurysm. This article presents the first reported case in the English literature of a lingual artery aneurysm after penetrating trauma managed successfully with ultrasound-guided percutaneous thrombin injection. PMID:24365080

Masella, Pamela C; Hanson, Megan M; Hall, Brian T; Verghese, John J; Kellicut, Dwight C

2014-07-01

207

Does thrombin stimulation of human platelets proceed via a simultaneous Na+-H+ exchange?  

International Nuclear Information System (INIS)

Thrombin stimulation of human platelets initiates a membrane depolarization attributable to a Na+ influx into, and an alkalinization of, the cytoplasm, both of which follow a similar rapid time scale and thrombin dose dependence. These responses precede secretion of the contents of dense granules (serotonin) and, after 1 min, of lysosomes (?-glucuronidase). These markers have been used to determine whether the Na+ influx and H+ efflux are sequential or simultaneous. They have examined these parameters in D2O-Hepes buffers. NMR evidence indicates that equilibration is rapid, and virtually complete within the 3 minute pre-stimulation platelets equilibration period. The rate of depolarization is 70-80% slower in D2O than in H2O. The time to reach maximal depolarization is 5-10 sec longer, the extent of depolarization 60% inhibited, and the [H+] change 85-100% inhibited. The serotonin secretion is unaltered, and the ?-glucuronidase secretion is 130-180% enhanced. 10-4 M amiloride inhibits Na+ influx, i.e. depolarization, and the pH change completely. Adjustment to pH/sub i/ 7.3 with NH4Cl led to a 30-80% enhanced ?-glucuronidase release upon thrombin exposure. These results suggest that the Na+ and H+ fluxes across the platelet membrane occur sequentially, the Na+ occurring first. Furthermore, granule secretion, previously shown by us to be independent of the existent Na+ gradient, depends on the cytoplasmic K+ and H+ concentrations

208

Topic Tracking with Dynamic Topic Model and Topic-based Weighting Method  

Directory of Open Access Journals (Sweden)

Full Text Available In topic tracking, a topic is usually described by several stories. How to represent a topic is always an issue and a difficult problem in the research on topic tracking. To emphasis the topic in stories, we provide an improved topic-based tf*idf weighting method to measure the topical importance of the features in the representation model. To overcome the topic drift problem and filter the noise existed in the tracked topic description, a dynamic topic model is proposed based on the static model. It extends the initial topic model with the information from the incoming related stories and filters the noise using the latest unrelated story. The topic tracking systems are implemented on the TDT4 Chinese corpus. The experimental results indicate that both the new weighting method and the dynamic model can improve the tracking performance.

Xiaoyan Zhang

2010-05-01

209

Selected topics in magnetism  

CERN Document Server

Part of the ""Frontiers in Solid State Sciences"" series, this volume presents essays on such topics as spin fluctuations in Heisenberg magnets, quenching of spin fluctuations by high magnetic fields, and kondo effect and heavy fermions in rare earths amongst others.

Gupta, L C

1993-01-01

210

Topical anesthesia in phacoemulsification  

Directory of Open Access Journals (Sweden)

Full Text Available Purpose : To evaluate the efficacy of topical anesthesia; topical Benoxinate 0.4% (Oxybuprocaine and Xylocaine (Lidocaine gel, in selected cataract patients as an alternative to peribulbar or retrobulbar block anesthesia during cataract surgery. Materials and Methods : Prospective non-comparative evaluation of patients? and surgeon?s satisfaction at the end of the procedure. Three hundred patients (300 eyes were included in the study. The procedure was explained to patients with details regarding what will happen and what to expect during surgery. All patients received topical anesthesia with Benoxinate 0.4% eye drops and Xylocaine gel 2%. All surgeries were done by the same surgeon using the same machine (updated LEGACY phacoemulsifier, Alcon and approach (clear corneal incision and followed by a foldable intraocular lens (IOL implantation. Results : None of the patients had severe pain during the procedure; only 2% (six of 300 required use of intravenous sedation (Propofol, both the surgeon?s and the patients? satisfaction were high. Eye movements and blepharospasm were not significant problems, and no serious complications occurred. Rate of vitreous loss due to posterior capsule tear/rupture was within literature reported range and not different from our previous experience. Conclusion : Topical anesthesia is a satisfactory and safe alternative to retrobulbar and peribulbar anesthesia for clear corneal phacoemulsification and intraocular lens implantation in selected cataract patients in the hands of experienced cataract surgeon.

Waheeb Saad

2010-01-01

211

Topic in Depth - Antimatter  

Science.gov (United States)

For every action there is a reaction, for every plus there is a minus, and for every matter there is wellâ¦antimatter. This topic in depth tackles the complex concept of antimatter, from the Big Bang to solar explosions and the technology to detect it.

2010-09-15

212

Topical immunomodulators in dermatology  

Directory of Open Access Journals (Sweden)

Full Text Available Topical immunomodulators are agents that regulate the local immune response of the skin. They are now emerging as the therapy of choice for several immune-mediated dermatoses such as atopic dermatitis, contact allergic dermatitis, alopecia areata, psoriasis, vitiligo, connective tissue disorders such as morphea and lupus erythematosus, disorders of keratinization and several benign and malignant skin tumours, because of their comparable efficacy, ease of application and greater safety than their systemic counterparts. They can be used on a domiciliary basis for longer periods without aggressive monitoring. In this article, we have discussed the mechanism of action, common indications and side-effects of the commonly used topical immunomodulators, excluding topical steroids. Moreover, newer agents, which are still in the experimental stages, have also been described. A MEDLINE search was undertaken using the key words "topical immunomodulators, dermatology" and related articles were also searched. In addition, a manual search for many Indian articles, which are not indexed, was also carried out. Wherever possible, the full article was reviewed. If the full article could not be traced, the abstract was used.

Khandpur Sujay

2004-04-01

213

Activated recombinant adenovirus proteinases  

Science.gov (United States)

This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying said peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described.

Anderson, Carl W. (Stony Brook, NY); Mangel, Walter F. (Shoreham, NY)

1999-08-10

214

Investigation of the thrombin-generating capacity, evaluated by thrombogram, and clot formation evaluated by thrombelastography of platelets stored in the blood bank for up to 7 days  

DEFF Research Database (Denmark)

BACKGROUND AND OBJECTIVES: Transfusion based on the Thrombelastograph (TEG) results reduces transfusion requirements in cardiac surgery and in liver transplantation. Taking the pivotal role of thrombin generation in the coagulation process into consideration, the clinical utility of the TEG may, in part, depend on its reflection of the dynamics of thrombin generation. MATERIAL AND METHODS: The kinetics of thrombin generation of platelets stored for 2 and 7 days, respectively, was assessed by calibrated automated thrombogram (CAT) and the lag time (min), time to peak (ttPeak; min), peak (nm thrombin) and endogenous thrombin potential (ETP; nm thrombin*min) were registered. Clot formation was evaluated by TEG and the R time (min), maxial amplitude (MA; mm), time to maximum thrombus generation (TMG; min) and maximum thrombus generation (MTG; dynes cm(-2) s(-1)) and total thrombus generation (TTG; dyne cm(-2)) were registered. RESULTS: Platelets become more procoagulant, evaluated both by TEG and CAT during storage. The reduction in CAT lag time and the ttPeak correlated with a decrease in the TEG R time and TMG (P < 0.0001) as did the CAT peak thrombin generation and the TEG MTG (P = 0.0035). No correlation between ETP and TTG was found (P = 0.65). CONCLUSION: The kinetics of thrombin generation, as evaluated by CAT, correlates with the thrombus generation, as evaluated by thrombelastography and this may in part explain the clinical utility of the TEG in identifying clinically relevant coagulopathies, secondary to impaired thrombin generation Udgivelsesdato: 2008/2

Johansson, Per Ingemar; Svendsen, M.S.

2008-01-01

215

Protein C inhibits endocytosis of thrombin-thrombomodulin complexes in A549 lung cancer cells and human umbilical vein endothelial cells  

International Nuclear Information System (INIS)

We investigated the effect of protein C on the endocytosis of thrombin-thrombomodulin complexes. We previously showed that exposure of umbilical vein endothelial cells to thrombin stimulated the internalization and degradation of thrombin. A similar internalization was stimulated by a monoclonal antithrombomodulin antibody. We have repeated these studies in the presence of protein C and found that endocytosis of 125I-thrombin-thrombomodulin complexes, but not 125I-antithrombomodulin-thrombomodulin complexes, is inhibited. Activated protein C did not inhibit endocytosis of thrombin-thrombomodulin complexes. Protein C inhibited both internalization and degradation of 125I-thrombin and diisopropylphosphoryl (DIP) 125I-thrombin in human lung cancer cells (A549). These effects were observed at protein C concentrations found in human plasma. Protein S had no effect on the inhibition of endocytosis of thrombin-thrombomodulin complexes by protein C. We propose that protein C may regulate the rate of endocytosis of thrombin-thrombomodulin complexes in vivo and thereby control the capacity for endothelium to activate protein C

216

New topical antifungal drugs.  

Science.gov (United States)

The new antifungal drugs used for topical treatment of superficial, skin and mucosal mycoses are reviewed. Amorolfine and allylamines (naftifine and terbinafine) are promising original molecules with new and different modes of action against fungi. Rilopirox is a new pyridone derivative under study. A great number of azole derivatives, such as oxiconazole, isoconazole, sulconazole, and terconazole, are used as topical antifungals. Three of them are synthesized in Barcelona by pharmaceutical laboratories: sertaconazole, flutrimazole and eberconazole. All of them are now in the register process for commercialization. The combination of antifungals with active products, such as keratoplastics, is used mainly for the treatment of onychomycoses; 40% urea associated with 1% bifonazole has shown high efficacy for this indication. PMID:8118161

Torres-Rodríguez, J M

1993-01-01

217

Topics in String Phenomenology  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We consider two questions in string ``phenomenology.'' First, are there any generic string predictions? Second, are there any general lessons which string theory suggests for thinking about low energy models, particularly in the framework of supersymmetry? Among the topics we consider are the squark and slepton spectrum, flavor symmetries, discrete symmetries including $CP$, and Peccei-Quinn symmetries. We also note that in some cases, discrete symmetries can be used to co...

Dine, Michael

1993-01-01

218

Topics on String Phenomenology  

Digital Repository Infrastructure Vision for European Research (DRIVER)

These lectures present some topics of string phenomenology and contain two parts. In the first part, I review the possibility of lowering the string scale in the TeV region, that provides a theoretical framework for solving the mass hierarchy problem and unifying all interactions. The apparent weakness of gravity can then be accounted by the existence of large internal dimensions, in the submillimeter region, and transverse to a braneworld where our universe must be confined...

Antoniadis, I.

2007-01-01

219

Superconductivity elementary topics  

CERN Document Server

This book describes the elementary concepts of superconductivity and discusses the topics of flux-lattice melting, magnetization including the para-Meissner effect, microwave absorption, a.c. resistivity along with the London penetration depth, the Mössbauer effect, levitation, fractals and nuclear magnetic resonance. There are appendices covering superconducting compounds, the isotope effect, symmetries, the pseudogap, relativistic superconductivity, the Cherenkov effect and soft vortices. Also included is an appendix on the quantum Hall effect. In all of the chapters, the theoretical descrip

Shrivastava, KN

2000-01-01

220

Topics in industrial mathematics  

International Nuclear Information System (INIS)

Mathematical methods are widely used to solve practical problems arising in modern industry. This article outlines some of the topics relevant to AECL programmes. This covers the applications of transmission and neutron transport tomography to determine density distributions in rocks and two phase flow situations. Another example covered is the use of variational methods to solve the problems of aerosol migration and control theory. (author). 7 refs

 
 
 
 
221

Topical Methotrexate In Localized Psoriasis  

Directory of Open Access Journals (Sweden)

Full Text Available A comparative study of topical methotrexate and topical tar in eighteen patients of localized psoriasis with bilateral involvement of both lower legs, equal in area and disease severity was carried out. The patients were asked to apply topical methorexate 0.25% in a hydrophilic gel twice daily on right leg. The test sites were score, before therapy, after one month and after two months. The result with topical methotrexate preparation was promising but was comparable to topical tar formulation.

Rath Namita

2004-01-01

222

Thrombin- and histamine-induced signal transduction in human endothelial cells. Stimulation and agonist-dependent desensitization of protein phosphorylation.  

Science.gov (United States)

Treatment of human endothelial cells with thrombin, histamine, or dioctanoylglycerol (DiC8), a synthetic diacylglycerol, resulted in the rapid and transient phosphorylation of a Mr = 29,000 protein (P29) in a dose-dependent manner. Various tumor promoters also promoted P29 phosphorylation while the adenylate cyclase activator, forskolin, did not. The level of phosphorylation with all three agonists was similar (2.5-4 fold), and analysis of P29 by two-dimensional gel electrophoresis revealed identical patterns in each case. Receptor specificity was demonstrated for the histamine-stimulated changes; pyrilamine (10(-6) M; H1) but not cimetidine (10(-4); H2) blocked the response. The thrombin effect was active site-dependent. Phosphorylation induced by thrombin and histamine occurred within 1 min, peaked between 5 and 10 min, and returned to control levels by 1 h. DiC8-induced phosphorylation occurred more slowly but was also reduced by 1 h while phorbol ester treatment prolonged phosphorylation for at least 4 h. Treatment of these cells with thrombin or histamine for 1 h desensitized P29 to further phosphorylation by the homologous agonist although secondary phosphorylation could occur with heterologous compounds. However, if the primary agonist was removed following the onset of a desensitized state, secondary phosphorylation of P29 could be stimulated by the same compound. These same results were observed with two other phosphoproteins Mr = 18,000 (P18) and 80,000 (P80) which became more highly phosphorylated in response to thrombin treatment and with histamine/thrombin-stimulated prostaglandin I2 production. In contrast, homologous down-regulation of P29 phosphorylation was not observed with DiC8-treated cells, and the decline in phosphorylated P29 was associated with the loss of functional DiC8. The protein kinase inhibitors staurosporine and H-7 blocked P18 and P80 phosphorylation by thrombin but had no effect on P29 phosphorylation by histamine, thrombin, or DiC8 suggesting distinct pathways leading to the phosphorylation of these different proteins. These data suggest that multiple and independent thrombin/histamine-induced events are susceptible to receptor occupancy-dependent homologous down-regulation. PMID:1985891

Levin, E G; Santell, L

1991-01-01

223

Regulation of Meiotic Recombination  

Energy Technology Data Exchange (ETDEWEB)

Meiotic recombination results in the heritable rearrangement of DNA, primarily through reciprocal exchange between homologous chromosome or gene conversion. In plants these events are critical for ensuring proper chromosome segregation, facilitating DNA repair and providing a basis for genetic diversity. Understanding this fundamental biological mechanism will directly facilitate trait mapping, conventional plant breeding, and development of genetic engineering techniques that will help support the responsible production and conversion of renewable resources for fuels, chemicals, and the conservation of energy (1-3). Substantial progress has been made in understanding the basal recombination machinery, much of which is conserved in organisms as diverse as yeast, plants and mammals (4, 5). Significantly less is known about the factors that regulate how often and where that basal machinery acts on higher eukaryotic chromosomes. One important mechanism for regulating the frequency and distribution of meiotic recombination is crossover interference - or the ability of one recombination event to influence nearby events. The MUS81 gene is thought to play an important role in regulating the influence of interference on crossing over. The immediate goals of this project are to use reverse genetics to identify mutants in two putative MUS81 homologs in the model plant Arabidopsis thaliana, characterize those mutants and initiate a novel forward genetic screen for additional regulators of meiotic recombination. The long-term goal of the project is to understand how meiotic recombination is regulated in higher eukaryotes with an emphasis on the molecular basis of crossover interference. The ability to monitor recombination in all four meiotic products (tetrad analysis) has been a powerful tool in the arsenal of yeast geneticists. Previously, the qrt mutant of Arabidopsis, which causes the four pollen products of male meiosis to remain attached, was developed as a facile system for assaying recombination using tetrad analysis in a higher eukaryotic system (6). This system enabled the measurement of the frequency and distribution of recombination events at a genome wide level in wild type Arabidopsis (7), construction of genetic linkage maps which include positions for each centromere (8), and modeling of the strength and pattern of interference (9). This proposal extends the use of tetrad analysis in Arabidopsis by using it as the basis for assessing the phenotypes of mutants in genes important for recombination and the regulation of crossover interference and performing a novel genetic screen. In addition to broadening our knowledge of a classic genetic problem - the regulation of recombination by crossover interference - this proposal also provides broader impact by: generating pedagogical tools for use in hands-on classroom experience with genetics, building interdisciplinary collegial partnerships, and creating a platform for participation by junior scientists from underrepresented groups. There are three specific aims: (1) Isolate mutants in Arabidopsis MUS81 homologs using T-DNA and TILLING (2) Characterize recombination levels and interference in mus81 mutants (3) Execute a novel genetic screen, based on tetrad analysis, for genes that regulate meiotic recombination

Gregory p. Copenhaver

2011-11-09

224

Thrombin stimulation of synthesis and secretion of fibronectin by human A549 epithelial cells and mouse LB fibroblasts  

International Nuclear Information System (INIS)

Thrombin, a serine protease generated at wound sites, takes part in multiple biological functions, including wound healing. The present report elucidates the effect of thrombin on fibronectin (FN) synthesis and secretion in fibroblasts and epithelial cells. Subconfluent cultures of mouse LB fibroblasts and human A549 epithelial cells were exposed to various concentrations of bovine plasma thrombin at 37 degrees C for 16 hr. After exposure, cells were processed for determination of cell-associated and secreted FN by metabolic labeling, immunoprecipitation, immunofluorescence, and peroxidase immunocytochemistry. The correlation of FN production with cell growth was studied by a combined procedure of peroxidase immunocytochemistry and light microscopic autoradiography. The amounts of cell-associated and secreted FN were significantly increased with dose increments of thrombin. The increases were most evident in secreted FN. The increase of cell-associated FN was also evidenced by results from immunofluorescence and immunocytochemical studies. Ultrastructurally, the intracellular FN was localized in rough endoplasmic reticulum, Golgi complexes, and secretory granules, whereas non-released extracellular FN was localized in the plasma membrane of cell-to-cell contacts and in the extracellular fibrils. More intense cytoplasmic FN staining was observed in cells that were not labeled with [3H]-thymidine, indicating that FN production may vary with different phases of cell growth. The results imply that thrombin may play an important role in the early phases of wound healing

225

[US-guided percutaneous management of femoral artery pseudoaneurysms by thrombin injection: personal experience and review of the literature].  

Science.gov (United States)

Treatment of femoral artery pseudoaneurysms includes US-guided compression, endovascular coil embolisation or stenting, thrombin injection and open surgical repair. Thrombin injection is currently the standard approach to all non-complicated pseudoaneurysms of the peripheral arteries. Between January 2001 and December 2004, 59 pseudoaneurysms of the femoral artery were submitted to percutaneous US-guided thrombin injection in our surgery division. Complicated pseudoaneurysms, patients in whom percutaneous treatment failed, and chronic pseudoaneurysms with large necks were excluded from this kind of treatment and underwent surgical repair. Eighteen patients were on therapeutic anticoagulation (30.5%) and none of the pseudoaneurysms had an associated arteriovenous fistula. All patients were submitted to clinical and ultrasound follow-up at 24 hours and at 1, 6 and 12 months, and thereafter at yearly intervals for evaluation of recurrences. The thrombin injection was successful in 57/59 patients (96.6%) while two patients (3.4%) were submitted to surgical treatment after failure of the procedure. In 1 patient (1.7%) an additional injection was necessary to achieve complete thrombosis of the pseudoaneurysm. No complications occurred; none of the patients reported discomfort and there was no request for sedation. No recurrences were seen at follow-up. Nowadays US-guided percutaneous thrombin injection is the treatment of choice for femoral artery pseudoaneurysms. Surgical treatment is reserved to rare selected cases. More studies are necessary to evaluate further indications for treatment of visceral or small distal artery pseudoaneurysms. PMID:16400767

Veraldi, Gian Franco; Tasselli, Sebastiano; Firpo, Maria; Pedrazzani, Corrado; Ruzzenente, Andrea

2005-01-01

226

Magnetic relaxation switch and colorimetric detection of thrombin using aptamer-functionalized gold-coated iron oxide nanoparticles  

International Nuclear Information System (INIS)

We describe a sensitive biosensing system combining magnetic relaxation switch diagnosis and colorimetric detection of human ?-thrombin, based on the aptamer-protein interaction induced aggregation of Fe3O4-Au nanoparticles. To demonstrate the concept, gold-coated iron oxide nanoparticle was synthesized by iterative reduction of HAuCl4 onto the dextran-coated Fe3O4 nanoparticles. The resulting core-shell structure had a flowerlike shape with pretty narrow size distribution (referred to as 'nanorose'). The two aptamers corresponding to human ?-thrombin were conjugated separately to two distinct nanorose populations. Once a solution containing human ?-thrombin was introduced, the nanoroses switched from a well dispersed state to an aggregated one, leading to a change in the spin-spin relaxation time (T2) as well as the UV-Vis absorption spectra of the solution. Thus the qualitative and quantitative detection method for human ?-thrombin was established. The dual-mode detection is clearly advantageous in obtaining a more reliable result; the detection range is widened as well. By using the dual-mode detection method, a detectable T2 change is observed with 1.0 nM human ?-thrombin, and the detection range is from 1.6 nM to 30.4 nM.

227

Aptamer conjugated Mo(6)S(9-x)I(x) nanowires for direct and highly sensitive electrochemical sensing of thrombin.  

Science.gov (United States)

We demonstrate the use of a novel electrochemical sensing platform based on aptamer conjugated Mo(6)S(9-x)I(x) nanowires (MoSI NWs) for the highly sensitive detection of the blood clotting enzyme thrombin. MoSI NWs nanowires were self-assembled on a gold electrode to which thrombin binding aptamers were covalently attached. The modification and immobilization steps of the electrodes were characterised by cyclic voltammetry along with high-resolution transmission electron microscopy and X-ray photoelectron spectroscopy. The platform is based on the creation of a self-assembled MoSI MW layer via the sulfur-gold affinity followed by the creation of MoSI-thiolated aptamer conjugates via the sulfur-sulfur affinity. Using this system, sensitive quantitative detection of thrombin is realized by monitoring differences of differential pulse voltammetric responses of electrostatically trapped [Ru(NH(3))(6)](3+) cations to the aptamer before and after thrombin binding. The sensitivity limit for the detection of thrombin is 10 pM. This value is 10-fold better than all currently reported one step label free electrochemical strategies. Given the direct label free nature of the approach and the simplicity of the electronic detection, the aptamer conjugated MoSI NWs biosensor appears well suited for implementation in portable point of care microdevices directed at the rapid and sensitive detection of proteins and pathogens. PMID:20176468

McMullan, Martin; Sun, Nijuan; Papakonstantinou, Pagona; Li, Meixian; Zhou, Wuzong; Mihailovic, Dragan

2011-01-15

228

Isolation and properties of two forms of thrombin inhibitor from the nymphs of the camel tick Hyalomma dromedarii (Acari: Ixodidae).  

Science.gov (United States)

Two forms of the nymphal thrombin inhibitors (NTI) 3.2 kDa and 14.9 kDa were purified by chromatography on CM-cellulose. Sephacryl S-300 and Sephadex G-50 columns and designated NTI- 1 and NTI-2 respectively. The NTI-2 turned out to be homogenous monomeric protein in both native-PAGE and denatured SDS-PAGE with M(r) value of 14.9 kDa approximately and its pI value ranged from 7.2 to 7.5. The NTI-1 and NTI-2 displayed anticoagulant activity since they prolonged both the activated partial thromboplastin time (APTT) and the prothrombin time (PT) of the camel plasma in a concentration-dependent manner. The potency of NTI-I toward thrombin was 5-fold higher than that toward FXa, while NTI-2 was 3-fold active toward FXa than thrombin. However, both of them did not inhibit any of the other examined proteases. The types of inhibition of thrombin by NTI-1 and NTI-2 were non-competitive and competitive with inhibition constants (Ki) values of 11.7 microM and 211 nM respectively. One binding site was deduced on thrombin for each inhibitor. PMID:12171275

Ibrahim, M A; Ghazy, A H; Maharem, T; Khalil, M

2001-01-01

229

Key role of integrin ?(IIb)? (3) signaling to Syk kinase in tissue factor-induced thrombin generation.  

Science.gov (United States)

The fibrin(ogen) receptor, integrin ?(IIb)?(3), has a well-established role in platelet spreading, aggregation and clot retraction. How ?(IIb)?(3) contributes to platelet-dependent coagulation is less well resolved. Here, we demonstrate that the potent suppressing effect of clinically used ?(IIb)?(3) blockers on tissue factor-induced thrombin generation is linked to diminished platelet Ca(2+) responses and phosphatidylserine (PS) exposure. The same blockers suppress these responses in platelets stimulated with collagen and thrombin receptor agonists, whereas added fibrinogen potentiates these responses. In platelets spreading on fibrinogen, outside-in ?(IIb)?(3) signaling similarly enhances thrombin-induced Ca(2+) rises and PS exposure. These responses are reduced in ?(IIb)?(3)-deficient platelets from patients with Glanzmann's thrombasthenia. Furthermore, the contribution of ?(IIb)?(3) to tissue factor-induced platelet Ca(2+) rises, PS exposure and thrombin generation in plasma are fully dependent on Syk kinase activity. Tyrosine phosphorylation analysis confirms a key role of Syk activation, which is largely but not exclusively dependent on ?(IIb)?(3) activation. It is concluded that the majority of tissue factor-induced procoagulant activity of platelets relies on Syk activation and ensuing Ca(2+) signal generation, and furthermore that a considerable part of Syk activation relies on ?(IIb)?(3) signaling. These results hence point to a novel role of Syk in integrin-dependent thrombin generation. PMID:22669259

van der Meijden, Paola E J; Feijge, Marion A H; Swieringa, Frauke; Gilio, Karen; Nergiz-Unal, Reyhan; Hamulyák, Karly; Heemskerk, Johan W M

2012-10-01

230

Thrombin inhibits HMGB1-mediated proinflammatory signaling responses when endothelial protein C receptor is occupied by its natural ligand  

Directory of Open Access Journals (Sweden)

Full Text Available High mobility group box 1 (HMGB1 is involved in thepathogenesis of vascular diseases. Unlike activated protein C(APC, the activation of PAR-1 by thrombin is known to elicitproinflammatory responses. To determine whether the occupancyof EPCR by the Gla-domain of APC is responsible for thePAR-1-dependent antiinflammatory activity of the protease, wepretreated HUVECs with the PC zymogen and then activatedPAR-1 with thrombin. It was found that thrombin downregulatesthe HMGB1-mediated induction of both TNF-? andIL-6 and inhibits the activation of both p38 MAPK and NF-?B inHUVECs pretreated with PC. Furthermore, thrombin inhibitedHMGB1-mediated hyperpermeability and leukocyte adhesion/migration by inhibiting the expression of cell adhesion moleculesin HUVECs if EPCR was occupied. Collectively, theseresults suggest the concept that thrombin can initiate proinflammatoryresponses in vascular endothelial cells through theactivation of PAR-1 may not hold true for normal vesselsexpressing EPCR under in vivo conditions. [BMB Reports 2013;46(11: 544-549

Jong-Sup Bae

2013-11-01

231

Apoferritin Protein Nanoparticles Dually labeled with Aptamer and Horseradish Peroxidase as a Sensing Probe for Thrombin Detection  

Energy Technology Data Exchange (ETDEWEB)

A sandwich-type electrochemical aptasensor has been developed for the detection of thrombin, based on dual signal-amplification using HRP and apoferritin. Aptamer1 (Apt1) loaded on core/shell Fe3O4/Au magnetic nanoparticle (AuMNP) was used as recognition elements, and apoferritin dually labeled with Aptamer2 (Apt2) and HRP was used as a detection probe. Sandwich-type complex, Apt1/thrombin/Apt2–apoferritin NPs–HRP was formed by the affinity reactions between AuMNPs–Apt1, thrombin, and Apt2–apoferritin–HRP. The complex was anchored on a screen-printed carbon electrode (SPCE). Differential pulse voltammetry (DPV) was used to monitor the electrode response. The proposed aptasensor yielded a linear current response to thrombin concentrations over a broad range of 0.5 pM to 100 pM with a detection limit of 0.07 pM (S/N = 3). The detection signal was amplified by using apoferritin and HRP. This nanoparticle-based aptasensor offers a new method for rapid, sensitive, selective, and inexpensive quantification of thrombin, and offers a promising potential in biomarker detection and disease diagnosis. --------------------------------------------------------------------------------

Zhao, Jie; Liu, Meiling; Zhang, Youyu; Li, Haitao; Lin, Yuehe; Yao, Shouzhuo

2013-01-08

232

Topics in CP violation  

International Nuclear Information System (INIS)

Given the varied backgrounds of the members of this audience this talk will be a grab bag of topics related to the general theme of CP Violation. I do not have time to dwell in detail on any of them. First, for the astronomers and astrophysicists among you, I want to begin by reviewing the experimental status of evidence for CP violation in particle processes. There is only one system where this has been observed, and that is in the decays of neutral K mesons

233

Differential proteolytic activation of factor VIII-von Willebrand factor complex by thrombin  

International Nuclear Information System (INIS)

Blood coagulation factor VIII (fVIII) is a plasma protein that is decreased or absent in hemophilia A. It is isolated as a mixture of heterodimers that contain a variably sized heavy chain and a common light chain. Thrombin catalyzes the activation of fVIII in a reaction that is associated with cleavages in both types of chain. The authors isolated a serine protease from Bothrops jararacussu snake venom that catalyzes thrombin-like heavy-chain cleavage but not light-chain cleavage in porcine fVIII as judged by NaDodSO4/PAGE and N-terminal sequence analysis. Using a plasma-free assay of the ability of activated 125I-fVIII to function as a cofactor in the activation of factor X by factor IXa, they found that fVIII is activated by the venom enzyme. The venom enzyme-activated fVIII was isolated in stable form by cation-exchange HPLC. von Willebrand factor inhibited venom enzyme-activated fVIII but not thrombin-activated fVIII. These results suggest that the binding of fVIII to von Willebrand factor depends on the presence of an intact light chain and that activated fVIII must dissociate from von Willebrand factor to exert its cofactor effect. Thus, proteolytic activation of fVIII-von Willebrand factor complex appears to be differentially regulated by light-chain cleavage to dissociate the complex and heavy-chain cleavage to activate the cofactor function

234

Structure of saxthrombin, a thrombin-like enzyme from Gloydius saxatilis.  

Science.gov (United States)

Snake-venom thrombin-like enzymes (SVTLEs) are serine proteases that are widely distributed in snakes from the Crotalinae subfamily of the Viperidae. In contrast to other snake-venom serine proteases, they have a biochemical activity similar to that of thrombin and play an important role in the process of blood coagulation. However, SVTLEs cannot activate factor VIII, which is essential in blood-clot stabilization. Consequently, blood clots produced by SVTLEs are not stable and are cleared rapidly. This characteristic makes SVTLEs attractive as potential candidates for antithrombotic therapy. Saxthrombin, an SVTLE from Gloydius saxatilis, was purified and crystallized to obtain a high-quality crystal, from which data were acquired to 1.43?Å resolution. Preliminary X-ray diffraction analysis showed that the crystal belonged to space group C2, with unit-cell parameters a = 94.2, b = 52.2, c = 50.1?Å, ? = 96.7°. The crystal structure was determined by molecular replacement and the final R factor was 18.69%; the R(free) was 20.01%. This is the first report of a crystal structure of an SVTLE. Saxthrombin belongs to the typical ?/?-hydrolase fold of serine proteases. Its structure was compared with those of thrombin and other snake-venom serine proteases. The observed differences in the amino-acid composition of the loops surrounding the active site appear to contribute to different surface-charge distributions and thus alter the shape of the active-site cleft, which may explain the differences in substrate affinity. PMID:21821882

Huang, Kai; Zhao, Wei; Gao, Yongxiang; Wei, Wenqing; Teng, Maikun; Niu, Liwen

2011-08-01

235

BjussuSP-I: a new thrombin-like enzyme isolated from Bothrops jararacussu snake venom.  

Science.gov (United States)

A thrombin-like enzyme named BjussuSP-I, isolated from B. jararacussu snake venom, is an acidic single chain glycoprotein with approximately 6% sugar, Mr=61,000 under reducing conditions and pI approximately 3.8, representing 1.09% of the chromatographic A(280) recovery. BjussuSP-I is a glycosylated serine protease containing both N-linked carbohydrates and sialic acid in its structure. BjussuSP-I showed a high clotting activity upon human plasma, which was inhibited by PMSF, leupeptin, heparin and 1,10-phenantroline. This enzyme showed high stability regarding coagulant activity when analyzed at different temperatures (-70 to 37 degrees C), pHs (4.5 to 8.0), and presence of two divalent metal ions (Ca(2+) and Mg(2+)). It also displayed TAME esterase and proteolytic activities toward natural (fibrinogen and fibrin) and synthetic (BAPNA) substrates, respectively, being also inhibited by PMSF and leupeptin. BjussuSP-I can induce production of polyclonal antibodies able to inhibit its clotting activity, but unable to inhibit its proteolytic activity on fibrinogen. The enzyme also showed crossed immunoreactivity against 11 venom samples of Bothrops, 1 of Crotalus, and 1 of Calloselasma snakes, in addition of LAAO isolated from B. moojeni venom. It displayed neither hemorrhagic, myotoxic, edema-inducing profiles nor proteolytic activity on casein. BjussuSP-I showed an N-terminal sequence (VLGGDECDINEHPFLA FLYS) similar to other thrombin-like enzymes from snake venoms. Based on its biochemical, enzymatic and pharmacological characteristics, BjussuSP-I was identified as a new thrombin-like enzyme isoform from Bothrops jararacussu snake venom. PMID:17466550

Sant' Ana, Carolina D; Ticli, Fabio K; Oliveira, Leandro L; Giglio, Jose R; Rechia, Carem G V; Fuly, André L; Selistre de Araújo, Heloisa S; Franco, João J; Stabeli, Rodrigo G; Soares, Andreimar M; Sampaio, Suely V

2008-11-01

236

Thrombin related peptide TP508 promoted fracture repair in a mouse high energy fracture model  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Thrombin related peptide (TP508 is a 23 amino-acid synthetic peptide that represents a portion of the receptor-binding domain of thrombin molecule. Previous studies have shown that TP508 can accelerate musculoskeletal tissue repair including fracture healing. Objectives The aim of this study was to investigate the effect of TP508 on fracture healing in a murine fracture model representing high energy fracture situation. Methods Eighty CD 1 mice underwent controlled quadriceps muscle crush and open transverse mid diaphyseal femoral fracture that was then fixed with an external fixator. Animals were randomised into four groups to receive an intra-operative dose of either 100 ?g TP508 into the fracture gap; 100 ?g TP508 into the surrounding damaged muscle tissues; 10 ?g TP508 into the fracture gap, or control equal amount of saline into the fracture gap. Radiographic assessment was performed weekly for 5 weeks; histological analysis was at 3 and 5 weeks post fracture and biomechanical testing of the fractured bone was performed at 5 weeks post fracture. Results Mechanical testing data showed that the fracture stiffness was significantly higher in the group receiving 100 ?g TP508 into the fracture gap than other groups. Histological and radiographic analysis revealed a trend of increase in bone formation in the 100 ?g TP508 injected into the fracture gap group compared to the saline control group. It was noted that the scar tissues was significantly less in Group II comparing with the saline control group and there was increased blood vessel formation in the crushed muscles and fracture gap areas in the groups receiving TP508 comparing to the saline control group. Conclusion The results from this study demonstrated the use of thrombin related peptide TP508 in the situation of a high energy fracture can promote fracture healing and reduce the potential complications such as muscle fibrosis and fracture delayed or non-union.

Pan Xiao-Hua

2009-01-01

237

Use of a Thrombin-gelatin Hemostatic Matrix (Surgiflo) in Spinal Surgery.  

Science.gov (United States)

A variety of techniques have been used to stop venous bleeding from the spinal epidural space. These generally consist of packing with Surgicel®, fibrillar collagen or Gelfoam®. Bipolar coagulation may also be used to control bleeding from spinal venous plexus, but it may bear the risk of healthy nervous tissue injury: dissipation of heat from the tips of the bipolar forceps may induce thermal injury to adjacent neural structures. In the case of intraspinal bleeding, quick and safe hemostasis is mandatory to ensure adequate visualization and safe preparation so as to avoid damaging nerves and spinal medulla. In addition, quick and safe hemostasis reduces the duration of surgery. Efficient control of bleeding can thereby reduce perioperative morbidity. During 6 months, the authors performed more than 170 major spinal surgeries, and in 67 procedures they used injection of thrombin-gelatin hemostatic matrix (Surgiflo, Johnson & Johnson Wound Management, Somerville, NJ) into spinal epidural space to assist in hemostasis. When the venous bleeding continued from the epidural space after packing with hemostatic agents as Surgicel and fibrillar collagen, gelatin matrix was used to stop venous bleeding. In all cases, the results were judged to be excellent, with immediate stoppage of epidural bleeding, or good. No complications related to the thrombin-gelatin hemostatic matrix were encountered. The thrombin-gelatin matrix could represent a valuable tool when other hemostatic strategies are ineffective or suboptimal. It is safe and biocompatible when compared with hemostatic agents currently in use. This is the first study reporting the use of Surgiflo hemostatic matrix in spinal surgery. PMID:25419955

Gazzeri, Roberto; De Bonis, Costanzo; Galarza, Marcelo

2014-11-01

238

Topic and Topic-Comment Constructions in Mandarin Chinese.  

Science.gov (United States)

Attempts to provide a precise definition for topic and to derive most of the properties of topic from this definition. The main assumption is that the topic-comment construction is a syntactic device employed to fulfill certain discourse functions. (Author/VWL)

Shi, Dingxu

2000-01-01

239

Improving the thrombin inhibitory activity of glycyrrhizin, a triterpenic saponin, through a molecular simplification of the carbohydrate moiety.  

Science.gov (United States)

Glycyrrhizin, a saponin, and its aglycone glycyrrhetinic acid are natural products found in the Liquorice (Glycyrrhiza glabra L.) root extract. This saponin is known for its in vitro and in vivo thrombin inhibitory activity. The design and synthesis of five glycyrrhizin derivatives were carried out to improve the natural product activity. Compound 3b, a phthalic ester derivative of glycyrrhizin, presented a more pronounced thrombin inhibition (IC50  = 114.4 ± 1.3 ?m) than the saponin (IC50  = 235.7 ± 1.4 ?m). Molecular docking simulations performed to investigate the molecular interaction between compound 3b and the enzyme indicate that this product is, as previously determined for glycyrrhizin, an allosteric thrombin inhibitor. PMID:23964664

de Paula, Fernando T; Frauches, Petrina Q; Pedebos, Conrado; Berger, Markus; Gnoatto, Simone C B; Gossmann, Grace; Verli, Hugo; Guimarães, Jorge A; Graebin, Cedric S

2013-12-01

240

A proposal for dose-adjustment of dabigatran etexilate in atrial fibrillation guided by thrombin time.  

Science.gov (United States)

Dabigatran is an oral anticoagulant that is increasingly used for atrial fibrillation (AF). Presently, many authorities state that routine laboratory coagulation monitoring is not required. However, data have recently been published demonstrating that higher trough plasma dabigatran concentrations are associated with lower thromboembolic and higher haemorrhagic event rates. Using these data, we simulate a range of AF patients with varying risks for these events and derive a target range of trough plasma dabigatran concentrations (30-130??g?l(-1) ). Finally, we propose that a conventional screening coagulation assay, the thrombin time (TT), can be used to discern whether or not patients are within this range of dabigatran concentrations. PMID:24592851

Chin, Paul K L; Wright, Daniel F B; Patterson, David M; Doogue, Matthew P; Begg, Evan J

2014-09-01

 
 
 
 
241

[Specific inhibition of thrombin activity during cardiopulmonary bypass reduces ischemia-reperfusion injury of the lung].  

Science.gov (United States)

The pathophysiologic role of thrombin in the development of lung injury after the normothermic cardiopulumonary bypass (CPB) was studied in the rabbit model. A control group (group D) was subjected to the pericardiotomy without institution of CPB. Group A rabbits (n = 6) underwent left heart bypass (80 ml/kg/min) for 60 minutes without occlusion of the systemic or pulmonary artery and a succeeding reduced flow (20-30 ml/kg/min) for another 30 minutes, group B rabbits (n = 6) underwent complete CPB (80 ml/kg/min) for 60 minutes in the working mode with occlusion of the pulmonary arterial trunk and a succeeding reduced flow without occlusion of the pulmonary artery for another 30 minutes, group C rabbits (n = 6) underwent the same CPB technique as group B in conjunction with continuous intravenous infusion of argatroban (60 micrograms/kg/min), the specific thrombin inhibitor. In this group, infusion of argatroban was initiated 60 minutes prior to institution of CPB and terminated at the end of the experiment. We sacrificed rabbits four hours after the experiment began, and assessed not only morphometrically thrombus formation, leukocytic infiltration and luminal narrowing of small-sized pulmonary arteries but also immunohistochemically the expression of tissue factor (TF) and IL-1 beta, and physico-functionally respiratory index (RI) and pulmonary vascular resistance (PVR). Rabbits in group A showed multiple occurrence of lung thrombi, luminal narrowing of small arteries, and mild infiltration of macrophages and neutrophils positive for TF, and, in addition, their RI and PVR became mildly worse. In group B, all these morphological and physico-functional parameters became much worse than those observed in group A rabbits (p argatroban treatment could significantly improve these parameters (p < .01). The expression of TF and IL-1 beta, however, was not significantly different in group A, B and C. These findings indicate that thrombin function intimately participates in the development of pulmonary ischemia-reperfusion injury during CPB. In addition, the anti-thrombin treatment would be an effective therapeutical tool for the prevention of not only activation of extrinsic coagulation pathway but also its sequential inflammatory and circulatory disturbance in ischemia-reperfusion injury of lung during CPB. PMID:11246985

Tanaka, K

2001-01-01

242

Effects of tissue factor, thrombomodulin and elevated clotting factor levels on thrombin generation in the calibrated automated thrombogram.  

Science.gov (United States)

Elevated procoagulant levels have been correlated with increased thrombin generation in vitro and with increased venous thromboembolism (VTE) risk in epidemiological studies. Thrombin generation tests are increasingly being employed as a high throughput method to provide a global measure of procoagulant activity in plasma samples. The objective of this study was to distinguish the effects of assay conditions [tissue factor (TF), thrombomodulin, platelets/lipids] and factor levels on thrombin generation parameters, and determine the conditions and parameters with the highest sensitivity and specificity for detecting elevated factor levels. Thrombin generation was measured using calibrated automated thrombography (CAT) in corn trypsin inhibitor (CTI)-treated platelet-free plasma (PFP) and platelet-rich plasma (PRP). Statistical analysis was performed using logarithms of observed values with analysis of variance that accounted for experiment and treatment. The relative sensitivity of lag time (LT), time to peak (TTP), peak height and endogenous thrombin potential (ETP) to elevated factors XI, IX, VIII, X, and prothrombin was as follows: PFP initiated with 1 pM TF > PFP initiated with 5 pM TF > PRP initiated with 1 pM TF. For all conditions, inclusion of thrombomodulin prolonged the LT and decreased the peak and ETP; however, addition of thrombomodulin did not increase the ability of CAT to detect elevated levels of individual procoagulant factors. In conclusion, CAT conditions differentially affected the sensitivity of thrombin generation to elevated factor levels. Monitoring the peak height and/or ETP following initiation of clotting in PFP with 1 pM TF was most likely to detect hypercoagulability due to increased procoagulant factor levels. PMID:19888532

Machlus, Kellie R; Colby, Emily A; Wu, Jogin R; Koch, Gary G; Key, Nigel S; Wolberg, Alisa S

2009-11-01

243

MicroRNA-146a Decreases High Glucose/Thrombin-Induced Endothelial Inflammation by Inhibiting NAPDH Oxidase 4 Expression  

Science.gov (United States)

Diabetes is associated with hyperglycemia and increased thrombin production. However, it is unknown whether a combination of high glucose and thrombin can modulate the expression of NAPDH oxidase (Nox) subtypes in human aortic endothelial cells (HAECs). Moreover, we investigated the role of a diabetes-associated microRNA (miR-146a) in a diabetic atherothrombosis model. We showed that high glucose (HG) exerted a synergistic effect with thrombin to induce a 10.69-fold increase in Nox4 mRNA level in HAECs. Increased Nox4 mRNA expression was associated with increased Nox4 protein expression and ROS production. Inflammatory cytokine kit identified that the treatment increased IL-8 and IL-6 levels. Moreover, HG/thrombin treatment caused an 11.43-fold increase of THP-1 adhesion to HAECs. In silico analysis identified the homology between miR-146a and the 3?-untranslated region of the Nox4 mRNA, and a luciferase reporter assay confirmed that the miR-146a mimic bound to this Nox4 regulatory region. Additionally, miR-146a expression was decreased to 58% of that in the control, indicating impaired feedback restraint of HG/thrombin-induced endothelial inflammation. In contrast, miR-146a mimic transfection attenuated HG/thrombin-induced upregulation of Nox4 expression, ROS generation, and inflammatory phenotypes. In conclusion, miR-146a is involved in the regulation of endothelial inflammation via modulation of Nox4 expression in a diabetic atherothrombosis model. PMID:25298619

Wang, Huang-Joe; Huang, Yuan-Li; Shih, Ya-Yun; Wu, Hsing-Yu; Peng, Ching-Tien; Lo, Wan-Yu

2014-01-01

244

Topical tretinoin in acanthosis nigricans  

Directory of Open Access Journals (Sweden)

Full Text Available Efficacy of topical tretinoin was assessed in 30 cases of idiopathic acanthosis nigricans which were recalcitrant to conventional modalities of treatment. Topical tretinoin once at night application was found to be very effective both clinically and histologically.

Lahiri Koushik

1996-01-01

245

Health Topics: MedlinePlus  

Science.gov (United States)

... features on this page, please enable JavaScript. Health Topics Read about symptoms, causes, treatment and prevention for ... illnesses, health conditions and wellness issues. MedlinePlus health topics are regularly reviewed, and links are updated daily. ...

246

Amino acids 225-235** of the protein C serine-protease domain are important for the interaction with the thrombin-thrombomodulin complex.  

Science.gov (United States)

Protein C (PC) is a vitamin K-dependent zymogen that inactivates factors Va and VIIIa after its activation by thrombin complexed to thrombomodulin. We characterized a monoclonal antibody (mAb) against PC, whose only influence on PC functions was to inhibit PC activation by the thrombin-thrombomodulin complex. It recognized an epitope in the PC heavy chain, the conformation of which is calcium-dependent. The mAb did not recognize a natural PC variant that was not activated by the thrombin-thrombomodulin complex (mutation R229Q) and did recognize a synthetic peptide corresponding to PC amino acids 225-235 in an Elisa assay. The peptide inhibited PC activation by the thrombin-thrombomodulin complex. These data confirm that the calcium-binding loop of the serine-protease domain is involved in the interaction of PC with the thrombin-thrombomodulin complex. PMID:7540990

Vincenot, A; Gaussem, P; Pittet, J L; Debost, S; Aiach, M

1995-06-26

247

Recent Developments in Dissociative Recombination  

International Nuclear Information System (INIS)

There have been a number of recent developments in dissociative recombination research as it relates to ITER, that should be highlighted. These concern primarily experimental and modelling issues and this document will not touch upon the topics of the other scientists involved in DR studies that are present at the meeting. The topic of branching ratios in general is a topic fundamental to DR especially how it influences the formation of radical and stable neutral molecules that again might play a role in particle formation. It should be remembered that the reactions of neutral radicals to form cyclic compounds are responsible for the formation of soot in combustion, though the role played by ions in flames is at best uncertain. In the near wall plasma environment, ion processes may well be more important since neutral species are rarer. Modelling studies by Pernot and collaborators at the Universite de Paris-Sud have shown that if one compares the yields of individual neutral species in ion-chemistry models (in this particular case, the ionosphere of Titan), and if one assumes that DR reactions of hydrocarbon ions primarily decay via the ejection of a hydrogen ion (which is assumed by most Titan ionospheric models) and if one compares these predictions with those coming from a model where actual measured branching ratios are used, differences of up to 5 orders of magnitude are found. This shows very clearly the need for branching ratio studies. In early merged beam studies of DR performed in Canada in the 1970's, it was noticed that cross sections for polyatomic species typically displayed a sharp fall-off above 0.1 eV. This has since been seen in many storage ring studies and clearly this has important consequences for ITER chemistry where plasma temperatures are likely to be well above ambient. In a recent analysis, Jungen and Pratt have explained this phenomenon on the basis that the recombination is dominated by the indirect process (initial capture into a vibrationally excited, neutral Rydberg state) in which the propensity rule (+?v=1) dominates the capture. When the electron energy exceeds that between the v'=0 and v'=1 levels of the ion, where the capture must now involve a ?v=2 transition, this will be much less effective and so the cross section drops precipitously. This assumes of course that the recombining ion is primarily in the ground v=0 level. H3+ continues to be an active subject of research and a very recent experiment at the TSR ring in Heidelberg has examined the influence of rotational excitation on the rate of the recombination. This is a very beautiful study but an important outcome is that even though a cryogenically cooled storage trap was used to produce the ions, the internal rotational temperature of the ions was never found to be below 150K. This suggests that ion cooling by storage in the ring leads eventually to an equilibrium value for the internal energy of the ions as they are de-excited/re-excited by passage through the electron cooler. As observed in earlier merged beam experiments in Canada, the extraction field in the ion source plays an important role in determining the excitation state of the ions as collisions outside the source can lead to re-heating. Indeed in the TSR experiments using a conventional Penning source and a normal extraction field, the ions were found to have a rotational temperature of several thousands of degrees. This clearly has important significance for earlier measurements taken in storage rings. Finally, the world will soon have a new storage ring facility for dissociative recombination research and this will be in Langzhou in China. This machine will have a higher magnetic rigidity that previous rings used for DR and so heavier ions and higher mass resolution experiments can be performed there. Experimental operation of this new ring is expected to commence in 2012/2013. (author)

248

The Use of Direct Thrombin Injection to Treat a Type II Endoleak Following Endovascular Repair of Abdominal Aortic Aneurysm  

International Nuclear Information System (INIS)

This report describes the use of thrombin to treat a type II endoleak which was causing continued abdominal aortic aneurysm expansion in a patient who had undergone endovascular repair. A small quantity of thrombin was injected into the leak by a percutaneous approach directly into the aneurysm sac using color doppler ultrasound. The procedure was successful and required only a few minutes to perform. We believe this procedure is an alternative to some of the more complex and technically challenging means of treating this lesion

249

The dissociative recombination of ?  

Science.gov (United States)

The dissociative recombination of 0953-4075/31/8/021/img2 has been measured using a single-pass merged-beam apparatus. It is found that the cross section displays resonant structure and falls-off abruptly above 0.3 eV.

LePadellec, A.; Sheehan, C.; Mitchell, J. B. A.

1998-04-01

250

Recombinant DNA for Teachers.  

Science.gov (United States)

A science teacher describes his experience at a workshop to learn to teach the Cold Spring Harbor DNA Science Laboratory Protocols. These protocols lead students through processes for taking E. coli cells and transforming them into a new antibiotic resistant strain. The workshop featured discussions of the role of DNA recombinant technology in…

Duvall, James G., III

1992-01-01

251

Recombinant hormones in osteoporosis  

DEFF Research Database (Denmark)

For the last 10 years, bone anabolic therapy with the recombinant human parathyroid hormone (rhPTH) analogue, teriparatide (rhPTH[1 - 34]), or full-length rhPTH(1 - 84) has been an option in the treatment of osteoporosis. Both drugs are given as a daily subcutaneous injection. In the USA, only teriparatide is marketed.

Rejnmark, Lars; Rejnmark, Lars

2013-01-01

252

Testing for recombinant human erythropoietin  

Science.gov (United States)

ERYTHROPOIETIN (Epo) may have effects on exercise capacity and physiological regulation beyond a simple increase in red cell mass and the associated improvement in oxygen transport (4). In the context of a larger study on this topic, Lundby and colleagues (11) also asked questions about the reliability of urine testing for recombinant human Epo (rHuEpo). They studied eight healthy male subjects during a 4-wk "loading" and 2-wk "boosting" phase of Epo use followed by a 2-wk maintenance phase. In the parent study they showed that the effects of Epo on exercise performance were confined to its impact on red cell mass and not to other physiological effects of the hormone. These results were consistent with ideas about the relationship between maximal oxygen uptake and red cell mass or total body hemoglobin that emerged in the 1950s. The findings are timely and have implications for public policy relating to the control of doping practices. In this short report a number of challenges related to urine testing for Epo are highlighted.

Joris R Delanghe (Ghent University Hospital Clinical Chemistry); Michael J Joyner (Mayo Clinic Anesthesiology)

2008-08-10

253

Role of protein kinase Czeta in thrombin-induced RhoA activation and inter-endothelial gap formation of human dermal microvessel endothelial cell monolayers.  

Science.gov (United States)

We studied the potential involvement of the Ca(2+)-independent atypical protein kinase C isoform PKCzeta in mediating the thrombin-induced increase in endothelial permeability. Studies were done using human dermal microvessel endothelial cells (HMEC), which we showed constitutively expressed PKCzeta. We quantified the patency of inter-endothelial junctions (IEJs) and endothelial barrier function by measuring transendothelial electrical resistance (TER) in confluent HMEC monolayers. In control monolayers, thrombin decreased TER by approximately 50%, indicating thrombin-dependent opening of IEJs. Thrombin also elicited increases in cytosolic Ca(2+) concentration [Ca(2+)](i), actin stress fiber formation, and myosin light chain (MLC) phosphorylation. Pan-PKC inhibitors, calphostin C and chelerythrine, abrogated these responses. Thrombin also decreased TER after depletion of conventional and novel Ca(2+)-dependent PKC isoforms using phorbol 12-myristate 13-acetate (PMA). In these PMA-treated cells, thrombin induced inter-endothelial gap formation, MLC phosphorylation, and actin stress fiber formation, but failed to increase [Ca(2+)](i). Inhibition of PKCzeta activation using the PKCzeta pseudosubstrate peptide (PSI), depletion of PKCzeta protein with siRNA, and competitive inhibition of PKCzeta activity using dominant-negative (dn) PKCzeta mutant all prevented the thrombin-induced decrease in TER and MLC phosphorylation. Expression of dn-PKCzeta also inhibited thrombin-induced RhoA activation. These findings reveal a novel Ca(2+)-independent, PKCzeta-dependent mechanism of thrombin-induced increase in endothelial permeability. The results raise the possibility that inhibition of PKCzeta may be a novel drug target for thrombin-induced inflammatory hyperpermeability. PMID:20417648

Minshall, Richard D; Vandenbroucke, Emily E; Holinstat, Michael; Place, Aaron T; Tiruppathi, Chinnaswamy; Vogel, Stephen M; van Nieuw Amerongen, Geerten P; Mehta, Dolly; Malik, Asrar B

2010-09-01

254

Topics in broken supersymmetry  

International Nuclear Information System (INIS)

Studies on two topics in the framework of broken supersymmetry are presented. Chapter I is a brief introduction in which the motivation and the background of this work are discussed. In Chapter II, the author studies the decay K+ ? ?+ ?? in models with spontaneous supersymmetry breaking and find that it is generally suppressed relative to the decay K+ ? ?+ anti nu nu of the conventional model, except possibly for a class of models where the scalar quark masses are generated by radiative corrections from a much larger supersymmetry breaking scale. For a small range of scalar quark and photino mass parameters, the cascade decay process K+ ? ?+ ?0 ? ?+ ?? will become dominant over the anti nu nu mode. The author also comments on the possibility of probing the neutrino mass through the K+ ? ?+ ?0 ? ?+ anti nu nu cascade decay. Chapter III is concerned with the implications of explicit lepton number violating soft operators in a general low energy effective theory with softly broken supersymmetry

255

Superconcentration and related topics  

CERN Document Server

A certain curious feature of random objects, introduced by the author as “super concentration,” and two related topics, “chaos” and “multiple valleys,” are highlighted in this book. Although super concentration has established itself as a recognized feature in a number of areas of probability theory in the last twenty years (under a variety of names), the author was the first to discover and explore its connections with chaos and multiple valleys. He achieves a substantial degree of simplification and clarity in the presentation of these findings by using the spectral approach. Understanding the fluctuations of random objects is one of the major goals of probability theory and a whole subfield of probability and analysis, called concentration of measure, is devoted to understanding these fluctuations. This subfield offers a range of tools for computing upper bounds on the orders of fluctuations of very complicated random variables. Usually, concentration of measure is useful when more direct prob...

Chatterjee, Sourav

2014-01-01

256

Topics on String Phenomenology  

CERN Document Server

These lectures present some topics of string phenomenology and contain two parts. In the first part, I review the possibility of lowering the string scale in the TeV region, that provides a theoretical framework for solving the mass hierarchy problem and unifying all interactions. The apparent weakness of gravity can then be accounted by the existence of large internal dimensions, in the submillimeter region, and transverse to a braneworld where our universe must be confined. I review the main properties of this scenario and its implications for observations at both particle colliders, and in non-accelerator gravity experiments. In the second part, I discuss a simple framework of toroidal string models with magnetized branes, that offers an interesting self-consistent setup for string phenomenology. I will present an algorithm for fixing the geometric parameters of the compactification, build calculable particle physics models such as a supersymmetric SU(5) Grand Unified Theory with three generations of quark...

Antoniadis, Ignatios

2008-01-01

257

Perspective and research topics  

International Nuclear Information System (INIS)

In the preceding chapters various aspects of the theory of turbulent reacting flows are discussed in detail. The fundamental features of such flows, some of the relevant practical problems connected with turbulent combustion, approaches appropriate for the limiting cases of nonpremixed and premixed reactants, and the direct probability-density approach to the description of such flows are treated in successive chapters. There results a relatively complete view of the present status of the fundamental theory and its application to somewhat idealized flows. In this chapter we attempt to provide an overview and perspective of the field and call attention to research topics of greatest interest from both practical and fundamental points of view. (orig.)

258

Computational study of some benzamidine-based inhibitors of thrombin-like snake venom proteinases  

Science.gov (United States)

Pit viper venoms contain a number of serine proteinases that, despite their observed coagulant thrombin-like action in vitro, exhibit a paradoxical benign defibrinogenating (anticoagulant) action in vivo, with clinical applications in preventing thrombi and improved blood circulation. Considering that several benzamidine-based inhibitors, some highly selective to thrombin, also inhibit the enzymatic activity of such venombins, the modeling of their enzyme-inhibitor interactions could provide valuable information on the topological factors that determine the divergences in activity. The first step, and the object of the present study, was to derive the necessary set of parameters, consistent with the CHARMM force field, and to perform molecular dynamics (MD) simulations on a few selected representatives of the inhibitors in question under physiological conditions. Bonding and van der Waals parameters were derived by analogy to similar ones in the existing force field. Net atomic charges were obtained with a restrained fitting to the molecular electrostatic potential generated at B3LYP/6-31G(d) level. The parameters were refined to reproduce the available experimental geometries and crystal data, and the MD simulations of the free inhibitors in aqueous solution at 298 K provided an insightful description of their available conformational space.

Henriques, Elsa S.; Nascimento, Marco A. C.; Ramos, Maria João

259

A systems approach to hemostasis: 3. Thrombus consolidation regulates intrathrombus solute transport and local thrombin activity.  

Science.gov (United States)

Hemostatic thrombi formed after a penetrating injury have a distinctive structure in which a core of highly activated, closely packed platelets is covered by a shell of less-activated, loosely packed platelets. We have shown that differences in intrathrombus molecular transport emerge in parallel with regional differences in platelet packing density and predicted that these differences affect thrombus growth and stability. Here we test that prediction in a mouse vascular injury model. The studies use a novel method for measuring thrombus contraction in vivo and a previously characterized mouse line with a defect in integrin ?IIb?3 outside-in signaling that affects clot retraction ex vivo. The results show that the mutant mice have a defect in thrombus consolidation following vascular injury, resulting in an increase in intrathrombus transport rates and, as predicted by computational modeling, a decrease in thrombin activity and platelet activation in the thrombus core. Collectively, these data (1) demonstrate that in addition to the activation state of individual platelets, the physical properties of the accumulated mass of adherent platelets is critical in determining intrathrombus agonist distribution and platelet activation and (2) define a novel role for integrin signaling in the regulation of intrathrombus transport rates and localization of thrombin activity. PMID:24951426

Stalker, Timothy J; Welsh, John D; Tomaiuolo, Maurizio; Wu, Jie; Colace, Thomas V; Diamond, Scott L; Brass, Lawrence F

2014-09-11

260

Aptamer-based electrochemical approach to the detection of thrombin by modification of gold nanoparticles.  

Science.gov (United States)

This paper presents a simple electrochemical approach for the detection of thrombin, using aptamer-modified electrodes. The use of gold nanoparticles results in significant signal enhancement for subsequent detection. 1,6-Hexanedithiol was used as the medium to link Au nanoparticles to a bare gold electrode. Anti-thrombin aptamers were immobilized on the gold nanoparticles' surfaces by self-assembly. The packing density of aptamers was determined by cyclic voltammetric (CV) studies of redox cations (e.g., [Ru(NH(3))(6)](3+)) which were electrostatically bound to the DNA phosphate backbones. The results indicate that the total amount of aptamer probes immobilized on the gold nanoparticle surface is sixfold higher than that on the bare electrode, leading to increased sensitivity of the aptasensor and a detection limit of 1 pmol L(-1). Based on the Langmuir model, the sensor signal displayed an almost perfect linear relationship over the range of 1 pmol L(-1) to 30 nmol L(-1). Moreover, the proposed aptasensor is highly selective and stable. In summary, this biosensor is simple, highly sensitive, and selective, which is beneficial to the ever-growing interest in fabricating portable bio-analytical devices with simple electrical readout procedures. PMID:20607523

Li, Lidong; Zhao, Hongtao; Chen, Zhengbo; Mu, Xiaojiao; Guo, Lin

2010-09-01

 
 
 
 
261

Tissue factor expression in newt iris coincides with thrombin activation and lens regeneration.  

Science.gov (United States)

Lens regeneration in adult salamanders occurs at the pupillary margin of the mid-dorsal iris where pigmented epithelial cells (PEC) re-enter the cell cycle and transdifferentiate into lens. It is not understood how the injury caused by removal of the lens (lentectomy) in one location is linked to initiating the response in a different spatial location (dorsal iris) and to this particular sector. We propose that the blood provides a link between the localised coagulation and signal transduction pathways that lead to regeneration. A transmembrane protein (tissue factor) is expressed in a striking patch-like domain in the dorsal iris of the newt that localises coagulation specifically to this location, but is not expressed in the axolotl, a related species that does not show thrombin activation after lentectomy and cannot regenerate its lens. Our hypothesis is that tissue factor expression localises the initiation of regeneration through the activation of thrombin and the recruitment of blood cells, leading to local growth factor release. This is the first example of gene expression in a patch of cells that prefigures the location of a regenerative response, and links the immune system with the initiation of a regenerative program. PMID:20420902

Godwin, James W; Liem, Karel F; Brockes, Jeremy P

2010-01-01

262

Dual nanoparticle amplified surface plasmon resonance detection of thrombin at subattomolar concentrations.  

Science.gov (United States)

A novel dual nanoparticle amplification approach is introduced for the enhanced surface plasmon resonance (SPR) detection of a target protein at subattomolar concentrations. Thrombin was used as a model target protein as part of a sandwich assay involving an antithrombin (anti-Th) modified SPR chip surface and a thrombin specific DNA aptamer (Th-aptamer) whose sequence also includes a polyadenine (A30) tail. Dual nanoparticle (NP) enhancement was achieved with the controlled hybridization adsorption of first polythymine-NP conjugates (T20-NPs) followed by polyadenine-NPs (A30-NPs). Two different nanoparticle shapes (nanorod and quasi-spherical) were explored resulting in four different NP pair combinations being directly compared. It was found that both the order and NP shape were important in optimizing the assay performance. The use of real-time SPR measurements to detect target concentrations as low as 0.1 aM is a 10-fold improvement compared to single NP-enhanced SPR detection methods. PMID:25186782

Baek, Seung Hee; Wark, Alastair W; Lee, Hye Jin

2014-10-01

263

Evidence of recombination within human alpha-papillomavirus  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Human papillomavirus (HPV has a causal role in cervical cancer with almost half a million new cases occurring each year. Presence of the carcinogenic HPV is necessary for the development of the invasive carcinoma of the genital tract. Therefore, persistent infection with carcinogenic HPV causes virtually all cervical cancers. Some aspects of the molecular evolution of this virus, as the putative importance of recombination in its evolutionary history, are an opened current question. In addition, recombination could also be a significant issue nowadays since the frequency of co-infection with more than one HPV type is not a rare event and, thus, new recombinant types could be currently being generated. Results We have used human alpha-PV sequences from the public database at Los Alamos National Laboratory to report evidence that recombination may exist in this virus. A model-based population genetic approach was used to infer the recombination signal from the HPV DNA sequences grouped attending to phylogenetic and epidemiological information, as well as to clinical manifestations. Our results agree with recently published ones that use a different methodology to detect recombination associated to the gene L2. In addition, we have detected significant recombination signal in the genes E6, E7, L2 and L1 at different groups, and importantly within the high-risk type HPV16. The method used has recently been shown to be one of the most powerful and reliable procedures to detect the recombination signal. Conclusion We provide new support to the recent evidence of recombination in HPV. Additionally, we performed the recombination estimation assuming the best-fit model of nucleotide substitution and rate variation among sites, of the HPV DNA sequence sets. We found that the gene with recombination in most of the groups is L2 but the highest values were detected in L1 and E6. Gene E7 was recombinant only within the HPV16 type. The topic deserves further study because recombination is an important evolutionary mechanism that could have high impact both in pharmacogenomics (i.e. on the influence of genetic variation on the response to drugs and for vaccine development.

Carvajal-Rodríguez Antonio

2007-03-01

264

Oxygen-hydrogen recombiner  

International Nuclear Information System (INIS)

Purpose: To reduce the amount of a catalyst and improve the hydrogen removing performance of a catalyst type oxygen-hydrogen recombiner in a radioactive gaseous wastes processing facility of BWR type nuclear power plants. Constitution: The catalyst (of palladium or platinum group) has a low activity at a low temperature. In view of the above, hydrogen gases injected from the outside for improving the catalyst activity even under a low temperature state such as reactor start-up and shut-down. When the hydrogen gas concentration increases, chemical combination between oxygen and hydrogen in the recombiner is increased, by which the amount of heat generation is increased to rise the temperature of the catalyst. In this way, the catalyst activity can be improved. Corresponding to the injection of the hydrogen gas, oxygen is also injected so as to attain 1 : 2 ratio. (Ikeda, J.)

265

AECL passive autocatalytic recombiners  

International Nuclear Information System (INIS)

Atomic Energy of Canada Limited's (AECL) Passive Autocatalytic Recombiner (PAR) is a passive device used for hydrogen mitigation under post-accident conditions in nuclear reactor containment. The PAR employs a proprietary AECL catalyst which promotes the exothermal reaction between hydrogen and oxygen to form water vapour. The heat of reaction combined with the PAR geometry establishes a convective flow through the recombiner, where ambient hydrogen-rich gas enters the PAR inlet and hot, humid, hydrogen-depleted gas exits the outlet. AECL's PAR has been extensively qualified for CANDU and light water reactors (LWRs), and has been supplied to France, Finland, Ukraine, South Korea and is currently being deployed in Canadian nuclear power plants. (author)

266

SUMO Wrestles with Recombination  

Directory of Open Access Journals (Sweden)

Full Text Available DNA double-strand breaks (DSBs comprise one of the most toxic DNA lesions, as the failure to repair a single DSB has detrimental consequences on the cell. Homologous recombination (HR constitutes an error-free repair pathway for the repair of DSBs. On the other hand, when uncontrolled, HR can lead to genome rearrangements and needs to be tightly regulated. In recent years, several proteins involved in different steps of HR have been shown to undergo modification by small ubiquitin-like modifier (SUMO peptide and it has been suggested that deficient sumoylation impairs the progression of HR. This review addresses specific effects of sumoylation on the properties of various HR proteins and describes its importance for the homeostasis of DNA repetitive sequences. The article further illustrates the role of sumoylation in meiotic recombination and the interplay between SUMO and other post-translational modifications.

Lumír Krej?í

2012-07-01

267

Site directed recombination  

Science.gov (United States)

Enhanced homologous recombination is obtained by employing a consensus sequence which has been found to be associated with integration of repeat sequences, such as Alu and ID. The consensus sequence or sequence having a single transition mutation determines one site of a double break which allows for high efficiency of integration at the site. By introducing single or double stranded DNA having the consensus sequence flanking region joined to a sequence of interest, one can reproducibly direct integration of the sequence of interest at one or a limited number of sites. In this way, specific sites can be identified and homologous recombination achieved at the site by employing a second flanking sequence associated with a sequence proximal to the 3'-nick.

Jurka, Jerzy W. (Los Altos, CA)

1997-01-01

268

Topics in statistical mechanics  

International Nuclear Information System (INIS)

This thesis deals with four independent topics in statistical mechanics: (1) the dimer problem is solved exactly for a hexagonal lattice with general boundary using a known generating function from the theory of partitions. It is shown that the leading term in the entropy depends on the shape of the boundary; (2) continuum models of percolation and self-avoiding walks are introduced with the property that their series expansions are sums over linear graphs with intrinsic combinatorial weights and explicit dimension dependence; (3) a constrained SOS model is used to describe the edge of a simple cubic crystal. Low and high temperature results are derived as well as the detailed behavior near the crystal facet; (4) the microscopic model of the lambda-transition involving atomic permutation cycles is reexamined. In particular, a new derivation of the two-component field theory model of the critical behavior is presented. Results for a lattice model originally proposed by Kikuchi are extended with a high temperature series expansion and Monte Carlo simulation. 30 references

269

Intrachromosomal recombination in plants.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Molecular evidence for intrachromosomal recombination between closely linked DNA repeats within the plant genome is presented. The non-overlapping complementary deletion derivatives of the selectable neomycin phosphotransferase gene (nptII), when intact conferring kanamycin resistance, were inserted into the genome of Nicotiana tabacum. The functional marker gene was restored with frequencies between 10(-4) and 10(-6) per proliferating cell clone. Prolonged tissue culture prior to kanamycin s...

Peterhans, A.; Schlu?pmann, H.; Basse, C.; Paszkowski, J.

1990-01-01

270

A Simplified Recombinant PSO  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Simplified forms of the particle swarm algorithm are very beneficial in contributing to understanding how a particle swarm optimization (PSO) swarm functions. One of these forms, PSO with discrete recombination, is extended and analyzed, demonstrating not just improvements in performance relative to a standard PSO algorithm, but also significantly different behavior, namely, a reduction in bursting patterns due to the removal of stochastic components from the update equations.

Bratton, Daniel; Blackwell, Tim M.

2008-01-01

271

Relativistic dielectronic recombination theory  

International Nuclear Information System (INIS)

Dielectronic recombination (DR) is an inverse Auger process in which a free electron is captured by a recombining ion to form a doubly excited autoionizing state. The subsequent decay of the autoionizing state to a stabilized bound state by emitting photons completes the recombination process. DR is an important recombination process for high temperature plasmas. It can affect the ionization balance and level kinetics of the hot plasmas. In addition, the dielectronic satellite lines observed in the emission spectra are frequently used as plasmas diagnostic tools. In the past decade, intense theoretical and experimental studies on the DR process have been carried out. Most of the earlier theoretical calculations on the DR rate coefficients were done either by using a term average approximation or in LS coupling without including the effects of relativity and configuration interaction. The early experimental investigations were concentrated on few times ionized low-Z ions. Recently, the development of electron beam ion trap (EBIT), electron beam ion source (EBIS) and heavy ion storage ring has become possible to produce very highly-charged heavy ions (e.g. U82+ and Xe53+)and to study the interaction between electrons and these ions. For highly-charged heavy ions, one excepts that the nonrelativistic method would be inadequate and a relativistic treatment is necessary. To meet this challenge we have developed a relativistic package based on the multiconfiguration Dirac-Fock method and have carried out systematic relativistic calculations of DR cross sections and rate coefficients and resonant transfer and excitation cross sections in ion-atom collisions. In this paper, we will briefly discuss the relativistic calculations of atomic structure and transition rates and will focus for attention on the effects of relativity and intermediate coupling on the DR cross sections and rate coefficients

272

Discovering health topics in social media using topic models.  

Science.gov (United States)

By aggregating self-reported health statuses across millions of users, we seek to characterize the variety of health information discussed in Twitter. We describe a topic modeling framework for discovering health topics in Twitter, a social media website. This is an exploratory approach with the goal of understanding what health topics are commonly discussed in social media. This paper describes in detail a statistical topic model created for this purpose, the Ailment Topic Aspect Model (ATAM), as well as our system for filtering general Twitter data based on health keywords and supervised classification. We show how ATAM and other topic models can automatically infer health topics in 144 million Twitter messages from 2011 to 2013. ATAM discovered 13 coherent clusters of Twitter messages, some of which correlate with seasonal influenza (r = 0.689) and allergies (r = 0.810) temporal surveillance data, as well as exercise (r = ?.534) and obesity (r = ?-.631) related geographic survey data in the United States. These results demonstrate that it is possible to automatically discover topics that attain statistically significant correlations with ground truth data, despite using minimal human supervision and no historical data to train the model, in contrast to prior work. Additionally, these results demonstrate that a single general-purpose model can identify many different health topics in social media. PMID:25084530

Paul, Michael J; Dredze, Mark

2014-01-01

273

Recombinant peptides in thrombolysis.  

Science.gov (United States)

Recombinant thrombolytic peptides are mainly represented by recombinant forms of tissue plasminogen activator (t-PA), a proteolytic enzyme that catalyzes the conversion of plasminogen into active plasmin, which then functions to dissolve clots. The three clinically relevant recombinant thrombolytic peptides are alteplase (t-PA), reteplase (r-PA), and tenecteplase (TNK). r-PA and TNK have been structurally modified from native t-PA to increase their half-life and fibrin specificity. Thrombolytics play an important role in several diseases, including ST-segment elevation myocardial infarction (STEMI), deep vein thrombosis (DVT) and pulmonary embolism (PE), ischemic stroke, and peripheral arterial disease. Thrombolytic therapy has evolved into an alternative treatment for STEMI, reserved predominantly for patients who do not have access to timely percutaneous coronary intervention. In patients with DVT/PE or arterial related critical limb ischemia, the use of thrombolytic therapy is limited to specific patient populations. Thrombolytic therapy is the treatment of choice for ischemic stroke in patients who present

Campbell, Jennifer; Hilleman, Daniel

2010-07-01

274

Recombinant human milk proteins.  

Science.gov (United States)

Human milk provides proteins that benefit newborn infants. They not only provide amino acids, but also facilitate the absorption of nutrients, stimulate growth and development of the intestine, modulate immune function, and aid in the digestion of other nutrients. Breastfed infants have a lower prevalence of infections than formula-fed infants. Since many women in industrialized countries choose not to breastfeed, and an increasing proportion of women in developing countries are advised not to breastfeed because of the risk of HIV transmission, incorporation of recombinant human milk proteins into infant foods is likely to be beneficial. We are expressing human milk proteins known to have anti-infective activity in rice. Since rice is a normal constituent of the diet of infants and children, limited purification of the proteins is required. Lactoferrin has antimicrobial and iron-binding activities. Lysozyme is an enzyme that is bactericidal and also acts synergistically with lactoferrin. These recombinant proteins have biological activities identical to their native counterparts. They are equally resistant to heat processing, which is necessary for food applications, and to acid and proteolytic enzymes which are needed to maintain their biological activity in the gastrointestinal tract of infants. These recombinant human milk proteins may be incorporated into infant formulas, baby foods and complementary foods, and used with the goal to reduce infectious diseases. PMID:16902336

Lönnerdal, Bo

2006-01-01

275

GDP beta S enhances the activation of phospholipase C caused by thrombin in human platelets: evidence for involvement of an inhibitory GTP-binding protein  

International Nuclear Information System (INIS)

Guanosine 5'-O-thiotriphosphate (GTP gamma S) and thrombin stimulate the activity of phospholipase C in platelets that have been permeabilized with saponin and whose inositol phospholipids have been prelabeled with [3H]inositol. Ca2+ has opposite effects on the formation of [3H]inositol phosphates induced by thrombin or GTP gamma S. While the action of GTP gamma S on the formation of [3H]inositol phosphates is inhibited by Ca2+, action of thrombin is stimulated by Ca2+. Guanosine 5'-O-(2-thiodiphosphate) (GDP beta S), which inhibits the function of GTP-binding proteins, also inhibits the effect of GTP gamma S on phospholipase C stimulation but, surprisingly, increases the effect of thrombin. Ca2+ increases the inhibitory effect of GDP beta S on GTP gamma S activation of phospholipase C, but Ca2+ further enhances the stimulatory effect of GDP beta S on the thrombin activation of phospholipase C. This indicates that two mechanisms are responsible for the activation of phospholipase C in platelets. A GTP-binding protein is responsible for regulation of phospholipase C induced by GTP gamma S, while the effect of thrombin on the stimulation of phospholipase C is independent of GTP-binding proteins. However, the effect of thrombin may be modulated by the action of an inhibitory GTP-binding protein

276

Development of aptamer-conjugated magnetic graphene/gold nanoparticle hybrid nanocomposites for specific enrichment and rapid analysis of thrombin by MALDI-TOF MS.  

Science.gov (United States)

Simple, rapid and sensitive analysis of thrombin (a tumor biomarker) in complex samples is quite clinical relevant and essential for the development of disease diagnosis and pharmacotherapy. Herein, we developed a novel method based on aptamer-conjugated magnetic graphene/gold nanoparticles nanocomposites (MagG@Au) for specific enrichment and rapid analysis of thrombin in biological samples using MALDI-TOF-MS. At first, gold nanoparticles were compactly deposited on PDDA functionalized magnetic graphene through electrostatic interaction. Afterwards, aptamer was easily conjugated to gold nanoparticles via Au-S bond formation. The as-made aptamer-conjugated nanocomposites took advantage of the magnetism of magnetic graphene, the high affinity and specificity of aptamer, facilitating a high-efficient separation and enrichment of thrombin. More importantly, due to the large surface area of the hybrid substrate, the average coverage density of aptamer achieved 0.34 nmol/mg, which enhanced the thrombin binding capacity and the recovery of thrombin in real samples. In turn, the enriched thrombin attributed to the sensitive output of MALDI-TOF mass spectrometry signal, 0.085 ng ?L(-1) (2.36 nM) thrombin could be detected. This proposed method has a relatively wide linear relation ranging from 0.1 ng ?L(-1) to 10 ng ?L(-1), and satisfactory specificity. The proposed high-throughput method based on MALDI-TOF MS is expected to the application in the disease biomarker detection and clinical diagnosis. PMID:25127596

Xiong, Ya; Deng, Chunhui; Zhang, Xiangmin

2014-11-01

277

Effect of Locked-Nucleic Acid on a Biologically Active G-Quadruplex. A Structure-Activity Relationship of the Thrombin Aptamer  

Directory of Open Access Journals (Sweden)

Full Text Available Here we tested the ability to augment the biological activity of the thrombin aptamer, d(GGTTGGTGTGGTTGG, by using locked nucleic acid (LNA to influence its G-quadruplex structure. Compared to un-substituted control aptamer, LNA-containing aptamers displayed varying degrees of thrombin inhibition. Aptamers with LNA substituted in either positions G5, T7, or G8 showed decreased thrombin inhibition, whereas LNA at position G2 displayed activity comparable to un-substituted control aptamer. Interestingly, the thermal stability of the substituted aptamers does not correlate to activity – the more stable aptamers with LNA in position G5, T7, or G8 showed the least thrombin inhibition, while a less stable aptamer with LNA at G2 was as active as the un-substituted aptamer. These results suggest that LNA substitution at sites G5, T7, and G8 directly perturbs aptamer-thrombin affinity. This further implies that for the thrombin aptamer, activity is not dictated solely by the stability of the G-quadruplex structure, but by specific interactions between the central TGT loop and thrombin and that LNA can be tolerated in a biologically active nucleic acid structure albeit in a position dependent fashion.

Michael B. Jarstfer

2008-03-01

278

A electrochemiluminescence aptasensor for detection of thrombin incorporating the capture aptamer labeled with gold nanoparticles immobilized onto the thio-silanized ITO electrode  

International Nuclear Information System (INIS)

A novel electrochemiluminescence (ECL) aptasensor was proposed for sensitive and cost-effective detection of the target thrombin adopted an aptamer-based sandwich format. To detect thrombin, capture aptamers labeled with gold nanoparticles (AuNPs) were first immobilized onto the thio-silanized ITO electrode surface through strong Au-S bonds. After catching the target thrombin, signal aptamers tagged with ECL labels were attached to the assembled electrode surface. As a result, an AuNPs-capture-aptamer/thrombin/ECL-tagged-signal-aptamer sandwich type was formed. Treating the resulting electrode surface with tri-n-propylamine (TPA) and applying a swept potential to the electrode, ECL response was generated which realized the detection of target protein. Spectroscopy and electrochemical impedance techniques were used to characterize and confirm the fabrication of the ECL aptasensor. AuNPs amplification and smart sensor fabrication art were implemented for the sensitive and cost-effective detection purpose. Signal-to-dose curve excellently followed a sandwich format equation and could be used to quantify the protein, and the detection limit was estimated to be 10 nM. Other forms of thrombin such as ?- and ?-thrombins had negligible response, which indicated a high specificity of ?-thrombin detection. The aptasensor opened up new fields of aptamer applications in ECL domain, a highly sensitive technique, and had a promising perspective to be applied in microarray analysis

279

Personal experiences in direct ultrasound-guided injection of thrombin into the lumen of pseudoaneurysm as a method of treatment in case of iatrogenic femoral artery damage  

International Nuclear Information System (INIS)

Background: Pseudoaneurysms constitute a quite common complication of procedures requiring puncture of the common femoral artery. The risk factors of the condition include: obesity, arterial hypertension, sex (more prevalent in males) as well as antithrombotic therapy. Material/Methods: The US-guided injection of thrombin into the pseudoaneurysm lumen was performed in patients referred from the Department of Invasive Cardiology who had undergone coronarography or coronary angioplasty. Pseudoaneurysms constituted the complication of common femoral artery canulation. After setting the diagnosis of pseudoaneurysm by means of Doppler ultrasound, patients with large pseudoaneurysms of volume exceeding 10 mm were qualified for thrombin injection. Generally, 33 patients underwent the treatment. In 3 cases - due to the presence of multiocular pseudoaneurysm - thrombin was administered twice. Results: Taking into account the safety of the procedure, ultimately 33 patients were qualified for thrombin administration, in whom aneurism of diameter exceeding 10 mm was diagnosed. In 3 patients with aneurysm of less than 10 mm, only a compression band was used prophylactically. In one case, because of a considerable oedema surrounding the tissue, as well as deep location of the aneurysm in the groin, thrombin treatment was not given due to technical reasons. In 30 cases, single administration of thrombin was effective and resulted in a complete thrombosis of the pseudoaneurism lumen within a couple of seconds following thrombin injection. In 3 patients with multicellular aneurysm, thrombin was given twice, resulting in a total obliteration of the pseudoaneurysm in two cases only. No complications were observed after the performed procedures. No recanalisation of pseudoaneurysms was demonstrated in follow-up examinations. Conclusions: 1. Direct thrombin injection into the pseudoaneurysm lumen can constitute an alternative method of treatment for open surgical techniques. 2. The procedure is highly effective, cheap and minimally invasive. (authors)

280

An Automatic Topic Identification Algorithm  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Problem statement: Topic is a stream of words which stands for the content of a text. Knowing the topic of a document can help people to be aware from its content and facilitate their searching process. Approach: This paper proposes an automatic algorithm to identify the topic for a textual document based on the chunks corresponding to each sentences in the document. Results and conclusion: We achieved 86% matching for both total and partial matching i...

Baghdadi, Hossein S.; Bali Ranaivo-Malançon

2011-01-01

 
 
 
 
281

Topic of the Moment - Supernovae  

...Topic of the Moment - Supernovae Topic of the Moment - Supernovae Topic of the Moment - Supernovae This site uses cookies. By continuing ...Other IOP websites Energy harvesting On this day Topic of the Moment - Supernovae A new supernova has been spotted in the galaxy M82. ... On 22 January, it was announced that astronomers had found a new supernova in the galaxy M82, about 12 million lightyears away. ...The discovery is exciting astronomers since the last supernova observed within our own galaxy was seen by Kepler in 1604, but what is ...

282

[E. coli-based production of recombinant HMG-17 and its antibacterial domain].  

Science.gov (United States)

Total RNA was extracted from human LAK cell, and a cDNA encoding mature peptide HMG-17 and its alpha helix domain was amplified by RT-PCR. The recombinant prokaryotic expression vector pGEX-1lambdaT-HMG-17 and pGEX-1lambdaT HMG-17alpha helix was constructed. Using affinity chromatography, thrombin cleaving and AU-PAGE elution, we obtained the purified HMG-17. Analyses of MIC, MEC and MBC indicated that HMG-17 and HMG-17alpha had strong antibacterial activity. MIC of the alpha-helic domain was almost the same as that of HMG17, suggesting that the alpha-helic structure would be essential for the antibacterial activity of HMG-17. PMID:16156270

Feng, Yun; Yang, Huarong; Huangning; Wu, Qi; Bao, Lang; Wang, Boyao

2005-08-01

283

Hot topic [editorial  

International Nuclear Information System (INIS)

There is strong evidence for the human impact on climate change, but we should not ignore those who think otherwise. Unseasonably warm weather in many parts of Europe and North America last month will probably have added to the impression in many people's minds that climate change is a reality and that humans are guilty of warming our planet. The several hundred members of the United Nations' Intergovernmental Panel on Climate Change (IPCC) certainly think that the evidence for anthropogenic climate change is solid. Although Physics World was unable to obtain a copy of the IPCC's latest report on the science of climate change before its release date of 2 February - a clear sign of how sensitive its findings are - hints from those involved in writing the report suggest that the IPCC will have strengthened its conclusions, previously stated in 2001, that humans are heating up the Earth. While most scientists probably share this view, there are some who think otherwise. Many of those are either scientifically ill-informed or have dubious links with the energy industry. But some have genuine doubts. One bona fide sceptic is Richard Lindzen, a climate physicist from the Massachusetts Institute of Technology in the US, who was involved in preparing the IPCC's 2001 scientific report. While he does not dispute that the Earth is getting hotter, Lindzen thinks that, in all probability, the warming is largely the result of natural variations in the Earth's climate. Lindzen believes that climate models, although rooted in physics, contain far too many uncertainties to provide accurate forecasts. Indeed, mainstream climate physicists admit their computer models are far from perfect. Writing in their feature, for example, the chief scientist of the UK's Meteorological Office and colleagues describe how hard it is to incorporate the impact of clouds, which are much smaller than the resolution of the best models. They also warn that if clouds were modelled incorrectly, climate simulations 'would be seriously in error'. One may ask if this magazine should give space to Lindzen or those involved in geoengineering to air their views. Given the uncertainties still present within climate models and the potential costs of dealing with global warming, it would be wrong for Physics World to ignore those outside the mainstream. After all, as Richard Feynman once wrote: 'There is no harm in doubt and scepticism, for it is through these that new discoveries are made'. Physicists should never take anything at face value, not least a topic as important as climate change. (U.K.)

284

[Influence of new N,N'-substituted piperazines on thrombin-induced platelet activation].  

Science.gov (United States)

We have studied the influence of new N,N'-substituted piperazines with variable nature of the linker between piperazine and aromatic cycles (carbonyl group in VR-0411 versus sulfonyl group in VR-0511) on thrombin-induced platelet aggregation, cytoplasmic Ca2+ mobilization in platelets, and P-selectin exposure on the platelet plasma membrane. The inhibitory effect of VR-0511 on platelet aggregation exceeds the effects of the reference compound aspirin and VR-0411 by 35 and 42%, respectively (p compounds on the mobilization of cytoplasmic Ca2+ in platelets and P-selectin exposure indicate that VR-0411 and VR-0511 inhibit platelet activation in these tests by 28 and 61%, (p < or = 0.01) and 34 and 58% (p < or = 0.01), respectively. The possible targets for VR-0411 and VR-0511 are thromboxane and inositol triphosphate-dependent (IP3 formation) pathways of activation signal transfer. PMID:25335388

Veselkina, O C; Petrishchev, N N; Vasina, L V; Borovitov, M E; Seliutin, A V; Chepanov, S V; Sel'kov, S A

2014-01-01

285

[Method of simultaneous determination of kallikrein, plasmin and thrombin precursors and inhibitors in human blood plasma].  

Science.gov (United States)

Prekallikrein, plasminogen and prothrombin of human blood plasma have been separately activated by caolin streptokinase and thromboplastin. By measuring the TAME-esterase (N-d Tozy-L-arginine methyl ester) activity of each enzyme and its changes in the course of plasma incubation with the activator, it was possible to estimate the values of precursors of kallikrein, plasmin, thrombin and their inhibitors. Evidence is given that under conditions described the activation is specific of each enzyme and does not affect the level of the two other percursors. The method has been developed in two modifications, permitting to obtain the value of seven parameters in 0.4--0.7 ml of blood plasma. PMID:132976

Gomazkov, O A; Komissarova, N V

1976-05-01

286

Ultrasensitive electrochemiluminescence detection of thrombin based on aptamer and cystamine modified gold nanoparticle probe  

Science.gov (United States)

Recently, our group showed that one can detect specific oligonucleotides at low femtomolar levels with the electrochemiluminescence (ECL) biobarcode approach based on tris-(2, 2'-bipyridyl) ruthenium (TBR)-labeled cysteamine. It would be a significant advance to use the cysteamine assisted ECL biobarcode assay to detect protein targets in addition to DNA targets. Taking advantage of sandwich binding of two affinity aptamers for increased specificity, TBR-cysteamine as biobarcode for signal amplification and magnetic beads based ECL technology for rapid detection, a promising assay for thrombin quantification is developed. The sandwich complex could be selectively captured by micromagnetic particles and then quantified by ECL signals. Current cysteamine-Gold nanoparticle (GNP) conjugates based ECL biobarcode assay is expected to become a powerful tool for protein analysis.

Duan, Ruixue; Zhou, Xiaoming

2012-03-01

287

Hemoperitoneum presenting with the use of a topical hemostatic agent in oocyte retrieval: a case report  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Introduction Hemoperitoneum may occur from an ovarian puncture point after oocyte retrieval. Case presentation We report a case of massive hemoperitoneum following transvaginal ultrasound-guided oocyte retrieval in a 33-year-old Caucasian woman. The bleeding required emergency laparoscopy because of active bleeding from the ovarian puncture point. Hemostasis was very difficult to achieve, and traditional operative procedures were not efficient. The only way to stop the bleeding was by using an absorbable fibrinogen and thrombin sealant sponge, which was applied around the ovary. During laparoscopy three pints of packed red blood were administered. No specific alteration of screening coagulation tests was found one month later. Conclusions Hemostasis can be very difficult to achieve with traditional operative procedures. Topical hemostatic agents can be useful to preserve the ovary wherever possible.

Chatrian Amélie

2012-11-01

288

Purification and properties of a coagulant thrombin-like enzyme from the venom of Bothrops leucurus.  

Science.gov (United States)

A thrombin-like enzyme from Bothrops leucurus venom, named leucurobin (leuc), was purified by gel filtration, affinity and ion exchange chromatographies. Physicochemical studies indicated that the purified enzyme is a 35 kDa monomeric glycoprotein on SDS-PAGE under reducing conditions, which decreased to 29 kDa after deglycosylation with N-glycosidase F (PNGase F). The amino acid sequence of leuc was determined by automated sequencing of the intact native protein and peptides produced by digestion of the S-pyridyl-ethylated protein with trypsin. The protein sequence exhibits significant similarities with other serine proteases reported from snake venoms, and contains two potential sites of N-linked glycosylation. The proteinase split off fibrinopeptide A (FPA) rapidly from human fibrinogen; however, only negligible traces of fibrinopeptide B (FPB) were observed. In addition, the enzyme released the N-terminal peptide (Mr=4572) containing the first 42 residues from the Bbeta-chain. Leuc could neither activate factor XIII nor release kinins from heat-treated bovine plasma. Its specific clotting activity was equivalent to 198 NIH thrombin U/mg on human fibrinogen. Kinetic properties of leuc were determined using representative chromogenic substrates. The enzyme evoked the gyroxin syndrome when injected into the tail veins of mice at levels of 0.143 microg/g mouse. The inhibitory effects of PMSF and benzamidine on the amidolytic activity suggest that leuc is a serine proteinase, and inhibition by beta-mercaptoethanol revealed the important role of the disulfide bonds in the stabilization of the native structure. Antibothropic serum, SBTI and EDTA had little or no effect on its amidolytic activity. However, the clotting effect of the enzyme was strongly inhibited by antibothropic serum. A Dixon plot showed that the hydrolysis of Bz-L-Arg-pNA by leuc was competitively inhibited by benzamidine (Ki=1.61+/-0.25 mM). PMID:16481207

Magalhães, Arinos; Magalhães, Henrique P B; Richardson, Michael; Gontijo, Silea; Ferreira, Rodrigo N; Almeida, Alvair P; Sanchez, Eladio F

2007-04-01

289

Resolution of radiolabeled molecular species of phospholipid in human platelets: effect of thrombin  

International Nuclear Information System (INIS)

Resolution of individual molecular species of human platelet 1,2-diradyl-sn-glycero-3-phosphocholines and 1,2-diradyl-sn-glycero-3-phosphoethanolamines by reverse phase high pressure liquid chromatography (HPLC) allowed a thorough analysis of those phospholipids labeled with [3H]arachidonic acid. Approximately 54% and 16% of the total incorporated radiolabel was found in choline glycerophospholipids and ethanolamine glycerophospholipids, respectively, with ca. 90% of this being found in the 1,2-diacyl molecular species. Eighty percent of [3H]-arachidonic acid incorporated into 1-acyl-2-arachidonoyl-sn-glycero-3-phosphocholine in resting platelets was equally distributed between 1-palmitoyl-2-arachidonoyl and 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphocholine, while 70% of the radiolabel in 1-acyl-2-arachidonoyl-sn-glycero-3-phosphoethanolamine was found in 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphoethanolamine. Thrombin stimulation (5 U/ml for 5 min) resulted in deacylation of all 1-acyl-2-[3H]arachidonoyl molecular species of 1-acyl-2-arachidonoyl-sn-glycero-3-phosphocholine and 1-acyl-2-arachidonoyl-sn-glycero-3-ethanolamine. There was also a slight increase in 1-O-alkyl-2-[3H]arachidonoyl-sn-glycero-3-phosphocholine and a significant increase in 1-O-alk-1'-enyl-2-[3H]arachidonoyl-sn-glycero-3-phosphoethanolamine molecular species of over 300%. Thus, HPLC methodology indicates that arachidonoyl-containingogy indicates that arachidonoyl-containing molecular species of phosphatidylcholine and phosphatidylethanolamine are the major source of arachidonic acid in thrombin-stimulated human platelets, while certain ether phospholipid molecular species become enriched in arachidonate

290

Recombinant Collagenlike Proteins  

Science.gov (United States)

A group of collagenlike recombinant proteins containing high densities of biologically active sites has been invented. The method used to express these proteins is similar to a method of expressing recombinant procollagens and collagens described in U. S. Patent 5,593,859, "Synthesis of human procollagens and collagens in recombinant DNA systems." Customized collagenous proteins are needed for biomedical applications. In particular, fibrillar collagens are attractive for production of matrices needed for tissue engineering and drug delivery. Prior to this invention, there was no way of producing customized collagenous proteins for these and other applications. Heretofore, collagenous proteins have been produced by use of such biological systems as yeasts, bacteria, and transgenic animals and plants. These products are normal collagens that can also be extracted from such sources as tendons, bones, and hides. These products cannot be made to consist only of biologically active, specific amino acid sequences that may be needed for specific applications. Prior to this invention, it had been established that fibrillar collagens consist of domains that are responsible for such processes as interaction with cells, binding of growth factors, and interaction with a number of structural proteins present in the extracellular matrix. A normal collagen consists of a sequence of domains that can be represented by a corresponding sequence of labels, e.g., D1D2D3D4. A collagenlike protein of the present invention contains regions of collagen II that contain multiples of a single domain (e.g., D1D1D1D1 or D4D4D4D4) chosen for its specific biological activity. By virtue of the multiplicity of the chosen domain, the density of sites having that specific biological activity is greater than it is in a normal collagen. A collagenlike protein according to this invention can thus be made to have properties that are necessary for tissue engineering.

Fertala, Andzej

2007-01-01

291

Linguistic Extensions of Topic Models  

Science.gov (United States)

Topic models like latent Dirichlet allocation (LDA) provide a framework for analyzing large datasets where observations are collected into groups. Although topic modeling has been fruitfully applied to problems social science, biology, and computer vision, it has been most widely used to model datasets where documents are modeled as exchangeable…

Boyd-Graber, Jordan

2010-01-01

292

Clindamycin and Benzoyl Peroxide Topical  

Science.gov (United States)

... peroxide are in a class of medications called topical antibiotics. The combination of clindamycin and benzoyl peroxide works ... erythromycin (E.E.S., E-Mycin, Erythrocin) and other topical medications ... diarrhea caused by antibiotics.tell your doctor if you are pregnant, plan ...

293

Topics: From Myths to Objectives.  

Science.gov (United States)

This paper outlines the development of the topic approach to learning in Great Britain and the tradition of objectives-based teaching; presents integrated schemes of work which merge the two approaches for students with severe learning difficulties; and presents an example, using the topic of the school garden. (JDD)

Byers, Richard

1990-01-01

294

Comparative studies on the topical administration of mucopolysaccharide and heparin ointments in nonhuman primates.  

Science.gov (United States)

Mucopolysaccharide polysulfate (MPS) represents a mammalian-derived sulfated polysaccharide. Because the origin and structure of heparins is similar to MPS, this study was conducted to compare 2 ointment formulations containing MPS or heparin with a placebo ointment on tissue factor pathway inhibitor (TFPI) released in nonhuman primates (Macaca mulatta). A primate colony composed of 18 animals, housed at Loyola University Medical Center, was used in compliance with an Institutional Animal Care and Use Committee (IACUC)-approved protocol. Mucopolysaccharide polysulfate (4.5%), heparin (4.5%), and a placebo ointment were topically applied to individual groups of primates in a crossover study for periods of up to 2 weeks. Blood samples were drawn on days 1, 2, 5, 7, and 10. The anticoagulant effects (activated partial thromboplastin time [APTT], Heptest, thrombin time [TT]), TFPI antigen and functional levels, thrombin activatable fibrinolytic inhibitor (TAFI), and antiheparin platelet factor 4 antibodies (AHPF4 abs) were measured in citrated plasma. All data were compiled as mean +/- 1 standard deviation and compared in groups. Topical administration of both the MPS and heparin ointments resulted in no measurable anticoagulant effects in the primate model; however, MPS produced a concentration-dependent release of TFPI antigen and a functional activity that was stronger than the effects observed with heparin. A decrease in TAFI activation was also observed in the MPS-treated primates. In addition, in the heparin-treated group, a slight increase in AHPF4 abs was observed. In conclusion, MPS showed a stronger release of TFPI than heparin that was not associated with a strong anticoagulant effect. Moreover, MPS downregulated TAFI, resulting in an enhanced fibrinolytic effect. PMID:19959490

Hoppensteadt, Debra A; Neville, Brian; Schultz, Christopher; Jeske, Walter; Raake, Wolfram; Fareed, Jawed

2010-02-01

295

Thrombin induces ICAM-1 expression in human lung epithelial cells via c-Src/PDGFR/PI3K/Akt-dependent NF-?B/p300 activation.  

Science.gov (United States)

Up-regulation of ICAM-1 (intercellular adhesion molecule-1) is frequently implicated in lung inflammation and lung diseases, such as IPF (idiopathic pulmonary fibrosis). Thrombin has been shown to play a key role in inflammation via the induction of adhesion molecules, which then causes lung injury. However, the mechanisms underlying thrombin-induced ICAM-1 expression in HPAEpiCs (human pulmonary alveolar epithelial cells) remain unclear. In the present study, we have shown that thrombin induced ICAM-1 expression in HPAEpiCs. Pre-treatment with the inhibitor of thrombin [PPACK (D-Phe-Pro-Arg-chloromethyl ketone)], c-Src (PP1), PDGFR (platelet-derived growth factor receptor) (AG1296), PI3K (phosohinositide 3-kinase) (LY294002), NF-?B (nuclear factor ?B) (Bay11-7082) or p300 (GR343) and transfection with siRNAs of c-Src, PDGFR, Akt, p65 and p300 markedly reduced thrombin-induced ICAM-1 expression and monocyte adherence to HPAEpiCs challenged with thrombin. In addition, we established that thrombin stimulated the phosphorylation of c-Src, PDGFR, Akt and p65, which were inhibited by pre-treatment with their respective inhibitors PP1, AG1296, LY294002 or Bay11-7082. In addition, thrombin also enhanced Akt and NF-?B translocation from the cytosol to the nucleus, which was reduced by PP1, AG1296 or LY294002. Thrombin induced NF-?B promoter activity and the formation of the p65-Akt-p300 complex, which were inhibited by AG1296, LY294002 or PP1. Finally, we have shown that thrombin stimulated in vivo binding of p300, Akt and p65 to the ICAM-1 promoter, which was reduced by AG1296, LY294002, SH-5 or PP1. These results show that thrombin induced ICAM-1 expression and monocyte adherence via a c-Src/PDGFR/PI3K/Akt/NF-?B-dependent pathway in HPAEpiCs. Increased understanding of the signalling mechanisms underlying ICAM-1 gene regulation will create opportunities for the development of anti-inflammatory therapeutic strategies. PMID:24506791

Cheng, Shin-Ei; Lee, I-Ta; Lin, Chih-Chung; Hsiao, Li-Der; Yang, Chuen-Mao

2014-08-01

296

Dual-colored graphene quantum dots-labeled nanoprobes/graphene oxide: functional carbon materials for respective and simultaneous detection of DNA and thrombin  

Science.gov (United States)

Convenient and simultaneous detection of multiple biomarkers such as DNA and proteins with biocompatible materials and good analytical performance still remains a challenge. Herein, we report the respective and simultaneous detection of DNA and bovine ?-thrombin (thrombin) entirely based on biocompatible carbon materials through a specially designed fluorescence on-off-on process. Colorful fluorescence, high emission efficiency, good photostability and excellent compatibility enables graphene quantum dots (GQDs) as the best choice for fluorophores in bioprobes, and thus two-colored GQDs as labeling fluorophores were chemically bonded with specific oligonucleotide sequence and aptamer to prepare two probes targeting the DNA and thrombin, respectively. Each probe can be assembled on the graphene oxide (GO) platform spontaneously by ?–? stacking and electrostatic attraction; as a result, fast electron transfer in the assembly efficiently quenches the fluorescence of probe. The presence of DNA or thrombin can trigger the self-recognition between capturing a nucleotide sequence and its target DNA or between thrombin and its aptamer due to their specific hybridization and duplex DNA structures or the formation of apatamer–substrate complex, which is taken advantage of in order to achieve a separate quantitative analysis of DNA and thrombin. A dual-functional biosensor for simultaneous detection of DNA and thrombin was also constructed by self-assembly of two probes with distinct colors and GO platform, and was further evaluated with the presence of various concentrations of DNA and thrombin. Both biosensors serving as a general detection model for multiple species exhibit outstanding analytical performance, and are expected to be applied in vivo because of the excellent biocompatibility of their used materials.

Qian, Zhao Sheng; Shan, Xiao Yue; Chai, Lu Jing; Chen, Jian Rong; Feng, Hui

2014-10-01

297

Dual-colored graphene quantum dots-labeled nanoprobes/graphene oxide: functional carbon materials for respective and simultaneous detection of DNA and thrombin.  

Science.gov (United States)

Convenient and simultaneous detection of multiple biomarkers such as DNA and proteins with biocompatible materials and good analytical performance still remains a challenge. Herein, we report the respective and simultaneous detection of DNA and bovine ?-thrombin (thrombin) entirely based on biocompatible carbon materials through a specially designed fluorescence on-off-on process. Colorful fluorescence, high emission efficiency, good photostability and excellent compatibility enables graphene quantum dots (GQDs) as the best choice for fluorophores in bioprobes, and thus two-colored GQDs as labeling fluorophores were chemically bonded with specific oligonucleotide sequence and aptamer to prepare two probes targeting the DNA and thrombin, respectively. Each probe can be assembled on the graphene oxide (GO) platform spontaneously by ?-? stacking and electrostatic attraction; as a result, fast electron transfer in the assembly efficiently quenches the fluorescence of probe. The presence of DNA or thrombin can trigger the self-recognition between capturing a nucleotide sequence and its target DNA or between thrombin and its aptamer due to their specific hybridization and duplex DNA structures or the formation of apatamer-substrate complex, which is taken advantage of in order to achieve a separate quantitative analysis of DNA and thrombin. A dual-functional biosensor for simultaneous detection of DNA and thrombin was also constructed by self-assembly of two probes with distinct colors and GO platform, and was further evaluated with the presence of various concentrations of DNA and thrombin. Both biosensors serving as a general detection model for multiple species exhibit outstanding analytical performance, and are expected to be applied in vivo because of the excellent biocompatibility of their used materials. PMID:25248862

Sheng Qian, Zhao; Yue Shan, Xiao; Jing Chai, Lu; Rong Chen, Jian; Feng, Hui

2014-10-17

298

Molecular design and structure--activity relationships leading to the potent, selective, and orally active thrombin active site inhibitor BMS-189664.  

Science.gov (United States)

A series of structurally novel small molecule inhibitors of human alpha-thrombin was prepared to elucidate their structure-activity relationships (SARs), selectivity and activity in vivo. BMS-189664 (3) is identified as a potent, selective, and orally active reversible inhibitor of human alpha-thrombin which is efficacious in vivo in a mouse lethality model, and at inhibiting both arterial and venous thrombosis in cynomolgus monkey models. PMID:11738570

Das, Jagabandhu; Kimball, S David; Hall, Steven E; Han, Wen Ching; Iwanowicz, Edwin; Lin, James; Moquin, Robert V; Reid, Joyce A; Sack, John S; Malley, Mary F; Chang, Chiehying Y; Chong, Saeho; Wang-Iverson, David B; Roberts, Daniel G M; Seiler, Steven M; Schumacher, William A; Ogletree, Martin L

2002-01-01

299

Experiência inicial com o uso de adesivo tissular contendo trombina para tratamento do pseudo-aneurisma femoral Treatment of femoral pseudoaneurysm with thrombin tissue adhesive: initial experience  

Directory of Open Access Journals (Sweden)

Full Text Available O pseudo-aneurisma (PSA após cateterização femoral tem sido diagnosticado com regularidade em serviços com grande movimento de intervenções percutâneas, com incidência variando de 0,05 a 6%. PSA femorais pequenos podem ser acompanhados até a resolução espontânea. As opções de tratamento são: compressão guiada por ultra-som, injeção de trombina para trombose do PSA e tratamento cirúrgico. A injeção percutânea de trombina tem a vantagem de ser um procedimento indolor e rápido. Podem ser utilizados trombina isolada ou preparados contendo trombina associada a fibrinogênio e fatores de coagulação. A experiência inicial dos autores de cinco casos tratados com injeção de adesivo tissular contendo trombina mostrou resultado satisfatório em quatro; um caso necessitou tratamento cirúrgico. Não houve sucesso com uso isolado de trombina humana, porém, ocorreu trombose imediata após injeção de preparado de trombina associada a fibrinogênio/fator XIII. Neste artigo, são discutidas as opções de tratamento dos PSA femorais e a técnica do uso de trombina percutânea.Pseudoaneurysms caused by femoral artery catheterization have been regularly diagnosed in medical units with a great number of percutaneous interventions, with a documented incidence between 0.05 and 6%. Small femoral pseudoaneurysms undergo spontaneous resolution. Treatment options are: ultrasound-guided compression, thrombin injection to induce pseudoaneurysm thrombosis and surgical treatment. Percutaneous thrombin injection has the advantage of being a fast and painless procedure. Both isolated thrombin and thrombin preparations with fibrinogen and coagulation factors can be used. The authors' initial experience with five cases treated with thrombin tissue adhesive showed successful results in four; one case required surgery. There was no success with isolated human thrombin, but immediate thrombosis was achieved after injection of thrombin associated to fibrinogen and factor XIII. In this article, the treatment options for femoral pseudoaneurysms and the technique of percutaneous thrombin are discussed.

Daniel Mendes Pinto

2006-03-01

300

Experiência inicial com o uso de adesivo tissular contendo trombina para tratamento do pseudo-aneurisma femoral / Treatment of femoral pseudoaneurysm with thrombin tissue adhesive: initial experience  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese O pseudo-aneurisma (PSA) após cateterização femoral tem sido diagnosticado com regularidade em serviços com grande movimento de intervenções percutâneas, com incidência variando de 0,05 a 6%. PSA femorais pequenos podem ser acompanhados até a resolução espontânea. As opções de tratamento são: compre [...] ssão guiada por ultra-som, injeção de trombina para trombose do PSA e tratamento cirúrgico. A injeção percutânea de trombina tem a vantagem de ser um procedimento indolor e rápido. Podem ser utilizados trombina isolada ou preparados contendo trombina associada a fibrinogênio e fatores de coagulação. A experiência inicial dos autores de cinco casos tratados com injeção de adesivo tissular contendo trombina mostrou resultado satisfatório em quatro; um caso necessitou tratamento cirúrgico. Não houve sucesso com uso isolado de trombina humana, porém, ocorreu trombose imediata após injeção de preparado de trombina associada a fibrinogênio/fator XIII. Neste artigo, são discutidas as opções de tratamento dos PSA femorais e a técnica do uso de trombina percutânea. Abstract in english Pseudoaneurysms caused by femoral artery catheterization have been regularly diagnosed in medical units with a great number of percutaneous interventions, with a documented incidence between 0.05 and 6%. Small femoral pseudoaneurysms undergo spontaneous resolution. Treatment options are: ultrasound- [...] guided compression, thrombin injection to induce pseudoaneurysm thrombosis and surgical treatment. Percutaneous thrombin injection has the advantage of being a fast and painless procedure. Both isolated thrombin and thrombin preparations with fibrinogen and coagulation factors can be used. The authors' initial experience with five cases treated with thrombin tissue adhesive showed successful results in four; one case required surgery. There was no success with isolated human thrombin, but immediate thrombosis was achieved after injection of thrombin associated to fibrinogen and factor XIII. In this article, the treatment options for femoral pseudoaneurysms and the technique of percutaneous thrombin are discussed.

Daniel Mendes, Pinto; José Olimpio, Dias Júnior; Bernardo Lopes Cançado, Fonseca; Rodrigo Daniel, Moreialvar; Leonardo Ghizoni, Bez; Caetano de Sousa, Lopes.

 
 
 
 
301

Experiência inicial com o uso de adesivo tissular contendo trombina para tratamento do pseudo-aneurisma femoral / Treatment of femoral pseudoaneurysm with thrombin tissue adhesive: initial experience  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: Portuguese Abstract in portuguese O pseudo-aneurisma (PSA) após cateterização femoral tem sido diagnosticado com regularidade em serviços com grande movimento de intervenções percutâneas, com incidência variando de 0,05 a 6%. PSA femorais pequenos podem ser acompanhados até a resolução espontânea. As opções de tratamento são: compre [...] ssão guiada por ultra-som, injeção de trombina para trombose do PSA e tratamento cirúrgico. A injeção percutânea de trombina tem a vantagem de ser um procedimento indolor e rápido. Podem ser utilizados trombina isolada ou preparados contendo trombina associada a fibrinogênio e fatores de coagulação. A experiência inicial dos autores de cinco casos tratados com injeção de adesivo tissular contendo trombina mostrou resultado satisfatório em quatro; um caso necessitou tratamento cirúrgico. Não houve sucesso com uso isolado de trombina humana, porém, ocorreu trombose imediata após injeção de preparado de trombina associada a fibrinogênio/fator XIII. Neste artigo, são discutidas as opções de tratamento dos PSA femorais e a técnica do uso de trombina percutânea. Abstract in english Pseudoaneurysms caused by femoral artery catheterization have been regularly diagnosed in medical units with a great number of percutaneous interventions, with a documented incidence between 0.05 and 6%. Small femoral pseudoaneurysms undergo spontaneous resolution. Treatment options are: ultrasound- [...] guided compression, thrombin injection to induce pseudoaneurysm thrombosis and surgical treatment. Percutaneous thrombin injection has the advantage of being a fast and painless procedure. Both isolated thrombin and thrombin preparations with fibrinogen and coagulation factors can be used. The authors' initial experience with five cases treated with thrombin tissue adhesive showed successful results in four; one case required surgery. There was no success with isolated human thrombin, but immediate thrombosis was achieved after injection of thrombin associated to fibrinogen and factor XIII. In this article, the treatment options for femoral pseudoaneurysms and the technique of percutaneous thrombin are discussed.

Daniel Mendes, Pinto; José Olimpio, Dias Júnior; Bernardo Lopes Cançado, Fonseca; Rodrigo Daniel, Moreialvar; Leonardo Ghizoni, Bez; Caetano de Sousa, Lopes.

2006-03-01

302

Toehold strand displacement-driven assembly of G-quadruplex DNA for enzyme-free and non-label sensitive fluorescent detection of thrombin.  

Science.gov (United States)

Based on a new signal amplification strategy by the toehold strand displacement-driven cyclic assembly of G-quadruplex DNA, the development of an enzyme-free and non-label aptamer sensing approach for sensitive fluorescent detection of thrombin is described. The target thrombin associates with the corresponding aptamer of the partial dsDNA probes and liberates single stranded initiation sequences, which trigger the toehold strand displacement assembly of two G-quadruplex containing hairpin DNAs. This toehold strand displacement reaction leads to the cyclic reuse of the initiation sequences and the production of DNA assemblies with numerous G-quadruplex structures. The fluorescent dye, N-Methyl mesoporphyrin IX, binds to these G-quadruplex structures and generates significantly amplified fluorescent signals to achieve highly sensitive detection of thrombin down to 5pM. Besides, this method shows high selectivity towards the target thrombin against other control proteins. The developed thrombin sensing method herein avoids the modification of the probes and the involvement of any enzyme or nanomaterial labels for signal amplification. With the successful demonstration for thrombin detection, our approach can be easily adopted to monitor other target molecules in a simple, low-cost, sensitive and selective way by choosing appropriate aptamer/ligand pairs. PMID:25240130

Xu, Yunying; Zhou, Wenjiao; Zhou, Ming; Xiang, Yun; Yuan, Ruo; Chai, Yaqin

2015-02-15

303

Characterization of a thrombin-like serine protease, Kangshuanmei, isolated from the venom of a Chinese snake, Agkistrodon halys brevicaudus stejneger.  

Science.gov (United States)

An enzyme, referred to as Kangshuanmei, was isolated from the venom of the Chinese snake Agkistrodon halys brevicaudus stejneger by gel filtration chromatography followed by affinity chromatography. Kangshuanmei is composed of a single polypeptide chain with a molecular weight of approximately 34,000, estimated by SDS-PAGE. The enzyme hydrolyzed both benzoyl-arginine ethyl ester and H-D-Phe-Pip-Arg-p-nitroanilide, specific substrates for thrombin. The protease activity of Kangshuanmei was inhibited by 4-(2-aminoethyl)-benzensulfonyl fluoride, but was not affected by EDTA. The enzyme acted on human fibrinogen to form a fibrin clot and released three fragments. These fragments were shown to be fibrinopeptide A, fibrinopeptide B, and the Bbeta1-42 peptide of fibrinogen, respectively. These results indicate that Kangshuanmei is a thrombin-like serine protease with coagulant activity. However, the enzyme did not induce activation of blood coagulation factor XIII, unlike thrombin. Moreover, antithrombin-III, the specific thrombin inhibitor in plasma, had no inhibitory effect on the thrombin-like amidolytic activity of Kangshuanmei. The N-terminal amino acid sequence of the enzyme up to 50 residues was determined by a peptide sequencer. The N-terminal sequence of Kangshuanmei was highly homologous to most thrombin-like serine proteases from the venom of the snakes of the crotalidae family. PMID:11491462

Zhang, S; Ma, B; Sakai, J; Shiono, H; Matsui, T; Sugie, I; Okada, T

2001-08-01

304

Recombinant glucose uptake system  

Science.gov (United States)

Recombinant organisms are disclosed that contain a pathway for glucose uptake other than the pathway normally utilized by the host cell. In particular, the host cell is one in which glucose transport into the cell normally is coupled to PEP production. This host cell is transformed so that it uses an alternative pathway for glucose transport that is not coupled to PEP production. In a preferred embodiment, the host cell is a bacterium other than Z. mobilis that has been transformed to contain the glf and glk genes of Z. mobilis. By uncoupling glucose transport into the cell from PEP utilization, more PEP is produced for synthesis of products of commercial importance from a given quantity of biomass supplied to the host cells.

Ingrahm, Lonnie O. (Gainesville, FL); Snoep, Jacob L. (Groede, NL); Arfman, Nico (Delft, NL)

1997-01-01

305

Dielectronic Recombination Lines of C+  

CERN Document Server

The current paper presents atomic data generated to investigate the recombination lines of C II in the spectra of planetary nebulae. These data include energies of bound and autoionizing states, oscillator strengths and radiative transition probabilities, autoionization probabilities, and recombination coefficients. The R-matrix method of electron scattering theory was used to describe the C2+ plus electron system.

Sochi, Taha

2012-01-01

306

Dielectronic recombination lines of C+  

Science.gov (United States)

The present paper presents atomic data generated to investigate the recombination lines of C II in the spectra of planetary nebulae. These data include energies of bound and autoionizing states, oscillator strengths and radiative transition probabilities, autoionization probabilities, and recombination coefficients. The R-matrix method of electron scattering theory was used to describe the C2+ plus electron system.

Sochi, Taha; Storey, Peter J.

2013-11-01

307

Purification and characterization of jararassin-I, A thrombin-like enzyme from Bothrops jararaca snake venom.  

Science.gov (United States)

A thrombin-like serine protease, jararassin-I, was isolated from the venom of Bothrops jararaca. The protein was obtained in high yield and purity by a single chromatographic step using the affinity resin Benzamidine-Sepharose CL-6B. SDS-PAGE and dynamic light scattering analyses indicated that the molecular mass of the enzyme was about 30 kD. The enzyme possessed fibrinogenolytic and coagulant activities. The jararassin-I degraded the Bb chain of fibrinogen while the Aa chain and g chain were unchanged. Proteases inhibitors, PMSF and benzamidine inhibited the coagulant activity. These results showed jararassin-I is a serine protease similar to coagulating thrombin-like snake venom proteases, but it specifically cleaves Bb chain of bovine fibrinogen. Single crystals of enzyme were obtained (0.2 mm x 0.2 mm x 0.2 mm) and used for X-ray diffraction experiments. PMID:15592646

Vieira, Dëora F; Watanabe, Leandra; Sant'ana, Carolina D; Marcussi, Silvana; Sampaio, Suely V; Soares, Andreimar M; Arni, Raghuvir K

2004-12-01

308

Rapid purification and revised amino-terminal sequence of hirudin: a specific thrombin inhibitor of the bloodsucking leech.  

Science.gov (United States)

Hirudin is a specific polypeptide thrombin inhibitor consisting of 65 amino acids that is produced by the leech, Hirudo medicinalis. We describe a rapid method for the purification of hirudin from a leech extract. Crude hirudin, purchased from a commercial source, was first fractionated on a DEAE-HPLC column using a salt gradient. Hirudin activity was monitored by inhibition of the thrombin-mediated hydrolysis of a synthetic substrate H-D-Phenylalanyl-Pipecolyl-Arginine-p-Nitroanilide. The fractions containing antithrombin activity were pooled and further purified by reverse-phase HPLC. The homogeneity of purified hirudin was confirmed by a single amino-terminal sequence for 43 residues with Val-Val as the first two amino acids. Residue 33 was Asn rather than Asp as reported previously. PMID:3578808

Mao, S J; Yates, M T; Blankenship, D T; Cardin, A D; Krstenansky, J L; Lovenberg, W; Jackson, R L

1987-03-01

309

Separate induction of human blood platelet aggregation or cytotoxicity by different concentrations of PAF-acether and thrombin.  

Science.gov (United States)

Using decreasing concentrations of PAF-acether or thrombin, it was possible to observe on human platelets, first, aggregation, classically associated to activation, then , below a threshold, cytotoxicity towards Schistosoma mansoni larvae, proposed here as stimulation. These two activities appeared as distinct and antithetic. However, their induction might be the consequence of triggering of the same receptors with different intensity, since PAF-induced, but not thrombin-induced, cytotoxicity could be inhibited with specific PAF-antagonists BN 52021 and BN 52024 also known to inhibit PAF-induced aggregation. These results give credit to the hypothesis that haemostatic and cytotoxic properties of platelets are two distinct functions of these blood elements. PMID:1414687

Tran, A; Vanhée, D; Capron, A; Vorng, H; Braquet, P; Joseph, M

1992-05-01

310

5-hydroxymethyl-2'-deoxyuridine residues in the thrombin binding aptamer: investigating anticoagulant activity by making a tiny chemical modification.  

Science.gov (United States)

We report an investigation into analogues of the thrombin binding aptamer (TBA). Individual thymidines were replaced by the unusual residue 5-hydroxymethyl-2'-deoxyuridine (hmU). This differs from the canonical thymidine by a hydroxyl group on the 5-methyl group. NMR and CD data clearly indicate that all TBA derivatives retain the ability to fold into the "chair-like" quadruplex structure. The presence of the hmU residue does not significantly affect the thermal stability of the modified aptamers compared to the parent, except for analogue H9, which showed a marked increase in melting temperature. Although all TBA analogues showed decreased affinities to thrombin, H3, H7, and H9 proved to have improved anticoagulant activities. Our data open up the possibility to enhance TBA biological properties, simply by introducing small chemical modifications. PMID:25214456

Virgilio, Antonella; Petraccone, Luigi; Scuotto, Maria; Vellecco, Valentina; Bucci, Mariarosaria; Mayol, Luciano; Varra, Michela; Esposito, Veronica; Galeone, Aldo

2014-11-01

311

Iodine Absorption After Topical Administration  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Absorption from povidone-iodine preparations after topical administration has been reported to be negligible, but an elderly woman had increased serum iodine levels with possible metabolic complications after povidone-iodine solution was applied to decubitus ulcers.

Cruz, Francine Dela; Brown, Deborah Harper; Leikin, Jerrold B.; Franklin, Cory; Hryhorczuk, Daniel O.

1987-01-01

312

Present topics of nuclear energy  

International Nuclear Information System (INIS)

The report is discussing the topics: Reprocessing of spent fuel elements; Final storage of radioactive wastes; Effects of thermal power plants upon the climate; Safeguarding of nuclear facilities and fissionable materials; Properties and possibilities of plutonium. (orig./HP)

313

Erythromycin and Benzoyl Peroxide Topical  

Science.gov (United States)

The combination of erythromycin and benzoyl peroxide is used to treat acne. Erythromycin and benzoyl peroxide are in a class of medications called topical antibiotics. The combination of erythromycin ...

314

Topics of Bioengineering in Wikipedia  

Directory of Open Access Journals (Sweden)

Full Text Available The present report aims to give a snapshot of how topics from the field of bioengineering (bioinformatics, bioprocess systems, biomedical engineering, biotechnology, etc. are currently covered in the free electronic encyclopedia Wikipedia. It also offers insights and information about what Wikipedia is, how it functions, how and when to cite Wikipedian articles, if necessary. Several external wikis, devoted to topics of bioengineering, are also listed and reviewed.

Vassia Atanassova

2009-10-01

315

KEY TOPICS IN SPORTS MEDICINE  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Key Topics in Sports Medicine is a single quick reference source for sports and exercise medicine. It presents the essential information from across relevant topic areas, and includes both the core and emerging issues in this rapidly developing field. It covers: 1) Sports injuries, rehabilitation and injury prevention, 2) Exercise physiology, fitness testing and training, 3) Drugs in sport, 4) Exercise and health promotion, 5) Sport and exercise for special and clinical populations, 6) The ps...

Amir Ali Narvani; Panagiotis Thomas; Burce Lynn

2006-01-01

316

Incorporation of thrombin cleavage peptide into a protein cage for constructing a protease-responsive multifunctional delivery nanoplatform.  

Science.gov (United States)

Protein cages are spherical hollow supramolecules that are attractive nanoscale platforms for constructing cargo delivery vehicles. Using ferritin isolated from the hyperthermophilic archaeon Pyrococcus furiosus (Pf_Fn), we developed a multifunctional protein cage-based delivery nanoplatform that can hold cargo molecules securely, deliver them to the targeted sites, and release them to the targeted cells. The release is triggered by cleavage induced by the protease, thrombin. The thrombin cleavage peptide (GGLVPR/GSGAS) was inserted into the flexible loop region of Pf_Fn, which is located at a 4-fold axis of symmetry exposed on the surface of protein cages (Thr-Pf_Fn). Subsequently, the C-terminal glycine, which is situated in the interior cavity, was substituted with cysteine (G173C) to permit site-specific conjugation of cargo molecules. The introduced cysteine (G173C) was labeled with a fluorescent probe (F5M-Thr-Pf_Fn) for cell imaging and cargo release monitoring. The surface of F5M-Thr-Pf_Fn was further modified with biotins (F5M-Thr-Pf_Fn-NPB) as targeting ligands. The specific binding of dual functionalized F5M-Thr-Pf_Fn-NPB to the MDA MB 231 cell line, which overexpresses biotin-specific receptors on its surface, was confirmed by fluorescence microscopic analyses. The inserted thrombin cleavage peptides were effectively cleaved by thrombin, resulting in the release of the C-terminal helix in buffer and on the targeted cells without disruption of the cage architecture. Protein cage scaffolds that combine genetic and chemical modifications may serve as stimulus-responsive delivery nanoplatforms and provide opportunities for developing new types of theranostic nanoplatforms. PMID:23163509

Kang, Young Ji; Park, Dae Cheul; Shin, Hyun-Hee; Park, Jongnam; Kang, Sebyung

2012-12-10

317

Effects of the direct thrombin inhibitor dabigatran on ex vivo coagulation time in orthopaedic surgery patients: a population model analysis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Aims To describe the pharmacokinetic–pharmacodynamic (PK–PD) characteristics of the direct thrombin inhibitor dabigatran in hip replacement patients by assessing coagu- lation parameters activated partial thromboplastin time (aPTT) and ecarin clotting time (ECT), interindividual variability and factors affecting PD responses. Methods BISTRO I patients received oral dabigatran etexilate postsurgery for 6–10 days. Dabig- atran plasma concentrations and aPTT/ECT were measured on t...

Liesenfeld, K. H.; Scha?fer, G. H.; Troconiz, I. F.; Tillmann, C. C.; Eriksson, B. I.; Stangier, J.

2006-01-01

318

Successful direct thrombin injection of a femoral artery pseudoaneurysm in an anticoagulated patient with a mechanical mitral valve  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Femoral pseudoaneurysm is a complication of cardiac catheterization and may be related to the use of anticoagulants, antiplatelet agents, larger diameter sheaths and prolonged duration of sheath insertion. The treatment ranges from direct compression with or without direct thrombin injection to surgical repair. The present report describes a unique scenario of postcardiac catheterization femoral artery pseudoaneursym in a patient with a pre-existing mechanical mitral valve requiring anticoagu...

Gelwix, Christopher; Harrison, Melissa; Berman, Lorraine; Prinz, Andreas W.; Ali, Asghar; Jovin, Ion S.

2010-01-01

319

The Acute Coagulopathy of Trauma is due to Impaired Initial Thrombin Generation but not Clot Formation or Clot Strength  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The Acute Coagulopathy of Trauma (ACOT) has been described as a very early hypocoagulable state, but the mechanism remains controversial. One proposed mechanism is tissue hypoperfusion leading to protein C activation, with subsequent inhibition of Factors V and VIII. Variability in trauma has impeded the use of clinical data towards the elucidation of the mechanisms of ACOT, but thrombelastography (TEG) may provide insight by assessing hemostatic function from initial thrombin activation to f...

Harr, Jeffrey N.; Moore, Ernest E.; Wohlauer, Max V.; Droz, Nathan; Fragoso, Miguel; Banerjee, Anirban; Silliman, Christopher C.

2011-01-01

320

Isolation and characterization of the thrombin-like enzyme from Cryptelytrops albolabris (white-lipped tree viper) venom.  

Science.gov (United States)

A thrombin-like enzyme (termed albolabrase) was isolated in purified form from the venom of Cryptelytrops albolabris (white-lipped tree viper) using high performance anion ion exchange and gel filtration chromatography. The molecular mass of albolabrase was 33.7 kDa as determined by SDS-PAGE and 35.8 kDa as determined by Superose gel filtration chromatography. The N-terminal sequence was determined to be VVGGDECNINE which is homologous to many snake venom thrombin-like enzymes. Albolabrase exhibits both arginine ester hydrolase and arginine amidase activities and the enzyme is fastidious towards tripeptide chromogenic anilide substrates. The fibrinogen clotting activity was optimum at 3mg/mL bovine fibrinogen, and showed distinct species differences in the following decreasing order: bovine fibrinogen>dog fibrinogen?human fibrinogen>goat fibrinogen. The enzyme failed to clot both rabbit and cat fibrinogens. Reversed-phase HPLC analysis on the breakdown products of fibrinogenolytic action of albolabrase indicated that the enzyme belongs to the AB class of snake venom thrombin-like enzyme. In the indirect ELISA, IgG anti-albolabrase reacted extensively with most crotalid venoms, except with Tropidolaemus wagleri and Calloselasma rhodostoma venoms. The double sandwich ELISA, however, showed that anti-albolabrase reacted strongly only with venoms from the Trimeresurus complex, and that the results support the proposed new taxonomy changes concerning the Trimeresurus complex. PMID:21983189

Tan, Nget Hong; Fung, Shin Yee; Yap, Yeannie Hui Yeng

2012-01-01

 
 
 
 
321

A "turn-off" SERS-based detection platform for ultrasensitive detection of thrombin based on enzymatic assays.  

Science.gov (United States)

Herein we describe a novel "turn-off" biosensing strategy for thrombin detection based on Surface Enhanced Raman scattering (SERS) and the mediation of spacing between 4-mercaptobenzoic acid (4-MBA) labeled gold nanoparticles (AuNPs). The multiple arginine peptides that initiated gold nano-aggregates incorporating Raman reporter molecules due to the formation of "Hot Spots", are induced to disband by the addition of thrombin in which the peptides are catalytically cleaved into fragments and thus SERS signals of 4-MBA are sharply declined because of the weakened ability of fragments to induce aggregation. Through this strategy, a novel "turn-off" SERS biosensor for thrombin based on enzymatic amplification is established with sensitivity, selectivity and simplicity as AuNPs and peptides are easily accessible. Compared with non-enzymatic amplification based methods, this newly proposed method has improved sensitivity. The limit of detection was 160 fM (at the ratio of signal to noise, S/N=3:1). Further, controlled experiments showed that the method exhibited good selectivity over other proteases. The method demonstrated the capability and the potential for application in complex matrix and future biomarker development. The results also presented the potential and superiority of SERS biosensor based on signal amplification. PMID:23380645

Wu, Zitong; Liu, Yizhen; Zhou, Xiaodong; Shen, Aiguo; Hu, Jiming

2013-06-15

322

A sensitive electrochemical aptasensor based on water soluble CdSe quantum dots (QDs) for thrombin determination  

Energy Technology Data Exchange (ETDEWEB)

A novel aptamer biosensor with easy operation and good sensitivity, specificity, stability and reproducibility was developed by immobilizing the aptamer on water soluble CdSe quantum dots (QDs) modified on the top of the glassy carbon electrode (GCE). Methylene blue (MB) was intercalated into the aptamer sequence and used as an electrochemical marker. CdSe QDs improved the electrochemical signal because of their larger surface area and ion centers of CdSe QDs may also had a major role on amplifying the signal. The higher ion concentration caused more combination of aptamer which caused larger signal. The thrombin was detected by differential pulse voltammetry (DPV) quantitatively. Under optimal conditions, the two linear ranges were obtained from 3 to 13 {mu}g mL{sup -1} and from 14 to 31 {mu}g mL{sup -1}, respectively. The detection limit was 0.08 {mu}g mL{sup -1} at 3{sigma}. The constructed biosensor had better responses compared with that in the absence of the CdSe QDs immobilizing. The control experiment was also carried out by using BSA, casein and IgG in the absence of thrombin. The results showed that the aptasensor had good specificity, stability and reproducibility to the thrombin. Moreover, the aptasensor could be used for detection of real sample with consistent results in comparison with those obtained by fluorescence method which could provide a promising platform for fabrication of aptamer based biosensors.

Li Yanfen; Han Min [Jiangsu Laboratory of New Power Batteries, Jiangsu Key Laboratory of Biofuctional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210097 (China); Bai Hongyan [Jiangsu Laboratory of New Power Batteries, Jiangsu Key Laboratory of Biofuctional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210097 (China); College of Biological and Chemical Engineering, Jiaxing College, Jiaxing 314001 (China); Wu Yong [Jiangsu Laboratory of New Power Batteries, Jiangsu Key Laboratory of Biofuctional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210097 (China); Dai Zhihui, E-mail: daizhihuii@njnu.edu.cn [Jiangsu Laboratory of New Power Batteries, Jiangsu Key Laboratory of Biofuctional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210097 (China); Bao Jianchun, E-mail: baojianchun@njnu.edu.cn [Jiangsu Laboratory of New Power Batteries, Jiangsu Key Laboratory of Biofuctional Materials, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210097 (China)

2011-08-01

323

Thrombin and Its Receptor Enhance ST-Segment Elevation in Acute Myocardial Infarction by Activating the KATP Channel  

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ST-segment elevation is the major clinical criterion for committing patients with chest pain to have emergent coronary revascularizations; however, the mechanism responsible for ST-segment elevation is unknown. In a guinea pig model of ST-segment elevation acute myocardial infarction (AMI), local application of hirudin, a thrombin antagonist, significantly decreased AMI-induced ST-segment elevation in a dose-dependent manner. Hirudin-induced (5 antithrombin units [ATU]) decrease in ST elevation was reversed by 250 nmol/L thrombin receptor activator peptide (TRAP). TRAP (250 nmol/L [100 ?L]) significantly induced ST-segment elevation in hearts without AMI. The TRAP effect was blocked by 4 mg/kg glibenclamide and 4 mg/kg HMR1098 and partially blocked by 3 mg/kg 5HD. Pinacidil (0.45 mg/kg) simulated the effect of TRAP (250 nmol/L [100 ?L]) on hearts without AMI. Moreover, single-channel recordings showed that TRAP induced ATP-sensitive K+ channel (KATP channel) activity, and this effect was blocked by HMR1098 but not 5HD. Finally, TRAP significantly shortened the monophasic action potential (MAP) at 90% repolarization (MAP90) and epicardial MAP (EpiMAP) duration. These effects of TRAP were completely reversed by HMR1098 and partially reversed by 5HD. Thrombin and its receptor activation enhanced ST-segment elevation in an AMI model by activating the sarcolemmal KATP channel. PMID:20386871

Long, Ming; Yang, Lei; Huang, Genya; Liu, Liping; Dong, Yugang; Du, Zhimin; Tang, Anli; Hu, Chenghen; Gu, Ruimin; Gao, Xiuren; Tang, Lilong

2010-01-01

324

Clinical and pharmacological properties of new oral anticoagulants for the prevention of cerebral thromboembolism: Factor Xa and thrombin inhibitors  

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Full Text Available Vitamin K antagonists, such as warfarin and phen-procoumon, are the first-line oral anticoagulants for primary and secondary prevention of cerebral embo-lism in patients with atrial fibrillation. Although vitamin K antagonists can significantly decrease the risk of stroke, their use is limited by several important drawbacks, such as a narrow therapeutic window, the risk of intracranial and gastrointestinal bleeding, interactions with a number of drugs and nutrients, and the need for regular laboratory tests for therapy adjustment. Currently, new oral anticoagulants, such as direct thrombin inhibitors (e.g., dabigatran and direct factor Xa inhibitors (e.g., apixaban, rivaroxaban, are being developed and tested in clinical trials. Dabigatran and rivaroxaban were recently approved for prevention of cerebral embolism in patients with atrial fibrillation. The ad-vantages of dabigatran in comparison to warfarin are a lower rate of major bleedings with dabigatran 110mg bid, a better efficacy with dabigatran 150mg bid, no clinically relevant interactions with other drugs and no need for routine coagulation monitoring. The disadvantages are the absence of antidote and the absence of routine laboratory tests for precise mea-surements of anticoagulant effect of direct thrombin/ factor Xa inhibitors. This review will focus on throm-bin and factor Xa inhibitors, which are new and promising oral anticoagulants for the prevention of cerebral embolism. We will discuss their pharmacol-ogical and clinical properties and provide the most recent updates on their clinical trials.

Wolfgang H. Oertel

2012-02-01

325

Effect of thrombin and endotoxin on the in vivo metabolism of antithrombin III (AT III) in dogs  

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Effect of thrombin and endotoxin on the metabolism of I-125-labelled canine AT III was studied in mongrel dogs. Under control condition, mean total amount of intravascular AT III with standard deviation was 23.4 +/- 2.4 mg/kg, plasma half life of i.v. injected I-125-AT III was 1.7 +/- 0.2 days, and the fractional catabolic flux (j3x) was 16.3 +/- 1.6 mg/kg/day. The total amount of intra- and extra-vascular AT III was 36.0 +/- 0.34 mg/kg. Neither a 3 hour infusion of a small dose (30 units/kg/hr) of thrombin nor i.v. injection of a large amount of thrombin (5,000-15,000 units/day) with heparin significantly affected AT III metabolism except for a transient decrease in AT III concentration in the latter case, although decrease in plasma fibrinogen concentration and platelet count was observed in both cases. Two injections with 200 micrograms/kg of endotoxin resulted in an evident acceleration of AT III metabolism with significant decrease in the plasma AT III, fibrinogen concentrations and platelet count. More marked changes in AT III metabolism were induced by a single infusion with 1 mg/kg of endotoxin. Changes in hemostatic system coincided with those observed in DIC

326

Hydroxyethyl starch enhances argatroban-mediated decreases in clot propagation and strength by diminishing thrombin-fibrinogen interactions.  

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Direct thrombin inhibitors (DTIs) have been administered for anticoagulation during cardiopulmonary bypass for patients with heparin-induced thrombocytopenia. While DTIs prolonged clot initiation and decreased clot propagation, clot strength did not change. Hydroxyethyl starches (HES), however, significantly decreased clot propagation and strength. We hypothesized that DTI with HES could significantly decrease hemostasis more than DTI alone. Plasma was exposed to 0 or 5 microg/ml argatroban with 0 or 30% dilution with 0.9% NaCl, 10% pentastarch or 6% Voluven. Additional argatroban-exposed samples diluted with HES had addition of alpha-thrombin (0.25 U/ml) and fibrinogen (150 mg/ml). Clot kinetics were determined via thrombelastography. While dilution with 0.9% NaCl significantly (P argatroban, dilution with pentastarch and Voluven significantly (P argatroban. Addition of alpha-thrombin/fibrinogen restored clot strength. DTI/HES administration diminished hemostasis to a greater extent than DTI exposure alone. Further investigation is warranted to determine whether this therapeutic approach can improve the safety of anticoagulation during cardiopulmonary bypass in patients with heparin-induced thrombocytopenia. PMID:17179827

Nielsen, Vance G; Kirklin, James K

2007-01-01

327

Anti-thrombin III, Protein C, and Protein S deficiency in acute coronary syndrome  

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Full Text Available The final most common pathway for the majority of coronary artery disease is occlusion of a coronary vessel. Under normal conditions, antithrombin III (AT III, protein C, and protein S as an active protein C cofactor, are natural anticoagulants (hemostatic control that balances procoagulant activity (thrombin antithrombin complex balance to prevent thrombosis. If the condition becomes unbalanced, natural anticoagulants and the procoagulants can lead to thrombosis. Thirty subjects with acute coronary syndrome (ACS were studied for the incidence of antithrombin III (AT III, protein C, and protein S deficiencies, and the result were compare to the control group. Among patients with ACS, the frequency of distribution of AT-III with activity < 75% were 23,3% (7 of 30, and only 6,7% ( 2 of 30 in control subject. No one of the 30 control subject have protein C activity deficient, in ACS with activity < 70% were 13,3% (4 of 30. Fifteen out of the 30 (50% control subjects had protein S activity deficiency, while protein S deficiency activity < 70% was found 73.3.% (22 out of 30. On linear regression, the deterministic coefficient of AT-III activity deficiency to the development ACS was 13,25 %, and the deterministic coefficient of protein C activity deficient to the development of ACS was 9,06 %. The cut-off point for AT-III without protein S deficiency expected to contribute to the development of vessel disease was 45%. On discriminant analysis, protein C activity deficiency posed a risk for ACS of 4,5 greater than non deficient subjects, and AT-III activity deficiency posed a risk for ACS of 3,5 times greater than non deficient subjects. On binary logistic regression, protein S activity acted only as a reinforcing factor of AT-III activity deficiency in the development of ACS. Protein C and AT III deficiency can trigger ACS, with determinant coefficients of 9,06% and 13,25% respectively. Low levels of protein C posed a greater risk of ACS than low levels of AT III. Protein S deficiency was a reinforcing factor on AT-III deficient to development of ACS. The cut-off point of AT-III without protein S deficiency expected to give single vessel disease was 45%, and 9,5% for the development of triple vessel disease. (Med J Indones 2002; 11: 87-92Keywords: acute coronary syndrome, Anti-thrombin III, Protein C, Protein S

Dasnan Ismail

2002-05-01

328

Impact of moderate blast exposures on thrombin biomarkers assessed by calibrated automated thrombography in rats.  

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Severe blast exposures are frequently complicated with fatal intracranial hemorrhages. However, many more sustain low level blasts without tissue damage detectable by brain imaging. To investigate effects of nonlethal blast on thrombin-related biomarkers, rats were subjected to two different types of head-directed blast: 1) moderate "composite" blast with strong head acceleration or 2) moderate primary blast, without head acceleration. Thrombin generation (TG) ex vivo after blast was studied by calibrated automated thrombography (CAT). In the same blood samples, we assessed maximal concentration of TG (TGmax), start time, peak time, mean time, and concentrations of protein markers for vascular/hemostatic dysfunctions: integrin ?/?, soluble endothelial selectin (sE-selectin), soluble intercellular cell adhesion molecule-1 (sICAM-1), and matrix metalloproteinases (MMP)-2, MMP-8, and MMP-13. Blast remarkably affected all TG indices. In animals exposed to "composite" blast, TGmax peaked at 6?h (?4.5-fold vs. control), sustained at day 1 (?3.8-fold increase), and declined to a 2-fold increase over control at day 7 post-blast. After primary blast, TGmax also rose to ?4.2-fold of control at 6?h, dropped to ?1.7-fold of control at day 1, and then exhibited a slight secondary increase at 2-fold of control at day 7. Other TG indices did not differ significantly between two types of blast exposure. The changes were also observed in other microvascular/inflammatory/hemostatic biomarkers. Integrin ?/? and sICAM-1 levels were elevated after both "composite" and primary blast at 6?h, 1 day, and 7 days. sE-selectin exhibited near normal levels after "composite" blast, but increased significantly at 7 days after primary blast; MMP-2, MMP-8, and MMP-13 slightly rose after "composite" blast and significantly increased (?2-4-fold) after primary blast. In summary, CAT may have a clinical diagnostic utility in combination with selected set of microvascular/inflammatory biomarkers in patients subjected to low/moderate level blast exposures. PMID:23805797

Prima, Victor; Serebruany, Victor L; Svetlov, Artem; Hayes, Ronald L; Svetlov, Stanislav I

2013-11-15

329

Thrombin generation during cardiopulmonary bypass: the possible role of retransfusion of blood aspirated from the surgical field  

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Full Text Available Abstract Background In spite of using heparin-coated extracorporeal circuits, cardiopulmonary bypass (CPB is still associated with an extensive thrombin generation, which is only partially suppressed by the use of high dosages of heparin. Recent studies have focused on the origins of this thrombotic stimulus and the possible role of retransfused suctioned blood from the thoracic cavities on the activation of the extrinsic coagulation pathway. The present study was designed to find during CPB an association between retransfusion of suctioned blood from the pericardium and pleural space, containing activated factor VIIa and systemic thrombin generation. Methods Blood samples taken from 12 consenting patients who had elective cardiac surgery were assayed for plasma factor VIIa, prothrombin fragment 1+2 (F1+2, and thrombin-antithrombin (TAT concentrations. Blood aspirated from the pericardium and pleural space was collected separately, assayed for F1+2, TAT, and factor VIIa and retransfused to the patient after the aorta occlusion. Results After systemic heparinization and during CPB thrombin generation was minimal, as indicated by the lower than base line plasma levels of F1+2, and TAT after correction for hemodilution. In contrast, blood aspirated from the thoracic cavities had significantly higher levels of factor VIIa, F1+2, and TAT compared to the simultaneous samples from the blood circulation (P 1+2, and TAT rose significantly from 1.6 to 2.9 nmol/L (P = 0.002 and from 5.1 to 37.5 ?g/L (P = 0.01, respectively. The increase in both F1+2, and TAT levels correlated significantly with the amount of retransfused suctioned blood (r = 0.68, P = 0.021 and r = 0.90, P = 0.001, respectively. However, the circulating factor VIIa levels did not correlate with TAT and F1+2 levels. Conclusions These data suggest that blood aspirated from the thoracic cavities during CPB is highly thrombogenic. Retransfusion of this blood may, therefore, promote further systemic thrombin generation during CPB.

de Jong Dick S

2003-07-01

330

Topical agents in burn care  

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Full Text Available Introduction Understanding of fluid shifts and recognition of the importance of early and appropriate fluid replacement therapy have significantly reduced mortality in the early post burn period. After the bum patient successfully passes the resuscitation period, the burn wound represents the greatest threat to survival. History Since the dawn of civilization, man has been trying to find an agent which would help burn wounds heal, and at the same time, not harm general condition of the injured. It was not until the XX century, after the discovery of antibiotics, when this condition was fulfilled. In 1968, combining silver and sulfadiazine, fox made silver-sulfadiazine, which is a 1% hydro-soluble cream and a superior agent in topical treatment of burns today. Current topical agents None of the topical antimicrobial agents available today, alone or combined, have the characteristics of ideal prophylactic agents, but they eliminate colonization of burn wound, and invasive infections are infrequent. With an excellent spectrum of activity, low toxicity, and ease of application with minimal pain, silver-sulfadiazine is still the most frequently used topical agent. Conclusion The incidence of invasive infections and overall mortality have been significantly reduced after introduction of topical burn wound antimicrobial agents into practice. In most burn patients the drug of choice for prophylaxis is silver sulfadiazine. Other agents may be useful in certain clinical situations.

Mom?ilovi? Dragan

2002-01-01

331

Unifying the mechanism of recombinant FVIIa action: dose dependence is regulated differently by tissue factor and phospholipids.  

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Recombinant factor VIIa (rFVIIa) is used for treatment of hemophilia patients with inhibitors, as well for off-label treatment of severe bleeding in trauma and surgery. Effective bleeding control requires supraphysiological doses of rFVIIa, posing both high expense and uncertain thrombotic risk. Two major competing theories offer different explanations for the supraphysiological rFVIIa dosing requirement: (1) the need to overcome competition between FVIIa and FVII zymogen for tissue factor (TF) binding, and (2) a high-dose-requiring phospholipid-related pathway of FVIIa action. In the present study, we found experimental conditions in which both mechanisms contribute simultaneously and independently to rFVIIa-driven thrombin generation in FVII-deficient human plasma. From mathematical simulations of our model of FX activation, which were confirmed by thrombin-generation experiments, we conclude that the action of rFVIIa at pharmacologic doses is dominated by the TF-dependent pathway with a minor contribution from a phospholipid-dependent mechanism. We established a dose-response curve for rFVIIa that is useful to explain dosing strategies. In the present study, we present a pathway to reconcile the 2 major mechanisms of rFVIIa action, a necessary step to understanding future dose optimization and evaluation of new rFVIIa analogs currently under development. PMID:22563088

Shibeko, Alexey M; Woodle, Samuel A; Lee, Timothy K; Ovanesov, Mikhail V

2012-07-26

332

Dynamic affinity chromatography in the separation of sulfated lignins binding to thrombin.  

Science.gov (United States)

Sulfated low molecular weight lignins (LMWLs), a mixture of chemo-enzymatically prepared oligomers, have been found to be potent antagonists of coagulation. However, structures that induce anticoagulation remain unidentified. The highly polar sulfate groups on these molecules and the thousands of different structures present in these mixtures make traditional chromatographic resolution of sulfated LMWLs difficult. We performed dynamic thrombin affinity chromatography monitored using chromogenic substrate hydrolysis assay to isolate sulfated LMWL fractions that differed significantly in their biophysical and biochemical properties. Three fractions, I(35), I(55) and Peak II, were isolated from the starting complex mixture. Independent plasma clotting assays suggested that I(35) possessed good anticoagulation potential (APTT=4.2?M; PT=6.8?M), while I(55) and Peak II were approximately 10- and 100-fold less potent. The ESI-MS spectrum of this oligomeric fraction showed multiple peaks at 684.8, 610.6, 557.4, 541.4, 536.5, and 519.4m/z, which most probably arise from variably functionalized ?-O4?-?-linked trimers and/or a ?-O4?-O4-linked dimers. The first direct observation of these structures in sulfated LMWLs will greatly assist in the discovery of more potent sulfated LMWL-based anticoagulants. PMID:23122400

Liang, Aiye; Thakkar, Jay N; Hindle, Michael; Desai, Umesh R

2012-11-01

333

Peptide affinity labels for thrombin and other trypsin-like proteases  

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A peptide affinity label of the formula (I): ##STR1## wherein X is a radical capable of acting as a leaving group in a nucleophilic substitution reaction; A is an aromatic amino acid residue; B is H, or a C.sub.1 -C.sub.4 alkyl group, or aryl; Y is selected from the group consisting of hydrogen, aroyl, C.sub.1 -C.sub.6 acyl, and Q--(A)--.sub.n, wherein Q=hydrogen, aroyl, or C.sub.1 -C.sub.6 acyl, n=1-10, A is an amino acid residue selected from the aliphatic, hydroxy-containing, carboxylic acid group, and amide-thereof-containing, aromatic, sulfur-containing and imino-containing amino acids; and wherein J is selected from the group consisting of --CH.sub.2 --, --CH.sub.2 --CH.sub.2 --,--CH.sub.2 --CH.sub.2 --CH.sub.2 --, --CH.dbd.CH-- and --CH(OH)--CH.sub.2. The affinity label is useful for irreversibly inactivating thrombin and trypsin-like enzymes and may be used as a potential anticlotting agent.

Shaw, Elliott N. (Shoreham, NY); Kettner, Charles A. (Yaphank, NY)

1982-03-09

334

The role of oral direct thrombin inhibitors in the prophylaxis of venous thromboembolism.  

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Venous thromboembolism, which is manifested as deep vein thrombosis (DVT) and pulmonary embolism (PE), represents a significant cause of death, disability, and discomfort. Two million people/year are affected by VTE, making it the third most common cardiovascular disease after coronary heart disease and stroke. The rationale for VTE prophylaxis stems from the clinically silent presentation of the disease and its prevalence among hospitalized patients. At greatest risk are patients undergoing major orthopedic surgery and those admitted to the intensive care unit with acute myocardial infarction, heart failure, ischemic stroke, respiratory disease, systemic infection, or other medical conditions that immobilize patients for 5 days or longer. Several anticoagulant regimens have been effective in reducing the risk of VTE after major orthopedic surgery. For patients undergoing total hip or knee replacement, treatment with adjusted-dose warfarin, low-molecular-weight heparins, or fondaparinux may be used. Warfarin, which has been around for more than 50 years, is the only oral anticoagulant available for VTE prophylaxis. Ximelagatran, a new low-molecular-weight oral prodrug of the direct thrombin inhibitor melagatran, has advantages over warfarin that may make it the drug of choice for prevention of VTE. PMID:15624337

Hawkins, David

2004-10-01

335

Two related thrombin-like enzymes present in Bothrops atrox venom  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english This article describes the presence of two new forms of a thrombin-like enzyme, both with apparent molecular masses of 38 kDa, in Bothrops atrox venom. Both share the ability to cleave fibrinogen into fibrin and to digest casein. Both present identical Km on the substrate BApNA. Their N-terminal ami [...] no acid sequences are identical for 26 residues, sharing 80% homology with batroxobin and flavoxobin. Two groups of monoclonal antibodies (mAbs) raised against the purified enzyme forms recognized different epitopes of the putative corresponding enzymes present in B. atrox crude venom. On Western blotting analysis of B. atrox crude venom, mAbs 5DB2C8, 5AA10 and 5CF11, but not mAbs 6CC5 and 6AD2-G5, revealed two or more protein bands ranging from 25 to 38 kDa. By immunoprecipitation assays, the 6AD2-G5 mAb was able to precipitate protein bands of 36-38 kDa from B. atrox, B. leucurus, B. pradoi, B. moojeni, B. jararaca and B. neuwiedii crude venoms. Fibrinogen-clotting activity was inhibited when the same venom specimens were pre-incubated with mAb 6AD2-G5, except for B. jararaca and B. neuwiedii.

J.H., Petretski; M., Kanashiro; C.P., Silva; E.W., Alves; T.L., Kipnis.

336

Two related thrombin-like enzymes present in Bothrops atrox venom  

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Full Text Available This article describes the presence of two new forms of a thrombin-like enzyme, both with apparent molecular masses of 38 kDa, in Bothrops atrox venom. Both share the ability to cleave fibrinogen into fibrin and to digest casein. Both present identical Km on the substrate BApNA. Their N-terminal amino acid sequences are identical for 26 residues, sharing 80% homology with batroxobin and flavoxobin. Two groups of monoclonal antibodies (mAbs raised against the purified enzyme forms recognized different epitopes of the putative corresponding enzymes present in B. atrox crude venom. On Western blotting analysis of B. atrox crude venom, mAbs 5DB2C8, 5AA10 and 5CF11, but not mAbs 6CC5 and 6AD2-G5, revealed two or more protein bands ranging from 25 to 38 kDa. By immunoprecipitation assays, the 6AD2-G5 mAb was able to precipitate protein bands of 36-38 kDa from B. atrox, B. leucurus, B. pradoi, B. moojeni, B. jararaca and B. neuwiedii crude venoms. Fibrinogen-clotting activity was inhibited when the same venom specimens were pre-incubated with mAb 6AD2-G5, except for B. jararaca and B. neuwiedii.

Petretski J.H.

2000-01-01

337

Two related thrombin-like enzymes present in Bothrops atrox venom.  

Science.gov (United States)

This article describes the presence of two new forms of a thrombin-like enzyme, both with apparent molecular masses of 38 kDa, in Bothrops atrox venom. Both share the ability to cleave fibrinogen into fibrin and to digest casein. Both present identical K(m) on the substrate BApNA. Their N-terminal amino acid sequences are identical for 26 residues, sharing 80% homology with batroxobin and flavoxobin. Two groups of monoclonal antibodies (mAbs) raised against the purified enzyme forms recognized different epitopes of the putative corresponding enzymes present in B. atrox crude venom. On Western blotting analysis of B. atrox crude venom, mAbs 5DB2C8, 5AA10 and 5CF11, but not mAbs 6CC5 and 6AD2-G5, revealed two or more protein bands ranging from 25 to 38 kDa. By immunoprecipitation assays, the 6AD2-G5 mAb was able to precipitate protein bands of 36-38 kDa from B. atrox, B. leucurus, B. pradoi, B. moojeni, B. jararaca and B. neuwiedii crude venoms. Fibrinogen-clotting activity was inhibited when the same venom specimens were pre-incubated with mAb 6AD2-G5, except for B. jararaca and B. neuwiedii. PMID:11050658

Petretski, J H; Kanashiro, M; Silva, C P; Alves, E W; Kipnis, T L

2000-11-01

338

Serum thrombin activatable fibrinolysis inhibitor levels in patients with newly diagnosed multiple myeloma.  

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Multiple myeloma has been associated with the development of thromboembolic events. Thrombin activatable fibrinolysis inhibitor (TAFI) is a carboxypeptidase B-like proenzyme, which potently inhibits fibrinolysis. The purpose of the present study was to assess the TAFI levels in patients with newly diagnosed multiple myeloma. Twenty-seven newly diagnosed multiple myeloma patients (16 women and 11 men) and 27 age-matched healthy individuals (14 women and 13 men) were included in the study. Serum TAFI levels were significantly increased in patients with multiple myeloma (46 ± 13. 3 vs. 36. 6 ± 9.7 ?g/ml) compared with healthy individuals. Serum TAFI levels were negatively correlated with serum albumin (CC: -0.453, P < 0.05) and hemoglobin levels (CC: -0.392, P < 0.05) and positively correlated with the ?-2 microglobulin levels (CC: 0.524, P < 0.05). In this study, we observed significantly elevated TAFI levels in patients with multiple myeloma and higher serum TAFI levels were suggested to be associated with higher disease stage. With these results, a possible role of elevated TAFI levels in thromboembolic manifestations in the course of multiple myeloma can be suggested. PMID:21297448

Balcik, Ozlem S; Albayrak, Murat; Uyar, Mehtap E; Dagdas, Simten; Yokus, Osman; Ceran, Funda; Cipil, Handan; Kosar, Ali; Ozet, Gulsum

2011-06-01

339

Plasma levels of thrombin-activatable fibrinolysis inhibitor in primary and secondary thrombocytosis.  

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An elevated platelet count is a common finding in both hospitalized and ambulatory patients. Thrombosis and bleeding complications are more frequently observed in patients with clonal thrombocytosis than secondary thrombocytosis. The aim of this study was to investigate the behaviors of thrombin-activatable fibrinolysis inhibitor (TAFI) activity, the inhibitor of fibrinolysis, and also prothrombin time (PT), active partial thromboplastin time, and D-dimer and fibrinogen levels in 21 patients affected with clonal thrombocytemia as compared with 21 patients with reactive thrombocytosis and 21 healthy controls. In the clonal thrombocytemia group, plasma levels of TAFI activity were significantly higher than in both the reactive thrombocytosis and the control group. Plasma levels of leukocyte and platelet counts were significantly higher in the clonal thrombocytemia group than in the other two groups and also higher in the reactive thrombocytosis group than in the control group, which was also significant. Fibrinogen and D-dimer levels were higher in patients than in the control group but showed no significant difference between the clonal and secondary thrombocytosis groups. Plasma levels of PT and aPTT were higher in secondary thrombocytosis group than the clonal thrombocytosis group. The results of this study showed for the first time that TAFI activity is increased in patients with clonal thrombocytosis. These increased levels in clonal thrombocytosis can be considered a factor to explain the thrombotic tendency in myeloproliferative disorders. PMID:16244771

Kaftan, O; Balcik, O S; Cipil, H; Ozet, G; Bavbek, N; Ko?ar, A; Dagdas, S

2005-10-01

340

Thrombin antithrombin complex and IL-18 serum levels in stroke patients  

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Full Text Available The complex picture of inflammation and coagulation alterations comes to life in acute stroke phases. Increasing evidence points to a strong interaction and extensive crosstalk between the inflammation and coagulation systems: the interest towards this relationship has increased since recent experimental research showed that the early administration of antithrombin III (ATIII decreases the volume of ischemia in mice and might be neuroprotective, playing an antiinflammatory role. We aimed to establish the extent of the relationship among markers of inflammation (S100B and IL-18 and procoagulant and fibrinolytic markers (ATIII, thrombin-antithrombin III complex (TAT, Fibrin Degradation Products (FDP, D-dimer in 13 comatose patients affected by focal cerebral ischemia. Plasma levels of TAT, D-dimer and FDP, IL18 and S100B were increased. IL-18 and S100B high serum levels in ischemic patients suggest an early activation of the inflammatory cascade in acute ischemic injury. The basic principles of the interaction between inflammatory and coagulation systems are revised, from the perspective that simultaneous modulation of both coagulation and inflammation, rather than specific therapies aimed at one of these systems could be more successful in stroke therapy.

Rosalba Tufano

2010-06-01

 
 
 
 
341

Inhibitory effects of total saponin from Korean red ginseng via vasodilator-stimulated phosphoprotein-Ser(157) phosphorylation on thrombin-induced platelet aggregation.  

Science.gov (United States)

In this study, we have investigated the effects of total saponin from Korean red ginseng (TSKRG) on thrombin-induced platelet aggregation. TSKRG dose-dependently inhibited thrombin-induced platelet aggregation with IC50 value of about 81.1 ?g/mL. In addition, TSKRG dose-dependently decreased thrombin-elevated the level of cytosolic-free Ca(2+) ([Ca(2+)]i), one of aggregation-inducing molecules. Of two Ca(2+)-antagonistic cyclic nucleotides as aggregation-inhibiting molecules, cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), TSKRG significantly dose-dependently elevated intracellular level of cAMP, but not cGMP. In addition, TSKRG dose-dependently inhibited thrombin-elevated adenosine triphosphate (ATP) release from platelets. These results suggest that the suppression of [Ca(2+)]i elevation, and of ATP release by TSKRG are associated with upregulation of cAMP. TSKRG elevated the phosphorylation of vasodilator-stimulated phosphoprotein (VASP)-Ser(157), a cAMP-dependent protein kinase (A-kinase) substrate, but not the phosphorylation of VASP-Ser(239), a cGMPdependent protein kinase substrate, in thrombin-activated platelets. We demonstrate that TSKRG involves in increase of cAMP level and subsequent elevation of VASP-Ser(157) phosphorylation through A-kinase activation to inhibit [Ca(2+)]i mobilization and ATP release in thrombin-induced platelet aggregation. These results strongly indicate that TSKRG is a beneficial herbal substance elevating cAMP level in thrombin-platelet interaction, which may result in preventing of platelet aggregation-mediated thrombotic diseases. PMID:23717170

Lee, Dong-Ha; Cho, Hyun-Jeong; Kim, Hyun-Hong; Rhee, Man Hee; Ryu, Jin-Hyeob; Park, Hwa-Jin

2013-04-01

342

Affinity labeling of lysine-149 in the anion-binding exosite of human ?-thrombin with an N?-(dinitrofluorobenzyl)hirudin C-terminal peptide  

International Nuclear Information System (INIS)

In order to define structural regions in thrombin that interact with hirudin, the N?-dinitrofluorobenzyl analogue of an undecapeptide was synthesized corresponding to residues 54-64 of hirudin [GDFEEIPEEY(O35SO3)L (DNFB-[35S]Hir54-64)]. DNFB-[35S]Hir54-64 was reacted at a 10-fold molar excess with human ?-thrombin in phosphate-buffered saline at pH 7.4 and 23 degree C for 18 h. Autoradiographs of the product in reducing SDS-polyacrylamide gels revealed a single 35S-labeled band of Mr ?32,500. The labeled product was coincident with a band on Coomassie Blue stained gels migrating slightly above an unlabeled thrombin band at Mr ?31,000. Incorporation of the 35S affinity reagent peptide was found markedly reduced when reaction with thrombin was performed in the presence of 5- and 20-fold molar excesses of unlabeled hirudin peptide, showing that a specific site was involved in complex formation. The human ?-thrombin-DNFB-Hir54-64 complex was reduced, S-carboxymethylated, and treated with pepsin. Peptic fragments were separated by reverse-phase HPLC revealing two major peaks containing absorbance at 310 nm. Automated Edman degradation of the peptide fragments allowed identification of Lys-149 of human thrombin as the major site of DNFB-Hir54-64 derivatization. These data suggest that the anionic C-terminal tail of hirudin intonic C-terminal tail of hirudin interacts with an anion-binding exosite in human thrombin removed 18-20 angstrom from the catalytic apparatus

343

Recombineering: genetic engineering in bacteria using homologous recombination.  

Science.gov (United States)

The bacterial chromosome and bacterial plasmids can be engineered in vivo by homologous recombination using PCR products and synthetic oligonucleotides as substrates. This is possible because bacteriophage-encoded recombination proteins efficiently recombine sequences with homologies as short as 35 to 50 bases. Recombineering allows DNA sequences to be inserted or deleted without regard to location of restriction sites. This unit first describes preparation of electrocompetent cells expressing the recombineering functions and their transformation with dsDNA or ssDNA. It then presents support protocols that describe several two-step selection/counter-selection methods of making genetic alterations without leaving any unwanted changes in the targeted DNA, and a method for retrieving onto a plasmid a genetic marker (cloning by retrieval) from the Escherichia coli chromosome or a co-electroporated DNA fragment. Additional protocols describe methods to screen for unselected mutations, removal of the defective prophage from recombineering strains, and other useful techniques. Curr. Protoc. Mol. Biol. 106:1.16.1-1.16.39. © 2014 by John Wiley & Sons, Inc. PMID:24733238

Thomason, Lynn C; Sawitzke, James A; Li, Xintian; Costantino, Nina; Court, Donald L

2014-01-01

344

Topical timolol and lipid profile  

Directory of Open Access Journals (Sweden)

Full Text Available To evaluate the effects of topical timolol on lipid profile, blood samples from 25 patients (16 males and 9 females were analysed before topical instillation of 0.5% timolol maleate and thereafter at one month and two months. After two months of therapy, the level of high density lipoprotein (HDL decreased significantly (p < 0.02. The levels of low density lipoprotein (LDL, very low density lipoprotein (VLDL and triglyceride increased but the changes were not statistically significant. Since low level of HDL is strongly associated with increased risk of myocardial infarction, our study cautions use of topical timolol in patients with low levels of HDL or with previous history of coronary heart disease.

Manoher J

1995-01-01

345

Topical corticosteroid addiction and phobia.  

Science.gov (United States)

Corticosteroids, one of the most widely prescribed topical drugs, have been used for about six decades till date. However, rampant misuse and abuse down the years has given the drug a bad name. Topical steroid abuse may lead to two major problems which lie at the opposing ends of the psychosomatic spectrum. Topical steroid addiction, a phenomenon that came to be recognized about a decade after the introduction of the molecule is manifested as psychological distress and rebound phenomenon on stoppage of the drug. The rebound phenomenon, which can affect various parts of the body particularly the face and the genitalia has been reported by various names in the literature. TC phobia which lies at the opposite end of the psychiatric spectrum of steroid abuse has been reported particularly among parents of atopic children. Management of both conditions is difficult and frustrating. Psychological counseling and support can be of immense help in both the conditions. PMID:25284851

Ghosh, Aparajita; Sengupta, Sujata; Coondoo, Arijit; Jana, Amlan Kusum

2014-09-01

346

[Topical therapy of ulcerative colitis].  

Science.gov (United States)

The availability of new topical preparations for the treatment of left sided ulcerative colitis ulcerosa offers a therapy optimization for many patients. Rectal application of steroids and 5-aminosalicylic acid (5-ASA) is associated with fewer side effects and has a higher therapeutic efficacy in mild to moderate-active left-sided colitis as compared to a systemic therapy. Often it is argued that the patients' compliance is insufficient with a rectal therapy. However, with sufficient information on the proven advantages this is usually not the case. The rectal application of drugs in distal ulcerative colitis is suitable also for the maintenance of remission. Therefore the new therapy guidelines recommend topical therapy more than in former times. Subsequently, these manuscripts focussed specifically on the topical therapy of distal colitis, to elucidate that clear treatment advantages are present in daily practice. PMID:22086380

Rogler, G; Beglinger, C; Mottet, C; Seibold, F; Gross, V

2011-11-16

347

Topical Acne Medication Can Clear Acne  

Science.gov (United States)

... irritate your skin. Apply these medications — benzoyl peroxide, topical antibiotics, and topical retinoids — to the entire acne-prone area, not ... see redness, drying, or peeling after applying a topical acne medication? These indicate that your skin is ...

348

Special Topics in Water Science (Water Pollution)  

Science.gov (United States)

... A Teachers Contact Back to previous page Special Topics in Water Science Our Special Topics section lets you explore other water-science topic areas, such as water quality, urbanization and water, ...

349

Selected topics in nuclear structure  

International Nuclear Information System (INIS)

The Fourth International Conference on selected topics in nuclear structure was held at Dubna in July 1994 on recent experimental and theoretical investigations in nuclear structure. Topics discussed were the following: nuclear structure at low-energy excitations (collective quasiparticle phenomena, proton-neutron interactions, microscopic and phenomenological theories of nuclear structure; nuclear structure studies with charged particles. heavy ions, neutrons and photons; nuclei at high angular momenta and superdeformation, structure and decay properties of giant resonances, charge-exchange resonances and ?-decay; semiclassical approach of large amplitude collective motion and structure of hot nuclei

350

Hydrogen recombiner development at AECL  

International Nuclear Information System (INIS)

Catalytic recombiners have been developed at AECL for the purpose of hydrogen removal in post-accident nuclear containment buildings. The recombiners are based on a particular catalyst designed by AECL which has extraordinary resistance to fouling from water and water vapour and a large thermodynamic range of operation. The catalysts were developed, originally, for the purpose of heavy water manufacturing by way of a catalytic exchange process. Application of these catalyst materials in recombiners for containment applications began in the late 1980's. The first application was a passive recombiner, qualified for use in control of radiolytic hydrogen in the headspace of a pool-type experimental reactor of AECL design in 1988. The passive, or natural convection recombiner concept has continued development to commercial stage for application in power reactor containments. This paper reviews the AECL recombiner development, describes the current model and shows results from tests of full-scale recombiners in the Large Scale Vented Combustion Test Facility at AECL-WL. The AECL recombiner is designed for compactness and ease of engineering into containment. The design is a simple, open-ended rectangular enclosure with catalyst elements arranged inside to promote optimum convective flow driven by heat of recombination at the catalyst surface. Self start, as evidenced by catalyst heating and initiation of flow, is achieved in less than 1% hydrogen, with available oxygen, at room temperature and 100% relative humidity. This low temperature start-up in condensing atmospheres is viewed as the most challenging condition for wet-proofing effectiveness. Cold start-up is a vital performance requirement in containments, such as CANDU, where engineered air-cooling systems are operating and where long-term hydrogen control is required, after containment atmospheres have cooled. Once started, the removal capacity scales linearly with the inlet cross-section area and the partial pressure of hydrogen. The recombiner also reacts carbon monoxide, in the presence of hydrogen, at approximately the same rate as the hydrogen. The catalyst materials and wet-proofing are unaffected by radiation or high temperatures. Large scale tests confirm self-start behavior and demonstrate strong mixing, irrespective of recombiner placement. (author)

351

Research topics in statistical database management  

Energy Technology Data Exchange (ETDEWEB)

This report identifies research topics in statistical database management. These topics are grouped into four major areas: characteristics of statistical databases, functionality/usage, metadata, and logical models.

Denning, D.; Nicholson, W.; Sande, B.; Shoshani, A.

1983-09-01

352

Two-center dielectronic recombination  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In the presence of a neighboring atom, electron-ion recombination can proceed resonantly via excitation of an electron in the atom, with subsequent relaxation through radiative decay. It is shown that this two-center dielectronic process can largely dominate over single-center radiative recombination at internuclear distances as large as several nanometers. The relevance of the predicted process is demonstrated by using examples of water-dissolved alkali cations and warm den...

Mu?ller, C.; Voitkiv, A. B.; Lo?pez-urrutia, J. R. Crespo; Harman, Z.

2010-01-01

353

Two-center dielectronic recombination  

CERN Document Server

In the presence of a neighboring atom, electron-ion recombination can proceed resonantly via excitation of an electron in the atom, with subsequent relaxation through radiative decay. It is shown that this two-center dielectronic process can largely dominate over single-center radiative recombination at internuclear distances as large as several nanometers. The relevance of the predicted process is demonstrated by using examples of water-dissolved alkali cations and warm dense matter.

Müller, C; López-Urrutia, J R Crespo; Harman, Z

2010-01-01

354

Ethanol production by recombinant hosts  

Science.gov (United States)

Novel plasmids comprising genes which code for the alcohol dehydrogenase and pyruvate decarboxylase are described. Also described are recombinant hosts which have been transformed with genes coding for alcohol dehydrogenase and pyruvate. By virtue of their transformation with these genes, the recombinant hosts are capable of producing significant amounts of ethanol as a fermentation product. Also disclosed are methods for increasing the growth of recombinant hosts and methods for reducing the accumulation of undesirable metabolic products in the growth medium of these hosts. Also disclosed are recombinant host capable of producing significant amounts of ethanol as a fermentation product of oligosaccharides and plasmids comprising genes encoding polysaccharases, in addition to the genes described above which code for the alcohol dehydrogenase and pyruvate decarboxylase. Further, methods are described for producing ethanol from oligomeric feedstock using the recombinant hosts described above. Also provided is a method for enhancing the production of functional proteins in a recombinant host comprising overexpressing an adhB gene in the host. Further provided are process designs for fermenting oligosaccharide-containing biomass to ethanol.

Ingram, Lonnie O. (Gainesville, FL); Beall, David S. (Gainesville, FL); Burchhardt, Gerhard F. H. (Gainesville, FL); Guimaraes, Walter V. (Vicosa, BR); Ohta, Kazuyoshi (Miyazaki, JP); Wood, Brent E. (Gainesville, FL); Shanmugam, Keelnatham T. (Gainesville, FL)

1995-01-01

355

Topical Knowledge and ESL Writing  

Science.gov (United States)

This study investigates the effects of topical knowledge on ESL (English as a Second Language) writing performance in the English Language Proficiency Index (LPI), a standardized English proficiency test used by many post-secondary institutions in western Canada. The participants were 50 students with different levels of English proficiency…

He, Ling; Shi, Ling

2012-01-01

356

Seven topics in perturbative QCD  

Energy Technology Data Exchange (ETDEWEB)

The following topics of perturbative QCD are discussed: (1) deep inelastic scattering; (2) higher order corrections to e/sup +/e/sup -/ annihilation, to photon structure functions and to quarkonia decays; (3) higher order corrections to fragmentation functions and to various semi-inclusive processes; (4) higher twist contributions; (5) exclusive processes; (6) transverse momentum effects; (7) jet and photon physics.

Buras, A.J.

1980-09-01

357

Research topics in coal utilization  

Energy Technology Data Exchange (ETDEWEB)

Some specific research topics are described that would improve the understanding of coal utilization and match facilities and expertise that currently exist at LLNL. Areas discussed include: fouling, slagging, and metal corrosion; and the characterization, pyrolysis and combination of sulfur species in coal. (LEW)

Truhan, J.J.; Lucht, L.M.; Piwinskii, A.J.; Taylor, R.W.

1983-12-12

358

Topics in optics and music  

Science.gov (United States)

While the use of optics in the playback of music has been a tremendously successful technology and laser light shows are a common occurrence, other intersections of optics and music tend to be less well known. Topics such as optics-based instruments, performance tools and effects, instrument characterization and manufacturing, recording, playback, and signal processing are explored.

Sparks, Andrew W.

2012-10-01

359

Selected topics in nuclear structure  

International Nuclear Information System (INIS)

19. winter school in Zakopane was devoted to selected topics in nuclear structure such as: production of spin resonances, heavy ions reactions and their applications to the investigation of high spin states, octupole deformations, excited states and production of new elements etc. The experimental data are ofen compared with theoretical predictions. Report contains 28 papers. (M.F.W.)

360

Seven topics in perturbative QCD  

International Nuclear Information System (INIS)

The following topics of perturbative QCD are discussed: (1) Deep inelastic scattering; (2) Higher order corrections to e+e- annihilation, to photon structure functions and to quarkonia decays; (3) Higher order corrections to fragmentation functions and to various semi-inclusive processes; (4) Higher twist contributions; (5) Exclusive processes; (6) p(transverse) effects; (7) Jet and photon physics. (Auth.)

 
 
 
 
361

Seven topics in perturbative QCD  

International Nuclear Information System (INIS)

The following topics of perturbative QCD are discussed: (1) deep inelastic scattering; (2) higher order corrections to e+e- annihilation, to photon structure functions and to quarkonia decays; (3) higher order corrections to fragmentation functions and to various semi-inclusive processes; (4) higher twist contributions; (5) exclusive processes; (6) transverse momentum effects; (7) jet and photon physics

362

Optimization of the purification methods for recovery of recombinant growth hormone from Paralichthys olivaceus  

Science.gov (United States)

This study aimed to optimize the purification of recombinant growth hormone from Paralichthys olivaceus. Recombinant flounder growth hormone (r-fGH) was expressed by Escherichia coli in form of inclusion body or as soluble protein under different inducing conditions. The inclusion body was renatured using two recovery methods, i.e., dilution and dialysis. Thereafter, the refolded protein was purified by Glutathione Sepharase 4B affinity chromatography and r-fGH was obtained by cleavage of thrombin. For soluble products, r-fGH was directly purified from the lysates by Glutathione Sepharase 4B affinity chromatography. ELISA-receptor assay demonstrated that despite its low receptor binding activity, the r-fGH purified from refolded inclusion body had a higher yield (2.605 mg L-1) than that from soluble protein (1.964 mg L-1). Of the tested recovery methods, addition of renaturing buffer (pH 8.5) into denatured inclusion body yielded the best recovery rate (17.9%). This work provided an optimized purification method for high recovery of r-fGH, thus contributing to the application of r-fGH to aquaculture.

Zang, Xiaonan; Zhang, Xuecheng; Mu, Xiaosheng; Liu, Bin

2013-03-01

363

Existence of beta-methylnorleucine in recombinant hirudin produced by Escherichia coli.  

Science.gov (United States)

A gene encoding for hirudin, a potent thrombin inhibitor, was expressed in Escherichia coli, which is the most widely used host. When the recombinant hirudin analog, CX-397, was overproduced by E. coli (600 mg l(-1)) in the absence of nutrient amino acids in the culture medium, the presence of two derivatives in the final product was observed with extremely increased retention times on reverse-phase high-performance liquid chromatography. Each derivative was due to methylation of an isoleucine residue at Ile29 or Ile59 in the CX-397. The structure was deducible as beta-methylnorleucine (beta MeNle; (2S,3S)-2-amino-3-methylhexanoic acid). The modification pathway of beta MeNle is not thought to be a post-translational modification of the protein because Ile has no functional group in its side-chain. Additionally, beta MeNle is synthesized by mutants of Serratia marcescens that belong to the same family, Enterobacteriaceae, as E. coli (J. Antibiot. 34 (1981a) 1278). These findings suggest that the lack of nutrient amino acids in the culture medium leads to the synthesis of beta MeNle in E. coli, which is then activated by E. coli isoleucyl-tRNA synthetase and incorporated into the overproduced recombinant protein. PMID:11738720

Muramatsu, Ryo; Negishi, Toru; Mimoto, Tsutomu; Miura, Akira; Misawa, Satoru; Hayashi, Hideya

2002-02-14

364

A label-free aptasensor for highly sensitive detection of ATP and thrombin based on metal-enhanced PicoGreen fluorescence.  

Science.gov (United States)

A label-free fluorescence aptasensor for highly selective and sensitive detection of ATP and thrombin was developed by using PicoGreen (PG) as signal molecule and surface-bound metal-enhanced fluorescence (MEF) substrates (silver island films, SIFs) as signal enhancers. On binding with ATP or thrombin, aptamers undergo structure switching, leading to a reduction of fluorescence intensity of PG. Chang of fluorescence intensity can be magnified by SIFs. The limit of detection for ATP and thrombin is 1.3 nM and 0.073 nM, respectively. The fluorescence quenching efficiency is linear in the logarithmic scale with ATP concentration range from 10 nM to 100 ?M (R(2)=0.995) and thrombin concentration range from 0.1 nM to 100 nM (R(2)=0.997). The coefficients of variation of the intra-assay reproducibility and inter-assay reproducibility for ATP (10 ?M) assay are 3.8% and 5.2%, respectively. In addition, the aptasensor is stable and can be reliably used for ATP measurement in biological samples. Overall, the aptasensor can be a useful and cost effective tool for the specific detection of ATP, thrombin and potentially other biomolecules in biological samples. PMID:25086329

Wang, Kaiyu; Liao, Jian; Yang, Xiangyue; Zhao, Meng; Chen, Min; Yao, Weirong; Tan, Weihong; Lan, Xiaopeng

2015-01-15

365

[Three-step column chromatographic method for separation and purification of thrombin-like enzyme from venom of Agkistrodon halys pallas].  

Science.gov (United States)

A three-step column chromatographic method, utilizing CM Sepharose CL-6B, DEAE Sepharose Fast Flow and Sephadex G-75 in sequence, for the separation and purification of thrombin-like enzyme from crude venom of Agkistrodon halys pallas was developed. Based on the separation results on lab-scale chromatographic column, this isolation and purification process was amplified according to the diameter of the chromatographic columns used in labscale. It was found that when the amount of the sample loaded on the large-scale columns was 25 times as that loaded on the lab-scale columns, the quality level of the thrombin-like enzyme obtained with the large-scale columns was almost the same as that obtained with lab-scale columns. The relative molecular mass of thrombin-like enzyme obtained was about 33500, and its purity was about 96%. When using this method to isolate thrombin-like enzyme from crude venom of Agkistrodon halys pallas, the total protein yield and total activity yield of thrombin-like enzyme were about 0.3% and 64% respectively, and specific activity was more than 2000 U/mg. PMID:16830459

Bian, Liujiao; Yang, Xiaoyan; Liu, Li

2006-03-01

366

The 29-kDa proteins phosphorylated ion thrombin-activated human platelets are forms of the estrogen receptor-related 27-kDa heat shock protein  

Energy Technology Data Exchange (ETDEWEB)

Thrombin plays a critical role in platelet activation, hemostasis, and thrombosis. Cellular activation by thrombin leads to the phosphorylation of multiple proteins, most of which are unidentified. The authors have characterized several 29-kDa proteins that are rapidly phosphorylated following exposure of intact human platelets to thrombin. A murine monoclonal antibody raised to an unidentified estrogen receptor-related 29-kDa protein selectively recognized these proteins as well as a more basic, unphosphorylated 27-kDa protein. Cellular activation by thrombin led to a marked shift in the proportion of protein from the 27-kDa unphosphorylated form to the 29-kDa phosphoprotein species. Using this antibody, they isolated and sequenced a human cDNA clone encoding a protein that was identical to the mammalian 27-kDa heat shock protein (HSP27), a protein of uncertain function that is known to be phosphorylated to several forms and to be transcriptionally induced by estrogen. The 29-kDa proteins were confirmed to be phosphorylated forms of HSP27 by immunoprecipitation studies. Thus, the estrogen receptor-related protein is HSP27, and the three major 20-kDa proteins phosphorylated in thrombin-activated platelets are forms of HSP27. These data suggest a role for HSP27 in the signal transduction events of platelet activation.

Mendelsohn, M.E.; Yan Zhu; O' Neill, S. (Harvard Medical School, Boston, MA (United States))

1991-12-15

367

Recombination of H and He in Yang-Mills Gravity  

CERN Document Server

We investigate some aspects of the thermal history of the early universe according to Yang-Mills Gravity (YMG); a gauge theory of gravity set in flat spacetime. Specifically, equations for the ionization fractions of hydrogen and singly ionized helium during the recombination epoch are deduced analytically and then solved numerically. By considering several approximations we find that the presence of primordial helium and its interaction with Lyman series photons has a much stronger effect on the overall free electron density in YMG than it does in the standard, General Relativity (GR) based, model. Compared to the standard model recombination happens over a much larger range of temperatures, although there is still a very sharp temperature of last scattering around 2000 K. Since the ionization history of the universe is not directly observable we discuss how one may use it to predict the CMB power spectrum and thus test YMG. This topic will be explored in detail in an upcoming paper.

Katz, Daniel

2014-01-01

368

Dabigatran Etexilate, A Novel Oral Direct Thrombin Inhibitor, for Preventing Thromboembolic Events After Knee Replacement Arthroplasty  

Directory of Open Access Journals (Sweden)

Full Text Available Background: Dabigatran etexilate is one of the few direct thrombin inhibitors with anti-coagulant activities and the following distinctive features: taken orally, no need to closely monitor for complications, and no need for regular dose adjustments. Relying on the above mentioned valuable advantages, dabigatran etexilate can be considered as a premier choice for the prevention of venous thromboembolism after knee replacement arthroplasty. Methods: Forty five patients undergoing 50 knee replacement surgeries were included in this case-series study undertaken in Hazrat Rasool Akram and Khatam-alanbia Hospitals during 2010. Dabigatran etexilate was administered for the prevention of venous thromboembolism after knee arthroplasty in doses of 110 mg in the first 1-4 h after surgery followed by daily doses of 220 mg for 10 days. Patients were examined 3 times and a color Doppler sonography was performed on the 11th day to check for venous thrombosis. Finally, the patients were re-examined at the end of the 1st and the 3rd months postoperatively. Results: Only one out of 45 patients was diagnosed to have venous thrombosis on sonography done on the 11th day but the patient did not have any symptoms and repeat sonographies at the end of the 1st and the 3rd months postoperatively showed no venous thrombosis either. No complications were witnessed in the patients in the 3-month follow-up period. Conclusion: Dabigatran etexilate (220 mg/d for 10 days can be an effective drug against venous thrombosis after total knee replacement surgeries.

Yegane A

2012-02-01

369

Investigations for designing catalytic recombiners  

International Nuclear Information System (INIS)

In case of a severe accident in pressurised water reactors (PWR) a high amount of hydrogen up to about 20,000 m3 might be generated and released into the containments. The mixture consisting of hydrogen and oxygen may either burn or detonate, if ignited. In case of detonation the generated shock wave may endanger the components of the plant or the plant itself. Consequently, effective removal of hydrogen is required. The fact that hydrogen and oxygen react exo-thermally on catalytically acting surfaces already at low temperatures generating steam and heat is made use of in catalytic recombiners. They consist of substrates coated with catalyst (mainly platinum or palladium) which are arranged inside a casing. Being passively acting measures, recombiners do not need any additional energy supply. Experimental investigations on catalytic hydrogen recombination are conducted at FZJ (Forschungszentrum Juelich) using three test facilities. The results yield insight in the development potential of contemporary recombiner systems as well as of innovative systems. Detailed investigations on a recombiner section show strong temperature gradients over the surface of a catalytically coated sample. Dependent on the flow velocity, ignition temperature may be reached at the leading edge already at an inlet hydrogen concentration of about 5 vol.-%. The thermal strain of the substrate leads to considerable detachment of catalyst particles probably causing unintended ignition of the flammable mixture. Temperature peaks can be prevented effectively by leaving the first part of the plate uncoated. In order to avoid overheating of the catalyst elements of a recombiner even at high hydrogen concentrations a modular system of porous substrates is proposed. The metallic substrates are coated with platinum at low catalyst densities thus limiting the activity of the single specimen. A modular arrangement of these elements provides high recombination rates over a large hydrogen concentration range without igniting the mixture

370

Increased preoperative thrombin generation and low protein S level associated with unfavorable postoperative hemodynamics after coronary artery bypass grafting.  

Science.gov (United States)

In a previous study, preoperative levels of activated protein C (APC) were associated with unfavorable postoperative hemodynamics after coronary artery bypass grafting (CABG). Protein C is activated by thrombin. Protein S, the cofactor of activated protein C, has activated protein C-independent anticoagulant activity and cytoprotective effects. Therefore, the objective of this study was to test whether preoperative, baseline levels of either thrombin or protein S were associated with hemodynamic performance or markers of myocardial damage after CABG. One hundred patients undergoing elective on-pump CABG were prospectively studied. Prothrombin fragment F1+2 (a marker of thrombin generation) and free protein S were measured preoperatively and cardiac index, systemic vascular resistance index (SVRI), and pulmonary vascular resistance index (PVRI) were measured serially thereafter at fixed time points. Cardiac biomarkers CK-MBm and TnT were measured postoperatively. There was an inverse correlation between preoperative F1+2 and free protein S levels (r= -0.30, p=0.003). High preoperative F1+2 and low preoperative protein S levels were associated with a less favorable hemodynamic profile postoperatively. Patients with F1+2 in the highest decile (?0.85 nmol/l) and patients with preoperative protein S in the lowest decile (?63%) had lower CI values, and higher pulmonary and systemic vascular resistance index values postoperatively than comparison patients. Preoperative F1+2 or protein S did not correlate with postoperative cardiac biomarker levels. Baseline activation of coagulation and the balance between pro-coagulant and anti-coagulant factors preoperatively might have implications for postoperative hemodynamic recovery after CABG. PMID:21177725

Raivio, Peter M; Lassila, Riitta; Kuitunen, Anne H; Eriksson, Heidi; Suojaranta-Ylinen, Raili T; Petäjä, Jari

2011-03-01

371

Roles of platelet STIM1 and Orai1 in glycoprotein VI- and thrombin-dependent procoagulant activity and thrombus formation.  

Science.gov (United States)

In platelets, STIM1 has been recognized as the key regulatory protein in store-operated Ca(2+) entry (SOCE) with Orai1 as principal Ca(2+) entry channel. Both proteins contribute to collagen-dependent arterial thrombosis in mice in vivo. It is unclear whether STIM2 is involved. A key platelet response relying on Ca(2+) entry is the surface exposure of phosphatidylserine (PS), which accomplishes platelet procoagulant activity. We studied this response in mouse platelets deficient in STIM1, STIM2, or Orai1. Upon high shear flow of blood over collagen, Stim1(-/-) and Orai1(-/-) platelets had greatly impaired glycoprotein (GP) VI-dependent Ca(2+) signals, and they were deficient in PS exposure and thrombus formation. In contrast, Stim2(-/-) platelets reacted normally. Upon blood flow in the presence of thrombin generation and coagulation, Ca(2+) signals of Stim1(-/-) and Orai1(-/-) platelets were partly reduced, whereas the PS exposure and formation of fibrin-rich thrombi were normalized. Washed Stim1(-/-) and Orai1(-/-) platelets were deficient in GPVI-induced PS exposure and prothrombinase activity, but not when thrombin was present as co-agonist. Markedly, SKF96365, a blocker of (receptor-operated) Ca(2+) entry, inhibited Ca(2+) and procoagulant responses even in Stim1(-/-) and Orai1(-/-) platelets. These data show for the first time that: (i) STIM1 and Orai1 jointly contribute to GPVI-induced SOCE, procoagulant activity, and thrombus formation; (ii) a compensating Ca(2+) entry pathway is effective in the additional presence of thrombin; (iii) platelets contain two mechanisms of Ca(2+) entry and PS exposure, only one relying on STIM1-Orai1 interaction. PMID:20519511

Gilio, Karen; van Kruchten, Roger; Braun, Attila; Berna-Erro, Alejandro; Feijge, Marion A H; Stegner, David; van der Meijden, Paola E J; Kuijpers, Marijke J E; Varga-Szabo, David; Heemskerk, Johan W M; Nieswandt, Bernhard

2010-07-30

372

Roles of Platelet STIM1 and Orai1 in Glycoprotein VI- and Thrombin-dependent Procoagulant Activity and Thrombus Formation*  

Science.gov (United States)

In platelets, STIM1 has been recognized as the key regulatory protein in store-operated Ca2+ entry (SOCE) with Orai1 as principal Ca2+ entry channel. Both proteins contribute to collagen-dependent arterial thrombosis in mice in vivo. It is unclear whether STIM2 is involved. A key platelet response relying on Ca2+ entry is the surface exposure of phosphatidylserine (PS), which accomplishes platelet procoagulant activity. We studied this response in mouse platelets deficient in STIM1, STIM2, or Orai1. Upon high shear flow of blood over collagen, Stim1?/? and Orai1?/? platelets had greatly impaired glycoprotein (GP) VI-dependent Ca2+ signals, and they were deficient in PS exposure and thrombus formation. In contrast, Stim2?/? platelets reacted normally. Upon blood flow in the presence of thrombin generation and coagulation, Ca2+ signals of Stim1?/? and Orai1?/? platelets were partly reduced, whereas the PS exposure and formation of fibrin-rich thrombi were normalized. Washed Stim1?/? and Orai1?/? platelets were deficient in GPVI-induced PS exposure and prothrombinase activity, but not when thrombin was present as co-agonist. Markedly, SKF96365, a blocker of (receptor-operated) Ca2+ entry, inhibited Ca2+ and procoagulant responses even in Stim1?/? and Orai1?/? platelets. These data show for the first time that: (i) STIM1 and Orai1 jointly contribute to GPVI-induced SOCE, procoagulant activity, and thrombus formation; (ii) a compensating Ca2+ entry pathway is effective in the additional presence of thrombin; (iii) platelets contain two mechanisms of Ca2+ entry and PS exposure, only one relying on STIM1-Orai1 interaction. PMID:20519511

Gilio, Karen; van Kruchten, Roger; Braun, Attila; Berna-Erro, Alejandro; Feijge, Marion A. H.; Stegner, David; van der Meijden, Paola E. J.; Kuijpers, Marijke J. E.; Varga-Szabo, David; Heemskerk, Johan W. M.; Nieswandt, Bernhard

2010-01-01

373

Novel magnetic fibrin hydrogel scaffolds containing thrombin and growth factors conjugated iron oxide nanoparticles for tissue engineering  

Directory of Open Access Journals (Sweden)

Full Text Available Ofra Ziv-Polat1, Hadas Skaat1, Abraham Shahar2, Shlomo Margel11Department of Chemistry, Bar-Ilan Institute of Nanotechnology and Advanced Materials, Ramat-Gan 52900, Israel; 2NVR Research Ltd, Nes-Ziona 74031, IsraelAbstract: Novel tissue-engineered magnetic fibrin hydrogel scaffolds were prepared by the interaction of thrombin-conjugated iron oxide magnetic nanoparticles with fibrinogen. In addition, stabilization of basal fibroblast growth factor (bFGF was achieved by the covalent and physical conjugation of the growth factor to the magnetic nanoparticles. Adult nasal olfactory mucosa (NOM cells were seeded in the transparent fibrin scaffolds in the absence or presence of the free or conjugated bFGF-iron oxide nanoparticles. The conjugated bFGF enhanced significantly the growth and differentiation of the NOM cells in the fibrin scaffolds, compared to the same or even five times higher concentration of the free bFGF. In the presence of the bFGF-conjugated magnetic nanoparticles, the cultured NOM cells proliferated and formed a three-dimensional interconnected network composed mainly of tapered bipolar cells. The magnetic properties of these matrices are due to the integration of the thrombin- and bFGF-conjugated magnetic nanoparticles within the scaffolds. The magnetic properties of these scaffolds may be used in future work for various applications, such as magnetic resonance visualization of the scaffolds after implantation and reloading the scaffolds via magnetic forces with bioactive agents, eg, growth factors bound to the iron oxide magnetic nanoparticles.Keywords: thrombin, fibroblast growth factor, fibrin scaffold, iron oxide nanoparticles, tissue engineering, magnetism, bioactive nanoparticle

Ziv-Polat O

2012-03-01

374

Recombinant protein scaffolds for tissue engineering  

International Nuclear Information System (INIS)

New biological materials for tissue engineering are now being developed using common genetic engineering capabilities to clone and express a variety of genetic elements that allow cost-effective purification and scaffold fabrication from these recombinant proteins, peptides or from chimeric combinations of these. The field is limitless as long as the gene sequences are known. The utility is dependent on the ease, product yield and adaptability of these protein products to the biomedical field. The development of recombinant proteins as scaffolds, while still an emerging technology with respect to commercial products, is scientifically superior to current use of natural materials or synthetic polymer scaffolds, in terms of designing specific structures with desired degrees of biological complexities and motifs. In the field of tissue engineering, next generation scaffolds will be the key to directing appropriate tissue regeneration. The initial period of biodegradable synthetic scaffolds that provided shape and mechanical integrity, but no biological information, is phasing out. The era of protein scaffolds offers distinct advantages, particularly with the combination of powerful tools of molecular biology. These include, for example, the production of human proteins of uniform quality that are free of infectious agents and the ability to make suitable quantities of proteins that are found in low quantity or are hard to isolate from tissue. For the particular needs of from tissue. For the particular needs of tissue engineering scaffolds, fibrous proteins like collagens, elastin, silks and combinations of these offer further advantages of natural well-defined structural scaffolds as well as endless possibilities of controlling functionality by genetic manipulation. (topical review)

375

A novel low-cost and easy to develop functionalization platform. Case study: aptamer-based detection of thrombin by surface plasmon resonance.  

Science.gov (United States)

A novel low-cost platform to assess biomolecular interactions was investigated using surface plasmon resonance and an aptamer-based assay for thrombin detection. Gold SPR surface functionalized with a carboxylated cross-linked BSA film (cBSA) and commercially available carboxymethylated dextran chip (CM5) were used as immobilization platforms for the thrombin binding aptamer. The high end commercial instrument Biacore 3000 and a custom made FIA set-up involving TI Spreeta sensor (TSPR2K23) were used to assess different concentrations of thrombin within the range 0.1-150 nM both in buffer and in a complex matrix (plasma) using the obtained aptasensors. Based on data derived from both CM5 and cBSA platforms, the cBSA aptasensor exhibited good selectivity, stability and regeneration ability, both in buffer and in complex matrices (plasma), comparable with CM5. PMID:20152466

Polonschii, Cristina; David, Sorin; Tombelli, Sara; Mascini, Marco; Gheorghiu, Mihaela

2010-03-15

376

Major research topics in combustion  

Energy Technology Data Exchange (ETDEWEB)

The Institute for Computer Applications in Science and Engineering (ICASE) and NASA Langley Research Center (LaRC) hosted a workshop on October 2--4, 1989 to discuss some combustion problems of technological interest to LaRC and to foster interaction with the academic community in these research areas. The topics chosen for this purpose were flame structure, flame holding/extinction, chemical kinetics, turbulence-kinetics interaction, transition to detonation, and reacting free shear layers. This document contains the papers and edited versions of general discussions on these topics. The lead paper set the stage for the meeting by discussing the status and issues of supersonic combustion relevant to the scramjet engine. Experts were then called upon to review the current knowledge in the aforementioned areas, to focus on how this knowledge can be extended and applied to high-speed combustion, and to suggest future directions of research in these areas.

Hussaini, M.Y.; Kumar, A.; Voigt, R.G. (eds.)

1992-01-01

377

Do scientists trace hot topics?  

Science.gov (United States)

Do scientists follow hot topics in their scientific investigations? In this paper, by performing analysis to papers published in the American Physical Society (APS) Physical Review journals, it is found that papers are more likely to be attracted by hot fields, where the hotness of a field is measured by the number of papers belonging to the field. This indicates that scientists generally do follow hot topics. However, there are qualitative differences among scientists from various countries, among research works regarding different number of authors, different number of affiliations and different number of references. These observations could be valuable for policy makers when deciding research funding and also for individual researchers when searching for scientific projects.

Wei, Tian; Li, Menghui; Wu, Chensheng; Yan, Xiao-Yong; Fan, Ying; di, Zengru; Wu, Jinshan

2013-07-01

378

Do scientists trace hot topics?  

CERN Document Server

Do scientists follow hot topics in their scientific investigations? In this paper, by performing analysis to papers published in the American Physical Society (APS) Physical Review journals, it is found that papers are more likely to be attracted by hot fields, where the hotness of a field is measured by the number of papers belonging to the field. This indicates that scientists generally do follow hot topics. However, there are qualitative differences among scientists from various countries, among research works regarding different number of authors, different number of affiliations and different number of references. These observations could be valuable for policy makers when deciding research funding and also for individual researchers when searching for scientific projects.

Wei, Tian; Wu, Chensheng; Yan, XiaoYong; Fan, Ying; Di, Zengru; Wu, Jinshan

2013-01-01

379

Ethical use of topical corticosteroids.  

Science.gov (United States)

Dermatologists rely very heavily on corticosteroids for treating many common dermatoses. Concerns about their incorrect use are widely expressed both in lay public and specialist discourse. From the point of view of medical ethics, issues of autonomy, beneficence and non-maleficence are all raised frequently when we prescribe topical corticosteroids to our patients. We need to be aware of situations when conflicts between these issues arise and have a clear thought process about resolving them. This can only be achieved if we have a thorough understanding of the skin disease being treated coupled with expertise in the use of the varied potencies and available dosage forms of topical corticosteroids. A good understanding of human psychology and effective communication is also needed to use these agents optimally. PMID:25284852

Saraswat, Abir

2014-09-01

380

In vitro assessment of Tc-99m labeled bovine thrombin and streptokinase-activated human plasmin: concise communication  

International Nuclear Information System (INIS)

Bovine thrombin and streptokinase-activated human plasmin have been labeled with Tc-99m using stannous reduction of pertechnetate under physiological conditions (pH 7.4). The binding efficiency of radiotechnetium to these enzymes is greater than 94%, with less than 5% of reduced but unbound Tc-99m (Sn) complex as assayed by ascending paper radiochromatography using ITLC silica gel plate. Free or unbound pertechnetate is less than 1%. In vitro enzymatic analyses of the Tc-99m-labeled enzymes demonstrate no evidence of protein denaturation or significant loss of enzymatic activity after labeling. Both labeled enzymes are biochemically active in vitro with their respective substrates

 
 
 
 
381

The heparin-binding exosite of factor IXa is a critical regulator of plasma thrombin generation and venous thrombosis  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The role of the factor IXa heparin-binding exosite in coagulation was assessed with mutations that enhance (R170A) or reduce (R233A) stability of the protease-factor VIIIa A2 domain interaction. After tissue factor (TF) addition to reconstituted factor IX-deficient plasma, factor IX R170A supported a 2-fold increase in velocity index (slope) and peak thrombin concentration, whereas factor IX R233A had a 4- to 10-fold reduction relative to factor IX wild-type. In the absence of TF, 5 to 100 pM...

Buyue, Yang; Whinna, Herbert C.; Sheehan, John P.

2008-01-01

382

Mathematics: Essays on Mathematical Topics  

Science.gov (United States)

High school and college students are the target audience of this outstanding math site. The main topics addressed in the nine essays are trigonometry, algebra, and basic calculus. The author does a good job of introducing new concepts and working through examples; however, statements are occasionally made without much justification. Students at the intended level probably do not need detailed proofs for these rare instances, and for the most part, the material is presented in a very organized and comprehensive matter.

383

Hot topics from the Tevatron  

International Nuclear Information System (INIS)

The Tevatron Run-II began in March 2001. To date, both the CDF and D0 experiments have collected 1 fb-1 of data each. The results obtained from this data set were summarized at this conference in 39 parallel session presentations covering a wide range of topics. The author summarizes the most important of those results here and comments on some of the prospects for the future

384

Current topics in shock waves  

International Nuclear Information System (INIS)

The report contains papers on the following topics: shock formation, focusing and implosion; shock reflection and diffraction; turbulence; laser produced plasmas and waves; ionization and shock-plasma interaction; chemical kinetics, pyrolysis, and soot formation; laser-based diagnostics and techniques; experimental facilities, techniques, and applications; ignition of detonation and combustion; particle entrainment and shock propagation through particle suspension; boundary layers and blast simulation; and computational methods and numerical simulation

385

Topics in clinical oncology. 15  

International Nuclear Information System (INIS)

The monograph comprising primarily papers on topical subjects of oncology and cancer research, contains also a selection of papers presented at the 2. Congress of the Czechoslovak Society of Nuclear Medicine and Radiation Hygiene. Seven papers were selected on behalf of their subject related to clinical oncology. All of them were iputted in INIS; five of them deal with the scintiscanning of the skeleton of cancer patients, one with radioimmunodetection of tumors, and one with radionuclide lymphography. (A.K.)

386

Topics on electricity transmission pricing  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The efficiency of the power market is heavily affected by the operation and pricing of the transmission system and the topic of this thesis concerns the interaction of the transmission network and the energy markets. After describing different power flow models, we provide an overview of models developed to efficiently coordinate the allocation of transmission resources. The focus is mainly on short-term efficiency, and the survey is only partial, but provides an integrated overview of some o...

Bjørndal, Mette

2000-01-01

387

Stochastic Analysis and Related Topics  

CERN Document Server

The Silvri Workshop was divided into a short summer school and a working conference, producing lectures and research papers on recent developments in stochastic analysis on Wiener space. The topics treated in the lectures relate to the Malliavin calculus, the Skorohod integral and nonlinear functionals of white noise. Most of the research papers are applications of these subjects. This volume addresses researchers and graduate students in stochastic processes and theoretical physics.

Ustunel, Ali

1988-01-01

388

Hot topics from the Tevatron  

Energy Technology Data Exchange (ETDEWEB)

The Tevatron Run-II began in March 2001. To date, both the CDF and D0 experiments have collected 1 fb{sup -1} of data each. The results obtained from this data set were summarized at this conference in 39 parallel session presentations covering a wide range of topics. The author summarizes the most important of those results here and comments on some of the prospects for the future.

Glenzinski, D.; /Fermilab

2008-01-01

389

Plasma membrane associated phospholipase C from human platelets: Synergistic stimulation of phosphatidylinositol 4,5-bisphosphate hydrolysis by thrombin and guanosine 5'-O-(3-thiotriphosphate)  

International Nuclear Information System (INIS)

The effects of thrombin and GTP?S on the hydrolysis of phosphoinositides by membrane-associated phospholipase C (PLC) from human platelets were examined with endogenous [3H]inositol-labeled membranes or with lipid vesicles containing either [3H]phosphatidylinositol or [3H]phosphatidylinositol 4,5-bisphosphate. GTP?S (1 ?M) or thrombin (1 unit/mL) did not stimulate release of inositol trisphosphate (IP3), inositol bisphosphate (IP2), or inositol phosphate (IP) from [3H]inositol-labeled membranes. IP2 and IP3, but not IP, from [3H]inositol-labeled membranes were, however, stimulated 3-fold by GTP?S (1 ?M) plus thrombin (1 unit/mL). A higher concentration of GTP?S (100 ?M) alone also stimulated IP2 and IP3, but not IP, release. In the presence of 1 mM calcium, release of IP2 and IP3 was increased 6-fold over basal levels; however, formation of IP was not observed. At submicromolar calcium concentration, hydrolysis of exogenous phosphatidylinositol 4,5-bisphosphate (PIP2) by platelet membrane associated PLC was also markedly enhanced by GTP?S (100 ?M) or GTP?S (1 ?M) plus thrombin (1 unit/mL). Under identical conditions, exogenous phosphatidylinositol (PI) was not hydrolyzed. The same substrate specificity was observed when the membrane-associated PLC was activated with 1 mM calcium. Thrombin-induced hydrolysiium. Thrombin-induced hydrolysis of PIP2 was inhibited by treatment of the membranes with pertussis toxin or pretreatment of intact platelets with 12-O-tetradecanoyl-13-acetate (TPA) prior to preparation of membranes. Pertussis toxin did not inhibit GTP?S (100 ?M) or calcium (1 mM) dependent PIP2 breakdown, while TPA inhibited GTP?S-dependent but not calcium-dependent phospholipase C activity

390

Recombining plasmas as laser sources  

International Nuclear Information System (INIS)

The recombination of a dense plasma into a hydrogen-like ion introduces a potential medium for the amplification of radiation over a wide spectral range from 1R to X-Rays. The recombination is a three-body process and hence favours the higher quantum levels. A time dependent calculation shows very large gains for relatively cold temperatures. The time histories of inversion densities and hence the termination of the laser action are controlled by the return of the plasma into equilibrium condition. (Auth.)

391

Isolation and characterization of a serine proteinase with thrombin-like activity from the venom of the snake Bothrops asper  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english A serine proteinase with thrombin-like activity was isolated from the venom of the Central American pit viper Bothrops asper. Isolation was performed by a combination of affinity chromatography on aminobenzamidine-Sepharose and ion-exchange chromatography on DEAE-Sepharose. The enzyme accounts for a [...] pproximately 0.13% of the venom dry weight and has a molecular mass of 32 kDa as determined by SDS-PAGE, and of 27 kDa as determined by MALDI-TOF mass spectrometry. Its partial amino acid sequence shows high identity with snake venom serine proteinases and a complete identity with a cDNA clone previously sequenced from this species. The N-terminal sequence of the enzyme is VIGGDECNINEHRSLVVLFXSSGFL CAGTLVQDEWVLTAANCDSKNFQ. The enzyme induces clotting of plasma (minimum coagulant dose = 4.1 µg) and fibrinogen (minimum coagulant dose = 4.2 µg) in vitro, and promotes defibrin(ogen)ation in vivo (minimum defibrin(ogen)ating dose = 1.0 µg). In addition, when injected intravenously in mice at doses of 5 and 10 µg, it induces a series of behavioral changes, i.e., loss of the righting reflex, opisthotonus, and intermittent rotations over the long axis of the body, which closely resemble the `gyroxin-like' effect induced by other thrombin-like enzymes from snake venoms.

A.V, Pérez; A, Rucavado; L, Sanz; J.J, Calvete; J.M, Gutiérrez.

392

Isolation and characterization of a serine proteinase with thrombin-like activity from the venom of the snake Bothrops asper  

Directory of Open Access Journals (Sweden)

Full Text Available A serine proteinase with thrombin-like activity was isolated from the venom of the Central American pit viper Bothrops asper. Isolation was performed by a combination of affinity chromatography on aminobenzamidine-Sepharose and ion-exchange chromatography on DEAE-Sepharose. The enzyme accounts for approximately 0.13% of the venom dry weight and has a molecular mass of 32 kDa as determined by SDS-PAGE, and of 27 kDa as determined by MALDI-TOF mass spectrometry. Its partial amino acid sequence shows high identity with snake venom serine proteinases and a complete identity with a cDNA clone previously sequenced from this species. The N-terminal sequence of the enzyme is VIGGDECNINEHRSLVVLFXSSGFL CAGTLVQDEWVLTAANCDSKNFQ. The enzyme induces clotting of plasma (minimum coagulant dose = 4.1 µg and fibrinogen (minimum coagulant dose = 4.2 µg in vitro, and promotes defibrin(ogenation in vivo (minimum defibrin(ogenating dose = 1.0 µg. In addition, when injected intravenously in mice at doses of 5 and 10 µg, it induces a series of behavioral changes, i.e., loss of the righting reflex, opisthotonus, and intermittent rotations over the long axis of the body, which closely resemble the `gyroxin-like' effect induced by other thrombin-like enzymes from snake venoms.

A.V Pérez

2008-01-01

393

3D photonic crystal-based biosensor functionalized with quantum dot-based aptamer for thrombine detection  

Science.gov (United States)

In this paper, we propose a new technique for protein detection by using the enhancement of intensity in quantum dots (Qdot) whose emission is guided by 3D photonic crystal (PC) structures. For easy to use, we design the emitted light from the sensor can be recovered, when the chemical antibody (aptamer) conjugated with guard DNA (g-DNA) labeled with a quencher (Black FQ) hybridizes with the target proteins. In detail, we synthesis a Qdot-aptamer complex and then immobilize these complex on the PC surfaces. Next, we perform the hybridization of the Qdot-aptamer complex with g-DNA labeled with the quencher. It induces the quenching effect of fluoresce intensity in the Qdot-aptamer. In presence of target protein (thrombin), the Qdot-aptamer complex prefers to form the thrombin-aptamer complex: this results in the release of Black FQ-g-DNA and the quenched light intensity recovers into the original high intensity with Qdot. The intensity recovery varies quantitatively according to the level of the target protein concentration. This proposed sensor shows much higher detection sensitivity than the general fluorescent detection mechanism, which is functionalized on the flat surfaces because of the light guiding effect from 3D photonic crystal structures.