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Sample records for thrombin topical recombinant

  1. Topical thrombin preparations and their use in cardiac surgery

    Directory of Open Access Journals (Sweden)

    Brianne L Dunn

    2009-10-01

    Full Text Available Brianne L Dunn1, Walter E Uber1, John S Ikonomidis21Department of Pharmacy Services and 2Division of Cardiothoracic Surgery, Medical University of South Carolina, Charleston, South Carolina, USAAbstract: Coagulopathic bleeding may lead to increased morbidity and mortality after cardiac surgery. Topical bovine thrombin has been used to promote hemostasis after surgical procedures for over 60 years and is used frequently as a topical hemostatic agent in cardiac surgery. Recently, use of bovine thrombin has been reported to be associated with increased risk for anaphylaxis, thrombosis, and immune-mediated coagulopathy thought secondary to the production of antifactor V and antithrombin antibodies. In patients who develop bovine thrombin-induced immune-mediated coagulopathy, clinical manifestations may range from asymptomatic alterations in coagulation tests to severe hemorrhage and death. Patients undergoing cardiac surgical procedures may be at increased risk for development of antibodies to bovine thrombin products and associated complications. This adverse immunologic profile has led to the development of alternative preparations including a human and a recombinant thrombin which have been shown to be equally efficacious to bovine thrombin and have reduced antigenicity. However, the potential benefit associated with reduced antigenicity is not truly known secondary to the lack of long-term experience with these products. Given the potentially higher margin of safety and less stringent storage concerns compared to human thrombin, recombinant thrombin may be the most reasonable approach in cardiac surgery.Keywords: bovine thrombin, human thrombin, recombinant thrombin, immune-mediated coagulopathy, topical hemostatic agents, thrombin 

  2. Clinical use of topical thrombin as a surgical hemostat

    Directory of Open Access Journals (Sweden)

    Wesley K Lew

    2008-08-01

    Full Text Available Wesley K Lew1, Fred A Weaver21University of Southern California, Department of Surgery, Los Angeles, CA, USA; 2CardioVascular Thoracic Institute at the University of Southern California, Los Angeles, CA, USAAbstract: When surgical ligation of bleeding fails, or is not possible, surgeons rely on a number of hemostatic aids, including thrombin. This review discusses the history, pharmacology and clinical application of thrombin as a surgical hemostat. The initial thrombin was bovine in origin, but its use has been complicated by the formation of antibodies that cross react with human coagulation factors. This has been associated with life threatening bleeding and in some circumstances anaphylaxis and death. Human thrombin, isolated from pooled plasma of donors, has been developed in an effort to minimize these risks, but its downside is the potential of transmitting blood-borne pathogens and limited availability. Recently a recombinant thrombin has been developed and approved for use by the FDA. It has the advantage of being minimally antigenic and devoid of the risk if viral transmission. Thrombin is often used in conjunction with other hemostatic aids, including absorbable agents (like gelfoam, collagen, and cellulose, and with fibrinogen in fibrin glues. The last part of this review will discuss these agents in detail, and review their clinical applications.Keywords: bovine, recombinant, human, thrombin, antigenicity, antibodies

  3. Development of gel-forming lyophilized formulation with recombinant human thrombin.

    Science.gov (United States)

    Murányi, Andrej; Bartoš, Peter; Tichý, Eduard; Lazová, Jana; Pšenková, Jana; Žabka, Marián

    2015-01-01

    The objective of this work was development and evaluation of gel-forming lyophilized formulation with recombinant human thrombin for topical administration. The influence of pH, ionic strength and buffer type on protein stability was evaluated as part of the pre-formulation screening studies. Results indicated an optimal pH from 6.0 to 7.0 and increased stability with increasing content of sodium chloride. The tested buffer types had no significant effect on thrombin stability. For further development, thermosensitive Pluronic® F-127 was employed as a bulking and gelling agent. Physical and mechanical characterization and viscosity measurement confirmed the gel-forming properties of the formulation at the application temperature of 32 °C. Several techniques (addition of well-soluble polyols, different freezing protocols and reconstitution under vacuum) were tested to decrease the reconstitution time. The obtained results revealed that a vacuum in the vial headspace is crucial for acceptable reconstitution. The freeze drying process has no negative impact on recombinant thrombin stability, and this was confirmed by reverse-phase-HPLC, activity assay and optical density measurements.

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

    Science.gov (United States)

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

    2004-07-30

    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

  5. Recombinant DNA Technology. A Topics Course for Undergraduates.

    Science.gov (United States)

    Parson, Kathleen A.

    1988-01-01

    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)

  6. Topics in cosmology: Structure formation, dark energy and recombination

    Science.gov (United States)

    Alizadeh, Esfandiar

    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

  7. Topics in cosmology: Structure formation, dark energy and recombination

    Science.gov (United States)

    Alizadeh, Esfandiar

    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

  8. Thrombin during cardiopulmonary bypass.

    Science.gov (United States)

    Edmunds, L Henry; Colman, Robert W

    2006-12-01

    Cardiopulmonary bypass (CPB) ignites a massive defense reaction that stimulates all blood cells and five plasma protein systems to produce a myriad of vasoactive and cytotoxic substances, cell-signaling molecules, and upregulated cellular receptors. Thrombin is the key enzyme in the thrombotic portion of the defense reaction and is only partially suppressed by heparin. During CPB, thrombin is produced by both extrinsic and intrinsic coagulation pathways and activated platelets. The routine use of a cell saver and the eventual introduction of direct thrombin inhibitors now offer the possibility of completely suppressing thrombin production and fibrinolysis during cardiac surgery with CPB. PMID:17126170

  9. Single amino acid substitutions dissociate fibrinogen-clotting and thrombomodulin-binding activities of human thrombin.

    OpenAIRE

    Wu, Q Y; Sheehan, J P; Tsiang, M; Lentz, S R; Birktoft, J J; Sadler, J E

    1991-01-01

    Thrombin is a serine protease that acts as a procoagulant by clotting fibrinogen and activating platelets and as an anticoagulant by activating protein C in a thrombomodulin-dependent reaction. Fibrinogen and thrombomodulin bind competitively to an anion-binding exosite on thrombin. We prepared recombinant normal human thrombin and mutant thrombins with single amino acid substitutions in order to localize and distinguish the fibrinogen- and thrombomodulin-binding sites. Normal and mutant thro...

  10. The use of thrombin in the radiology department.

    LENUS (Irish Health Repository)

    Ward, E

    2009-03-01

    Thrombin is a naturally occurring coagulation protein that converts soluble fibrinogen into insoluble fibrin and plays a vital role in the coagulation cascade and in turn haemostasis. Thrombin also promotes platelet activation. In the last few years, there has been a rapid increase in the use of thrombin by radiologists in a variety of clinical circumstances. It is best known for its use in the treatment of pseudoaneurysms following angiography. However, there are now a variety of cases in the literature describing the treatment of traumatic, inflammatory and infected aneurysms with thrombin in a variety of locations within the human body. There have even been recent reports describing the use of thrombin in conventional aneurysms as well as ruptured aneurysms. Its use has also been described in the treatment of endoleaks (type II) following aneurysm repair. In nearly all of these cases, treatment with thrombin requires imaging guidance. Recently, thrombin has also been used as a topical treatment post-percutaneous intervention to reduce or stop bleeding. Most radiologists have only a limited knowledge of the pharmacodynamics of thrombin, its wide range of utilisation and its limitations. Apart from a few case reports and case series, there is little in the radiological literature encompassing the wide range of applications that thrombin may have in the radiology department. In this review article, we comprehensively describe the role and pathophysiology of thrombin, describing with examples many of its potential uses. Techniques of usage as well as pitfalls and limitations are also described.

  11. "Mirror image" antagonists of thrombin-induced platelet activation based on thrombin receptor structure.

    OpenAIRE

    Hung, D. T.; Vu, T K; Wheaton, V I; Charo, I F; Nelken, N A; Esmon, N; Esmon, C T; Coughlin, S R

    1992-01-01

    Platelet activation by thrombin plays a critical role in hemostasis and thrombosis. Based on structure-activity studies of a cloned platelet thrombin receptor, we designed two "mirror image" antagonists of thrombin and thrombin receptor function. First, "uncleavable" peptides mimicking the receptor domain postulated to interact with thrombin were found to be potent thrombin inhibitors. Second, proteolytically inactive mutant thrombins designed to bind but not cleave the thrombin receptor were...

  12. Thrombin Inhibitors from Different Animals

    OpenAIRE

    Tanaka-Azevedo, A. M.; Morais-Zani, K.; Torquato, R. J. S.; A. S. Tanaka

    2010-01-01

    Venous and arterial thromboembolic diseases are still the most frequent causes of death and disability in high-income countries. Clinical anticoagulants are inhibitors of enzymes involved in the coagulation pathway, such as thrombin and factor Xa. Thrombin is a key enzyme of blood coagulation system, activating the platelets, converting the fibrinogen to the fibrin net, and amplifying its self-generation by the activation of factors V, VIII, and XI. Thrombin has long been a target for the dev...

  13. Recombinant snake venom prothrombin activators

    OpenAIRE

    Lövgren, Ann

    2012-01-01

    Three prothrombin activators; ecarin, which was originally isolated from the venom of the saw-scaled viper Echis carinatus, trocarin from the rough-scaled snake Tropidechis carinatus, and oscutarin from the Taipan snake Oxyuranus scutellatus, were expressed in mammalian cells with the purpose to obtain recombinant prothrombin activators that could be used to convert prothrombin to thrombin. We have previously reported that recombinant ecarin can efficiently generate thrombin without the need ...

  14. Ximelagatran: direct thrombin inhibitor

    Directory of Open Access Journals (Sweden)

    Shir-Jing Ho

    2006-03-01

    Full Text Available Shir-Jing Ho1, Tim A Brighton2,31St George Hospital, Sydney, NSW, Australia; 2University of New South Wales, Sydney, Australia; 3SEALS (Randwick, Prince of Wales Hospital, Sydney, AustraliaAbstract: Warfarin sodium is an effective oral anticoagulant drug. However, warfarin has a narrow therapeutic window with significant risks of hemorrhage at therapeutic concentrations. Dosing is difficult and requires frequent monitoring. New oral anticoagulant agents are required to improve current anticoagulant therapy. Furthermore, while warfarin is effective in venous disease, it does not provide more than 60% risk reduction compared with placebo in venous thrombosis prophylaxis and considerably lower risk reduction in terms of arterial thrombosis. Ximelagatran is an oral pro-drug of melagatran, a synthetic small peptidomimetic with direct thrombin inhibitory actions and anticoagulant activity. As an oral agent, ximelagatran has a number of desirable properties including a rapid onset of action, fixed dosing, stable absorption, apparent low potential for medication interactions, and no requirement for monitoring of drug levels or dose adjustment. It has a short plasma elimination half-life of about 4 hours in cases of unexpected hemorrhage or need for reversal. Its main toxicity relates to the development of abnormal liver biochemistry and/or liver dysfunction with “long-term” use of the drug. This usually occurs within the first 6 months of commencing therapy, with a small percentage of patients developing jaundice. The biochemical abnormality usually resolves despite continuation of the drug. The cause of this toxicity remains unknown. Clinical studies to date have shown that ximelagatran is noninferior to warfarin in stroke prevention in patients with nonvalvular atrial fibrillation, noninferior to standard therapy as acute and extended therapy of deep vein thrombosis (DVT, and superior to warfarin for the prevention of venous thromboembolism post

  15. [Monitoring of Oral Thrombin Inhibitor].

    Science.gov (United States)

    Matsuno, Kazuhiko; Usami, Takayuki; Hatuse, Masanao; Shimizu, Chikara

    2014-10-01

    Dabigatran etexilate is a prodrug that is converted into its active metabolite, dabigatran, by hydrolysis. Dabigatran is a selective thrombin inhibitor that has been approved for the prevention of stroke in patients with non-valvular atrial fibrillation in Japan. Laboratory monitoring is not needed, but an assessment of its anticoagulant effect in certain clinical settings, such as emergency surgery, suspected overdose, or the occurrence of bleeding, is desirable. We overview the special coagulation assays, such as Hemoclot Thrombin Inhibitor (HTI), the thrombin generation assay (TGA), ecarin clotting time (ECT), ecarin chromogenic assay (ECA), prothrombinase-induced clotting time (PiCT), and activated clotting time (ACT). We also examined the relationship between dabigatran levels as determined by HTI, and the activated partial thromboplastin time (APTT) and prothrombin time (PT). APTT and PT demonstrated a positive correlation with the dabigatran levels. APTT, PT, and the combination of APTT and PT may estimate the dabigatran levels in plasma. PMID:27526541

  16. Aptamer Based Microsphere Biosensor for Thrombin Detection

    OpenAIRE

    Xudong Fan; White, Ian M.; Suter, Jonathan D.; Hongying Zhu

    2006-01-01

    We have developed an optical microsphere resonator biosensor using aptamer as receptor for the measurement of the important biomolecule thrombin. The sphere surface is modified with anti-thrombin aptamer, which has excellent binding affinity and selectivity for thrombin. Binding of the thrombin at the sphere surface is monitored by the spectral position of the microsphere's whispering gallery mode resonances. A detection limit on the order of 1 NIH Unit/mL is demonstrated. Control experiments...

  17. Thrombin interaction with fibrin polymerization sites.

    Science.gov (United States)

    Hsieh, K

    1997-05-15

    Thrombin is central to hemostasis, and postclotting fibrinolysis and wound healing. During clotting, thrombin transforms plasma fibrinogen into polymerizing fibrin, which selectively adsorbs the enzyme into the clot. This protects thrombin from heparin-antithrombin inactivation, thus preserving the enzyme for postclotting events. To determine how the fibrin N-terminal polymerization sites of A alpha 17-23 (GPRVVER) and B beta 15-25 (GHRPLDKKREE) and their analogs may interact with thrombin, amidolysis vs. plasma- and fibrinogen-clotting assays were used to differentiate blockade of catalytic site vs. other thrombin domains. Amidolysis studies suggest GPRVVER inhibition of thrombin catalytic site through hydrophobic interaction, and GPRVVER inhibited clotting. Neither GPRP nor VVER nor the B beta 15-25 homologs inhibited amidolysis. Contrary to heparin, acyl-DKKREE promoted plasma-clotting, but inhibited fibrinogen-clotting. In addition, acyl-DKKREE reversed the anticoagulant effect of heparin (0.1 U/ml) in plasma. The results suggest fibrin B beta 15-25 interaction with thrombin, possibly by blocking the heparin-binding site. Together with the reported fibrin A alpha 27-50 binding to thrombin, polymerizing fibrin appears to initially bind to thrombin catalytic site and exosite-1 through A alpha 17-50, and to another thrombin site through B beta 15-25. As these fibrin sites are also involved in polymerization, competition of the polymerization process with thrombin-binding could subsequently dislodge thrombin from fibrin alpha-chain. This may re-expose the catalytic site and exosite-1, thus explaining the thrombogenicity of clot-bound thrombin. The implications of these findings in polymerization mechanism and anticoagulant design are discussed.

  18. PAR1-dependent and independent increases in COX-2 and PGE2 in human colonic myofibroblasts stimulated by thrombin.

    Science.gov (United States)

    Seymour, Michelle L; Zaidi, Nosheen F; Hollenberg, Morley D; MacNaughton, Wallace K

    2003-05-01

    Subepithelial myofibroblast-derived prostaglandin E(2) (PGE(2)) regulates epithelial chloride secretion in the intestine. Thrombin is elevated in inflammatory conditions of the bowel. Therefore, we sought to determine a role for thrombin in regulating PGE(2) synthesis by colonic myofibroblasts. Incubation of cultured CCD-18Co colonic myofibroblasts with thrombin, the proteinase-activated receptor 1 (PAR(1))-activating peptide (Cit-NH(2)), and peptides corresponding to 2 noncatalytic regions of thrombin (TP367 and TP508) for 18 h increased both cyclooxygenase (COX)-2 expression (immunocytochemistry) and PGE(2) synthesis (enzyme immunoassay). Inhibition of thrombin by D-Phe-Pro-Arg-chloromethylketone (PPACK) did not significantly reduce PGE(2) synthesis, which remained elevated compared with control. We also investigated the basic fibroblast growth factor (bFGF) dependence of thrombin-induced PGE(2) elevations. Recombinant human bFGF concentration dependently increased PGE(2) synthesis, and a bFGF neutralizing antibody inhibited PGE(2) synthesis induced by TP367 and TP508 (approximately 40%) and by thrombin (approximately 20%) (but not Cit-NH(2)). Thrombin, therefore, upregulates COX-2-derived PGE(2) synthesis by both catalytic cleavage of PAR(1) and bFGF-dependent noncatalytic activity. This presents a novel mechanism by which intestinal myofibroblasts might regulate epithelial chloride secretion. PMID:12505789

  19. Recombinant snake venom prothrombin activators.

    Science.gov (United States)

    Lövgren, Ann

    2013-01-01

    Three prothrombin activators; ecarin, which was originally isolated from the venom of the saw-scaled viper Echis carinatus, trocarin from the rough-scaled snake Tropidechis carinatus, and oscutarin from the Taipan snake Oxyuranus scutellatus, were expressed in mammalian cells with the purpose to obtain recombinant prothrombin activators that could be used to convert prothrombin to thrombin. We have previously reported that recombinant ecarin can efficiently generate thrombin without the need for additional cofactors, but does not discriminate non-carboxylated prothrombin from biologically active γ-carboxylated prothrombin. Here we report that recombinant trocarin and oscutarin could not efficiently generate thrombin without additional protein co-factors. We confirm that both trocarin and oscutarin are similar to human coagulation Factor X (FX), explaining the need for additional cofactors. Sequencing of a genomic fragment containing 7 out of the 8 exons coding for oscutarin further confirmed the similarity to human FX. PMID:23111318

  20. Investigation of the selectivity of thrombin-binding aptamers for thrombin titration in murine plasma.

    Science.gov (United States)

    Trapaidze, Ana; Hérault, Jean-Pascal; Herbert, Jean-Marc; Bancaud, Aurélien; Gué, Anne-Marie

    2016-04-15

    Detection of thrombin in plasma raises timely challenges to enable therapeutic management of thrombosis in patients under vital threat. Thrombin binding aptamers represent promising candidates as sensing elements for the development of real-time thrombin biosensors; however implementation of such biosensor requires the clear understanding of thrombin-aptamer interaction properties in real-like environment. In this study, we used Surface Plasmon Resonance technique to answer the questions of specificity and sensitivity of thrombin detection by the thrombin-binding aptamers HD1, NU172 and HD22. We systematically characterized their properties in the presence of thrombin, as well as interfering molecular species such as the thrombin precursor prothrombin, thrombin in complex with some of its natural inhibitors, nonspecific serum proteins, and diluted plasma. Kinetic experiments show the multiple binding modes of HD1 and NU172, which both interact with multiple sites of thrombin with low nanomolar affinities and show little specificity of interaction for prothrombin vs. thrombin. HD22, on the other hand, binds specifically to thrombin exosite II and has no affinity to prothrombin at all. While thrombin in complex with some of its inhibitors could not be recognized by any aptamer, the binding of HD1 and NU172 properties is compromised by thrombin inhibitors alone, as well as with serum albumin. Finally, the complex nature of plasma was overwhelming for HD1, but we define conditions for the thrombin detection at 10nM range in 100-fold diluted plasma by HD22. Consequently HD22 showed key advantage over HD1 and NU172, and appears as the only alternative to design an aptasensor.

  1. Inhibitory effect of apixaban compared with rivaroxaban and dabigatran on thrombin generation assay.

    Science.gov (United States)

    Wong, Pancras C; White, Andrew; Luettgen, Joseph

    2013-02-01

    The effect of the oral direct activated factor X (factor Xa) inhibitor apixaban on tissue factor-induced thrombin generation in human plasma was investigated in vitro using the calibrated automated thrombogram (CAT) method and compared with the oral direct factor Xa inhibitor rivaroxaban and the direct thrombin inhibitor dabigatran. Pooled citrated, anticoagulated, platelet-poor human plasma was spiked with apixaban, rivaroxaban, or dabigatran at concentrations of 0.01 to 10 μM. The inhibitory potencies of the compounds were quantified by 5 CAT parameters: the control thrombin lag time (LT) and time to thrombin peak (TTP) for the doubling of inhibitor concentration (IC2x); and the control endogenous thrombin potential (ETP), thrombin peak, and maximum rate of thrombin generation (Vmax) for the inhibitor concentration, which inhibited 50% (IC50). The inhibitors modified CAT concentration dependently. Their inhibitory potencies, expressed as IC2x LT, IC2x TTP, IC50 ETP, IC50 peak thrombin, and IC50 Vmax, were as follows: 0.10 ± 0.01, 0.19 ± 0.02, 0.65 ± 0.11, 0.089 ± 0.019, and 0.049 ± 0.007 μM for apixaban; 0.049 ± 0.007, 0.070 ± 0.009, 0.43 ± 0.07, 0.048 ± 0.008, and 0.022 ± 0.005 μM for rivaroxaban; and 0.063 ± 0.019, 0.18 ± 0.06, 0.50 ± 0.08, 0.55 ± 0.06, and 0.57 ± 0.27 μM for dabigatran. In summary, apixaban, rivaroxaban, and dabigatran have similar potencies in the prolongation of LT and TTP. The CAT parameters that are related to the rate of thrombin generation during the propagation phase (ie, peak thrombin and Vmax) are more sensitive to activities of apixaban and rivaroxaban than dabigatran. The ETP is the least sensitive parameter for measuring the activities of these inhibitors. Recombinant activated factor VII at 5 and 50 μg/mL reversed the anticoagulant effects of apixaban more at 0.2 μM than at 2 μM. Our study suggests that the CAT method is a sensitive assay to monitor the pharmacodynamic and pharmacokinetic properties of

  2. DABIGATRAN ETEXILATE: NEW DIRECT THROMBIN INHIBITORS ANTICOAGULANTS

    Directory of Open Access Journals (Sweden)

    Patel Kinjal B

    2011-04-01

    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.

  3. [Direct oral thrombin inhibitor, "dabigatran"].

    Science.gov (United States)

    Yasaka, Masahiro

    2013-01-01

    Dabigatran is an oral, direct, and competitive inhibitor of thrombin, which is administered to patients with non-valvular atrial fibrillation for prevention of stroke at a dose of 110 mg twice daily or 150 mg twice daily. Anticoagulation by dabigatran is "hybrid anticoagulation", consisting of action of both dabigatran and physiological coagulation inhibitors because warfarin inhibits production of protein C and protein S but dabigatran does not. Management of dabigatran is easier than that of warfarin because food restriction is unnecessary, drug interaction is small, and absorption time is short and serum concentration corresponds to the anticoagulatory effect in dabigatran treatment. The RE-LY trial confirmed effectiveness and safety of both doses of dabigatran for prevention of stroke and both doses of dabigatran had much lower risks of intracranial bleeding compared with warfarin. Compliance to guidance of dabigatran treatment is essential for avoidance of severe hemorrhagic complications. PMID:23631181

  4. Skin regeneration in deep second-degree scald injuries either by infusion pumping or topical application of recombinant human erythropoietin gel

    Directory of Open Access Journals (Sweden)

    Giri P

    2015-05-01

    Full Text Available Priya Giri,1 Sabine Ebert,1 Ulf-Dietrich Braumann,2 Mathias Kremer,3 Shibashish Giri,1 Hans-Günther Machens,4 Augustinus Bader1 1Department of Cell Techniques and Applied Stem Cell Biology, Center for Biotechnology and Biomedicine (BBZ, Faculty of Medicine, University of Leipzig, Leipzig, Germany; 2Interdisciplinary Center for Bioinformatics (IZBI, University of Leipzig, Leipzig, Germany; 3Department of Plastic and Hand Surgery, University of Lübeck, Lübeck, Germany; 4Department of Plastic and Hand Surgery, Technical University of Munich, Munich, Germany Abstract: Large doses of recombinant growth factors formulated in solution form directly injected into the body is usual clinical practice in treating second-degree scald injuries, with promising results, but this approach creates side effects; furthermore, it may not allow appropriate levels of the factor to be sensed by the target injured tissue/organ in the specific time frame, owing to complications arising from regeneration. In this research, two delivery methods (infusion pumping and local topical application were applied to deliver recombinant human erythropoietin (rHuEPO for skin regeneration. First, rHuEPO was given in deep second-degree scald injury sites in mice by infusion pump. Vascularization was remarkably higher in the rHuEPO pumping group than in controls. Second, local topical application of rHuEPO gel was given in deep second-degree scald injury sites in rats. Histological analysis showed that epithelialization rate was significantly higher in the rHuEPO gel-treated group than in controls. Immunohistochemical studies showed that the rHuEPO gel-treated group showed remarkably higher expression of skin regeneration makers than the control group. An accurate method for visualization and quantification of blood vessel networks in target areas has still not been developed up to this point, because of technical difficulties in detecting such thin blood vessels. A method which

  5. Endogenous thrombin potential in polycystic ovary syndrome

    DEFF Research Database (Denmark)

    Aziz, Mubeena; Sidelmann, Johannes J; Wissing, Marie Louise Muff;

    2015-01-01

    OBJECTIVES: The objective of this study is to investigate plasma endogenous thrombin generation in four different phenotypes of polycystic ovary syndrome (PCOS) defined by Body Mass Index (BMI) and insulin resistance (IR). PCOS is diagnosed according to the Rotterdam criteria. DESIGN: Multicenter...... cross-sectional study. SETTING: Two major University Hospitals in the Capital region of Denmark. PATIENTS: Hundred forty-eight European women with PCOS were consecutively recruited during April 2010-February 2012. Clinical examination, blood sampling, and DEXA scan were performed. MAIN OUTCOME MEASURES......: Endogenous thrombin potential (ETP). RESULTS: PCOS women with phenotype BMI > 25 + IR have increased potential of thrombin generation. ETP is associated with total body fat mass, IR, and CRP. CONCLUSIONS: Obese and insulin resistant women with PCOS have elevated level of ETP corresponding to increased risk...

  6. Thrombin Hydrolysis of Human Osteopontin Is Dependent on Thrombin Anion-binding Exosites*

    OpenAIRE

    Myles, Timothy; Leung, Lawrence L. K.

    2008-01-01

    The cytokine osteopontin (OPN) can be hydrolyzed by thrombin exposing a cryptic α4β1/α9β1 integrin-binding motif (SVVYGLR), thereby acting as a potent cytokine for cells bearing these activated integrins. We show that purified milk OPN is a substrate for thrombin with a kcat/Km value of 1.14 × 105 m–1 s–1. Thrombin cleavage of OPN was inhibited by unsulfated hirugen (IC50 = 1.2 ± 0.2 μm), unfractionated heparin (IC50 = 56.6 ± 8.4 μg/ml) and low molecular ...

  7. Discovery of thrombin activatable fibrinolysis inhibitor (TAFI)

    NARCIS (Netherlands)

    Bertina, R.M.; Tilburg, N.H. van; Haverkate, F.; Bouma, B.N.; Borne, P.A.K. von dem; Meijers, J.C.M.; Campbell, W.; Eaton, D.; Hendriks, D.F.; Willemse, J.L.

    2006-01-01

    CAS: blood clotting factor 11, 9013-55-2; thrombin, 9002-04-4; tissue plasminogen activator, 105913-11-9; protein C, 60202-16-6; Carboxypeptidase U, 3.4.17.20; Protein C; Tissue Plasminogen Activator, 3.4.21.68

  8. Thrombin-Mediated Direct Activation of Proteinase-Activated Receptor-2: Another Target for Thrombin Signaling.

    Science.gov (United States)

    Mihara, Koichiro; Ramachandran, Rithwik; Saifeddine, Mahmoud; Hansen, Kristina K; Renaux, Bernard; Polley, Danny; Gibson, Stacy; Vanderboor, Christina; Hollenberg, Morley D

    2016-05-01

    Thrombin is known to signal to cells by cleaving/activating a G-protein-coupled family of proteinase-activated receptors (PARs). The signaling mechanism involves the proteolytic unmasking of an N-terminal receptor sequence that acts as a tethered receptor-activating ligand. To date, the recognized targets of thrombin cleavage and activation for signaling are PAR1 and PAR4, in which thrombin cleaves at a conserved target arginine to reveal a tethered ligand. PAR2, which like PAR1 is also cleaved at an N-terminal arginine to unmask its tethered ligand, is generally regarded as a target for trypsin but not for thrombin signaling. We now show that thrombin, at concentrations that can be achieved at sites of acute injury or in a tumor microenvironment, can directly activate PAR2 vasorelaxation and signaling, stimulating calcium and mitogen-activated protein kinase responses along with triggeringβ-arrestin recruitment. Thus, PAR2 can be added alongside PAR1 and PAR4 to the targets, whereby thrombin can affect tissue function.

  9. Amino acid residues of heparin cofactor II required for stimulation of thrombin inhibition by sulphated polyanions.

    Science.gov (United States)

    Colwell, N S; Grupe, M J; Tollefsen, D M

    1999-04-12

    A variety of sulphated polyanions in addition to heparin and dermatan sulphate stimulate the inhibition of thrombin by heparin cofactor II (HCII). Previous investigations indicated that the binding sites on HCII for heparin and dermatan sulphate overlap but are not identical. In this study we determined the concentrations (IC50) of various polyanions required to stimulate thrombin inhibition by native recombinant HCII in comparison with three recombinant HCII variants having decreased affinity for heparin (Lys-173-->Gln), dermatan sulphate (Arg-189-->His), or both heparin and dermatan sulphate (Lys-185-->Asn). Pentosan polysulphate, sulphated bis-lactobionic acid amide, and sulphated bis-maltobionic acid amide resembled dermatan sulphate, since their IC50 values were increased to a much greater degree (>/=8-fold) by the mutations Arg-189-->His and Lys-185-->Asn than by Lys-173-->Gln (Gln and Lys-185-->Asn (>/=6-fold) than by Arg-189-->His (thrombin by an N-terminal deletion mutant of HCII (Delta1-74). These results suggest that, like dermatan sulphate and heparin, other polyanions stimulate HCII primarily by an allosteric mechanism requiring the N-terminal acidic domain. PMID:10209287

  10. Heparin enhances the catalytic activity of des-ETW-thrombin.

    Science.gov (United States)

    Goodwin, C A; Deadman, J J; Le Bonniec, B F; Elgendy, S; Kakkar, V V; Scully, M F

    1996-04-01

    The thrombin mutant, des-ETW-thrombin, lacking Glu(146), Thr(147), and Trp(148) within a unique insertion loop located at the extreme end of the primary specificity pocket, has been shown previously to exhibit reduced catalytic activity with respect to macromolecular and synthetic thrombin substrates and reduced or enhanced susceptibility to inhibition. Investigation of the hydrolysis of peptidyl p-nitroanilide substrates by des-ETW-thrombin showed increased activity in the presence of heparin and other sulphated glycosaminoglycans. No effect was observed upon the activity of wild-type thrombin. Heparin was found to decrease the K(m) for cleavage of four thrombin-specific substrates by des-ETW-thrombin by 3-4-fold. Similarly, pentosan polysulphate (PPS) decreased the K(m) with these substrates by 8-10-fold. Heparin also increased the rate of inhibition of des-ETW-thrombin by antithrombin III and D-phenylalanyl-prolyl-arginylchloromethane (PPACK). The inhibition of des-ETW-thrombin by a number of thrombin-specific peptide boronic acids also showed significant reduction in the final K(i) in the presence of heparin, due to reduction in the off-rate. A peptide analogue of a sequence of hirudin which binds thrombin tightly to exosite I (fibrinogen recognition site) potentiated the activity of des-ETW-thrombin against peptide p-nitroanilide substrates in a manner similar to heparin. The K(i) for the inhibition of des-ETW-thrombin by p-aminobenzamidine was decreased by these ligands from 9.7 mM to 7.5 mM, 5.1 mM, and 2.5 mM in the presence of heparin, hirudin peptide and PPS respectively, suggesting the increased catalytic activity is due to enhanced access to the primary specificity pocket. The positive influence of these ligands on des-ETW-thrombin was reversed in the presence of ATP or ADP; the latter has previously been shown to inhibit thrombin activity by blocking initial interaction with fibrinogen at exosite 1. Because the effect of heparin and PPS is similar to

  11. Crystal structure of two new bifunctional nonsubstrate type thrombin inhibitors complexed with human alpha-thrombin.

    Science.gov (United States)

    Féthière, J.; Tsuda, Y.; Coulombe, R.; Konishi, Y.; Cygler, M.

    1996-01-01

    The crystal structures of two new thrombin inhibitors, P498 and P500, complexed with human alpha-thrombin have been determined at 2.0 A resolution and refined to crystallographic R-factors of 0.170 and 0.169, respectively. These compounds, with picomolar binding constants, belong to a family of potent bifunctional inhibitors that bind thrombin at two remote sites: the active site and the fibrinogen recognition exosite (FRE). The inhibitors incorporate a nonsubstrate type active site binding fragment: Dansyl-Arg-(D)Pipecolic acid (Dns-Arg-(D)Pip), reminiscent of the active-site directed inhibitors MD-805 and MQPA, rendering them resistant to thrombin-induced hydrolysis. The FRE binding fragment of these inhibitors corresponds to the hirudin55-65 sequence. They differ in the chemical nature of the nonpeptidyl linker bridging these two functional activities. In both cases, the active site binding fragment is well defined in the electron density. The DnsH1, ArgH2, and (D)PipH3 groups occupy the S3, S1, and S2 subsites of thrombin, respectively, in a way similar to that observed in the thrombin-MQPA complexes. Binding in the active site of thrombin is characterized by numerous van der Waals contacts and ring-ring system interactions. Unlike in the substrate-like inhibitors, ArgH2 enters the S1 specificity pocket from the P2 position and adopts a bent conformation to make an hydrogen bond to the carboxylate of Asp189. In this noncanonical position, its carbonyl points away from the oxyanion hole, which is now occupied by well-ordered solvent molecules. The linkers fit in the groove extending from the active site to the FRE. The C-terminal fragments of both inhibitors bind in the same way as analogous FRE binding elements in previously described complexes. PMID:8762149

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

    Directory of Open Access Journals (Sweden)

    Eliseo Portilla-de Buen

    2014-04-01

    Full Text Available 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.

  13. A Kazal-type inhibitor with thrombin specificity from Rhodnius prolixus.

    Science.gov (United States)

    Friedrich, T; Kröger, B; Bialojan, S; Lemaire, H G; Höffken, H W; Reuschenbach, P; Otte, M; Dodt, J

    1993-08-01

    A thrombin-specific inhibitor with an apparent molecular mass of 11 kDa has been purified from the insect Rhodnius prolixus. Amino-terminal protein sequence analysis allowed the molecular cloning of the corresponding cDNA. The open reading frame codes for a protein of about 103 amino acid residues and displays an internal sequence homology of residues 6-48 with residues 57-101 indicating a two-domain structure. Based on the amino acid sequence the two domains exhibit high homology to protease inhibitors belonging to the Kazal-type family. Model building suggests that the first domain binds to the active site with residue His10 pointing into the specificity pocket. From gel filtration and tight-binding inhibition experiments the inhibitor appears to form 1:1 complexes with thrombin. Periplasma-directed heterologous expression of the rhodniin cDNA in Escherichia coli yields the intact thrombin inhibitor. Natural and recombinant rhodniin both display inhibition constants of about 2 x 10(-13) M.

  14. XIMELAGATRAN: A NEW DIRECT THROMBIN INHIBITOR

    Directory of Open Access Journals (Sweden)

    Mehta Hiren R

    2011-04-01

    Full Text Available 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. Warfarin sodium is an effective oral anticoagulant drug. However, warfarin has a narrow therapeutic window with significant risks of hemorrhage at therapeutic concentrations. Dosing is difficult and requires frequent monitoring. New oral anticoagulant agents are required to improve current anticoagulant therapy. Furthermore, while warfarin is effective in venous disease, it does not provide more than 60% risk reduction compared with placebo in venous thrombosis prophylaxis and considerably lower risk reduction in terms of arterial thrombosis. 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 is an oral pro-drug of melagatran, a synthetic small peptidomimetic with direct thrombin inhibitory actions and anticoagulant activity. As an oral agent, ximelagatran has a number of desirable properties including a rapid onset of action, fixed dosing, stable absorption, apparent low potential for medication interactions, and no requirement for monitoring of drug levels or dose adjustment. It has a short plasma elimination half-life of about 4 hours in cases of unexpected hemorrhage or need for reversal.

  15. Hypersensitivity to thrombin of platelets from hypercholesterolemic rats

    International Nuclear Information System (INIS)

    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

  16. The Toxic Effect of Thrombin on Periventricular Tissue

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective To investigate the effect of thrombin on periventricular tissue. Methods Thrombin solution (10U/10μl) was infused into the ventricles of rats. Then, the local cerebral blood flow (rCBF) and blood-brain barrier (BBB) permeability in the area of basal ganglia and thalamus were measured. In separate experiments neurons were cultured in the medium with thrombin, and lactate dehydrogenase was determined. Results The thrombin induced blood-brain barrier disruption and neuron death in the culture, whereas cerebral blood flow did not drop to the injury level. Conclusion Periventricular tissue damage suffering from the thrombin and blood-brain barrier disruption and cell toxicity could elicit pathological mechanisms.

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

    Science.gov (United States)

    Frense, D.; Kang, S.; Schieke, K.; Reich, P.; Barthel, A.; Pliquett, U.; Nacke, T.; Brian, C.; Beckmann, D.

    2013-04-01

    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).

  18. Aptamer-based SERRS sensor for thrombin detection.

    Science.gov (United States)

    Cho, Hansang; Baker, Brian R; Wachsmann-Hogiu, Sebastian; Pagba, Cynthia V; Laurence, Ted A; Lane, Stephen M; Lee, Luke P; Tok, Jeffrey B H

    2008-12-01

    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 alpha-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'-capped, 3'-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. PMID:19367849

  19. APTAMER-BASED SERRS SENSOR FOR THROMBIN DETECTION

    Energy Technology Data Exchange (ETDEWEB)

    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

    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.

  20. Test of hirudin activity by tracking the binding of hirudin to thrombin in the presence of BS3 cross-linking.

    Science.gov (United States)

    Liu, Yanfang; Yang, Jian; Wang, Jiangmin; Huang, Qingmei; Yang, Xiaohong; Zhang, Jianhua

    2015-10-01

    Hirudin has a great potential in inhibiting thrombin, and its antithrombin activity has direct bearing on its clinical application. Using bovine alpha-thrombin and recombinant hirudin of Poecilobdella javanica purified from Phichia pastoris as materials, this study introduced a novel method to testing antithrombin activity of hirudin visually and dynamically by tracking the binding of hirudin to thrombin. After incubating the mixture of thrombin and hirudin at 37 °C for 5 min, the binding of hirudin to thrombin was cross-linked by bis[sulfosuccinimidyl] suberate for 30 min and visualized by SDS-polyacrylamide gel electrophoresis. With the aid of image analysis on the basis of INRA-Noésis E1D analysis software, antithrombin activity of hirudin was calculated through intensity variations of protein bands of either thrombin-hirudin compound, unbound thrombin, or unbound hirudin. In this regard, activity of the given hirudin was tested to be 5625 ATU/mg based on a single reaction, and 5675.3 ATU/mg based on a series of reactions in a stepwise manner, close to the result of 6000 ATU/mg concluded by titration method. The superiorities of the method include good accuracy (the minimum testable concentration of hirudin is 1.5 μg/ml) and little sample consumption (sample consumption of hirudin is generally 1-11.5 μl using the apparatus of Mini Protean 3 Cell). Easy operation, low input, and equipment requirement also grant it as an effective way. PMID:26332983

  1. Facile fabrication of an aptasensor for thrombin based on graphitic carbon nitride/TiO2 with high visible-light photoelectrochemical activity.

    Science.gov (United States)

    Fan, Dawei; Guo, Cuijuan; Ma, Hongmin; Zhao, Di; Li, Yina; Wu, Dan; Wei, Qin

    2016-01-15

    A novel aptasensor for thrombin with high visible-light activity was facilely fabricated based on graphitic carbon nitride/TiO2 (g-C3N4/TiO2) photoelectrochemical (PEC) composite. Crystallization of TiO2 nanoparticles (NPs) and their strong interaction with g-C3N4 sheet were confirmed by high-resolution transmission electron microscope (HR-TEM), both of which contributed to the high photocurrent intensity under visible-light irradiation. Carboxyl functionalized thrombin aptamers were first successfully bound to the g-C3N4/TiO2 modified electrode as proven by photoelectrochemical test and electrochemical impedance spectroscopy (EIS) analysis. Ascorbic acid was utilized as the electron donor for scavenging photo-generated holes and inhibiting light driven electron-hole pair recombination. The specific recognition between thrombin aptamer and thrombin led to the linear decrease of photocurrent with the increase of logarithm of thrombin concentration in the range of 5.0×10(-13)molL(-1) to 5.0×10(-9)molL(-1) with a detection limit of 1.2×10(-13)molL(-1). This proposed low-cost, convenient and sensitive aptasensor showed promising applications in biosensor and photoelectrochemical analysis.

  2. Activation of pro-(matrix metalloproteinase-2) (pro-MMP-2) by thrombin is membrane-type-MMP-dependent in human umbilical vein endothelial cells and generates a distinct 63 kDa active species.

    Science.gov (United States)

    Lafleur, M A; Hollenberg, M D; Atkinson, S J; Knäuper, V; Murphy, G; Edwards, D R

    2001-01-01

    Thrombin, a critical enzyme in the coagulation cascade, has also been associated with angiogenesis and activation of the zymogen form of matrix metalloproteinase-2 (MMP-2 or gelatinase-A). We show that thrombin activated pro-MMP-2 in a dose- and time-dependent manner in cultured human umbilical-vein endothelial cells (HUVECs) to generate a catalytically active 63 kDa protein that accumulated as the predominant form in the conditioned medium. This 63 kDa thrombin-activated MMP-2 is distinct from the 62 kDa species found following concanavalin A or PMA stimulated pro-MMP-2 activation. Hirudin and leupeptin blocked thrombin-induced pro-MMP-2 activation, demonstrating that the proteolytic activity of thrombin is essential. However, activation was also dependent upon membrane-type-MMP (MT-MMP) action, since it was blocked by EDTA, o-phenanthroline, hydroxamate metalloproteinase inhibitors, tissue inhibitor of metalloproteinase-2 (TIMP-2) and TIMP-4, but not TIMP-1. Thrombin inefficiently cleaved recombinant 72 kDa pro-MMP-2, but efficiently cleaved the 64 kDa MT-MMP-processed intermediate form in the presence of cells. Thrombin also rapidly (within 1 h) increased cellular MT-MMP activity, and at longer time points (>6 h) it increased expression of MT1-MMP mRNA and protein. Thus signalling via proteinase-activated receptors (PARs) may play a role in thrombin-induced MMP-2 activation, though this does not appear to involve PAR1, PAR2, or PAR4 in HUVECs. These results indicate that in HUVECs the activation of pro-MMP-2 by thrombin involves increased MT-MMP activity and preferential cleavage of the MT-MMP-processed 64 kDa MMP-2 form in the presence of cells. The integration of these proteinase systems in the vascular endothelium may be important during thrombogenesis and tissue remodelling associated with neovascularization. PMID:11415441

  3. Struktur und kinetische Charakterisierung vom Blutegel-Inhibitor Haemadin im Komplex mit humanem alpha;-Thrombin

    OpenAIRE

    Richardson, John Lee

    2007-01-01

    Haemadin, a peptide from the Indian land living leech Haemadipsa sylvestris was expressed, purified and crystallised in complex with human alpha-thrombin, and the structure solved to 3.1 Å. The inhibitor was thoroughly characterised with thrombin and ten thrombin mutants, which revealed which residues were important for binding. Synthetic tsetse thrombin inhibitor originally isolated from the tsetse fly was crystallised in complex with bovine alpha-thrombin and the structure was solved after ...

  4. The Characteristics of Thrombin in Osteoarthritic Pathogenesis and Treatment

    Directory of Open Access Journals (Sweden)

    Pei-Yu Chou

    2014-01-01

    Full Text Available Osteoarthritis (OA is a mechanical abnormality associated with degradation of joints. It is characterized by chronic, progressive degeneration of articular cartilage, abnormalities of bone, and synovial change. The most common symptom of OA is local inflammation resulting from exogenous stress or endogenous abnormal cytokines. Additionally, OA is associated with local and/or systemic activation of coagulation and anticoagulation pathways. Thrombin plays an important role in the stimulation of fibrin deposition and the proinflammatory processes in OA. Thrombin mediates hemostatic and inflammatory responses and guides the immune response to tissue damage. Thrombin activates intracellular signaling pathways by interacting with transmembrane domain G protein coupled receptors (GPCRs, known as protease-activated receptors (PARs. In pathogenic mechanisms, PARs have been implicated in the development of acute and chronic inflammatory responses in OA. Therefore, discovery of thrombin signaling pathways would help us to understand the mechanism of OA pathogenesis and lead us to develop therapeutic drugs in the future.

  5. Anticoagulants and the propagation phase of thrombin generation.

    Directory of Open Access Journals (Sweden)

    Thomas Orfeo

    Full Text Available The view that clot time-based assays do not provide a sufficient assessment of an individual's hemostatic competence, especially in the context of anticoagulant therapy, has provoked a search for new metrics, with significant focus directed at techniques that define the propagation phase of thrombin generation. Here we use our deterministic mathematical model of tissue-factor initiated thrombin generation in combination with reconstructions using purified protein components to characterize how the interplay between anticoagulant mechanisms and variable composition of the coagulation proteome result in differential regulation of the propagation phase of thrombin generation. Thrombin parameters were extracted from computationally derived thrombin generation profiles generated using coagulation proteome factor data from warfarin-treated individuals (N = 54 and matching groups of control individuals (N = 37. A computational clot time prolongation value (cINR was devised that correlated with their actual International Normalized Ratio (INR values, with differences between individual INR and cINR values shown to derive from the insensitivity of the INR to tissue factor pathway inhibitor (TFPI. The analysis suggests that normal range variation in TFPI levels could be an important contributor to the failure of the INR to adequately reflect the anticoagulated state in some individuals. Warfarin-induced changes in thrombin propagation phase parameters were then compared to those induced by unfractionated heparin, fondaparinux, rivaroxaban, and a reversible thrombin inhibitor. Anticoagulants were assessed at concentrations yielding equivalent cINR values, with each anticoagulant evaluated using 32 unique coagulation proteome compositions. The analyses showed that no anticoagulant recapitulated all features of warfarin propagation phase dynamics; differences in propagation phase effects suggest that anticoagulants that selectively target fXa or

  6. Thrombomodulin Binding Selects the Catalytically Active Form of Thrombin.

    Science.gov (United States)

    Handley, Lindsey D; Treuheit, Nicholas A; Venkatesh, Varun J; Komives, Elizabeth A

    2015-11-01

    Human α-thrombin is a serine protease with dual functions. Thrombin acts as a procoagulant, cleaving fibrinogen to make the fibrin clot, but when bound to thrombomodulin (TM), it acts as an anticoagulant, cleaving protein C. A minimal TM fragment consisting of the fourth, fifth, and most of the sixth EGF-like domain (TM456m) that has been prepared has much improved solubility, thrombin binding capacity, and anticoagulant activity versus those of previous TM456 constructs. In this work, we compare backbone amide exchange of human α-thrombin in three states: apo, D-Phe-Pro-Arg-chloromethylketone (PPACK)-bound, and TM456m-bound. Beyond causing a decreased level of amide exchange at their binding sites, TM and PPACK both cause a decreased level of amide exchange in other regions including the γ-loop and the adjacent N-terminus of the heavy chain. The decreased level of amide exchange in the N-terminus of the heavy chain is consistent with the historic model of activation of serine proteases, which involves insertion of this region into the β-barrel promoting the correct conformation of the catalytic residues. Contrary to crystal structures of thrombin, hydrogen-deuterium exchange mass spectrometry results suggest that the conformation of apo-thrombin does not yet have the N-terminus of the heavy chain properly inserted for optimal catalytic activity, and that binding of TM allosterically promotes the catalytically active conformation. PMID:26468766

  7. Rapid Detection of Thrombin and Other Protease Activity Directly in Whole Blood

    Science.gov (United States)

    Yu, Johnson Chung Sing

    Thrombin is a serine protease that plays a key role in the clotting cascade to promote hemostasis following injury to the endothelium. From a clinical diagnostic perspective, in-vivo thrombin activity is linked to various blood clotting disorders, as well as cardiovascular disease (DVT, arteriosclerosis, etc). Thus, the ability to rapidly measure protease activity directly in whole blood will provide important new diagnostics, and clinical researchers with a powerful tool to further elucidate the relationship between circulating protease levels and disease. The ultimate goal is to design novel point of care (POC) diagnostic devices that are capable of monitoring protease activities directly in whole blood and biological sample. A charge-changing substrate specific to the thrombin enzyme was engineered and its functionality was confirmed by a series of experiments. This led to the preliminary design, construction, and testing of two device platforms deemed fully functional for the electrophoretic separation and focusing of charged peptide fragments. The concept of using the existing charge-changing substrate platform for bacterial protease detection was also investigated. Certain strains of E coli are associated with severe symptoms such as abdominal cramps, bloody diarrhea, and vomiting. The OmpT protease is expressed on the outer membrane of E coli and plays a role in the cleavage of antimicrobial peptides, the degradation of recombinant heterologous proteins, and the activation of plasminogen in the host. Thus, a synthetic peptide substrate specific to the OmpT protease was designed and modeled for the purpose of detecting E coli in biological sample.

  8. Fluorouracil Topical

    Science.gov (United States)

    ... 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 ...

  9. Bimatoprost Topical

    Science.gov (United States)

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

  10. Bexarotene Topical

    Science.gov (United States)

    ... condition may improve as soon as 4 weeks after you begin to use topical bexarotene, or it may ... notice any improvement. Continue to use topical bexarotene after you notice improvement; your condition may continue to improve. ...

  11. Testosterone Topical

    Science.gov (United States)

    ... in which the body does not produce enough natural testosterone). Testosterone is used only for men with ... topical may control your symptoms but will not cure your condition. Continue to use testosterone topical even ...

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

    Directory of Open Access Journals (Sweden)

    Kristensen Torsten

    2009-05-01

    Full Text Available Abstract 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.

  13. Thrombin, a mediator of cerebrovascular inflammation in AD and hypoxia

    Directory of Open Access Journals (Sweden)

    Debjani eTripathy

    2013-05-01

    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 1α (HIF-1α, 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-1α, 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-1α, 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.

  14. Potentiation of thrombin generation in hemophilia A plasma by coagulation factor VIII and characterization of antibody-specific inhibition.

    Directory of Open Access Journals (Sweden)

    Bhavya S Doshi

    Full Text Available Development of inhibitory antibodies to coagulation factor VIII (fVIII is the primary obstacle to the treatment of hemophilia A in the developed world. This adverse reaction occurs in 20-30% of persons with severe hemophilia A treated with fVIII-replacement products and is characterized by the development of a humoral and neutralizing immune response to fVIII. Patients with inhibitory anti-fVIII antibodies are treated with bypassing agents including recombinant factor VIIa (rfVIIa. However, some patients display poor hemostatic response to bypass therapy and improved treatment options are needed. Recently, we demonstrated that fVIII inhibitors display widely variable kinetics of inhibition that correlate with their respective target epitopes. Thus, it was hypothesized that for antibodies that display slow rates of inhibition, supplementation of rfVIIa with fVIII would result in improved thrombin generation and be predictive of clinical responses to this novel treatment regimen. In order to test this hypothesis, 10 murine monoclonal antibodies (MAbs with non-overlapping epitopes spanning fVIII, differential inhibition titers, and inhibition kinetics were studied using a thrombin generation assay. Of the 3 MAbs with high inhibitory titers, only the one with fast and complete (classically defined as "type I" kinetics displayed significant inhibition of thrombin generation with no improvement upon supplementation of rfVIIa with fVIII. The other two MAbs that displayed incomplete (classically defined as "type II" inhibition did not suppress the potentiation of thrombin generation by fVIII. All antibodies that did not completely inhibit fVIII activity demonstrated potentiation of thrombin generation by the addition of fVIII as compared to rfVIIa alone. In conclusion, fVIII alone or in combination with rfVIIa corrects the thrombin generation defect produced by the majority of anti-fVIII MAbs better than single agent rfVIIa. Therefore, combined f

  15. Calibrated automated thrombin generation measurement in clotting plasma.

    Science.gov (United States)

    Hemker, H Coenraad; Giesen, Peter; Al Dieri, Raed; Regnault, Véronique; de Smedt, Eric; Wagenvoord, Rob; Lecompte, Thomas; Béguin, Suzette

    2003-01-01

    Calibrated automated thrombography displays the concentration of thrombin in clotting plasma with or without platelets (platelet-rich plasma/platelet-poor plasma, PRP/PPP) in up to 48 samples by monitoring the splitting of a fluorogenic substrate and comparing it to a constant known thrombin activity in a parallel, non-clotting sample. Thus, the non-linearity of the reaction rate with thrombin concentration is compensated for, and adding an excess of substrate can be avoided. Standard conditions were established at which acceptable experimental variation accompanies sensitivity to pathological changes. The coefficients of variation of the surface under the curve (endogenous thrombin potential) are: within experiment approximately 3%; intra-individual: AVK, heparin(-likes), direct inhibitors]. In PRP, it is diminished in von Willebrand's disease, but it also shows the effect of platelet inhibitors (e.g. aspirin and abciximab). Addition of activated protein C (APC) or thrombomodulin inhibits thrombin generation and reflects disorders of the APC system (congenital and acquired resistance, deficiencies and lupus antibodies) independent of concomitant inhibition of the procoagulant pathway as for example by anticoagulants.

  16. A Novel Photoelectrochemical Biosensor for Tyrosinase and Thrombin Detection

    Directory of Open Access Journals (Sweden)

    Jiexia Chen

    2016-01-01

    Full Text Available A novel photoelectrochemical biosensor for step-by-step assay of tyrosinase and thrombin was fabricated based on the specific interactions between the designed peptide and the target enzymes. A peptide chain with a special sequence which contains a positively charged lysine-labeled terminal, tyrosine at the other end and a cleavage site recognized by thrombin between them was designed. The designed peptide can be fixed on surface of the CdTe quantum dots (QDs-modified indium-tin oxide (ITO electrode through electrostatic attraction to construct the photoelectrochemical biosensor. The tyrosinase target can catalyze the oxidization of tyrosine by oxygen into ortho-benzoquinone residues, which results in a decrease in the sensor photocurrent. Subsequently, the cleavage site could be recognized and cut off by another thrombin target, restoring the sensor photocurrent. The decrease or increase of photocurrent in the sensor enables us to assay tyrosinase and thrombin. Thus, the detection of tyrosinase and thrombin can be achieved in the linear range from 2.6 to 32 μg/mL and from 4.5 to 100 μg/mL with detection limits of 1.5 μg/mL and 1.9 μg/mL, respectively. Most importantly, this strategy shall allow us to detect different classes of enzymes simultaneously by designing various enzyme-specific peptide substrates.

  17. A plasmin-activatable thrombin inhibitor reduces experimental thrombosis and assists experimental thrombolysis in murine models.

    Science.gov (United States)

    Sheffield, W P; Eltringham-Smith, L J; Gataiance, S; Bhakta, V

    2015-05-01

    The leech protein hirudin is a potent natural thrombin inhibitor. Its potential as an antithrombotic agent is limited by its promotion of bleeding. We attempted to modify this profile by positioning albumin and a plasmin cleavage site on its N-terminus, in recombinant protein HSACHV3 [comprising hirudin variant 3 (HV3) fused to the C-terminus of human serum albumin (HSA) via a plasmin cleavage site (C)], Previously we showed that HSACHV3 inhibited thrombin in a plasmin-dependent manner, and that, unlike HV3, it did not increase bleeding in vivo when administered to mice. Here we tested HSACHV3 for the ability to reduce thrombosis and assist enzymatic thrombolysis in animal models. Intravenous administration of HSACHV3, but not a control protein lacking the plasmin cleavage site (HSAHV3), reduced thrombus weight by 2.1-fold in the ferric chloride-injured mouse vena cava. Similarly, thrombi formed in a rabbit jugular vein stasis model were 1.7-fold lighter in animals treated with HSACHV3 compared to those receiving HSAHV3. Administration of 60 mg/kg body weight HSACHV3 prolonged the time to occlusion in the ferric chloride-injured mouse carotid artery by threefold compared to vehicle controls, while equimolar HSAHV3 had no effect. HSACHV3 had no ability to restore flow to the murine carotid arteries occluded by ferric chloride treatment, but combining HSACHV3 (60 mg/kg) with recombinant mutant tissue plasminogen activator (TNKase) significantly reduced the time to restore patency to the artery compared to TNKase alone. Unlike unfused HV3, HSACHV3 did not increase bleeding in a mouse liver laceration model. Our results show that HSACHV3 acts as an antithrombotic agent that does not promote bleeding and which speeds the time to flow restoration when used as an adjunct to pharmacological thrombolysis in animal models. PMID:25481811

  18. Thrombin-Responsive Gated Silica Mesoporous Nanoparticles As Coagulation Regulators.

    Science.gov (United States)

    Bhat, Ravishankar; Ribes, Àngela; Mas, Núria; Aznar, Elena; Sancenón, Félix; Marcos, M Dolores; Murguía, Jose R; Venkataraman, Abbaraju; Martínez-Máñez, Ramón

    2016-02-01

    The possibility of achieving sophisticated actions in complex biological environments using gated nanoparticles is an exciting prospect with much potential. We herein describe new gated mesoporous silica nanoparticles (MSN) loaded with an anticoagulant drug and capped with a peptide containing a thrombin-specific cleavage site. When the coagulation cascade was triggered, active thrombin degraded the capping peptidic sequence and induced the release of anticoagulant drugs to delay the clotting process. The thrombin-dependent response was assessed and a significant increase in coagulation time in plasma from 2.6 min to 5 min was found. This work broadens the application of gated silica nanoparticles and demonstrates their ability to act as controllers in a complex scenario such as hemostasis. PMID:26794474

  19. Human thrombin for the treatment of gastric and ectopic varices

    Institute of Scientific and Technical Information of China (English)

    Norma C McAvoy; John N Plevris; Peter C Hayes

    2012-01-01

    AIM:To evaluate the efficacy of human thrombin in the treatment of bleeding gastric and ectopic varices.METHODS:Retrospective observational study in a Tertiary Referral Centre.Between January 1999-October 2005,we identified 37 patients who were endoscopically treated with human thrombin injection therapy for bleeding gastric and ectopic varices.Patient details including age,gender and aetiology of liver disease/ segmental portal hypertension were documented.The thrombin was obtained from the Scottish National Blood Transfusion Service and prepared to give a solution of 250 IU/mL which was injected via a standard injection needle.All patient case notes were reviewed and the total dose of thrombin given along with the number of endoscopy sessions was recorded.Initial haemostasis rates,rebleeding rates and mortality were catalogued along with the incidence of any immediate complications which could be attributable to the thrombin therapy.The duration of follow up was also listed.The study was conducted according to the United Kingdom research ethics guidelines.RESULTS:Thirty-seven patients were included.33 patients (89%) had thrombin (250 U/mL) for gastric varices,2 (5.4%) for duodenal varices,1 for rectal varices and 1 for gastric and rectal varices.(1) Gastric varices,an average of 15.2 mL of thrombin was used per patient.Re-bleeding occurred in 4 patients (10.8%),managed in 2 by a transjugular intrahepatic portosystemic shunt (TIPSS) (one unsuccessfully who died) and in other 2 by a distal splenorenal shunt; (2) Duodenal varices (or type 2 isolated gastric varices),an average of 12.5 mL was used per patient over 2-3 endoscopy sessions.Re-bleeding occurred in one patient,which was treated by TIPSS; and (3) Rectal varices,an average of 18.3 mL was used per patient over 3 endoscopy sessions.No re-bleeding occurred in this group.CONCLUSION:Human thrombin is a safe,easy to use and effective therapeutic option to control haemorrhage from gastric and ectopic varices.

  20. Thrombin-like enzymes from venom gland of Deinagkistrodon acutus:cDNA cloning, mechanism of diversity and phylogenetic tree construction

    Institute of Scientific and Technical Information of China (English)

    Xiang-dong ZHA; He-sheng HUANG; Li-zhi ZHOU; Jing LIU; Kang-sen XU

    2006-01-01

    Aim: To clone cDNAs of thrombin-like enzymes (TLEs) from venom gland of Deinagkistrodon acutus and analyze the mechanisms by which their structural diversity arose. Methods: Reverse transcription-polymerase chain reaction and gene cloning techniques were used, and the cloned sequences were analyzed by using bioinformatics tools. Results: Novel cDNAs of snake venom TLEs were cloned. The possibilities of post-transcriptional recombination and horizontal gene transfer are discussed. A phylogenetic tree was constructed. Conclusion:The cDNAs of snake venom TLEs exhibit great diversification. There are several types of structural variations. These variations may be attributable to certain mechanisms including recombination.

  1. New Approaches to the Role of Thrombin in Acute Coronary Syndromes: Quo Vadis Bivalirudin, a Direct Thrombin Inhibitor?

    Directory of Open Access Journals (Sweden)

    María Asunción Esteve-Pastor

    2016-02-01

    Full Text Available The pathophysiology of acute coronary syndrome (ACS involves platelet activation and thrombus formation after the rupture of atherosclerotic plaques. Thrombin is generated at the blood-plaque interface in association with cellular membranes on cells and platelets. Thrombin also amplifies the response to the tissue injury, coagulation and platelet response, so the treatment of ACS is based on the combined use of both antiplatelet (such as aspirin, clopidogrel, prasugrel and ticagrelor and antithrombotic drugs (unfractionated heparin, enoxaparin, fondaparinux and bivalirudin. Bivalirudin competitively inhibits thrombin with high affinity, a predictable response from its linear pharmacokinetics and short action. However, a present remarkable controversy exists between the latest main Guidelines in Clinical Practice and the key trials evaluating the use of bivalirudin in ACS. The aim of this review is to update the development of bivalirudin, including pharmacological properties, obtained information from clinical trials evaluating efficacy and safety of bivalirudin in ACS; as well as the recommendations of clinical Guidelines.

  2. AI Topics

    OpenAIRE

    Buchanan, Bruce G; Glick, Jonathan

    2002-01-01

    The debut of the AI in the News column elsewhere in this issue of AI Magazine created a good opportunity to introduce the professional community to the AI Topics web site, home of the AI in the news virtual page. Although AI Topics is designed for the lay public, it serves a much larger audience.

  3. Increased anticoagulant activity of thrombin-binding DNA aptamers by nanoscale organization on DNA nanostructures

    DEFF Research Database (Denmark)

    Rangnekar, Abhijit; Zhang, Alex M.; Shiyuan Li, Susan;

    2012-01-01

    Control over thrombin activity is much desired to regulate blood clotting in surgical and therapeutic situations. Thrombin-binding RNA and DNA aptamers have been used to inhibit thrombin activity and thus the coagulation cascade. Soluble DNA aptamers, as well as two different aptamers tethered by...

  4. A Guided Mode Resonance Aptasensor for Thrombin Detection

    Directory of Open Access Journals (Sweden)

    Wen-Yih Chen

    2011-09-01

    Full Text Available Recent developments in aptamers have led to their widespread use in analytical and diagnostic applications, particularly for biosensing. Previous studies have combined aptamers as ligands with various sensors for numerous applications. However, merging the aptamer developments with guided mode resonance (GMR devices has not been attempted. This study reports an aptasensor based home built GMR device. The 29-mer thrombin aptamer was immobilized on the surface of a GMR device as a recognizing ligand for thrombin detection. The sensitivity reported in this first trial study is 0.04 nm/μM for thrombin detection in the concentration range from 0.25 to 1 μM and the limit of detection (LOD is 0.19 μM. Furthermore, the binding affinity constant (Ka measured is in the range of 106 M−1. The investigation has demonstrated that such a GMR aptasensor has the required sensitivity for the real time, label-free, in situ detection of thrombin and provides kinetic information related to the binding.

  5. Translational success stories: development of direct thrombin inhibitors.

    Science.gov (United States)

    Coppens, Michiel; Eikelboom, John W; Gustafsson, David; Weitz, Jeffrey I; Hirsh, Jack

    2012-09-14

    Anticoagulants are the cornerstone of therapy for conditions associated with arterial and venous thrombosis. Direct thrombin inhibitors (DTIs) are anticoagulants that bind to thrombin and block its enzymatic activity. The bivalent parenteral DTIs hirudin and bivalirudin were based on the observation that the salivary extracts of medicinal leeches prevented blood from clotting. Key events that facilitated the subsequent development of small molecule active site inhibitors, such as argatroban, were the observation that fibrinopeptide A had antithrombotic properties and determination of the crystal structure of thrombin. Hirudin and argatroban have found their niche for the treatment of patients with heparin-induced thrombocytopenia, whereas bivalirudin is approved as an alternative to heparin for patients undergoing percutaneous coronary intervention. The development of orally active direct thrombin inhibitors was challenging because of the need to convert water-soluble, poorly absorbable, active site inhibitors into fat-soluble prodrugs that were then transformed back to the active drug after intestinal absorption. Dabigatran etexilate was the first new oral anticoagulant to be approved for long-term anticoagulant treatment in 6 decades. This Review highlights the development of DTIs as a translational success story; an example in which the combination of scientific ingenuity, structure-based design, and rigorous clinical trials has created a new class of anticoagulants that has improved patient care. PMID:22982873

  6. Recent Developments in Thrombin-Activatable Fibrinolysis Inhibitor Research

    NARCIS (Netherlands)

    P.F. Marx; C.J.N. Verkleij; M.V. Seron; J.C.M. Meijers

    2009-01-01

    Thrombin-activatable fibrinolysis inhibitor (TAFI) provides an important molecular link between the coagulation and fibrinolytic systems. In this review, recent major advances in TAFI research, including the elucidation of crystal structures, the development of small inhibitors and the role of TAFI

  7. 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

  8. Clindamycin Topical

    Science.gov (United States)

    ... topical clindamycin.you should know that abrasive or medicated soaps and skin products that contain alcohol may ... in, tightly closed, and out of reach of children. Store it at room temperature and away from ...

  9. Estradiol Topical

    Science.gov (United States)

    ... Estradiol topical gel is also used to treat vaginal dryness, itching, and burning in women who are experiencing menopause. However, women whose only bothersome symptoms are vaginal burning, itching, and dryness may benefit more from a medication that is ...

  10. Tretinoin Topical

    Science.gov (United States)

    ... lotions, astringents, and perfumes); they can sting your skin, especially when you first use tretinoin.Do not use any other topical medications, especially benzoyl peroxide, salicylic acid (wart remover), and dandruff shampoos containing sulfur or ...

  11. Thrombin mutant W215A/E217A treatment improves neurological outcome and attenuates central nervous system damage in experimental autoimmune encephalomyelitis.

    Science.gov (United States)

    Verbout, Norah G; Yu, Xiaolin; Healy, Laura D; Phillips, Kevin G; Tucker, Erik I; Gruber, András; McCarty, Owen J T; Offner, Halina

    2015-02-01

    Multiple sclerosis (MS) is a neuroinflammatory disease characterized by demyelination and axonal damage of the central nervous system. The pathogenesis of MS has also been linked to vascular inflammation and local activation of the coagulation system, resulting in perivascular fibrin deposition. Treatment of experimental autoimmune encephalomyelitis (EAE), a model of human MS, with antithrombotic and antiinflammatory activated protein C (APC) reduces disease severity. Since recombinant APC (Drotecogin alfa), originally approved for the treatment of severe sepsis, is not available for human MS studies, we tested the hypothesis that pharmacologic activation of endogenous protein C could likewise improve the outcome of EAE. Mice were immunized with murine myelin oligodendrocyte glycoprotein (MOG) peptides and at the onset of EAE symptoms, were treated every other day with either WE thrombin (25 μg/kg; i.v.), a selective recombinant protein C activator thrombin analog, or saline control. Mice were monitored for changes in disease score until euthanized for ex vivo analysis of inflammation. Administration of WE thrombin significantly ameliorated clinical severity of EAE, reduced inflammatory cell infiltration and demyelination, suppressed the activation of macrophages comprising the CD11b + population and reduced accumulation of fibrin (ogen) in the spinal cord. These data suggest that symptomatic MS may respond to a treatment strategy that involves temporal pharmacological enhancement of endogenous APC generation. PMID:24810631

  12. Homologous desensitization of HEL cell thrombin receptors. Distinguishable roles for proteolysis and phosphorylation.

    Science.gov (United States)

    Brass, L F

    1992-03-25

    Loss of sensitivity to thrombin following an initial response is characteristic of a number of cell types, including platelets. It has recently been proposed that thrombin receptors resemble other G protein-coupled receptors, but that activation involves a novel mechanism in which thrombin cleaves the receptor, exposing a new N terminus that serves as the ligand for the receptor. Based upon this model, we have examined the mechanism of thrombin receptor desensitization by comparing the effects of thrombin with those of a peptide corresponding to the N-terminal sequence of the receptor following proteolysis by thrombin: SFLLRNPNDKYEPF or TRP42/55. Like thrombin, TRP42/55 stimulated pertussis toxin-sensitive inositol 1,4,5-trisphosphate formation, raised cytosolic Ca2+, and inhibited cAMP formation in the megakaryoblastic HEL cell line. Exposure to either thrombin or TRP42/55 desensitized the cells to both, but not to a third agonist, neuropeptide Y. The rate of recovery after desensitization depended upon the order of agonist addition. Resensitization of the cell to thrombin following a brief exposure to thrombin required up to 24 h and could be inhibited with cycloheximide. Resensitization to TRP42/55 after exposure to thrombin, or to thrombin after exposure to TRP42/55, on the other hand, was detectable within 30 min and could be inhibited by serine/threonine phosphatase inhibitors, but not by cycloheximide. Loss of responsiveness to thrombin and TRP42/55 was also observed following addition of the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA). However, while the protein kinase inhibitor staurosporine completely prevented the desensitization caused by TPA, it had only a limited effect on the desensitization caused by TRP42/55. These results demonstrate that the G protein-mediated effects of thrombin can be reproduced by a receptor-derived peptide and suggest that desensitization occurs by at least two mechanisms. The first, which is seen with thrombin

  13. Spotlight Topics

    Science.gov (United States)

    A Spotlight Topic consists of a set of two or more review articles focused on a specific subject in surface science. The topics are recommended by the Board of Editors. A topic may be chosen because it is particularly new or fast-breaking, thus deserving introduction to the general readership. Or, it may be because a topic is especially controversial or confusing, requiring clarification by experts. Each review will give a critical assessment rather than an encyclopedic report. While our editors always will insist on fairness and accuracy, any review which forwards an opinion is bound to be somewhat subjective. Therefore, it is the editors' wish that the set of reviews written by different authors on the same subject matter will provide a broad and balanced viewpoint. It is often the case that an author who is an expert in a technique or method may be especially enthusiastic or critical about this technique or method. A companion review in the set may provide a different viewpoint. We are hopeful that the reader, after studying these reviews and checking some of the key references, will obtain an informed opinion of the subject. We think the set of reviews in a spotlight area will considerably shorten the ``learning time'' that a nonexpert would otherwise need to become knowledgeable about a subject. In this issue, we feature a spotlight topic on oxide surfaces. The set contains an overview article by Jacques Jupille, and four articles written by G. Pacchioni, F. Cosandey and T. E. Madey, B. G. Daniels, R. Lindsay and G. Thornton, and C. Noguera respectively. Of these, the article by Pacchioni has already appeared in SRL 7, 277 (2000). The other three articles appear in this issue. A reader who wishes to suggest a spotlight topic or recommend authors to write such reviews should contact the Editor-in-Chief. We would like to hear from you.

  14. Increased thrombin generation in women with polycystic ovary syndrome

    DEFF Research Database (Denmark)

    Glintborg, Dorte; Sidelmann, Johannes Jakobsen; Lambaa Altinok, Magda;

    2015-01-01

    Objective. Polycystic ovary syndrome (PCOS) is associated with risk factors for cardiovascular disease (CVD) which may be modified by the use of metformin and oral contraceptives (OC). Thrombin generation (TG) measures are risk markers of CVD and address the composite of multiple factors...... randomized to 12 months treatment with metformin, metformin + OC or OC alone. C-reactive protein (CRP), fibrinogen, total cholesterol, trunk fat mass, body mass index, estradiol, testosterone, sex hormone binding globulin (SHBG) as well as TG measures, i.e. the lag time for formation of thrombin...... significantly. Conclusions. PCOS is associated with increase in TG measures independent of other risk factors of CVD. OC increases TG measures further and may thus add to the increased risk of CVD already present in women with PCOS....

  15. A novel histochemical method for the visualization of thrombin activity in the nervous system.

    Science.gov (United States)

    Bushi, D; Gera, O; Kostenich, G; Shavit-Stein, E; Weiss, R; Chapman, J; Tanne, D

    2016-04-21

    Although thrombin has an important role in both central and peripheral nerve diseases, characterization of the anatomical distribution of its proteolytic activity has been limited by available methods. This study presents the development, challenges, validation and implementation of a novel histochemical method for visualization of thrombin activity in the nervous system. The method is based on the cleavage of the substrate, Boc-Asp(OBzl)-Pro-Arg-4MβNA by thrombin to liberate free 4-methoxy-2-naphthylamine (4MβNA). In the presence of 5-nitrosalicylaldehyde, free 4MβNA is captured, yielding an insoluble yellow fluorescent precipitate which marks the site of thrombin activity. The sensitivity of the method was determined in vitro using known concentrations of thrombin while the specificity was verified using a highly specific thrombin inhibitor. Using this method we determined the spatial distribution of thrombin activity in mouse brain following transient middle cerebral artery occlusion (tMCAo) and in mouse sciatic nerve following crush injury. Fluorescence microscopy revealed well-defined thrombin activity localized to the right ischemic hemisphere in cortical areas and in the striatum compared to negligible thrombin activity contralaterally. The histochemical localization of thrombin activity following tMCAo was in good correlation with the infarct areas per triphenyltetrazolium chloride staining and to thrombin activity measured biochemically in tissue punches (85 ± 35 and 20 ± 3 mU/ml, in the cortical and striatum areas respectively, compared to 7 ± 2 and 13 ± 2 mU/ml, in the corresponding contralateral areas; mean ± SEM; p<0.05). In addition, 24 h following crush injury, focal areas of highly elevated thrombin activity were detected in teased sciatic fibers. This observation was supported by the biochemical assay and western blot technique. The histochemical method developed in this study can serve as an important tool for studying the role of thrombin

  16. An evaluation of different enzymatic cleavage methods for recombinant fusion proteins, applied on des(1-3)insulin-like growth factor I.

    Science.gov (United States)

    Forsberg, G; Baastrup, B; Rondahl, H; Holmgren, E; Pohl, G; Hartmanis, M; Lake, M

    1992-04-01

    Different enzymatic methods for cleavage of recombinant fusion proteins were compared. To find an efficient cleavage method, five different fusion proteins were produced. The fusion proteins differed only in the linker region between the fusion partner and the desired product, human des(1-3)insulin-like growth factor I. A cleavage study was performed with enterokinase, plasmin, thrombin, urokinase, and recombinant H64A subtilisin. Significant cleavage was obtained using thrombin, H64A subtilisin, and enterokinase. Thrombin cleavage was studied on a larger scale and des(1-3)IGF-I was recovered at a final yield of 3 mg/L growth medium. Thrombin and enterokinase were also studied as immobilized proteases and they cleaved the fusion proteins with retained activity. To further improve thrombin cleavage, a continuous reactor was constructed, consisting of a closed system with a thrombin column and an ion exchange column in series. Here, the fusion protein circulated while free des(1-3)IGF-I was bound to the ion exchange column after release from the fusion protein. In the reactor, thrombin was as efficient as the free enzyme but gave a diminished rate of product degradation. PMID:1388667

  17. Metabolic Plasticity in Resting and Thrombin Activated Platelets

    OpenAIRE

    Ravi, Saranya; Chacko, Balu; Sawada, Hirotaka; Kramer, Philip A.; Johnson, Michelle S.; Benavides, Gloria A.; O’DONNELL, Valerie; Marques, Marisa B.; Darley-Usmar, Victor M.

    2015-01-01

    Platelet thrombus formation includes several integrated processes involving aggregation, secretion of granules, release of arachidonic acid and clot retraction, but it is not clear which metabolic fuels are required to support these events. We hypothesized that there is flexibility in the fuels that can be utilized to serve the energetic and metabolic needs for resting and thrombin-dependent platelet aggregation. Using platelets from healthy human donors, we found that there was a rapid throm...

  18. An electrochemical aptasensor electrocatalyst for detection of thrombin.

    Science.gov (United States)

    Tian, Rong; Chen, Xiaojun; Li, Qingwen; Yao, Cheng

    2016-05-01

    This work reports a novel signal amplification strategy based on three-dimensional ordered macroporous C60-poly(3,4-ethylenedioxythiophene)-1-butyl-3-methylimidazolium hexafluorophosphate (3DOM C60-PEDOT-[BMIm][BF6]) for ultrasensitive detection of thrombin by cascade catalysis of Au-PEDOT@SiO2 microspheres and alcohol dehydrogenase (ADH). Au-PEDOT@SiO2 microspheres were constructed not only as nanocarriers to anchor the large amounts of secondary thrombin aptamers but also as nanocatalysts to catalyze the oxidation of ethanol efficiently. Significantly, the electrochemical signal was greatly enhanced based on cascade catalysis: First, ADH catalyzed the oxidation of ethanol to acetaldehyde with the concomitant generation of NADH in the presence of β-nicotinamide adenine dinucleotide hydrate (NAD(+)). Then, gold nanoparticles (AuNPs) as nanocatalysts could effectively catalyze NADH to produce NAD(+) with the help of PEDOT as redox probe. Under the optimal conditions, the proposed aptasensor exhibits a linear range of 2 × 10(-13) to 2 × 10(-8) M with a low detection limit of 2 × 10(-14) M for thrombin detection and shows high sensitivity and good specificity. PMID:26869084

  19. Metabolic plasticity in resting and thrombin activated platelets.

    Directory of Open Access Journals (Sweden)

    Saranya Ravi

    Full Text Available Platelet thrombus formation includes several integrated processes involving aggregation, secretion of granules, release of arachidonic acid and clot retraction, but it is not clear which metabolic fuels are required to support these events. We hypothesized that there is flexibility in the fuels that can be utilized to serve the energetic and metabolic needs for resting and thrombin-dependent platelet aggregation. Using platelets from healthy human donors, we found that there was a rapid thrombin-dependent increase in oxidative phosphorylation which required both glutamine and fatty acids but not glucose. Inhibition of fatty acid oxidation or glutamine utilization could be compensated for by increased glycolytic flux. No evidence for significant mitochondrial dysfunction was found, and ATP/ADP ratios were maintained following the addition of thrombin, indicating the presence of functional and active mitochondrial oxidative phosphorylation during the early stages of aggregation. Interestingly, inhibition of fatty acid oxidation and glutaminolysis alone or in combination is not sufficient to prevent platelet aggregation, due to compensation from glycolysis, whereas inhibitors of glycolysis inhibited aggregation approximately 50%. The combined effects of inhibitors of glycolysis and oxidative phosphorylation were synergistic in the inhibition of platelet aggregation. In summary, both glycolysis and oxidative phosphorylation contribute to platelet metabolism in the resting and activated state, with fatty acid oxidation and to a smaller extent glutaminolysis contributing to the increased energy demand.

  20. Stabilization of the E* Form Turns Thrombin into an Anticoagulant

    Energy Technology Data Exchange (ETDEWEB)

    Bah, Alaji; Carrell, Christopher J.; Chen, Zhiwei; Gandhi, Prafull S.; Di Cera, Enrico; (WU-MED)

    2009-07-31

    Previous studies have shown that deletion of nine residues in the autolysis loop of thrombin produces a mutant with an anticoagulant propensity of potential clinical relevance, but the molecular origin of the effect has remained unresolved. The x-ray crystal structure of this mutant solved in the free form at 1.55 {angstrom} resolution reveals an inactive conformation that is practically identical (root mean square deviation of 0.154 {angstrom}) to the recently identified E* form. The side chain of Trp215 collapses into the active site by shifting >10 {angstrom} from its position in the active E form, and the oxyanion hole is disrupted by a flip of the Glu192-Gly193 peptide bond. This finding confirms the existence of the inactive form E* in essentially the same incarnation as first identified in the structure of the thrombin mutant D102N. In addition, it demonstrates that the anticoagulant profile often caused by a mutation of the thrombin scaffold finds its likely molecular origin in the stabilization of the inactive E* form that is selectively shifted to the active E form upon thrombomodulin and protein C binding.

  1. Tissue factor-independent effects of recombinant factor VIIa on hemostasis

    NARCIS (Netherlands)

    Weeterings, Cees; Lisman, Ton; de Groot, Philip G.

    2008-01-01

    The molecular mechanisms responsible for the hemostatic efficacy of recombinant activated factor VII (rFVIIa; NovoSeven (R), Novo Nordisk, Bagsvaerd, Denmark) in platelet-related bleeding disorders remain unclear. The general concept is that rFVIIa locally enhances thrombin generation at the site of

  2. Increased anticoagulant activity of thrombin-binding DNA aptamers by nanoscale organization on DNA nanostructures

    OpenAIRE

    Rangnekar, Abhijit; Zhang, Alex M.; Shiyuan Li, Susan; M. Bompiani, Kristin; Hansen, Majken Nørgaard; Gothelf, Kurt Vesterager; Sullenger, Bruce A; LaBean, Thomas H.

    2012-01-01

    Control over thrombin activity is much desired to regulate blood clotting in surgical and therapeutic situations. Thrombin-binding RNA and DNA aptamers have been used to inhibit thrombin activity and thus the coagulation cascade. Soluble DNA aptamers, as well as two different aptamers tethered by a flexible single-strand linker, have been shown to possess anticoagulant activity. Here, we link multiple aptamers at programmed positions on DNA nanostructures to optimize spacing and orientation o...

  3. Regulation of tissue inflammation by thrombin-activatable carboxypeptidase B (or TAFI)

    OpenAIRE

    Leung, Lawrence L. K.; Myles, Timothy; Nishimura, Toshihiko; Song, Jason J.; Robinson, William H.

    2008-01-01

    Thrombin-activatable procarboxypeptidase B (proCPB or thrombin-activatable fibrinolysis inhibitor or TAFI) is a plasma procarboxypeptidase that is activated by the thrombin-thrombomodulin complex on the vascular endothelial surface. The activated CPB removes the newly exposed carboxyl terminal lysines in the partially digested fibrin clot, diminishes tissue plasminogen activator and plasminogen binding, and protects the clot from premature lysis. We have recently shown that CPB is catalytical...

  4. 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)

    Herrera Maria

    2006-04-01

    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.

  5. Thrombin Cleavage of Osteopontin Modulates Its Activities in Human Cells In Vitro and Mouse Experimental Autoimmune Encephalomyelitis In Vivo

    Directory of Open Access Journals (Sweden)

    Elena Boggio

    2016-01-01

    Full Text Available Osteopontin is a proinflammatory cytokine and plays a pathogenetic role in multiple sclerosis and its animal model, experimental autoimmune encephalomyelitis (EAE, by recruiting autoreactive T cells into the central nervous system. Osteopontin functions are modulated by thrombin cleavage generating N- and C-terminal fragment, whose individual roles are only partly known. Published data are difficult to compare since they have been obtained with heterogeneous approaches. Interestingly, thrombin cleavage of osteopontin unmasks a cryptic domain of interaction with α4β1 integrin that is the main adhesion molecule involved in lymphocyte transmigration to the brain and is the target for natalizumab, the most potent drug preventing relapses. We produced recombinant osteopontin and its N- and C-terminal fragments in an eukaryotic system in order to allow their posttranslational modifications. We investigated, in vitro, their effect on human cells and in vivo in EAE. We found that the osteopontin cleavage plays a key role in the function of this cytokine and that the two fragments exert distinct effects both in vitro and in vivo. These findings suggest that drugs targeting each fragment may be used to fine-tune the pathological effects of osteopontin in several diseases.

  6. Binding modes of thrombin binding aptamers investigated by simulations and experiments

    Science.gov (United States)

    Trapaidze, A.; Bancaud, A.; Brut, M.

    2015-01-01

    Thrombin binding aptamers HD1 and HD22 are the most studied aptamers, both for therapeutic and sensing purposes. Yet, there is still no commercialized aptamer-based sensor device for thrombin detection, suggesting that the binding modes of these aptamers remain to be precisely described. Here, we investigate thrombin-aptamer interactions with molecular dynamics simulations, and show that the different solved structures of HD1-thrombin complex are energetically similar and consequently possibly co-existing. Conversely, HD22 folding is much more stable, and its binding energy with thrombin is significantly larger than that of HD1 complexes. These results are confronted to experiments, which consist in monitoring aggregation of aptamer-functionalized gold nanoparticles triggered by thrombin. HD1 alone, but not HD22, can trigger aggregation, meaning that this aptamer has multiple sites of interactions with thrombin. Furthermore, pre-incubation of HD22 with thrombin impedes HD1 aggregation, suggesting that HD1 and HD22 have competing affinities for the same binding site. Altogether, this study shows that the characterization of aptamer-thrombin interactions by structural and kinetic experiments joined to simulations is necessary for the development of biosensors.

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

    Energy Technology Data Exchange (ETDEWEB)

    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

    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

  8. Low paediatric thrombin generation is caused by an attenuation of prothrombin conversion.

    Science.gov (United States)

    Kremers, Romy M W; Wagenvoord, Rob J; de Laat, H Bas; Monagle, Paul; Hemker, H Coenraad; Ignjatovic, Vera

    2016-06-01

    Thrombin generation (TG) is decreased in children. TG is determined by two underlying processes: the conversion of prothrombin to thrombin and the inactivation of thrombin. Therefore, lower TG capacity in children can either be caused by a reduction of prothrombin conversion, an increase of thrombin inactivation, or both. In 36 children and 8 adults, TG and the factors that determine thrombin inactivation (antithrombin, α2Macroglobulin (α2M) and fibrinogen) were measured. Prothrombin conversion, thrombin inhibitor complex formation, and the overall thrombin decay capacity were determined. In silico modelling was performed to determine the contribution prothrombin conversion and thrombin inactivation to deviant paediatric TG. Both the amount of prothrombin converted and the maximal prothrombin conversion rate are significantly reduced in children as compared to adults. This is partly due to the prothrombin levels being lower and partly to a lower prothrombin conversion rate. The overall thrombin decay capacity is not significantly different in children, but α2Macroglobulin plays a more important role than it does in adults. In silico experiments demonstrate that reduced prothrombin conversion and to a lesser extent elevated α2M levels provide an explanation for low TG in children. Young age has a dual effect on prothrombin conversion. Lower plasma prothrombin levels result in decreased prothrombin conversion but the rate of prothrombin conversion is also decreased, i. e. the development of prothrombinase is lower than in adults.

  9. Recombination instability

    DEFF Research Database (Denmark)

    D'Angelo, N.

    1967-01-01

    A recombination instability is considered which may arise in a plasma if the temperature dependence of the volume recombination coefficient, alpha, is sufficiently strong. Two cases are analyzed: (a) a steady-state plasma produced in a neutral gas by X-rays or high energy electrons; and (b...

  10. Interaction of C1 inhibitor with thrombin on the endothelial surface.

    Science.gov (United States)

    Caccia, Sonia; Castelli, Roberto; Maiocchi, Diana; Bergamaschini, Luigi; Cugno, Massimo

    2011-10-01

    Thrombin, the central bioregulatory enzyme of haemostasis, also has a potent vasopermeability effect on the surface of endothelial cells, and has therefore been considered a major link between the activation of the coagulation pathway and inflammation. C1 inhibitor inhibits thrombin with a low second-order rate constant that can be increased by heparin. The aim of this study was to investigate whether the C1 inhibitor-induced inhibition of thrombin is potentiated on the endothelial surface. The interaction of C1 inhibitor and thrombin was evaluated in an in-vitro system of human umbilical vein endothelial cells (HUVECs) to which purified C1 inhibitor and thrombin have been added. The role of heparins and selectins has been tested by adding heparinase and Mab to selectins. Kinetic analysis under pseudo-first-order conditions showed that the inhibitory effect of C1 inhibitor on thrombin is greater on the surface of endothelial cells. After incubating nanomolar concentrations of thrombin and micromolar concentrations of C1 inhibitor in a purified system, thrombin activity remained significant, but was almost totally suppressed in the presence of HUVECs. The abolition of such suppression by heparinase and Mab to selectins supports the involvement of heparin and selectins in C1 inhibitor-thrombin interaction. Furthermore, the second-order rate constant was 25 ± 3 /s per mol/l in our purified system, but increased to 100 ± 9 /s per mol/l in the presence of HUVECs. Our results indicate that C1 inhibitor can inhibit thrombin activity on vascular endothelium via binding to selectins and potentiation by heparins. This may contribute to the modulation of thrombin activity on vasopermeability and on coagulation especially when the major natural anticoagulant pathways are impaired. PMID:21959589

  11. Poor prognosis of hypocoagulability assessed by thrombin generation assay in disseminated intravascular coagulation.

    Science.gov (United States)

    Lee, Kyunghoon; Kim, Ji-Eun; Kwon, Jihyun; Kim, Inho; Yoon, Sung-Soo; Park, Seonyang; Han, Kyou-Sup; Kim, Hyun Kyung

    2014-04-01

    Overall assessment of the hemostatic system including procoagulant and anticoagulant changes may help assess the clinical status and prognosis of disseminated intravascular coagulation (DIC). The thrombin generation assay provides useful information about the global hemostatic status. Therefore, we measured several parameters of global hemostatic potential by the thrombin generation assay in patients suspected of having DIC. A total of 114 patients with suspected DIC were included. The thrombin generation assay was performed on the calibrated automated thrombogram using tissue factor with or without the addition of thrombomodulin, showing three parameters: lag time, endogenous thrombin potential (ETP), and peak thrombin. Both 1 and 5 pmol/l tissue factor-stimulated ETP and peak thrombin were well correlated with DIC severity. Interestingly, antithrombin level greatly affected ETP, whereas protein C influenced lag time. Prognostic analysis revealed that the area under the curve of peak thrombin stimulated by 1 pmol/l tissue factor was superior to that of D-dimer. Moreover, multivariate Cox analysis showed that the lag time and time to peak with both 1 and 5 pmol/l tissue factor were independent prognostic markers. ETP and peak thrombin well reflect DIC severity. Hypocoagulability manifesting as prolonged lag time and time to peak is expected to be an independent prognostic marker in DIC.

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

    Science.gov (United States)

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

    2014-11-01

    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 EGFR phosphorylation by 8.5 ± 1.3-fold (p EGFR tyrosine kinase irreversible inhibitor, 10 µM PD169540 (PD), and 20 µM anti-HB-EGF antibody significantly reduced thrombin-stimulated EGFR and ERK1/2 phosphorylation by 81, 72 % and by 48 and 61 %, respectively. Furthermore, the same pretreatments with PD or anti-HB-EGF antibody reduced thrombin-induced VSMC proliferation by 44 and 45 %, respectively. In addition, 30-min pretreatments with 10 µM specific MMP-2 inhibitor significantly reduced thrombin-stimulated phosphorylation of both EGFR and ERK1/2 by 25 %. Moreover, the same pretreatment with MMP-2 inhibitor reduced thrombin-induced VSMC proliferation by 45 %. These results show that the thrombin-induced 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.

  13. 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

  14. The thrombin E192Q-BPTI complex reveals gross structural rearrangements: implications for the interaction with antithrombin and thrombomodulin.

    OpenAIRE

    van de Locht, A; Bode, W.; Huber, R; Le Bonniec, B F; Stone, S R; Esmon, C T; Stubbs, M T

    1997-01-01

    Previous crystal structures of thrombin indicate that the 60-insertion loop is a rigid moiety that partially occludes the active site, suggesting that this structural feature plays a decisive role in restricting thrombin's specificity. This restricted specificity is typified by the experimental observation that thrombin is not inhibited by micromolar concentrations of basic pancreatic trypsin inhibitor (BPTI). Surprisingly, a single atom mutation in thrombin (E192Q) results in a 10(-8) M affi...

  15. [Topical contraceptives].

    Science.gov (United States)

    Alipov, V I; Korkhov, V V

    1982-02-01

    Recently there has been little interest in topical contraceptives. The most popular are the cervical cap and the diaphragm. Other types of mechanical contraceptive devices are being investigated. Standley and Kessler have developed a device for introduction into the cervical canal with a reservoir of spermatocide, it does not block the flow of blood during menstruation. New models of vaginal rings are also being developed which are simple enough for self-insertion and also contain a reservoir of spermatocide. Work is being done on spermatocide-containing sponges in many countries. Another project being investigated is the possibility of using natural proteins, collagens, and other substances which absorb spermatozoids. The ancients used various vaginal suppositories to kill spermatozoids; in the late 19th century quinine sulfate was used for this, and a variety of substances have been used recently. These spermicidal creams also have the advantage of acting as anti-infectious agents in many cases. But they do have some negative effects. They are about 85% effective, are local irritants, and some cause discomfort during intercourse. And it is possible that some are resorbed by the body and act on the liver and other organs. Vaginal globules and suppositories are also popular. The "Kontraceptin-T" brand contains quinosol, boric acid, and tannin. There are also foaming tablets which are mixed with water and then introduced. New locally-active chemical substances are being developed in Japan, West Germany, and the USSR. Kontraceptin-E contains paranonyl-phenoxypolyethylene glycol and sodium dioctylsulfosuccinate. The "Norforks" and other preparations contain mercurial compounds which may turn out to be harmful. The future promises the development of products which will act to prevent fertilization by acting on the hyaluronidase and the acrosine of the spermatozoid, thus preventing it from penetrating the ovum. It would be best to find enzyme inhibitors which are

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

    Science.gov (United States)

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

    2013-06-01

    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 Aα- and Bβ-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 γA/γA isoform of fibrin(ogen) and the γA/γ' variant with an extended γ-chain. Thrombin binds to the γ'-chain and forms a higher affinity interaction with γA/γ'-fibrin(ogen) than γA/γA-fibrin(ogen). In contrast, batroxobin binds both fibrin(ogen) isoforms with similar high affinity (Kd values of about 0.5 μM) even though it does not interact with the γ'-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 γA/γA-fibrinogen with batroxobin. Furthermore, although both thrombin and batroxobin bind to the central E-region of fibrinogen with a Kd value of 2-5 μM, the α(17-51) and Bβ(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

  17. Safety and efficacy of recombinant Hirudin (CGP 39 393) versus Heparin in patients with stable angina undergoing coronary angioplasty

    NARCIS (Netherlands)

    A.A. van den Bos (Arjan); G.R. Heyndrickx (Guy); G-J. Laarman (GertJan); H. Suryapranata (Harry); F. Zijlstra (Felix); P. Close (Philip); J.J.M.M. Rijnierse (Joep); H.R. Büller (Harry); P.W.J.C. Serruys (Patrick); J.W. Deckers (Jaap)

    1993-01-01

    textabstractBACKGROUND. Enhanced thrombin activity has been associated with acute and long-term complications following balloon angioplasty (percutaneous transluminal coronary angioplasty (PTCA). We evaluated, in a 2-to-1 randomized, double-blind trial, the effects of recombinant hirudin, CGP 39 393

  18. Loop Electrostatics Asymmetry Modulates the Preexisting Conformational Equilibrium in Thrombin.

    Science.gov (United States)

    Pozzi, Nicola; Zerbetto, Mirco; Acquasaliente, Laura; Tescari, Simone; Frezzato, Diego; Polimeno, Antonino; Gohara, David W; Di Cera, Enrico; De Filippis, Vincenzo

    2016-07-19

    Thrombin exists as an ensemble of active (E) and inactive (E*) conformations that differ in their accessibility to the active site. Here we show that redistribution of the E*-E equilibrium can be achieved by perturbing the electrostatic properties of the enzyme. Removal of the negative charge of the catalytic Asp102 or Asp189 in the primary specificity site destabilizes the E form and causes a shift in the 215-217 segment that compromises substrate entrance. Solution studies and existing structures of D102N document stabilization of the E* form. A new high-resolution structure of D189A also reveals the mutant in the collapsed E* form. These findings establish a new paradigm for the control of the E*-E equilibrium in the trypsin fold. PMID:27347732

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

    LENUS (Irish Health Repository)

    Ni Ainle, Fionnuala

    2009-08-20

    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.

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

    Science.gov (United States)

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

    1998-07-01

    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

  1. The modification of the thrombin generation test for the clinical assessment of dabigatran etexilate efficiency.

    Science.gov (United States)

    Gribkova, Irina V; Lipets, Elena N; Rekhtina, Irina G; Bernakevich, Alex I; Ayusheev, Dorzho B; Ovsepyan, Ruzanna A; Ataullakhanov, Fazoil I; Sinauridze, Elena I

    2016-01-01

    A new oral anticoagulant, dabigatran etexilate (DE, a prodrug of direct thrombin inhibitor (DTI) dabigatran), has been used clinically to prevent thrombosis. The assessment of dabigatran efficiency is necessary in some clinical cases, such as renal insufficiency, risk of bleeding, and drug interactions. However, a specific thrombin generation test (TGT) that is one of the most informative and sensitive to anticoagulant therapy (calibrated automated thrombinography (САТ)) shows a paradoxical increase of test parameters, such as endogenous thrombin potential (ETP) and peak thrombin, in patients receiving DE. The paradoxical behaviour of ETP and peak thrombin in these patients in the presence of DTIs is mostly caused by a decrease in the activity of thrombin in the α2-macroglobulin-thrombin complex that is used as a calibrator in CAT. For a correct estimation of the TGT parameters in patient's plasma containing DTIs we proposed to use our previously described alternative calibration method that is based on the measurement of the fluorescence signal of a well-known concentration of the reaction product (7-amino-4-methylcoumarin). In this study, the validity of such approach was demonstrated in an ex vivo study in patients with knee replacement and two special patients with multiple myeloma, who received DE for thrombosis prophylaxis. PMID:27377013

  2. The modification of the thrombin generation test for the clinical assessment of dabigatran etexilate efficiency

    Science.gov (United States)

    Gribkova, Irina V.; Lipets, Elena N.; Rekhtina, Irina G.; Bernakevich, Alex I.; Ayusheev, Dorzho B.; Ovsepyan, Ruzanna A.; Ataullakhanov, Fazoil I.; Sinauridze, Elena I.

    2016-01-01

    A new oral anticoagulant, dabigatran etexilate (DE, a prodrug of direct thrombin inhibitor (DTI) dabigatran), has been used clinically to prevent thrombosis. The assessment of dabigatran efficiency is necessary in some clinical cases, such as renal insufficiency, risk of bleeding, and drug interactions. However, a specific thrombin generation test (TGT) that is one of the most informative and sensitive to anticoagulant therapy (calibrated automated thrombinography (САТ)) shows a paradoxical increase of test parameters, such as endogenous thrombin potential (ETP) and peak thrombin, in patients receiving DE. The paradoxical behaviour of ETP and peak thrombin in these patients in the presence of DTIs is mostly caused by a decrease in the activity of thrombin in the α2-macroglobulin-thrombin complex that is used as a calibrator in CAT. For a correct estimation of the TGT parameters in patient’s plasma containing DTIs we proposed to use our previously described alternative calibration method that is based on the measurement of the fluorescence signal of a well-known concentration of the reaction product (7-amino-4-methylcoumarin). In this study, the validity of such approach was demonstrated in an ex vivo study in patients with knee replacement and two special patients with multiple myeloma, who received DE for thrombosis prophylaxis. PMID:27377013

  3. Quantitative phosphoproteomics unveils temporal dynamics of thrombin signaling in human endothelial cells.

    Science.gov (United States)

    van den Biggelaar, Maartje; Hernández-Fernaud, Juan Ramon; van den Eshof, Bart L; Neilson, Lisa J; Meijer, Alexander B; Mertens, Koen; Zanivan, Sara

    2014-03-20

    Thrombin is the key serine protease of the coagulation cascade and a potent trigger of protease-activated receptor 1 (PAR1)-mediated platelet aggregation. In recent years, PAR1 has become an appealing target for anticoagulant therapies. However, the inhibitors that have been developed so far increase bleeding risk in patients, likely because they interfere with endogenous PAR1 signaling in the endothelium. Because of its complexity, thrombin-induced signaling in endothelial cells has remained incompletely understood. Here, we have combined stable isotope amino acids in cell culture, affinity-based phosphopeptide enrichment, and high-resolution mass spectrometry and performed a time-resolved analysis of the thrombin-induced signaling in human primary endothelial cells. We identified 2224 thrombin-regulated phosphorylation sites, the majority of which have not been previously related to thrombin. Those sites were localized on proteins that are novel to thrombin signaling, but also on well-known players such as PAR1, Rho-associated kinase 2, phospholipase C, and proteins related to actin cytoskeleton, cell-cell junctions, and Weibel-Palade body release. Our study provides a unique resource of phosphoproteins and phosphorylation sites that may generate novel insights into an intimate understanding of thrombin-mediated PAR signaling and the development of improved PAR1 antagonists that affect platelet but not endothelial cell function. PMID:24501219

  4. STIM1 and Orai1 mediate thrombin-induced Ca(2+) influx in rat cortical astrocytes.

    Science.gov (United States)

    Moreno, Claudia; Sampieri, Alicia; Vivas, Oscar; Peña-Segura, Claudia; Vaca, Luis

    2012-12-01

    In astrocytes, thrombin leads to cytoplasmic Ca(2+) elevations modulating a variety of cytoprotective and cytotoxic responses. Astrocytes respond to thrombin stimulation with a biphasic Ca(2+) increase generated by an interplay between ER-Ca(2+) release and store-operated Ca(2+) entry (SOCE). In many cell types, STIM1 and Orai1 have been demonstrated to be central components of SOCE. STIM1 senses the ER-Ca(2+) depletion and binds Orai1 to activate Ca(2+) influx. Here we used immunocytochemistry, overexpression and siRNA assays to investigate the role of STIM1 and Orai1 in the thrombin-induced Ca(2+) response in primary cultures of rat cortical astrocytes. We found that STIM1 and Orai1 are endogenously expressed in cortical astrocytes and distribute accordingly with other mammalian cells. Importantly, native and overexpressed STIM1 reorganized in puncta under thrombin stimulation and this reorganization was reversible. In addition, the overexpression of STIM1 and Orai1 increased by twofold the Ca(2+) influx evoked by thrombin, while knockdown of endogenous STIM1 and Orai1 significantly decreased this Ca(2+) influx. These results indicate that STIM1 and Orai1 underlie an important fraction of the Ca(2+) response that astrocytes exhibit in the presence of thrombin. Thrombin stimulation in astrocytes leads to ER-Ca(2+) release which causes STIM1 reorganization allowing the activation of Orai1 and the subsequent Ca(2+) influx.

  5. Topical haemostatic agents for skin wounds: a systematic review

    Directory of Open Access Journals (Sweden)

    Ubbink Dirk T

    2011-07-01

    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.

  6. Thrombostatin FM compounds: direct thrombin inhibitors - mechanism of action in vitro and in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Nieman, M T; Burke, F; Warnock, M; Zhou, Y; Sweigart, J; Chen, A; Ricketts, D; Lucchesi, B R; Chen, Z; Cera, E Di; Hilfinger, J; Kim, J S; Mosberg, H I; Schmaier, A H [Case Western; (Michigan); (TSRL); (WU-MED)

    2008-04-29

    Novel pentapeptides called Thrombostatin FM compounds consisting mostly of D-isomers and unusual amino acids were prepared based upon the stable angiotensin converting enzyme breakdown product of bradykinin - RPPGF. These peptides are direct thrombin inhibitors prolonging the thrombin clotting time, activated partial thromboplastin time, and prothrombin time at ≥0.78, 1.6, and 1.6 μm, respectively. They competitively inhibit α-thrombin-induced cleavage of a chromogenic substrate at 4.4--8.2 μm. They do not significantly inhibit plasma kallikrein, factor (F) XIIa, FXIa, FIXa, FVIIa-TF, FXa, plasmin or cathepsin G. One form, FM19 [rOicPaF(p-Me)], blocks α-thrombin-induced calcium flux in fibroblasts with an IC50 of 6.9 ± 1.2 μm. FM19 achieved 100% inhibition of threshold α- or γ-thrombin-induced platelet aggregation at 8.4 ± 4.7 μm and 16 ± 4 μm, respectively. The crystal structure of thrombin in complex with FM19 shows that the N-terminal D-Arg retrobinds into the S1 pocket, its second residue Oic interacts with His-57, Tyr-60a and Trp-60d, and its C-terminal p-methyl Phe engages thrombin's aryl binding site composed of Ile-174, Trp-215, and Leu-99. When administered intraperitoneal, intraduodenal, or orally to mice, FM19 prolongs thrombin clotting times and delays carotid artery thrombosis. FM19, a low affinity reversible direct thrombin inhibitor, might be useful as an add-on agent to address an unmet need in platelet inhibition in acute coronary syndromes in diabetics and others who with all current antiplatelet therapy still have reactive platelets.

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

    DEFF Research Database (Denmark)

    Pasternak, Anna; Hernandez, Frank J; Rasmussen, Lars Melholt;

    2011-01-01

    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...... 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...

  8. Fluorescent reporters of thrombin, heparin cofactor II, and heparin binding in a ternary complex

    OpenAIRE

    Verhamme, Ingrid M.

    2011-01-01

    Thrombin inactivation by heparin cofactor II (HCII) is accelerated by ternary complex formation with heparin. The novel active-site-labeled thrombins, [4′F]FPR-T and [6F]FFR-T, and the exosite I probe, Hir-(54–65)( SO3−), characterized thrombin exosite I and II interactions with HCII and heparin in the complex. HCII binding to exosite I of heparin-bound [4′F]FPR-T caused a saturable fluorescence increase, absent with antithrombin. Heparin binding to exosite II and a second weaker site caus...

  9. Crystallization and preliminary X-ray analysis of the complex of human α-thrombin with a modified thrombin-binding aptamer

    International Nuclear Information System (INIS)

    The complex between human α-thrombin and a modified thrombin-binding aptamer has been crystallized. Diffraction data were collected to 2.15 Å resolution, the structure was solved by molecular replacement and refinement of the model is in progress. The thrombin-binding aptamer (TBA) is a consensus DNA 15-mer that binds specifically to human α-thrombin at nanomolar concentrations and inhibits its procoagulant functions. Recently, a modified TBA (mTBA) containing a 5′–5′ inversion-of-polarity site has been shown to be more stable and to possess a higher thrombin affinity than its unmodified counterpart. The structure of the thrombin–TBA complex has previously been determined at low resolution, but did not provide a detailed picture of the aptamer conformation or of the protein–DNA assembly, while that of the complex with mTBA is unknown. Crystallographic analysis of the thrombin–mTBA complex has been attempted. The crystals diffracted to 2.15 Å resolution and belonged to space group I222

  10. Fibrinolytic action of an enzyme preparation covalently bound with modified thrombin

    Energy Technology Data Exchange (ETDEWEB)

    Maksimenko, A.V.; Rusetskii, A.N.; Torchilin, V.P.

    1987-06-01

    It was pointed out previously that modification of thrombin at the tryptophan, tyrosine, arginine and lysine residues impairs its affinity for fibrinogen. Since a long binding site of macromolecular substrate in the thrombin molecule is responsible for binding of the enzyme with the platelet membrane also, it is probably preferable to carry out the modification at other amino acid residues. The purpose of this paper was to confirm experimentally the validity of this approach to the targeted modification of alpha-thrombin in order to obtain a protein polymer matrix with affinity for centers of thrombus formation. Technetium 99m was used as a label in assessing the ability of the modified thrombin to destroy the fibrin clot.

  11. Reduced activity of TAFI (thrombin-activatable fibrinolysis inhibitor) in acute promyelocytic leukaemia

    NARCIS (Netherlands)

    Meijers, JCM; Oudijk, EJD; Mosnier, LO; Nieuwenhuis, HK; Fijnheer, R; Bouma, Bonno N.; Bos, R

    2000-01-01

    Acute promyelocytic leukaemia (APL) is a disease that is distinguished from other leukaemias by the high potential for early haemorrhagic death. Several processes are involved, such as disseminated intravascular coagulation and hyperfibrinolysis. Recently, TAFI (thrombin-activatable fibrinolysis inh

  12. 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)

  13. Modeling the microscopic electrical properties of thrombin binding aptamer (TBA) for label-free biosensors

    CERN Document Server

    Alfinito, Eleonora; Cataldo, Rosella; De Nunzio, Giorgio; Giotta, Livia; Guascito, Maria Rachele

    2016-01-01

    Aptamers are chemically produced oligonucleotides, able to bind a variety of targets such as drugs, proteins and pathogens with high sensitivity and selectivity. Therefore, aptamers are largely employed for producing label-free biosensors, with significant applications in diagnostics and drug delivery. In particular, the anti-thrombin aptamers are biomolecules of high interest for clinical use, because of their ability to recognize and bind the thrombin enzyme. Among them, the DNA 15-mer thrombin-binding aptamer (TBA), has been widely explored concerning both its structure, which was resolved with different techniques, and its function, especially about the possibility of using it as the active part of biosensors. This paper proposes a microscopic model of the electrical properties of TBA and the aptamer-thrombin complex, combining information from both structure and function. The novelty consists in describing both the aptamer alone and the complex as an impedance network, thus going deeper inside the issues...

  14. Membrane lipid peroxidation in neurodegeneration: Role of thrombin and proteinase-activated receptor-1.

    Science.gov (United States)

    Citron, Bruce A; Ameenuddin, Syed; Uchida, K; Suo, William Z; SantaCruz, Karen; Festoff, Barry W

    2016-07-15

    Thrombin and membrane lipid peroxidation (MLP) have been implicated in various central nervous system (CNS) disorders from CNS trauma to stroke, Alzheimer's (AD) and Parkinson's (PD) diseases. Because thrombin also induces MLP in platelets and its involvement in neurodegenerative diseases we hypothesized that its deleterious effects might, in part, involve formation of MLP in neuronal cells. We previously showed that thrombin induced caspase-3 mediated apoptosis in motor neurons, via a proteinase-activated receptor (PAR1). We have now investigated thrombin's influence on the oxidative state of neurons leading to induction of MLP-protein adducts. Translational relevance of thrombin-induced MLP is supported by increased levels of 4-hydroxynonenal-protein adducts (HNEPA) in AD and PD brains. We now report for the first time that thrombin dose-dependently induces formation of HNEPA in NSC34 mouse motor neuron cells using anti-HNE and anti-acrolein monoclonal antibodies. The most prominent immunoreactive band, in SDS-PAGE, was at ∼54kDa. Membrane fractions displayed higher amounts of the protein-adduct than cytosolic fractions. Thrombin induced MLP was mediated, at least in part, through PAR1 since a PAR1 active peptide, PAR1AP, also elevated HNEPA levels. Of interest, glutamate and Fe2SO4 also increased the ∼54kDa HNEPA band in these cells but to a lesser extent. Taken together our results implicate the involvement of thrombin and MLP in neuronal cell loss observed in various CNS degenerative and traumatic pathologies. PMID:27138068

  15. Gold nanoparticles doped conducting polymer nanorod electrodes: ferrocene catalyzed aptamer-based thrombin immunosensor.

    Science.gov (United States)

    Rahman, Md Aminur; Son, Jung Ik; Won, Mi-Sook; Shim, Yoon-Bo

    2009-08-15

    Au nanoparticles-doped conducting polymer nanorods electrodes (AuNPs/CPNEs) were prepared by coating Au nanorods (AuNRs) with a conducting polymer layer. The AuNRs were prepared through an electroless deposition method using the polycarbonate membrane (pore diameter, 50 nm, pore density, 6 x 10(8) pores/cm(2)) as a template. The AuNPs/CPNEs combining catalytic activity of ferrocene to ascorbic acid were used for the fabrication of an ultrasensitive aptamer sensor for thrombin detection. The AuNPs/3D-CPNEs were characterized employing cyclic voltammetry (CV), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and atomic force microscopy (AFM). Sandwiched immunoassay for alpha-human thrombin with NH(2)-functionalized-thrombin binding aptamer (Apt) immobilized on AuNPs/3D-CPNEs was studied through the electrocatalytic oxidation of ascorbic acid by the ferrocene moiety that was bound with an antithrombin antibody and attached with the Apt/3D-CPNEs probe through target binding. Various experimental parameters affecting thrombin detection were optimized, and the performance of the thrombin aptamer sensor was examined. The Apt/AuNPs/3D-CPNEs based thrombin sensor exhibited a wide dynamic range of 5-2000 ng L(-1) and a low detection limit of 5 ng L(-1) (0.14 pM). The selectivity and the stability of the proposed thrombin aptamer sensor were excellent, and it was tested in a real human serum sample for the detection of spiked concentrations of thrombin. PMID:20337374

  16. Thrombin Binding Aptamer, More than a Simple Aptamer: Chemically Modified Derivatives and Biomedical Applications

    OpenAIRE

    Aviñó, Anna Maria; Eritja Casadellà, Ramón; Fàbrega, Carme; Tintoré, María

    2012-01-01

    The thrombin binding aptamer (TBA) is a well characterized chair-like, antiparallel quadruplex structure that binds specifically to thrombin at nanomolar concentrations and therefore it has interesting anticoagulant properties. In this article we review the research involved in the development of new TBA derivatives with improved anticoagulant properties as well as the use of the TBA as a model compound for the study of quadruplex structures. Specifically, we describe the impact of modified n...

  17. The modification of the thrombin generation test for the clinical assessment of dabigatran etexilate efficiency

    OpenAIRE

    Gribkova, Irina V.; Lipets, Elena N; Rekhtina, Irina G.; Alex I. Bernakevich; Ayusheev, Dorzho B.; Ruzanna A. Ovsepyan; Ataullakhanov, Fazoil I.; Elena I Sinauridze

    2016-01-01

    A new oral anticoagulant, dabigatran etexilate (DE, a prodrug of direct thrombin inhibitor (DTI) dabigatran), has been used clinically to prevent thrombosis. The assessment of dabigatran efficiency is necessary in some clinical cases, such as renal insufficiency, risk of bleeding, and drug interactions. However, a specific thrombin generation test (TGT) that is one of the most informative and sensitive to anticoagulant therapy (calibrated automated thrombinography (САТ)) shows a paradoxical i...

  18. Thrombin stimulates albumin transcytosis in lung microvascular endothelial cells via activation of acid sphingomyelinase.

    Science.gov (United States)

    Kuebler, Wolfgang M; Wittenberg, Claudia; Lee, Warren L; Reppien, Eike; Goldenberg, Neil M; Lindner, Karsten; Gao, Yizhuo; Winoto-Morbach, Supandi; Drab, Marek; Mühlfeld, Christian; Dombrowsky, Heike; Ochs, Matthias; Schütze, Stefan; Uhlig, Stefan

    2016-04-15

    Transcellular albumin transport occurs via caveolae that are abundant in lung microvascular endothelial cells. Stimulation of albumin transcytosis by proinflammatory mediators may contribute to alveolar protein leak in lung injury, yet the regulation of albumin transport and its underlying molecular mechanisms are so far incompletely understood. Here we tested the hypothesis that thrombin may stimulate transcellular albumin transport across lung microvascular endothelial cells in an acid-sphingomyelinase dependent manner. Thrombin increased the transport of fluorescently labeled albumin across confluent human lung microvascular endothelial cell (HMVEC-L) monolayers to an extent that markedly exceeds the rate of passive diffusion. Thrombin activated acid sphingomyelinase (ASM) and increased ceramide production in HMVEC-L, but not in bovine pulmonary artery cells, which showed little albumin transport in response to thrombin. Thrombin increased total caveolin-1 (cav-1) content in both whole cell lysates and lipid rafts from HMVEC-L, and this effect was blocked by inhibition of ASM or de novo protein biosynthesis. Thrombin-induced uptake of albumin into lung microvascular endothelial cells was confirmed in isolated-perfused lungs by real-time fluorescence imaging and electron microscopy of gold-labeled albumin. Inhibition of ASM attenuated thrombin-induced albumin transport both in confluent HMVEC-L and in intact lungs, whereas HMVEC-L treatment with exogenous ASM increased albumin transport and enriched lipid rafts in cav-1. Our findings indicate that thrombin stimulates transcellular albumin transport in an acid sphingomyelinase-dependent manner by inducing de novo synthesis of cav-1 and its recruitment to membrane lipid rafts. PMID:26851257

  19. Thrombin-linked aptamer assay for detection of platelet derived growth factor BB on magnetic beads in a sandwich format.

    Science.gov (United States)

    Guo, Limin; Zhao, Qiang

    2016-09-01

    Here we describe a thrombin-linked aptamer assay (TLAA) for protein by using thrombin as an enzyme label, harnessing enzyme activity of thrombin and aptamer affinity binding. TLAA converts detection of specific target proteins to the detection of thrombin by using a DNA sequence that consists of two aptamers with the first aptamer binding to the specific target protein and the second aptamer binding to thrombin. Through the affinity binding, the thrombin enzyme is labeled on the protein target, and thrombin catalyzes the hydrolysis of small peptide substrate into product, generating signals for quantification. As a proof of principle, we show a sandwich TLAA for platelet derived growth factor BB (PDGF-BB) by using anti-PDGF-BB antibody coated on magnetic beads and an oligonucleotide containing the aptamer for PDGF-BB and the aptamer for thrombin. The binding of PDGF-BB to both the antibody and the aptamer results in labeling the complex with thrombin. We achieved detection of PDGF-BB at 16 pM. This TLAA contributes a new application of thrombin and its aptamer in bioanalysis, and shows potentials in assay developments. PMID:27343590

  20. Study on an electrochemical biosensor for thrombin recognition based on aptamers and nano particles

    Institute of Scientific and Technical Information of China (English)

    ZHENG Jing; LIN Li; CHENG GuiFang; WANG AnBao; TAN XueLian; HE PinGang; FANG YuZhi

    2007-01-01

    This paper presents a high specific, sensitive electrochemical biosensor for recognition of protein such as thrombin based on aptamers and nano particles. Two different aptamers were chosen to construct a sandwich manner for detecting thrombin. Aptamer Ⅰ was immobilized on nano magnetic particle for capturing thrombin, and aptamer Ⅱ labled with nano gold was used for detection. The electrical current generated from gold after the formation of the complex of magnetic particle, thrombin and nano gold,and then an electrochemical cell designed by ourselves was used for separating, gathering, and electrochemical detecting. Through magnetic separation, high specific and sensitive detection of the target protein, thrombin, was achieved. Linear response was observed over the range 5.6×10-12-1.12×10-9mol/L, with a detection limit of 1.42×10-12 mol/L. The presence of other protein as BSA did not affect the detection, which indicates that high selective recognition of thrombin can be achieved in complex biological samples such as human plasma.

  1. Thrombin Production and Human Neutrophil Elastase Sequestration by Modified Cellulosic Dressings and Their Electrokinetic Analysis

    Directory of Open Access Journals (Sweden)

    Nicolette Prevost

    2011-12-01

    Full Text Available Wound healing is a complex series of biochemical and cellular events. Optimally, functional material design addresses the overlapping acute and inflammatory stages of wound healing based on molecular, cellular, and bio-compatibility issues. In this paper the issues addressed are uncontrolled hemostasis and inflammation which can interfere with the orderly flow of wound healing. In this regard, we review the serine proteases thrombin and elastase relative to dressing functionality that improves wound healing and examine the effects of charge in cotton/cellulosic dressing design on thrombin production and elastase sequestration (uptake by the wound dressing. Thrombin is central to the initiation and propagation of coagulation, and elastase is released from neutrophils that can function detrimentally in a stalled inflammatory phase characteristic of chronic wounds. Electrokinetic fiber surface properties of the biomaterials of this study were determined to correlate material charge and polarity with function relative to thrombin production and elastase sequestration. Human neutrophil elastase sequestration was assessed with an assay representative of chronic wound concentration with cotton gauze cross-linked with three types of polycarboxylic acids and one phosphorylation finish; thrombin production, which was assessed in a plasma-based assay via a fluorogenic peptide substrate, was determined for cotton, cotton-grafted chitosan, chitosan, rayon/polyester, and two kaolin-treated materials including a commercial hemorrhage control dressing (QuickClot Combat Gauze. A correlation in thrombin production to zeta potential was found. Two polycarboxylic acid cross linked and a phosphorylated cotton dressing gave high elastase sequestration.

  2. Targeting thrombin long-term after an acute coronary syndrome: Opportunities and challenges.

    Science.gov (United States)

    De Caterina, Raffaele; Goto, Shinya

    2016-06-01

    Patients after an acute coronary syndrome (ACS) are at increased risk of recurrent thrombotic events, justifying the search for additional antithrombotic treatments. The pathophysiology of ACS involves arterial thrombus formation, in turn occurring because of a combination of platelet activation and fibrin formation, with thrombin playing a key role in both. Antiplatelet therapy, targeting the thromboxane pathway and the ADP P2Y12 receptor has been widely accepted for secondary prevention after an ACS. Now, data from recent clinical trials in such patients also encourage the pursuit of inhibiting thrombin formation or thrombin-mediated platelet activation in addition to antiplatelet therapy. This "triple pathway inhibition", including inhibition of thrombin activity or thrombin receptor(s), is currently an option in pure ACS, but already a must in the setting of ACS accompanied by atrial fibrillation (AF), where anticoagulants have been shown to be much more effective than antiplatelet agents in preventing stroke. We here discuss the challenges of managing combined thrombin activity or receptor inhibition and antiplatelet therapy in all such patients. Translating this into practice still requires further studies and patient tailoring to fully exploit its potential. PMID:26994821

  3. Study on an electrochemical biosensor for thrombin recognition based on aptamers and nano particles

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    This paper presents a high specific, sensitive electrochemical biosensor for recognition of protein such as thrombin based on aptamers and nano particles. Two different aptamers were chosen to construct a sandwich manner for detecting thrombin. Aptamer I was immobilized on nano magnetic particle for capturing thrombin, and aptamer II labled with nano gold was used for detection. The electrical current generated from gold after the formation of the complex of magnetic particle, thrombin and nano gold, and then an electrochemical cell designed by ourselves was used for separating, gathering, and electrochemical detecting. Through magnetic separation, high specific and sensitive detection of the target protein, thrombin, was achieved. Linear response was observed over the range 5.6×10-12―1.12×10-9 mol/L, with a detection limit of 1.42×10-12 mol/L. The presence of other protein as BSA did not affect the detection, which indicates that high selective recognition of thrombin can be achieved in complex biological samples such as human plasma.

  4. Expression of thrombin and its associated protein in cerebellum of human and rat after intracerebral hemorrhage

    Institute of Scientific and Technical Information of China (English)

    ZHANG Zhi-yi; QI Ji-ping; ZHU Hong; SONG Yue-jia; WU He; JIA Ying; ZHANG Guang-mei

    2010-01-01

    Background Intracerebral hemorrhage (ICH) can cause brain damage through a number of pathways.The purpose of the study was to explore the effect of thrombin, protease nexin-1 (PN-1) and protease activated receptor-1 (PAR-1) in rat and human cerebellum after ICH.Methods A model of ICH was produced in adult Sprague-Dawley rats by direct injection of autologous blood (50 μl) into caudate nucleus.Patients with injured hemorrhage were also enrolled in this study.Different expressions of thrombin,PAR-1, PN-1 were detected in rat and human cerebellum by immunohistochemistry and in situ hybridization.Results In rat cerebellum, thrombin protein significantly increased at 6 hours and reached the maximum 2 days afterICH.The expression of PAR-1 protein reached the maximum at 24-48 hours, and then began to decrease.The expression of PN-1 protein reached the maximum at 3 hours, decreased somewhat after that and increased a little at 5days after ICH.While in human cerebellum, the changing tendency of thrombin, PAR-1 and PN-1 was almost conform to the rat.Conclusion In cerebellum, thrombin can activate PAR-1 expression after ICH, and PN-1 appears quickly after ICH in order to control the deleterious effect of thrombin.

  5. The direct thrombin inhibitors (argatroban, bivalirudin and lepirudin) and the indirect Xa-inhibitor (danaparoid) increase fibrin network porosity and thus facilitate fibrinolysis.

    Science.gov (United States)

    He, Shu; Blombäck, Margareta; Bark, Niklas; Johnsson, Hans; Wallén, N Hakan

    2010-05-01

    The present study aimed to assess whether the fibrin network structure is modified by the direct thrombin-inhibitors lepirudin, argatroban or bivalirudin and by the indirect Xa-inhibitor danaparoid. Using an in vitro assay that imitates the physiological process of coagulation from thrombin generation to fibrin formation, we examined a normal plasma pool spiked with one of the inhibitors. At concentrations considered to be the plasma levels observed during therapy, almost no influence was detected for lepirudin despite clear-cut effects on "clotting time". However, argatroban, bivalirudin and danaparoid increased the fibrin gel permeability (Ks) to a similar extent. At concentrations higher than the "therapeutic" levels, the dose-response curve in the Ks assay became very steep for lepirudin while those were shallow for the others. In parallel with the drug-induced increases of Ks, larger network pores in 3D-microscopic images and significant shortenings in "clot lysis time" induced by addition of rtPA were observed. Recombinant factor VIII (rFVIII) added to danaparoid-treated samples profoundly counteracted the increase of Ks but had only a slight or no effect on the other drugs. Thus, in vitro, argatroban, bivalirudin and danaparoid have comparable anticoagulating effects, rendering the fibrin network more permeable and less resistant to fibrinolysis. For lepirudin, the steep dose-response curve supports previous clinical findings, i.e. this thrombin inhibitor has a narrow therapeutic window. Furthermore, our data suggest that the haemostatic agent, rFVIII, might be effective in treatment of bleeding complications induced by danaparoid. PMID:20216982

  6. Genetic and pharmacological modifications of thrombin formation in apolipoprotein e-deficient mice determine atherosclerosis severity and atherothrombosis onset in a neutrophil-dependent manner.

    Directory of Open Access Journals (Sweden)

    Julian I Borissoff

    Full Text Available BACKGROUND: Variations in the blood coagulation activity, determined genetically or by medication, may alter atherosclerotic plaque progression, by influencing pleiotropic effects of coagulation proteases. Published experimental studies have yielded contradictory findings on the role of hypercoagulability in atherogenesis. We therefore sought to address this matter by extensively investigating the in vivo significance of genetic alterations and pharmacologic inhibition of thrombin formation for the onset and progression of atherosclerosis, and plaque phenotype determination. METHODOLOGY/PRINCIPAL FINDINGS: We generated transgenic atherosclerosis-prone mice with diminished coagulant or hypercoagulable phenotype and employed two distinct models of atherosclerosis. Gene-targeted 50% reduction in prothrombin (FII(-/WT:ApoE(-/- was remarkably effective in limiting disease compared to control ApoE(-/- mice, associated with significant qualitative benefits, including diminished leukocyte infiltration, altered collagen and vascular smooth muscle cell content. Genetically-imposed hypercoagulability in TM(Pro/Pro:ApoE(-/- mice resulted in severe atherosclerosis, plaque vulnerability and spontaneous atherothrombosis. Hypercoagulability was associated with a pronounced neutrophilia, neutrophil hyper-reactivity, markedly increased oxidative stress, neutrophil intraplaque infiltration and apoptosis. Administration of either the synthetic specific thrombin inhibitor Dabigatran etexilate, or recombinant activated protein C (APC, counteracted the pro-inflammatory and pro-atherogenic phenotype of pro-thrombotic TM(Pro/Pro:ApoE(-/- mice. CONCLUSIONS/SIGNIFICANCE: We provide new evidence highlighting the importance of neutrophils in the coagulation-inflammation interplay during atherogenesis. Our findings reveal that thrombin-mediated proteolysis is an unexpectedly powerful determinant of atherosclerosis in multiple distinct settings. These studies suggest that

  7. Thrombin-Binding Aptamer Quadruplex Formation: AFM and Voltammetric Characterization

    Directory of Open Access Journals (Sweden)

    Victor Constantin Diculescu

    2010-01-01

    Full Text Available The adsorption and the redox behaviour of thrombin-binding aptamer (TBA and extended TBA (eTBA were studied using atomic force microscopy and voltammetry at highly oriented pyrolytic graphite and glassy carbon. The different adsorption patterns and degree of surface coverage were correlated with the sequence base composition, presence/absence of K+, and voltammetric behaviour of TBA and eTBA. In the presence of K+, only a few single-stranded sequences present adsorption, while the majority of the molecules forms stable and rigid quadruplexes with no adsorption. Both TBA and eTBA are oxidized and the only anodic peak corresponds to guanine oxidation. Upon addition of K+ ions, TBA and eTBA fold into a quadruplex, causing the decrease of guanine oxidation peak and occurrence of a new peak at a higher potential due to the oxidation of G-quartets. The higher oxidation potential of G-quartets is due to the greater difficulty of electron transfer from the inside of the quadruplex to the electrode surface than electron transfer from the more flexible single strands.

  8. Quantification of thymidine kinase (TK1) mRNA in normal and leukemic cells and investigation of structure-function relatiosnhip of recombinant TK1enzyme

    DEFF Research Database (Denmark)

    Kristensen, Tina

    protein was used. TKI protein is cleaved from the GST-part with thrombin. Two TKI mutants, TKI-l 93 and TKI-l 76, with deletions from the C-terminal were constructed by the recombinant PCR method. Deletion of 57 amino acids from the C-terminal (TKI- 176) results in an inactive enzyme. Deletion of 40 amino...

  9. [Sodium ions as the effector of catalytic action of alpha-thrombin].

    Science.gov (United States)

    Kolodzeĭskaia, M V; Volkov, G L

    2007-01-01

    A process of thrombin interaction with synthetic and natural substrates in the presence of Na+ ions has been analyzed in the survey. Molecular bases of this interaction have been presented, interrelation between the structure and function of thrombin has been noted; the nature of the unique site of its active centre which determines high thrombin affinity for the substrates and increase of its catalytic activity defined by the term of "specificity to univalent cations" have been considered in detail. Na+ ions play the role of allosteric effector in realization of two informational states of thrombin which penform, respectively, two fundamental and competing functions in the process of hemostasis. The molecular basis of the process of Na+ binding with thrombin is rather simple and depends only on the single site which importance for the enzyme function is marked by numerous investigations of a number of authors, and it is shown that Na(+)-binding site is distributed in the other zone of thrombin molecule as compared to exosites I and II, which do not take part in Na(+)-binding and allosteric transduction. Considerable attention was given to conformational conversions of a thrombin molecule caused by Na+ ions binding. It was shown that the transition slow fast of the enzyme forms leads to formation of the ion pair Arg-187: Asp-222, optimal orientation of Asp-189 and Ser-195 for binding of substrates and considerable shift of the lateral chain Glu-192 determined by the disturbance of the lattice of water molecules which connects Na(+)-binding site with aminoacid Ser-195 of the active centre of the enzyme. New data have been presented which indicate that the changes in the lattice of water molecules and allosteric nucleus of Na(+)-binding site of the enzyme are the basic link of raising the affinity between the thrombin and substrate and mechanism of the enzyme activation by Na(+)-ions. The survey touches some problems of creation of allosteric inhibitors of thrombin

  10. An electrochemical label-free and sensitive thrombin aptasensor based on graphene oxide modified pencil graphite electrode.

    Science.gov (United States)

    Ahour, F; Ahsani, M K

    2016-12-15

    In this work, we tactfully constructed a novel label-free electrochemical aptasensor for rapid and facile detection of thrombin using graphene oxide (GO) and thrombin binding aptamer (TBA). The strategy relies on the preferential adsorption of single-stranded DNA (ssDNA) to GO over aptamer-target complexes. The TBA-thrombin complex formation was monitored by differential pulse voltammetry (DPV) using the guanine oxidation signal. In the absence of thrombin, the aptamers adsorbed onto the surface of GO leading to a strong background guanine oxidation signal. Conversely, in the presence of thrombin, the conformational transformation of TBA after incubating with the thrombin solution and formation of the aptamer-thrombin complexes which had weak binding ability to GO, leads to the desorption of TBA-thrombin complex from electrode surface and significant oxidation signal decrease. The selectivity of the biosensor was studied using other biological substances. The biosensor's signal was proportional to the thrombin concentration from 0.1 to 10nM with a detection limit of 0.07nM. Particularly, the proposed method could be widely applied to the aptamer-based determination of other target analytes. PMID:27476058

  11. Ultrasound-guided Thrombin Injection: An Alternative Treatment for Femoral Artery Pseudoaneurysm with Better Efficiency and Safety

    Institute of Scientific and Technical Information of China (English)

    Qinghai YAO; Hongliang CONG; Shangqin WU; Shan SUN; Qike DONG; Dongmei CHEN; Peng LI

    2008-01-01

    The aim of this study was to evaluate the efficiency and safety of ultrasound-guided thrombin injection on femoral pseudoaneurysm (FPA) as compared to ultrasound-guided local oppression. Eleven cases of FPA were enrolled and 7 cases received ultrasound-guide thrombin injection (injection group), and the remaining 4 cases were treated with local oppression (oppression group). Efficiency and safety were analyzed by ultrasound and subsequent follow-up. The results showed that 1 case relapsed in oppression group while no relapse occurred in thrombin injection group. Ultrasound-guided thrombin injection is better for treatment of FPA in terms of effectiveness and safety.

  12. Differential effects on glial activation by a direct versus an indirect thrombin inhibitor.

    Science.gov (United States)

    Marangoni, M Natalia; Braun, David; Situ, Annie; Moyano, Ana L; Kalinin, Sergey; Polak, Paul; Givogri, Maria I; Feinstein, Douglas L

    2016-08-15

    Thrombin is a potent regulator of brain function in health and disease, modulating glial activation and brain inflammation. Thrombin inhibitors, several of which are in clinical use as anti-coagulants, can reduce thrombin-dependent neuroinflammation in pathological conditions. However, their effects in a healthy CNS are largely unknown. In adult healthy mice, we compared the effects of treatment by the direct thrombin inhibitor dabigatran etexilate (DE), to those of warfarin, which acts by preventing vitamin K recycling essential for coagulation. After 4weeks, warfarin increased both astrocyte GFAP and microglia Iba-1 staining throughout the CNS; whereas DE reduced expression of both markers. Warfarin, but not DE, reduced sulfatide levels; and warfarin showed longer lasting changes in cerebellar gene expression. DE also reduced glial activation in a mouse model of Alzheimer's disease, although no changes in amyloid plaque burden were observed. These results suggest that treatment with direct thrombin inhibitors may be preferable to those agents which reduce vitamin K levels and have the potential to increase glial activation. PMID:27397090

  13. Random Forests Are Able to Identify Differences in Clotting Dynamics from Kinetic Models of Thrombin Generation.

    Science.gov (United States)

    Arumugam, Jayavel; Bukkapatnam, Satish T S; Narayanan, Krishna R; Srinivasa, Arun R

    2016-01-01

    Current methods for distinguishing acute coronary syndromes such as heart attack from stable coronary artery disease, based on the kinetics of thrombin formation, have been limited to evaluating sensitivity of well-established chemical species (e.g., thrombin) using simple quantifiers of their concentration profiles (e.g., maximum level of thrombin concentration, area under the thrombin concentration versus time curve). In order to get an improved classifier, we use a 34-protein factor clotting cascade model and convert the simulation data into a high-dimensional representation (about 19000 features) using a piecewise cubic polynomial fit. Then, we systematically find plausible assays to effectively gauge changes in acute coronary syndrome/coronary artery disease populations by introducing a statistical learning technique called Random Forests. We find that differences associated with acute coronary syndromes emerge in combinations of a handful of features. For instance, concentrations of 3 chemical species, namely, active alpha-thrombin, tissue factor-factor VIIa-factor Xa ternary complex, and intrinsic tenase complex with factor X, at specific time windows, could be used to classify acute coronary syndromes to an accuracy of about 87.2%. Such a combination could be used to efficiently assay the coagulation system. PMID:27171403

  14. Ru(bpy)32+-doped Silica Nanoparticle Aptasensor for Detection of Thrombin Based on Electrogenerated Chemiluminescence

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-Ying; YUN Wen; ZHOU Jing-Ming; DONG Ping; HE Pin-Gang; FANG Yu-Zhi

    2008-01-01

    A new electrogenerated chemiluminescent (ECL) detection system for protein using the aptamers was developed.Two different aptamers, which recognize different positions of thrombin, were chosen to construct sandwich type sensing system for protein, one was immobilized onto the gold electrode for capturing thrombin onto the electrode and the other was used for detection.To obtain the signal, the aptamer for detection was labeled with tris(2,2'-bipyridyl)ruthenium(Ⅱ)-doped silica nanoparticles ( Ru(bpy)32+-doped SNPs).The increase of the ECL signal generated by Ru(bpy)32+-doped SNPs was observed in dependent manner on the concentration of thrombin added.Bovine serum albumin and bovine hemoglobin had almost negligible responses.The assay allows detection at levels as low as 1.0 fmol·L-1 of the thrombin due to that there are a large number of Ru(bpy)32+ molecules inside SNPs labeled on the aptamer used for detection.The ECL signal was linearly related to the concentration of the thrombin analyte in the range of 2.0 fmol·L-1 to 2.0 pmol·L-1.

  15. Random Forests Are Able to Identify Differences in Clotting Dynamics from Kinetic Models of Thrombin Generation.

    Directory of Open Access Journals (Sweden)

    Jayavel Arumugam

    Full Text Available Current methods for distinguishing acute coronary syndromes such as heart attack from stable coronary artery disease, based on the kinetics of thrombin formation, have been limited to evaluating sensitivity of well-established chemical species (e.g., thrombin using simple quantifiers of their concentration profiles (e.g., maximum level of thrombin concentration, area under the thrombin concentration versus time curve. In order to get an improved classifier, we use a 34-protein factor clotting cascade model and convert the simulation data into a high-dimensional representation (about 19000 features using a piecewise cubic polynomial fit. Then, we systematically find plausible assays to effectively gauge changes in acute coronary syndrome/coronary artery disease populations by introducing a statistical learning technique called Random Forests. We find that differences associated with acute coronary syndromes emerge in combinations of a handful of features. For instance, concentrations of 3 chemical species, namely, active alpha-thrombin, tissue factor-factor VIIa-factor Xa ternary complex, and intrinsic tenase complex with factor X, at specific time windows, could be used to classify acute coronary syndromes to an accuracy of about 87.2%. Such a combination could be used to efficiently assay the coagulation system.

  16. A Universal Base in a Specific Role: Tuning up a Thrombin Aptamer with 5-Nitroindole

    Science.gov (United States)

    Tsvetkov, Vladimir B.; Varizhuk, Anna M.; Pozmogova, Galina E.; Smirnov, Igor P.; Kolganova, Natalia A.; Timofeev, Edward N.

    2015-11-01

    In this study we describe new modified analogs of the thrombin binding aptamer (TBA) containing 5-nitroindole residues. It has been shown that all modified TBAs form an anti-parallel G-quadruplex structure and retain the ability to inhibit thrombin. The most advanced TBA variant (TBA-N8) has a substantially increased clotting time and two-fold lower IC50 value compared to the unmodified prototype. Molecular modelling studies suggest that the improved anticoagulant properties of TBA-N8 result from changes in the binding mode of the analog. A modified central loop in TBA-N8 is presumed to participate in the binding of the target protein. Studies of FAM labelled TBA and TBA-N8 showed an improved binding affinity of the modified aptamer and provided evidence of a direct interaction between the modified central loop and thrombin. Our findings have implications for the design of new aptamers with improved binding affinities.

  17. Placental vascular pathology and increased thrombin generation as mechanisms of disease in obstetrical syndromes

    Directory of Open Access Journals (Sweden)

    Salvatore Andrea Mastrolia

    2014-11-01

    Full Text Available Obstetrical complications including preeclampsia, fetal growth restriction, preterm labor, preterm prelabor rupture of membranes and fetal demise are all the clinical endpoint of several underlying mechanisms (i.e., infection, inflammation, thrombosis, endocrine disorder, immunologic rejection, genetic, and environmental, therefore, they may be regarded as syndromes. Placental vascular pathology and increased thrombin generation were reported in all of these obstetrical syndromes. Moreover, elevated concentrations of thrombin-anti thrombin III complexes and changes in the coagulation as well as anticoagulation factors can be detected in the maternal circulation prior to the clinical development of the disease in some of these syndromes. In this review, we will assess the changes in the hemostatic system during normal and complicated pregnancy in maternal blood, maternal–fetal interface and amniotic fluid, and describe the contribution of thrombosis and vascular pathology to the development of the great obstetrical syndromes.

  18. A nanoresonant gold-aptamer probe for rapid and sensitive detection of thrombin

    International Nuclear Information System (INIS)

    Resonance light scattering (RLS) is a sensitive technique for monitoring scattered light induced by extended aggregates of chromophores. It has been widely used to study aggregations for its simple manipulation, high sensitivity and great versatility. Gold nanoparticles generate colorful light-scattering signals due to their unique surface plasmon resonances, hence extraordinary light scattering upon aggregation. In this paper we report a rapid and sensitive method based on gold nanoparticles and DNA aptamer to detect protein biomarkers by RLS. Thiol modified thrombin aptamer was covalently assembled to the surface of gold nanoparticles as nanobio probes. As thrombin has two specific binding sites for its aptamer, it can bridge the well dispersed nanoparticles and lead to a network of particle aggregations. The formation of aggregation ia measured by RLS, and the specific detection of thrombin at nM level is achieved. The method has good specificity. (authors)

  19. Systems biology of coagulation initiation: kinetics of thrombin generation in resting and activated human blood.

    Directory of Open Access Journals (Sweden)

    Manash S Chatterjee

    Full Text Available Blood function defines bleeding and clotting risks and dictates approaches for clinical intervention. Independent of adding exogenous tissue factor (TF, human blood treated in vitro with corn trypsin inhibitor (CTI, to block Factor XIIa will generate thrombin after an initiation time (T(i of 1 to 2 hours (depending on donor, while activation of platelets with the GPVI-activator convulxin reduces T(i to ∼20 minutes. Since current kinetic models fail to generate thrombin in the absence of added TF, we implemented a Platelet-Plasma ODE model accounting for: the Hockin-Mann protease reaction network, thrombin-dependent display of platelet phosphatidylserine, VIIa function on activated platelets, XIIa and XIa generation and function, competitive thrombin substrates (fluorogenic detector and fibrinogen, and thrombin consumption during fibrin polymerization. The kinetic model consisting of 76 ordinary differential equations (76 species, 57 reactions, 105 kinetic parameters predicted the clotting of resting and convulxin-activated human blood as well as predicted T(i of human blood under 50 different initial conditions that titrated increasing levels of TF, Xa, Va, XIa, IXa, and VIIa. Experiments with combined anti-XI and anti-XII antibodies prevented thrombin production, demonstrating that a leak of XIIa past saturating amounts of CTI (and not "blood-borne TF" alone was responsible for in vitro initiation without added TF. Clotting was not blocked by antibodies used individually against TF, VII/VIIa, P-selectin, GPIb, protein disulfide isomerase, cathepsin G, nor blocked by the ribosome inhibitor puromycin, the Clk1 kinase inhibitor Tg003, or inhibited VIIa (VIIai. This is the first model to predict the observed behavior of CTI-treated human blood, either resting or stimulated with platelet activators. CTI-treated human blood will clot in vitro due to the combined activity of XIIa and XIa, a process enhanced by platelet activators and which proceeds

  20. Thrombin stimulation of inflammatory breast cancer cells leads to aggressiveness via the EGFR-PAR1-Pak1 pathway.

    Science.gov (United States)

    Ohshiro, Kazufumi; Bui-Nguyen, Tri M; Divijendra Natha, Reddy S; Schwartz, Arnold M; Levine, Paul; Kumar, Rakesh

    2012-12-27

    Inflammatory breast cancer (IBC) accounts for a small fraction but aggressive form of epithelial breast cancer. Although the role of thrombin in cancer is beginning to be unfolded, its impact on the biology of IBC remains unknown. The purpose of this study was to establish the role of thrombin on the invasiveness of IBC cells. The IBC SUM149 cell line was treated with thrombin in the absence or presence of the epidermal growth factor receptor (EGFR) inhibitor erlotinib and protease-activated receptor 1 (PAR1) inhibitor. The effects of pharmacological inhibitors on the ability of thrombin to stimulate the growth rate and invasiveness were examined. We found that the inhibition of putative cellular targets of thrombin action suppresses both the growth and invasiveness of SUM149 cells in a concentration-dependent manner. In addition, thrombin-mediated increased invasion of SUM149 cells was routed through EGFR phosphorylation, and in turn, stimulation of the p21-activated kinase (Pak1) activity in a EGFR-sensitive manner. Interestingly, thrombin-mediated activation of the Pak1 pathway stimulation was blocked by erlotinib and PAR1 inhibitor. For proof-of-principle studies, we found immunohistochemical evidence of Pak1 activation as well as expression of PAR1 in IBC. Thrombin utilizes EGFR to relay signals promoting SUM149 cell growth and invasion via the Pak1 pathway. The study provides the rationale for future therapeutic approaches in mitigating the invasive nature of IBC by targeting Pak1 and/or EGFR.

  1. Computer based screening of compound databases: 1. Preselection of benzamidine-based thrombin inhibitors.

    Science.gov (United States)

    Fox, T; Haaksma, E E

    2000-07-01

    We present a computational protocol which uses the known three-dimensional structure of a target enzyme to identify possible ligands from databases of compounds with low molecular weight. This is accomplished by first mapping the essential interactions in the binding site with the program GRID. The resulting regions of favorable interaction between target and ligand are translated into a database query, and with UNITY a flexible 3D database search is performed. The feasibility of this approach is calibrated with thrombin as the target. Our results show that the resulting hit lists are enriched with thrombin inhibitors compared to the total database.

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

    DEFF Research Database (Denmark)

    Valnickova, Zuzana; Thaysen-Andersen, Morten; Højrup, Peter;

    2009-01-01

    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 fib......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...

  3. Topic and Textual Coherence

    Institute of Scientific and Technical Information of China (English)

    何明烈; 戢焕奇

    2008-01-01

    Based on the investigation of the origin and definition of topic as well as the classification of textual coherence, this paper ana-lyzes the function of topic to non-coherence, imphcit coherence and explicit coherence, and concludes that topic relevance is the primary cri-terion for textual coherence.

  4. In vitro study of the role of thrombin in platelet rich plasma (PRP) preparation: utility for gel formation and impact in growth factors release

    OpenAIRE

    Huber, Stephany Cares; Cunha Júnior, José Luiz Rosenberis; Montalvão, Silmara; da Silva, Letícia Queiroz; Paffaro, Aline Urban; da Silva, Francesca Aparecida Ramos; Rodrigues, Bruno Lima; Lana, José Fabio Santos Duarte; Annichino-Bizzacchi, Joyce Maria

    2016-01-01

    Introduction: The use of PRP has been studied for different fields, with promising results in regenerative medicine. Until now, there is no study in the literature evaluating thrombin levels in serum, used as autologous thrombin preparation. Therefore, in the present study we evaluated the role played by different thrombin concentrations in PRP and the impact in the release of growth factors. Also, different activators for PRP gel formation were evaluated. Methods: Thrombin levels were measur...

  5. Longitudinal assessment of thrombin generation potential in response to alteration of antiplatelet therapy after TIA or ischaemic stroke.

    LENUS (Irish Health Repository)

    Tobin, W O

    2013-02-01

    The impact of changing antiplatelet therapy on thrombin generation potential in patients with ischaemic cerebrovascular disease (CVD) is unclear. We assessed patients within 4 weeks of TIA or ischaemic stroke (baseline), and then 14 days (14d) and >90 days (90d) after altering antiplatelet therapy. Thrombin generation was assessed in platelet poor plasma. Ninety-one patients were recruited. Twenty-four were initially assessed on no antiplatelet therapy, and then after 14d (N = 23) and 90d (N = 8) on aspirin monotherapy; 52 were assessed on aspirin monotherapy, and after 14 and 90 days on aspirin and dipyridamole combination therapy; 21 patients were assessed on aspirin and after 14 days (N = 21) and 90 days (N = 19) on clopidogrel. Peak thrombin generation and endogenous thrombin potential were reduced at 14 and 90 days (p ≤ 0.04) in the overall cohort. We assessed the impact of individual antiplatelet regimens on thrombin generation parameters to investigate the cause of this effect. Lag time and time-to-peak thrombin generation were unchanged at 14 days, but reduced 90 days after commencing aspirin (p ≤ 0.009). Lag time, peak thrombin generation and endogenous thrombin potential were reduced at both 14 and 90 days after adding dipyridamole to aspirin (p ≤ 0.01). Lag time was reduced 14 days after changing from aspirin to clopidogrel (p = 0.045), but this effect was not maintained at 90 days (p = 0.2). This pilot study did not show any consistent effects of commencing aspirin, or of changing from aspirin to clopidogrel on thrombin generation potential during follow-up. The addition of dipyridamole to aspirin led to a persistent reduction in peak and total thrombin generation ex vivo, and illustrates the diverse, potentially beneficial, newly recognised \\'anti-coagulant\\' effects of dipyridamole in ischaemic CVD.

  6. Large-scale molecular dynamics simulation: Effect of polarization on thrombin-ligand binding energy.

    Science.gov (United States)

    Duan, Li L; Feng, Guo Q; Zhang, Qing G

    2016-01-01

    Molecular dynamics (MD) simulations lasting 500 ns were performed in explicit water to investigate the effect of polarization on the binding of ligands to human α-thrombin based on the standard nonpolarizable AMBER force field and the quantum-derived polarized protein-specific charge (PPC). The PPC includes the electronic polarization effect of the thrombin-ligand complex, which is absent in the standard force field. A detailed analysis and comparison of the results of the MD simulation with experimental data provided strong evidence that intra-protein, protein-ligand hydrogen bonds and the root-mean-square deviation of backbone atoms were significantly stabilized through electronic polarization. Specifically, two critical hydrogen bonds between thrombin and the ligand were broken at approximately 190 ns when AMBER force field was used and the number of intra-protein backbone hydrogen bonds was higher under PPC than under AMBER. The thrombin-ligand binding energy was computed using the molecular mechanics Poisson-Boltzmann surface area (MM/PBSA) method, and the results were consistent with the experimental value obtained using PPC. Because hydrogen bonds were unstable, it was failed to predict the binding affinity under the AMBER force field. Furthermore, the results of the present study revealed that differences in the binding free energy between AMBER and PPC almost comes from the electrostatic interaction. Thus, this study provides evidence that protein polarization is critical to accurately describe protein-ligand binding. PMID:27507430

  7. Regulation of Thrombin-Induced Lung Endothelial Cell Barrier Disruption by Protein Kinase C Delta

    Science.gov (United States)

    Xie, Lishi; Chiang, Eddie T.; Kelly, Gabriel T.; Kanteti, Prasad; Singleton, Patrick A.; Camp, Sara M.; Zhou, Tingting; Dudek, Steven M.; Natarajan, Viswanathan; Wang, Ting; Black, Steven M.; Garcia, Joe G. N.; Jacobson, Jeffrey R.

    2016-01-01

    Protein Kinase C (PKC) plays a significant role in thrombin-induced loss of endothelial cell (EC) barrier integrity; however, the existence of more than 10 isozymes of PKC and tissue–specific isoform expression has limited our understanding of this important second messenger in vascular homeostasis. In this study, we show that PKCδ isoform promotes thrombin-induced loss of human pulmonary artery EC barrier integrity, findings substantiated by PKCδ inhibitory studies (rottlerin), dominant negative PKCδ construct and PKCδ silencing (siRNA). In addition, we identified PKCδ as a signaling mediator upstream of both thrombin-induced MLC phosphorylation and Rho GTPase activation affecting stress fiber formation, cell contraction and loss of EC barrier integrity. Our inhibitor-based studies indicate that thrombin-induced PKCδ activation exerts a positive feedback on Rho GTPase activation and contributes to Rac1 GTPase inhibition. Moreover, PKD (or PKCμ) and CPI-17, two known PKCδ targets, were found to be activated by PKCδ in EC and served as modulators of cytoskeleton rearrangement. These studies clarify the role of PKCδ in EC cytoskeleton regulation, and highlight PKCδ as a therapeutic target in inflammatory lung disorders, characterized by the loss of barrier integrity, such as acute lung injury and sepsis. PMID:27442243

  8. Regulation of Thrombin-Induced Lung Endothelial Cell Barrier Disruption by Protein Kinase C Delta.

    Directory of Open Access Journals (Sweden)

    Lishi Xie

    Full Text Available Protein Kinase C (PKC plays a significant role in thrombin-induced loss of endothelial cell (EC barrier integrity; however, the existence of more than 10 isozymes of PKC and tissue-specific isoform expression has limited our understanding of this important second messenger in vascular homeostasis. In this study, we show that PKCδ isoform promotes thrombin-induced loss of human pulmonary artery EC barrier integrity, findings substantiated by PKCδ inhibitory studies (rottlerin, dominant negative PKCδ construct and PKCδ silencing (siRNA. In addition, we identified PKCδ as a signaling mediator upstream of both thrombin-induced MLC phosphorylation and Rho GTPase activation affecting stress fiber formation, cell contraction and loss of EC barrier integrity. Our inhibitor-based studies indicate that thrombin-induced PKCδ activation exerts a positive feedback on Rho GTPase activation and contributes to Rac1 GTPase inhibition. Moreover, PKD (or PKCμ and CPI-17, two known PKCδ targets, were found to be activated by PKCδ in EC and served as modulators of cytoskeleton rearrangement. These studies clarify the role of PKCδ in EC cytoskeleton regulation, and highlight PKCδ as a therapeutic target in inflammatory lung disorders, characterized by the loss of barrier integrity, such as acute lung injury and sepsis.

  9. The role of thrombin-activatable fibrinolysis inhibitor in diabetic wound healing

    NARCIS (Netherlands)

    C.J.N. Verkleij; J.J.T.H. Roelofs; S.R. Havik; J.C.M. Meijers; P.F. Marx

    2010-01-01

    Introduction: One of the major complications in patients with diabetes mellitus is impaired wound healing. The fibrinolytic system is involved in parts of the wound healing process and deficiency of thrombin-activatable fibrinolysis inhibitor (TAFI) results in delayed wound closure. Moreover, levels

  10. Quantitative phosphoproteomics unveils temporal dynamics of thrombin signaling in human endothelial cells

    NARCIS (Netherlands)

    van den Biggelaar, Maartje; Hernández-Fernaud, Juan Ramon; van den Eshof, Bart L; Neilson, Lisa J; Meijer, Alexander B; Mertens, Koenraad; Zanivan, Sara

    2014-01-01

    Thrombin is the key serine protease of the coagulation cascade and a potent trigger of protease-activated receptor 1 (PAR1)-mediated platelet aggregation. In recent years, PAR1 has become an appealing target for anticoagulant therapies. However, the inhibitors that have been developed so far increas

  11. Targeting the thrombin receptor modulates inflammation and astrogliosis to improve recovery after spinal cord injury.

    Science.gov (United States)

    Radulovic, Maja; Yoon, Hyesook; Wu, Jianmin; Mustafa, Karim; Scarisbrick, Isobel A

    2016-09-01

    The deregulation of serine protease activity is a common feature of neurological injury, but little is known regarding their mechanisms of action or whether they can be targeted to facilitate repair. In this study we demonstrate that the thrombin receptor (Protease Activated Receptor 1, (PAR1)) serves as a critical translator of the spinal cord injury (SCI) proteolytic microenvironment into a cascade of pro-inflammatory events that contribute to astrogliosis and functional decline. PAR1 knockout mice displayed improved locomotor recovery after SCI and reduced signatures of inflammation and astrogliosis, including expression of glial fibrillary acidic protein (GFAP), vimentin, and STAT3 signaling. SCI-associated elevations in pro-inflammatory cytokines such as IL-1β and IL-6 were also reduced in PAR1-/- mice and co-ordinate improvements in tissue sparing and preservation of NeuN-positive ventral horn neurons, and PKCγ corticospinal axons, were observed. PAR1 and its agonist's thrombin and neurosin were expressed by perilesional astrocytes and each agonist increased the production of IL-6 and STAT3 signaling in primary astrocyte cultures in a PAR1-dependent manner. In turn, IL-6-stimulated astrocytes increased expression of PAR1, thrombin, and neurosin, pointing to a model in which PAR1 activation contributes to increased astrogliosis by feedforward- and feedback-signaling dynamics. Collectively, these findings identify the thrombin receptor as a key mediator of inflammation and astrogliosis in the aftermath of SCI that can be targeted to reduce neurodegeneration and improve neurobehavioral recovery. PMID:27145117

  12. Pleiotropic effects of factor Xa and thrombin : what to expect from novel anticoagulants

    NARCIS (Netherlands)

    Spronk, Henri M. H.; de Jong, Anne Margreet; Crijns, Harry J.; Schotten, Ulrich; Van Gelder, Isabelle C.; ten Cate, Hugo

    2014-01-01

    Factor Xa and thrombin are well-known components of the coagulation cascade and have been proven to be viable targets for effective anticoagulation treatment. However, accumulating evidence suggests that these serine proteases are also crucial modulators of other cellular mechanisms through the acti

  13. Generation and characterization of a highly stable form of activated thrombin-activable fibrinolysis inhibitor

    NARCIS (Netherlands)

    Marx, PF; Havik, [No Value; Marquart, JA; Meijers, JCM; Bouma, Bonno N.

    2004-01-01

    Activated thrombin-activable fibrinolysis inhibitor (TAFIa) is a carboxypeptidase B that can down-regulate fibrinolysis. TAFIa is a labile enzyme that can be inactivated by conformational instability or proteolysis. TAFI is similar to 40% identical to pancreatic carboxypeptidase B (CPB). In contrast

  14. Effects of fused hirudin on activity of thrombin and function of platelets

    Institute of Scientific and Technical Information of China (English)

    SHEN Li; CHEN Shao-ping; CAI Zai-long; YANG Sheng-sheng; QIN Yong-wen

    2005-01-01

    Objective: To investigate whether fused hirudin peptide has both antithrombin and antiplatelet functions. Methods: The core region of fused hirudin was the C-terminal tail of hirudin(hirudin53-64),which could bind to the anion binding exosite (ABE) of thrombin.Arg-Pro-Pro-Gly-Phe(RPPGF) amino acid sequence,a metabolite of bradykinin,was added to the N-terminus of hirudin53-64.It bound to the active site of thrombin.Additionally,Arg-Gly-Asp(RGD)amino acid sequence,an inibitor of glycoprotein Ⅱb/Ⅲa( GP Ⅱb/Ⅲa) receptor,was linked to C-terminus of hirudin53-64.This 26-animo acid-fused hirudin peptide was artificially synthesized,purified and analysed. Results: Fused hirudin peptide significantly lengthened the activated partial thromboplastin time(APTT),thrombin time(TT)and prothrombin time(PT) and inhibited the amidolytic activity of thrombin.The ADP-induced platelet aggregation was markedly inhibited by fused hirudin peptide. Conclusion: Fused hirudin peptide has activity of antithrombin as well as antiplatelet.Therefore bifunctional anticoagulation peptide has capacity to target various components of haemostatic process and may become more powerful antithrombosis agent.

  15. Thrombin action decreases acetylcholine receptor aggregate number and stability in cultured mouse myotubes.

    Science.gov (United States)

    Davenport, R W; Lanuza, M; Kim, S; Jia, M; Snyder, E; Nelson, P G

    2000-08-30

    Neurons develop and make very stable, long-term synaptic connections with other nerve cells and with muscle. Synaptic stability at the neuromuscular junction changes over development in that a proliferation of synaptic input are made to individual myotubes and synapses from all but one neuron are lost during development. In an established co-culture paradigm in which spinal motoneurons synaptically contact myotubes, thrombin and associated protease inhibitors have been shown to affect the loss of functional synaptic contacts [6]. Evidence has not been provided which clearly demonstrate whether protease/protease inhibitors affect either the pre- or postsynaptic terminal, or both. In an effort to determine whether these reagents directly affect postsynaptic receptors on myotubes, myotubes were cultured in the absence of neurons and the spontaneous presence and stability of aggregates of acetylcholine receptors (AChR) in control and thrombin-containing media were evaluated. In dishes fixed after treatment and in dishes in which individual aggregates were observed live, thrombin action appeared to increase loss of AChR aggregates over time. Hirudin, a specific inhibitor of the thrombin protease, diminished this loss. Neither reagent affected the overall incorporation or degradation of AChR; therefore, it appears these protease/protease inhibitors affect the state of AChR aggregation. PMID:10960680

  16. Hypercoagulability following major partial liver resection - detected by thrombomodulin-modified thrombin generation testing

    NARCIS (Netherlands)

    Potze, W.; Alkozai, E. M.; Adelmeijer, J.; Porte, R. J.; Lisman, T.

    2015-01-01

    BackgroundConventional coagulation tests are frequently prolonged after liver surgery, suggesting a post-operative hypocoagulability. However, these tests are unreliable for assessment of the haemostatic status in these patients. In contrast, thrombin generation testing measures the true balance bet

  17. Large-scale molecular dynamics simulation: Effect of polarization on thrombin-ligand binding energy

    Science.gov (United States)

    Duan, Li L.; Feng, Guo Q.; Zhang, Qing G.

    2016-01-01

    Molecular dynamics (MD) simulations lasting 500 ns were performed in explicit water to investigate the effect of polarization on the binding of ligands to human α-thrombin based on the standard nonpolarizable AMBER force field and the quantum-derived polarized protein-specific charge (PPC). The PPC includes the electronic polarization effect of the thrombin-ligand complex, which is absent in the standard force field. A detailed analysis and comparison of the results of the MD simulation with experimental data provided strong evidence that intra-protein, protein-ligand hydrogen bonds and the root-mean-square deviation of backbone atoms were significantly stabilized through electronic polarization. Specifically, two critical hydrogen bonds between thrombin and the ligand were broken at approximately 190 ns when AMBER force field was used and the number of intra-protein backbone hydrogen bonds was higher under PPC than under AMBER. The thrombin-ligand binding energy was computed using the molecular mechanics Poisson-Boltzmann surface area (MM/PBSA) method, and the results were consistent with the experimental value obtained using PPC. Because hydrogen bonds were unstable, it was failed to predict the binding affinity under the AMBER force field. Furthermore, the results of the present study revealed that differences in the binding free energy between AMBER and PPC almost comes from the electrostatic interaction. Thus, this study provides evidence that protein polarization is critical to accurately describe protein-ligand binding. PMID:27507430

  18. Thrombin inhibition by dabigatran attenuates atherosclerosis in ApoE deficient mice

    OpenAIRE

    Pingel, Simon; Tiyerili, Vedat; Mueller, Jens; Werner, Nikos; Nickenig, Georg; Mueller, Cornelius

    2014-01-01

    Introduction Atherosclerosis is a chronic inflammatory disease characterized by endothelial cell damage, infiltration, proliferation and accumulation of macrophages, lymphocytes and transformed vascular smooth muscle cells within the vascular wall and procoagulation processes involving activation of plasmatic coagulation events and platelets. Numerous studies suggested a close interaction between thrombin action and atherogenesis, but possibly underlying mechanisms are multiple and specific t...

  19. (-)-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)

    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

    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.

  20. Recombinant Technology and Probiotics

    Directory of Open Access Journals (Sweden)

    Icy D’Silva

    2011-09-01

    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.

  1. 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

  2. Thrombography reveals thrombin generation potential continues to deteriorate following cardiopulmonary bypass surgery despite adequate hemostasis.

    Science.gov (United States)

    Wong, Raymond K; Sleep, Joseph R; Visner, Allison J; Raasch, David J; Lanza, Louis A; DeValeria, Patrick A; Torloni, Antonio S; Arabia, Francisco A

    2011-03-01

    The intrinsic and extrinsic activation pathways of the hemostatic system converge when prothrombin is converted to thrombin. The ability to generate an adequate thrombin burst is the most central aspect of the coagulation cascade. The thrombin-generating potential in patients following cardiopulmonary bypass (CPB) may be indicative of their hemostatic status. In this report, thrombography, a unique technique for directly measuring the potential of patients' blood samples to generate adequate thrombin bursts, is used to characterize the coagulopathic profile in post-CPB patients. Post-CPB hemostasis is typically achieved with protamine reversal of heparin anticoagulation and occasionally supplemented with blood product component transfusions. In this pilot study, platelet poor plasma samples were derived from 11 primary cardiac surgery patients at five time points: prior to CPB, immediately post-protamine, upon arrival to the intensive care unit (ICU), 3 hours post-ICU admission, and 24 hours after ICU arrival. Thrombography revealed that the Endogenous Thrombin Potential (ETP) was not different between [Baseline] and [PostProtamine] but proceeded to deteriorate in the immediate postoperative period. At the [3HourPostICU] time point, the ETP was significantly lower than the [Baseline] values, 1233 +/- 591 versus 595 +/- 379 nM.min (mean +/- SD; n=9, p coagulation testing results, and blood loss volumes are also presented. Despite adequate hemostasis, thrombography reveals an underlying coagulopathic process that could put some cardiac surgical patients at risk for postoperative bleeding. Thrombography is a novel technique that could be developed into a useful tool for perfusionists and physicians to identify coagulopathies and optimize blood management following CPB. PMID:21449230

  3. Modulation of 3D Fibrin Matrix Stiffness by Intrinsic Fibrinogen–Thrombin Compositions and by Extrinsic Cellular Activity

    Science.gov (United States)

    Duong, Haison; Wu, Benjamin

    2009-01-01

    Fibrin is a substance formed through catalytic conversion of coagulation constituents: fibrinogen and thrombin. The kinetics of the two constituents determines the structural properties of the fibrin architecture. We have shown previously that changing the fibrinogen and thrombin concentrations in the final three-dimensional (3D) fibrin matrix influenced cell proliferation and differentiation. In this study, we further examined the effect of changing fibrinogen and thrombin concentrations in the absence or presence of fibroblasts on the structural modulus or stiffness of 3D fibrin matrices. We have prepared fibroblast-free and fibroblast-embedded 3D fibrin matrices of different fibrinogen and thrombin formulations, and tested the stiffness of these constructs using standard mechanical testing assays. Results showed that there was a corresponding increase in stiffness with increasing thrombin and fibrinogen concentrations; the increase was more notable with fibrinogen and to a lesser degree with thrombin. The effect of fibroblasts on the stiffness of the fibrin construct was also examined. We have observed a small increase in the stiffness of the fibroblast-incorporated fibrin construct as they proliferated and exhibited spreading morphology. To our knowledge, this is the first comprehensive report detailing the relationship between fibrinogen and thrombin concentrations, cell proliferation, and stiffness in 3D fibrin matrices. The data obtained may lead to optimally design suitable bioscaffolds where we can control both cell proliferation and structural integrity for a variety of tissue engineering applications. PMID:19309239

  4. [Topical therapy of rosacea].

    Science.gov (United States)

    Schöfer, H

    2013-07-01

    Metronidazole and azelaic acid are the only topical medications approved for rosacea. All other topical treatments for rosacea and its special forms are used off-label. Topical steroids are not indicated in rosacea, because of their side effects (induction of steroid rosacea, high risk of facial skin atrophy, and high risk of rebound after cessation of therapy). Topical as well as systemic steroids are allowed only as initial and short term therapy for acute forms of rosacea (e.g. rosacea fulminans). Papular and pustular rosacea is the major indication for topical therapy. Sebaceous gland and connective tissue hyperplasia in glandular-hypertrophic rosacea as well as erythema in erythematous rosacea do not respond well to topical measures. A new active substance, the alpha-2-adrenoreceptor agonist brimonidine, will be approved soon for the topical treatment of erythema in rosacea. All severe forms of rosacea should initially be treated with a combination of topical and systemic agents. After improvement of the clinical symptoms, topical treatment alone is usually adequate to maintain the control.

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

    OpenAIRE

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

    2014-01-01

    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 leve...

  6. Syntacticized topics in Kurmuk

    DEFF Research Database (Denmark)

    Andersen, Torben

    2015-01-01

    . The grammatical relation of the topic is expressed by a voice-like inflection of the verb, here called orientation. While subject-orientation is morphologically unmarked, object-oriented and adjunct-oriented verbs are marked by a subject suffix or by a suffix indicating that the topic is not subject, and adjunct...

  7. Salicylic Acid Topical

    Science.gov (United States)

    ... skin blemishes in people who have acne. Topical salicylic acid is also used to treat skin conditions that involve scaling or overgrowth of skin ... water for 15 minutes.Do not apply topical salicylic acid to skin that is broken, red, swollen, irritated, or infected. ...

  8. Topic Identification in Discourse

    CERN Document Server

    Chen, K

    1995-01-01

    This paper proposes a corpus-based language model for topic identification. We analyze the association of noun-noun and noun-verb pairs in LOB Corpus. The word association norms are based on three factors: 1) word importance, 2) pair co-occurrence, and 3) distance. They are trained on the paragraph and sentence levels for noun-noun and noun-verb pairs, respectively. Under the topic coherence postulation, the nouns that have the strongest connectivities with the other nouns and verbs in the discourse form the preferred topic set. The collocational semantics then is used to identify the topics from paragraphs and to discuss the topic shift phenomenon among paragraphs.

  9. Factor Xa generation by computational modeling: an additional discriminator to thrombin generation evaluation.

    Directory of Open Access Journals (Sweden)

    Kathleen E Brummel-Ziedins

    Full Text Available Factor (fXa is a critical enzyme in blood coagulation that is responsible for the initiation and propagation of thrombin generation. Previously we have shown that analysis of computationally generated thrombin profiles is a tool to investigate hemostasis in various populations. In this study, we evaluate the potential of computationally derived time courses of fXa generation as another approach for investigating thrombotic risk. Utilizing the case (n = 473 and control (n = 426 population from the Leiden Thrombophilia Study and each individual's plasma protein factor composition for fII, fV, fVII, fVIII, fIX, fX, antithrombin and tissue factor pathway inhibitor, tissue factor-initiated total active fXa generation was assessed using a mathematical model. FXa generation was evaluated by the area under the curve (AUC, the maximum rate (MaxR and level (MaxL and the time to reach these, TMaxR and TMaxL, respectively. FXa generation was analyzed in the entire populations and in defined subgroups (by sex, age, body mass index, oral contraceptive use. The maximum rates and levels of fXa generation occur over a 10- to 12- fold range in both cases and controls. This variation is larger than that observed with thrombin (3-6 fold in the same population. The greatest risk association was obtained using either MaxR or MaxL of fXa generation; with an ∼2.2 fold increased risk for individuals exceeding the 90(th percentile. This risk was similar to that of thrombin generation(MaxR OR 2.6. Grouping defined by oral contraceptive (OC use in the control population showed the biggest differences in fXa generation; a >60% increase in the MaxR upon OC use. FXa generation can distinguish between a subset of individuals characterized by overlapping thrombin generation profiles. Analysis of fXa generation is a phenotypic characteristic which may prove to be a more sensitive discriminator than thrombin generation among all individuals.

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

    Directory of Open Access Journals (Sweden)

    R. Anand

    2014-10-01

    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

  11. Recombinant DNA in Medicine

    OpenAIRE

    Cederbaum, Stephen D.; Fareed, George C.; Lovett, Michael A.; Shapiro, Larry J.

    1984-01-01

    Studies in bacteria and bacterial viruses have led to methods to manipulate and recombine DNA in unique and reproducible ways and to amplify these recombined molecules millions of times. Once properly identified, the recombinant DNA molecules can be used in various ways useful in medicine and human biology. There are many applications for recombinant DNA technology. Cloned complementary DNA has been used to produce various human proteins in microorganisms. Insulin and growth hormone have been...

  12. Improving baculovirus recombination

    OpenAIRE

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

    2003-01-01

    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...

  13. Mutations in the fourth EGF-like domain affect thrombomodulin-induced changes in the active site of thrombin

    OpenAIRE

    Koeppe, Julia R.; Beach, Muneera A.; Baerga-Ortiz, Abel; Jordan Kerns, S.; Komives, Elizabeth A.

    2008-01-01

    A number of alanine and more conservative mutants of residues in the fourth domain of thrombomodulin (TM) were prepared and assayed for protein C activation and for thrombin binding. Several of the alanine mutations appeared to cause misfolding or structural defects as assessed by poor expression and/or NMR HSQC experiments, while more conservative mutations at the same site appeared to fold correctly and retain activity. Several of the conservative mutants bound more weakly to thrombin despi...

  14. Thrombin generation by exposure of blood to endotoxin: a simple model to study disseminated intravascular coagulation.

    Science.gov (United States)

    Stief, T W

    2006-04-01

    Pathologic disseminated intravascular coagulation (PDIC) is a serious complication in sepsis. In an in-vitro system consisting of incubation of fresh citrated blood with lipopolysaccharides (LPS) or glucans and subsequent plasma recalcification plasmatic thrombin was quantified. Five hundred microliters of freshly drawn citrated blood of healthy donors were incubated with up to 800 ng/mL LPS (Escherichia coli) or up to 80 microg/mL Zymosan A (ZyA; Candida albicans) for 30 minutes at room temperature (RT). The samples were centrifuged, and 30 microL plasma were recalcified with 1 volume or less of CaCl(2) (25 micromoles Ca(2+)/mL plasma). After 0 to 12 minutes (37 degrees C), 20 microL 2.5 M arginine, pH 8.6, were added. Thirty microliters 0.9 mM HD-CHG-Ala-Arg-pNA in 2.3 M arginine were added, and the absorbance increase at 405 nm was determined. Fifty microliters plasma were also incubated with 5 microL 250 mM CaCl2 for 5, 10, or 15 minutes (37 degrees C). Fifty microliters 2.5 M arginine stops coagulation, and 50 microL 0.77 mM HD-CHG-Ala-Arg-pNA in 2.3 M arginine starts the thrombin detection. The standard was 1 IU/mL thrombin in 7% human albumin instead of plasma. Arginine was also added in the endotoxin exposure time (EET) or in the plasma coagulation reaction time (CRT). Tissue factor (TF)-antigen and soluble CD14 were determined. LPS at blood concentrations greater than 10 ng/mL or ZyA at greater than 1 microg/mL severalfold enhance thrombin generation, when the respective plasmas are recalcified. After 30 minutes EET at RT, the thrombin activity at 12 minutes CRT generated by the addition of 200 ng/mL LPS or 20 microg/mL ZyA is approximately 200 mIU/mL compared to approximately 20 mIU/mL without addition of endotoxin, or compared to about 7 mIU/mL thrombin at 0 minutes CRT. Arginine added to blood or to plasma inhibits thrombin generation; the inhibitory concentration 50% (IC 50) is approximately 15 mM plasma concentration. Endotoxin incubation of blood

  15. Endoscopic ultrasound guided thrombin injection of angiographically occult pancreatitis associated visceral artery pseudoaneurysms:Case series

    Institute of Scientific and Technical Information of China (English)

    Shivanand; Gamanagatti; Usha; Thingujam; Pramod; Garg; Surajkumar; Nongthombam; Nihar; Ranjan; Dash

    2015-01-01

    Pseudoaneurysm is a known complication of pancreatitis associated with significant mortality and morbidity. Imaging plays an important role in the diagnosis and management. Computed tomography(CT) helps localize the lesion and the severity of the background pancreatitis but digital subtraction angiography with coil embolization is recommended to avoid bleeding and inadvertent surgery. However, in cases where angiographic coil embolization is not feasible due to technical reasons, thrombin injection via CT or ultrasound guidance remains a viable option and often described in literature. In this series, effort has been made to highlight the role of endoscopic ultrasound guided thrombin instillation especially in patients with poorly visualized pseudoaneurysm on ultrasound thereby avoiding surgery and the associated mortality and morbidity.

  16. Implications of Dabigatran, a direct thrombin inhibitor, for oral surgery practice.

    Science.gov (United States)

    Davis, Clayton; Robertson, Chad; Shivakumar, Sudeep; Lee, Min

    2013-01-01

    Direct thrombin inhibitors, specifically orally administered dabigatran etexilate, are emerging as alternatives to warfarin for anticoagulation in the management of atrial fibrillation and venous thromboembolism. The risk associated with bleeding events while taking dabigatran has been documented in multiple randomized controlled trials, but to date, no studies have focused on the risk of bleeding after dental extraction. Extraction of teeth is one of the most common surgical procedures and may cause significant bleeding, so a thorough understanding of the pharmacology of anticoagulant medications is required to prevent complications. With the increasing use of direct thrombin inhibitors, the safe management of patients taking these anticoagulants must be delineated. This review compares dabigatran and warfarin, especially in terms of their effects on dental and oral surgery practice, and examines best management of these patients in light of the existing literature. PMID:23920075

  17. 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.)

  18. Diclofenac Topical (osteoarthritis pain)

    Science.gov (United States)

    ... should know that you should not apply sunscreens, cosmetics, lotions, moisturizers, insect repellents, or other topical medications ... that is bloody or looks like coffee grounds slow, shallow, or irregular breathing hives itching difficulty breathing ...

  19. Advanced Topics in Aerodynamics

    DEFF Research Database (Denmark)

    Filippone, Antonino

    1999-01-01

    "Advanced Topics in Aerodynamics" is a comprehensive electronic guide to aerodynamics,computational fluid dynamics, aeronautics, aerospace propulsion systems, design and relatedtechnology. We report data, tables, graphics, sketches,examples, results, photos, technical andscientific literature, for...

  20. Sensitive electrochemical aptasensor for thrombin detection based on graphene served as platform and graphene oxide as enhancer.

    Science.gov (United States)

    He, Chun; Xu, Zenghong; Sun, Tao; Wang, Li

    2014-01-01

    A sensitive electrochemical aptasensor was developed with conductive graphene served as platform and inert graphene oxide (GO) as enhancer. An electrodeposited nano-Au layer was firstly formed on conductive graphene modified glass carbon electrode surface for further immobilizing of electrochemical redox probe hexacyanoferrates nanoparticles (NiHCFNPs). Subsequently, another nano-Au layer was formed for immobilizing of thrombin aptamer (TBA). In the presence of thrombin, the TBA on the electrode surface could bind with thrombin, which made a barrier for electrons and inhibited the electro-transfer, resulting in the decreased electrochemical signals of NiHCFNPs. Owing to the non-conductivity property of graphene oxide, further decreased electrochemical signals of NiHCFNPs could be obtained via the sandwich reaction with GO-labeled TBA. According to the signal changes before the thrombin recognition and after sandwich reaction, trace detection of thrombin could be achieved. As a result, the proposed approach showed a high sensitivity and a wider linearity to thrombin in the range from 0.005 nM to 50 nM with a detection limit of 1 pM. PMID:24142359

  1. Large-scale preparation of active caspase-3 in E. coli by designing its thrombin-activatable precursors

    Directory of Open Access Journals (Sweden)

    Park Sung

    2008-12-01

    Full Text Available Abstract Background Caspase-3, a principal apoptotic effector that cleaves the majority of cellular substrates, is an important medicinal target for the treatment of cancers and neurodegenerative diseases. Large amounts of the protein are required for drug discovery research. However, previous efforts to express the full-length caspase-3 gene in E. coli have been unsuccessful. Results Overproducers of thrombin-activatable full-length caspase-3 precursors were prepared by engineering the auto-activation sites of caspase-3 precursor into a sequence susceptible to thrombin hydrolysis. The engineered precursors were highly expressed as soluble proteins in E. coli and easily purified by affinity chromatography, to levels of 10–15 mg from 1 L of E. coli culture, and readily activated by thrombin digestion. Kinetic evaluation disclosed that thrombin digestion enhanced catalytic activity (kcat/KM of the precursor proteins by two orders of magnitude. Conclusion A novel method for a large-scale preparation of active caspase-3 was developed by a strategic engineering to lack auto-activation during expression with amino acid sequences susceptible to thrombin, facilitating high-level expression in E. coli. The precursor protein was easily purified and activated through specific cleavage at the engineered sites by thrombin, generating active caspase-3 in high yields.

  2. Deletion of the thrombin cleavage domain of osteopontin mediates breast cancer cell adhesion, proteolytic activity, tumorgenicity, and metastasis

    Directory of Open Access Journals (Sweden)

    Postenka Carl O

    2011-01-01

    Full Text Available Abstract Background Osteopontin (OPN is a secreted phosphoprotein often overexpressed at high levels in the blood and primary tumors of breast cancer patients. OPN contains two integrin-binding sites and a thrombin cleavage domain located in close proximity to each other. Methods To study the role of the thrombin cleavage site of OPN, MDA-MB-468 human breast cancer cells were stably transfected with either wildtype OPN (468-OPN, mutant OPN lacking the thrombin cleavage domain (468-ΔTC or an empty vector (468-CON and assessed for in vitro and in vivo functional differences in malignant/metastatic behavior. Results All three cell lines were found to equivalently express thrombin, tissue factor, CD44, αvβ5 integrin and β1 integrin. Relative to 468-OPN and 468-CON cells, 468-ΔTC cells expressing OPN with a deleted thrombin cleavage domain demonstrated decreased cell adhesion (p in vitro. Furthermore, injection of 468-ΔTC cells into the mammary fat pad of nude mice resulted in decreased primary tumor latency time (p Conclusions The results presented here suggest that expression of thrombin-uncleavable OPN imparts an early tumor formation advantage as well as a metastatic advantage for breast cancer cells, possibly due to increased proteolytic activity and decreased adhesion and apoptosis. Clarification of the mechanisms responsible for these observations and the translation of this knowledge into the clinic could ultimately provide new therapeutic opportunities for combating breast cancer.

  3. Spontaneous pseudoaneurysm of the uterine artery during pregnancy treated by direct thrombin injection: A case report

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Jung Hee; Kim, See Hyung; Kim, Young Hwan [Dept. Radiology, Keimyung University School of Medicine, Dongsan Medical Center, Daegu (Korea, Republic of)

    2016-04-15

    Pseudoaneurysm of uterine artery during pregnancy is a very rare disease. It is mostly associated with uterine artery injury, usually occurring after proceeding conditions such as history of gynecologic operation and infection. However, the best treatment modality has not been established yet. Herein, we reported a case of spontaneous formation of uterine artery pseudoaneurysm during pregnancy treated by direct thrombin injection without any complication or recurrence.

  4. A novel method of screening thrombin-inhibiting DNA aptamers using an evolution-mimicking algorithm

    OpenAIRE

    Ikebukuro, Kazunori; Okumura, Yuji; SUMIKURA, Koichi; Karube, Isao

    2005-01-01

    Thrombin-inhibiting DNA aptamers have already been obtained through the systematic evolution of ligands by exponential enrichment (SELEX). However, SELEX is a method that screens DNA aptamers that bind to their target molecules, and it sometimes fails to screen good inhibitors. Therefore, it is necessary to develop a method of screening DNA aptamers based on their inhibitory effects on the target molecules. We developed a novel method of detecting aptamers using an evolution-mimicking algorit...

  5. A New Potent Inhibitor of Thrombin from the Leech Haemendipsa Yanyuanensis

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Two components of anticoagulant protein were isolated from the leech Haemendipsa yanyuanensis by heparin agarose affinity chromatography and ultracentrifugation. The determination of anticoagulant activity and characterization analysis of the pro tein using the method of chromogenic substrate indicates that the anticoagulant protein is thrombin-specific but not factor Xa-specific. The results lay a foundation for the research of the anticoagulant mechanism and application of anticoagulant protein from H. yanyua nensis.

  6. Endoscopic ultrasound guided thrombin injection of angiographically occult pancreatitis associated visceral artery pseudoaneurysms: Case series

    OpenAIRE

    Gamanagatti, Shivanand; Thingujam, Usha; Garg, Pramod; Nongthombam, Surajkumar; Dash, Nihar Ranjan

    2015-01-01

    Pseudoaneurysm is a known complication of pancreatitis associated with significant mortality and morbidity. Imaging plays an important role in the diagnosis and management. Computed tomography (CT) helps localize the lesion and the severity of the background pancreatitis but digital subtraction angiography with coil embolization is recommended to avoid bleeding and inadvertent surgery. However, in cases where angiographic coil embolization is not feasible due to technical reasons, thrombin in...

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

    LENUS (Irish Health Repository)

    Keeling, Aoife N

    2011-07-25

    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.

  8. Raman and surface-enhanced Raman scattering (SERS) studies of the thrombin-binding aptamer.

    Science.gov (United States)

    Wu, Tsai-Chin; Vasudev, Milana; Dutta, Mitra; Stroscio, Michael A

    2013-06-01

    Surface-enhanced Raman scattering is used to study the Raman spectra and peak shifts the thrombin-binding aptamer (TBA) on substrates having two different geometries; one with a single stranded sequence and one with double stranded sequence. The Raman signals of the deoxyribonucleic acids on both substrates are enhanced and specific peaks of bases are identified. These results are highly reproducible and have promising applications in low cost nucleic acid detection.

  9. Allosteric Partial Inhibition of Monomeric Proteases. Sulfated Coumarins Induce Regulation, not just Inhibition, of Thrombin

    OpenAIRE

    Stephen Verespy III; Mehta, Akul Y.; Daniel Afosah; Al-Horani, Rami A.; Desai, Umesh R.

    2016-01-01

    Allosteric partial inhibition of soluble, monomeric proteases can offer major regulatory advantages, but remains a concept on paper to date; although it has been routinely documented for receptors and oligomeric proteins. Thrombin, a key protease of the coagulation cascade, displays significant conformational plasticity, which presents an attractive opportunity to discover small molecule probes that induce sub-maximal allosteric inhibition. We synthesized a focused library of some 36 sulfated...

  10. Direct thrombin inhibitors in acute coronary syndromes: effect in patients undergoing early percutaneous coronary intervention

    OpenAIRE

    Sinnaeve, Peter; Simes, John; Yusuf, Salim; Garg, Jyotsna; Mehta, Shamir; Eikelboom, John; Bittl, John A; Serruys, Patrick; Topol, Eric J.; Granger, Christopher B

    2005-01-01

    AIMS: We evaluated the effect of direct thrombin inhibitors (DTIs) in patients undergoing early percutaneous coronary intervention (PCI), using the DTI Trialists' Collaboration database of 35,970 patients from 11 randomized trials of DTIs vs. heparin. METHODS AND RESULTS: We performed a Cox proportional hazards regression analysis with PCI as a time-dependent covariate to assess the independent impact of DTIs according to the performance of early PCI. PCI was performed in 7049 patients in the...

  11. Supplemental topics on voids

    Energy Technology Data Exchange (ETDEWEB)

    Rood, H.J.

    1988-09-01

    Several topics concerning voids are presented, supplementing the report of Rood (1988). The discovery of the Coma supercluster and void and the recognition of the cosmological significance of superclusters and voids are reviewed. Galaxy redshift surveys and redshift surveys for the Abell clusters and very distant objects are discussed. Solar system and extragalactic dynamics are examined. Also, topics for future observational research on voids are recommended. 50 references.

  12. Topical treatment of melasma

    Directory of Open Access Journals (Sweden)

    Bandyopadhyay Debabrata

    2009-01-01

    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.

  13. Electrochemiluminescence biosensor based on CdSe quantum dots for the detection of thrombin

    International Nuclear Information System (INIS)

    A novel QDs electrochemiluminescence (ECL) biosensor for the determination of thrombin was described. The CdSe QDs solution was dripped onto the clear surface of the ITO and then immersed in PBS which contained EDC and NHS as a coupling agent to activate the carboxyl-terminated surface of the CdSe QDs. The ITO electrode was immersed in the PBS containing 0.4 μM aptamer, followed by rinsing with PBS and dried with N2 again, then dipped in the BSA solution for 30 min to decrease the non-specific binding. After that, the aptamer modified ITO was soaked in PBS to remove unbound aptamer. Under optimal conditions, the linear range was obtained from 0 to 64 μg mL−1 with a correlation coefficient of 0.9986 (n = 16). The control experiment was also carried out by using BSA, lysozyme 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 electrochemical method which could provide a promising platform for fabrication of aptamer based biosensors.

  14. Molecular modeling studies, synthesis and biological evaluation of dabigatran analogues as thrombin inhibitors.

    Science.gov (United States)

    Dong, Ming-Hui; Chen, Hai-Feng; Ren, Yu-Jie; Shao, Fang-Ming

    2016-01-15

    In this work, 48 thrombin inhibitors based on the structural scaffold of dabigatran were analyzed using a combination of molecular modeling techniques. We generated three-dimensional quantitative structure-activity relationship (3D-QSAR) models based on three alignments for both comparative molecular field analysis (CoMFA) and comparative molecular similarity index analysis (CoMSIA) to highlight the structural requirements for thrombin protein inhibition. In addition to the 3D-QSAR study, Topomer CoMFA model also was established with a higher leave-one-out cross-validation q(2) and a non-cross-validation r(2), which suggest that the three models have good predictive ability. The results indicated that the steric, hydrophobic and electrostatic fields play key roles in QSAR model. Furthermore, we employed molecular docking and re-docking simulation explored the binding relationship of the ligand and the receptor protein in detail. Molecular docking simulations identified several key interactions that were also indicated through 3D-QSAR analysis. On the basis of the obtained results, two compounds were designed and predicted by three models, the biological evaluation in vitro (IC50) demonstrated that these molecular models were effective for the development of novel potent thrombin inhibitors.

  15. Mutant N143P Reveals How Na[superscript +] Activates Thrombin

    Energy Technology Data Exchange (ETDEWEB)

    Niu, Weiling; Chen, Zhiwei; Bush-Pelc, Leslie A.; Bah, Alaji; Gandhi, Prafull S.; Di Cera, Enrico; (WU-MED)

    2010-01-12

    The molecular mechanism of thrombin activation by Na{sup +} remains elusive. Its kinetic formulation requires extension of the classical Botts-Morales theory for the action of a modifier on an enzyme to correctly account for the contribution of the E*, E, and E:Na{sup +} forms. The extended scheme establishes that analysis of k{sub cat} unequivocally identifies allosteric transduction of Na{sup +} binding into enhanced catalytic activity. The thrombin mutant N143P features no Na{sup +}-dependent enhancement of k{sub cat} yet binds Na{sup +} with an affinity comparable to that of wild type. Crystal structures of the mutant in the presence and absence of Na{sup +} confirm that Pro{sup 143} abrogates the important H-bond between the backbone N atom of residue 143 and the carbonyl O atom of Glu{sup 192}, which in turn controls the orientation of the Glu{sup 192}-Gly{sup 193} peptide bond and the correct architecture of the oxyanion hole. We conclude that Na{sup +} activates thrombin by securing the correct orientation of the Glu{sup 192}-Gly{sup 193} peptide bond, which is likely flipped in the absence of cation. Absolute conservation of the 143-192 H-bond in trypsin-like proteases and the importance of the oxyanion hole in protease function suggest that this mechanism of Na{sup +} activation is present in all Na{sup +}-activated trypsin-like proteases.

  16. Elevated Cytokines, Thrombin and PAI-1 in Severe HCPS Patients Due to Sin Nombre Virus

    Directory of Open Access Journals (Sweden)

    Virginie Bondu

    2015-02-01

    Full Text Available Sin Nombre Hantavirus (SNV, Bunyaviridae Hantavirus is a Category A pathogen that causes Hantavirus Cardiopulmonary Syndrome (HCPS with case fatality ratios generally ranging from 30% to 50%. HCPS is characterized by vascular leakage due to dysregulation of the endothelial barrier function. The loss of vascular integrity results in non-cardiogenic pulmonary edema, shock, multi-organ failure and death. Using Electric Cell-substrate Impedance Sensing (ECIS measurements, we found that plasma samples drawn from University of New Mexico Hospital patients with serologically-confirmed HCPS, induce loss of cell-cell adhesion in confluent epithelial and endothelial cell monolayers grown in ECIS cultureware. We show that the loss of cell-cell adhesion is sensitive to both thrombin and plasmin inhibitors in mild cases, and to thrombin only inhibition in severe cases, suggesting an increasing prothrombotic state with disease severity. A proteomic profile (2D gel electrophoresis and mass spectrometry of HCPS plasma samples in our cohort revealed robust antifibrinolytic activity among terminal case patients. The prothrombotic activity is highlighted by acute ≥30 to >100 fold increases in active plasminogen activator inhibitor (PAI-1 which, preceded death of the subjects within 48 h. Taken together, this suggests that PAI-1 might be a response to the severe pathology as it is expected to reduce plasmin activity and possibly thrombin activity in the terminal patients.

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

    Science.gov (United States)

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

    2014-07-01

    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

  18. Allosteric Partial Inhibition of Monomeric Proteases. Sulfated Coumarins Induce Regulation, not just Inhibition, of Thrombin

    Science.gov (United States)

    Verespy III, Stephen; Mehta, Akul Y.; Afosah, Daniel; Al-Horani, Rami A.; Desai, Umesh R.

    2016-01-01

    Allosteric partial inhibition of soluble, monomeric proteases can offer major regulatory advantages, but remains a concept on paper to date; although it has been routinely documented for receptors and oligomeric proteins. Thrombin, a key protease of the coagulation cascade, displays significant conformational plasticity, which presents an attractive opportunity to discover small molecule probes that induce sub-maximal allosteric inhibition. We synthesized a focused library of some 36 sulfated coumarins to discover two agents that display sub-maximal efficacy (~50%), high potency (150-fold). Michaelis-Menten, competitive inhibition, and site-directed mutagenesis studies identified exosite 2 as the site of binding for the most potent sulfated coumarin. Stern-Volmer quenching of active site-labeled fluorophore suggested that the allosteric regulators induce intermediate structural changes in the active site as compared to those that display ~80–100% efficacy. Antithrombin inactivation of thrombin was impaired in the presence of the sulfated coumarins suggesting that allosteric partial inhibition arises from catalytic dysfunction of the active site. Overall, sulfated coumarins represent first-in-class, sub-maximal inhibitors of thrombin. The probes establish the concept of allosteric partial inhibition of soluble, monomeric proteins. This concept may lead to a new class of anticoagulants that are completely devoid of bleeding. PMID:27053426

  19. Topical Drugs for Pain Relief

    Directory of Open Access Journals (Sweden)

    Anjali Srinivasan

    2015-03-01

    Full Text Available Topical therapy helps patients with oral and perioral pain problems such as ulcers, burning mouth syndrome, temporomandibular disorders, neuromas, neuropathies and neuralgias. Topical drugs used in the field of dentistry are topical anaesthetics, topical analgesics, topical antibiotics and topical corticosteroids. It provides symptomatic/curative effect. Topical drugs are easy to apply, avoids hepatic first pass metabolism and more sites specific. But it can only be used for medications that require low plasma concentrations to achieve a therapeutic effect.

  20. Implicit stage topics

    Directory of Open Access Journals (Sweden)

    Karen Lahousse

    2008-04-01

    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.

  1. Photoionization and Recombination

    Science.gov (United States)

    Nahar, Sultana N.

    2000-01-01

    Theoretically self-consistent calculations for photoionization and (e + ion) recombination are described. The same eigenfunction expansion for the ion is employed in coupled channel calculations for both processes, thus ensuring consistency between cross sections and rates. The theoretical treatment of (e + ion) recombination subsumes both the non-resonant recombination ("radiative recombination"), and the resonant recombination ("di-electronic recombination") processes in a unified scheme. In addition to the total, unified recombination rates, level-specific recombination rates and photoionization cross sections are obtained for a large number of atomic levels. Both relativistic Breit-Pauli, and non-relativistic LS coupling, calculations are carried out in the close coupling approximation using the R-matrix method. Although the calculations are computationally intensive, they yield nearly all photoionization and recombination parameters needed for astrophysical photoionization models with higher precision than hitherto possible, estimated at about 10-20% from comparison with experimentally available data (including experimentally derived DR rates). Results are electronically available for over 40 atoms and ions. Photoionization and recombination of He-, and Li-like C and Fe are described for X-ray modeling. The unified method yields total and complete (e+ion) recombination rate coefficients, that can not otherwise be obtained theoretically or experimentally.

  2. Rational design and characterization of D-Phe-Pro-D-Arg-derived direct thrombin inhibitors.

    Directory of Open Access Journals (Sweden)

    Ana C Figueiredo

    Full Text Available The tremendous social and economic impact of thrombotic disorders, together with the considerable risks associated to the currently available therapies, prompt for the development of more efficient and safer anticoagulants. Novel peptide-based thrombin inhibitors were identified using in silico structure-based design and further validated in vitro. The best candidate compounds contained both L- and D-amino acids, with the general sequence D-Phe(P3-Pro(P2-D-Arg(P1-P1'-CONH₂. The P1' position was scanned with L- and D-isomers of natural or unnatural amino acids, covering the major chemical classes. The most potent non-covalent and proteolysis-resistant inhibitors contain small hydrophobic or polar amino acids (Gly, Ala, Ser, Cys, Thr at the P1' position. The lead tetrapeptide, D-Phe-Pro-D-Arg-D-Thr-CONH₂, competitively inhibits α-thrombin's cleavage of the S2238 chromogenic substrate with a K(i of 0.92 µM. In order to understand the molecular details of their inhibitory action, the three-dimensional structure of three peptides (with P1' L-isoleucine (fPrI, L-cysteine (fPrC or D-threonine (fPrt in complex with human α-thrombin were determined by X-ray crystallography. All the inhibitors bind in a substrate-like orientation to the active site of the enzyme. The contacts established between the D-Arg residue in position P1 and thrombin are similar to those observed for the L-isomer in other substrates and inhibitors. However, fPrC and fPrt disrupt the active site His57-Ser195 hydrogen bond, while the combination of a P1 D-Arg and a bulkier P1' residue in fPrI induce an unfavorable geometry for the nucleophilic attack of the scissile bond by the catalytic serine. The experimental models explain the observed relative potency of the inhibitors, as well as their stability to proteolysis. Moreover, the newly identified direct thrombin inhibitors provide a novel pharmacophore platform for developing antithrombotic agents by exploring the

  3. Rational design and characterization of D-Phe-Pro-D-Arg-derived direct thrombin inhibitors.

    Science.gov (United States)

    Figueiredo, Ana C; Clement, Cristina C; Zakia, Sheuli; Gingold, Julian; Philipp, Manfred; Pereira, Pedro J B

    2012-01-01

    The tremendous social and economic impact of thrombotic disorders, together with the considerable risks associated to the currently available therapies, prompt for the development of more efficient and safer anticoagulants. Novel peptide-based thrombin inhibitors were identified using in silico structure-based design and further validated in vitro. The best candidate compounds contained both L- and D-amino acids, with the general sequence D-Phe(P3)-Pro(P2)-D-Arg(P1)-P1'-CONH₂. The P1' position was scanned with L- and D-isomers of natural or unnatural amino acids, covering the major chemical classes. The most potent non-covalent and proteolysis-resistant inhibitors contain small hydrophobic or polar amino acids (Gly, Ala, Ser, Cys, Thr) at the P1' position. The lead tetrapeptide, D-Phe-Pro-D-Arg-D-Thr-CONH₂, competitively inhibits α-thrombin's cleavage of the S2238 chromogenic substrate with a K(i) of 0.92 µM. In order to understand the molecular details of their inhibitory action, the three-dimensional structure of three peptides (with P1' L-isoleucine (fPrI), L-cysteine (fPrC) or D-threonine (fPrt)) in complex with human α-thrombin were determined by X-ray crystallography. All the inhibitors bind in a substrate-like orientation to the active site of the enzyme. The contacts established between the D-Arg residue in position P1 and thrombin are similar to those observed for the L-isomer in other substrates and inhibitors. However, fPrC and fPrt disrupt the active site His57-Ser195 hydrogen bond, while the combination of a P1 D-Arg and a bulkier P1' residue in fPrI induce an unfavorable geometry for the nucleophilic attack of the scissile bond by the catalytic serine. The experimental models explain the observed relative potency of the inhibitors, as well as their stability to proteolysis. Moreover, the newly identified direct thrombin inhibitors provide a novel pharmacophore platform for developing antithrombotic agents by exploring the conformational

  4. Recombineering Homologous Recombination Constructs in Drosophila

    OpenAIRE

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

    2013-01-01

    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 recombineeri...

  5. PREFACE: CEWQO Topical Issue CEWQO Topical Issue

    Science.gov (United States)

    Bozic, Mirjana; Man'ko, Margarita

    2009-09-01

    This topical issue of Physica Scripta collects selected peer-reviewed contributions based on invited and contributed talks and posters presented at the 15th Central European Workshop on Quantum Optics (CEWQO) which took place in Belgrade 29 May-3 June 2008 (http://cewqo08.phy.bg.ac.yu). On behalf of the whole community took place in Belgrade 29 May-3 June 2008 (http://cewqo08.phy.bg.ac.yu, cewqo08.phy.bg.ac.yu). On behalf of the whole community of the workshop, we thank the referees for their careful reading and useful suggestions which helped to improve all of the submitted papers. A brief description of CEWQO The Central European Workshop on Quantum Optics is a series of conferences started informally in Budapest in 1992. Sometimes small events transform into important conferences, as in the case of CEWQO. Professor Jozsef Janszky, from the Research Institute of Solid State Physics and Optics, is the founder of this series. Margarita Man'ko obtained the following information from Jozsef Janszky during her visit to Budapest, within the framework of cooperation between the Russian and Hungarian Academies of Sciences in 2005. He organized a small workshop on quantum optics in Budapest in 1992 with John Klauder as a main speaker. Then, bearing in mind that a year before Janszky himself was invited by Vladimir Buzek to give a seminar on the same topic in Bratislava, he decided to assign the name 'Central European Workshop on Quantum Optics', considering the seminar in Bratislava to be the first workshop and the one in Budapest the second. The third formal workshop took place in Bratislava in 1993 organized by Vladimir Buzek, then in 1994 (Budapest, by Jozsef Janszky), 1995 and 1996 (Budmerice, Slovakia, by Vladimir Buzek), 1997 (Prague, by Igor Jex), 1999 (Olomouc, Czech Republic, by Zdenek Hradil), 2000 (Balatonfüred, Hungary, by Jozsef Janszky ), 2001 (Prague, by Igor Jex), 2002 (Szeged, Hungary, by Mihaly Benedict), 2003 (Rostock,Germany, by Werner Vogel and

  6. Activated thrombin-activatable fibrinolysis inhibitor (TAFIa) attenuates breast cancer cell metastatic behaviors through inhibition of plasminogen activation and extracellular proteolysis

    OpenAIRE

    Bazzi, Zainab A.; Lanoue, Danielle; El-Youssef, Mouhanned; Romagnuolo, Rocco; Tubman, Janice; Cavallo-Medved, Dora; Porter, Lisa A.; Boffa, Michael B.

    2016-01-01

    Background Thrombin activatable fibrinolysis inhibitor (TAFI) is a plasma zymogen, which can be converted to activated TAFI (TAFIa) through proteolytic cleavage by thrombin, plasmin, and most effectively thrombin in complex with the endothelial cofactor thrombomodulin (TM). TAFIa is a carboxypeptidase that cleaves carboxyl terminal lysine and arginine residues from protein and peptide substrates, including plasminogen-binding sites on cell surface receptors. Carboxyl terminal lysine residues ...

  7. Topical ketoprofen patch.

    Science.gov (United States)

    Mazières, Bernard

    2005-01-01

    Although oral nonsteroidal anti-inflammatory drugs (NSAIDs) are effective in the treatment of a variety of acute and chronic pain conditions, their use may be associated with serious systemic adverse effects, particularly gastrointestinal disorders. In order to minimise the incidence of systemic events related to such agents, topical NSAIDs have been developed. Topical NSAIDs, applied as gels, creams or sprays, penetrate the skin, subcutaneous fatty tissue and muscle in amounts that are sufficient to exert a therapeutic effect on peripheral and central mechanisms in the absence of high plasma concentrations. Data indicate that topical NSAIDs are effective at relieving pain in a number of acute and chronic pain indications. This review article discusses the pharmacokinetics, efficacy and tolerability of a new formulation of ketoprofen available as a topical patch. The topical patch containing ketoprofen 100mg as the active principle has been developed using a novel delivery system that dispenses therapeutic doses of the drug directly to the site of injury. Pharmacokinetic data indicate that although plasma levels of ketoprofen are higher when the drug is administered as a patch versus a gel, the total systemic bioavailability of ketoprofen 100 mg administered via a patch is no more than 10% of that reported for ketoprofen 100 mg administered orally. Because the patch facilitates ketoprofen delivery over a 24-hour period, the drug remains continually present in the tissue subjacent to the site of application. High tissue but low plasma ketoprofen concentrations mean that while tissue concentrations are high enough to exert a therapeutic effect, plasma concentrations remain low enough to not result in systemic adverse events caused by elevated serum NSAID levels. Phase III clinical trials in patients with non-articular rheumatism and traumatic painful soft tissue injuries showed that the topical ketoprofen patch was significantly more effective than placebo at

  8. Topics in Physical Mathematics

    CERN Document Server

    Marathe, Kishore

    2010-01-01

    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

  9. A label-free and high sensitive aptamer biosensor based on hyperbranched polyester microspheres for thrombin detection

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Chong [Jiangsu Key Laboratory of Biofunctional Materials, Biomedical Functional Materials Collaborative Innovation Center, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); Institute of Agricultural Products Processing, Jiangsu Academy of Agricultural Sciences, Nanjing 210014 (China); Han, Qiaorong [Jiangsu Key Laboratory of Biofunctional Materials, Biomedical Functional Materials Collaborative Innovation Center, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); Wang, Daoying; Xu, Weimin [Institute of Agricultural Products Processing, Jiangsu Academy of Agricultural Sciences, Nanjing 210014 (China); Wang, Weijuan [Jiangsu Key Laboratory of Biofunctional Materials, Biomedical Functional Materials Collaborative Innovation Center, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); Zhao, Wenbo, E-mail: zhaowenbo@njnu.edu.cn [Jiangsu Key Laboratory of Biofunctional Materials, Biomedical Functional Materials Collaborative Innovation Center, College of Chemistry and Materials Science, Nanjing Normal University, Nanjing 210023 (China); Zhou, Min, E-mail: zhouminnju@126.com [Department of Vascular Surgery, the Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing 210008 (China)

    2014-11-19

    Highlights: • A label-free thrombin aptamer biosensor applied in whole blood has been developed. • The aptamer biosensor showed a wide detection range and a low detection limit. • The antibiofouling idea utilized for biosensor is significant for diagnostics. - Abstract: In this paper, we have synthesized hyperbranched polyester microspheres with carboxylic acid functional groups (HBPE-CA) and developed a label-free electrochemical aptamer biosensor using thrombin-binding aptamer (TBA) as receptor for the measurement of thrombin in whole blood. The indium tin oxide (ITO) electrode surface modified with HBPE-CA microspheres was grafted with TBA, which has excellent binding affinity and selectivity for thrombin. Binding of the thrombin at the modified ITO electrode surface greatly restrained access of electrons for a redox probe of [Fe(CN){sub 6}]{sup 3−/4−}. Moreover, the aptamer biosensor could be used for detection of thrombin in whole blood, a wide detection range (10 fM–100 nM) and a detection limit on the order of 0.90 fM were demonstrated. Control experiments were also carried out by using bull serum albumin (BSA) and lysozyme in the absence of thrombin. The good stability and repeatability of this aptamer biosensor were also proved. We expect that this demonstration will lead to the development of highly sensitive label-free sensors based on aptamer with lower cost than current technology. The integration of the technologies, which include anticoagulant, sensor and nanoscience, will bring significant input to high-performance biosensors relevant to diagnostics and therapy of interest for human health.

  10. A label-free and high sensitive aptamer biosensor based on hyperbranched polyester microspheres for thrombin detection

    International Nuclear Information System (INIS)

    Highlights: • A label-free thrombin aptamer biosensor applied in whole blood has been developed. • The aptamer biosensor showed a wide detection range and a low detection limit. • The antibiofouling idea utilized for biosensor is significant for diagnostics. - Abstract: In this paper, we have synthesized hyperbranched polyester microspheres with carboxylic acid functional groups (HBPE-CA) and developed a label-free electrochemical aptamer biosensor using thrombin-binding aptamer (TBA) as receptor for the measurement of thrombin in whole blood. The indium tin oxide (ITO) electrode surface modified with HBPE-CA microspheres was grafted with TBA, which has excellent binding affinity and selectivity for thrombin. Binding of the thrombin at the modified ITO electrode surface greatly restrained access of electrons for a redox probe of [Fe(CN)6]3−/4−. Moreover, the aptamer biosensor could be used for detection of thrombin in whole blood, a wide detection range (10 fM–100 nM) and a detection limit on the order of 0.90 fM were demonstrated. Control experiments were also carried out by using bull serum albumin (BSA) and lysozyme in the absence of thrombin. The good stability and repeatability of this aptamer biosensor were also proved. We expect that this demonstration will lead to the development of highly sensitive label-free sensors based on aptamer with lower cost than current technology. The integration of the technologies, which include anticoagulant, sensor and nanoscience, will bring significant input to high-performance biosensors relevant to diagnostics and therapy of interest for human health

  11. Mutant B-Raf(V600E) Promotes Melanoma Paracellular Transmigration by Inducing Thrombin-mediated Endothelial Junction Breakdown.

    Science.gov (United States)

    Zhang, Pu; Feng, Shan; Liu, Gentao; Wang, Heyong; Zhu, Huifeng; Ren, Qiao; Bai, Huiyuan; Fu, Changliang; Dong, Cheng

    2016-01-29

    Tumor invasiveness depends on the ability of tumor cells to breach endothelial barriers. In this study, we investigated the mechanism by which the adhesion of melanoma cells to endothelium regulates adherens junction integrity and modulates tumor transendothelial migration (TEM) by initiating thrombin generation. We found that the B-Raf(V600E) mutation in metastatic melanoma cells up-regulated tissue factor (TF) expression on cell membranes and promoted thrombin production. Co-culture of endothelial monolayers with metastatic melanoma cells mediated the opening of inter-endothelial spaces near melanoma cell contact sites in the presence of platelet-free plasma (PFP). By using small interfering RNA (siRNA), we demonstrated that B-Raf(V600E) and TF silencing attenuated the focal disassembly of adherens junction induced by tumor contact. Vascular endothelial-cadherin (VE-cadherin) disassembly was dependent on phosphorylation of p120-catenin on Ser-879 and VE-cadherin on Tyr-658, Tyr-685, and Tyr-731, which can be prevented by treatment with the thrombin inhibitor, hirudin, or by silencing the thrombin receptor, protease-activated receptor-1, in endothelial cells. We also provided strong evidence that tumor-derived thrombin enhanced melanoma TEM by inducing ubiquitination-coupled VE-cadherin internalization, focal adhesion formation, and actin assembly in endothelium. Confocal microscopic analysis of tumor TEM revealed that junctions transiently opened and resealed as tumor cells accomplished TEM. In addition, in the presence of PFP, tumor cells preferentially transmigrated via paracellular routes. PFP supported melanoma transmigration under shear conditions via a B-Raf(V600E)-thrombin-dependent mechanism. We concluded that the activation of thrombin generation by cancer cells in plasma is an important process regulating melanoma extravasation by disrupting endothelial junction integrity. PMID:26504080

  12. METHODS OF TREATING OR PREVENTING DEMYELINATION USING THROMBIN INHIBITORS AND METHODS OF DETECTING DEMYELINATION USING NEUROFASCIN 155 | NCI Technology Transfer Center | TTC

    Science.gov (United States)

    Researchers at the Eunice Kennedy Shriver National Institute of Child Health and Human Development (“NICHD”), seek CRADA partner or collaboration for development of agents to treat multiple sclerosis or other conditions associated with myelin remodeling by administering an agent that inhibits cleavage of Neurofascin 155 or Caspr1. The agent could be a thrombin inhibitor, an agent that inhibits thrombin expression, an anti-thrombin antibody that specifically inhibits thrombin mediated cleavage of Neurofascin 155, a mutated version or fragment of Neurofascin 155 or Caspr1, or antibodies to Neurofascin 155 or Caspr1.

  13. Topics in Nonlinear Dynamics

    DEFF Research Database (Denmark)

    Mosekilde, Erik

    Through a significant number of detailed and realistic examples this book illustrates how the insights gained over the past couple of decades in the fields of nonlinear dynamics and chaos theory can be applied in practice. Aomng the topics considered are microbiological reaction systems, ecological...

  14. Historical Topics in Algebra.

    Science.gov (United States)

    National Council of Teachers of Mathematics, Inc., Reston, VA.

    This is a reprint of the historical capsules dealing with algebra from the 31st Yearbook of NCTM,"Historical Topics for the Mathematics Classroom." Included are such themes as the change from a geometric to an algebraic solution of problems, the development of algebraic symbolism, the algebraic contributions of different countries, the origin and…

  15. Transportation: Topic Paper E.

    Science.gov (United States)

    National Council on the Handicapped, Washington, DC.

    As one of a series of topic papers assessing federal laws and programs affecting persons with disabilities, this paper reviews the issue of transportation services. In the area of urban mass transit, four relevant pieces of legislation and public transportation accessibility regulations are cited, and cost issues are explored. Paratransit systems,…

  16. Differential Topic Models.

    Science.gov (United States)

    Chen, Changyou; Buntine, Wray; Ding, Nan; Xie, Lexing; Du, Lan

    2015-02-01

    In applications we may want to compare different document collections: they could have shared content but also different and unique aspects in particular collections. This task has been called comparative text mining or cross-collection modeling. We present a differential topic model for this application that models both topic differences and similarities. For this we use hierarchical Bayesian nonparametric models. Moreover, we found it was important to properly model power-law phenomena in topic-word distributions and thus we used the full Pitman-Yor process rather than just a Dirichlet process. Furthermore, we propose the transformed Pitman-Yor process (TPYP) to incorporate prior knowledge such as vocabulary variations in different collections into the model. To deal with the non-conjugate issue between model prior and likelihood in the TPYP, we thus propose an efficient sampling algorithm using a data augmentation technique based on the multinomial theorem. Experimental results show the model discovers interesting aspects of different collections. We also show the proposed MCMC based algorithm achieves a dramatically reduced test perplexity compared to some existing topic models. Finally, we show our model outperforms the state-of-the-art for document classification/ideology prediction on a number of text collections. PMID:26353238

  17. Selected topics in magnetism

    CERN Document Server

    Gupta, L C

    1993-01-01

    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.

  18. Topic: Mastery Learning.

    Science.gov (United States)

    Killoran, James, Ed.

    1983-01-01

    This journal issue addresses the topic of mastery learning at both the elementary and secondary school levels. The first article, "The Theory and Practice of Mastery Learning" (Guskey), gives a definition of and information about the development, operation, and application of mastery learning, based on the theories of Benjamin Bloom. In addition,…

  19. Topical Research: Africa.

    Science.gov (United States)

    Lynn, Karen

    This lesson plan can be used in social studies, language arts, or library research. The instructional objective is for students to select a topic of study relating to Africa, write a thesis statement, collect information from media sources, and develop a conclusion. The teacher may assign the lesson for written or oral evaluation. The teacher…

  20. Contrastive topics decomposed

    Directory of Open Access Journals (Sweden)

    Michael Wagner

    2012-12-01

    Full Text Available The analysis of contrastive topics introduced in Büring 1997b and further developed in Büring 2003 relies on distinguishing two types of constituents that introduce alternatives: the sentence focus, which is marked by a FOC feature, and the contrastive topic, which is marked by a CT feature. A non-compositional rule of interpretation that refers to these features is used to derive a topic semantic value, a nested set of sets of propositions. This paper presents evidence for a correlation between the restrictive syntax of nested focus operators and the syntax of contrastive topics, a correlation which is unexpected under this analysis. A compositional analysis is proposed that only makes use of the flatter focus semantic values introduced by focus operators. The analysis aims at integrating insights from the original analysis while at the same time capturing the observed syntactic restrictions. http://dx.doi.org/10.3765/sp.5.8 BibTeX info

  1. Characters and Topical Diversity

    DEFF Research Database (Denmark)

    Eriksson, Rune

    2014-01-01

    The purpose of this article is to contribute to our understanding of the difference between the bestseller and the non-bestseller in nonfiction. It is noticed that many bestsellers in nonfiction belongs to the sub-genre of creative nonfiction, but also that the topics in this kind of literature i...

  2. Topics and topic prominence in two sign languages

    NARCIS (Netherlands)

    V. Kimmelman

    2015-01-01

    In this paper we describe topic marking in Russian Sign Language (RSL) and Sign Language of the Netherlands (NGT) and discuss whether these languages should be considered topic prominent. The formal markers of topics in RSL are sentence-initial position, a prosodic break following the topic, and non

  3. Enhancement of aptamer immobilization using egg shell-derived nano-sized spherical hydroxyapatite for thrombin detection in neuroclinic.

    Science.gov (United States)

    Derkus, Burak; Arslan, Yavuz Emre; Emregul, Kaan C; Emregul, Emel

    2016-09-01

    In the present study, we describe the sonochemical isolation of nano-sized spherical hydroxyapatite (nHA) from egg shell and application towards thrombin aptasensing. In addition to the sonochemical method, two conventional methods present in literature were carried out to perform a comparative study. Various analysis methods including Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Fourier Transform Infrared Spectroscopy (FTIR), X-Ray Diffraction (XRD), Energy-Dispersive Analysis of X-Rays (EDAX), and Thermal Gravimetric Analysis (TGA) have been applied for the characterization of nHA and its nanocomposite with marine-derived collagen isolated from Rhizostoma pulmo jellyfish. TEM micrographs revealed the sonochemically synthesized nHA nanoparticles to have a unique porous spherical shape with a diameter of approximately 60-80nm when compared to hydroxyapatite nanoparticles synthesized using the other two methods which had a typical needle shaped morphology. EDAX, XRD and FTIR results demonstrated that the obtained patterns belonged to hydroxyapatite. Electrochemical impedance spectroscopy (EIS) is the main analyzing technique of the developed thrombin aptasensor. The proposed aptasensor has a detection limit of 0.25nM thrombin. For clinical application of the developed aptasensor, thrombin levels in blood and cerebrospinal fluid (CSF) samples obtained from patients with Multiple Sclerosis, Myastenia Gravis, Epilepsy, Parkinson, polyneuropathy and healthy donors were analyzed using both the aptasensor and commercial ELISA kit. The results showed that the proposed system is a promising candidate for clinical analysis of thrombin. PMID:27343583

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

    Energy Technology Data Exchange (ETDEWEB)

    Liang Guohai; Cai Shaoyu; Zhang Peng [Department of Chemistry and Institutes of Biomedical Sciences, Fudan University, Shanghai 200433 (China); Peng Youyuan [Department of Chemistry, Quanzhou Normal University, Quanzhou 362000 (China); Chen Hui; Zhang Song [Department of Chemistry and Institutes of Biomedical Sciences, Fudan University, Shanghai 200433 (China); Kong Jilie, E-mail: jlkong@fudan.edu.cn [Department of Chemistry and Institutes of Biomedical Sciences, Fudan University, Shanghai 200433 (China)

    2011-03-18

    We describe a sensitive biosensing system combining magnetic relaxation switch diagnosis and colorimetric detection of human {alpha}-thrombin, based on the aptamer-protein interaction induced aggregation of Fe{sub 3}O{sub 4}-Au nanoparticles. To demonstrate the concept, gold-coated iron oxide nanoparticle was synthesized by iterative reduction of HAuCl{sub 4} onto the dextran-coated Fe{sub 3}O{sub 4} 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 {alpha}-thrombin were conjugated separately to two distinct nanorose populations. Once a solution containing human {alpha}-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 (T{sub 2}) as well as the UV-Vis absorption spectra of the solution. Thus the qualitative and quantitative detection method for human {alpha}-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 T{sub 2} change is observed with 1.0 nM human {alpha}-thrombin, and the detection range is from 1.6 nM to 30.4 nM.

  5. Endoscopic injection sclerotherapy in non-variceal upper gastrointestinal bleeding. A comparative study of polidocanol and thrombin.

    Science.gov (United States)

    Benedetti, G; Sablich, R; Lacchin, T

    1991-01-01

    To date several agents have been used to achieve haemostasis in patients with non-variceal upper gastrointestinal bleeding using endoscopic sclerotherapy techniques. Polidocanol has been widely used but local complications have been reported after treatment. We have compared the efficacy and safety of thrombin and polidocanol in 82 consecutive patients with ongoing or recent bleeding from duodenal, gastric, or anastomotic ulcers. Primary control of haemostasis from spurting vessels was achieved in 90% of cases using polidocanol and in 86.6% using thrombin. Definitive haemostasis was obtained in 80% of patients in both groups. When a non-bleeding vessel was visible, injection of polidocanol or thrombin effectively prevented rebleeding in 90.9% and 85.7% of cases, respectively. When a non-bleeding sentinel clot was present, injection of polidocanol or thrombin provided definitive haemostasis in 100% and 92.8% of cases, respectively. No statistically significant difference was evident between the two agents. In the polidocanol group, one local haemorrhagic complication was noted. No general or local complications were recorded in the thrombin group.

  6. Effects of thrombin, PAR-1 activating peptide and a PAR-1 antagonist on umbilical artery resistance in vitro

    Directory of Open Access Journals (Sweden)

    Elliott John T

    2005-02-01

    Full Text Available Abstract Background The non-thrombotic effects of thrombin in cardiovascular tissues, as mediated via the protease activated receptors (PARs, and particularly PAR-1, have been the focus of much recent research. The aims of this study were to evaluate the effects of thrombin, a specific PAR-1 activating peptide (PAR1-AP, and a PAR-1 antagonist on human umbilical artery tone in vitro. Methods Human umbilical artery samples were obtained from 17 women at term. Arterial rings were suspended under physiologic conditions for isometric recording. The in vitro effects of thrombin (0.5 units/mL to 3 units/mL, PAR1-AP TFLLR-NH2 [10(-9 to 10(-6 M], and PAR-1 antagonist (N-trans cinnamoyl- p-fluoroPhe-p-guanidinoPhe-Leu-Arg-Orn-NH2 [10(-9 M to 10(-5 M] on umbilical artery tone were measured. Results Both thrombin and TFLLR-NH2 exerted a potent cumulative vasodilatory effect on human umbilical artery resistance (P 0.05. Conclusion These findings highlight a potential role for thrombin and PAR-1 receptors in vascular regulation of feto-placental blood flow in normal pregnancy, and in association with the vascular lesions associated with IUGR and pre-eclampsia.

  7. Thrombin in combination with intensive nursing in treating upper gastrointestinal bleeding in children.

    Science.gov (United States)

    Yang, F; Xiang, M L; Liu, Y M

    2016-01-01

    Pediatric upper gastrointestinal bleeding, a commonly seen pediatric emergency, needs timely symptomatic treatment to avoid a worse outcome. To discuss the clinical effect of thrombin treatment in combination with intensive nursing on pediatric upper gastrointestinal bleeding, this study analyzed 128 children who were treated in the second ward of the Children’s Internal Medical Department in the First Affiliated Hospital of Zhengzhou University between February 2012 and December 2014. The patients were divided into two groups, an experimental group and a control group. Besides thrombin, the experimental group was given intensive nursing, consisting of regular nursing and targeted nursing, while the control group was given regular nursing only. Clinical indexes of the two groups, such as effective rate, nursing satisfaction and side effect rate, were compared. Relevant clinical indexes such as duration of hospital stay, time to stopping of bleeding and Self-Rating Anxiety Scale (SAS) score, as well as overall satisfaction level of the observation group were all better than those of the control group and differences between the two groups had statistical significance (P less than 0.05). Furthermore, difference of overall effective rate between the experimental group (90.63%) and the control group (68.75%) was significant. Difference of incidence of side effects between the two groups was statistically significant. Thus thrombin treatment in combination with intensive nursing proved to have a remarkable clinical effect and high safety level in treating pediatric upper gastrointestinal bleeding and, moreover, it shortens treatment time and enhances the patients’ quality of life. PMID:27358137

  8. High-level expression and preparation of recombinant human fibrinogen as biopharmaceuticals.

    Science.gov (United States)

    Hirashima, Masaki; Imamura, Takayuki; Yano, Kentaro; Kawamura, Ryoichi; Meta, Akihiro; Tokieda, Yoshiyuki; Nakashima, Toshihiro

    2016-02-01

    Fibrinogen is a large and complex glycoprotein containing two sets of each of three different chains (α, β and γ). There have been no reports of high-level expression of fibrinogen at commercial levels using mammalian cultured cells such as CHO cells because of the difficulty in highly expressing a protein with such a complex structure. We achieved high-level (1.3 g/l or higher) expression of recombinant human fibrinogen using CHO DG44 cells by optimizing the expression system and culture conditions. We also succeeded in establishing a high-recovery preparation method for recombinant fibrinogen that rarely yields degraded products. To characterize the properties of the recombinant human fibrinogen, we performed SDS-PAGE; western blotting of the α, β and γ chains using specific antibodies and scanning electron microscopy observations of fibrin fibres. We also evaluated the functional equivalence between recombinant fibrinogen and plasma fibrinogen with respect to the release of fibrinopeptides initiated by thrombin and its cross-linking properties. The basic properties of recombinant fibrinogen showed no apparent differences from those of plasma fibrinogen. Here, we report the development of methods for the culture and preparation of recombinant human fibrinogen of satisfactory quality that can be scaled up to the commercial level.

  9. Structure-based organic synthesis of unnatural aeruginosin hybrids as potent inhibitors of thrombin.

    Science.gov (United States)

    Hanessian, Stephen; Ersmark, Karolina; Wang, Xiaotian; Del Valle, Juan R; Blomberg, Niklas; Xue, Yafeng; Fjellström, Ola

    2007-06-15

    Based on X-ray crystallographic data of complexes of chlorodysinosin A with the enzyme thrombin, a series of analogs were synthesized varying the nature of the P(1), P(2), and P(3) pharmacophoric sites and the central octahydroindole carboxyamide core. In general, introduction of a hydrophobic substituent on the d-leucine amide residue dramatically improved the inhibition of the enzyme. This is rationalized based on a better fit of the P(3) subunit in the hydrophobic S(3) enzyme site. Single digit nanomolar inhibition expressed as IC(50) was observed for several analogs. PMID:17428662

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

    OpenAIRE

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

    2000-01-01

    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 ...

  11. Topical retinoids for acne.

    Science.gov (United States)

    Yeh, Lindsey; Bonati, Lauren Meshkov; Silverberg, Nanette B

    2016-06-01

    Topical retinoids are currently approved by the US Food and Drug Administration for the treatment of acne vulgaris in nonpregnant, nonlactating patients 12 years of age and older. Their efficacy, safety, and tolerability are well documented for inflammatory and noninflammatory acne with studies repeatedly demonstrating a decrease in the number of lesions, significant improvement in acne severity, improvement in the cosmetic appearance of acne, and the prevention of acne lesions through microcomedone formation. There is some variability between prescription retinoid products regarding efficacy, safety, and tolerability; with erythema, peeling, and dryness being common, potential side effects. Due to their efficacious and safe profile, topical retinoids remain the first-line treatment for acne vulgaris. PMID:27416308

  12. pHAST (pH-Driven Aptamer Switch for Thrombin) Catch-and-Release of Target Protein.

    Science.gov (United States)

    McConnell, E M; Bolzon, R; Mezin, P; Frahm, G; Johnston, M; DeRosa, M C

    2016-06-15

    A pH-driven DNA nanomachine based on the human α-thrombin binding aptamer was designed for the specific catch-and-release of human α-thrombin at neutral and acidic pH, respectively. In neutral conditions, the thrombin aptamer component of the nanomachine is exposed and exists in the G-quadruplex conformation required to bind to the target protein. At slightly acidic pH, the polyadenine tail of the nanomachine becomes partially protonated and A+(anti)•G(syn) mispairing results in a conformational change, causing the target protein to be released. Förster resonance energy transfer (FRET) was used to monitor conformational switching over multiple pH cycles. Electrophoretic mobility shift assay (EMSA) and fluorescence anisotropy were used to show pH dependent protein binding and release by the nanomachine. This approach could be applied generally to existing G-rich aptamers to develop novel biosensors, theranostics, and nanoswitches. PMID:27115292

  13. Protein kinase C is differentially regulated by thrombin, insulin, and epidermal growth factor in human mammary tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Gomez, M.L.; Tellez-Inon, M.T. (Instituto de Ingenieria Genetica y Biologia Molecular, Buenos Aires (Argentina)); Medrano, E.E.; Cafferatta, E.G.A. (Instituto de Investigaciones Bioquimicas Fundacion Campomar, Buenos Aires (Argentina))

    1988-03-01

    The exposure of serum-deprived mammary tumor cells MCF-7 and T-47D to insulin, thrombin, and epidermal growth factor (EGF) resulted in dramatic modifications in the activity and in the translocation capacity of protein kinase C from cytosol to membrane fractions. Insulin induces a 600% activation of the enzyme after 5 h of exposure to the hormone in MCF-7 cells; thrombin either activates (200% in MCF-7) or down-regulates (in T-47D), and EGF exerts only a moderate effect. Thus, the growth factors studied modulate differentially the protein kinase C activity in human mammary tumor cells. The physiological significance of the results obtained are discussed in terms of the growth response elicited by insulin, thrombin, and EGF.

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

    Directory of Open Access Journals (Sweden)

    Huang-Joe Wang

    2014-01-01

    Full Text Available 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.

  15. MicroRNA-146a decreases high glucose/thrombin-induced endothelial inflammation by inhibiting NAPDH oxidase 4 expression.

    Science.gov (United States)

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

    2014-01-01

    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

  16. Deletion of the thrombin cleavage domain of osteopontin mediates breast cancer cell adhesion, proteolytic activity, tumorgenicity, and metastasis

    International Nuclear Information System (INIS)

    Osteopontin (OPN) is a secreted phosphoprotein often overexpressed at high levels in the blood and primary tumors of breast cancer patients. OPN contains two integrin-binding sites and a thrombin cleavage domain located in close proximity to each other. To study the role of the thrombin cleavage site of OPN, MDA-MB-468 human breast cancer cells were stably transfected with either wildtype OPN (468-OPN), mutant OPN lacking the thrombin cleavage domain (468-ΔTC) or an empty vector (468-CON) and assessed for in vitro and in vivo functional differences in malignant/metastatic behavior. All three cell lines were found to equivalently express thrombin, tissue factor, CD44, αvβ5 integrin and β1 integrin. Relative to 468-OPN and 468-CON cells, 468-ΔTC cells expressing OPN with a deleted thrombin cleavage domain demonstrated decreased cell adhesion (p < 0.001), decreased mRNA expression of MCAM, maspin and TRAIL (p < 0.01), and increased uPA expression and activity (p < 0.01) in vitro. Furthermore, injection of 468-ΔTC cells into the mammary fat pad of nude mice resulted in decreased primary tumor latency time (p < 0.01) and increased primary tumor growth and lymph node metastatic burden (p < 0.001) compared to 468-OPN and 468-CON cells. The results presented here suggest that expression of thrombin-uncleavable OPN imparts an early tumor formation advantage as well as a metastatic advantage for breast cancer cells, possibly due to increased proteolytic activity and decreased adhesion and apoptosis. Clarification of the mechanisms responsible for these observations and the translation of this knowledge into the clinic could ultimately provide new therapeutic opportunities for combating breast cancer

  17. Thermodynamic and biological evaluation of a thrombin binding aptamer modified with several unlocked nucleic acid (UNA) monomers and a 2′-C-piperazino-UNA monomer

    DEFF Research Database (Denmark)

    Jensen, Troels B.; Henriksen, Jonas Rosager; Rasmussen, Bjarne E.;

    2011-01-01

    Thrombin binding aptamer is a DNA 15-mer which forms a G-quadruplex structure and possess promising anticoagulant properties due to specific interactions with thrombin. Herein we present the influence of a single 2′-C-piperazino-UNA residue and UNA residues incorporated in several positions on th...

  18. Aptamer-based fluorescent solid-phase thrombin assay using a silver-coated glass substrate and signal amplification by glucose oxidase

    International Nuclear Information System (INIS)

    We describe an aptamer-based solid-state biosensor for the fluorometric determination of thrombin. The surface of silver-coated glass was modified with the thrombin-binding aptamer 1 (TBA 1) of the sequence 5′-HS-TTT TTT TTT TTT TTT GGT TGG TGT GGT TGG-3′. A second (and biotinylated) thrombin -binding aptamer (TBA 2) with the sequence 5′-biotin-AGT CCG TGG TAG GGC AGG TTG GGG TGA CT-3′ was applied as the signaling aptamer. Following binding of thrombin by TBA 1 on the surface, TBA 2 is added and then binds to the thrombin on the surface of the silver-coated glass to form the thrombin-aptamer complex. The biotin groups on TBA 2 are then coated with streptavidin, and biotin-labeled glucose oxidase (biotin-GOx) is added to bind to streptavidin. The quantity of GOx immobilized in this way is directly related to the quantity of thrombin bound on the surface. Following cleavage of the aptamer with DNase I, glucose is added and oxidized by GOx to yield H2O2. Horseradish peroxidase is added and causes the oxidation of 3-p-hydroxyphenylpropanoic acid to yield a fluorescent product. The intensity of the blue fluorescence is directly related to the thrombin concentration in the 300 pM to 6500 pM range, and the detection limit is as low as 82 pM. The assay has good selectivity and practicability. (author)

  19. Aptamer-based luminescence energy transfer from near-infrared-to-near-infrared upconverting nanoparticles to gold nanorods and its application for the detection of thrombin.

    Science.gov (United States)

    Yuan, Fei; Chen, Hongqi; Xu, Juan; Zhang, Yiyan; Wu, Yong; Wang, Lun

    2014-03-01

    A new luminescence energy transfer (LET) system has been designed for the detection of thrombin in the near-infrared (NIR) region by utilizing NIR-to-NIR upconversion lanthanide nanophosphors (UCNPs) as the donor and gold nanorods (Au NRs) as the acceptor. The use of upconverting NaYF4 :Yb(3+) ,Tm(3+) nanoparticles with sharp NIR emission peaks upon NIR excitation by an inexpensive infrared continuous wave laser diode provided large spectral overlap between the donor and the acceptor. Both the Au NRs and carboxyl-terminated NaYF4 :Yb(3+) ,Tm(3+) UCNPs were first modified with different thrombin aptamers. When thrombin was added, a LET system was then formed because of the specific recognition between the thrombin aptamers and thrombin. The LET system was used to monitor thrombin concentrations in aqueous buffer and human blood samples. The limits of detection for thrombin are as low as 0.118 nM in buffer solution and 0.129 nM in human serum. The method was also successfully applied to thrombin detection in blood samples. PMID:24501010

  20. Inactivation of active thrombin-activable fibrinolysis inhibitor takes place by a process that involves conformational instability rather than proteolytic cleavage

    NARCIS (Netherlands)

    Marx, PF; Hackeng, TM; Dawson, PE; Griffin, JH; Meijers, JCM; Bouma, Bonno N.

    2000-01-01

    Thrombin-activable fibrinolysis inhibitor (TAFI) is present in the circulation as an inactive zymogen. Thrombin converts TAFI to a carboxypeptidase B-like enzyme (TAFIa) by cleaving at Arg(92) in a process accelerated by the cofactor, thrombomodulin. TAFIa attenuates fibrinolysis. TAFIa can be inact

  1. Effect of long-term hormone replacement therapy on tissue factor pathway inhibitor and thrombin activatable fibrinolysis inhibitor in healthy postmenopausal women: a randomized controlled study

    DEFF Research Database (Denmark)

    Bladbjerg, E-M; Madsen, J S; Kristensen, S R;

    2003-01-01

    arterial tissue factor expression and higher thrombin formation, and changes in tissue factor pathway coagulation inhibitor (TFPI) and thrombin activatable fibrinolysis inhibitor (TAFI) may be deleterious. Healthy postmenopausal women (n = 719) were randomized to hormone therapy [n = 357; opposed (n = 290...

  2. Prelabeled glycoprotein Ib/IX receptors are not cleared from exposed surfaces of thrombin-activated platelets.

    OpenAIRE

    White, J. G.; Krumwiede, M. D.; Cocking-Johnson, D.; Escolar, G.

    1996-01-01

    The present investigation has re-examined the hypothesis proposing that glycoprotein (GP)Ib/IX receptors for von Willebrand factor are rapidly cleared from exposed surfaces to internal membrane systems after activation of platelets by thrombin in suspension. Platelets were prelabeled with either a polyclonal antibody to GPIb alpha, antiglycocalicin (A-Gl), or a cocktail of two monoclonal antibodies, AP1 and 6D1, exposed to 0.1 or 0.2 U/ml thrombin for 5 or 10 minutes, fixed and stained with S...

  3. Phosphoproteomic analysis of platelets activated by pro-thrombotic oxidized phospholipids and thrombin.

    Directory of Open Access Journals (Sweden)

    Alejandro Zimman

    Full Text Available Specific oxidized phospholipids (oxPCCD36 promote platelet hyper-reactivity and thrombosis in hyperlipidemia via the scavenger receptor CD36, however the signaling pathway(s induced in platelets by oxPCCD36 are not well defined. We have employed mass spectrometry-based tyrosine, serine, and threonine phosphoproteomics for the unbiased analysis of platelet signaling pathways induced by oxPCCD36 as well as by the strong physiological agonist thrombin. oxPCCD36 and thrombin induced differential phosphorylation of 115 proteins (162 phosphorylation sites and 181 proteins (334 phosphorylation sites respectively. Most of the phosphoproteome changes induced by either agonist have never been reported in platelets; thus they provide candidates in the study of platelet signaling. Bioinformatic analyses of protein phosphorylation dependent responses were used to categorize preferential motifs for (dephosphorylation, predict pathways and kinase activity, and construct a phosphoproteome network regulating integrin activation. A putative signaling pathway involving Src-family kinases, SYK, and PLCγ2 was identified in platelets activated by oxPCCD36. Subsequent ex vivo studies in human platelets demonstrated that this pathway is downstream of the scavenger receptor CD36 and is critical for platelet activation by oxPCCD36. Our results provide multiple insights into the mechanism of platelet activation and specifically in platelet regulation by oxPCCD36.

  4. A sensitive HIV-1 envelope induced fusion assay identifies fusion enhancement of thrombin

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, De-Chun; Zhong, Guo-Cai; Su, Ju-Xiang [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Liu, Yan-Hong [Second Affiliated Hospital of Harbin Medical University, 246 Xuefu Road, Harbin, Heilongjiang 150081 (China); Li, Yan; Wang, Jia-Ye [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Hattori, Toshio [Department of Emerging Infectious Diseases, Division of Internal Medicine, Graduate School of Medicine, Tohoku University, Sendai 9808574 (Japan); Ling, Hong, E-mail: lingh@ems.hrbmu.edu.cn [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Department of Parasitology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Key Lab of Heilongjiang Province for Infection and Immunity, Key Lab of Heilongjiang Province Education Bureau for Etiology, Harbin, Heilongjiang 150081 (China); Zhang, Feng-Min, E-mail: fengminzhang@yahoo.com.cn [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Key Lab of Heilongjiang Province for Infection and Immunity, Key Lab of Heilongjiang Province Education Bureau for Etiology, Harbin, Heilongjiang 150081 (China)

    2010-01-22

    To evaluate the interaction between HIV-1 envelope glycoprotein (Env) and target cell receptors, various cell-cell-fusion assays have been developed. In the present study, we established a novel fusion system. In this system, the expression of the sensitive reporter gene, firefly luciferase (FL) gene, in the target cells was used to evaluate cell fusion event. Simultaneously, constitutively expressed Renilla luciferase (RL) gene was used to monitor effector cell number and viability. FL gave a wider dynamic range than other known reporters and the introduction of RL made the assay accurate and reproducible. This system is especially beneficial for investigation of potential entry-influencing agents, for its power of ruling out the false inhibition or enhancement caused by the artificial cell-number variation. As a case study, we applied this fusion system to observe the effect of a serine protease, thrombin, on HIV Env-mediated cell-cell fusion and have found the fusion enhancement activity of thrombin over two R5-tropic HIV strains.

  5. Mechanism of the Anticoagulant Activity of Thrombin Mutant W215A/E217A

    Energy Technology Data Exchange (ETDEWEB)

    Gandhi, Prafull S.; Page, Michael J.; Chen, Zhiwei; Bush-Pelc, Leslie; Di Cera, Enrico; (WU-MED)

    2009-09-15

    The thrombin mutant W215A/E217A (WE) is a potent anticoagulant both in vitro and in vivo. Previous x-ray structural studies have shown that WE assumes a partially collapsed conformation that is similar to the inactive E* form, which explains its drastically reduced activity toward substrate. Whether this collapsed conformation is genuine, rather than the result of crystal packing or the mutation introduced in the critical 215-217 {beta}-strand, and whether binding of thrombomodulin to exosite I can allosterically shift the E* form to the active E form to restore activity toward protein C are issues of considerable mechanistic importance to improve the design of an anticoagulant thrombin mutant for therapeutic applications. Here we present four crystal structures of WE in the human and murine forms that confirm the collapsed conformation reported previously under different experimental conditions and crystal packing. We also present structures of human and murine WE bound to exosite I with a fragment of the platelet receptor PAR1, which is unable to shift WE to the E form. These structural findings, along with kinetic and calorimetry data, indicate that WE is strongly stabilized in the E* form and explain why binding of ligands to exosite I has only a modest effect on the E*-E equilibrium for this mutant. The E* {yields} E transition requires the combined binding of thrombomodulin and protein C and restores activity of the mutant WE in the anticoagulant pathway.

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

    Science.gov (United States)

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

    2014-09-11

    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 αIIbβ3 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

  7. The role of structural information in the discovery of direct thrombin and factor Xa inhibitors.

    Science.gov (United States)

    Nar, Herbert

    2012-05-01

    The quest for novel medications to treat thromboembolic disorders such as venous thrombosis, pulmonary embolism and stroke received a boost when the 3D structures of two major players in the blood coagulation cascade were determined in 1989 and 1993. Structure-guided design of inhibitors of thrombin (factor IIa, fIIa) and factor Xa (fXa) eventually led to the discovery of potent, selective, efficacious, orally active and safe compounds that proved successful in clinical studies. In 2008, the direct thrombin inhibitor dabigatran etexilate developed by Boehringer Ingelheim became the first novel antithrombotic molecular entity to enter the market in 50 years. Additional compounds targeting factor Xa were subsequently granted marketing authorization or are in late-stage clinical studies. In this review, I use selected case studies to describe the discovery of novel fIIa and fXa inhibitors, with a particular emphasis on the pre-eminent role that structural information played in this process. PMID:22503439

  8. Recombinant methods and materials

    Energy Technology Data Exchange (ETDEWEB)

    Roizman, B.; Post, L.E.

    1988-09-06

    This patent describes a method for stably effecting the insertion or deletion of a selected DNA sequence at a specific site in a viral genome. The method consists of: (1) isolating from the genome a linear DNA fragment comprising both (a) the specific site determined for insertion or deletion of selected DNA sequence and (b) flanking DNA sequences normally preceding and following the site; (2) preparing first and second altered genome fragments from the fragment isolated in step (1). (a) the first altered fragment comprising the fragment comprising a thymidine kinase gene in a position intermediate the ends of the fragment, and (b) the second altered fragment comprising the fragment having the selected DNA sequence inserted therein or deleted therefrom; (3) contacting the genome with the first altered fragment under conditions permitting recombination at sites of DNA sequence homology, selecting for a recombinant genome comprising the thymidine kinase gene, and isolating the recombinant genome; and (4) contacting the recombinant genome isolated in step (3) with the second altered fragment under conditions permitting recombination at sites of DNA sequence homology, selecting for a recombinant genome lacking the thymidine kinase gene, and isolating the recombinant genome product.

  9. Topics in Operator Theory

    CERN Document Server

    Ball, Joseph A; Helton, JWilliam; Rodman, Leiba; Spitkovsky, Iiya

    2010-01-01

    This is the first volume of a collection of original and review articles on recent advances and new directions in a multifaceted and interconnected area of mathematics and its applications. It encompasses many topics in theoretical developments in operator theory and its diverse applications in applied mathematics, physics, engineering, and other disciplines. The purpose is to bring in one volume many important original results of cutting edge research as well as authoritative review of recent achievements, challenges, and future directions in the area of operator theory and its applications.

  10. Topics in field theory

    CERN Document Server

    Karpilovsky, G

    1989-01-01

    This monograph gives a systematic account of certain important topics pertaining to field theory, including the central ideas, basic results and fundamental methods.Avoiding excessive technical detail, the book is intended for the student who has completed the equivalent of a standard first-year graduate algebra course. Thus it is assumed that the reader is familiar with basic ring-theoretic and group-theoretic concepts. A chapter on algebraic preliminaries is included, as well as a fairly large bibliography of works which are either directly relevant to the text or offer supplementary material of interest.

  11. Topical Acne Treatments and Pregnancy

    Science.gov (United States)

    Topical Acne Treatments and Pregnancy In every pregnancy, a woman starts out with a 3-5% chance of having ... This sheet talks about whether exposure to topical acne treatments may increase the risk for birth defects ...

  12. 蛇毒类凝血酶基因工程的研究进展%Research Progress on Cloning of Snake Venom Thrombin-like Enzymes

    Institute of Scientific and Technical Information of China (English)

    李家祺; 李谦; 唐松山; 李红枝

    2013-01-01

    蛇毒类凝血酶(Thrombin-like enzyme,TLE)是蛇毒中与血浆凝血酶性质相似的一类丝氨酸蛋白酶,所不同的是在蛇毒类凝血酶结构中已经没有纤维蛋白稳定因子激活组分.由于蛇毒类凝血酶在新药研究中扮演重要角色,比如来自Bothrops jararaca和Bothrops atrox蛇毒的立芷雪、来自东北白眉蝮蛇毒的邦停和来自尖吻蝮蛇毒的苏灵,它们是具有止血作用的新药;来自Gloydius shedaoensis和Gloydius ussuriensis蛇毒的东菱克栓酶、来自白眉蝮蛇和尖吻蝮蛇毒的降纤酶、来自Calloselasma rhodostoma蛇毒的安克洛酶和来自Crotalus adamanteus蛇毒的Crotalase,它们是具有溶栓作用的新药.这些药用蛋白质的来源和产量因有限的蛇毒原料而有很大限制,通过基因克隆可解决资源问题.文章综述了蛇毒类凝血酶的基因结构、糖基化特点和各种重组表达体系,为大量制备供临床和基础研究使用奠定基础.%Snake venom thrombin-like enzymes (snTLE) are one kind of serine proteases similar to thrombin from blood, but the basic difference between them is that snTLE has no XIII-activated domain to lead thrombosis in vivo or in vitro. Because snake venoms play very important role in pharmaceutics,for example Reptilase from Bothrops jararaca or Bothrops atrox snake venom, Bangtin from Agkistrodon halys snake venom,or Siding from Agkistrodon acutus snake venom,are hemostatic drugs. Defibrase from Gloydius shedaoensis ,Cloydius ussuriensis,Agkistrodon halys,or Agkistrodon acutus snake venom,Ancrod from Calloselasma rhodostoma snake venom, or Crotalase from Crotalus adamanteus snake venom, is thrombolytic drug. So the shortage of snake venom source can be overcome by using gene recombination method. The gene structures, protein glycosylation, and all kinds of cloning expression systems for snTLE protein are summarized in the paper,which will provide a basis for large-scale production of some snTLE proteins.

  13. Expression of Recombinant Antibodies

    OpenAIRE

    Frenzel, André; Hust, Michael; Schirrmann, Thomas

    2013-01-01

    Recombinant antibodies are highly specific detection probes in research, diagnostics, and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transg...

  14. INVOLVEMENT OF BACTERICIDAL FACTORS FROM THROMBIN-STIMULATED PLATELETS IN CLEARANCE OF ADHERENT VIRIDANS STREPTOCOCCI IN EXPERIMENTAL INFECTIVE ENDOCARDITIS

    NARCIS (Netherlands)

    VANDERWERFF, J; ZAAT, SAJ; JOLDERSMA, W; HESS, J

    1995-01-01

    Platelets activated with thrombin release bactericidal factors. We studied the role of the susceptibility of viridans streptococci to these bactericidal factors in the development of infective endocarditis (IE). By using the experimental endocarditis rabbit model, the initial adherence and the devel

  15. In vitro effect of hemodilution on activated clotting time and high-dose thrombin time during cardiopulmonary bypass

    NARCIS (Netherlands)

    Huyzen, RJ; vanOeveren, W; Wei, FY; Stellingwerf, P; Boonstra, PW; Gu, YJ

    1996-01-01

    Background. Extreme dilution of clotting factors, as may occur during pediatric or neonatal cardiopulmonary bypass, often leads to inadequate monitoring of anticoagulation with activated dotting time (ACT). In this study we postulate that the high-dose thrombin time (HiTT) is less influenced by extr

  16. Binding characteristics of thrombin-activatable fibrinolysis inhibitor to streptococcal surface collagen-like proteins A and B

    NARCIS (Netherlands)

    Seron, Mercedes Valls; Plug, Tom; Marquart, J. Arnoud; Marx, Pauline F.; Herwald, Heiko; de Groot, Philip G.; Meijers, Joost C. M.

    2011-01-01

    Streptococcus pyogenes is the causative agent in a wide range of diseases in humans. Thrombin-activatable fibrinolysis inhibitor (TAFI) binds to collagen-like proteins ScIA and ScIB at the surface of S. pyogenes. Activation of TAFI at this surface redirects inflammation from a transient to chronic s

  17. Binding characteristics of thrombin-activatable fibrinolysis inhibitor to streptococcal surface collagen-like proteins A and B

    NARCIS (Netherlands)

    M.V. Serón; T. Plug; J.A. Marquart; P.F. Marx; H. Herwald; P.G. de Groot; J.C.M. Meijers

    2011-01-01

    Streptococcus pyogenes is the causative agent in a wide range of diseases in humans. Thrombin-activatable fibrinolysis inhibitor (TAFI) binds to collagen-like proteins SclA and SclB at the surface of S. pyogenes. Activation of TAFI at this surface redirects inflammation from a transient to chronic s

  18. Mechanism of poly(acrylic acid) acceleration of antithrombin inhibition of thrombin: implications for the design of novel heparin mimics.

    Science.gov (United States)

    Monien, Bernhard H; Cheang, Kai I; Desai, Umesh R

    2005-08-11

    The bridging mechanism of antithrombin inhibition of thrombin is a dominant mechanism contributing a massive approximately 2500-fold acceleration in the reaction rate and is also a key reason for the clinical usage of heparin. Our recent study of the antithrombin-activating properties of a carboxylic acid-based polymer, poly(acrylic acid) (PAA), demonstrated a surprisingly high acceleration in thrombin inhibition (Monien, B. H.; Desai, U. R. J. Med. Chem. 2005, 48, 1269). To better understand this interesting phenomenon, we have studied the mechanism of PAA-dependent acceleration in antithrombin inhibition of thrombin. Competitive binding studies with low-affinity heparin and a heparin tetrasaccharide suggest that PAA binds antithrombin in both the pentasaccharide- and the extended heparin-binding sites, and these results are corroborated by molecular modeling. The salt-dependence of the K(D) of the PAA-antithrombin interaction shows the formation of five ionic interactions. In contrast, the contribution of nonionic forces is miniscule, resulting in an interaction that is significantly weaker than that observed for heparins. A bell-shaped profile of the observed rate constant for antithrombin inhibition of thrombin as a function of PAA concentration was observed, suggesting that inhibition proceeds through the "bridging" mechanism. The knowledge gained in this mechanistic study highlights important rules for the rational design of orally available heparin mimics. PMID:16078853

  19. The Expression of the Thrombin Receptors PAR-3 and PAR-4 is Downregulated in Pancreatic Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Claudia Rudroff

    2015-05-01

    Full Text Available Background: Patients with pancreatic cancer frequently suffer from thrombosis as a consequence of excess thrombin generation. In addition to its role in the plasmatic coagulation cascade, thrombin induces numerous cellular effects by activating a unique group of G-protein-coupled receptors on the cell membrane, the proteinase-activated receptors (PARs. At present, PAR-1, PAR-3 and PAR-4 are known to be activated by thrombin. We previously demonstrated a putative role for PAR-1 in pancreatic cancer progression, but little is known about the physiological and pathophysiological roles of PAR-3 and PAR-4. In the present study, we examined the expression patterns of PAR-3 and PAR-4 in pancreatic tissue and pancreatic cancer cells. Methods: Tissue samples from three patients with pancreatic adenocarcinoma and six human pancreatic carcinoma cell lines were examined. Gene expression was analysed by RT-PCR and quantified by HPLC. Protein expression was determined by Western blot analysis. Data analysis was performed using ANOVA in SPSS. Results and Conclusion: In contrast to PAR-1, both PAR-3 and PAR-4 were expressed in healthy pancreases but downregulated in pancreatic cancer. The contrasting expression patterns of PAR-3 and PAR-4 compared with PAR-1 indicate that the mechanism that regulates the cellular effects of thrombin on tumor progression remains to be fully elucidated.

  20. Topics in atomic physics

    CERN Document Server

    Burkhardt, Charles E

    2006-01-01

    The study of atomic physics propelled us into the quantum age in the early twentieth century and carried us into the twenty-first century with a wealth of new and, in some cases, unexplained phenomena. Topics in Atomic Physics provides a foundation for students to begin research in modern atomic physics. It can also serve as a reference because it contains material that is not easily located in other sources. A distinguishing feature is the thorough exposition of the quantum mechanical hydrogen atom using both the traditional formulation and an alternative treatment not usually found in textbooks. The alternative treatment exploits the preeminent nature of the pure Coulomb potential and places the Lenz vector operator on an equal footing with other operators corresponding to classically conserved quantities. A number of difficult to find proofs and derivations are included as is development of operator formalism that permits facile solution of the Stark effect in hydrogen. Discussion of the classical hydrogen...

  1. Rapid Upregulation of Orai1 Abundance in the Plasma Membrane of Platelets Following Activation with Thrombin and Collagen Related Peptide

    Directory of Open Access Journals (Sweden)

    Guilai Liu

    2015-11-01

    Full Text Available Background: Blood platelets accomplish primary hemostasis following vascular injury and contribute to the orchestration of occlusive vascular disease. Platelets are activated by an increase of cytosolic Ca2+-activity ([Ca2+]i, which is accomplished by Ca2+-release from intracellular stores and subsequent store operated Ca2+ entry (SOCE through Ca2+ release activated Ca2+ channel moiety Orai1. Powerful activators of platelets include thrombin and collagen related peptide (CRP, which are in part effective by activation of small G- protein Rac1. The present study explored the influence of thrombin and CRP on Orai1 protein abundance and cytosolic Ca2+-activity ([Ca2+]i in platelets drawn from wild type mice. Methods: Orai1 protein surface abundance was quantified utilizing CF™488A conjugated antibodies, and [Ca2+]i was determined with Fluo3-fluorescence. Results: In resting platelets, Orai1 protein abundance and [Ca2+]i were low. Thrombin (0.02 U/ml and CRP (5ug/ml within 2 min increased [Ca2+]i and Orai1 protein abundance at the platelet surface. [Ca2+]i was further increased by Ca2+ ionophore ionomycin (1 µM and by store depletion with the sarcoendoplasmatic Ca2+ ATPase inhibitor thapsigargin (1 µM. However, Orai1 protein abundance at the platelet surface was not significantly affected by ionomycin and only slightly increased by thapsigargin. The effect of thrombin and CRP on Orai1 abundance and [Ca2+]i was significantly blunted by Rac1 inhibitor NSC23766 (50 µM. Conclusion: The increase of [Ca2+]i following stimulation of platelets with thrombin and collagen related peptide is potentiated by ultrarapid Rac1 sensitive translocation of Orai1 into the cell membrane.

  2. Antithrombotic effects of LB30870, a potent, orally active, selective and direct thrombin inhibitor, and pharmacokinetics of its prodrug.

    Science.gov (United States)

    Park, Hee Dong; Lee, Sung-Hack; Kim, Tae Hoon; Lee, Sun Hwa; Cho, Kwan Hyung; Kim, Aeri

    2013-09-01

    Antithrombotic activity and bleeding complication of a new potent, selective, and direct thrombin inhibitor, LB30870, were evaluated in comparison with other anticoagulants. In order to improve oral absorption of LB30870, pharmacokinetics of LB30889, which is a double prodrug with blocking groups in both amidine and carboxyl groups, was studied in rats and dogs. LB30870 was more potent than melagatran or argatroban with thrombin inhibition constants of 0.02, 1.3 and 4.5 nM, respectively. All three direct thrombin inhibitors were selective towards other serine proteases with selectivity ratio greater than 1000, except for trypsin. Thrombin binding kinetics of LB30870 showed rapid association and slow dissociation rate constants, demonstrating its potential as anticoagulant. LB30870 was more effective than melagatran or argatroban in plasma clot-bound thrombin inhibition. In the rat venous stasis model of the caval vein, LB30870 reduced wet clot weights in a dose dependent manner after the intravenous bolus with infusion administration. The ED50 of LB30870, melagatran and enoxaparin were 50 μg/kg+2 μg/kg/min, 35 μg/kg+1.4 μg/kg/min and 200 μg/kg+8.3 μg/kg/min, respectively. No significant bleeding problem was observed with LB30870 at the dose up to two times ED80 in rats. LB30889, a double prodrug of LB30870, showed species difference in pharmacokinetics. Its oral bioavailability in rats or dogs was not better than that of LB30870. In conclusion, LB30870 has the potential to be useful as an effective oral anticoagulant for the prevention and treatment of thromboembolic diseases. PMID:23899618

  3. Design, synthesis and antithrombotic evaluation of novel dabigatran etexilate analogs, a new series of non-peptides thrombin inhibitors.

    Science.gov (United States)

    Chen, Dongxing; Wang, Shaochi; Diao, Xiaojuan; Zhu, Qihua; Shen, Huiliang; Han, Xueqing; Wang, Yiwei; Gong, Guoqing; Xu, Yungen

    2015-12-01

    Thrombin is a serine protease that plays a key role in blood clotting, which makes it a promising target for the treatment of thrombotic diseases. Dabigatran is direct potent thrombin inhibitor. Based on bioisosteric and scaffold hopping principle, two dabigatran mimics (I-1 and II-1) in which the benzamidine moiety of dabigatran was replaced by a tricyclic fused scaffold were designed, synthesized and evaluated for their in vitro activities for inhibiting thrombin. The results reveal that compounds I-1 (IC50=9.20nM) and II-1 (IC50=7.48nM) are potent direct thrombin inhibitors and the activity is in the range of reference drug. On this basis, twenty-two ester and carbamate derivatives of I-1 or II-1 were prepared and evaluated for their anticoagulant activity. Prodrugs I-4a (IC50=0.73μM), I-4b (IC50=0.75μM), II-2a (IC50=1.44μM) and II-2b (IC50=0.91μM) display excellent effects of inhibiting thrombin induced-platelet aggregation. Moreover, compounds I-9 and II-4, which contain a cleavable moiety with anti-platelet activity, show the best anticoagulant efficacy among the tested compounds in the rat venous thrombosis model. The compounds which have better in vitro and in vivo activity were subjected to rat tail bleeding test, and the result demonstrates that compound I-9 is less likely to have bleeding risk than dabigatran etexilate.

  4. Thrombin-activatable fibrinolysis inhibitor activity in healthy and diseased dogs

    DEFF Research Database (Denmark)

    Jessen, Lisbeth Rem; Wiinberg, Bo; Kjelgaard-Hansen, Mads;

    2010-01-01

    Background: In people, increased thrombin-activatable fibrinolysis inhibitor (TAFI) antigen has been associated with increased risk of thrombosis, and decreased TAFI may contribute to bleeding diathesis. TAFI activity in dogs has been described in experimental models, but not in dogs...... with spontaneous disease. Objective: The aim of this study was to compare TAFI activity in healthy dogs with TAFI activity in dogs with spontaneous disease. Methods: Plasma samples from 20 clinically healthy Beagles and from 35 dogs with various diseases were analyzed using a commercial chromogenic assay...... that measured TAFI activity relative to activity in standardized pooled human plasma. Results: Median TAFI activity for the 20 Beagles was 46.1% (range 32.2-70.8%) compared with 62.6% (29.1-250%) for the 35 diseased dogs, and 14/35 (40%) had TAFI activities >the upper limit for controls. The highest individual...

  5. Dynamics Govern Specificity of a Protein-Protein Interface: Substrate Recognition by Thrombin.

    Directory of Open Access Journals (Sweden)

    Julian E Fuchs

    Full Text Available Biomolecular recognition is crucial in cellular signal transduction. Signaling is mediated through molecular interactions at protein-protein interfaces. Still, specificity and promiscuity of protein-protein interfaces cannot be explained using simplistic static binding models. Our study rationalizes specificity of the prototypic protein-protein interface between thrombin and its peptide substrates relying solely on binding site dynamics derived from molecular dynamics simulations. We find conformational selection and thus dynamic contributions to be a key player in biomolecular recognition. Arising entropic contributions complement chemical intuition primarily reflecting enthalpic interaction patterns. The paradigm "dynamics govern specificity" might provide direct guidance for the identification of specific anchor points in biomolecular recognition processes and structure-based drug design.

  6. Thrombin inhibition with melagatran does not attenuate renal ischaemia-reperfusion injury in rats

    DEFF Research Database (Denmark)

    Nitescu, Nicoletta; Grimberg, Elisabeth; Ricksten, Sven-Erik;

    2007-01-01

    BACKGROUND: Renal ischaemia-reperfusion (IR) is associated with activation of the coagulation system and inflammation within the kidney. The aim of the present study was to examine the effects of selective thrombin inhibition with melagatran on kidney morphology and function in rats subjected...... of saline vehicle or melagatran [0.5 mumol/kg, subcutaneously (s.c.)] followed by a continuous infusion throughout (0.08 micromol/kg/h, s.c.). Forty-eight hours after IR, renal function was assessed in anaesthetized animals and kidney histology was analysed semi-quantitatively. RESULTS: Rats in group IR......-Saline showed an approximate 85% reduction in glomerular filtration rate, 5-fold increases in fractional urinary excretion rates of sodium, potassium and water, and marked renal histological abnormalities, compared with sham (P histopathological changes in the cortex and outer medulla were...

  7. A study of the conditions and accuracy of the thrombin time assay of plasma fibrinogen

    DEFF Research Database (Denmark)

    Jespersen, J; Sidelmann, Johannes Jakobsen

    1982-01-01

    The conditions, accuracy, precision and possible error of the thrombin time assay of plasma fibrinogen are determined. Comparison with an estimation of clottable protein by absorbance at 280 nm gave a correlation coefficient of 0.96 and the regression line y = 1.00 x + 0.56 (n = 34). Comparison...... with a radial immunodiffusion method yielded the correlation coefficient 0.97 and the regression line y = 1.18 x = 2.47 (n = 26). The presence of heparin in clinically applied concentrations produced a slight shortening of the clotting times. The resulting error in the estimated concentrations of fibrinogen...... was too small to affect the clinical usefulness of the determinations. The influence of fibrin(ogen) degradation products was significant only in excessive amounts in samples containing low levels of fibrinogen....

  8. Dynamics Govern Specificity of a Protein-Protein Interface: Substrate Recognition by Thrombin.

    Science.gov (United States)

    Fuchs, Julian E; Huber, Roland G; Waldner, Birgit J; Kahler, Ursula; von Grafenstein, Susanne; Kramer, Christian; Liedl, Klaus R

    2015-01-01

    Biomolecular recognition is crucial in cellular signal transduction. Signaling is mediated through molecular interactions at protein-protein interfaces. Still, specificity and promiscuity of protein-protein interfaces cannot be explained using simplistic static binding models. Our study rationalizes specificity of the prototypic protein-protein interface between thrombin and its peptide substrates relying solely on binding site dynamics derived from molecular dynamics simulations. We find conformational selection and thus dynamic contributions to be a key player in biomolecular recognition. Arising entropic contributions complement chemical intuition primarily reflecting enthalpic interaction patterns. The paradigm "dynamics govern specificity" might provide direct guidance for the identification of specific anchor points in biomolecular recognition processes and structure-based drug design. PMID:26496636

  9. Historical perspective and contemporary management of acute coronary syndromes: from MONA to THROMBINS2.

    Science.gov (United States)

    Kline, Kristopher P; Conti, C Richard; Winchester, David E

    2015-01-01

    Acute coronary syndrome (ACS) remains a major burden on morbidity and mortality in the United States. Medical professionals and students often use the mnemonic 'MONA' (morphine, oxygen, nitroglycerin and aspirin) to recall treatments for ACS; however, this list of therapies is outdated. We provide a historical perspective on 'MONA,' attempt to uncover its origin in the medical literature, and demonstrate the myriad changes that have occurred over the last 50 years of ACS management. We have developed a novel mnemonic, 'THROMBINS2' (thienopyridines, heparin/enoxaparin, renin-angiotensin system blockers, oxygen, morphine, beta blocker, intervention, nitroglycerin, statin/salicylate) to help bedside clinicians recall all the elements of contemporary ACS management. We demonstrate the mortality benefit for each component of contemporary ACS management, correlating the continued improvement with historical data on mortality after myocardial infarction. We encourage providers to utilize this mnemonic to explore options and guide treatments in ACS patients. PMID:26457728

  10. Historical perspective and contemporary management of acute coronary syndromes: from MONA to THROMBINS2.

    Science.gov (United States)

    Kline, Kristopher P; Conti, C Richard; Winchester, David E

    2015-01-01

    Acute coronary syndrome (ACS) remains a major burden on morbidity and mortality in the United States. Medical professionals and students often use the mnemonic 'MONA' (morphine, oxygen, nitroglycerin and aspirin) to recall treatments for ACS; however, this list of therapies is outdated. We provide a historical perspective on 'MONA,' attempt to uncover its origin in the medical literature, and demonstrate the myriad changes that have occurred over the last 50 years of ACS management. We have developed a novel mnemonic, 'THROMBINS2' (thienopyridines, heparin/enoxaparin, renin-angiotensin system blockers, oxygen, morphine, beta blocker, intervention, nitroglycerin, statin/salicylate) to help bedside clinicians recall all the elements of contemporary ACS management. We demonstrate the mortality benefit for each component of contemporary ACS management, correlating the continued improvement with historical data on mortality after myocardial infarction. We encourage providers to utilize this mnemonic to explore options and guide treatments in ACS patients.

  11. 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)

    Michael B. Jarstfer

    2008-03-01

    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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Fang Lanyun [State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, Jilin (China); Graduate School of the Chinese Academy of Sciences, Beijing 100049 (China); Ningbo Municipal Center for Disease Control and Prevention, Ningbo 315010, Zhejiang (China); Lue Zhaozi [State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, Jilin (China); Wei Hui [State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, Jilin (China); Graduate School of the Chinese Academy of Sciences, Beijing 100049 (China); Wang Erkang [State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022, Jilin (China)], E-mail: ekwang@ciac.jl.cn

    2008-10-17

    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 {beta}- and {gamma}-thrombins had negligible response, which indicated a high specificity of {alpha}-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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Oberdisse, E.; Lapetina, E.G.

    1987-05-14

    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 (/sup 3/H)inositol. Ca/sup 2 +/ has opposite effects on the formation of (/sup 3/H)inositol phosphates induced by thrombin or GTP gamma S. While the action of GTP gamma S on the formation of (/sup 3/H)inositol phosphates is inhibited by Ca/sup 2 +/, action of thrombin is stimulated by Ca/sup 2 +/. 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. Ca/sup 2 +/ increases the inhibitory effect of GDP beta S on GTP gamma S activation of phospholipase C, but Ca/sup 2 +/ 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.

  14. 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)

    Xiong, Ya; Deng, Chunhui; Zhang, Xiangmin

    2014-11-01

    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 μL(-1) (2.36 nM) thrombin could be detected. This proposed method has a relatively wide linear relation ranging from 0.1 ng μL(-1) to 10 ng μL(-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

  15. Ultrasensitive electrochemical aptasensor for the detection of thrombin based on dual signal amplification strategy of Au@GS and DNA-CoPd NPs conjugates.

    Science.gov (United States)

    Wang, Yaoguang; Zhang, Yong; Yan, Tao; Fan, Dawei; Du, Bin; Ma, Hongmin; Wei, Qin

    2016-06-15

    In this work, an ultrasensitive electrochemical aptasensor for the detection of thrombin was developed based on Au nanoparticles decorated graphene sheet (Au@GS) and CoPd binary nanoparticles (CoPd NPs). A sulfydryl-labeled thrombin capture probe (Apt1) and a biotin-labeled thrombin reporter probe (Apt2) were designed to achieve a sandwich-type strategy. Au@GS was used as a sensing platform for the facile immobilization of Apt1 through Au-S bond, forming a sensing interface for thrombin. The specific recognition of thrombin induced the attachment of Apt2-CoPd NPs to the electrode. The labeled CoPd NPs showed good catalytic properties toward the reduction of H2O2, resulting in an amperometric signal. The amperometric response was correlated to the thrombin concentration in sample solutions. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) confirmed the successful fabrication of the aptasensor. A linear response to thrombin in the range of 0.01-2.00 ng mL(-1) with a low detection limit (5 pg mL(-1)) was achieved. The proposed aptasensor showed good selectivity, good reproducibility and acceptable stability. This proposed strategy may find many potential applications in the detection of other biomolecules.

  16. 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 β- and γ-thrombins had negligible response, which indicated a high specificity of α-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

  17. Characterization of Ixophilin, a thrombin inhibitor from the gut of Ixodes scapularis.

    Directory of Open Access Journals (Sweden)

    Sukanya Narasimhan

    Full Text Available Ixodes scapularis, the black-legged tick, vectors several human pathogens including Borrelia burgdorferi, the agent of Lyme disease in North America. Pathogen transmission to the vertebrate host occurs when infected ticks feed on the mammalian host to obtain a blood meal. Efforts to understand how the tick confronts host hemostatic mechanisms and imbibes a fluid blood meal have largely focused on the anticoagulation strategies of tick saliva. The blood meal that enters the tick gut remains in a fluid state for several days during the process of feeding, and the role of the tick gut in maintaining the blood-meal fluid is not understood. We now demonstrate that the tick gut produces a potent inhibitor of thrombin, a key enzyme in the mammalian coagulation cascade. Chromatographic fractionation of engorged tick gut proteins identified one predominant thrombin inhibitory activity associated with an approximately 18 kDa protein, henceforth referred to as Ixophilin. The ixophilin gene was preferentially transcribed in the guts of feeding nymphs. Expression began after 24 hours of feeding, coincident with the flow of host blood into the tick gut. Immunity against Ixophilin delayed tick feeding, and decreased feeding efficiency significantly. Surprisingly, immunity against Ixophilin resulted in increased Borrelia burgdorferi transmission to the host, possibly due to delayed feeding and increased transmission opportunity. These observations illuminate the potential drawbacks of targeting individual tick proteins in a functional suite. They also underscore the need to identify the "anticoagulome" of the tick gut, and to prioritize a critical subset of anticoagulants that could be targeted to efficiently thwart tick feeding, and block pathogen transmission to the vertebrate host.

  18. Management of major bleedings during anticoagulant treatment with the oral direct thrombin inhibitor ximelagatran or warfarin.

    Science.gov (United States)

    Fernlöf, Gunilla; Sjöström, Britta M; Lindell, Klas M; Wall, Ulrika E

    2009-12-01

    Several new oral anticoagulants are currently investigated in phase III programmes, mainly with inhibition of factor Xa or thrombin as their pharmacological target. Advantages are expected with these new drugs compared with vitamin K antagonists, but one potential drawback is the lack of specific antidotes. During the clinical studies with ximelagatran, an oral direct thrombin inhibitor withdrawn due to hepatic side effects, investigators were instructed to manage bleedings with routine measures. We have retrospectively tried to assess whether this was sufficient or whether there was a need for reversal strategies. The study population consisted of patients with major bleedings in three long-term studies (104 ximelagatran, 155 warfarin). All individual patient narratives were reviewed with respect to management of the bleeding. Complementary data were retrieved from the data-based case report forms. Approximately, two of three of the patients in both groups were subject to some kind of treatment. One-third (1/3) in both groups had transfusions documented and/or received specific medication. Vitamin K was given more often to warfarin patients. Two ximelagatran patients received prothrombin complex (four-factor concentrate), but one was a patient with a severe hepatopathy suspected to be drug-induced. Overall, the case descriptions did not reveal any apparent differences in the course of events between groups. We found no indications that the lack of an antidote posed a clinical problem in patients treated with ximelagatran as compared with warfarin. The relatively short half-life of melagatran, the active metabolite of ximelagatran, may have contributed to these results.

  19. Thrombin induced connective tissue growth factor expression in rat vascular smooth muscle cells via the PAR-1/JNK/AP-1 pathway

    Institute of Scientific and Technical Information of China (English)

    Wen-chin KO; Bing-chang CHEN; Ming-jen HSU; Chia-ti TSAI; Chuang-ye HONG; Chien-huang LIN

    2012-01-01

    To investigate the signaling pathways involved in thrombin-induced connective tissue growth factor (CTGF) expression in rat vascular smooth muscle cells (VSMCs).Methods:Experiments were preformed on primary rat aortic smooth muscle cells (RASMCs) and a rat VSMC line (A10).CTGF protein levels were measured using Western blotting.Luciferase reporter genes and dominant negative mutants (DNs) were used to investigate the signaling pathways mediating the induction of CTGF expression by thrombin.Results:Thrombin (0.3-3.0 U/mL) caused a concentration- and time-dependent increase in CTGF expression in both RASMCs and A10 cells.Pretreating A10 cells with the protease-activated receptor 1 (PAR-1) antagonist SCH79797 (0.1 μmol/L) significantly blocked thrombin-induced CTGF expression,while the PAR-4 antagonist tcY-NH2 (30 μmol/L) had no effect.The PAR-1 agonist SFLLRN-NH2 (300 μmol/L) induced CTGF expression,while the PAR-4 agonist GYPGQV-NH2 (300 μmol/L) had no effect.Thrombin (1 U/mL) caused time-dependent phosphorylation of c-Jun N-terminal kinase (JNK).Pretreating with the JNK inhibitor SP600125 (3-30 μmol/L) or transfection with DNs of JNK1/2 significantly attenuated thrombin-induced CTGF expression.Thrombin (0.3-3.0 U/mL) increased activator protein-1 (AP-1)-Iuciferase activity,which was inhibited by the JNK inhibitor SP600125.The AP-1 inhibitor curcumin (1-10 μmol/L) concentration-dependently attenuated thrombin-induced CTGF expression.Conclusion:Thrombin acts on PAR-1 to activate the JNK signaling pathway,which in turn initiates AP-1 activation and ultimately induces CTGF expression in VSMCs.

  20. Regulation of Meiotic Recombination

    Energy Technology Data Exchange (ETDEWEB)

    Gregory p. Copenhaver

    2011-11-09

    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

  1. Thrombin-anti-thrombin levels and patency of arterio-venous fistula in patients undergoing haemodialysis compared to healthy volunteers: a prospective analysis.

    Directory of Open Access Journals (Sweden)

    James A Milburn

    Full Text Available BACKGROUND: Patients on haemodialysis (HD are at an increased risk of sustaining thrombotic events especially to their vascular access which is essential for maintenance of HD. OBJECTIVES: To assess whether 1 markers of coagulation, fibrinolysis or endothelial activation are increased in patients on HD compared to controls and 2 if measurement of any of these factors could help to identify patients at increased risk of arteriovenous (AVF access occlusion. PATIENTS/METHODS: Venous blood samples were taken from 70 patients immediately before a session of HD and from 78 resting healthy volunteers. Thrombin-antithrombin (TAT, D-dimer, von Willebrand factor (vWF, plasminogen activator inhibitor-1 antigen (PAI-1 and soluble p-selectin were measured by ELISA. C-reactive protein (hsCRP was measured by an immunonephelometric kinetic assay. Determination of the patency of the AVF was based upon international standards and was prospectively followed up for a minimum of four years or until the AVF was non-functioning. RESULTS: A total of 70 patients were studied with a median follow-up of 740 days (range 72-1788 days. TAT, D-dimer, vWF, p-selectin and hsCRP were elevated in patients on HD compared with controls. At one year follow-up, primary patency was 66% (46 patients. In multivariate analysis TAT was inversely associated with primary assisted patency (r = -0.250, p = 0.044 and secondary patency (r = -0.267, p= 0.031. CONCLUSIONS: The novel finding of this study is that in patients on haemodialysis, TAT levels were increased and inversely correlated with primary assisted patency and secondary patency. Further evaluation is required into the possible role of TAT as a biomarker of AVF occlusion.

  2. Advanced verification topics

    CERN Document Server

    Bhattacharya, Bishnupriya; Hall, Gary; Heaton, Nick; Kashai, Yaron; Khan Neyaz; Kirshenbaum, Zeev; Shneydor, Efrat

    2011-01-01

    The Accellera Universal Verification Methodology (UVM) standard is architected to scale, but verification is growing and in more than just the digital design dimension. It is growing in the SoC dimension to include low-power and mixed-signal and the system integration dimension to include multi-language support and acceleration. These items and others all contribute to the quality of the SOC so the Metric-Driven Verification (MDV) methodology is needed to unify it all into a coherent verification plan. This book is for verification engineers and managers familiar with the UVM and the benefits it brings to digital verification but who also need to tackle specialized tasks. It is also written for the SoC project manager that is tasked with building an efficient worldwide team. While the task continues to become more complex, Advanced Verification Topics describes methodologies outside of the Accellera UVM standard, but that build on it, to provide a way for SoC teams to stay productive and profitable.

  3. Superconcentration and related topics

    CERN Document Server

    Chatterjee, Sourav

    2014-01-01

    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...

  4. Researching Distressing Topics

    Directory of Open Access Journals (Sweden)

    Sharon Jackson

    2013-05-01

    Full Text Available Qualitative researchers who explore sensitive topics may expose themselves to emotional distress. Consequently, researchers are often faced with the challenge of maintaining emotional equilibrium during the research process. However, discussion on the management of difficult emotions has occupied a peripheral place within accounts of research practice. With rare exceptions, the focus of published accounts is concentrated on the analysis of the emotional phenomena that emerge during the collection of primary research data. Hence, there is a comparative absence of a dialogue around the emotional dimensions of working with secondary data sources. This article highlights some of the complex ways in which emotions enter the research process during secondary analysis, and the ways in which we engaged with and managed emotional states such as anger, sadness, and horror. The concepts of emotional labor and emotional reflexivity are used to consider the ways in which we “worked with” and “worked on” emotion. In doing so, we draw on our collective experiences of working on two collaborative projects with ChildLine Scotland in which a secondary analysis was conducted on children’s narratives of distress, worry, abuse, and neglect.

  5. 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

  6. Detection of Embryonic Research Topics by Analysing Semantic Topic Networks

    OpenAIRE

    Salatino, Angelo Antonio; Motta, Enrico

    2016-01-01

    Being aware of new research topics is an important asset for anybody involved in the research environment, including researchers, academic publishers and institutional funding bodies. In recent years, the amount of scholarly data available on the web has increased steadily, allowing the development of several approaches for detecting emerging research topics and assessing their trends. However, current methods focus on the detection of topics which are already associated with a label or a sub...

  7. Thrombin induces Egr-1 expression in fibroblasts involving elevation of the intracellular Ca2+ concentration, phosphorylation of ERK and activation of ternary complex factor

    Directory of Open Access Journals (Sweden)

    Thiel Gerald

    2009-05-01

    Full Text Available Abstract Background The serine protease thrombin catalyzes fibrin clot formation by converting fibrinogen into fibrin. Additionally, thrombin stimulation leads to an activation of stimulus-responsive transcription factors in different cell types, indicating that the gene expression pattern is changed in thrombin-stimulated cells. The objective of this study was to analyze the signaling cascade leading to the expression of the zinc finger transcription factor Egr-1 in thrombin-stimulated lung fibroblasts. Results Stimulation of 39M1-81 fibroblasts with thrombin induced a robust and transient biosynthesis of Egr-1. Reporter gene analysis revealed that the newly synthesized Egr-1 was biologically active. The signaling cascade connecting thrombin stimulation with Egr-1 gene expression required elevated levels of cytosolic Ca2+, the activation of diacylgycerol-dependent protein kinase C isoenzymes, and the activation of extracellular signal-regulated protein kinase (ERK. Stimulation of the cells with thrombin triggered the phosphorylation of the transcription factor Elk-1. Expression of a dominant-negative mutant of Elk-1 completely prevented Egr-1 expression in stimulated 39M1-81 cells, indicating that Elk-1 or related ternary complex factors connect the intracellular signaling cascade elicited by activation of protease-activated receptors with transcription of the Egr-1 gene. Lentiviral-mediated expression of MAP kinase phosphatase-1, a dual-specific phosphatase that dephosphorylates and inactivates ERK in the nucleus, prevented Elk-1 phosphorylation and Egr-1 biosynthesis in thrombin stimulated 39M1-81 cells, confirming the importance of nuclear ERK and Elk-1 for the upregulation of Egr-1 expression in thrombin-stimulated lung fibroblasts. 39M1-81 cells additionally express M1 muscarinic acetylcholine receptors. A comparison between the signaling cascades induced by thrombin or carbachol showed no differences, except that signal transduction via M

  8. Discovering health topics in social media using topic models.

    Directory of Open Access Journals (Sweden)

    Michael J Paul

    Full Text Available 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.

  9. Recombinant Helicobacter pylori catalase

    Institute of Scientific and Technical Information of China (English)

    Yang Bai; Ya-Li Zhang; Jian-Feng Jin; Ji-De Wang; Zhao-Shan Zhang

    2003-01-01

    AIM: To construct a recombinant strain which highly expresses catalase of Helicobacter pylori(H.pylori) and assay the activity of H. pylori catalase.METHODS: The catalase DNA was amplified from H. pylori chromosomal DNA with PCR techniques and inserted into the prokaryotie expression vector pET-22b (+), and then was transformed into the BL21 (DE3) E. coli strain which expressed catalase recombinant protein. The activity of H.pylori catalase was assayed by the Beers & Sizers.RESULTS: DNA sequence analysis showed that the sequence of catalase DNA was the same as GenBank's research. The catalase recombinant protein amounted to 24.4 % of the total bacterial protein after induced with IPTG for 3 hours at 37 ℃ and the activity of H. pylori catalase was high in the BL21 (DE3) E. coli strain.CONCLUSION: A clone expressing high activity H. pylori catalase is obtained, laying a good foundation for further studies.

  10. Recombination in ionized gases

    International Nuclear Information System (INIS)

    In this paper it is shown how capture-stabilized methodology (both macroscopic and microscopic) can provide a generic basis for a unified treatment of all of the above recombination mechanisms. A new semiclassical theory of dissociative recombination is also presented in an effort to gain further insight into the physics not included in the first-order treatment and difficult to extract from numerical quantal treatments based on configuration mixing and on multichannel quantum defect theory. A simple analytical expression more accurate than the standard first-order result is obtained for the cross section σ and rate coefficient α. (author)

  11. Topic Modeling of Hierarchical Corpora /

    OpenAIRE

    Kim, Do-kyum

    2014-01-01

    The sizes of modern digital libraries have grown beyond our capacity to comprehend manually. Thus we need new tools to help us in organizing and browsing large corpora of text that do not require manually examining each document. To this end, machine learning researchers have developed topic models, statistical learning algorithms for automatic comprehension of large collections of text. Topic models provide both global and local views of a corpus; they discover topics that run through the co...

  12. Identification of human liver cytochrome P450 enzymes involved in the metabolism of SCH 530348 (Vorapaxar), a potent oral thrombin protease-activated receptor 1 antagonist.

    Science.gov (United States)

    Ghosal, Anima; Lu, Xiaowen; Penner, Natalia; Gao, Lan; Ramanathan, Ragu; Chowdhury, Swapan K; Kishnani, Narendra S; Alton, Kevin B

    2011-01-01

    Vorapaxar (SCH 530348), a potent oral thrombin protease-activated receptor 1 antagonist, is being developed as an antiplatelet agent for patients with established vascular disease. The objective of this study was to identify the human liver cytochrome P450 (P450) enzyme(s) responsible for the metabolism of SCH 530348. Human liver microsomes metabolized SCH 530348 to M19, an amine metabolite formed via carbamate cleavage, and M20 (monohydroxy-SCH 530348). Recombinant human CYP3A4 exhibited the most activity (11.5% profiled radioactivity) for the formation of M19, followed by markedly less substrate conversion with CYP1A1 and CYP2C19. Trace levels of M19, a major excreted human metabolite, were detected with CYP1A2, CYP3A5, and CYP4F3A. Formation of M19 by human liver microsomes was inhibited 89% by ketoconazole (IC(50), 0.73 μM), 34% by tranylcypromine, and 89% by anti-CYP3A4 monoclonal antibody. There was a significant correlation between the rate of M19 formation and midazolam 1'-hydroxylation (r = 0.75) or M19 formation and testosterone 6β-hydroxylation (r = 0.92). The results of screening, inhibition, and correlation studies confirmed that CYP3A4 is the major P450 enzyme responsible for M19 formation from SCH 530348. In contrast, formation of M20, a major circulating human metabolite at steady state, was primarily catalyzed by CYP3A4 and CYP2J2. M20 is pharmacologically equipotent to SCH 530348, whereas M19 is an inactive metabolite. Formation of M20 by human liver microsomes was inhibited 89% by ketoconazole, 75% by astemizole (a CYP2J2 inhibitor), and 43% by CYP3A4 monoclonal antibody. These results suggest that CYP3A4 and CYP2J2 are both involved in the formation of M20 metabolite. PMID:20926621

  13. Hot topic [editorial

    International Nuclear Information System (INIS)

    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.)

  14. Recombinant DNA for Teachers.

    Science.gov (United States)

    Duvall, James G., III

    1992-01-01

    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…

  15. Recombineering Pseudomonas syringae

    Science.gov (United States)

    Here we report the identification of functions that promote genomic recombination of linear DNA introduced into Pseudomonas cells by electroporation. The genes encoding these functions were identified in Pseudomonas syringae pv. syringae B728a based on similarity to the lambda Red Exo/Beta and RecE...

  16. Preparation of brightly fluorescent silica nanoparticles modified with lucigenin and chitosan, and their application to an aptamer-based sandwich assay for thrombin

    International Nuclear Information System (INIS)

    We report on the preparation of fluorescent silica nanoparticles (SiNPs) modified with chitosan and lucigenin by using a reverse microemulsion method. The introduction of chitosan to the lucigenin doped SiNPs is shown to improve the fluorescence quantum yield. The modified SiNPs were used as fluorescent markers in an aptamer-based method for selective determination of thrombin. In this protocol, thrombin was sandwiched between streptavidin-coated magnetic beads and the fluorescent SiNPs modified with a thrombin-binding aptamer. The method was successfully applied to the determination of thrombin in human serum and showed a detection limit as low as 0.02 nM. In our perception, the protocol presented here is promising in that such SiNPs may be applied to the sensitive fluorescent detection of other analytes by changing the corresponding aptamer. (author)

  17. Thrombin mediates migration of rat brain astrocytes via PLC, Ca²⁺, CaMKII, PKCα, and AP-1-dependent matrix metalloproteinase-9 expression.

    Science.gov (United States)

    Lin, Chih-Chung; Lee, I-Ta; Wu, Wen-Bin; Liu, Chiung-Ju; Hsieh, Hsi-Lung; Hsiao, Li-Der; Yang, Chien-Chung; Yang, Chuen-Mao

    2013-12-01

    Matrix metalloproteinase-9 (MMP-9) plays a crucial role in pathological processes of brain inflammation, injury, and neurodegeneration. Thrombin has been known as a regulator of MMP-9 expression and cells migration. However, the mechanisms underlying thrombin-induced MMP-9 expression in rat brain astrocytes (RBA-1 cells) remain unclear. Here, we demonstrated that thrombin induced the expression of pro-form MMP-9 and migration of RBA-1 cells, which were inhibited by pretreatment with the inhibitor of Gq-coupled receptor (GPAnt2A), Gi/o-coupled receptor (GPAnt2), PC-PLC (D609), PI-PLC (U73122), Ca(2+)-ATPase (thapsigargin, TG), calmodulin (CaMI), CaMKII (KN62), PKC (Gö6976 or GF109203X), MEK1/2 (PD98059), p38 MAPK (SB202190), JNK1/2 (SP600125), or AP-1 (Tanshinone IIA) or the intracellular calcium chelator (BAPTA/AM) and transfection with siRNA of PKCα, Erk2, JNK1, p38 MAPK, c-Jun, or c-Fos. In addition, thrombin-induced elevation of intracellular Ca(2+) concentration was attenuated by PPACK (a thrombin inhibitor). Thrombin further induced CaMKII phosphorylation and PKCα translocation, which were inhibited by U73122, D609, KN62, TG, or BAPTA/AM. Thrombin also induced PKCα-dependent p42/p44 MAPK and JNK1/2, but not p38 MAPK activation. Finally, we showed that thrombin enhanced c-Fos expression and c-Jun phosphorylation. c-Fos mRNA levels induced by thrombin were reduced by PD98059, SP600125, and Gö6976, but not SB202190. Thrombin stimulated in vivo binding of c-Fos to the MMP-9 promoter, which was reduced by pretreatment with SP600125 or PD98059, but not SB202190. These results concluded that thrombin activated a PLC/Ca(2+)/CaMKII/PKCα/p42/p44 MAPK and JNK1/2 pathway, which in turn triggered AP-1 activation and ultimately induced MMP-9 expression in RBA-1 cells.

  18. An electrochemical aptasensor based on TiO2/MWCNT and a novel synthesized Schiff base nanocomposite for the ultrasensitive detection of thrombin.

    Science.gov (United States)

    Heydari-Bafrooei, Esmaeil; Amini, Maryam; Ardakani, Mehdi Hatefi

    2016-11-15

    A sensitive aptasensor based on a robust nanocomposite of titanium dioxide nanoparticles, multiwalled carbon nanotubes (MWCNT), chitosan and a novel synthesized Schiff base (SB) (TiO2/MWCNT/CHIT/SB) on the surface of a glassy carbon electrode (GCE) was developed for thrombin detection. The resultant nanocomposite can provide a large surface area, excellent electrocatalytic activity, and high stability, which would improve immobilization sites for biological molecules, allow remarkable amplification of the electrochemical signal and contribute to improved sensitivity. Thrombin aptamers were simply immobilized onto the TiO2-MWCNT/CHIT-SB nanocomposite matrix through simple π - π stacking and electrostatic interactions between CHIT/SB and aptamer strands. The electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were used to analyze the surface characterization of unmodified GCE and TiO2-MWCNT/CHIT-SB modified GCE, and also the interaction between aptamer and thrombin. In the presence of thrombin, the aptamer on the adsorbent layer captures the target on the electrode interface, which makes a barrier for electrons and inhibits electron transfer, thereby resulting in decreased DPV and increased impedance signals of the TiO2-MWCNT/CHIT-SB modified GCE. Furthermore, the proposed aptasensor has a very low LOD of 1.0fmolL(-1) thrombin within the detection range of 0.00005-10nmolL(-1). The aptasensor also presents high specificity and reproducibility for thrombin, which is unaffected by the coexistence of other proteins. Clinical application was performed with analysis of the thrombin levels in blood and CSF samples obtained from patients with MS, Parkinson, Epilepsy and Polyneuropathy using both the aptasensor and commercial ELISA kit. The results revealed the proposed system to be a promising candidate for clinical analysis of thrombin. PMID:27295570

  19. Dabigatran, a direct thrombin inhibitor, blocks differentiation of normal fibroblasts to a myofibroblast phenotype and demonstrates anti-fibrotic effects on scleroderma lung fibroblasts

    OpenAIRE

    Bogatkevich, Galina S.; Ludwicka-Bradley, Anna; Silver, Richard M.

    2009-01-01

    Myofibroblasts are the principal mesenchymal cells responsible for tissue remodeling, collagen deposition, and the restrictive nature of lung parenchyma associated with pulmonary fibrosis. We previously reported that thrombin activates protease-activated receptor (PAR)-1 thereby inducing normal lung fibroblasts to differentiate to a myofibroblast phenotype resembling scleroderma lung myofibroblasts. Here we demonstrate that the thrombin inhibitor dabigatran inhibits in a dose-dependant manner...

  20. The topical treatment of psoriasis.

    NARCIS (Netherlands)

    Kerkhof, P.C.M. van de; Vissers, W.H.P.M.

    2003-01-01

    According to the patients, improvement of efficacy, long-term safety and improvement of compliance are needed. The topical treatment has been innovated during the last decade. Most important are the introduction of two new classes of treatments: topical vitamin D(3) analogues and the retinoid tazaro

  1. Topics of Evolutionary Computation 2001

    DEFF Research Database (Denmark)

    Ursem, Rasmus Kjær

    This booklet contains the student reports from the course: Topics of Evolutionary Computation, Fall 2001, given by Thiemo Krink, Rene Thomsen and Rasmus K. Ursem......This booklet contains the student reports from the course: Topics of Evolutionary Computation, Fall 2001, given by Thiemo Krink, Rene Thomsen and Rasmus K. Ursem...

  2. Linguistic Extensions of Topic Models

    Science.gov (United States)

    Boyd-Graber, Jordan

    2010-01-01

    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…

  3. A colorimetric sandwich-type assay for sensitive thrombin detection based on enzyme-linked aptamer assay.

    Science.gov (United States)

    Park, Jun Hee; Cho, Yea Seul; Kang, Sungmuk; Lee, Eun Jeong; Lee, Gwan-Ho; Hah, Sang Soo

    2014-10-01

    A colorimetric sandwich-type assay based on enzyme-linked aptamer assay has been developed for the fast and sensitive detection of as low as 25 fM of thrombin with high linearity. Aptamer-immobilized glass was used to capture the target analyte, whereas a second aptamer, functionalized with horseradish peroxidase (HRP), was employed for the conventional 3,5,3',5'-tetramethylbenzidine (TMB)-based colorimetric detection. Without the troublesome antibody requirement of the conventional enzyme-linked immunosorbent assay (ELISA), as low as 25 fM of thrombin could be rapidly and reproducibly detected. This assay has superior, or at least equal, recovery and accuracy to that of conventional antibody-based ELISA.

  4. Purification and Characterization of Jararassin-I,A Thrombin-like Enzyme from Bothrops jararaca Snake Venom

    Institute of Scientific and Technical Information of China (English)

    Débora F. VIEIRA; Leandra WATANABE; Carolina D. SANT'ANA; Silvana MARCUSSI; Suely V. SAMPAIO; Andreimar M. SOARES; Raghuvir K. ARNI

    2004-01-01

    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 Bβ chain of fibrinogen while the Aα chain and γ chain were unchanged.Proteases inhibitors, PMSF and benzamidine inhibited the coagulant activity. These results showed jararassinI is a serine protease similar to coagulating thrombin-like snake venom proteases, but it specifically cleaves Bβ chain of bovine fibrinogen. Single crystals of enzyme were obtained (0.2 mm×0.2 mm×0.2 mm) and used for X-ray diffraction experiments.

  5. Three months of strictly controlled daily endurance exercise reduces thrombin generation and fibrinolytic risk markers in younger moderately overweight men

    DEFF Research Database (Denmark)

    Gram, Anne Sofie; Bladbjerg, Else-Marie; Skov, Jane;

    2015-01-01

    PURPOSE: Physical activity is associated with a decreased risk of cardiovascular disease, but dose dependency of long-term physical exercise on biomarkers within coagulation and fibrinolysis is unknown. We aimed to investigate effects of two doses of daily endurance exercise on biomarkers...... of 600 kcal/day (HIGH), 300 kcal/day (MOD), or to maintain their habitual lifestyle (CON). Fasting blood samples were collected before and after the intervention and analysed for thrombin generation (endogenous thrombin potential, ETP) and concentrations of tissue-type plasminogen activator (t...... of the haemostatic balance in overweight men. METHODS: Haemostatic variables were investigated in 53 healthy, younger (20-40 years), moderately overweight (BMI 25-30 kg/m(2)) men randomly assigned to 3 months of strictly controlled endurance exercise at two different doses corresponding to an energy expenditure...

  6. Advancements in Topical Antifungal Vehicles.

    Science.gov (United States)

    Kircik, Leon H

    2016-02-01

    The primary treatment for superficial fungal infections is antifungal topical formulations, and allylamines and azoles represent the two major classes of topical formulations that are used to treat these infections. The stratum corneum (SC) is composed of keratinocytes that are surrounded by a matrix of lipids. The efficacy of topically applied formulations depends on their ability to penetrate this lipid matrix, and the vehicle plays an integral role in the penetration of active molecule into skin. There are several challenges to formulating topical drugs, which include the biotransformation of the active molecules as they pass through the SC and the physical changes that occur to the vehicle itself when it is applied to the skin. This article will review current and emerging topical antifungal vehicles. PMID:26885798

  7. Spectroscopic and Electrochemical Detection of Thrombin/5'-SH or 3'-SH Aptamer Immobilized on (porous) Gold Substrates

    Energy Technology Data Exchange (ETDEWEB)

    Park, Buem Jin; Sa, Young Seung; Kim, Yong Hwan; Kim, Young Hun [Kwangwoon University, Seoul (Korea, Republic of)

    2012-01-15

    Thrombin is a serine protease that catalyzes the conversion of soluble fibrinogen to insoluble fibrin, and thus induces physiological and pathological blood coagulation. Therefore, it is important to detect thrombin in blood serum for purposes of diagnosis. To achieve this goal, it has been suggested that a 15-mer aptamer strongly binds with thrombin to form a G-quartet structure of the aptamer. Generally, 5'-end thiol-functionalized aptamer has been used as an anti-thrombin binder. Herein, we evaluate the possibility of utilizing a 3'-SH aptasensor for thrombin detection using SPR spectroscopy, and compare the enhancement of the electrochemical signal of the thrombin-aptamer bound on a porous gold substrate. Although the two aptamers have similar configurations, in SPR analysis, the 3'-SH aptamer was a effective aptasensor as well as 5'-SH aptamer. Results from electrochemical analysis showed that the porous gold substrate acted as a good substrate for an aptasensor and demonstrated 5-fold enhancement of current change, as compared to gold thin film.

  8. The Selection of DNA Aptamers for Two Different Epitopes of Thrombin Was Not Due to Different Partitioning Methods

    OpenAIRE

    Wilson, Robert; Cossins, Andrew; Nicolau, Dan V.; Missailidis, Sotiris

    2013-01-01

    Nearly all aptamers identified so far for any given target molecule have been specific for the same binding site (epitope). The most notable exception to the 1 aptamer per target molecule rule is the pair of DNA aptamers that bind to different epitopes of thrombin. This communication refutes the suggestion that these aptamers exist because different partitioning methods were used when they were selected. The possibility that selection of these aptamers was biased by conflicting secondary stru...

  9. Shc adaptor proteins are key transducers of mitogenic signaling mediated by the G protein-coupled thrombin receptor

    DEFF Research Database (Denmark)

    Chen, Y; Grall, D; Salcini, A E;

    1996-01-01

    The serine protease thrombin activates G protein signaling systems that lead to Ras activation and, in certain cells, proliferation. Whereas the steps leading to Ras activation by G protein-coupled receptors are not well defined, the mechanisms of Ras activation by receptor tyrosine kinases have...... kinase activation, gene induction and cell growth. From these data, we conclude that Shc represents a crucial point of convergence between signaling pathways activated by receptor tyrosine kinases and G protein-coupled receptors....

  10. Comparison of ultrasound-guided thrombin injection and compression repair in treatment of iatrogenic femoral arterial pseudoaneurysms

    Institute of Scientific and Technical Information of China (English)

    QIN Jun; GAO Yun-hua; ZHUO Zhong-xiong; HUANG Lan; LI Ai-min; SONG Yao-ming; JIN Jun; YU Xue-jun; GENG Zhao-hua; ZHOU Xia-bo; LIN Chun-mei

    2006-01-01

    Objective:To retrospectively compare the efficacy and safety of ultrasound-guided thrombin injection (UGTI) with ultrasound-guided compression repair (UGCR) in patients with postcatheterizational femoral arterial pseudoaneurysms (PSA). Methods: Thirty patients of this iatrogenic PSA [8males, 22 females, average age (66.5± 5.2) years] in our nstitution from 1997 to 2004 were retrospectively analyzed. Among them, 11 patients were treated with UGCR, 2 under continuous ultrasonographic (US) guidance and 9 under the guidance of femoral arterial bruit auscultation and dorsalis pedis artery palpation. Because UGCR was failed in 5 patients, consecutively 24 patients were treated with UGTI. Wine thrombin solution at a concentration of 200 U/ml was injected percutaneously using 22-25 gauge needles under color Doppler US. Demographics, clinical variables, pseudoaneurysm characteristics, and results of the 2 groups were compared by using Fisher's exact test and Student's t test. Results: The initial success rate of UGCR was 36.4% (4/11) nd the overall success rate was 45.5% (5/11). Ten of 11 patients suffered from local pain during the compression, but there was no any complication in UGTI group. The average dose of injected thrombin was (180±82) U for PSA of a single loculus and (315±150) U for multiloculated PSA. The initial success rate of UGTI was 89.5% (17/19) and the verall uccess rate was 100% (24/24). Conclusion:UGTI offers a safe, quick and effective means of definitively treating femoral pseudoaneurysms and seems superior to UGCR. The amount of thrombin applied on our people seems smaller compared with others' work.

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

    OpenAIRE

    Ziv-Polat O; Skaat H; Shahar A; Margel S

    2012-01-01

    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 phys...

  12. Anti-thrombin therapy during warm ischemia and cold preservation prevents chronic kidney graft fibrosis in a DCD model

    OpenAIRE

    Favreau, F.; Thuillier, R.; Cau, J; S. Milin; Manguy, E; Mauco, G; Zhu, X.; Lerman, LO; Hauet, T.

    2009-01-01

    Ischemia reperfusion injury (IRI) is pivotal for renal fibrosis development via peritubular capillaries injury. Coagulation represents a key mechanism involved in this process. Melagatran® (M), a thrombin inhibitor, was evaluated in an autotransplanted kidney model, using Large White pigs. To mimic deceased after cardiac death donor conditions, kidneys underwent warm ischemia (WI) for 60 min before cold preservation for 24 hours in University of Wisconsin solution. Treatment with M before WI ...

  13. Evidence supporting the use of recombinant activated factor VII in congenital bleeding disorders

    Directory of Open Access Journals (Sweden)

    Pär I Johansson

    2010-06-01

    Full Text Available Pär I Johansson, Sisse R OstrowskiCapital Region Blood Bank, Section for Transfusion Medicine, Rigshospitalet, University of Copenhagen, Copenhagen, DenmarkBackground: Recombinant activated factor VII (rFVIIa, NovoSeven® was introduced in 1996 for the treatment of hemophilic patients with antibodies against coagulation factor VIII or IX.Objective: To review the evidence supporting the use of rFVIIa for the treatment of patients with congenital bleeding disorders.Patients and methods: English-language databases were searched in September 2009 for reports of randomized controlled trials (RCTs evaluating the ability of rFVIIa to restore hemostasis in patients with congenital bleeding disorders.Results: Eight RCTs involving 256 hemophilic patients with antibodies against coagulation factors, also known as inhibitors, were identified. The evidence supporting the use of rFVIIa in these patients was weak with regard to dose, clinical setting, mode of administration, efficacy, and adverse events, given the limited sample size of each RCT and the heterogeneity of the studies.Conclusion: The authors suggest that rFVIIa therapy in hemophilic patients with inhibitors should be based on the individual’s ability to generate thrombin and form a clot, and not on the patient’s weight alone. Therefore, assays for thrombin generation, such as whole-blood thromboelastography, have the potential to significantly improve the treatment of these patients.Keywords: hemophilia, inhibitors, coagulation factor VIII, coagulation factor IX, rFVIIa, NovoSeven, FEIBA, hemostasis, RCT

  14. A signal amplification strategy using the cascade catalysis of gold nanoclusters and glucose dehydrogenase for ultrasensitive detection of thrombin.

    Science.gov (United States)

    Han, Jing; Zhuo, Ying; Chai, Yaqin; Gui, Guofeng; Zhao, Min; Zhu, Qiang; Yuan, Ruo

    2013-12-15

    This work reports a novel signal amplification strategy for ultrasensitive detection of thrombin by cascade catalysis of gold nanoclusters (AuNCs) and glucose dehydrogenase (GDH). Herein, the AuNCs prepared by using polyamidoamine dendrimer as template were constructed not only as nanocarriers for anchoring the large amounts of secondary thrombin aptamers but also as nanocatalysts to catalyze the oxidation of NADH efficiently. Moreover, a large amount of GDH was loaded through the immobilization technology of DNA hybridization and a large amount of toluidine blue (Tb) was intercalated into the DNA grooves via electrostatic interaction. Significantly, the electrochemical signal was greatly enhanced based on cascade catalysis: firstly, GDH catalyzed the oxidation of glucose to gluconolactone with the concomitant generation of NADH in the presence of NAD(+). Then, AuNCs as nanocatalysts could effectively catalyze NADH to produce NAD(+) with the help of Tb as redox probe. Under the optimal conditions, the proposed aptasensor exhibits a linear range of 1.0×10(-14)-5×10(-9) M with a low detection limit of 3.3×10(-15) M for thrombin detection and shows high sensitivity and good specificity. PMID:23850783

  15. Microfluidic Chip-Based Online Screening Coupled to Mass Spectrometry: Identification of Inhibitors of Thrombin and Factor Xa.

    Science.gov (United States)

    Iyer, Janaki Krishnamoorthy; Otvos, Reka A; Kool, Jeroen; Kini, R Manjunatha

    2016-02-01

    Thrombin and factor Xa (FXa) are critical enzymes of the blood coagulation cascade and are excellent targets of anticoagulant agents. Natural sources present an array of anticoagulants that can be developed as antithrombotic drugs. High-resolution, online screening techniques have been developed for the identification of drug leads from complex mixtures. In this study, we have developed and optimized a microfluidic online screening technique coupled to nano-liquid chromatography (LC) and in parallel with a mass spectrometer for the identification of thrombin and FXa inhibitors in mixtures. Inhibitors eluting from the nano-LC were split postcolumn in a 1:1 ratio; half was fed into a mass spectrometer (where its mass is detected), and the other half was fed into a microfluidic chip (which acts as a microreactor for the online assays). With our platform, thrombin and FXa inhibitors were detected in the assay in parallel with their mass identification. These methods are suitable for the identification of inhibitors from sample amounts as low as sub-microliter volumes. PMID:26323281

  16. Thrombin-cleaved COOH(-) terminal osteopontin peptide binds with cyclophilin C to CD147 in murine breast cancer cells.

    Science.gov (United States)

    Mi, Zhiyong; Oliver, Tim; Guo, Hongtao; Gao, Chengjiang; Kuo, Paul C

    2007-05-01

    Osteopontin is a glycoprotein that has been linked to metastatic function in breast, lung, and prostate cancers. However, the mechanism by which osteopontin acts to induce metastatic properties is largely unknown. One intriguing feature of osteopontin is the presence of a conserved thrombin cleavage site that is COOH-terminal from a well-characterized RGD domain. Although the COOH-terminal fragment may bind to cell surface CD44 receptors, little is known about the COOH-terminal osteopontin fragment. In the current study, we use the murine mammary epithelial tumor cell lines 4T1 and 4T07; these cells are thioguanine-resistant sublines derived from the parental population of 410.4 cells from Balb/cfC3H mice. Using flow cytometry and Forster resonance energy transfer, we show that the COOH-terminal fragment of osteopontin binds with another marker of metastatic function (cyclophilin C or rotamase) to the CD147 cell surface glycoprotein (also known as extracellular matrix metalloproteinase inducer), to activate Akt1/2 and matrix metalloproteinase-2. In in vitro assays, thrombin cleavage of osteopontin to generate short COOH-terminal osteopontin in the presence of cyclophilin C increases migration and invasion of both 4T07 and 4T1 cells. This interaction between osteopontin peptide and cyclophilin C has not been previously described but assigns a heretofore unknown function for the thrombin-cleaved osteopontin COOH-terminal fragment.

  17. 3,4,9,10-Perylenetetracarboxylic dianhydride functionalized graphene sheet as labels for ultrasensitive electrochemiluminescent detection of thrombin.

    Science.gov (United States)

    Gan, Xianxue; Yuan, Ruo; Chai, Yaqin; Yuan, Yali; Cao, Yaling; Liao, Yuhong; Liu, Huijing

    2012-05-13

    A novel tracer, 3,4,9,10-perylenetetracarboxylic dianhydride (PTCDA) functionalized graphene sheet (GS) composite (GS-TCDA), is employed to label the secondary anti-thrombin aptamer (TBA) to construct an ultrasensitive electrochemiluminescent sandwich-type aptasensor. The GS provided large surface area for loading abundant PTCDA and TBA with good stability and biocompatibility. Because of the excellent electroconductivity of GS and the desirable optical properties of PTCDA, the as-formed Apt II bioconjugate considerably amplified the electrochmiluminescence (ECL) signal of peroxydisulfate (S(2)O(8)(2-)) and worked as the desirable label for Apt II. On the basis of the considerably amplified ECL signal and sandwich format, an extremely wide range from 1 fM to 1 nM with an ultralow detection limit of 0.33 fM for thrombin was obtained. Additionally, the selectivity and stability of the proposed aptasensor were also excellent. Thus, this procedure has great promise for detection of thrombin present at ultra-trace levels during early stage of diseases. PMID:22541015

  18. A sensitive sandwich-type electrochemical aptasensor for thrombin detection based on platinum nanoparticles decorated carbon nanocages as signal labels.

    Science.gov (United States)

    Gao, Fenglei; Du, Lili; Zhang, Yu; Zhou, Fuyi; Tang, Daoquan

    2016-12-15

    In this work, a novel and sensitive sandwich-type electrochemical aptasensor has been developed for thrombin detection based on platinum nanoparticles (Pt NPs) decorated carbon nanocages (CNCs) as signal tags. The morphological and compositional of the Pt NPs/CNCs were examined using transmission electron microscopy, X-ray diffraction, and Raman spectroscopy. The results showed that the Pt NPs with about 3-5nm in diameter were well dispersed on the surface of CNCs. The thiolated aptamer was firstly immobilized on the gold electrode to capture the thrombin molecules, and then aptamer functionalized Pt NPs/CNCs nanocomposites were used to fabricate a sandwich sensing platform. Then, the high-content Pt NPs on carbon nanocages acting as hydrogen peroxide-mimicking enzyme catalyzed the reduction of H2O2, resulting in significant electrochemical signal amplification. Differential pulse voltammetry is employed to detect thrombin with different concentrations. Under optimized conditions, the approach provided a good linear response range from 0.05 pM to 20nM with a low detection limit of 10fM. This Pt NPs/CNCs-based aptasensor shows good precision, acceptable stability and reproducibility, which provided a promising strategy for electrochemical aptamer-based detection of other biomolecules. PMID:27376191

  19. Effect of the oral thrombin inhibitor dabigatran on allergic lung inflammation induced by repeated house dust mite administration in mice.

    Science.gov (United States)

    de Boer, Johannes D; Berkhout, Lea C; de Stoppelaar, Sacha F; Yang, Jack; Ottenhoff, Roelof; Meijers, Joost C M; Roelofs, Joris J T H; van't Veer, Cornelis; van der Poll, Tom

    2015-10-15

    Asthma is a chronic disease of the airways; asthma patients are hampered by recurrent symptoms of dyspnoea and wheezing caused by bronchial obstruction. Most asthma patients suffer from chronic allergic lung inflammation triggered by allergens such as house dust mite (HDM). Coagulation activation in the pulmonary compartment is currently recognized as a feature of allergic lung inflammation, and data suggest that coagulation proteases further drive inflammatory mechanisms. Here, we tested whether treatment with the oral thrombin inhibitor dabigatran attenuates allergic lung inflammation in a recently developed HDM-based murine asthma model. Mice were fed dabigatran (10 mg/g) or placebo chow during a 3-wk HDM airway exposure model. Dabigatran treatment caused systemic thrombin inhibitory activity corresponding with dabigatran levels reported in human trials. Surprisingly, dabigatran did not lead to inhibition of HDM-evoked coagulation activation in the lung as measured by levels of thrombin-antithrombin complexes and D-dimer. Repeated HDM administration caused an influx of eosinophils and neutrophils into the lungs, mucus production in the airways, and a T helper 2 response, as reflected by a rise in bronchoalveolar IL-4 and IL-5 levels and a systemic rise in IgE and HDM-IgG1. Dabigatran modestly improved HDM-induced lung pathology (P dabigatran in spite of adequate plasma levels, these results argue against clinical evaluation of dabigatran in patients with asthma.

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

    Energy Technology Data Exchange (ETDEWEB)

    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

    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.

  1. Evidence of recombination within human alpha-papillomavirus

    Directory of Open Access Journals (Sweden)

    Carvajal-Rodríguez Antonio

    2007-03-01

    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.

  2. Topic Model for Graph Mining.

    Science.gov (United States)

    Xuan, Junyu; Lu, Jie; Zhang, Guangquan; Luo, Xiangfeng

    2015-12-01

    Graph mining has been a popular research area because of its numerous application scenarios. Many unstructured and structured data can be represented as graphs, such as, documents, chemical molecular structures, and images. However, an issue in relation to current research on graphs is that they cannot adequately discover the topics hidden in graph-structured data which can be beneficial for both the unsupervised learning and supervised learning of the graphs. Although topic models have proved to be very successful in discovering latent topics, the standard topic models cannot be directly applied to graph-structured data due to the "bag-of-word" assumption. In this paper, an innovative graph topic model (GTM) is proposed to address this issue, which uses Bernoulli distributions to model the edges between nodes in a graph. It can, therefore, make the edges in a graph contribute to latent topic discovery and further improve the accuracy of the supervised and unsupervised learning of graphs. The experimental results on two different types of graph datasets show that the proposed GTM outperforms the latent Dirichlet allocation on classification by using the unveiled topics of these two models to represent graphs. PMID:25616091

  3. Mental Mechanisms for Topics Identification

    Directory of Open Access Journals (Sweden)

    Louis Massey

    2014-01-01

    Full Text Available Topics identification (TI is the process that consists in determining the main themes present in natural language documents. The current TI modeling paradigm aims at acquiring semantic information from statistic properties of large text datasets. We investigate the mental mechanisms responsible for the identification of topics in a single document given existing knowledge. Our main hypothesis is that topics are the result of accumulated neural activation of loosely organized information stored in long-term memory (LTM. We experimentally tested our hypothesis with a computational model that simulates LTM activation. The model assumes activation decay as an unavoidable phenomenon originating from the bioelectric nature of neural systems. Since decay should negatively affect the quality of topics, the model predicts the presence of short-term memory (STM to keep the focus of attention on a few words, with the expected outcome of restoring quality to a baseline level. Our experiments measured topics quality of over 300 documents with various decay rates and STM capacity. Our results showed that accumulated activation of loosely organized information was an effective mental computational commodity to identify topics. It was furthermore confirmed that rapid decay is detrimental to topics quality but that limited capacity STM restores quality to a baseline level, even exceeding it slightly.

  4. Peptide affinity labels for thrombin and other trypsin-like proteases

    Science.gov (United States)

    Shaw, Elliott N.; Kettner, Charles A.

    1982-03-09

    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.

  5. Unfolding mechanism of thrombin-binding aptamer revealed by molecular dynamics simulation and Markov State Model

    Science.gov (United States)

    Zeng, Xiaojun; Zhang, Liyun; Xiao, Xiuchan; Jiang, Yuanyuan; Guo, Yanzhi; Yu, Xinyan; Pu, Xuemei; Li, Menglong

    2016-04-01

    Thrombin-binding aptamer (TBA) with the sequence 5‧GGTTGGTGTGGTTGG3‧ could fold into G-quadruplex, which correlates with functionally important genomic regionsis. However, unfolding mechanism involved in the structural stability of G-quadruplex has not been satisfactorily elucidated on experiments so far. Herein, we studied the unfolding pathway of TBA by a combination of molecular dynamics simulation (MD) and Markov State Model (MSM). Our results revealed that the unfolding of TBA is not a simple two-state process but proceeds along multiple pathways with multistate intermediates. One high flux confirms some observations from NMR experiment. Another high flux exhibits a different and simpler unfolding pathway with less intermediates. Two important intermediate states were identified. One is similar to the G-triplex reported in the folding of G-quadruplex, but lack of H-bonding between guanines in the upper plane. More importantly, another intermediate state acting as a connector to link the folding region and the unfolding one, was the first time identified, which exhibits higher population and stability than the G-triplex-like intermediate. These results will provide valuable information for extending our understanding the folding landscape of G-quadruplex formation.

  6. Thrombin Activatable Fibrinolysis Inhibitor in Preeclmapsia and Gestational Hypertension throughout the Gestation

    Institute of Scientific and Technical Information of China (English)

    Yinghong ZHANG; Yu HU; Tao GUO; Wenning WEI; Xiaoping ZHANG

    2008-01-01

    To clarify the role of TAFI in hypertensive disorders in pregnancy, 22 subjects, including 10 with pre-eclampsia (PE) and 12 with gestational hypertension were examined for the levels of TAFI and thrombin-antithrombin (TAT) complex. Thirty normal pregnant women served as controls. ELISA was employed for the detection. The results showed that the TAFI antigen levels in normal pregnancy group, gestational hypertension group and PE group were (85.35±24.69)%, (99.65±18.27)%, (110.12±23.36)%; (97.06±21.40)%, (114.08±27.76)%, (125.49±24.70)%; (106.6±19.21)%, (129.2±25.07)%, (139.1±30.12)%, in the 1st, 2nd and 3rd trimester respectively. No significant differences were found between the normal pregnancy group and gestational hypertension group but significant difference existed between normal pregnancy group and PE group in each tri- mester (P<0.05). TAT complexes were significantly higher in patients with PE than that in controls (P<0.05), but no correlation was found between TAT and TAFI. It is concluded that TAFI may con- tributed to the impairment of fibrinolysis in the patients with PE and may serves as a sensitive indi- cator for PE, but it may not help in the diagnosis of the gestational hypertension.

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

    Directory of Open Access Journals (Sweden)

    J.H. Petretski

    2000-11-01

    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.

  8. The future of topical analgesics.

    Science.gov (United States)

    Arnstein, Paul M

    2013-07-01

    Topically applied analgesic therapies have been used throughout history to treat a variety of patient conditions that present with pain. Before modem pharmaceuticals became readily available, mud-based emollients, salves, cold therapies, and other natural remedies were often used. Now we have effective therapies and are developing advanced topical analgesics as we learn more about the physiology and pathophysiology of pain. The use of topical analgesics may be associated with fewer patient systemic side effects than are seen with oral, parenteral, or transdermally administered agents, making the topical route of administration attractive to prescribers and patients. With further refinement of existing drugs and the development of novel agents, topical analgesics may offer relief for treating patient pain conditions that are currently challenging to treat, such as pain resulting from burns, wound debridement, and pressure ulcers. Recognizing the value of a multimodal approach, topical analgesics may offer a therapeutic option that can become part of a comprehensive treatment plan for the patient. With continued advancements in targeted drug-delivery systems, topical analgesics may be able to provide a method to prevent or reverse the phenomena of peripheral and central sensitization, or the neuroplastic changes believed to be responsible for the transition from acute to chronic pain states in patients. For those patients at risk for developing chronic pain states, such as complex regional pain syndrome, the combination of cutaneous stimulation (achieved through rubbing during application) and analgesic effects produced by the drug itself may prevent the disabling pain that often emerges during the subacute phase of disease. In summary, better utilization of currently available topical analgesics and continued research promise to ensure that topical analgesics are, and will continue to be, important tools in the treatment of patients with resistant pain. PMID

  9. AECL passive autocatalytic recombiners

    Energy Technology Data Exchange (ETDEWEB)

    Gardner, L.B.; Marcinkowska, K. [Atomic Energy of Canada Limited, Chalk River, Ontario (Canada)

    2012-03-15

    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)

  10. AECL passive autocatalytic recombiners

    Energy Technology Data Exchange (ETDEWEB)

    Gardner, L.B.; Marcinkowska, K. [Atomic Energy of Canada Limited, Chalk River, Ontario (Canada)

    2011-07-01

    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)

  11. RECOMBINANT INFLUENZA VACCINES

    OpenAIRE

    Sedova, E.; Shcherbinin, D.; Migunov, A.; Smirnov, Iu; Logunov, D.; Shmarov, M.; Tsybalova, L.; Naroditskiĭ, B.; O. Kiselev; Gintsburg, A.

    2012-01-01

    This review covers the problems encountered in the construction and production of new recombinant influenza vaccines. New approaches to the development of influenza vaccines are investigated; they include reverse genetics methods, production of virus-like particles, and DNA- and viral vector-based vaccines. Such approaches as the delivery of foreign genes by DNA- and viral vector-based vaccines can preserve the native structure of antigens. Adenoviral vectors are a promising gene-delivery pla...

  12. Soluble recombinant influenza vaccines.

    OpenAIRE

    Fiers, W; Neirynck, S; Deroo, T; Saelens, X; Jou, W M

    2001-01-01

    Soluble, recombinant forms of influenza A virus haemagglutinin and neuraminidase have been produced in cells of lower eukaryotes, and shown in a mouse model to induce complete protective immunity against a lethal virus challenge. Soluble neuraminidase, produced in a baculovirus system, consisted of tetramers, dimers and monomers. Only the tetramers were enzymatically active. The immunogenicity decreased very considerably in the order tetra > di > mono. Therefore, we fused the head part of the...

  13. Clindamycin and Benzoyl Peroxide Topical

    Science.gov (United States)

    ... benzoyl peroxide works by killing the bacteria that cause acne. ... E-Mycin, Erythrocin) and other topical medications for acne. Your doctor ... colitis (a condition which causes swelling and sores in the lining of the ...

  14. Do scientists trace hot topics?

    OpenAIRE

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

    2013-01-01

    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,...

  15. Key Topics in Sports Medicine

    OpenAIRE

    Amir Ali Narvani; Panagiotis Thomas; Burce Lynn

    2006-01-01

    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...

  16. Topics of Bioengineering in Wikipedia

    Directory of Open Access Journals (Sweden)

    Vassia Atanassova

    2009-10-01

    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.

  17. Dabigatran and its reversal with recombinant factor VIIa and prothrombin complex concentrate

    DEFF Research Database (Denmark)

    Sølbeck, Sacha; Nilsson, Caroline U; Engström, Martin;

    2014-01-01

    OBJECTIVE: Dabigatran is a new oral direct thrombin inhibitor. No specific antidote exists in the event of hemorrhage, but prothrombin complex concentrate (PCC) and recombinant activated factor VII (rFVIIa) are suggested therapies. Sonoclot is a bedside viscoelastic instrument for monitoring...... the coagulation process in whole blood. The aim of this study was to investigate the effect of dabigatran and reversal with PCC and rFVIIa, as monitored by the Sonoclot. METHODS: Citrated whole blood was drawn and mixed in vitro with dabigatran, dabigatran + PCC or dabigatran + rFVIIa and analyzed with three...... different Sonoclot cuvettes: Glassbead, kaolin and tissue factor (diluted) activated. RESULTS: The Sonoclot detected in vitro-induced anticoagulation due to dabigatran with the glassbead- and kaolin-activated cuvettes. There was no reversing effect of PCC, probably due to the presence of heparin in the PCC...

  18. A quantum dot-aptamer beacon using a DNA intercalating dye as the FRET reporter: application to label-free thrombin detection.

    Science.gov (United States)

    Chi, Chun-Wei; Lao, Yeh-Hsing; Li, Yi-Shan; Chen, Lin-Chi

    2011-03-15

    A new quantum dot (QD)-aptamer (apt) beacon that acts by folding-induced dissociation of a DNA intercalating dye, BOBO-3(B), is demonstrated with label-free thrombin detection. The beacon, denoted as QD-apt:B, is constructed by (1) coupling of a single-stranded thrombin aptamer to Qdot 565 via EDC/Sulfo-NHS chemistry and (2) staining the duplex regions of the aptamer on QD with excess BOBO-3 before thrombin binding. When mixing a thrombin sample with QD-apt:B, BOBO-3 is competed away from the beacon due to target-induced aptamer folding, which then causes a decrease in QD fluorescence resonance energy transfer (FRET)-mediated BOBO-3 emission and achieves thrombin quantitation. In this work, the effects of Mg(2+), coupling time, and aptamer type on the beacon's performances are investigated and discussed thoroughly with various methods, including transmission electron microscopy (TEM), dynamic light scattering (DLS), and two-color differential gel electrophoresis. Using the best aptamer beacon (HTQ37), we attain highly specific and wide-range detection (from nM to μM) of thrombin in buffer, and the beacon can sense nM-range thrombin in 15% diluted serum. Compared to the reported QD aptamer assays, our method is advantageous from the aspect of using a simple sensory unit design without losing the detection sensitivity. Therefore, we consider the QD-apt:B beacon a potential alternative to immuno-reagents and an effective tool to study nucleic acid folding on QD as well. PMID:21306887

  19. A quantum dot-aptamer beacon using a DNA intercalating dye as the FRET reporter: application to label-free thrombin detection.

    Science.gov (United States)

    Chi, Chun-Wei; Lao, Yeh-Hsing; Li, Yi-Shan; Chen, Lin-Chi

    2011-03-15

    A new quantum dot (QD)-aptamer (apt) beacon that acts by folding-induced dissociation of a DNA intercalating dye, BOBO-3(B), is demonstrated with label-free thrombin detection. The beacon, denoted as QD-apt:B, is constructed by (1) coupling of a single-stranded thrombin aptamer to Qdot 565 via EDC/Sulfo-NHS chemistry and (2) staining the duplex regions of the aptamer on QD with excess BOBO-3 before thrombin binding. When mixing a thrombin sample with QD-apt:B, BOBO-3 is competed away from the beacon due to target-induced aptamer folding, which then causes a decrease in QD fluorescence resonance energy transfer (FRET)-mediated BOBO-3 emission and achieves thrombin quantitation. In this work, the effects of Mg(2+), coupling time, and aptamer type on the beacon's performances are investigated and discussed thoroughly with various methods, including transmission electron microscopy (TEM), dynamic light scattering (DLS), and two-color differential gel electrophoresis. Using the best aptamer beacon (HTQ37), we attain highly specific and wide-range detection (from nM to μM) of thrombin in buffer, and the beacon can sense nM-range thrombin in 15% diluted serum. Compared to the reported QD aptamer assays, our method is advantageous from the aspect of using a simple sensory unit design without losing the detection sensitivity. Therefore, we consider the QD-apt:B beacon a potential alternative to immuno-reagents and an effective tool to study nucleic acid folding on QD as well.

  20. Effectiveness of three percent topical spray injection with recombinant human interleukin-11 on acute radioactive dermatitis%急性放射性皮炎外喷3%注射用重组人白介素-11的疗效观察

    Institute of Scientific and Technical Information of China (English)

    童芳; 刘群; 蔡博宁; 黄玉荣

    2015-01-01

    目的 观察放射性皮炎外用3%注射用重组人白介素-1 1(recombinant human interleukin-11,rhIL-11)的临床疗效.方法 将2013年1-9月在我科行放疗并发放射性皮炎患者140例随机分两组,试验组70例使用3% rhIL-11外喷皮炎创面,对照组70例使用康复新液外敷.结果 用药48 h后试验组疼痛缓解效果、皮炎康复效果均优于对照组(P<0.05).尤其体现在Ⅲ度放射性皮炎中,疼痛治疗效果试验组优于对照组(x2=4.792,P=0.041).实验组3d康复效果显效率明显高于对照组(80%vs32%,P=0.008).试验组7d康复效果有效率明显高于对照组(80%vs30%,P=0.004).结论 放疗后≥Ⅱ度放射性皮炎局部外喷3% rhIL-11溶液,可减轻急性放射性皮炎症状,促进康复.

  1. Recombinant Collagenlike Proteins

    Science.gov (United States)

    Fertala, Andzej

    2007-01-01

    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.

  2. Primordial magnetogenesis before recombination

    CERN Document Server

    Fabre, Ophélia

    2015-01-01

    The origin of large magnetic fields in the Universe remains currently unknown. We investigate here a mechanism before recombination based on known physics. The source of the vorticity is due to the changes in the photon distribution function caused by the fluctuations in the background photons. We show that the magnetic field generated in the MHD limit, due to the Coulomb scattering, is of the order $10^{-49}$ G. We explicitly show that the magnetic fields generated from this process are sustainable and are not erased by resistive diffusion. We compare the results with current observations and discuss the implications.

  3. Fibrinogen-thrombin collagen patch reinforcement of high-risk colonic anastomoses in rats

    Science.gov (United States)

    Suárez-Grau, Juan Manuel; Bernardos García, Carlos; Cepeda Franco, Carmen; Mendez García, Cristina; García Ruiz, Salud; Docobo Durantez, Fernando; Morales-Conde, Salvador; Padillo Ruiz, Javier

    2016-01-01

    AIM To evaluate the effectiveness of human fibrinogen-thrombin collagen patch (TachoSil®) in the reinforcement of high-risk colon anastomoses. METHODS A quasi-experimental study was conducted in Wistar rats (n = 56) that all underwent high-risk anastomoses (anastomosis with only two sutures) after colectomies. The rats were divided into two randomized groups: Control group (24 rats) and treatment group (24 rats). In the treatment group, high-risk anastomosis was reinforced with TachoSil® (a piece of TachoSil® was applied over this high-risk anastomosis, covering the gap). Leak incidence, overall survival, intra-abdominal adhesions, and histologic healing of anastomoses were analyzed. Survivors were divided into two subgroups and euthanized at 15 and 30 d after intervention in order to analyze the adhesions and histologic changes. RESULTS Overall survival was 71.4% and 57.14% in the TachoSil® group and control group, respectively (P = 0.29); four rats died from other causes and six rats in the treatment group and 10 in the control group experienced colonic leakage (P > 0.05). The intra-abdominal adhesion score was similar in both groups, with no differences between subgroups. We found non-significant differences in the healing process according to the histologic score used in both groups (P = 0.066). CONCLUSION In our study, the use of TachoSil® was associated with a non-statistically significant reduction in the rate of leakage in high-risk anastomoses. TachoSil® has been shown to be a safe product because it does not affect the histologic healing process or increase intra-abdominal adhesions. PMID:27721926

  4. [Laboratory assessment of haemostatic parameters in patients taking a direct thrombin inhibitor].

    Science.gov (United States)

    Beliavskaia, O O; Vavilova, T V

    2014-01-01

    The problem of prevention and treatment of thromboembolic complications has a significant place in clinical practice for many years. The gold-standard agents in long-term protection from embologenic strokes, secondary prevention of venous thromboses and embolisms still remain vitamin K antagonists (in Russia - warfarin). However, despite high efficacy, administration of warfarin is fraught with dangers and associated with a series of inconveniences. A direct thrombin inhibitor, dabigatran etexilate (hereinafter referred to as dabigatran) was approved in the Russian Federation for prevention of thromboembolic complications in orthopaedic practice (2009), for prevention of ischaemic embologenic stroke in atrial fibrillation (2011) and for treatment of recurrent thrombosis of deep veins and pulmonary artery thromboembolism (2014). A characteristic feature of a therapeutic agent possessing an anticoagulation effect is correlation between intensity of hypocoagulation and haemorrhage. The effect of dabigatran on the laboratory parameters of haemostasis has been studied insufficiently, with no practical guidelines on assessing these alterations for prediction of the risk for haemorrhagic and thromboembolic complications. The present study included a total of 65 patients with non-valvular aetiology atrial fibrillation, taking dabigatran during from 6 to 18 months. All patients underwent laboratory assessment of the coagulation level and measuring blood coagulation activation markers in dynamics 10-14 days, 1, 6, 12 and 18 months after taking the agent. Thromboembolic and haemorrhagic risks were also assessed. It was revealed that administration of dabigatran leads to alterations in the main parameters of coagulogram. Determination of prothrombin (in % according to Quick's method) and activated partial thromboplastin time may be used for qualitative assessment of hypocoagulation. During the follow up period no statistically significant changes in the coagulation activation

  5. Thrombin-dependent Incorporation of von Willebrand Factor into a Fibrin Network.

    Science.gov (United States)

    Miszta, Adam; Pelkmans, Leonie; Lindhout, Theo; Krishnamoorthy, Ganeshram; de Groot, Philip G; Hemker, Coenraad H; Heemskerk, Johan W M; Kelchtermans, Hilde; de Laat, Bas

    2014-12-26

    Attachment of platelets from the circulation onto a growing thrombus is a process involving multiple platelet receptors, endothelial matrix components, and coagulation factors. It has been indicated previously that during a transglutaminase reaction activated factor XIII (FXIIIa) covalently cross-links von Willebrand factor (VWF) to polymerizing fibrin. Bound VWF further recruits and activates platelets via interactions with the platelet receptor complex glycoprotein Ib (GPIb). In the present study we found proof for binding of VWF to a fibrin monomer layer during the process of fibrinogen-to-fibrin conversion in the presence of thrombin, arvin, or a snake venom from Crotalus atrox. Using a domain deletion mutant we demonstrated the involvement of the C domains of VWF in this binding. Substantial binding of VWF to fibrin monomers persisted in the presence of the FXIIIa inhibitor K9-DON, illustrating that cross-linking via factor XIII is not essential for this phenomenon and suggesting the identification of a second mechanism through which VWF multimers incorporate into a fibrin network. Under high shear conditions, platelets were shown to adhere to fibrin only if VWF had been incorporated. In conclusion, our experiments show that the C domains of VWF and the E domain of fibrin monomers are involved in the incorporation of VWF during the polymerization of fibrin and that this incorporation fosters binding and activation of platelets. Fibrin thus is not an inert end product but partakes in further thrombus growth. Our findings help to elucidate the mechanism of thrombus growth and platelet adhesion under conditions of arterial shear rate. PMID:25381443

  6. Antithrombotic properties of SSR182289A, a new, orally active thrombin inhibitor.

    Science.gov (United States)

    Lorrain, J; Millet, L; Lechaire, I; Lochot, S; Ferrari, P; Visconte, C; Sainte-Marie, M; Lunven, C; Berry, C N; Schaeffer, P; Herbert, J-M; O'Connor, S E

    2003-02-01

    N-[3-[[[(1S)-4-(5-Amino-2-pyridinyl)-1-[[4-difluoromethylene)-1-piperidinyl]carbonyl]butyl]amino]sulfonyl][1,1'-biphenyl]-2-yl]acetamide hydrochloride (SSR182289A) is a novel, potent, and selective thrombin inhibitor. We have examined the antithrombotic properties of SSR182289A administered by i.v. and p.o. routes in several different animal thrombosis models in comparison with reference antithrombotic agents. Oral administration of SSR182289A produced dose-related antithrombotic effects in the following models; rat venous thrombosis (ED(50) 0.9 mg/kg p.o.), rat silk thread arterio-venous (AV) shunt (ED(50) 3.8 mg/kg p.o.), rat thromboplastin-induced AV shunt (ED(50) 3.1 mg/kg p.o.), rat carotid artery thrombosis (ED(200) 5.9 mg/kg p.o.), and rabbit venous thrombosis (ED(50) 7.5 mg/kg p.o.). Administered as an i.v. bolus, SSR182289A showed antithrombotic activity in the above models with ED(50)/ED(200) values in the range of 0.2 to 1.9 mg/kg i.v. SSR182289A increased rat tail transection bleeding time at doses > or =10 mg/kg p.o. In the rat thromboplastin-induced AV shunt model, SSR182289A 10 mg/kg p.o. produced marked antithrombotic effects at 30, 60, 120, and 240 min after administration. Hence, SSR182289A demonstrates potent oral antithrombotic properties in animal venous, AV-shunt, and arterial thrombosis models.

  7. Selecting a Topic in Public Speaking

    Institute of Scientific and Technical Information of China (English)

    刘玉霞

    2013-01-01

    The first step in speechmaking is choosing a topic, which will decide the success of the speech. In the paper, we are go⁃ing to discuss how to select a topic in public speaking, and we’ll mainly talk about the following: the speaker, the occasion and the audience, selecting a topic, specific methods for choosing a topic and available topics.

  8. Topical agents in burn care

    Directory of Open Access Journals (Sweden)

    Momčilović Dragan

    2002-01-01

    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.

  9. Phage display of the serpin alpha-1 proteinase inhibitor randomized at consecutive residues in the reactive centre loop and biopanned with or without thrombin.

    Directory of Open Access Journals (Sweden)

    Benjamin M Scott

    Full Text Available In spite of the power of phage display technology to identify variant proteins with novel properties in large libraries, it has only been previously applied to one member of the serpin superfamily. Here we describe phage display of human alpha-1 proteinase inhibitor (API in a T7 bacteriophage system. API M358R fused to the C-terminus of T7 capsid protein 10B was directly shown to form denaturation-resistant complexes with thrombin by electrophoresis and immunoblotting following exposure of intact phages to thrombin. We therefore developed a biopanning protocol in which thrombin-reactive phages were selected using biotinylated anti-thrombin antibodies and streptavidin-coated magnetic beads. A library consisting of displayed API randomized at residues 357 and 358 (P2-P1 yielded predominantly Pro-Arg at these positions after five rounds of thrombin selection; in contrast the same degree of mock selection yielded only non-functional variants. A more diverse library of API M358R randomized at residues 352-356 (P7-P3 was also probed, yielding numerous variants fitting a loose consensus of DLTVS as judged by sequencing of the inserts of plaque-purified phages. The thrombin-selected sequences were transferred en masse into bacterial expression plasmids, and lysates from individual colonies were screening for API-thrombin complexing. The most active candidates from this sixth round of screening contained DITMA and AAFVS at P7-P3 and inhibited thrombin 2.1-fold more rapidly than API M358R with no change in reaction stoichiometry. Deep sequencing using the Ion Torrent platform confirmed that over 800 sequences were significantly enriched in the thrombin-panned versus naïve phage display library, including some detected using the combined phage display/bacterial lysate screening approach. Our results show that API joins Plasminogen Activator Inhibitor-1 (PAI-1 as a serpin amenable to phage display and suggest the utility of this approach for the selection

  10. Label-free electrochemical aptasensor for sensitive thrombin detection using layer-by-layer self-assembled multilayers with toluidine blue-graphene composites and gold nanoparticles.

    Science.gov (United States)

    Xie, Shunbi; Yuan, Ruo; Chai, Yaqin; Bai, Lijuan; Yuan, Yali; Wang, Yan

    2012-08-30

    In the present study, toluidine blue-graphene (Tb-Gra) nanocomposites were prepared to design a Lable-free electrochemical aptasensor for highly sensitive detection of thrombin based on layer-by-layer (LBL) technology. The nanocomposites with excellent redox electrochemical activities were first immobilized on the gold nanoparticles (nano-Au) modified glassy carbon electrodes (GCE). Then, the LBL structure was performed by electrostatic adsorption between the positively charged Tb-Gra and negatively charged nano-Au, which formed {Tb-Gra/nano-Au}(n) multilayer films for electroactive species enrichment and biomolecule immobilization. Subsequently, the thiolated thrombin binding aptamer (TBA) was assembled on the nano-Au surface through Au-S bond. In the presence of target thrombin (TB), the TBA on the multilayer could catch the thrombin onto the electrode surface, which resulted in a barrier for electro-transfer, leading to the decrease of the electrochemical signal of Tb-Gra nanocomposites. Under the optimal conditions, a wide detection range from 0.001 nM to 80 nM and a low detection limit of 0.33 pM (defined as S/N=3) for thrombin were obtained. In addition, the sensor exhibited excellent selectivity against other proteins. PMID:22939121

  11. Quantum mechanics II advanced topics

    CERN Document Server

    Rajasekar, S

    2015-01-01

    Quantum Mechanics II: Advanced Topics uses more than a decade of research and the authors’ own teaching experience to expound on some of the more advanced topics and current research in quantum mechanics. A follow-up to the authors introductory book Quantum Mechanics I: The Fundamentals, this book begins with a chapter on quantum field theory, and goes on to present basic principles, key features, and applications. It outlines recent quantum technologies and phenomena, and introduces growing topics of interest in quantum mechanics. The authors describe promising applications that include ghost imaging, detection of weak amplitude objects, entangled two-photon microscopy, detection of small displacements, lithography, metrology, and teleportation of optical images. They also present worked-out examples and provide numerous problems at the end of each chapter.

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

    Directory of Open Access Journals (Sweden)

    Ziv-Polat O

    2012-03-01

    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

  13. Acyclic identification of aptamers for human alpha-thrombin using over-represented libraries and deep sequencing.

    Directory of Open Access Journals (Sweden)

    Gillian V Kupakuwana

    Full Text Available BACKGROUND: Aptamers are oligonucleotides that bind proteins and other targets with high affinity and selectivity. Twenty years ago elements of natural selection were adapted to in vitro selection in order to distinguish aptamers among randomized sequence libraries. The primary bottleneck in traditional aptamer discovery is multiple cycles of in vitro evolution. METHODOLOGY/PRINCIPAL FINDINGS: We show that over-representation of sequences in aptamer libraries and deep sequencing enables acyclic identification of aptamers. We demonstrated this by isolating a known family of aptamers for human α-thrombin. Aptamers were found within a library containing an average of 56,000 copies of each possible randomized 15mer segment. The high affinity sequences were counted many times above the background in 2-6 million reads. Clustering analysis of sequences with more than 10 counts distinguished two sequence motifs with candidates at high abundance. Motif I contained the previously observed consensus 15mer, Thb1 (46,000 counts, and related variants with mostly G/T substitutions; secondary analysis showed that affinity for thrombin correlated with abundance (K(d = 12 nM for Thb1. The signal-to-noise ratio for this experiment was roughly 10,000∶1 for Thb1. Motif II was unrelated to Thb1 with the leading candidate (29,000 counts being a novel aptamer against hexose sugars in the storage and elution buffers for Concanavilin A (K(d = 0.5 µM for α-methyl-mannoside; ConA was used to immobilize α-thrombin. CONCLUSIONS/SIGNIFICANCE: Over-representation together with deep sequencing can dramatically shorten the discovery process, distinguish aptamers having a wide range of affinity for the target, allow an exhaustive search of the sequence space within a simplified library, reduce the quantity of the target required, eliminate cycling artifacts, and should allow multiplexing of sequencing experiments and targets.

  14. Pharmacokinetics, pharmacodynamics and food effect of LB30870, a novel direct thrombin inhibitor, after single oral doses in healthy men.

    Science.gov (United States)

    Kim, John; Lee, Sung-Hack; Boyce, Malcolm; Warrington, Steve; Cho, Kwan Hyung; Yoon, Suk Kyoon; Park, Hee Dong; Kim, Aeri

    2015-01-01

    1. The safety, tolerability, pharmacokinetics, pharmacodynamics, and food effect of LB30870, a new selective thrombin inhibitor, were studied in 16 healthy men. 2. A double-blind, placebo-controlled single ascending dose study was done at oral doses of 5, 15, 30, 60, 120, and 240 mg under fasting conditions. An open, randomized, balanced cross-over food effect study was done at 60 mg dose. Plasma and urinary concentrations were measured up to 48 h post-dose. Coagulation and thrombin activity markers were measured at selected time points. 3. Cmax of LB30870 was at 1.3-3.0 h post-dose with a mean apparent terminal half-life (t1/2) of 2.8-4.1 h. AUC after doses above 15 mg appeared greater than dose-proportional. In fed state, AUC showed 80% reduction relative to fasting condition. 4. At doses 60 and 120 mg, peak activated partial thromboplastin time (aPTT) increased by 1.5- and 2-fold, respectively, from baseline. The aPTT and international normalized ratio (INR) were concentration-dependent, with less within-individual variability than ecarin clotting time (ECT), prothrombin time (PT), or thrombin time (TT). 5. Single oral doses of LB30870 up to 240 mg were well tolerated. The food effect must be overcome if LB30870 is to be used as an oral anti-coagulant. PMID:25673087

  15. Topics in current aerosol research

    CERN Document Server

    Hidy, G M

    1971-01-01

    Topics in Current Aerosol Research deals with the fundamental aspects of aerosol science, with emphasis on experiment and theory describing highly dispersed aerosols (HDAs) as well as the dynamics of charged suspensions. Topics covered range from the basic properties of HDAs to their formation and methods of generation; sources of electric charges; interactions between fluid and aerosol particles; and one-dimensional motion of charged cloud of particles. This volume is comprised of 13 chapters and begins with an introduction to the basic properties of HDAs, followed by a discussion on the form

  16. Topics in millimeter wave technology

    CERN Document Server

    Button, Kenneth

    1988-01-01

    Topics in Millimeter Wave Technology, Volume 1 presents topics related to millimeter wave technology, including fin-lines and passive components realized in fin-lines, suspended striplines, suspended substrate microstrips, and modal power exchange in multimode fibers. A miniaturized monopulse assembly constructed in planar waveguide with multimode scalar horn feeds is also described. This volume is comprised of five chapters; the first of which deals with the analysis and synthesis techniques for fin-lines as well as the various passive components realized in fin-line. Tapers, discontinuities,

  17. 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 β-decay; semiclassical approach of large amplitude collective motion and structure of hot nuclei

  18. Challenges of the management of severe hemophilia A with inhibitors: two case reports emphasizing the potential interest of a high-purity human Factor VIII/von Willebrand factor concentrate and individually tailored prophylaxis guided by thrombin-generation test.

    Science.gov (United States)

    Mathieu, Sophie; Crampe, Carine; Dargaud, Yesim; Lavigne-Lissalde, Géraldine; Escuriola-Ettingshausen, Carmen; Tardy, Brigitte; Meley, Roland; Thouvenin, Sandrine; Stephan, Jean L; Berger, Claire

    2015-12-01

    Severe hemophilia A is an X-linked bleeding disorder. Immune tolerance induction (ITI) is the best strategy of treatment when patients develop inhibitors. The objective is to illustrate the benefit of a high-purity human factor VIII/von Willebrand factor (VWF) concentrate (Octanate) in the management of ITI. We also wanted to raise the potential interest of laboratory assays such as thrombin-generation test (TGT) and epitope mapping. Two patients were treated during ITI, first with a recombinant FVIII and then with plasma-derived factor VIII without success, and, finally, with Octanate. Bypassing agents were used based on the results of TGT. Epitope mapping was performed during ITI therapy. These observations suggest the potential contribution of Octanate in the management of ITI in difficult cases. The use of bypassing agents can be necessary in prophylaxis or to treat bleedings, and may be guided by TGT results. Epitope mapping is used to describe the inhibitor. This article shows a decrease of the inhibitor directed against the C2 domain after initiation of Octanate. A high-purity human factor VIII/von Willebrand factor concentrate (Octanate) may be a valuable therapeutical option for ITI therapy. TGT and epitope mapping could be of help in the management of ITI. PMID:26517064

  19. Challenges of the management of severe hemophilia A with inhibitors: two case reports emphasizing the potential interest of a high-purity human Factor VIII/von Willebrand factor concentrate and individually tailored prophylaxis guided by thrombin-generation test.

    Science.gov (United States)

    Mathieu, Sophie; Crampe, Carine; Dargaud, Yesim; Lavigne-Lissalde, Géraldine; Escuriola-Ettingshausen, Carmen; Tardy, Brigitte; Meley, Roland; Thouvenin, Sandrine; Stephan, Jean L; Berger, Claire

    2015-12-01

    Severe hemophilia A is an X-linked bleeding disorder. Immune tolerance induction (ITI) is the best strategy of treatment when patients develop inhibitors. The objective is to illustrate the benefit of a high-purity human factor VIII/von Willebrand factor (VWF) concentrate (Octanate) in the management of ITI. We also wanted to raise the potential interest of laboratory assays such as thrombin-generation test (TGT) and epitope mapping. Two patients were treated during ITI, first with a recombinant FVIII and then with plasma-derived factor VIII without success, and, finally, with Octanate. Bypassing agents were used based on the results of TGT. Epitope mapping was performed during ITI therapy. These observations suggest the potential contribution of Octanate in the management of ITI in difficult cases. The use of bypassing agents can be necessary in prophylaxis or to treat bleedings, and may be guided by TGT results. Epitope mapping is used to describe the inhibitor. This article shows a decrease of the inhibitor directed against the C2 domain after initiation of Octanate. A high-purity human factor VIII/von Willebrand factor concentrate (Octanate) may be a valuable therapeutical option for ITI therapy. TGT and epitope mapping could be of help in the management of ITI.

  20. Effects of thrombin inhibition with melagatran on renal hemodynamics and function and liver integrity during early endotoxemia

    DEFF Research Database (Denmark)

    Nitescu, Nicoletta; Grimberg, Elisabeth; Ricksten, Sven-Erik;

    2007-01-01

    . Kidney function, renal blood flow (RBF), and intrarenal cortical and outer medullary perfusion (OMLDF) measured by laser-Doppler flowmetry were analyzed throughout. Markers of liver injury and tumor necrosis factor (TNF)-alpha were measured in plasma after 4.5 h of endotoxemia. In addition, liver...... histology and gene expression were examined. Melagatran treatment prevented the decline in OMLDF observed in the LPS-Saline group (P .... Melagatran did not diminish histological abnormalities in the liver or the elevated hepatic gene expression of TNF-alpha, intercellular adhesion molecule-1, and inducible nitric oxide synthase in endotoxemic rats. In summary, thrombin inhibition with melagatran preserved renal OMLDF, attenuated liver...

  1. In vitro assessment of Tc-99m labeled bovine thrombin and streptokinase-activated human plasmin: concise communication. [Iodine 125

    Energy Technology Data Exchange (ETDEWEB)

    Wong, D.W.; Tanaka, T.; Mishkin, F.; Lee, T.

    1979-09-01

    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.

  2. Prospective evaluation of hemostatic system activation and thrombin potential in healthy pregnant women with and without factor V Leiden.

    Science.gov (United States)

    Eichinger, S; Weltermann, A; Philipp, K; Hafner, E; Kaider, A; Kittl, E M; Brenner, B; Mannhalter, C; Lechner, K; Kyrle, P A

    1999-10-01

    Normal pregnancy is associated with alterations of the hemostatic system towards a hypercoagulable state and an increased risk of venous thromboembolism. The risk of venous thrombosis is higher in pregnant women with factor V Leiden (FVL) than in those with wildtype factor V. Routine laboratory assays are not useful to detect hypercoagulable conditions. A prospective and systematic evaluation of hemostatic system activation in women with and without FVL during an uncomplicated pregnancy employing more sensitive markers of hypercoagulability, such as prothrombin fragment 1+2 (F1+2), thrombin-antithrombin complex (TAT), D-Dimer, or the endogenous thrombin potential (ETP), an indicator of the plasma's potential to generate thrombin, has not been performed. We prospectively followed 113 pregnant women with (n = 11) and without (n = 102) FVL and measured F1+2. TAT, D-Dimer and the ETP at the 12th, 22nd and 34th gestational week as well as 3 months after delivery (baseline) in each subject. None of the women developed clinical signs of venous thromboembolism during pregnancy or postpartum. Pregnant women with and without FVL exhibited substantial activation of the coagulation and fibrinolytic system as indicated by a gradual increase of F1+2, TAT and D-Dimer throughout uncomplicated pregnancy up to levels similar to those found in acute thromboembolic events (p < 0.0001 by analysis of variance for each parameters). Levels of F1+2 and TAT were comparable between women with and without FVL, but levels of D-Dimer were significantly higher in women with FVL than in those without the mutation (p = 0.0005). The ETP remained unchanged in both women with and without FVL at all timepoints. Our data demonstrate a substantial coagulation and fibrinolytic system activation in healthy women with and without FVL during uncomplicated pregnancy. An elevated F1+2, TAT or D-Dimer level during pregnancy is not necessarily indicative for an acute thromboembolic event. The normal ETP in both

  3. Differential effects of formoterol on thrombin- and PDGF-induced proliferation of human pulmonary arterial vascular smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Goncharova Elena A

    2012-11-01

    Full Text Available Abstract Background Increased pulmonary arterial vascular smooth muscle (PAVSM cell proliferation is a key pathophysiological component of pulmonary vascular remodeling in pulmonary arterial hypertension (PH. The long-acting β2-adrenergic receptor (β2AR agonist formoterol, a racemate comprised of (R,R- and (S,S-enantiomers, is commonly used as a vasodilator in chronic obstructive pulmonary disease (COPD. PH, a common complication of COPD, increases patients’ morbidity and reduces survival. Recent studies demonstrate that formoterol has anti-proliferative effects on airway smooth muscle cells and bronchial fibroblasts. The effects of formoterol and its enantiomers on PAVSM cell proliferation are not determined. The goals of this study were to examine effects of racemic formoterol and its enantiomers on PAVSM cell proliferation as it relates to COPD-associated PH. Methods Basal, thrombin-, PDGF- and chronic hypoxia-induced proliferation of primary human PAVSM cells was examined by DNA synthesis analysis using BrdU incorporation assay. ERK1/2, mTORC1 and mTORC2 activation were determined by phosphorylation levels of ERK1/2, ribosomal protein S6 and S473-Akt using immunoblot analysis. Results We found that (R,R and racemic formoterol inhibited basal, thrombin- and chronic hypoxia-induced proliferation of human PAVSM cells while (S,S formoterol had lesser inhibitory effect. The β2AR blocker propranolol abrogated the growth inhibitory effect of formoterol. (R,R, but not (S,S formoterol attenuated basal, thrombin- and chronic hypoxia-induced ERK1/2 phosphorylation, but had little effect on Akt and S6 phosphorylation levels. Formoterol and its enantiomers did not significantly affect PDGF-induced DNA synthesis and PDGF-dependent ERK1/2, S473-Akt and S6 phosphorylation in human PAVSM cells. Conclusions Formoterol inhibits basal, thrombin-, and chronic hypoxia-, but not PDGF-induced human PAVSM cell proliferation and ERK1/2, but has little effect on

  4. International Normalized Ratio (INR), coagulation factor activities and calibrated automated thrombin generation - influence of 24 h storage at ambient temperature

    DEFF Research Database (Denmark)

    Christensen, T D; Jensen, C; Larsen, T B;

    2010-01-01

    clotting activity of coagulation factors II, VII, IX, and X as well as CAT generation was recorded after 0 and 24 h respectively. Statistical analyses included Bland-Altman plot, 95% limits of agreement, and a variability test using a mixed effect model. The level of INR remained statistically unchanged......International Normalized Ratio (INR) measurements are used to monitor oral anticoagulation therapy with coumarins. Single coagulation factor activities and calibrated automated thrombin (CAT) generation are considered as more advanced methods for evaluating overall haemostatic capacity. The aims...

  5. Separation and Purification of Thrombin-like Enzymes by Affinity Adsorbents

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    An affinity adsorbent, benzamidineSepharose 4B, was used to separate and purify thrombinlike enzymes. The paminobenzamidine as a specific ligand was coupled to the matrix-Sepharose 4B. The recombinant thrombinlike enzyme-defibrase was used as a model in order to evaluate the efficiency of this biospecific affinity adsorbent. The homogeneity of the enzyme preparation was comfirmed as one band on sodium dodecyl sulfatepolyacrylamide gel electrophoresis.

  6. The Health Curriculum: 500 Topics.

    Science.gov (United States)

    Byrd, Oliver E.

    2001-01-01

    This 1958 paper divides 500 health topics into 20 categories: health as a social accomplishment/social problem; nutrition; physical fitness; mental health and disease; heredity/eugenics; infection/immunity; chronic and degenerative disease; substance abuse; skin care; vision, hearing, and speech; dental health; safety; physical environment; health…

  7. 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.)

  8. Topics in optics and music

    Science.gov (United States)

    Sparks, Andrew W.

    2012-10-01

    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.

  9. Effect of NaC1 on inactivation of bovine thrombin by antithrombin III in the presence of low affinity-heparin or dextran sulfate.

    Science.gov (United States)

    Oshima, G; Nagasawa, K

    1986-02-01

    Heparin with low affinity (LA-heparin) to antithrombin III (AT III) enhanced the rate of inactivation of thrombin by AT III. The enhancement of the rate was saturable with AT III and was proportional to the LA-heparin concentration. Although the rate-enhancement in the presence of LA-heparin decreased with increase in NaC1 concentration, it was comparable with that in the presence of high affinity-heparin (HA-heparin) in the absence of NaC1. Inactivation of thrombin by AT III in the presence of dextran sulfate (DS) was also sensitive to NaC1 concentration. These findings indicate that free AT III is favorable for binding to the complexes of thrombin and highly sulfated polysaccharides having low affinities to AT III in the absence of NaC1.

  10. Primordial magnetogenesis before recombination

    Science.gov (United States)

    Fabre, Ophélia; Shankaranarayanan, S.

    2016-04-01

    The origin of large magnetic fields in the Universe remains currently unknown. We investigate here a mechanism before recombination based on known physics. The source of the vorticity is due to the changes in the photon distribution function caused by the fluctuations in the background photons. We show that the magnetic field generated in the MHD limit, due to the Coulomb scattering, is of the order 10-49 G on a coherence scale of 10 kpc. We explicitly show that the magnetic fields generated from this process are sustainable and are not erased by resistive diffusion. We compare the results with current observations and discuss the implications. Our seed magnetic fields are generated on small scales whereas the main mechanisms studied in the literature are on scale bigger than 1 Mpc. However, compared to more exotic theories generating seed magnetic fields on similar scales, the strength of our fields are generally smaller.

  11. The pharmacology of topical analgesics.

    Science.gov (United States)

    Barkin, Robert L

    2013-07-01

    Pain management of patients continues to pose challenges to clinicians. Given the multiple dimensions of pain--whether acute or chronic, mild, moderate, or severe, nociceptive or neuropathic--a multimodal approach may be needed. Fortunately, clinicians have an array of nonpharmacologic and pharmacologic treatment choices; however, each modality must be chosen carefully, because some often used oral agents are associated with safety and tolerability issues that restrict their use in certain patients. In particular, orally administered nonsteroidal antiinflammatory drugs, opioids, antidepressants, and anticonvulsants are known to cause systemic adverse effects in some patients. To address this problem, a number of topical therapies in various therapeutic classes have been developed to reduce systemic exposure and minimize the risks of patients developing adverse events. For example, topical nonsteroidal anti-inflammatory drug formulations produce a site-specific effect (ie, cyclo-oxygenase inhibition) while decreasing the systemic exposure that may lead to undesired effects in patients. Similarly, derivatives of acetylsalicylic acid (ie, salicylates) are used in topical analgesic formulations that do not significantly enter the patient's systemic circulation. Salicylates, along with capsaicin, menthol, and camphor, compose the counterirritant class of topical analgesics, which produce analgesia by activating and then desensitizing epidermal nociceptors. Additionally, patches and creams that contain the local anesthetic lidocaine, alone or co-formulated with other local anesthetics, are also used to manage patients with select acute and chronic pain states. Perhaps the most common topical analgesic modality is the cautious application of cutaneous cold and heat. Such treatments may decrease pain not by reaching the target tissue through systemic distribution, but by acting more directly on the affected tissue. Despite the tolerability benefits associated with avoiding

  12. Soluble expression of Catalase/GST fusion protein of helicobacter pylori and thrombin cleavage of GST tag on column%幽门螺杆菌Catalas/GST融合蛋白的可溶性表达及凝血酶柱上切割GST标签

    Institute of Scientific and Technical Information of China (English)

    李妍; 姜茵; 奚月; 何殿殿; 宁云山

    2012-01-01

    目的 利用GST融合基因表达系统表达幽门螺杆菌Catalase/GST融合蛋白,并利用凝血酶柱上切割GST标签.方法 将重组表达质粒Catalase/pGEX-4T-1转化大肠杆菌BL21 (DE3)感受态中并用IPTG进行诱导表达,菌体经反复冻融、溶菌酶裂解及超声破菌进行裂解.采用谷胱甘肽琼脂糖树脂Glutathione Sepharose 4B对Catalase/GST融合蛋白可溶性表达上清进行纯化,并同时利用凝血酶柱上切割GST标签,用鼠抗Catalase抗体对纯化产物进行Western blot鉴定.结果 高效表达出相对分子质量约85 kDa的Catalase/GST融合蛋白,以部分可溶性的形式表达,凝血酶柱上成功地切割了GST标签,Catalase蛋白能被鼠抗Catalase单克隆抗体识别.结论 成功表达了Catalase/GST融合蛋白并柱上切割了GST标签,为深入研究Catalase的功能奠定了基础.%OBJECTIVE To express Catalase/GST fusion protein by GST gene expression system and cleave GST-tag on column using thrombin. METHODS The recombinant expression plasmid Catalase/pGEX4T-l was transformed into E. Coli BL21 (DE3) component cell and induced by IPTG. The bacterial sediment was lysed by repeating freezing and thawing, lysozyme lysis and ultrasonication. Catalase/GST fusion protein was purified by Glutathione Sepharose 4B and cleaved the GST-tag on column using thrombin. Purified Catalase was identified by anti-Catalase monoclonal antibody (mAb) using Western blot. RESULTS The fusion Catalase/GST was partly expressed in soluble form with relative molecular mass of 85kDa. Thrombin cleaved the GST tag on column and Catalase protein was recognized by mouse anti-Catalase mAb. CONCLUSION The recombinant Catalase/GST is successfully expressed and GST-tag is cleaved on column, which lays a foundation for further study on function of Catalase protein.

  13. Cell biology of mitotic recombination

    DEFF Research Database (Denmark)

    Lisby, Michael; Rothstein, Rodney

    2015-01-01

    Homologous recombination provides high-fidelity DNA repair throughout all domains of life. Live cell fluorescence microscopy offers the opportunity to image individual recombination events in real time providing insight into the in vivo biochemistry of the involved proteins and DNA molecules as w...

  14. The phosphorylation status of extracellular-regulated kinase 1/2 in astrocytes and neurons from rat hippocampus determines the thrombin-induced calcium release and ROS generation.

    Science.gov (United States)

    Zündorf, Gregor; Reiser, Georg

    2011-12-01

    Challenge of protease-activated receptors induces cytosolic Ca(2+) concentration ([Ca(2+) ](c)) increase, mitogen-activated protein kinase activation and reactive oxygen species (ROS) formation with a bandwidth of responses in individual cells. We detected in this study in situ the thrombin-induced [Ca(2+) ](c) rise and ROS formation in dissociated hippocampal astrocytes and neurons in a mixed culture. In identified cells, single cell responses were correlated with extracellular-regulated kinase (ERK)1/2 phosphorylation level. On average, in astrocytes, thrombin induced a transient [Ca(2+) ](c) rise with concentration-dependent increase in amplitude and extrusion rate and high ERK1/2 phosphorylation level. Correlation analysis of [Ca(2+) ](c) response characteristics of single astrocytes reveals that astrocytes with nuclear phosphoERK1/2 localization have a smaller Ca(2+) amplitude and extrusion rate compared with cells with a cytosolic phosphoERK1/2 localization. In naive neurons, without thrombin challenge, variable ERK1/2 phosphorylation patterns are observed. ROS were detected by hydroethidine. Only in neurons with increased ERK1/2 phosphorylation level, we see sustained intracellular rise in fluorescence of the dye lasting over several minutes. ROS formation was abolished by pre-incubation with the NADPH oxidase inhibitor apocynin. Additionally, thrombin induced an immediate, transient hydroethidine fluorescence increase. This was interpreted as NADPH oxidase-mediated O(2) (•-) -release into the extracellular milieu, because it was decreased by pre-incubation with apocynin, and could be eluted by superfusion. In conclusion, the phosphorylation status of ERK1/2 determines the thrombin-dependent [Ca(2+) ](c) increase and ROS formation and, thus, influences the capacity of thrombin to regulate neuroprotection or neurodegeneration. PMID:21988180

  15. On the specificity of heparin/heparan sulfate binding to proteins. Anion-binding sites on antithrombin and thrombin are fundamentally different.

    Directory of Open Access Journals (Sweden)

    Philip D Mosier

    Full Text Available BACKGROUND: The antithrombin-heparin/heparan sulfate (H/HS and thrombin-H/HS interactions are recognized as prototypic specific and non-specific glycosaminoglycan (GAG-protein interactions, respectively. The fundamental structural basis for the origin of specificity, or lack thereof, in these interactions remains unclear. The availability of multiple co-crystal structures facilitates a structural analysis that challenges the long-held belief that the GAG binding sites in antithrombin and thrombin are essentially similar with high solvent exposure and shallow surface characteristics. METHODOLOGY: Analyses of solvent accessibility and exposed surface areas, gyrational mobility, symmetry, cavity shape/size, conserved water molecules and crystallographic parameters were performed for 12 X-ray structures, which include 12 thrombin and 16 antithrombin chains. Novel calculations are described for gyrational mobility and prediction of water loci and conservation. RESULTS: The solvent accessibilities and gyrational mobilities of arginines and lysines in the binding sites of the two proteins reveal sharp contrasts. The distribution of positive charges shows considerable asymmetry in antithrombin, but substantial symmetry for thrombin. Cavity analyses suggest the presence of a reasonably sized bifurcated cavity in antithrombin that facilitates a firm 'hand-shake' with H/HS, but with thrombin, a weaker 'high-five'. Tightly bound water molecules were predicted to be localized in the pentasaccharide binding pocket of antithrombin, but absent in thrombin. Together, these differences in the binding sites explain the major H/HS recognition characteristics of the two prototypic proteins, thus affording an explanation of the specificity of binding. This provides a foundation for understanding specificity of interaction at an atomic level, which will greatly aid the design of natural or synthetic H/HS sequences that target proteins in a specific manner.

  16. Bimolecular recombination in organic photovoltaics.

    Science.gov (United States)

    Lakhwani, Girish; Rao, Akshay; Friend, Richard H

    2014-01-01

    The recombination of electrons and holes is a major loss mechanism in photovoltaic devices that controls their performance. We review scientific literature on bimolecular recombination (BR) in bulk heterojunction organic photovoltaic devices to bring forward existing ideas on the origin and nature of BR and highlight both experimental and theoretical work done to quantify its extent. For these systems, Langevin theory fails to explain BR, and recombination dynamics turns out to be dependent on mobility, temperature, electric field, charge carrier concentration, and trapped charges. Relationships among the photocurrent, open-circuit voltage, fill factor, and morphology are discussed. Finally, we highlight the recent emergence of a molecular-level picture of recombination, taking into account the spin and delocalization of charges. Together with the macroscopic picture of recombination, these new insights allow for a comprehensive understanding of BR and provide design principles for future materials and devices.

  17. Monitoring low molecular weight heparins at therapeutic levels: dose-responses of, and correlations and differences between aPTT, anti-factor Xa and thrombin generation assays.

    Directory of Open Access Journals (Sweden)

    Owain Thomas

    Full Text Available Low molecular weight heparins (LMWH's are used to prevent and treat thrombosis. Tests for monitoring LMWH's include anti-factor Xa (anti-FXa, activated partial thromboplastin time (aPTT and thrombin generation. Anti-FXa is the current gold standard despite LMWH's varying affinities for FXa and thrombin.To examine the effects of two different LMWH's on the results of 4 different aPTT-tests, anti-FXa activity and thrombin generation and to assess the tests' concordance.Enoxaparin and tinzaparin were added ex-vivo in concentrations of 0.0, 0.5, 1.0 and 1.5 anti-FXa international units (IU/mL, to blood from 10 volunteers. aPTT was measured using two whole blood methods (Free oscillation rheometry (FOR and Hemochron Jr (HCJ and an optical plasma method using two different reagents (ActinFSL and PTT-Automat. Anti-FXa activity was quantified using a chromogenic assay. Thrombin generation (Endogenous Thrombin Potential, ETP was measured on a Ceveron Alpha instrument using the TGA RB and more tissue-factor rich TGA RC reagents.Methods' mean aPTT at 1.0 IU/mL LMWH varied between 54s (SD 11 and 69s (SD 14 for enoxaparin and between 101s (SD 21 and 140s (SD 28 for tinzaparin. ActinFSL gave significantly shorter aPTT results. aPTT and anti-FXa generally correlated well. ETP as measured with the TGA RC reagent but not the TGA RB reagent showed an inverse exponential relationship to the concentration of LMWH. The HCJ-aPTT results had the weakest correlation to anti-FXa and thrombin generation (Rs0.62-0.87, whereas the other aPTT methods had similar correlation coefficients (Rs0.80-0.92.aPTT displays a linear dose-response to LMWH. There is variation between aPTT assays. Tinzaparin increases aPTT and decreases thrombin generation more than enoxaparin at any given level of anti-FXa activity, casting doubt on anti-FXa's present gold standard status. Thrombin generation with tissue factor-rich activator is a promising method for monitoring LMWH's.

  18. Association of thrombin generation potential with platelet PAR-1 regulation and P-selectin expression in patients on dual antiplatelet therapy.

    Science.gov (United States)

    Badr Eslam, Roza; Posch, Florian; Lang, Irene M; Gremmel, Thomas; Eichelberger, Beate; Ay, Cihan; Panzer, Simon

    2014-02-01

    We studied the association of thrombin generation potential with platelet protease activated receptor (PAR)-1 regulation and platelet activation in 52 stable coronary artery disease patients on continuous therapy with aspirin and clopidogrel (n = 42) or prasugrel (n = 10). Compared to controls, peak thrombin generation potential was elevated in only 11 patients (p > 0.05), while F1.2 was elevated in 26 patients (p P-selectin expression were significantly elevated in patients compared to controls (p P-selectin (p = 0.002), suggesting in vivo depletion of platelet alpha granules due to ongoing platelet activation.

  19. A label-free electrochemical aptasensor based on the catalysis of manganese porphyrins for detection of thrombin.

    Science.gov (United States)

    Zheng, Yingning; Yuan, Yali; Chai, Yaqin; Yuan, Ruo

    2015-04-15

    A novel manganese porphyrin (MnPP)-catalyzed aerobic oxidation of l-cysteine to disulfides (RSSR) was firstly found and applied into electrochemical aptasensor with a label-free technique for signal amplification. The possible catalytic mechanism of the catalytic reaction where MnPP catalyzed l-cysteine with thiol (RSH) structure to RSSR was discussed in detail. For fabrication of the aptasensor, thionine (Thi), which served as an electron mediator, was mixed with MnPP and immobilized on the nafion coated carbon electrode through ion exchange adsorption. Gold nanoparticle (nano-Au) was assembled on the Thi for immobilizing thrombin binding aptamer (TBA). In the presence of thrombin (TB), TBA will capture TB and form TBA-TB composite thus perturbed electron transfer, leading to decrease of the current for quantitatively detecting TB. Under optimal condition, the electrochemical aptasensor exhibited a linear range of 0.1-25nM with a detection limit of 0.02nM. This work opens a novel way for signal amplification study about porphyrins that served as mimetic enzyme to thiol in electrochemical aptasensor. PMID:25530538

  20. Influenza virus H1N1 activates platelets through FcγRIIA signaling and thrombin generation.

    Science.gov (United States)

    Boilard, Eric; Paré, Guillaume; Rousseau, Matthieu; Cloutier, Nathalie; Dubuc, Isabelle; Lévesque, Tania; Borgeat, Pierre; Flamand, Louis

    2014-05-01

    Platelets play crucial functions in hemostasis and the prevention of bleeding. During H1N1 influenza A virus infection, platelets display activation markers. The platelet activation triggers during H1N1 infection remain elusive. We observed that H1N1 induces surface receptor activation, lipid mediator synthesis, and release of microparticles from platelets. These activation processes require the presence of serum/plasma, pointing to the contribution of soluble factor(s). Considering that immune complexes in the H1N1 pandemic were reported to play a pathogenic role, we assessed their contribution in H1N1-induced platelet activation. In influenza-immunized subjects, we observed that the virus scaffolds with immunoglobulin G (IgG) to form immune complexes that promote platelet activation. Mechanistically, this activation occurs through stimulation of low-affinity type 2 receptor for Fc portion of IgG (FcγRIIA), a receptor for immune complexes, independently of thrombin. Using a combination of in vitro and in vivo approaches, we found that the antibodies from H3N2-immunized mice activate transgenic mouse platelets that express FcγRIIA when put in the presence of H1N1, suggesting that cross-reacting influenza antibodies suffice. Alternatively, H1N1 can activate platelets via thrombin formation, independently of complement and FcγRIIA. These observations identify both the adaptive immune response and the innate response against pathogens as 2 intertwined processes that activate platelets during influenza infections.

  1. 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)

    A.V Pérez

    2008-01-01

    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.

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

    Science.gov (United States)

    Ziv-Polat, Ofra; Skaat, Hadas; Shahar, Abraham; Margel, Shlomo

    2012-01-01

    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. PMID:22419873

  3. Do scientists trace hot topics?

    CERN Document Server

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

    2013-01-01

    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.

  4. Four (Somewhat Nonstandard) Research Topics

    OpenAIRE

    Paun, Gheorghe; Research Group on Natural Computing (Universidad de Sevilla) (Coordinador)

    2014-01-01

    Four research directions are suggested, dealing with the following four main ideas: computing along the axon (up to now, this topic was only preliminarily investi- gated), using pre-computed resources in order to solve computationally hard problems, considering in P systems both objects \\of matter" and \\of anti-matter" (which annihilate each other when meet), and considering the distance (naturally de ned in a membrane structure of a given type) as a support of information.

  5. Preface to Special Topic: Optofluidics

    OpenAIRE

    Liu, Ai-qun

    2010-01-01

    This Special Topic section of Biomicrofluidics is on optofluidics or micro-optofluidic systems (MOFS), a burgeoning technology that aims to manipulate light and fluid at microscale and exploits their interaction to create highly versatile devices and integrated systems. This special issue puts together various contributed articles focusing on optofluidics or MOFS, which help inspire new research ideas and innovation in the microfluidics and nanofluidics community.

  6. Probabilistic analysis and related topics

    CERN Document Server

    Bharucha-Reid, A T

    1983-01-01

    Probabilistic Analysis and Related Topics, Volume 3 focuses on the continuity, integrability, and differentiability of random functions, including operator theory, measure theory, and functional and numerical analysis. The selection first offers information on the qualitative theory of stochastic systems and Langevin equations with multiplicative noise. Discussions focus on phase-space evolution via direct integration, phase-space evolution, linear and nonlinear systems, linearization, and generalizations. The text then ponders on the stability theory of stochastic difference systems and Marko

  7. 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.)

  8. Hot topics from the Tevatron

    Energy Technology Data Exchange (ETDEWEB)

    Glenzinski, D.; /Fermilab

    2008-01-01

    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.

  9. Quantification of thymidine kinase (TK1) mRNA in normal and leukemic cells and investigation of structure-function relatiosnhip of recombinant TK1enzyme

    DEFF Research Database (Denmark)

    Kristensen, Tina

    lymphocytes. As the high TKI mRNA level is not translated into an active enzyme, these results indicate a defect in the regulation of TKI in CLL cells. For the studies of the structure-function relationship of TKI a recombinant TKI protein, which is expressed as a glutathione-S-transferase (GST) fusion...... protein was used. TKI protein is cleaved from the GST-part with thrombin. Two TKI mutants, TKI-l 93 and TKI-l 76, with deletions from the C-terminal were constructed by the recombinant PCR method. Deletion of 57 amino acids from the C-terminal (TKI- 176) results in an inactive enzyme. Deletion of 40 amino...

  10. Analysis of interchromosomal mitotic recombination.

    Science.gov (United States)

    McGill, C B; Shafer, B K; Higgins, D R; Strathern, J N

    1990-07-01

    A novel synthetic locus is described that provides a simple assay system for characterizing mitotic recombinants. The locus consists of the TRP1 and HIS3 genes inserted into chromosome III of S. cerevisiae between the CRY1 and MAT loci. Defined trp1 and his3 alleles have been generated that allow the selection of interchromosomal recombinants in this interval. Trp+ or His+ recombinants can be divided into several classes based on coupling of the other alleles in the interval. The tight linkage of the CRY1 and MAT loci, combined with the drug resistance and cell type phenotypes that they respectively control, facilitates the classification of the recombinants without resorting to tetrad dissection. We present the distribution of spontaneous recombinants among the classes defined by this analysis. The data suggest that the recombination intermediate can have regions of symmetric strand exchange and that co-conversion tracts can extend over 1-3 kb. Continuous conversion tracts are favored over discontinuous tracts. The distribution among the classes defined by this analysis is altered in recombinants induced by UV irradiation.

  11. Aspergillus: sex and recombination.

    Science.gov (United States)

    Varga, János; Szigeti, Gyöngyi; Baranyi, Nikolett; Kocsubé, Sándor; O'Gorman, Céline M; Dyer, Paul S

    2014-12-01

    The genus Aspergillus is one of the most widespread groups of fungi on Earth, comprised of about 300-350 species with very diverse lifestyles. Most species produce asexual propagula (conidia) on conidial heads. Despite their ubiquity, a sexual cycle has not yet been identified for most of the aspergilli. Where sexual reproduction is present, species exhibit either homothallic (self fertile) or heterothallic (obligate outcrossing) breeding systems. A parasexual cycle has also been described in some Aspergillus species. As in other fungi, sexual reproduction is governed by mating-type (MAT) genes, which determine sexual identity and are involved in regulating later stages of sexual development. Previous population genetic studies have indicated that some supposedly asexual aspergilli exhibit evidence of a recombining population structure, suggesting the presence of a cryptic sexual cycle. In addition, genome analyses have revealed networks of genes necessary for sexual reproduction in several Aspergillus species, again consistent with latent sexuality in these fungi. Knowledge of MAT gene presence has then successfully been applied to induce sexual reproduction between MAT1-1 and MAT1-2 isolates of certain supposedly asexual aspergilli. Recent progress in understanding the extent and significance of sexual reproduction is described here, with special emphasis on findings that are relevant to clinically important aspergilli. PMID:25118872

  12. Rational Design of a Carrier Protein for the Production of Recombinant Toxic Peptides in Escherichia coli

    Science.gov (United States)

    Pizzo, Elio; Varcamonti, Mario; Zanfardino, Anna; Sgambati, Valeria; Di Maro, Antimo; Carpentieri, Andrea; Izzo, Viviana; Di Donato, Alberto; Cafaro, Valeria; Notomista, Eugenio

    2016-01-01

    Commercial uses of bioactive peptides require low cost, effective methods for their production. We developed a new carrier protein for high yield production of recombinant peptides in Escherichia coli very well suited for the production of toxic peptides like antimicrobial peptides. GKY20, a short antimicrobial peptide derived from the C-terminus of human thrombin, was fused to the C-terminus of Onconase, a small ribonuclease (104 amino acids), which efficiently drove the peptide into inclusion bodies with very high expression levels (about 200–250 mg/L). After purification of the fusion protein by immobilized metal ion affinity chromatography, peptide was obtained by chemical cleavage in diluted acetic acid of an acid labile Asp-Pro sequence with more than 95% efficiency. To improve peptide purification, Onconase was mutated to eliminate all acid labile sequences thus reducing the release of unwanted peptides during the acid cleavage. Mutations were chosen to preserve the differential solubility of Onconase as function of pH, which allows its selective precipitation at neutral pH after the cleavage. The improved carrier allowed the production of 15–18 mg of recombinant peptide per liter of culture with 96–98% purity without the need of further chromatographic steps after the acid cleavage. The antimicrobial activity of the recombinant peptide, with an additional proline at the N-terminus, was tested on Gram-negative and Gram-positive strains and was found to be identical to that measured for synthetic GKY20. This finding suggests that N-terminal proline residue does not change the antimicrobial properties of recombinant (P)GKY20. The improved carrier, which does not contain cysteine and methionine residues, Asp-Pro and Asn-Gly sequences, is well suited for the production of peptides using any of the most popular chemical cleavage methods. PMID:26808536

  13. Rational Design of a Carrier Protein for the Production of Recombinant Toxic Peptides in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Katia Pane

    Full Text Available Commercial uses of bioactive peptides require low cost, effective methods for their production. We developed a new carrier protein for high yield production of recombinant peptides in Escherichia coli very well suited for the production of toxic peptides like antimicrobial peptides. GKY20, a short antimicrobial peptide derived from the C-terminus of human thrombin, was fused to the C-terminus of Onconase, a small ribonuclease (104 amino acids, which efficiently drove the peptide into inclusion bodies with very high expression levels (about 200-250 mg/L. After purification of the fusion protein by immobilized metal ion affinity chromatography, peptide was obtained by chemical cleavage in diluted acetic acid of an acid labile Asp-Pro sequence with more than 95% efficiency. To improve peptide purification, Onconase was mutated to eliminate all acid labile sequences thus reducing the release of unwanted peptides during the acid cleavage. Mutations were chosen to preserve the differential solubility of Onconase as function of pH, which allows its selective precipitation at neutral pH after the cleavage. The improved carrier allowed the production of 15-18 mg of recombinant peptide per liter of culture with 96-98% purity without the need of further chromatographic steps after the acid cleavage. The antimicrobial activity of the recombinant peptide, with an additional proline at the N-terminus, was tested on Gram-negative and Gram-positive strains and was found to be identical to that measured for synthetic GKY20. This finding suggests that N-terminal proline residue does not change the antimicrobial properties of recombinant (PGKY20. The improved carrier, which does not contain cysteine and methionine residues, Asp-Pro and Asn-Gly sequences, is well suited for the production of peptides using any of the most popular chemical cleavage methods.

  14. Rational Design of a Carrier Protein for the Production of Recombinant Toxic Peptides in Escherichia coli.

    Science.gov (United States)

    Pane, Katia; Durante, Lorenzo; Pizzo, Elio; Varcamonti, Mario; Zanfardino, Anna; Sgambati, Valeria; Di Maro, Antimo; Carpentieri, Andrea; Izzo, Viviana; Di Donato, Alberto; Cafaro, Valeria; Notomista, Eugenio

    2016-01-01

    Commercial uses of bioactive peptides require low cost, effective methods for their production. We developed a new carrier protein for high yield production of recombinant peptides in Escherichia coli very well suited for the production of toxic peptides like antimicrobial peptides. GKY20, a short antimicrobial peptide derived from the C-terminus of human thrombin, was fused to the C-terminus of Onconase, a small ribonuclease (104 amino acids), which efficiently drove the peptide into inclusion bodies with very high expression levels (about 200-250 mg/L). After purification of the fusion protein by immobilized metal ion affinity chromatography, peptide was obtained by chemical cleavage in diluted acetic acid of an acid labile Asp-Pro sequence with more than 95% efficiency. To improve peptide purification, Onconase was mutated to eliminate all acid labile sequences thus reducing the release of unwanted peptides during the acid cleavage. Mutations were chosen to preserve the differential solubility of Onconase as function of pH, which allows its selective precipitation at neutral pH after the cleavage. The improved carrier allowed the production of 15-18 mg of recombinant peptide per liter of culture with 96-98% purity without the need of further chromatographic steps after the acid cleavage. The antimicrobial activity of the recombinant peptide, with an additional proline at the N-terminus, was tested on Gram-negative and Gram-positive strains and was found to be identical to that measured for synthetic GKY20. This finding suggests that N-terminal proline residue does not change the antimicrobial properties of recombinant (P)GKY20. The improved carrier, which does not contain cysteine and methionine residues, Asp-Pro and Asn-Gly sequences, is well suited for the production of peptides using any of the most popular chemical cleavage methods. PMID:26808536

  15. Controlled release from recombinant polymers.

    Science.gov (United States)

    Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

    2014-09-28

    Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and temporal release. Unlike the majority of chemical synthetic strategies used, recombinant DNA technology has allowed for the production of monodisperse polymers with specifically defined sequences. Several classes of recombinant polymers have been used for controlled drug delivery. These include, but are not limited to, elastin-like, silk-like, and silk-elastinlike proteins, as well as emerging cationic polymers for gene delivery. In this article, progress and prospects of recombinant polymers used in controlled release will be reviewed.

  16. Cell encoding recombinant human erythropoietin

    Energy Technology Data Exchange (ETDEWEB)

    Beck, A.K.; Withy, R.M.; Zabrecky, J.R.; Masiello, N.C.

    1990-09-04

    This patent describes a C127 cell transformed with a recombinant DNA vector. It comprises: a DNA sequence encoding human erythropoietin, the transformed cell being capable of producing N-linked and O-linked glycosylated human erythropoietin.

  17. Binding of thrombin-activated platelets to a fibrin scaffold through α(IIbβ₃ evokes phosphatidylserine exposure on their cell surface.

    Directory of Open Access Journals (Sweden)

    Tomasz Brzoska

    Full Text Available Recently, by employing intra-vital confocal microscopy, we demonstrated that platelets expose phosphatidylserine (PS and fibrin accumulate only in the center of the thrombus but not in its periphery. To address the question how exposure of platelet anionic phospholipids is regulated within the thrombus, an in-vitro experiment using diluted platelet-rich plasma was employed, in which the fibrin network was formed in the presence of platelets, and PS exposure on the platelet surface was analyzed using Confocal Laser Scanning Microscopy. Almost all platelets exposed PS after treatment with tissue factor, thrombin or ionomycin. Argatroban abrogated fibrin network formation in all samples, however, platelet PS exposure was inhibited only in tissue factor- and thrombin-treated samples but not in ionomycin-treated samples. FK633, an α(IIbβ₃ antagonist, and cytochalasin B impaired platelet binding to the fibrin scaffold and significantly reduced PS exposure evoked by thrombin. Gly-Pro-Arg-Pro amide abrogated not only fibrin network formation, but also PS exposure on platelets without suppressing platelet binding to fibrin/fibrinogen. These results suggest that outside-in signals in platelets generated by their binding to the rigid fibrin network are essential for PS exposure after thrombin treatment.

  18. The effect of genetic variants in the thrombin activatable fibrinolysis inhibitor (TAFI) gene on TAFI-antigen levels, clot lysis time and the risk of venous thrombosis

    NARCIS (Netherlands)

    C.H. Martini (C.); A. Brandts (A.); E.L.E. de Bruijne; A. van Hylckama Vlieg (Astrid); F.W.G. Leebeek (Frank); T. Lisman (Ton); F.R. Rosendaal (Frits)

    2006-01-01

    textabstractThrombin activatable fibrinolysis inhibitor (TAFI) is an important inhibitor of fibrinolysis. High TAFI antigen levels are associated with an increased risk of deep venous thrombosis (DVT). Because TAFI levels are partly determined genetically, we assessed the association between three T

  19. Thrombomodulin is a determinant of metastasis through a mechanism linked to the thrombin binding domain but not the lectin-like domain

    NARCIS (Netherlands)

    N.A. Horowitz; E.A. Blevins; W.M. Miller; A.R. Perry; K.E. Talmage; E.S. Mullins; M.J. Flick; K.C.S. Queiroz; K. Shi; C.A. Spek; E.M. Conway; B.P. Monia; H. Weiler; J.L. Degen; J.S. Palumbo

    2011-01-01

    Thrombomodulin (TM) is a predominantly endothelial transmembrane glycoprotein that modulates hemostatic function through a domain that controls thrombin-mediated proteolysis and an N-terminal lectin-like domain that controls inflammatory processes. To test the hypothesis that TM is a determinant of

  20. Stimulation of Naive Monocytes and PBMCs with Coagulation Proteases Results in Thrombin-Mediated and PAR-1-Dependent Cytokine Release and Cell Proliferation in PBMCs Only

    NARCIS (Netherlands)

    Nieuwenhuizen, L.; Falkenburg, W. J. J.; Schutgens, R. E. G.; Roosendaal, G.; van Veghel, K.; Biesma, D. H.; Lafeber, F. P. J. G.

    2013-01-01

    Protease-activated receptors (PARs) are stimulated by proteolytic cleavage of their extracellular domain. Coagulation proteases, such as FVIIa, the binary TF-FVIIa complex, free FXa, the ternary TF-FVIIa-FXa complex and thrombin, are able to stimulate PARs. Whereas the role of PARs on platelets is w

  1. mRNA expression of genes involved in inflammation and haemostasis in equine fibroblast-like synoviocytes following exposure to lipopolysaccharide, fibrinogen and thrombin

    DEFF Research Database (Denmark)

    Andreassen, Stine Mandrup; Berg, Lise Charlotte; Nielsen, Søren Saxmose;

    2015-01-01

    ) and protease activator receptor 1 (PAR-1) was assessed using quantitative real time reverse transcriptase PCR. Results: LPS caused a significant increase in mRNA expression of SAA, IL-6, MCP-1 and uPA, and a decrease in TF, PAI-1 and PAR-1 when compared to non-treated cells. Treatment with thrombin resulted...

  2. 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)

    Johansson, Per Ingemar; Svendsen, M.S.; Salado, J.;

    2008-01-01

    .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...

  3. Progenitors of Recombining Supernova Remnants

    OpenAIRE

    Moriya, Takashi J.

    2012-01-01

    Usual supernova remnants have either ionizing plasma or plasma in collisional ionization equilibrium, i.e., the ionization temperature is lower than or equal to the electron temperature. However, the existence of recombining supernova remnants, i.e., supernova remnants with the ionization temperature higher than the electron temperature, is recently confirmed. One suggested way to have recombining plasma in a supernova remnant is to have a dense circumstellar medium at the time of the superno...

  4. Plasmid recombination in Haemophilus influenzae

    Energy Technology Data Exchange (ETDEWEB)

    McCarthy, D.

    1982-01-01

    DNA recombination in exponential phase and competent Haemophilus influenzae was measured by an electron microscopic assay that relies on the conversion of plasmid RSF0885 monomers into multimeric forms. Dimer circles were present at a frequency of 2% in plasmid preparations from competent Rd (wild-type) cells; multimers were present at a frequency of 0.2% in preparations from exponential phase cells. Thus, plasmid recombination was stimulated in competent cells. Multimer formation occurred efficiently in cells of the transformation defective mutant rec2, implying that the rec2 gene product is not required for plasmid recombination. However, the absence of multimer plasmids in preparations from competent cells of the transformation defective mutant rec1 suggests that the rec1 gene product is required. Digestion of purified plasmids with restriction endonuclease PvuII, which makes a single cut in the monomer, revealed the presence of recombination intermediates composed of two linear plasmids joined to form two pairs of arms resembling the Greek letter chi. Length measurements of these arms taken from a population of recombination intermediates gave evidence that the plasmids were joined at sites of homology. The distributions of individual DNA strands, at the intersections of the four arms, could be resolved in some recombination intermediates and were of two types. The first type of junction appeared as a single-stranded arm appended to each corner. The second type of junction consisted of a single strand of DNA linking the two linear plasmids at a site of homology. The single-stranded linker was frequently situated at the edge of a short gap on one of the plasmids in the pair. The fine structures of the recombinational joints have been interpreted in terms of previously proposed models of recombination.

  5. Topics in black hole evaporation

    International Nuclear Information System (INIS)

    Two major aspects of particle creation by gravitational fields of black holes are studied: the neutrino emission from rotating black holes; and interactions between scalar particles emitted by a black hole. Neutrino emission is investigated under three topics: The asymmetry of the angular dependence of neutrino emission from rotating black holes; the production of a local matter excess by rotating black holes in a baryon symmetric universe; and cosmological magnetic field generation by neutrinos from evaporating black holes. Finally the author studies the effects of interactions on the black hole evaporation process

  6. Topics in atomic collision theory

    CERN Document Server

    Geltman, Sydney; Brueckner, Keith A

    1969-01-01

    Topics in Atomic Collision Theory originated in a course of graduate lectures given at the University of Colorado and at University College in London. It is recommended for students in physics and related fields who are interested in the application of quantum scattering theory to low-energy atomic collision phenomena. No attention is given to the electromagnetic, nuclear, or elementary particle domains. The book is organized into three parts: static field scattering, electron-atom collisions, and atom-atom collisions. These are in the order of increasing physical complexity and hence necessar

  7. Topics in commutative ring theory

    CERN Document Server

    Watkins, John J

    2009-01-01

    Topics in Commutative Ring Theory is a textbook for advanced undergraduate students as well as graduate students and mathematicians seeking an accessible introduction to this fascinating area of abstract algebra. Commutative ring theory arose more than a century ago to address questions in geometry and number theory. A commutative ring is a set-such as the integers, complex numbers, or polynomials with real coefficients--with two operations, addition and multiplication. Starting from this simple definition, John Watkins guides readers from basic concepts to Noetherian rings-one of

  8. Potential theory—selected topics

    CERN Document Server

    Aikawwa, Hiroaki

    1996-01-01

    The first part of these lecture notes is an introduction to potential theory to prepare the reader for later parts, which can be used as the basis for a series of advanced lectures/seminars on potential theory/harmonic analysis. Topics covered in the book include minimal thinness, quasiadditivity of capacity, applications of singular integrals to potential theory, L(p)-capacity theory, fine limits of the Nagel-Stein boundary limit theorem and integrability of superharmonic functions. The notes are written for an audience familiar with the theory of integration, distributions and basic functional analysis.

  9. Thrombin cleavage of osteopontin disrupts a pro-chemotactic sequence for dendritic cells, which is compensated by the release of its pro-chemotactic C-terminal fragment.

    Science.gov (United States)

    Shao, Zhifei; Morser, John; Leung, Lawrence L K

    2014-09-26

    Thrombin cleavage alters the function of osteopontin (OPN) by exposing an integrin binding site and releasing a chemotactic C-terminal fragment. Here, we examined thrombin cleavage of OPN in the context of dendritic cell (DC) migration to define its functional domains. Full-length OPN (OPN-FL), thrombin-cleaved N-terminal fragment (OPN-R), thrombin- and carboxypeptidase B2-double-cleaved N-terminal fragment (OPN-L), and C-terminal fragment (OPN-CTF) did not have intrinsic chemotactic activity, but all potentiated CCL21-induced DC migration. OPN-FL possessed the highest potency, whereas OPNRAA-FL had substantially less activity, indicating the importance of RGD. We identified a conserved (168)RSKSKKFRR(176) sequence on OPN-FL that spans the thrombin cleavage site, and it demonstrated potent pro-chemotactic effects on CCL21-induced DC migration. OPN-FLR168A had reduced activity, and the double mutant OPNRAA-FLR168A had even lower activity, indicating that these functional domains accounted for most of the pro-chemotactic activity of OPN-FL. OPN-CTF also possessed substantial pro-chemotactic activity, which was fully expressed upon thrombin cleavage and its release from the intact protein, because OPN-CTF was substantially more active than OPNRAA-FLR168A containing the OPN-CTF sequence within the intact protein. OPN-R and OPN-L possessed similar potency, indicating that the newly exposed C-terminal SVVYGLR sequence in OPN-R was not involved in the pro-chemotactic effect. OPN-FL and OPN-CTF did not directly bind to the CD44 standard form or CD44v6. In conclusion, thrombin cleavage of OPN disrupts a pro-chemotactic sequence in intact OPN, and its loss of pro-chemotactic activity is compensated by the release of OPN-CTF, which assumes a new conformation and possesses substantial activity in enhancing chemokine-induced migration of DCs. PMID:25112870

  10. Study on the effects of thrombin on AQP4 mRNA and AQP4 protein expression in rat primary astrocytes

    Institute of Scientific and Technical Information of China (English)

    Jinghua Zhou; Xuebing Cao; Shenggang Sun

    2006-01-01

    Objective: To study the biologic effects of various concentrations of thrombin on aquaporin 4 (AQP4) expression in rat primary cultured astrocytes, and to explore the regulation mechanism of transmembrane water transportation in astrocytes after intracerebral hemorrhage (ICH). Methods: Primary cultured astrocytes were incubated in culture mediums containing various concentrations of thrombin for 24 h and harvested. AQP4 mRNA and AQP4 protein expression were determined by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemical technique. Cell apoptosis was detected by TdT-mediated dUTP nick end labeling (TUNEL) technique. Cell morphology was observed by phase contrast microscope, and cell viability was assayed by MTT. Results: AQP4 mRNA and AQP4 protein showed a low expression in normal astrocytes. The expression of AQP4 mRNA and AQP4 protein significantly increased in the astrocytes treated with 100 U/ml or 200 U/ml thrombin (P < 0.01),and these astrocytes swelled. The number of TUNEL positive cells significantly increased. On the other hand, AQP4 mRNA and AQP4 protein expression were down-regulated in the astrocytes treated with 0.5 U/ml or l U/ml thrombin (P < 0.05),and the cell morphology did not change. Few TUNEL positive cells were observed. Conclusion: AQP4 over-expression induced by high concentrations of thrombin causes an increased permeability of water in astrocytic membrane. On the contrary, the decreased AQP4 expression prevents the astrocytes from swelling and apoptosis.

  11. A multifunctional hemin@metal-organic framework and its application to construct an electrochemical aptasensor for thrombin detection

    Science.gov (United States)

    Xie, Shunbi; Ye, Jiawei; Yuan, Yali; Chai, Yaqin; Yuan, Ruo

    2015-10-01

    A new type of multifunctional metal-organic framework (MOF) has been synthesized by encapsulating hemin into the nano-sized Fe-MIL-88 MOFs (hemin@MOFs) and first applied in an electrochemical aptasensor to detect thrombin (TB) with the aid of an enzyme for signal amplification. The gold nanoparticle functionalized hemin@MOFs (Au/hemin@MOFs) have not only simultaneously served as redox mediators and solid electrocatalysts, but have also been utilized as an ideal loading platform to immobilize a large number of biomolecules. In this aptasensor, Au/hemin@MOFs conjugated with glucose oxidase (GOD) and thrombin binding aptamer (TBA II) were used as the secondary aptamer bioconjugates (Au/hemin@MOF-TBA II-GOD bioconjugates), and TB was sandwiched between Au/hemin@MOF-TBA II-GOD bioconjugates and the amino-terminated TBA I which was self-assembled on the gold nanoparticle (AuNP) modified electrode. The GOD could oxidize glucose into gluconic acid accompanied by the generation of H2O2. The generated H2O2 on the electrode surface was further electrocatalyzed by hemin@MOFs to amplify the electrochemical signal of hemin contained in hemin@MOFs. Therefore, the synthesized hemin@MOFs represented a new paradigm for multifunctional materials since it combined three different functions including serving as catalysts, redox mediators and loading platforms within a single material. With such an ingenious design, a wide linear range of 0.0001 nM to 30 nM was acquired with a relatively low detection limit of 0.068 pM for TB detection.A new type of multifunctional metal-organic framework (MOF) has been synthesized by encapsulating hemin into the nano-sized Fe-MIL-88 MOFs (hemin@MOFs) and first applied in an electrochemical aptasensor to detect thrombin (TB) with the aid of an enzyme for signal amplification. The gold nanoparticle functionalized hemin@MOFs (Au/hemin@MOFs) have not only simultaneously served as redox mediators and solid electrocatalysts, but have also been utilized as an

  12. Radioiodination and biodistribution study of a thrombin-like enzyme/gyroxin from Lachesis muta muta venom

    International Nuclear Information System (INIS)

    Recently, our group isolated and sequenced a thrombin-like enzyme (TLE) of 40kDa from the snake Lachesis muta muta. This protein hydrolyses synthetic substrates with specificity similar to that of trypsin and may be involved in the haemorrhagic, proteolytic and blood-clotting activities of the Lachesis venom. When injected into the tail veins of mice at levels of 0.015-0.130μg/g mouse, the TLE induce temporary episodes of opisthotonus and rapid rolling around the long axis of the animals and that is the reason why it is also called gyroxin. If this gyroxin activity is caused by direct interaction with the brain or by indirect effect remains to be investigated. We report in this work the radioiodination of TLE and the biodistribution of I-TLE in order to investigate its pharmacokinetics and verify if this enzyme are able to penetrate the blood brain barrier to evoke directly the gyroxin effects. (author)

  13. Meta-analysis of randomized controlled trials on risk of myocardial infarction from the use of oral direct thrombin inhibitors

    DEFF Research Database (Denmark)

    Artang, Ramin; Rome, Eric; Nielsen, Jørn Dalsgaard;

    2013-01-01

    . To address these questions, we systematically searched MEDLINE and performed a meta-analysis on randomized trials that compared oral DTIs with warfarin for any indication with end point of MIs after randomization. We furthermore performed a secondary meta-analysis on atrial fibrillation stroke prevention......Dabigatran has been associated with greater risk of myocardial infarction (MI) than warfarin. It is unknown whether the increased risk is unique to dabigatran, an adverse effect shared by other oral direct thrombin inhibitors (DTIs), or the result of a protective effect of warfarin against MI...... trials with alternative anticoagulants compared with warfarin with end point of MIs after randomization. A total of 11 trials (39,357 patients) that compared warfarin to DTIs (dabigatran, ximelagatran, and AZD0837) were identified. In these trials, patients treated with oral DTIs were more likely...

  14. Human-competitive automatic topic indexing

    CERN Document Server

    Medelyan, Olena

    2009-01-01

    Topic indexing is the task of identifying the main topics covered by a document. These are useful for many purposes: as subject headings in libraries, as keywords in academic publications and as tags on the web. Knowing a document’s topics helps people judge its relevance quickly. However, assigning topics manually is labor intensive. This thesis shows how to generate them automatically in a way that competes with human performance. Three kinds of indexing are investigated: term assignment, a task commonly performed by librarians, who select topics from a controlled vocabulary; tagging, a popular activity of web users, who choose topics freely; and a new method of keyphrase extraction, where topics are equated to Wikipedia article names. A general two-stage algorithm is introduced that first selects candidate topics and then ranks them by significance based on their properties. These properties draw on statistical, semantic, domain-specific and encyclopedic knowledge. They are combined using a machine learn...

  15. Recombinant PsbF from Synechococcus sp. PCC 7002 forms β:βhomodimeric cytochrome b559

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    All organisms with oxygenic photosynthesis contain two photosystems: photosystemⅠ(PSⅠ) and photosystem-Ⅱ-(PSⅡ). The minimal photosystem-Ⅱ-particles which are photochemically active contain three subunits: D1, D2 and cytochrome b559 (Cyt b559). The function of Cyt b559 remains unclear. We have successfully overexpressed the psbF gene, encoding the - subunit of Cyt b559, from a marine cyanobacterium Synechococcus sp. PCC 7002 as a fusion gene and obtained a redox-active form of Cyt b559. When the N-terminal GST protein of the fusion gene product was removed with thrombin, the PsbF protein was still redox-active, suggesting that the recombinant PsbF can form dimer in Escherichia coli. The absorption spectra of either the oxidized form or the reduced form of both GST fusion protein and the purified PsbF dimer and the difference spectra between the two forms are the same as that of the Cyt b559 isolated from the higher plants. Redox titration analysis of recombinant PsbF showed that the mid-point redox potential of the recombinant Cyt b559 was approximately 50 mV, which is close to the low potential of Cyt b559. The results are helpful to the understanding of localization and function of Cyt b559 on thylakoid membranes.

  16. Topics in Banach space theory

    CERN Document Server

    Albiac, Fernando

    2016-01-01

    This text provides the reader with the necessary technical tools and background to reach the frontiers of research without the introduction of too many extraneous concepts. Detailed and accessible proofs are included, as are a variety of exercises and problems. The two new chapters in this second edition are devoted to two topics of much current interest amongst functional analysts: Greedy approximation with respect to bases in Banach spaces and nonlinear geometry of Banach spaces. This new material is intended to present these two directions of research for their intrinsic importance within Banach space theory, and to motivate graduate students interested in learning more about them. This textbook assumes only a basic knowledge of functional analysis, giving the reader a self-contained overview of the ideas and techniques in the development of modern Banach space theory. Special emphasis is placed on the study of the classical Lebesgue spaces Lp (and their sequence space analogues) and spaces of continuous f...

  17. Synergetics introduction and advanced topics

    CERN Document Server

    Haken, Hermann

    2004-01-01

    This book is an often-requested reprint of two classic texts by H. Haken: "Synergetics. An Introduction" and "Advanced Synergetics". Synergetics, an interdisciplinary research program initiated by H. Haken in 1969, deals with the systematic and methodological approach to the rapidly growing field of complexity. Going well beyond qualitative analogies between complex systems in fields as diverse as physics, chemistry, biology, sociology and economics, Synergetics uses tools from theoretical physics and mathematics to construct an unifying framework within which quantitative descriptions of complex, self-organizing systems can be made. This may well explain the timelessness of H. Haken's original texts on this topic, which are now recognized as landmarks in the field of complex systems. They provide both the beginning graduate student and the seasoned researcher with solid knowledge of the basic concepts and mathematical tools. Moreover, they admirably convey the spirit of the pioneering work by the founder of ...

  18. KEY TOPICS IN SPORTS MEDICINE

    Directory of Open Access Journals (Sweden)

    Amir Ali Narvani

    2006-12-01

    Full Text Available 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 psychology of performance and injury. PURPOSE The Key Topics format provides extensive, concise information in an accessible, easy-to-follow manner. AUDIENCE The book is targeted the students and specialists in sports medicine and rehabilitation, athletic training, physiotherapy and orthopaedic surgery. The editors are authorities in their respective fields and this handbook depends on their extensive experience and knowledge accumulated over the years. FEATURES The book contains the information for clinical guidance, rapid access to concise details and facts. It is composed of 99 topics which present the information in an order that is considered logical and progressive as in most texts. Chapter headings are: 1. Functional Anatomy, 2. Training Principles / Development of Strength and Power, 3. Biomechanical Principles, 4. Biomechanical Analysis, 5. Physiology of Training, 6. Monitoring of Training Progress, 7. Nutrition, 8. Hot and Cold Climates, 9. Altitude, 10. Sport and Travelling, 11. Principles of Sport Injury Diagnosis, 12. Principles of Sport and Soft Tissue Management, 13. Principles of Physical Therapy and Rehabilitation, 14. Principles of Sport Injury Prevention, 15. Sports Psychology, 16. Team Sports, 17. Psychological Aspects of Injury in Sport, 18. Injury Repair Process, 19. Basic Biomechanics of Tissue Injury, 20. Plain Film Radiography in Sport, 21. Nuclear Medicine, 22. Diagnostic Ultrasound, 23. MRI Scan, 24. Other Imaging, 5. Head Injury, 26. Eye

  19. Topics in Open Topological Strings

    CERN Document Server

    Prudenziati, Andrea

    2010-01-01

    This thesis is based on some selected topics in open topological string theory which I have worked on during my Ph.D. It comprises an introductory part where I have focused on the points most needed for the later chapters, trading completeness for conciseness and clarity. Then, following [12], we discuss tadpole cancellation for topological strings where we mainly show how its implementation is needed for ensuring the same "odd" moduli decoupling encountered in the closed theory. Next we move to analyse how the open and closed effective field theories for the B model interact writing the complete Lagrangian. We first check it deriving some already known tree level amplitudes in term of target space quantities, and then we extend the recipe to new results; later we implement open closed duality from a target field theory perspective. This last subject is also analysed from a worldsheet point of view extending the analysis of [13]. Some ideas for future research are briefly reported.

  20. Multilingual Topic Models for Unaligned Text

    CERN Document Server

    Boyd-Graber, Jordan

    2012-01-01

    We develop the multilingual topic model for unaligned text (MuTo), a probabilistic model of text that is designed to analyze corpora composed of documents in two languages. From these documents, MuTo uses stochastic EM to simultaneously discover both a matching between the languages and multilingual latent topics. We demonstrate that MuTo is able to find shared topics on real-world multilingual corpora, successfully pairing related documents across languages. MuTo provides a new framework for creating multilingual topic models without needing carefully curated parallel corpora and allows applications built using the topic model formalism to be applied to a much wider class of corpora.

  1. Primary targeting of recombinant Fv-immunotoxin hscFv25-mTNFα against hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Jing Zhang; Yan-Fang Liu; Shou-Jing Yang; Qing Qiao; Hong Cheng; Chuan-Shan Zhang; Fu-Cheng Ma; Hua-Zhang Guo

    2004-01-01

    AIM: To obtain human recombinant Fv-immunotoxin hscFv25-mTNFα (mutant human TNFα fused to human scFv25) against hepatocellular carcinoma (HCC). METHODS: Two relevant sites of enzymatic digestion were added to mTNFα by PCR. mTNFα was linked to the 3' end of hscFv25 in pGEX4T-1 vector. This anti-HCC recombinant Fv-immunotoxin hscFv25-mTNFα was expressed in Escherichia coliand purified from inclusions. After purified by glutathione-S-transferase affinity chromatography and thrombin digestion, it was identified by electrophoresis and Western blot. And then, the purified recombinant Fv immunotoxin was injected into nude mice with HCC xenografts through their tail veins. mTNFα protein and PBS were used as control at the same time. After treated for two weeks, nude mice were executed. The bulk and weight of tumors were observed. The tumor tissues were stained by immunohistochemical method with TNFα antibody.RESULTS: The expression ratio of recombinant Fv-immunotoxin hscFv25-mTNFα was 12% of bacterial protein. The result of tumor restraining trials of hscFv25-mTNFα showed 2/5 complete remission and 3/5 partial remission. mTNFα restraining trials showed 5/5 partial remission. The therapeutic result of hscFv25- mTNFα was better than that of mTNFα (F=8.70, P<0.05). The hscFv25-mTNFα remedial tumor tissues were positive for TNFα by immunohistochemical staining. The positive granules mainly existed in the cytoplasm of tumor cell. CONCLUSION: Recombinant Fv-immunotoxin hscFv25-mTNFα has better therapeutic effect than mTNFα. Tt can inhibit the cellular growth of HCC and has some potential of clinical application.

  2. Heterogeneity in recombinant protein production

    DEFF Research Database (Denmark)

    Schalén, Martin; Johanson, Ted; Lundin, Luisa;

    2012-01-01

    . the cell physiology is affected. Cells are stressed, and this may severely affect growth, by-product accumulation, biomass yield and recombinant product yield. The stress caused by exposure to divergent microenvironments, genetic differences of individual cells, differing cell cycle stage and cell age, all...... contribute to make a population in a fermenter heterogeneous, resulting in cell-to-cell variation in physiological parameters of the microbial culture. Our study aims at investigating how population heterogeneity and recombinant protein production is affected by environmental gradients in bioreactors...... are simulated in small bioreactors and the population heterogeneity can be visualised by analysing single cells with flow cytometry. This can give new insights to cell physiology and recombinant protein production at the industrial scale....

  3. The effect of a single recombination event

    DEFF Research Database (Denmark)

    Schierup, Mikkel Heide; Jensen, Thomas Mailund; Wiuf, Carsten

    We investigate the variance in how visible a single recombination event is in a SNP data set as a function of the type of recombination event and its age. Data is simulated under the coalescent with recombination and inference is by the popular composite likelihood methods. The major determinant...... of the effect of a recombination event is the genealogical type of the event and whether SNP variation is present that can reveal the genealogical consequences of the recombination event. Recombination events that only change some branch lengths in the genealogy have a very small, but detectable, effect....... The more lineages left when the recombination event occurs, the larger effect it has, implying that it is mainly young recombination events that we detect when estimating the rate. If the population is growing, though, more lineages are present back in time and relatively more ancient recombination events...

  4. Identifying Topics in Microblogs Using Wikipedia.

    Science.gov (United States)

    Yıldırım, Ahmet; Üsküdarlı, Suzan; Özgür, Arzucan

    2016-01-01

    Twitter is an extremely high volume platform for user generated contributions regarding any topic. The wealth of content created at real-time in massive quantities calls for automated approaches to identify the topics of the contributions. Such topics can be utilized in numerous ways, such as public opinion mining, marketing, entertainment, and disaster management. Towards this end, approaches to relate single or partial posts to knowledge base items have been proposed. However, in microblogging systems like Twitter, topics emerge from the culmination of a large number of contributions. Therefore, identifying topics based on collections of posts, where individual posts contribute to some aspect of the greater topic is necessary. Models, such as Latent Dirichlet Allocation (LDA), propose algorithms for relating collections of posts to sets of keywords that represent underlying topics. In these approaches, figuring out what the specific topic(s) the keyword sets represent remains as a separate task. Another issue in topic detection is the scope, which is often limited to specific domain, such as health. This work proposes an approach for identifying domain-independent specific topics related to sets of posts. In this approach, individual posts are processed and then aggregated to identify key tokens, which are then mapped to specific topics. Wikipedia article titles are selected to represent topics, since they are up to date, user-generated, sophisticated articles that span topics of human interest. This paper describes the proposed approach, a prototype implementation, and a case study based on data gathered during the heavily contributed periods corresponding to the four US election debates in 2012. The manually evaluated results (0.96 precision) and other observations from the study are discussed in detail. PMID:26991442

  5. Identifying Topics in Microblogs Using Wikipedia.

    Directory of Open Access Journals (Sweden)

    Ahmet Yıldırım

    Full Text Available Twitter is an extremely high volume platform for user generated contributions regarding any topic. The wealth of content created at real-time in massive quantities calls for automated approaches to identify the topics of the contributions. Such topics can be utilized in numerous ways, such as public opinion mining, marketing, entertainment, and disaster management. Towards this end, approaches to relate single or partial posts to knowledge base items have been proposed. However, in microblogging systems like Twitter, topics emerge from the culmination of a large number of contributions. Therefore, identifying topics based on collections of posts, where individual posts contribute to some aspect of the greater topic is necessary. Models, such as Latent Dirichlet Allocation (LDA, propose algorithms for relating collections of posts to sets of keywords that represent underlying topics. In these approaches, figuring out what the specific topic(s the keyword sets represent remains as a separate task. Another issue in topic detection is the scope, which is often limited to specific domain, such as health. This work proposes an approach for identifying domain-independent specific topics related to sets of posts. In this approach, individual posts are processed and then aggregated to identify key tokens, which are then mapped to specific topics. Wikipedia article titles are selected to represent topics, since they are up to date, user-generated, sophisticated articles that span topics of human interest. This paper describes the proposed approach, a prototype implementation, and a case study based on data gathered during the heavily contributed periods corresponding to the four US election debates in 2012. The manually evaluated results (0.96 precision and other observations from the study are discussed in detail.

  6. Identifying Topics in Microblogs Using Wikipedia

    Science.gov (United States)

    Yıldırım, Ahmet; Üsküdarlı, Suzan; Özgür, Arzucan

    2016-01-01

    Twitter is an extremely high volume platform for user generated contributions regarding any topic. The wealth of content created at real-time in massive quantities calls for automated approaches to identify the topics of the contributions. Such topics can be utilized in numerous ways, such as public opinion mining, marketing, entertainment, and disaster management. Towards this end, approaches to relate single or partial posts to knowledge base items have been proposed. However, in microblogging systems like Twitter, topics emerge from the culmination of a large number of contributions. Therefore, identifying topics based on collections of posts, where individual posts contribute to some aspect of the greater topic is necessary. Models, such as Latent Dirichlet Allocation (LDA), propose algorithms for relating collections of posts to sets of keywords that represent underlying topics. In these approaches, figuring out what the specific topic(s) the keyword sets represent remains as a separate task. Another issue in topic detection is the scope, which is often limited to specific domain, such as health. This work proposes an approach for identifying domain-independent specific topics related to sets of posts. In this approach, individual posts are processed and then aggregated to identify key tokens, which are then mapped to specific topics. Wikipedia article titles are selected to represent topics, since they are up to date, user-generated, sophisticated articles that span topics of human interest. This paper describes the proposed approach, a prototype implementation, and a case study based on data gathered during the heavily contributed periods corresponding to the four US election debates in 2012. The manually evaluated results (0.96 precision) and other observations from the study are discussed in detail. PMID:26991442

  7. Main technical topics in 1999

    International Nuclear Information System (INIS)

    This Safety Authority annual report strives to present current organizational provisions and future trends in nuclear safety supervision in France and to describe the most outstanding occurrences during the past year. A first part presents nine documents concerning the main topics of 1999: aging of nuclear installations, the Offsite Emergency Plans (PPI), the impact of nuclear activities on man and the environment, the criticality hazards, EDF in 1999, the EPR project, the Andra in 1999, the transport incidents, the nuclear safety in eastern Europe. The second part presents the missions and actions of the Nuclear Installations Safety in the domains of the liabilities, the organization of the nuclear safety control, the regulations of the INB, the public information, the international relations, the crisis management, the radioactive materials transportation, the radioactive wastes. The equipment, the radiation protection and the exploitation of the pressurized water reactors are also treated just as the experimental reactors, the fuel cycle installations and the the nuclear installations dismantling. (A.L.B.)

  8. Topics in Number Theory Conference

    CERN Document Server

    Andrews, George; Ono, Ken

    1999-01-01

    From July 31 through August 3,1997, the Pennsylvania State University hosted the Topics in Number Theory Conference. The conference was organized by Ken Ono and myself. By writing the preface, I am afforded the opportunity to express my gratitude to Ken for beng the inspiring and driving force behind the whole conference. Without his energy, enthusiasm and skill the entire event would never have occurred. We are extremely grateful to the sponsors of the conference: The National Sci­ ence Foundation, The Penn State Conference Center and the Penn State Depart­ ment of Mathematics. The object in this conference was to provide a variety of presentations giving a current picture of recent, significant work in number theory. There were eight plenary lectures: H. Darmon (McGill University), "Non-vanishing of L-functions and their derivatives modulo p. " A. Granville (University of Georgia), "Mean values of multiplicative functions. " C. Pomerance (University of Georgia), "Recent results in primality testing. " C. ...

  9. Decision Point 1 Topical Report

    Energy Technology Data Exchange (ETDEWEB)

    Yablonsky, Al; Barsoumian, Shant; Legere, David

    2013-05-01

    This Topical Report addresses accomplishments achieved during Phase 2a of the SkyMine® Carbon Mineralization Pilot Project. The primary objectives of this project are to design, construct, and operate a system to capture CO2 from a slipstream of flue gas from a commercial coal-fired cement kiln, convert that CO2 to products having commercial value (i.e., beneficial use), show the economic viability of the CO2 capture and conversion process, and thereby advance the technology to the point of readiness for commercial scale demonstration and proliferation. The overall process is carbon negative, resulting in mineralization of CO2 that would otherwise be released into the atmosphere. The project will also substantiate market opportunities for the technology by sales of chemicals into existing markets, and identify opportunities to improve technology performance and reduce costs at the commercial scale. The project is being conducted in two phases. The primary objectives of Phase 1 were to elaborate proven SkyMine® process chemistry to commercial pilot-scale operation and complete the preliminary design for the pilot plant to be built and operated in Phase 2, complete a NEPA evaluation, and develop a comprehensive carbon life cycle analysis. The objective of the current Phase (2a) is to complete the detailed design of the pilot plant to be built in Phase 2b.

  10. The Virtual Robotics Laboratory; TOPICAL

    International Nuclear Information System (INIS)

    The growth of the Internet has provided a unique opportunity to expand research collaborations between industry, universities, and the national laboratories. The Virtual Robotics Laboratory (VRL) is an innovative program at Oak Ridge National Laboratory (ORNL) that is focusing on the issues related to collaborative research through controlled access of laboratory equipment using the World Wide Web. The VRL will provide different levels of access to selected ORNL laboratory secondary education programs. In the past, the ORNL Robotics and Process Systems Division has developed state-of-the-art robotic systems for the Army, NASA, Department of Energy, Department of Defense, as well as many other clients. After proof of concept, many of these systems sit dormant in the laboratories. This is not out of completion of all possible research topics. but from completion of contracts and generation of new programs. In the past, a number of visiting professors have used this equipment for their own research. However, this requires that the professor, and possibly his/her students, spend extended periods at the laboratory facility. In addition, only a very exclusive group of faculty can gain access to the laboratory and hardware. The VRL is a tool that enables extended collaborative efforts without regard to geographic limitations

  11. Recent Advances In Topical Therapy In Dermatology

    Directory of Open Access Journals (Sweden)

    Mohan Thappa Devinder

    2003-01-01

    Full Text Available With changing times various newer topical agents are introduced in the field of dermatology. Tacrolimus and pimecrolimus are immunisuppressants, which are effective topically and are tried in the management of atopic dermatitis as well as other disorders including allergic contact dermatitis, atrophic lichen planus, pyoderma gangrenosum. Imiquimod, an immune response modifier, is presently in use for genital warts but has potentials as anti- tumour agent and in various other dermatological conditions when used topically. Tazarotene is a newer addition to the list of topical reginoids, which is effective in psoriasis and has better effect in combination with calcipotriene, phototherapy and topical costicosteroids. Tazarotene and adapelene are also effective in inflammatory acne. Calcipotriol, a vitamin D analogue has been introduced as a topical agent in the treatment of psoriasis. Steroid components are also developed recently which will be devoid of the side effects but having adequate anti-inflammatory effect. Topical photodynamic therapy has also a wide range of use in dermatology. Newer topical agents including cidofovir, capsaicin, topical sensitizers, topical antifungal agents for onychomycosis are also of use in clinical practice. Other promising developments include skin substitutes and growth factors for wound care.

  12. Preparing Recombinant Gonad Organ Cultures

    OpenAIRE

    sprotocols

    2014-01-01

    Authors: Blanche Capel and Jordan Batchvarov Corresponding author ([]()) ### INTRODUCTION It can be useful to assay migration between any two adjacent tissues during development. This protocol assays cell migration between the gonad and mesonephros using tissue recombination between genetically marked and unmarked tissue, combined with an organ culture technique. First, agar blocks are prepared in a custom-built mold. The size and sh...

  13. Controlled Release from Recombinant Polymers

    OpenAIRE

    Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

    2014-01-01

    Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and tempor...

  14. Selected topics of fluid mechanics

    Science.gov (United States)

    Kindsvater, Carl E.

    1958-01-01

    the Euler, Froude, Reynolds, Weber, and Cauchy numbers are defined as essential tools for interpreting and using experimental data. The derivations of the energy and momentum equations are treated in detail. One-dimensional equations for steady nonuniform flow are developed, and the restrictions applicable to the equations are emphasized. Conditions of uniform and gradually varied flow are discussed, and the origin of the Chezy equation is examined in relation to both the energy and the momentum equations. The inadequacy of all uniform-flow equations as a means of describing gradually varied flow is explained. Thus, one of the definitive problems of river hydraulics is analyzed in the light of present knowledge. This report is the outgrowth of a series of short schools conducted during the spring and summer of 1953 for engineers of the Surface Water Branch, Water Resources Division, U. S. Geological Survey. The topics considered are essentially the same as the topics selected for inclusion in the schools. However, in order that they might serve better as a guide and outline for informal study, the arrangement of the writer's original lecture notes has been considerably altered. The purpose of the report, like the purpose of the schools which inspired it, is to build a simple but strong framework of the fundamentals of fluid mechanics. It is believed that this framework is capable of supporting a detailed analysis of most of the practical problems met by the engineers of the Geological Survey. It is hoped that the least accomplishment of this work will be to inspire the reader with the confidence and desire to read more of the recent and current technical literature of modern fluid mechanics.

  15. Turn-on near-infrared electrochemiluminescence sensing of thrombin based on resonance energy transfer between CdTe/CdS coresmall/shellthick quantum dots and gold nanorods.

    Science.gov (United States)

    Wang, Jing; Jiang, Xiaochun; Han, Heyou

    2016-08-15

    Here we designed a near-infrared electrochemiluminescence (NECL) aptasensor for turn-on ultrasensitive determination of thrombin. It was based on the ECL resonance energy transfer (ECL-RET) of CdTe/CdS coresmall/shellthick quantum dots (QDs) to gold nanorods (AuNRs). AuNRs which functioned as ECL acceptors were assembled onto CdTe/CdS film by DNA hybridization between aptamers and their complementary oligonucleotides. In the absence of thrombin, the NECL of QDs was quenched as a result of the ECL-RET of QDs to AuNRs. In the presence of thrombin, the NECL of the system was "turned on" because thrombin can replace the AuNRs onto the QDs film, owing to the specific aptamer-protein affinity interactions. In this way, the increment of ECL intensity and the concentration of thrombin showed a logarithmic linear correlation in the range of 100 aM to 10 fM with a detection limit of 31 aM (S/N=3). Importantly, the developed aptasensor was successfully applied to thrombin sensing in real serum samples. PMID:27031188

  16. Structural transformation induced by locked nucleic acid or 2′–O-methyl nucleic acid site-specific modifications on thrombin binding aptamer

    OpenAIRE

    Liu, Bo; Li, Da

    2014-01-01

    Background Locked nucleic acid (LNA) and 2'–O-methyl nucleic acid (OMeNA) are two of the most extensively studied nucleotide derivatives in the last decades. However, how they affect DNA quadruplex structures remains largely unknown. To explore their possible biological affinities for quadruplexes, we investigated how LNA- or OMeNA-substitutions affect G-quadruplex structure formation using a thrombin binding aptamer (TBA), the most studied extracorporal G-quadruplex-forming DNA sequence, whi...

  17. In Vitro Resistance of Staphylococcus aureus to Thrombin-Induced Platelet Microbicidal Protein Is Associated with Alterations in Cytoplasmic Membrane Fluidity

    OpenAIRE

    Bayer, Arnold S.; Prasad, Rajendra; Chandra, Jyotsna; Koul, Anjni; Smriti, M.; Varma, Archana; Skurray, Ronald A.; Firth, Neville; Brown, Melissa H.; Koo, Su-Pin; Yeaman, Michael R.

    2000-01-01

    Platelet microbicidal proteins (PMPs) are small, cationic peptides which possess potent microbicidal activities against common bloodstream pathogens, such as Staphylococcus aureus. We previously showed that S. aureus strains exhibiting resistance to thrombin-induced PMP (tPMP-1) in vitro have an enhanced capacity to cause human and experimental endocarditis (T. Wu, M. R. Yeaman, and A. S. Bayer, Antimicrob. Agents Chemother. 38:729–732, 1994; A. S. Bayer et al., Antimicrob. Agents Chemother. ...

  18. Live recombinant BHV/BRSV vaccine

    NARCIS (Netherlands)

    Keil, G.M.; Rijsewijk, F.A.M.

    1998-01-01

    The present invention refers to synthetic Bovine Respiratory Syncytium virus genes. Also the invention relates to live attenuated Bovine Herpesvirus recombinants carrying such synthetic genes. Furthermore, the invention relates to vaccines based on these live attenuated recombinants, for the protect

  19. Recombinant innovation and endogenous technological transitions

    NARCIS (Netherlands)

    K. Frenken; L.R. Izquierdo; P. Zeppini

    2012-01-01

    We propose a model of technological transitions based on two different types of innovations. Branching innovations refer to technological improvements along a particular path, while recombinant innovations represent fusions of multiple paths. Recombinant innovations create "short-cuts" which reduce

  20. Tracking topic birth and death in LDA.

    Energy Technology Data Exchange (ETDEWEB)

    Wilson, Andrew T.; Robinson, David Gerald

    2011-09-01

    Most topic modeling algorithms that address the evolution of documents over time use the same number of topics at all times. This obscures the common occurrence in the data where new subjects arise and old ones diminish or disappear entirely. We propose an algorithm to model the birth and death of topics within an LDA-like framework. The user selects an initial number of topics, after which new topics are created and retired without further supervision. Our approach also accommodates many of the acceleration and parallelization schemes developed in recent years for standard LDA. In recent years, topic modeling algorithms such as latent semantic analysis (LSA)[17], latent Dirichlet allocation (LDA)[10] and their descendants have offered a powerful way to explore and interrogate corpora far too large for any human to grasp without assistance. Using such algorithms we are able to search for similar documents, model and track the volume of topics over time, search for correlated topics or model them with a hierarchy. Most of these algorithms are intended for use with static corpora where the number of documents and the size of the vocabulary are known in advance. Moreover, almost all current topic modeling algorithms fix the number of topics as one of the input parameters and keep it fixed across the entire corpus. While this is appropriate for static corpora, it becomes a serious handicap when analyzing time-varying data sets where topics come and go as a matter of course. This is doubly true for online algorithms that may not have the option of revising earlier results in light of new data. To be sure, these algorithms will account for changing data one way or another, but without the ability to adapt to structural changes such as entirely new topics they may do so in counterintuitive ways.

  1. A cascade signal amplification strategy for surface enhanced Raman spectroscopy detection of thrombin based on DNAzyme assistant DNA recycling and rolling circle amplification.

    Science.gov (United States)

    Gao, Fenglei; Du, Lili; Tang, Daoquan; Lu, Yao; Zhang, Yanzhuo; Zhang, Lixian

    2015-04-15

    A sensitive protocol for surface enhanced Raman spectroscopy (SERS) detection of thrombin is designed with R6G-Ag NPs as a signal tag by combining DNAzyme assistant DNA recycling and rolling circle amplification (RCA). Molecular beacon (MB) as recognition probe immobilizes on the glass slides and performs the amplification procedure. After thrombin-induced structure-switching DNA hairpins of probe 1, the DNAzyme is liberated from the caged structure, which hybridizes with the MB for cleavage of the MB in the presence of cofactor Zn(2+) and initiates the DNA recycling process, leading to the cleavage of a large number of MB and the generation of numerous primers for triggering RCA reaction. The long amplified RCA product which contained hundreds of tandem-repeat sequences, which can bind with oligonucleotide functionalized Ag NPs reporters. The attached signal tags can be easily read out by SERS. Because of the cascade signal amplification, these newly designed protocols provides a sensitive SERS detection of thrombin down to the femolar level (2.3fM) with a linear range of 5 orders of magnitude (from 10(-14) to 10(-9)M) and have high selectivity toward its target protein. The proposed method is expected to be a good clinical tool for the diagnosis of a thrombotic disease.

  2. Updates of Topical and Local Anesthesia Agents.

    Science.gov (United States)

    Boyce, Ricardo A; Kirpalani, Tarun; Mohan, Naveen

    2016-04-01

    As described in this article, there are many advances in topical and local anesthesia. Topical and local anesthetics have played a great role in dentistry in alleviating the fears of patients, eliminating pain, and providing pain control. Many invasive procedures would not be performed without the use and advances of topical/local anesthetics. The modern-day dentist has the responsibility of knowing the variety of products on the market and should have at least references to access before, during, and after treatment. This practice ensures proper care with topical and local anesthetics for the masses of patients entering dental offices worldwide. PMID:27040295

  3. Topical Products for the Aging Face.

    Science.gov (United States)

    McCook, John P

    2016-07-01

    This article focuses on nonprescription home-use topical treatment technologies for the aging face and is intended to serve as a guide for the core cosmeceutical technologies currently used and to help educate and assist the selection of topical antiaging products by the professional staff and their patients. Antiaging topical treatments for patient home use should be nonirritating, compatible with the patient skin type, effective, and complementary to surgical and minimally invasive office procedures, and aesthetically elegant. New topical antiaging technologies, formulated as monotherapy or as combinations with well-known cosmeceuticals, should present adequate clinical studies to support their selection for use. PMID:27363774

  4. The treatment of rosacea with topical ivermectin.

    Science.gov (United States)

    Ali, S T; Alinia, H; Feldman, S R

    2015-04-01

    The treatment of rosacea is challenging because several pathophysiologic processes may be involved, including neurovascular dysregulation and alterations in innate immune status. Demodex mites may play a role in the latter mechanism. Topical ivermectin is a new therapeutic modality which demonstrates antiparasitic and anti-inflammatory properties. This article reviews published evidence related to the efficacy and safety of topical ivermectin. PubMed was utilized to search for key words "topical ivermectin", "ivermectin cream" and "rosacea". Three clinical trials were found that studied topical ivermectin as a treatment option for rosacea. Ivermectin was effective, safe and well tolerated. PMID:26020066

  5. Topical 5-azacytidine accelerates skin wound healing in rats.

    Science.gov (United States)

    Gomes, Fabiana S; de-Souza, Gabriela F; Nascimento, Lucas F; Arantes, Eva L; Pedro, Rafael M; Vitorino, Daniele C; Nunez, Carla E; Melo Lima, Maria H; Velloso, Lício A; Araújo, Eliana P

    2014-01-01

    The development of new methods to improve skin wound healing may affect the outcomes of a number of medical conditions. Here, we evaluate the molecular and clinical effects of topical 5-azacytidine on wound healing in rats. 5-Azacytidine decreases the expression of follistatin-1, which negatively regulates activins. Activins, in turn, promote cell growth in different tissues, including the skin. Eight-week-old male Wistar rats were submitted to 8.0-mm punch-wounding in the dorsal region. After 3 days, rats were randomly assigned to receive either a control treatment or the topical application of a solution containing 5-azacytidine (10 mM) once per day. Photo documentation and sample collection were performed on days 5, 9, and 15. Overall, 5-azacytidine promoted a significant acceleration of complete wound healing (99.7% ± 0.7.0 vs. 71.2% ± 2.8 on day 15; n = 10; p < 0.01), accompanied by up to threefold reduction in follistatin expression. Histological examination of the skin revealed efficient reepithelization and cell proliferation, as evaluated by the BrdU incorporation method. 5-Azacytidine treatment also resulted in increased gene expression of transforming growth factor-beta and the keratinocyte markers involucrin and cytokeratin, as well as decreased expression of cytokines such as tumor necrosis factor-alpha and interleukin-10. Lastly, when recombinant follistatin was applied to the skin in parallel with topical 5-azacytidine, most of the beneficial effects of the drug were lost. Thus, 5-azacytidine acts, at least in part through the follistatin/activin pathway, to improve skin wound healing in rodents. PMID:25039304

  6. Determination of recombination in Mycoplasma hominis

    DEFF Research Database (Denmark)

    Jacobsen, Iben Søgaard; Boesen, Thomas; Mygind, Tina;

    2002-01-01

    indicating the presence of recombination. In order to test for intergenic recombination, phylogenetic trees were reconstructed for each of the genes but no well-supported bifurcating phylogenetic trees could be obtained. The genes were tested for intragenic recombination using the correlation between linkage...

  7. Common variants in the human platelet PAR4 thrombin receptor alter platelet function and differ by race

    Science.gov (United States)

    Edelstein, Leonard C.; Simon, Lukas M.; Lindsay, Cory R.; Kong, Xianguo; Teruel-Montoya, Raúl; Tourdot, Benjamin E.; Chen, Edward S.; Ma, Lin; Coughlin, Shaun; Nieman, Marvin; Holinstat, Michael; Shaw, Chad A.

    2014-01-01

    Human platelets express 2 thrombin receptors: protease-activated receptor (PAR)-1 and PAR4. Recently, we reported 3.7-fold increased PAR4-mediated aggregation kinetics in platelets from black subjects compared with white subjects. We now show that platelets from blacks (n = 70) express 14% more PAR4 protein than those from whites (n = 84), but this difference is not associated with platelet PAR4 function. Quantitative trait locus analysis identified 3 common single nucleotide polymorphisms in the PAR4 gene (F2RL3) associated with PAR4-induced platelet aggregation. Among these single nucleotide polymorphisms, rs773902 determines whether residue 120 in transmembrane domain 2 is an alanine (Ala) or threonine (Thr). Compared with the Ala120 variant, Thr120 was more common in black subjects than in white subjects (63% vs 19%), was associated with higher PAR4-induced human platelet aggregation and Ca2+ flux, and generated greater inositol 1,4,5-triphosphate in transfected cells. A second, less frequent F2RL3 variant, Phe296Val, was only observed in blacks and abolished the enhanced PAR4-induced platelet aggregation and 1,4,5-triphosphate generation associated with PAR4-Thr120. PAR4 genotype did not affect vorapaxar inhibition of platelet PAR1 function, but a strong pharmacogenetic effect was observed with the PAR4-specific antagonist YD-3 [1-benzyl-3(ethoxycarbonylphenyl)-indazole]. These findings may have an important pharmacogenetic effect on the development of new PAR antagonists. PMID:25293779

  8. Isolation and Biochemical Characterization of a New Thrombin-Like Serine Protease from Bothrops pirajai Snake Venom

    Directory of Open Access Journals (Sweden)

    Kayena D. Zaqueo

    2014-01-01

    Full Text Available This paper presents a novel serine protease (SP isolated from Bothrops pirajai, a venomous snake found solely in Brazil that belongs to the Viperidae family. The identified SP, named BpirSP-39, was isolated by three chromatographic steps (size exclusion, bioaffinity, and reverse phase chromatographies. The molecular mass of BpirSP-39 was estimated by SDS-PAGE and confirmed by mass spectrometry (39,408.32 Da. The protein was able to form fibrin networks, which was not observed in the presence of serine protease inhibitors, such as phenylmethylsulfonyl fluoride (PMSF. Furthermore, BpirSP-39 presented considerable thermal stability and was apparently able to activate factor XIII of the blood coagulation cascade, unlike most serine proteases. BpirSP-39 was capable of hydrolyzing different chromogenic substrates tested (S-2222, S-2302, and S-2238 while Cu2+ significantly diminished BspirSP-39 activity on the three tested substrates. The enzyme promoted platelet aggregation and also exhibited fibrinogenolytic, fibrinolytic, gelatinolytic, and amidolytic activities. The multiple alignment showed high sequence similarity to other thrombin-like enzymes from snake venoms. These results allow us to conclude that a new SP was isolated from Bothrops pirajai snake venom.

  9. Variation in hospital resource use and cost among surgical procedures using topical absorbable hemostats

    Directory of Open Access Journals (Sweden)

    Martyn D

    2015-11-01

    Full Text Available Derek Martyn,1 Lisa M Meckley,1 Gavin Miyasato,1 Sangtaeck Lim,2 Jerome B Riebman,3 Richard Kocharian,3 Jillian G Scaife,1 Yajing Rao,1 Mitra Corral2 1Trinity Partners, LLC, Waltham, MA, USA; 2Global Health Economics and Market Access, Ethicon, Inc., Bridgewater, NJ, USA; 3Medical Affairs, Ethicon, Inc., Bridgewater, NJ, USA Background: Adjunctive hemostats are used to assist with the control of intraoperative bleeding. The most common types are flowables, gelatins, thrombins, and oxidized regenerated celluloses (ORCs. In the US, Surgicel® products are the only US Food and Drug Administration-approved ORCs. Objective: To compare the outcomes of health care resource utilization (HRU and costs associated with using ORCs compared to other adjunctive hemostats (OAHs are defined as flowables, gelatins, and topical thrombins for surgical procedures in the US inpatient setting. Patients and methods: A retrospective, US-based cohort study was conducted using hospital inpatient discharges from the 2011–2012 calendar years in the Premier Healthcare Database. Patients with either an ORC or an OAH who underwent a cardiovascular procedure (valve surgery and/or coronary artery bypass graft surgery, carotid endarterectomy, cholecystectomy, or hysterectomy were included. Propensity score matching was used to create comparable groups of ORC and OAH patients. Clinical, economic, and HRU outcomes were compared. Results: The propensity score matching created balanced patient cohorts for cardiovascular procedure (22,718 patients, carotid endarterectomy (10,890 patients, cholecystectomy (6,090 patients, and hysterectomy (9,348 patients. In all procedures, hemostatic agent costs were 28%–56% lower for ORCs, and mean hemostat units per discharge were 16%–41% lower for ORCs compared to OAHs. Length of stay and total procedure costs for patients treated with ORCs were lower for carotid endarterectomy patients (0.3 days and US$700 and for cholecystectomy patients

  10. Selection of Recombinant Human Antibodies.

    Science.gov (United States)

    Tomszak, Florian; Weber, Susanne; Zantow, Jonas; Schirrmann, Thomas; Hust, Michael; Frenzel, André

    2016-01-01

    Since the development of therapeutic antibodies the demand of recombinant human antibodies is steadily increasing. Traditionally, therapeutic antibodies were generated by immunization of rat or mice, the generation of hybridoma clones, cloning of the antibody genes and subsequent humanization and engineering of the lead candidates. In the last few years, techniques were developed that use transgenic animals with a human antibody gene repertoire. Here, modern recombinant DNA technologies can be combined with well established immunization and hybridoma technologies to generate already affinity maturated human antibodies. An alternative are in vitro technologies which enabled the generation of fully human antibodies from antibody gene libraries that even exceed the human antibody repertoire. Specific antibodies can be isolated from these libraries in a very short time and therefore reduce the development time of an antibody drug at a very early stage.In this review, we describe different technologies that are currently used for the in vitro and in vivo generation of human antibodies. PMID:27236551

  11. Mechanisms of sister chromatid recombination

    International Nuclear Information System (INIS)

    Studies using T948 as a model system have been carried out aimed at elucidating the mechanism of sister chromatid recombination (SCR). Characterization of U.V. light- and x-ray-induced SCR, the relationiship between SCR induction and DNA repair using rad mutations, and the relationship between SCR induction and the time of cell division using cdc mutations are presented. It has been supposed that SCR is induced at the phase of S-G2 following DNA replication, that postreplication break of DNA strands is strongly involved in the induction of SCR, and that induction type of SCR, i.e., conversion type or recombination type, is dependent upon the type of molecular damage of DNA. (Namekawa, K.)

  12. Corneal staining after treatment with topical tetracycline

    NARCIS (Netherlands)

    R. Lapid-Gortzak; C.P. Nieuwendaal; A.R. Slomovic; L. Spanjaard

    2006-01-01

    Purpose: The purpose of this paper is to report a case of corneal staining after treatment with topical tetracycline. Methods: A patient with crystalline keratopathy caused by Streptococcus viridans after corneal transplantation was treated topically with tetracycline eye drops, based on results of

  13. Psoriasis: improving adherence to topical therapy.

    NARCIS (Netherlands)

    Feldman, S.R.; Horn, E.J.; Balkrishnan, R.; Basra, M.K.; Finlay, A.Y.; McCoy, D.; Menter, A.; Kerkhof, P.C.M. van de

    2008-01-01

    Topical therapy has an important role in psoriasis treatment. It is efficacious and has a favorable safety profile as demonstrated in clinical trials. However, poor treatment outcomes from topical therapy regimens likely result from poor adherence and ineffective use of the medication. The Internati

  14. Fostering Topic Knowledge: Essential for Academic Writing

    Science.gov (United States)

    Proske, Antje; Kapp, Felix

    2013-01-01

    Several researchers emphasize the role of the writer's topic knowledge for writing. In academic writing topic knowledge is often constructed by studying source texts. One possibility to support that essential phase of the writing process is to provide interactive learning questions which facilitate the construction of an adequate situation…

  15. Infantile generalized hypertrichosis caused by topical minoxidil*

    OpenAIRE

    Rampon, Greice; Henkin, Caroline; de Souza, Paulo Ricardo Martins; de Almeida Jr, Hiram Larangeira

    2016-01-01

    Rare cases of hypertrichosis have been associated with topically applied minoxidil. We present the first reported case in the Brazilian literature of generalized hypertrichosis affecting a 5-year-old child, following use of minoxidil 5%, 20 drops a day, for hair loss. The laboratory investigation excluded hyperandrogenism and thyroid dysfunction. Topical minoxidil should be used with caution in children.

  16. Aerodynamics of wind turbines emerging topics

    CERN Document Server

    Amano, R S

    2014-01-01

    Focusing on Aerodynamics of Wind Turbines with topics ranging from Fundamental to Application of horizontal axis wind turbines, this book presents advanced topics including: Basic Theory for Wind turbine Blade Aerodynamics, Computational Methods, and Special Structural Reinforcement Technique for Wind Turbine Blades.

  17. Topic Maps Based Project Knowledge Management

    Institute of Scientific and Technical Information of China (English)

    Wu Xiaofan; Zhou Liang; Zhang Lei; Li Lingzhi; Ding Qiulin

    2006-01-01

    Based on topic maps, a preprocessing scheme using similarity comparision is presented and applied in knowledge management.Topic and occurrence-oriented merging algorithm is also introduced to implement knowledge integration for the sub-system. An Omnigator-supported example from an aeroaustic institute is utilised to validate the preprocessing method and the result indicates it can speed up the research schedule.

  18. Infantile generalized hypertrichosis caused by topical minoxidil*

    Science.gov (United States)

    Rampon, Greice; Henkin, Caroline; de Souza, Paulo Ricardo Martins; de Almeida Jr, Hiram Larangeira

    2016-01-01

    Rare cases of hypertrichosis have been associated with topically applied minoxidil. We present the first reported case in the Brazilian literature of generalized hypertrichosis affecting a 5-year-old child, following use of minoxidil 5%, 20 drops a day, for hair loss. The laboratory investigation excluded hyperandrogenism and thyroid dysfunction. Topical minoxidil should be used with caution in children. PMID:26982785

  19. Infantile generalized hypertrichosis caused by topical minoxidil.

    Science.gov (United States)

    Rampon, Greice; Henkin, Caroline; de Souza, Paulo Ricardo Martins; Almeida, Hiram Larangeira de

    2016-01-01

    Rare cases of hypertrichosis have been associated with topically applied minoxidil. We present the first reported case in the Brazilian literature of generalized hypertrichosis affecting a 5-year-old child, following use of minoxidil 5%, 20 drops a day, for hair loss. The laboratory investigation excluded hyperandrogenism and thyroid dysfunction. Topical minoxidil should be used with caution in children. PMID:26982785

  20. Histiocytosis X: treatment with topical nitrogen mustard.

    Science.gov (United States)

    Berman, B; Chang, D L; Shupack, J L

    1980-07-01

    The case histories of two elderly patients with cutaneous histiocytosis X treated topically with nitrogen mustard are presented. The cutaneous lesions cleared within 2 to 3 weeks, and remission was maintained with daily topical administration of nitrogen mustard. The clinical impression of improvement was substantiated by light and electron microscopic studies prior to and after therapy.

  1. Topics on Electricity Transmission Pricing

    International Nuclear Information System (INIS)

    -subsidization, the largest state-owned firm was split to form Statkraft SF, Norway's largest generation company, and Statnett SF, the major transmission company. Statnett is also the system operator of the entire Norwegian power system. Many distribution companies, usually publicly owned, are still part of vertically integrated companies, but there is strict separation of the financial accounts for transmission/distribution and generation/marketing. Trading is accomplished through several channels. The largest organized market is the common Norwegian-Swedish pool, NordPool. NordPool ASA is owned by Statnett Marked (which is a subsidiary of Statnett) and the Swedish grid-company, Svenska Kraftnaet. NordPool also supports the Finnish and Danish markets, thus through cooperation with the grid-companies of Denmark (Eltra) and Finland (Fingrid), locational markets are provided in these countries as well. Unlike England and Wales, only part of actual production is traded on this market. In addition, independent brokers offer various energy-contracts and facilitate bilateral contracts. In 1998 22% of the energy was traded on the hourly day-ahead (spot) market of NordPool. NordPool also organizes a forward and futures market and settles contracts from the regulation market organized by Statnett. The regulation market is a realtime market used to settle imbalances in real time. In total, close to 30% of the energy traded is organized through NordPool, and this share is increasing year by year. The efficiency of the 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. Throughout, the findings are illustrated by means of simple examples. This is to enhance readability and intuition. To start with, chapter 2 is devoted to power flow models, whereas chapter 3 provides an overview of models developed to efficiently coordinate the allocation of transmission

  2. Recombinant erythropoietin in clinical practice

    OpenAIRE

    Ng, T; Marx, G.; Littlewood, T; Macdougall, I

    2003-01-01

    The introduction of recombinant human erythropoietin (RHuEPO) has revolutionised the treatment of patients with anaemia of chronic renal disease. Clinical studies have demonstrated that RHuEPO is also useful in various non-uraemic conditions including haematological and oncological disorders, prematurity, HIV infection, and perioperative therapies. Besides highlighting both the historical and functional aspects of RHuEPO, this review discusses the applications of RHuEPO in clinical practice a...

  3. Recombinant antibodies and tumor targeting

    OpenAIRE

    Sheikholvaezin, Ali

    2006-01-01

    Different antibody derived constructs are rapidly advancing as putative tools for treatment of malignant diseases. Antibody engineering has added significant new technologies to modify size, affinities, solubility, stability and biodistribution properties for immunoconjugates. In the present thesis, the aim was to increase our knowledge on how new recombinant antibodies could be tailored to optimize localization to experimental tumors in mice. One hybridoma, producing the monoclonal antibody ...

  4. Workshop on Radio Recombination Lines

    CERN Document Server

    1980-01-01

    Since their first detection 15 years ago, radio recombination lines from several elements have been observed in a wide variety of objects including HII regions, planetary nebulae, molecular clouds, the diffuse interstellar medium, and recently, other galaxies. The observations span almost the entire range from 0.1 to 100 GHz, and employ both single­ djsh and aperture synthesis techniques. The theory of radio recombination lines has also advanced strongly, to the point where it is perhaps one of the best-understood in astro­ physics. In a parallel development, it has become possible over the last decade to study these same highly-excited atoms in the laboratory; this work provides further confirmation of the theoretical framework. However there has been continuing controversy over the astrophysical interpre­ tation of radio recombination line observations, especially regarding the role of stimulated emission. A workshop was held in Ottawa on 24-25 August, 1979, bringing together many of the active scientist...

  5. Topical antioxidants in radiodermatitis: a clinical review.

    Science.gov (United States)

    Kodiyan, Joyson; Amber, Kyle T

    2015-09-01

    Radiation-induced skin toxicity is the most prevalent side effect of radiation therapy. Not only does it have a significant effect on patients' quality of life, but it also results in poor follow-up and early termination of radiotherapy treatment. Several skin care practices and topical applications have been studied in the field of radiodermatitis, including skin washing, topical steroids, and mechanical skin barriers. Aside from these methods, many patients turn to complementary and alternative medicine for the prevention and treatment of radiodermatitis. Many of these alternative therapies are topically applied antioxidants. While the rationale behind the use of antioxidants in treating radiodermatitis is strong, clinical studies have been far less consistent. Even in large scale randomised controlled trials, findings have been limited by the inconsistent use of topical vehicles and placebos. In this article, the authors review the role of topical antioxidants to better help the practitioner navigate through different available skin directed antioxidants.

  6. Correlated Topic Model for Web Services Ranking

    Directory of Open Access Journals (Sweden)

    Mustapha AZNAG

    2013-07-01

    Full Text Available With the increasing number of published Web services providing similar functionalities, it’s very tedious for a service consumer to make decision to select the appropriate one according to her/his needs. In this paper, we explore several probabilistic topic models: Probabilistic Latent Semantic Analysis (PLSA, Latent Dirichlet Allocation (LDA and Correlated Topic Model (CTM to extract latent factors from web service descriptions. In our approach, topic models are used as efficient dimension reduction techniques, which are able to capture semantic relationships between word-topic and topic-service interpreted in terms of probability distributions. To address the limitation of keywords-based queries, we represent web service description as a vector space and we introduce a new approach for discovering and ranking web services using latent factors. In our experiment, we evaluated our Service Discovery and Ranking approach by calculating the precision (P@n and normalized discounted cumulative gain (NDCGn.

  7. Nondisjunction of chromosome 15: Origin and recombination

    Energy Technology Data Exchange (ETDEWEB)

    Robinson, W.P.; Bernasconi, F.; Schinzel, A.A.; Mutirangura, A.; Ledbetter, D.H. (Baylor College of Medicine, Houston, TX (United States)); Langlois, S. (Univ. of Britisch Columbia, Vancouver (Canada)); Morris, M.A.; Malcolm, S.

    1993-09-01

    Thirty-two cases of uniparental disomy (UPD), ascertained from Prader-Willi syndrome patients (N=27) and Angelman syndrome patients (N-5), are used to investigate the pattern of recombination associated with nondisjunction of chromosome 15. In addition, the meiotic stage of nondisjunction is inferred by using markers mapping near the centromere. Two basic approaches to the analysis of recombination in specific pairwise intervals along the chromosome. This method shows a significant reduction in recombination for two of five intervals examined. Second, the observed frequency of each recombinant class (i.e., zero, one, two, three, or more observable crossovers) is compared with expected values. This is useful for testing whether the reduction in recombination can be attributed solely to a proportion of cases with no recombination at all (because of asynapsis), with the remaining groups showing normal recombination (or even excess recombination), or whether recombination is uniformly reduced. Analysis of maternal UPD(15) data shows a slight reduction in the multiple-recombinant classes, with a corresponding increase in both the zero- and one-recombinant classes over expected values. The majority, more than 82%, of the extra chromosomes in maternal UPD(15) cases are due to meiotic I nondisjunction events. In contrast, more paternal UPD(15) cases so far examined appear to have a postzygotic origin of the extra paternal chromosome. 33 refs., 1 fig., 7 tabs.

  8. Topics in Electricity Transmission Pricing

    Energy Technology Data Exchange (ETDEWEB)

    Bjoerndal, Mette

    2000-02-01

    Directorate (NVE). To prevent cross-subsidization, the largest state-owned firm was split to form Statkraft SF, Norway's largest generation company, and Statnett SF, the major transmission company. Statnett is also the system operator of the entire Norwegian power system. Many distribution companies, usually publicly owned, are still part of vertically integrated companies, but there is strict separation of the financial accounts for transmission/distribution and generation/marketing. Trading is accomplished through several channels. The largest organized market is the common Norwegian-Swedish pool, NordPool. NordPool ASA is owned by Statnett Marked (which is a subsidiary of Statnett) and the Swedish grid-company, Svenska Kraftnaet. NordPool also supports the Finnish and Danish markets, thus through cooperation with the grid-companies of Denmark (Eltra) and Finland (Fingrid), locational markets are provided in these countries as well. Unlike England and Wales, only part of actual production is traded on this market. In addition, independent brokers offer various energy-contracts and facilitate bilateral contracts. In 1998 22% of the energy was traded on the hourly day-ahead (spot) market of NordPool. NordPool also organizes a forward and futures market and settles contracts from the regulation market organized by Statnett. The regulation market is a realtime market used to settle imbalances in real time. In total, close to 30% of the energy traded is organized through NordPool, and this share is increasing year by year. The efficiency of the 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. Throughout, the findings are illustrated by means of simple examples. This is to enhance readability and intuition. To start with, chapter 2 is devoted to power flow models, whereas chapter 3 provides an overview of models developed to efficiently

  9. Topics on Electricity Transmission Pricing

    Energy Technology Data Exchange (ETDEWEB)

    Bjoerndal, Mette

    2000-02-01

    Directorate (NVE). To prevent cross-subsidization, the largest state-owned firm was split to form Statkraft SF, Norway's largest generation company, and Statnett SF, the major transmission company. Statnett is also the system operator of the entire Norwegian power system. Many distribution companies, usually publicly owned, are still part of vertically integrated companies, but there is strict separation of the financial accounts for transmission/distribution and generation/marketing. Trading is accomplished through several channels. The largest organized market is the common Norwegian-Swedish pool, NordPool. NordPool ASA is owned by Statnett Marked (which is a subsidiary of Statnett) and the Swedish grid-company, Svenska Kraftnaet. NordPool also supports the Finnish and Danish markets, thus through cooperation with the grid-companies of Denmark (Eltra) and Finland (Fingrid), locational markets are provided in these countries as well. Unlike England and Wales, only part of actual production is traded on this market. In addition, independent brokers offer various energy-contracts and facilitate bilateral contracts. In 1998 22% of the energy was traded on the hourly day-ahead (spot) market of NordPool. NordPool also organizes a forward and futures market and settles contracts from the regulation market organized by Statnett. The regulation market is a realtime market used to settle imbalances in real time. In total, close to 30% of the energy traded is organized through NordPool, and this share is increasing year by year. The efficiency of the 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. Throughout, the findings are illustrated by means of simple examples. This is to enhance readability and intuition. Chapter 2 is devoted to power flow models, whereas chapter 3 provides an overview of models developed to efficiently coordinate the

  10. Topics in Electricity Transmission Pricing

    Energy Technology Data Exchange (ETDEWEB)

    Bjoerndal, Mette

    2000-02-01

    Directorate (NVE). To prevent cross-subsidization, the largest state-owned firm was split to form Statkraft SF, Norway's largest generation company, and Statnett SF, the major transmission company. Statnett is also the system operator of the entire Norwegian power system. Many distribution companies, usually publicly owned, are still part of vertically integrated companies, but there is strict separation of the financial accounts for transmission/distribution and generation/marketing. Trading is accomplished through several channels. The largest organized market is the common Norwegian-Swedish pool, NordPool. NordPool ASA is owned by Statnett Marked (which is a subsidiary of Statnett) and the Swedish grid-company, Svenska Kraftnaet. NordPool also supports the Finnish and Danish markets, thus through cooperation with the grid-companies of Denmark (Eltra) and Finland (Fingrid), locational markets are provided in these countries as well. Unlike England and Wales, only part of actual production is traded on this market. In addition, independent brokers offer various energy-contracts and facilitate bilateral contracts. In 1998 22% of the energy was traded on the hourly day-ahead (spot) market of NordPool. NordPool also organizes a forward and futures market and settles contracts from the regulation market organized by Statnett. The regulation market is a realtime market used to settle imbalances in real time. In total, close to 30% of the energy traded is organized through NordPool, and this share is increasing year by year. The efficiency of the 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. Throughout, the findings are illustrated by means of simple examples. This is to enhance readability and intuition. To start with, chapter 2 is devoted to power flow models, whereas chapter 3 provides an overview of models developed to efficiently

  11. Topics on Electricity Transmission Pricing

    Energy Technology Data Exchange (ETDEWEB)

    Bjoerndal, Mette

    2000-02-01

    Directorate (NVE). To prevent cross-subsidization, the largest state-owned firm was split to form Statkraft SF, Norway's largest generation company, and Statnett SF, the major transmission company. Statnett is also the system operator of the entire Norwegian power system. Many distribution companies, usually publicly owned, are still part of vertically integrated companies, but there is strict separation of the financial accounts for transmission/distribution and generation/marketing. Trading is accomplished through several channels. The largest organized market is the common Norwegian-Swedish pool, NordPool. NordPool ASA is owned by Statnett Marked (which is a subsidiary of Statnett) and the Swedish grid-company, Svenska Kraftnaet. NordPool also supports the Finnish and Danish markets, thus through cooperation with the grid-companies of Denmark (Eltra) and Finland (Fingrid), locational markets are provided in these countries as well. Unlike England and Wales, only part of actual production is traded on this market. In addition, independent brokers offer various energy-contracts and facilitate bilateral contracts. In 1998 22% of the energy was traded on the hourly day-ahead (spot) market of NordPool. NordPool also organizes a forward and futures market and settles contracts from the regulation market organized by Statnett. The regulation market is a realtime market used to settle imbalances in real time. In total, close to 30% of the energy traded is organized through NordPool, and this share is increasing year by year. The efficiency of the 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. Throughout, the findings are illustrated by means of simple examples. This is to enhance readability and intuition. Chapter 2 is devoted to power flow models, whereas chapter 3 provides an overview of models developed to efficiently coordinate the

  12. Activation of PAR-1/NADPH Oxidase/ROS Signaling Pathways is Crucial for the Thrombin-Induced sFlt-1 Production in Extravillous Trophoblasts: Possible Involvement in the Pathogenesis of Preeclampsia

    Directory of Open Access Journals (Sweden)

    Qi-tao Huang

    2015-03-01

    Full Text Available Backgrounds/Aims: Preeclampsia was characterized by excessive thrombin generation in placentas and previous researches showed that thrombin could enhance soluble Fms-like tyrosine kinase 1 (sFlt-1 expression in first trimester trophoblasts. However, the detailed mechanism for the sFlt-1 over-production induced by thrombin was largely unknown. The purpose of this study was to explore the possible signaling pathway of thrombin-induced sFlt-1 production in extravillous trophoblasts (EVT. Methods: An EVT cell line (HRT-8/SVneo was treated with various concentrations of thrombin. The mRNA expression and protein secretion of sFlt-1 in EVT were detected with real-time polymerase chain reaction and ELISA, respectively. The levels of intracellular reactive oxygen species (ROS production were determined by DCFH-DA. Results: Exposure of EVT to thrombin induced increased intracellular ROS generation and overexpression of sFlt-1 at both mRNA and protein levels in a dose dependent manner. Short interfering RNA (siRNA directed against PAR-1 or apocynin (an inhibitor of NADPH oxidase could decrease the intracellular ROS generation and subsequently suppressed the production of sFlt-1 at mRNA and protein levels. Conclusions: Our results suggested that thrombin increased sFlt-1 production in EVT via the PAR-1 /NADPH oxidase /ROS signaling pathway. This also highlights the PAR-1 / NADPH oxidase / ROS pathway might be a potential therapeutic target for the prevention of preeclampsia in the future.

  13. Recombinant Factor IX Fc Fusion Protein Maintains Full Procoagulant Properties and Exhibits Prolonged Efficacy in Hemophilia B Mice.

    Directory of Open Access Journals (Sweden)

    Garabet G Toby

    Full Text Available Hemophilia B is an inherited X chromosome-linked disorder characterized by impaired blood clotting owing to the absence of functional coagulation factor IX. Due to the relatively short half-life of factor IX, patients with hemophilia B require frequent factor IX infusions to maintain prophylaxis. We have developed a recombinant factor IX (rFIX fused to the Fc region of IgG (rFIXFc with an extended half-life in animals and humans.Procoagulant properties of rFIXFc and rFIX (BENEFIX® were compared to determine the effect of the Fc region on rFIXFc hemostatic function. Specifically, we assessed rFIXFc activation, intermolecular interactions within the Xase complex, inactivation by antithrombin III (AT and thrombin generation potential compared with rFIX. We also assessed the acute and prophylactic efficacy profiles of rFIXFc and rFIX in vivo in hemophilia B mouse bleeding models.The activation by factor XIa or factor VIIa/tissue factor, inhibition by AT, interaction profiles with phospholipids, affinities for factor VIIIa within the context of the Xase complex, and thrombin generation profiles were similar for rFIXFc and rFIX. Xase complexes formed with either molecule exhibited similar kinetic profiles for factor Xa generation. In acute efficacy models, mice infused with rFIXFc or rFIX were equally protected from bleeding. However, in prophylactic efficacy models, protection from bleeding was maintained approximately three times longer in rFIXFc-dosed mice than in those given rFIX; this prolonged efficacy correlates with the previously observed half-life extension. We conclude that rFIXFc retains critical FIX procoagulant attributes and that the extension in rFIXFc half-life translates into prolonged efficacy in hemophilia B mice.

  14. Analyses of Research Topics in the Field of Informetrics Based on the Method of Topic Modeling

    OpenAIRE

    Sung-Chien Lin

    2014-01-01

    In this study, we used the approach of topic modeling to uncover the possible structure of research topics in the field of Informetrics, to explore the distribution of the topics over years, and to compare the core journals. In order to infer the structure of the topics in the field, the data of the papers published in the Journal of Informetricsand Scientometrics during 2007 to 2013 are retrieved from the database of the Web of Science as input of the approach of topic modeling. The results ...

  15. Risk assessment of topically applied products

    DEFF Research Database (Denmark)

    Søborg, Tue; Basse, Line Hollesen; Halling-Sørensen, Bent

    2007-01-01

    parameters for estimating the risk. The immediate human risk of BADGE and derivatives in topical dosage forms was found to be low. However, local treatment of broken skin may lead to higher exposure of BADGE and derivatives compared to application to normal skin. 3-BC permeated skin at higher flux than 4-MBC......The human risk of harmful substances in semisolid topical dosage forms applied topically to normal skin and broken skin, respectively, was assessed. Bisphenol A diglycidyl ether (BADGE) and three derivatives of BADGE previously quantified in aqueous cream and the UV filters 3-BC and 4-MBC were used...

  16. Control of pain with topical plant medicines

    Institute of Scientific and Technical Information of China (English)

    James; David; Adams; Jr.; Xiaogang; Wang

    2015-01-01

    Pain is normally treated with oral nonsteroidal anti-inflammatory agents and opioids. These drugs are dangerous and are responsible for many hospitalizations and deaths. It is much safer to use topical preparations made from plants to treat pain, even severe pain. Topical preparations must contain compounds that penetrate the skin, inhibit pain receptors such as transient receptor potential cation channels and cyclooxygenase-2, to relieve pain. Inhibition of pain in the skin disrupts the pain cycle and avoids exposure of internal organs to large amounts of toxic compounds. Use of topical pain relievers has the potential to save many lives, decrease medical costs and improve therapy.

  17. Digital Social Network Mining for Topic Discovery

    Science.gov (United States)

    Moradianzadeh, Pooya; Mohi, Maryam; Sadighi Moshkenani, Mohsen

    Networked computers are expanding more and more around the world, and digital social networks becoming of great importance for many people's work and leisure. This paper mainly focused on discovering the topic of exchanging information in digital social network. In brief, our method is to use a hierarchical dictionary of related topics and words that mapped to a graph. Then, with comparing the extracted keywords from the context of social network with graph nodes, probability of relation between context and desired topics will be computed. This model can be used in many applications such as advertising, viral marketing and high-risk group detection.

  18. Bacteriophage recombination systems and biotechnical applications.

    Science.gov (United States)

    Nafissi, Nafiseh; Slavcev, Roderick

    2014-04-01

    Bacteriophage recombination systems have been widely used in biotechnology for modifying prokaryotic species, for creating transgenic animals and plants, and more recently, for human cell gene manipulation. In contrast to homologous recombination, which benefits from the endogenous recombination machinery of the cell, site-specific recombination requires an exogenous source of recombinase in mammalian cells. The mechanism of bacteriophage evolution and their coexistence with bacterial cells has become a point of interest ever since bacterial viruses' life cycles were first explored. Phage recombinases have already been exploited as valuable genetic tools and new phage enzymes, and their potential application to genetic engineering and genome manipulation, vectorology, and generation of new transgene delivery vectors, and cell therapy are attractive areas of research that continue to be investigated. The significance and role of phage recombination systems in biotechnology is reviewed in this paper, with specific focus on homologous and site-specific recombination conferred by the coli phages, λ, and N15, the integrase from the Streptomyces phage, ΦC31, the recombination system of phage P1, and the recently characterized recombination functions of Yersinia phage, PY54. Key steps of the molecular mechanisms involving phage recombination functions and their application to molecular engineering, our novel exploitations of the PY54-derived recombination system, and its application to the development of new DNA vectors are discussed.

  19. Bacteriophage recombination systems and biotechnical applications.

    Science.gov (United States)

    Nafissi, Nafiseh; Slavcev, Roderick

    2014-04-01

    Bacteriophage recombination systems have been widely used in biotechnology for modifying prokaryotic species, for creating transgenic animals and plants, and more recently, for human cell gene manipulation. In contrast to homologous recombination, which benefits from the endogenous recombination machinery of the cell, site-specific recombination requires an exogenous source of recombinase in mammalian cells. The mechanism of bacteriophage evolution and their coexistence with bacterial cells has become a point of interest ever since bacterial viruses' life cycles were first explored. Phage recombinases have already been exploited as valuable genetic tools and new phage enzymes, and their potential application to genetic engineering and genome manipulation, vectorology, and generation of new transgene delivery vectors, and cell therapy are attractive areas of research that continue to be investigated. The significance and role of phage recombination systems in biotechnology is reviewed in this paper, with specific focus on homologous and site-specific recombination conferred by the coli phages, λ, and N15, the integrase from the Streptomyces phage, ΦC31, the recombination system of phage P1, and the recently characterized recombination functions of Yersinia phage, PY54. Key steps of the molecular mechanisms involving phage recombination functions and their application to molecular engineering, our novel exploitations of the PY54-derived recombination system, and its application to the development of new DNA vectors are discussed. PMID:24442504

  20. Differential Contributions of Intrinsic and Extrinsic Pathways to Thrombin Generation in Adult, Maternal and Cord Plasma Samples

    Science.gov (United States)

    Rice, Nicklaus T.; Szlam, Fania; Varner, Jeffrey D.; Bernstein, Peter S.; Szlam, Arthur D.; Tanaka, Kenichi A.

    2016-01-01

    Background Thrombin generation (TG) is a pivotal process in achieving hemostasis. Coagulation profiles during pregnancy and early neonatal period are different from that of normal (non-pregnant) adults. In this ex vivo study, the differences in TG in maternal and cord plasma relative to normal adult plasma were studied. Methods Twenty consented pregnant women and ten consented healthy adults were included in the study. Maternal and cord blood samples were collected at the time of delivery. Platelet-poor plasma was isolated for the measurement of TG. In some samples, anti-FIXa aptamer, RB006, or a TFPI inhibitor, BAX499 were added to elucidate the contribution of intrinsic and extrinsic pathway to TG. Additionally, procoagulant and inhibitor levels were measured in maternal and cord plasma, and these values were used to mathematically simulate TG. Results Peak TG was increased in maternal plasma (393.6±57.9 nM) compared to adult and cord samples (323.2±38.9 nM and 209.9±29.5 nM, respectively). Inhibitory effects of RB006 on TG were less robust in maternal or cord plasma (52% vs. 12% respectively) than in adult plasma (81%). Likewise the effectiveness of BAX499 as represented by the increase in peak TG was much greater in adult (21%) than in maternal (10%) or cord plasma (12%). Further, BAX499 was more effective in reversing RB006 in adult plasma than in maternal or cord plasma. Ex vivo data were reproducible with the results of the mathematical simulation of TG. Conclusion Normal parturient plasma shows a large intrinsic pathway reserve for TG compared to adult and cord plasma, while TG in cord plasma is sustained by extrinsic pathway, and low levels of TFPI and AT. PMID:27196067

  1. Meta-analysis of randomized controlled trials on risk of myocardial infarction from the use of oral direct thrombin inhibitors.

    Science.gov (United States)

    Artang, Ramin; Rome, Eric; Nielsen, Jørn Dalsgaard; Vidaillet, Humberto J

    2013-12-15

    Dabigatran has been associated with greater risk of myocardial infarction (MI) than warfarin. It is unknown whether the increased risk is unique to dabigatran, an adverse effect shared by other oral direct thrombin inhibitors (DTIs), or the result of a protective effect of warfarin against MI. To address these questions, we systematically searched MEDLINE and performed a meta-analysis on randomized trials that compared oral DTIs with warfarin for any indication with end point of MIs after randomization. We furthermore performed a secondary meta-analysis on atrial fibrillation stroke prevention trials with alternative anticoagulants compared with warfarin with end point of MIs after randomization. A total of 11 trials (39,357 patients) that compared warfarin to DTIs (dabigatran, ximelagatran, and AZD0837) were identified. In these trials, patients treated with oral DTIs were more likely to experience an MI than their counterparts treated with warfarin (285 of 23,333 vs 133 of 16,024, odds ratio 1.35, 95% confidence interval 1.10 to 1.66, p = 0.005). For secondary analysis, 8 studies (69,615 patients) were identified that compared warfarin with alternative anticoagulant including factor Xa inhibitors, DTIs, aspirin, and clopidogrel. There was no significant advantage in the rate of MIs with the use of warfarin versus comparators (odds ratio 1.06, 95% confidence interval 0.85 to 1.34, p = 0.59). In conclusion, our data suggest that oral DTIs were associated with increased risk of MI. This increased risk appears to be a class effect of these agents, not a specific phenomenon unique to dabigatran or protective effect of warfarin. These findings support the need for enhanced postmarket surveillance of oral DTIs and other novel agents. PMID:24075284

  2. Inhibitor of thrombin, bivalirudin%凝血酶抑制剂比伐卢定的临床应用

    Institute of Scientific and Technical Information of China (English)

    何燕; 王良友

    2013-01-01

    比伐卢定是2000年被FDA批准上市的抗凝血药物,它通过直接抑制凝血酶发挥抗凝作用,已被批准应用于PCI/PTCA、HIT/HITTS、ACS等领域.现有研究表明比伐卢定在临床抗凝治疗中有较好的安全性,可以用于常规治疗方案(肝素与糖蛋白Ⅱb/Ⅲa抑制剂)的替代治疗,而且可以降低出血风险和死亡率.本文主要介绍凝血酶直接抑制剂比伐卢定的药理作用及临床研究,总结分析其安全性及经济效益,并且结合国内临床应用情况,分析其在国内的临床运用前景.%Bivalirudin was approved by US Food and Drug Administration as a new direct inhibitor of thrombin in 2000, the drug was currently approved the application to PCI / PTCA, HIT / HITTS, ACS etc. Existing research shows bivalirudin have a better security and can be used for the classic treatment regimens ( hep-arin and glycoprotein Ⅱb/Ⅲa inhibitors ) alternative treatment, Moreover, it can reduce the risk of bleeding and mortality. This paper analy- sis the mechanism of action, pharmacokinetics, pharmacodynamics, clinical application, safety and cost-effectiveness of bivalirudin. But it limited in application in the domestic market, thus strengthening the clinical trails research is promising.

  3. Differential Contributions of Intrinsic and Extrinsic Pathways to Thrombin Generation in Adult, Maternal and Cord Plasma Samples.

    Directory of Open Access Journals (Sweden)

    Nicklaus T Rice

    Full Text Available Thrombin generation (TG is a pivotal process in achieving hemostasis. Coagulation profiles during pregnancy and early neonatal period are different from that of normal (non-pregnant adults. In this ex vivo study, the differences in TG in maternal and cord plasma relative to normal adult plasma were studied.Twenty consented pregnant women and ten consented healthy adults were included in the study. Maternal and cord blood samples were collected at the time of delivery. Platelet-poor plasma was isolated for the measurement of TG. In some samples, anti-FIXa aptamer, RB006, or a TFPI inhibitor, BAX499 were added to elucidate the contribution of intrinsic and extrinsic pathway to TG. Additionally, procoagulant and inhibitor levels were measured in maternal and cord plasma, and these values were used to mathematically simulate TG.Peak TG was increased in maternal plasma (393.6±57.9 nM compared to adult and cord samples (323.2±38.9 nM and 209.9±29.5 nM, respectively. Inhibitory effects of RB006 on TG were less robust in maternal or cord plasma (52% vs. 12% respectively than in adult plasma (81%. Likewise the effectiveness of BAX499 as represented by the increase in peak TG was much greater in adult (21% than in maternal (10% or cord plasma (12%. Further, BAX499 was more effective in reversing RB006 in adult plasma than in maternal or cord plasma. Ex vivo data were reproducible with the results of the mathematical simulation of TG.Normal parturient plasma shows a large intrinsic pathway reserve for TG compared to adult and cord plasma, while TG in cord plasma is sustained by extrinsic pathway, and low levels of TFPI and AT.

  4. Statins but not aspirin reduce thrombotic risk assessed by thrombin generation in diabetic patients without cardiovascular events: the RATIONAL trial.

    Directory of Open Access Journals (Sweden)

    Alejandro Macchia

    Full Text Available BACKGROUND: The systematic use of aspirin and statins in patients with diabetes and no previous cardiovascular events is controversial. We sought to assess the effects of aspirin and statins on the thrombotic risk assessed by thrombin generation (TG among patients with type II diabetes mellitus and no previous cardiovascular events. METHODOLOGY/PRINCIPAL FINDINGS: Prospective, randomized, open, blinded to events evaluation, controlled, 2×2 factorial clinical trial including 30 patients randomly allocated to aspirin 100 mg/d, atorvastatin 40 mg/d, both or none. Outcome measurements included changes in TG levels after treatment (8 to 10 weeks, assessed by a calibrated automated thrombogram. At baseline all groups had similar clinical and biochemical profiles, including TG levels. There was no interaction between aspirin and atorvastatin. Atorvastatin significantly reduced TG measured as peak TG with saline (85.09±55.34 nmol vs 153.26±75.55 nmol for atorvastatin and control groups, respectively; p = 0.018. On the other hand, aspirin had no effect on TG (121.51±81.83 nmol vs 116.85±67.66 nmol, for aspirin and control groups, respectively; p = 0.716. The effects of treatments on measurements of TG using other agonists were consistent. CONCLUSIONS/SIGNIFICANCE: While waiting for data from ongoing large clinical randomized trials to definitively outline the role of aspirin in primary prevention, our study shows that among diabetic patients without previous vascular events, statins but not aspirin reduce thrombotic risk assessed by TG. TRIAL REGISTRATION: ClinicalTrials.gov NCT00793754.

  5. Recombinant factor XIII and congenital factor XIII deficiency: an update from human and animal studies

    Directory of Open Access Journals (Sweden)

    Inbal A

    2013-10-01

    Full Text Available Aida InbalThrombosis and Hemostasis Unit, Hematology Institute, Beilinson Hospital, Rabin Medical Center, Petach Tikva, and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, IsraelAbstract: Factor XIII (FXIII is a protransglutaminase composed of two catalytic A subunits and two carrier B subunits. An intracellular form of FXIII is present in monocytes/macrophages and platelets as a homodimer of two A subunits. Following activation by thrombin, FXIII becomes plasma transglutaminase, which crosslinks γ-glutamyl-ε-lysine residues of fibrin chains and thereby stabilizes the fibrin clot. FXIII deficiency results in a moderate to severe hemorrhagic disorder, abnormal wound healing in about 30% of patients, and recurrent abortion in homozygous females. More than 800 cases of FXIII deficiency have been reported, most of them due to mutation in the FXIII-A gene, resulting in FXIII-A deficiency. Among mutations causing FXIII-A deficiency, 50% are missense mutations. Only 16 mutations in the FXIII-B gene have been published. Routine laboratory tests are normal in patients with FXIII deficiency, and the diagnosis is established by demonstration of decreased FXIII activity and antigen. Plasma-derived, virus-inactivated factor XIII concentrate is the treatment of choice. The low plasma levels of FXIII (about 5% required to control bleeding and its long half-life make monthly prophylactic therapy feasible. Recently, recombinant FXIII concentrate with a half-life similar to that of native FXIII has been developed and tested in a multinational clinical study. This new product appears to be safe and appropriate for lifelong prophylactic treatment of patients with FXIII-A deficiency.Keywords: recombinant FXIII concentrate, FXIII deficiency

  6. X-ray spectroscopy diagnostics of a recombining plasma in laboratory astrophysics studies

    Science.gov (United States)

    Ryazantsev, S. N.; Skobelev, I. Yu.; Faenov, A. Ya.; Pikuz, T. A.; Grum-Grzhimailo, A. N.; Pikuz, S. A.

    2015-12-01

    The investigation of a recombining laser plasma is topical primarily because it can be used to simulate the interaction between plasma jets in astrophysical objects. It has been shown that the relative intensities of transitions of a resonance series of He-like multicharged ions can be used for the diagnostics of the recombining plasma. It has been found that the intensities of the indicated transitions for ions with the nuclear charge number Z n ~ 10 are sensitive to the plasma density in the range N e ~ 1016-1020 cm-3 at temperatures of 10-100 eV. The calculations performed for the F VIII ion have determined the parameters of plasma jets created at the ELFIE nanosecond laser facility (Ecole Polytechnique, France) in order to simulate astrophysical phenomena. The resulting universal calculation dependences can be used to diagnose different recombining plasmas containing helium-like fluorine ions.

  7. CRMAGE: CRISPR Optimized MAGE Recombineering

    DEFF Research Database (Denmark)

    Ronda, Carlotta; Pedersen, Lasse Ebdrup; Sommer, Morten Otto Alexander;

    2016-01-01

    A bottleneck in metabolic engineering and systems biology approaches is the lack of efficient genome engineering technologies. Here, we combine CRISPR/Cas9 and λ Red recombineering based MAGE technology (CRMAGE) to create a highly efficient and fast method for genome engineering of Escherichia coli...... that are assembled by a USER-cloning approach enabling quick and cost efficient gRNA replacement. CRMAGE furthermore utilizes CRISPR/Cas9 for efficient plasmid curing, thereby enabling multiple engineering rounds per day. To facilitate the design process, a web-based tool was developed to predict both the λ Red...

  8. Environmental Health Topics from A to Z

    Science.gov (United States)

    ... Topics Environmental Agents Acrylamide Air Pollution Allergens & Irritants Aloe Vera Arsenic Bisphenol A (BPA) Cell Phones Climate ... Cigarette Smoke Cockroaches Dust Mites Pets & Animals Pollen Aloe Vera Alternatives to Animal Testing Arsenic Asthma Autism ...

  9. MedlinePlus Health Topic Web Service

    Data.gov (United States)

    U.S. Department of Health & Human Services — A search-based Web service that provides access to disease, condition and wellness information via MedlinePlus health topic data in XML format. The service accepts...

  10. MedlinePlus Health Topic XML Files

    Data.gov (United States)

    U.S. Department of Health & Human Services — XML files containing information resources on diseases, conditions and wellness issues. MedlinePlus produces three types of health topic XML files – MedlinePlus...

  11. Mining Concurrent Topical Activity in Microblog Streams

    CERN Document Server

    Panisson, A; Quaggiotto, M; Cattuto, C

    2014-01-01

    Streams of user-generated content in social media exhibit patterns of collective attention across diverse topics, with temporal structures determined both by exogenous factors and endogenous factors. Teasing apart different topics and resolving their individual, concurrent, activity timelines is a key challenge in extracting knowledge from microblog streams. Facing this challenge requires the use of methods that expose latent signals by using term correlations across posts and over time. Here we focus on content posted to Twitter during the London 2012 Olympics, for which a detailed schedule of events is independently available and can be used for reference. We mine the temporal structure of topical activity by using two methods based on non-negative matrix factorization. We show that for events in the Olympics schedule that can be semantically matched to Twitter topics, the extracted Twitter activity timeline closely matches the known timeline from the schedule. Our results show that, given appropriate techn...

  12. A Survey On Interactivity in Topic Models

    Directory of Open Access Journals (Sweden)

    Patrik Ehrencrona Kjellin

    2016-04-01

    Full Text Available Trying to make sense and gain deeper insight from large sets of data is becoming a task very central to computer science in general. Topic models, capable of uncovering the semantic themes pervading through large collections of documents, have seen a surge in popularity in recent years. However, topic models are high level statistical tools; their output is given in terms of probability distributions, suited neither for simple interpretation nor deep analysis. Interpreting the fitted topic models in an intuitive manner requires visual and interactive tools. Additionally, some measure of human interaction is typically required for refining the output offered by such models. In the research, this area remains relatively unexplored – only recently has this aspect been receiving more attention. In this paper, the literature is surveyed as it pertains to interactivity and visualisation within the context of topic models, with the goal of finding current research trends in this area.

  13. Efficacious and orally bioavailable thrombin inhibitors based on a 2,5-thienylamidine at the P1 position: discovery of N-carboxymethyl-d-diphenylalanyl-l-prolyl[(5-amidino-2-thienyl)methyl]amide.

    Science.gov (United States)

    Lee, Koo; Park, Cheol Won; Jung, Won-Hyuk; Park, Hee Dong; Lee, Sun Hwa; Chung, Kyung Ha; Park, Su Kyung; Kwon, O Hwan; Kang, Myunggyun; Park, Doo-Hee; Lee, Sang Koo; Kim, Eunice E; Yoon, Suk Kyoon; Kim, Aeri

    2003-08-14

    Thrombin, a crucial enzyme in the blood coagulation, has been a target for antithrombotic therapy. Orally active thrombin inhibitors would provide effective and safe prophylaxis for venous and arterial thrombosis. We conducted optimization of a highly efficacious benzamidine-based thrombin inhibitor LB30812 (3, K(i) = 3 pM) to improve oral bioavailability. Of a variety of arylamidines investigated at the P1 position, 2,5-thienylamidine effectively replaced the benzamidine without compromising the thrombin inhibitory potency and oral absorption. The sulfamide and sulfonamide derivatization at the N-terminal position in general afforded highly potent thrombin inhibitors but with moderate oral absorption, while the well-absorbable N-carbamate derivatives exhibited limited metabolic stability in S9 fractions. The present work culminated in the discovery of the N-carboxymethyl- and 2,5-thienylamidine-containing compound 22 that exhibits the most favorable profiles of anticoagulant and antithrombotic activities as well as oral bioavilability (K(i) = 15 pM; F = 43%, 42%, and 15% in rats, dogs, and monkeys, respectively). This compound on a gravimetric basis was shown to be more effective than a low molecular weight heparin, enoxaparin, in the venous thrombosis models of rat and rabbit. Compound 22 (LB30870) was therefore selected for further preclinical and clinical development. PMID:12904065

  14. Recent Advances In Topical Therapy In Dermatology

    OpenAIRE

    Mohan Thappa Devinder

    2003-01-01

    With changing times various newer topical agents are introduced in the field of dermatology. Tacrolimus and pimecrolimus are immunisuppressants, which are effective topically and are tried in the management of atopic dermatitis as well as other disorders including allergic contact dermatitis, atrophic lichen planus, pyoderma gangrenosum. Imiquimod, an immune response modifier, is presently in use for genital warts but has potentials as anti- tumour agent and in various other dermatological co...

  15. Topical calcineurin inhibitors in systemic lupus erythematosus

    OpenAIRE

    Lampropoulos, Christos

    2010-01-01

    Christos E Lampropoulos, David P D’CruzLupus Research Unit, Rayne Institute, St. Thomas’ Hospital, London, UKAbstract: Cutaneous lupus erythematosus (CLE) encompasses a variety of lesions that may be refractory to systemic or topical agents. Discoid lupus erythematosus (DLE) and subacute cutaneous lupus erythematosus (SCLE) are the most common lesions in clinical practice. The topical calcineurin inhibitors, tacrolimus and pimecrolimus, have been used to treat resistant cu...

  16. The rational design of topical formulations

    OpenAIRE

    Duszynska-Krupa, A. M.

    2015-01-01

    This thesis addresses the development of topical formulations designed to treat atopic dermatitis (AD) using nicotinamide (NA). A rational approach to the development of topical formulations based on the physical and chemical properties of the drug and vehicle components is studied. This approach is an alternative to the model of formulation development where the drug is added into an existing vehicle without optimisation of the formulation in terms of the active delivery to its site of actio...

  17. Formulation, pharmacokinetics and pharmacodynamics of topical microbicides

    OpenAIRE

    Adams, Jessica L.; Kashuba, Angela D. M.

    2012-01-01

    The development of safe topical microbicides that effectively prevent human immunodeficiency virus (HIV) infection is a major goal in curbing the human immunodeficiency virus pandemic. A number of past failures resulting from mucosal toxicity or lack of efficacy have informed the field. Products that caused toxicity to the female genital tract mucosa, and thereby increased the likelihood of HIV acquisition, included nonoxynol 9, cellulose sulfate, and C31 G vaginal gel Savvy®. Topical product...

  18. Topical and oral therapeutic approach to rosacea.

    Science.gov (United States)

    Helfrich, Yolanda R; Maier, Lisa M

    2016-06-01

    Rosacea is an inflammatory condition of the skin, primarily affecting the central convexities of the face. Various topical and oral therapeutic approaches exist. Most have been developed to treat the papulopustular subtype of rosacea; however, other approaches can be used to treat the erythematotelangiectatic, ocular, and phymatous subtypes. This review provides a summary of available topical and oral approaches for the treatment of rosacea. PMID:27416312

  19. Pharmacogenetics of ophthalmic topical β-blockers

    OpenAIRE

    Sidjanin, Duska J; Catherine A McCarty; Patchett, Richard; Smith, Edward; Wilke, Russell A

    2008-01-01

    Glaucoma is the second leading cause of blindness worldwide. The primary glaucoma risk factor is elevated intraocular pressure. Topical β-blockers are affordable and widely used to lower intraocular pressure. Genetic variability has been postulated to contribute to interpersonal differences in efficacy and safety of topical β-blockers. This review summarizes clinically significant polymorphisms that have been identified in the β-adrenergic receptors (ADRB1, ADRB2 and ADRB3). The implications ...

  20. Topical calcineurin inhibitors in systemic lupus erythematosus

    Directory of Open Access Journals (Sweden)

    Christos E Lampropoulos

    2010-04-01

    Full Text Available Christos E Lampropoulos, David P D’CruzLupus Research Unit, Rayne Institute, St. Thomas’ Hospital, London, UKAbstract: Cutaneous lupus erythematosus (CLE encompasses a variety of lesions that may be refractory to systemic or topical agents. Discoid lupus erythematosus (DLE and subacute cutaneous lupus erythematosus (SCLE are the most common lesions in clinical practice. The topical calcineurin inhibitors, tacrolimus and pimecrolimus, have been used to treat resistant cutaneous lupus since 2002 and inhibit the proliferation and activation of T-cells and suppress immune-mediated cutaneous inflammation. This article reviews the mechanism of action, efficacy, adverse effects, and the recent concern about their possible carcinogenic effect. Although the total number of patients is small and there is only one relevant randomized controlled study, the data are encouraging. Many patients, previously resistant to systemic agents or topical steroids, improved after four weeks of treatment. DLE and SCLE lesions were less responsive, reflecting the chronicity of the lesions, although more than 50% of patients still showed improvement. Topical calcineurin inhibitors may be a safe and effective alternative to topical steroids for CLE although the only approved indication is for atopic dermatitis.Keywords: tacrolimus, pimecrolimus, cutaneous lupus erythematosus, topical calcineurin inhibitors