Treatment of proximal deep vein thrombosis with a novel synthetic compound (SR90107A/ORG31540) with pure anti-factor Xa activity - A phase II evaluation

NARCIS (Netherlands)

Buller, HR; Cariou, R; Gallus, A; Gent, M; Ginsberg, J; Prins, MH; Lensing, A; Levi, M; Nurmohammed, N; Hirsh, J; Roberts, R; ten Cate, JW; Decousus, H; Mismetti, P; Buchmuller, A; Charlet, [No Value; Viallon, A; van der Meer, J; Meinardi, [No Value; Meijer, K; Piovella, F; Barone, M; Beltrametti, C; Serafini, S; Kraaijenhagen, RA; Koopman, MMW; Jagt, HHT; Muller, MBR; Kooy, MV; Nauta, S; Gallus, AS; Coghlan, D; Rich, C; Prandoni, P; Scudeller, A; Scarano, L; Girolami, A; Baker, R; Tan, E; Cooney, J; Eikelboom, J; Ninet, J; Dolmazon, C; Madoux, MHG; Coppere, B; Nenci, GG; Agnelli, G; Falcinelli, F; Morini, M; d'Angelo, A

2000-01-01

Background-Patients with venous thromboembolism require initial treatment with an immediate-acting anticoagulant, low-molecular-weight heparin. We evaluated a novel synthetic factor Xa inhibitor (SR90107a/ORG31540) as an alternative treatment. Methods and Results-A randomized-parallel-group, phase

In vivo studies on the binding of heparin and its fractions with platelet factor 4

International Nuclear Information System (INIS)

Walz, D.A.; Hung, G.L.

1985-01-01

PF4 has a half-life in plasma of less than 3 minutes, and its rapid clearance appears to be a function of binding to the vascular endothelium. Once bound to the endothelium, PF4 can be released by heparin in a time-dependent manner; recovery is greater the sooner heparin is administered following PF4 infusion. This heparin-induced release of PF4 can be abolished if the heparin is first complexed with hexadimethrine bromide. Likewise, this heparin-induced release of PF4 is dependent upon the type of heparin used; low molecular weight heparin fractions and fragments do not cause the PF4 rebound seen with intact heparin. Thus, it would appear that low molecular weight forms of heparin are advantageous in that their in vivo administration would not be mediated by such platelet modulators as PF4

Structural and binding studies of SAP-1 protein with heparin.

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Yadav, Vikash K; Mandal, Rahul S; Puniya, Bhanwar L; Kumar, Rahul; Dey, Sharmistha; Singh, Sarman; Yadav, Savita

2015-03-01

SAP-1 is a low molecular weight cysteine protease inhibitor (CPI) which belongs to type-2 cystatins family. SAP-1 protein purified from human seminal plasma (HuSP) has been shown to inhibit cysteine and serine proteases and exhibit interesting biological properties, including high temperature and pH stability. Heparin is a naturally occurring glycosaminoglycan (with varied chain length) which interacts with a number of proteins and regulates multiple steps in different biological processes. As an anticoagulant, heparin enhances inhibition of thrombin by the serpin antithrombin III. Therefore, we have employed surface plasmon resonance (SPR) to improve our understanding of the binding interaction between heparin and SAP-1 (protease inhibitor). SPR data suggest that SAP-1 binds to heparin with a significant affinity (KD = 158 nm). SPR solution competition studies using heparin oligosaccharides showed that the binding of SAP-1 to heparin is dependent on chain length. Large oligosaccharides show strong binding affinity for SAP-1. Further to get insight into the structural aspect of interactions between SAP-1 and heparin, we used modelled structure of the SAP-1 and docked with heparin and heparin-derived polysaccharides. The results suggest that a positively charged residue lysine plays important role in these interactions. Such information should improve our understanding of how heparin, present in the reproductive tract, regulates cystatins activity. © 2014 John Wiley & Sons A/S.

Changes in heparin dose response slope during cardiac surgery: possible result in inaccuracy in predicting heparin bolus dose requirement to achieve target ACT.

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Ichikawa, Junko; Mori, Tetsu; Kodaka, Mitsuharu; Nishiyama, Keiko; Ozaki, Makoto; Komori, Makiko

2017-09-01

The substantial interpatient variability in heparin requirement has led to the use of a heparin dose response (HDR) technique. The accuracy of Hepcon-based heparin administration in achieving a target activated clotting time (ACT) using an HDR slope remains controversial. We prospectively studied 86 adult patients scheduled for cardiac surgery requiring cardiopulmonary bypass. The total dose of calculated heparin required for patient and pump priming was administered simultaneously to achieve a target ACT of 450 s for HDR on the Hepcon HMS system. Blood samples were obtained after the induction of anesthesia, at 3 min after heparin administration and after the initiation of CPB to measure kaolin ACT, HDR slope, whole-blood heparin concentration based on the HDR slope and anti-Xa heparin concentration, antithrombin and complete blood count. The target ACT of 450 s was not achieved in 68.6% of patients. Compared with patients who achieved the target ACT, those who failed to achieve their target ACT had a significantly higher platelet count at baseline. Correlation between the HDR slope and heparin sensitivity was poor. Projected heparin concentration and anti-Xa heparin concentration are not interchangeable based on the Bland-Altman analysis. It can be hypothesized that the wide discrepancy in HDR slope versus heparin sensitivity may be explained by an inaccurate prediction of the plasma heparin level and/or the change in HDR of individual patients, depending on in vivo factors such as extravascular sequestration of heparin, decreased intrinsic antithrombin activity level and platelet count and/or activity.

First steps in the direction of synthetic, allosteric, direct inhibitors of thrombin and factor Xa.

Science.gov (United States)

Verghese, Jenson; Liang, Aiye; Sidhu, Preet Pal Singh; Hindle, Michael; Zhou, Qibing; Desai, Umesh R

2009-08-01

Designing non-saccharide functional mimics of heparin is a major challenge. In this work, a library of small, aromatic molecules based on the sulfated DHP scaffold was synthesized and screened against thrombin and factor Xa. The results reveal that (i) selected monomeric benzofuran derivatives inhibit the two enzymes, albeit weakly; (ii) the two enzymes recognize different structural features in the benzofurans studied suggesting significant selectivity of recognition; and (iii) the mechanism of inhibition is allosteric. The molecules represent the first allosteric small molecule inhibitors of the two enzymes.

First Steps in the Direction of Synthetic, Allosteric, Direct Inhibitors of Thrombin and Factor Xa

Science.gov (United States)

Verghese, Jenson; Liang, Aiye; Sidhu, Preet Pal Singh; Hindle, Michael; Zhou, Qibing; Desai, Umesh R.

2009-01-01

Designing non-saccharide functional mimics of heparin is a major challenge. In this work, a library of small, aromatic molecules based on the sulfated DHP scaffold was synthesized and screened against thrombin and factor Xa. The results reveal that i) selected monomeric benzofuran derivatives inhibit the two enzymes, albeit weakly; ii) the two enzymes recognize different structural features in the benzofurans studied suggesting significant selectivity of recognition; and iii) the mechanism of inhibition is allosteric. The molecules represent the first allosteric small molecule inhibitors of the two enzymes. PMID:19540113

Human IGF-I propeptide A promotes articular chondrocyte biosynthesis and employs glycosylation-dependent heparin binding.

Science.gov (United States)

Shi, Shuiliang; Kelly, Brian J; Wang, Congrong; Klingler, Ken; Chan, Albert; Eckert, George J; Trippel, Stephen B

2018-03-01

Insulin-like growth factor I (IGF-I) is a key regulator of chondrogenesis, but its therapeutic application to articular cartilage damage is limited by rapid elimination from the repair site. The human IGF-I gene gives rise to three IGF-I propeptides (proIGF-IA, proIGF-IB and proIGF-IC) that are cleaved to create mature IGF-I. In this study, we elucidate the processing of IGF-I precursors by articular chondrocytes, and test the hypotheses that proIGF-I isoforms bind to heparin and regulate articular chondrocyte biosynthesis. Human IGF-I propeptides and mutants were overexpressed in bovine articular chondrocytes. IGF-I products were characterized by ELISA, western blot and FPLC using a heparin column. The biosynthetic activity of IGF-I products on articular chondrocytes was assayed for DNA and glycosaminoglycan that the cells produced. Secreted IGF-I propeptides stimulated articular chondrocyte biosynthetic activity to the same degree as mature IGF-I. Of the three IGF-I propeptides, only one, proIGF-IA, strongly bound to heparin. Interestingly, heparin binding of proIGF-IA depended on N-glycosylation at Asn92 in the EA peptide. To our knowledge, this is the first demonstration that N-glycosylation determines the binding of a heparin-binding protein to heparin. The biosynthetic and heparin binding abilities of proIGF-IA, coupled with its generation of IGF-I, suggest that proIGF-IA may have therapeutic value for articular cartilage repair. These data identify human pro-insulin-like growth factor IA as a bifunctional protein. Its combined ability to bind heparin and augment chondrocyte biosynthesis makes it a promising therapeutic agent for cartilage damage due to trauma and osteoarthritis. Copyright © 2017 Elsevier B.V. All rights reserved.

Metabolic engineering of Chinese hamster ovary cells: towards a bioengineered heparin.

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Baik, Jong Youn; Gasimli, Leyla; Yang, Bo; Datta, Payel; Zhang, Fuming; Glass, Charles A; Esko, Jeffrey D; Linhardt, Robert J; Sharfstein, Susan T

2012-03-01

Heparin is the most widely used pharmaceutical to control blood coagulation in modern medicine. A health crisis that took place in 2008 led to a demand for production of heparin from non-animal sources. Chinese hamster ovary (CHO) cells, commonly used mammalian host cells for production of foreign pharmaceutical proteins in the biopharmaceutical industry, are capable of producing heparan sulfate (HS), a related polysaccharide naturally. Since heparin and HS share the same biosynthetic pathway, we hypothesized that heparin could be produced in CHO cells by metabolic engineering. Based on the expression of endogenous enzymes in the HS/heparin pathways of CHO-S cells, human N-deacetylase/N-sulfotransferase (NDST2) and mouse heparan sulfate 3-O-sulfotransferase 1 (Hs3st1) genes were transfected sequentially into CHO host cells growing in suspension culture. Transfectants were screened using quantitative RT-PCR and Western blotting. Out of 120 clones expressing NDST2 and Hs3st1, 2 clones, Dual-3 and Dual-29, were selected for further analysis. An antithrombin III (ATIII) binding assay using flow cytometry, designed to recognize a key sugar structure characteristic of heparin, indicated that Hs3st1 transfection was capable of increasing ATIII binding. An anti-factor Xa assay, which affords a measure of anticoagulant activity, showed a significant increase in activity in the dual-expressing cell lines. Disaccharide analysis of the engineered HS showed a substantial increase in N-sulfo groups, but did not show a pattern consistent with pharmacological heparin, suggesting that further balancing the expression of transgenes with the expression levels of endogenous enzymes involved in HS/heparin biosynthesis might be necessary. Copyright © 2012 Elsevier Inc. All rights reserved.

Heparin-associated thrombocytopenia: antibody binding specificity to platelet antigens.

Science.gov (United States)

Lynch, D M; Howe, S E

1985-11-01

Sera from four patients with heparin-associated thrombocytopenia (HAT) were evaluated by a quantitative enzyme-linked immunosorbent assay (ELISA) to detect heparin-dependent serum platelet-bindable immunoglobulin (S-PBIg) and by Western blotting and immunoprecipitation to investigate the specificity of the antibody binding. All HAT sera showed mildly increased S-PBIg (mean, 7.8 fg per platelet; normal, less than 6.0 fg per platelet) to intact target platelets in the ELISA, which was markedly increased in the presence of heparin (mean, 20.9 fg per platelet). This increase was 20-fold greater than normal control sera, which showed a mean differential increase of only 0.5 fg per platelet. Immunoglobulin binding specificity to platelet antigens was investigated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis of platelet lysate with transfer of the platelet fractions onto nitrocellulose strips (Western blotting) and subsequent immunoassay using HAT and normal sera. In the presence of heparin, the four HAT patients demonstrated increased binding of immunoglobulin to platelet antigens of apparent molecular weights of 180, 124, and 82 kd. Radiolabeled heparin when incubated with HAT sera, normal sera, or albumin blanks bound to platelet proteins of the same apparent molecular weights. These observations are consistent with current hypotheses suggesting that HAT antibody is directed to heparin-platelet complexes or, alternatively, that heparin induces conformational change of antigenic sites on the platelet membrane.

Quantitative description of thermodynamic and kinetic properties of the platelet factor 4/heparin bonds

Science.gov (United States)

Nguyen, Thi-Huong; Greinacher, Andreas; Delcea, Mihaela

2015-05-01

Heparin is the most important antithrombotic drug in hospitals. It binds to the endogenous tetrameric protein platelet factor 4 (PF4) forming PF4/heparin complexes which may cause a severe immune-mediated adverse drug reaction, so-called heparin-induced thrombocytopenia (HIT). Although new heparin drugs have been synthesized to reduce such a risk, detailed bond dynamics of the PF4/heparin complexes have not been clearly understood. In this study, single molecule force spectroscopy (SMFS) is utilized to characterize the interaction of PF4 with heparins of defined length (5-, 6-, 8-, 12-, and 16-mers). Analysis of the force-distance curves shows that PF4/heparin binding strength rises with increasing heparin length. In addition, two binding pathways in the PF4/short heparins (=8-mers) are identified. We provide a model for the PF4/heparin complexes in which short heparins bind to one PF4 tetramer, while long heparins bind to two PF4 tetramers. We propose that the interaction between long heparins and PF4s is not only due to charge differences as generally assumed, but also due to hydrophobic interaction between two PF4s which are brought close to each other by long heparin. This complicated interaction induces PF4/heparin complexes more stable than other ligand-receptor interactions. Our results also reveal that the boundary between antigenic and non-antigenic heparins is between 8- and 12-mers. These observations are particularly important to understand processes in which PF4-heparin interactions are involved and to develop new heparin-derived drugs.Heparin is the most important antithrombotic drug in hospitals. It binds to the endogenous tetrameric protein platelet factor 4 (PF4) forming PF4/heparin complexes which may cause a severe immune-mediated adverse drug reaction, so-called heparin-induced thrombocytopenia (HIT). Although new heparin drugs have been synthesized to reduce such a risk, detailed bond dynamics of the PF4/heparin complexes have not been clearly

Click-coated, heparinized, decellularized vascular grafts.

Science.gov (United States)

Dimitrievska, Sashka; Cai, Chao; Weyers, Amanda; Balestrini, Jenna L; Lin, Tylee; Sundaram, Sumati; Hatachi, Go; Spiegel, David A; Kyriakides, Themis R; Miao, Jianjun; Li, Guoyun; Niklason, Laura E; Linhardt, Robert J

2015-02-01

A novel method enabling the engineering of a dense and appropriately oriented heparin-containing layer on decellularized aortas has been developed. Amino groups of decellularized aortas were first modified to azido groups using 3-azidobenzoic acid. Azide-clickable dendrons were attached onto the azido groups through "alkyne-azide" click chemistry, affording a tenfold amplification of adhesions sites. Dendron end groups were finally decorated with end-on modified heparin chains. Heparin chains were oriented like heparan sulfate groups on native endothelial cells surface. X-ray photoelectron spectroscopy, nuclear magnetic resonance imaging, mass spectrometry and Fourier transform infrared FTIR spectroscopy were used to characterize the synthesis steps, building the final heparin layered coatings. The continuity of the heparin coating was verified using fluorescent microscopy and histological analysis. The efficacy of heparin linkage was demonstrated with factor Xa anti-thrombogenic assay and platelet adhesion studies. The results suggest that oriented heparin immobilization to decellularized aortas may improve the in vivo blood compatibility of decellularized aortas and vessels. Copyright © 2014 Acta Materialia Inc. All rights reserved.

Improved assay for measuring heparin binding to bull sperm

International Nuclear Information System (INIS)

Miller, D.J.; Ax, R.L.

1988-01-01

The binding of heparin to sperm has been used to study capacitation and to rank relative fertility of bulls. Previous binding assays were laborious, used 10 7 sperm per assay point, and required large amounts of radiolabeled heparin. A modified heparin-binding assay is described that used only 5 x 10 4 cells per incubation well and required reduced amounts of [ 3 H] heparin. The assay was performed in 96-well Millititer plates, enabling easy incubation and filtering. Dissociation constants and concentrations of binding sites did not differ if analyzed by Scatchard plots, Woolf plots, or by log-logit transformed weighted nonlinear least squares regression, except in the case of outliers. In such cases, Scatchard analysis was more sensitive to outliers. Nonspecific binding was insignificant using nonlinear logistic fit regression and a proportion graph. The effects were tested of multiple free-thawing of sperm in either a commercial egg yolk extender, 40 mM Tris buffer with 8% glycerol, or 40 mM Tris buffer without glycerol. Freeze-thawing in extender did not affect the dissociation constant or the concentration of binding sites. However, freeze-thawing three times in 40 mM Tris reduced the concentration of binding sites and lowered the dissociation constant (raised the affinity). The inclusion of glycerol in the 40 mM Tris did not significantly affect the estimated dissociation constant or the concentration of binding sites as compared to 40 mM Tris without glycerol

Heparin-independent, PF4-dependent binding of HIT antibodies to platelets: implications for HIT pathogenesis.

Science.gov (United States)

Padmanabhan, Anand; Jones, Curtis G; Bougie, Daniel W; Curtis, Brian R; McFarland, Janice G; Wang, Demin; Aster, Richard H

2015-01-01

Antibodies specific for platelet factor 4 (PF4)/heparin complexes are the hallmark of heparin-induced thrombocytopenia and thrombosis (HIT), but many antibody-positive patients have normal platelet counts. The basis for this is not fully understood, but it is believed that antibodies testing positive in the serotonin release assay (SRA) are the most likely to cause disease. We addressed this issue by characterizing PF4-dependent binding of HIT antibodies to intact platelets and found that most antibodies testing positive in the SRA, but none of those testing negative, bind to and activate platelets when PF4 is present without any requirement for heparin (P HIT antibodies recognize PF4 in a complex with heparin, only a subset of these antibodies recognize more subtle epitopes induced in PF4 when it binds to CS, the major platelet glycosaminoglycan. Antibodies having this property could explain "delayed HIT" seen in some individuals after discontinuation of heparin and the high risk for thrombosis that persists for weeks in patients recovered from HIT. © 2015 by The American Society of Hematology.

Comparison of established and novel purity tests for the quality control of heparin by means of a set of 177 heparin samples.

Science.gov (United States)

Alban, Susanne; Lühn, Susanne; Schiemann, Simone; Beyer, Tanja; Norwig, Jochen; Schilling, Claudia; Rädler, Oliver; Wolf, Bernhard; Matz, Magnus; Baumann, Knut; Holzgrabe, Ulrike

2011-01-01

The widespread occurrence of heparin contaminated with oversulfated chrondroitin sulfate (OSCS) in 2008 initiated a comprehensive revision process of the Pharmacopoeial heparin monographs and stimulated research in analytical techniques for the quality control of heparin. Here, a set of 177 heparin samples from the market in 2008 as well as pure heparin sodium spiked with defined amounts of OSCS and DS were used to evaluate established and novel methods for the quality control of heparin. Besides (1)H nuclear magnetic resonance spectroscopy (NMR), the assessment included two further spectroscopic methods, i.e., attenuated total reflection-infrared spectroscopy (ATR-IR) and Raman spectroscopy, three coagulation assays, i.e., activated partial thromboplastin time (aPTT) performed with both sheep and human plasma and the prothrombin time (PT), and finally two novel purity assays, each consisting of an incubation step with heparinase I followed by either a fluorescence measurement (Inc-PolyH-assay) or by a chromogenic aXa-assay (Inc-aXa-assay). NMR was shown to allow not only sensitive detection, but also quantification of OSCS by using the peak-height method and a response factor determined by calibration. Chemometric evaluation of the NMR, ATR-IR, and Raman spectra by statistical classification techniques turned out to be best with NMR spectra concerning the detection of OSCS. The validity of the aPTT, the current EP assay, could be considerably improved by replacing the sheep plasma by human plasma. In this way, most of the contaminated heparin samples did not meet the novel potency limit of 180 IU/mg. However, also more than 50% of the uncontaminated samples had interpretation of the results.

Heparin-binding peptide amphiphile supramolecular architectures as platforms for angiogenesis and drug delivery

Science.gov (United States)

Chow, Lesleyann W.

A fascinating phenomenon in nature is the self-assembly of molecules into a functional, hierarchical structure. In the past decade, the Stupp Laboratory has developed several classes of self-assembling biomaterials, one of which is the synthetic peptide amphiphile (PA). Self-assembling PAs are attractive and versatile biomolecules that can be customized for specific applications in regenerative medicine. In particular, a heparin-binding peptide amphiphile (HBPA) containing a specific heparin-binding peptide sequence was used here to induce angiogenesis and serve as a delivery vehicle for growth factors and small hydrophobic molecules. Throughout this dissertation, the HBPA/heparin system is used in different architectures for a variety of regenerative medicine applications. In one aspect of this work, hybrid scaffolds made from HBPA/heparin gelled on a poly(L-lactic acid) (PLLA) fiber mesh were used to promote angiogenesis to facilitate pancreatic islet transplantation for the treatment of type 1 diabetes. Delivery of growth factors with HBPA/PLLA scafflolds increased vessel density in vivo and correlated with improved transplant outcomes in a streptozotocin-induced diabetic mouse model. Soluble HBPA nanofiber architectures were also useful for islet transplantation applications. These nanofibers were used at concentrations below gelation to deliver growth factors into the dense islet cell aggregate, promoting cell survival and angiogenesis in vitro. The nanostructures infiltrated the islets and promoted the retention of heparin and growth factors within the islet. Another interesting growth factor release system discussed here is the HBPA membrane structure. HBPA was found to self-assemble with hyaluronic acid, a large biopolymer found in the body, into macroscopic, hierarchically-ordered membranes. Heparin was incorporated into these membranes and affected the membrane's mechanical properties and growth factor release. Human mesenchymal stem cells were also shown

A Heparin Binding Motif Rich in Arginine and Lysine is the Functional Domain of YKL-40

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Nipaporn Ngernyuang

2018-02-01

Full Text Available The heparin-binding glycoprotein YKL-40 (CHI3L1 is intimately associated with microvascularization in multiple human diseases including cancer and inflammation. However, the heparin-binding domain(s pertinent to the angiogenic activity have yet been identified. YKL-40 harbors a consensus heparin-binding motif that consists of positively charged arginine (R and lysine (K (RRDK; residues 144–147; but they don't bind to heparin. Intriguingly, we identified a separate KR-rich domain (residues 334–345 that does display strong heparin binding affinity. A short synthetic peptide spanning this KR-rich domain successfully competed with YKL-40 and blocked its ability to bind heparin. Three individual point mutations, where alanine (A substituted for K or R (K337A, K342A, R344A, led to remarkable decreases in heparin-binding ability and angiogenic activity. In addition, a neutralizing anti-YKL-40 antibody that targets these residues and prevents heparin binding impeded angiogenesis in vitro. MDA-MB-231 breast cancer cells engineered to express ectopic K337A, K342A or R344A mutants displayed reduced tumor development and compromised tumor vessel formation in mice relative to control cells expressing wild-type YKL-40. These data reveal that the KR-rich heparin-binding motif is the functional heparin-binding domain of YKL-40. Our findings shed light on novel molecular mechanisms underlying endothelial cell angiogenesis promoted by YKL-40 in a variety of diseases.

Relationship of nonreturn rates of dairy bulls to binding affinity of heparin to sperm

International Nuclear Information System (INIS)

Marks, J.L.; Ax, R.L.

1985-01-01

The binding of the glycosaminoglycan [ 3 H] heparin to bull spermatozoa was compared with nonreturn rates of dairy bulls. Semen samples from five bulls above and five below an average 71% nonreturn rate were used. Samples consisted of first and second ejaculates on a single day collected 1 d/wk for up to 5 consecutive wk. Saturation binding assays using [ 3 H] heparin were performed to quantitate the binding characteristics of each sample. Scatchard plot analyses indicated a significant difference in the binding affinity for [ 3 H] heparin between bulls of high and low fertility. Dissociation constants were 69.0 and 119.3 pmol for bulls of high and low fertility, respectively. In contrast, the number of binding sites for [ 3 H] heparin did not differ significantly among bulls. Differences in binding affinity of [ 3 H] heparin to bull sperm might be used to predict relative fertility of dairy bulls

Removal of glycosaminoglycans from bovine granulosa cells contributes to increased binding of hydrogen-3 heparin

Energy Technology Data Exchange (ETDEWEB)

Ax, R.L.; Stodd, C.M.; Boehm, S.K.; Bellin, M.E.

1986-02-01

Granulosa cells from small or large bovine follicles were pretreated with enzymes that hydrolyze various glycosaminoglycans, and binding of (/sup 3/H)-heparin to the granulosa was measured. Binding of (/sup 3/H) heparin increased significantly after enzymatic pretreatments with chondroitinase ABC and fungal hyaluronidase, and similar results were obtained with granulosa from small and large follicles. No changes in binding of (/sup 3/H) heparin were detected after hydrolyses with chondroitinase AC and heparinase in either follicle size. Heparitinase, which hydrolyzes heparan sulfate, led to a significant 50% increase in binding of (/sup 3/H) heparin to granulosa from large follicles but was without effect in small follicles. These results suggest that the lower binding of (/sup 3/H) heparin, which has been reported with follicular enlargement, may be due to heparan sulfate occupying or obstructing binding sites for heparin on granulosa from large follicles.

Tissue factor-dependent activation of tritium-labeled factor IX and factor X in human plasma

International Nuclear Information System (INIS)

Morrison, S.A.; Jesty, J.

1984-01-01

A comparism was made of the tissue factor-dependent activation of tritium-labeled factor IX and factor X in a human plasma system and a study was made of the role of proteases known to stimulate factor VII activity. Plasma was defibrinated by heating and depleted of its factors IX and X by passing it through antibody columns. Addition of human brain thromboplastin, Ca2+, and purified 3H-labeled factor X to the plasma resulted, after a short lag, in burst-like activation of the factor X, measured as the release of radiolabeled activation peptide. The progress of activation was slowed by both heparin and a specific inhibitor of factor Xa but factor X activation could not be completely abolished by such inhibitors. In the case of 3H-factor IX activation, the rate also increased for approximately 3 min after addition of thromboplastin, but was not subsequently curtailed. A survey of proteases implicated as activators of factor VII in other settings showed that both factor Xa and factor IXa could accelerate the activation of factor IX. However, factor Xa was unique in obliterating activation when present at concentrations greater than approximately 1 nM. Heparin inhibited the tissue factor-dependent activation of factor IX almost completely, apparently through the effect of antithrombin on the feedback reactions of factors Xa and IXa on factor VII. These results suggest that a very tight, biphasic control of factor VII activity exists in human plasma, which is modulated mainly by factor Xa. At saturation of factor VIIa/tissue factor, factor IX activation was significantly more rapid than was previously found in bovine plasma under similar conditions. The activation of factor X at saturation was slightly more rapid than in bovine plasma, despite the presence of heparin

A sensitive competitive binding assay for exogenous and endogenous heparins

International Nuclear Information System (INIS)

Dawes, J.; Pepper, D.S.

1982-01-01

A new type of assay for heparins has been devised, in which the test material competes with 125 I-labelled heparin for binding to protamine-Sepharose. The assay is very sensitive and will measure heparin concentrations down to 10 ng ml-1. It responds to both the degree of sulphation and the molecular weight of acidic polysaccharides, but is independent of their biological activities. It can be used to quantitate heparins in biological fluids after pretreatment of the samples with protease. In this way endogenous heparins were measured in normal human serum, plasma and urine. The assay is extremely versatile and has great potential for the investigation of endogenous and exogenous heparins

Characterization of a human coagulation factor Xa-binding site on Viperidae snake venom phospholipases A2 by affinity binding studies and molecular bioinformatics

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Gowda Veerabasappa T

2007-12-01

Full Text Available Abstract Background The snake venom group IIA secreted phospholipases A2 (SVPLA2, present in the Viperidae snake family exhibit a wide range of toxic and pharmacological effects. They exert their different functions by catalyzing the hydrolysis of phospholipids (PL at the membrane/water interface and by highly specific direct binding to: (i presynaptic membrane-bound or intracellular receptors; (ii natural PLA2-inhibitors from snake serum; and (iii coagulation factors present in human blood. Results Using surface plasmon resonance (SPR protein-protein interaction measurements and an in vitro biological test of inhibition of prothrombinase activity, we identify a number of Viperidae venom SVPLA2s that inhibit blood coagulation through direct binding to human blood coagulation factor Xa (FXa via a non-catalytic, PL-independent mechanism. We classify the SVPLA2s in four groups, depending on the strength of their binding. Molecular electrostatic potentials calculated at the surface of 3D homology-modeling models show a correlation with inhibition of prothrombinase activity. In addition, molecular docking simulations between SVPLA2 and FXa guided by the experimental data identify the potential FXa binding site on the SVPLA2s. This site is composed of the following regions: helices A and B, the Ca2+ loop, the helix C-β-wing loop, and the C-terminal fragment. Some of the SVPLA2 binding site residues belong also to the interfacial binding site (IBS. The interface in FXa involves both, the light and heavy chains. Conclusion We have experimentally identified several strong FXa-binding SVPLA2s that disrupt the function of the coagulation cascade by interacting with FXa by the non-catalytic PL-independent mechanism. By theoretical methods we mapped the interaction sites on both, the SVPLA2s and FXa. Our findings may lead to the design of novel, non-competitive FXa inhibitors.

Factor Xa stimulates fibroblast procollagen production, proliferation, and calcium signaling via PAR1 activation

International Nuclear Information System (INIS)

Blanc-Brude, Olivier P.; Archer, Fabienne; Leoni, Patricia; Derian, Claudia; Bolsover, Steven; Laurent, Geoffrey J.; Chambers, Rachel C.

2005-01-01

Fibroblast proliferation and procollagen production are central features of tissue repair and fibrosis. In addition to its role in blood clotting, the coagulation cascade proteinase thrombin can contribute to tissue repair by stimulating fibroblasts via proteolytic activation of proteinase-activated receptor-1 (PAR 1 ). During hemostasis, the coagulation cascade proteinase factor X is converted into factor Xa. We have previously shown that factor Xa upregulates fibroblast proliferation via production of autocrine PDGF. In this study, we further examined the effects of factor Xa on fibroblast function and aimed to identify its signaling receptor. We showed that factor Xa stimulates procollagen promoter activity and protein production by human and mouse fibroblasts. This effect was independent of PDGF and thrombin production, but dependent on factor Xa proteolytic activity. We also showed that PAR 1 -deficient mouse fibroblasts did not upregulate procollagen production, mobilize cytosolic calcium, or proliferate in response to factor Xa. Desensitization techniques and PAR 1 -specific agonists and inhibitors were used to demonstrate that PAR 1 mediates factor Xa signaling in human fibroblasts. This is the first report that factor Xa stimulates extracellular matrix production. In contrast with endothelial cells and vascular smooth muscle cells, fibroblasts appear to be the only cell type in which the effects of factor Xa are mediated mainly via PAR 1 and not PAR 2 . These findings are critical for our understanding of tissue repair and fibrotic mechanisms, and for the design of novel approaches to inhibit the profibrotic effects of the coagulation cascade without compromising blood hemostasis

Histones Differentially Modulate the Anticoagulant and Profibrinolytic Activities of Heparin, Heparin Derivatives, and Dabigatran.

Science.gov (United States)

Ammollo, Concetta Tiziana; Semeraro, Nicola; Carratù, Maria Rosaria; Colucci, Mario; Semeraro, Fabrizio

2016-02-01

The antithrombin activity of unfractionated heparin (UFH) is offset by extracellular histones, which, along with DNA, represent a novel mediator of thrombosis and a structural component of thrombi. Here, we systematically evaluated the effect of histones, DNA, and histone-DNA complexes on the anticoagulant and profibrinolytic activities of UFH, its derivatives enoxaparin and fondaparinux, and the direct thrombin inhibitor dabigatran. Thrombin generation was assessed by calibrated automated thrombinography, inhibition of factor Xa and thrombin by synthetic substrates, tissue plasminogen activator-mediated clot lysis by turbidimetry, and thrombin-activatable fibrinolysis inhibitor (TAFI) activation by a functional assay. Histones alone delayed coagulation and slightly stimulated fibrinolysis. The anticoagulant activity of UFH and enoxaparin was markedly inhibited by histones, whereas that of fondaparinux was enhanced. Histones neutralized both the anti-Xa and anti-IIa activities of UFH and preferentially blocked the anti-IIa activity of enoxaparin. The anti-Xa activity of fondaparinux was not influenced by histones when analyzed by chromogenic substrates, but was potentiated in a plasma prothrombinase assay. Histones inhibited the profibrinolytic activity of UFH and enoxaparin and enhanced that of fondaparinux by acting on the modulation of TAFI activation by anticoagulants. Histone H1 was mainly responsible for these effects. Histone-DNA complexes, as well as intact neutrophil extracellular traps, impaired the activities of UFH, enoxaparin, and fondaparinux. Dabigatran was not noticeably affected by histones and/or DNA, whatever the assay performed. In conclusion, histones and DNA present in the forming clot may variably influence the antithrombotic activities of anticoagulants, suggesting a potential therapeutic advantage of dabigatran and fondaparinux over heparins. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

Andexanet Alfa for Acute Major Bleeding Associated with Factor Xa Inhibitors

NARCIS (Netherlands)

Connolly, Stuart J.; Milling, Truman J.; Eikelboom, John W.; Gibson, C. Michael; Curnutte, John T.; Gold, Alex; Bronson, Michele D.; Lu, Genmin; Conley, Pamela B.; Verhamme, Peter; Schmidt, Jeannot; Middeldorp, Saskia; Cohen, Alexander T.; Beyer-Westendorf, Jan; Albaladejo, Pierre; Lopez-Sendon, Jose; Goodman, Shelly; Leeds, Janet; Wiens, Brian L.; Siegal, Deborah M.; Zotova, Elena; Meeks, Brandi; Nakamya, Juliet; Lim, W. Ting; Crowther, Mark; Connolly, S. C.; Crowther, M.; Eikelboom, J.; Gibson, M.; Milling, T. J.; Albaladejo, P.; Cohen, A.; Lopez-Sendon, J.; Schmidt, J.; Verhamme, P.; Beyer-Westendorf, J.; Wyse, D. G.; Garcia, D.; Prins, M. [=Martin H.; Nakamya, J.; Büller, H. R.; Mahaffey, K.; Alexander, J.; Demchuk, A.; Raskob, G.; Schulman, S.; Meeks, B.; Zotova, E.; Holadyk-Gris, I.; Coppens, M.

2016-01-01

Background Andexanet alfa (andexanet) is a recombinant modified human factor Xa decoy protein that has been shown to reverse the inhibition of factor Xa in healthy volunteers. Methods In this multicenter, prospective, open-label, single-group study, we evaluated 67 patients who had acute major

Mapping the heparin-binding site of the BMP antagonist gremlin by site-directed mutagenesis based on predictive modelling.

Science.gov (United States)

Tatsinkam, Arnold Junior; Mulloy, Barbara; Rider, Christopher C

2015-08-15

Gremlin is a member of the CAN (cerberus and DAN) family of secreted BMP (bone morphogenetic protein) antagonists and also an agonist of VEGF (vascular endothelial growth factor) receptor-2. It is critical in limb skeleton and kidney development and is re-expressed during tissue fibrosis. Gremlin binds strongly to heparin and heparan sulfate and, in the present study, we sought to investigate its heparin-binding site. In order to explore a putative non-contiguous binding site predicted by computational molecular modelling, we substituted a total of 11 key arginines and lysines located in three basic residue sequence clusters with homologous sequences from cerberus and DAN (differential screening selected gene abberative in neuroblastoma), CAN proteins which lack basic residues in these positions. A panel of six Myc-tagged gremlin mutants, MGR-1-MGR-6 (MGR, mutant gremlin), each containing different combinations of targeted substitutions, all showed markedly reduced affinity for heparin as demonstrated by their NaCl elution on heparin affinity chromatography, thus verifying our predictions. Both MGR-5 and MGR-6 retained BMP-4-binding activity comparable to that of wild-type gremlin. Low-molecular-mass heparin neither promoted nor inhibited BMP-4 binding. Finally, glutaraldehyde cross-linking demonstrated that gremlin forms non-covalent dimers, similar behaviour to that of DAN and also PRDC (protein related to cerberus and DAN), another CAN protein. The resulting dimer would possess two heparin-binding sites, each running along an exposed surface on the second β-strand finger loop of one of the monomers. © 2015 Authors; published by Portland Press Limited.

Heparin binding domain of antithrombin III: Characterization using a synthetic peptide directed polyclonal antibody

International Nuclear Information System (INIS)

Smith, J.W.; Dey, B.; Knauer, D.J.

1990-01-01

Antithrombin III (ATIII) is a plasma-borne serine protease inhibitor that apparently forms covalent complexes with thrombin. The interaction between ATIII and thrombin is enhanced several thousandfold by the glycosaminoglycan, heparin. The authors have previously proposed that the heparin binding site of ATIII residues within a region extending from amino acid residues 114-156. Computer-assisted analysis of this region revealed the presence of a 22 amino acid domain (residues 124-145), part of which shows a strong potential for the formation of an amphipathic helix: hydrophobic on one face and highly positively charged on the other. In the presence studies, polyclonal antisera were generated against a synthetic peptide corresponding to residues 124-145 in native human ATIII. Affinity-purified IgG from these antisera, as well as monovalent Fab's derived from them, specifically blocked the binding of heparin to ATIII. Additionally, occupancy of the heparin binding site by these same monovalent and bivalent IgG's at least partially substituted for heparin, accelerating linkage formation between ATIII and thrombin. These results provide the first immunological evidence that region 124-145 is directly involved in the binding of heparin to ATIII and that an antibody-induced conformational change within this region can mediate ATIII activation

Layer-by-Layer Heparinization of the Cell Surface by Using Heparin-Binding Peptide Functionalized Human Serum Albumin.

Science.gov (United States)

Song, Guowei; Hu, Yaning; Liu, Yusheng; Jiang, Rui

2018-05-20

Layer-by-layer heparinization of therapeutic cells prior to transplantation is an effective way to inhibit the instant blood-mediated inflammatory reactions (IBMIRs), which are the major cause of early cell graft loss during post-transplantation. Here, a conjugate of heparin-binding peptide (HBP) and human serum albumin (HSA), HBP-HSA, was synthesized by using heterobifunctional crosslinker. After the first heparin layer was coated on human umbilical vein endothelial cells (HUVECs) by means of the HBP-polyethylene glycol-phospholipid conjugate, HBP-HSA and heparin were then applied to the cell surface sequentially to form multiple layers. The immobilization and retention of heparin were analyzed by confocal microscopy and flow cytometry, respectively, and the cytotoxity of HBP-HSA was further evaluated by cell viability assay. Results indicated that heparin was successfully introduced to the cell surface in a layer-by-layer way and retained for at least 24 h, while the cytotoxity of HBP-HSA was negligible at the working concentration. Accordingly, this conjugate provides a promising method for co-immobilization of heparin and HSA to the cell surface under physiological conditions with improved biocompatibility.

Silencing of the Rice Gene LRR1 Compromises Rice Xa21 Transcript Accumulation and XA21-Mediated Immunity.

Science.gov (United States)

Caddell, Daniel F; Park, Chang-Jin; Thomas, Nicholas C; Canlas, Patrick E; Ronald, Pamela C

2017-12-01

The rice immune receptor XA21 confers resistance to Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial leaf blight. We previously demonstrated that an auxilin-like protein, XA21 BINDING PROTEIN 21 (XB21), positively regulates resistance to Xoo. To further investigate the function of XB21, we performed a yeast two-hybrid screen. We identified 22 unique XB21 interacting proteins, including LEUCINE-RICH REPEAT PROTEIN 1 (LRR1), which we selected for further analysis. Silencing of LRR1 in the XA21 genetic background (XA21-LRR1Ri) compromises resistance to Xoo compared with control XA21 plants. XA21-LRR1Ri plants have reduced Xa21 transcript levels and reduced expression of genes that serve as markers of XA21-mediated activation. Overexpression of LRR1 is insufficient to alter resistance to Xoo in rice lines lacking XA21. Taken together, our results indicate that LRR1 is required for wild-type Xa21 transcript expression and XA21-mediated immunity.

Prolonged Activated Clotting Time after Protamine Administration Does Not Indicate Residual Heparinization after Cardiopulmonary Bypass in Pediatric Open Heart Surgery.

Science.gov (United States)

Yamamoto, Tomohiro; Wolf, Hans-Gerd; Sinzobahamvya, Nicodème; Asfour, Boulos; Hraska, Victor; Schindler, Ehrenfried

2015-08-01

In open heart surgery, heparinization is commonly neutralized using an empirical heparin:protamine ratio ranging between 1:1 and 1:1.5. However, these ratios may result in protamine overdose that should be avoided for its negative side effects on the coagulation system. This study aimed to indicate the appropriate treatment for prolonged activated clotting time (ACT) after protamine administration following cardiopulmonary bypass (CPB) in pediatric open heart surgery by investigating the underlying reasons for it. Twenty-seven children (open heart surgery were included. Heparin was administered only before CPB (400 IU/kg) and in the pump priming volume for CPB (2,000 IU) and was neutralized by 1:1 protamine after CPB. The blood heparin concentration was measured using anti-Xa assay. ACT and blood concentrations of heparin, coagulation factors, thrombin-antithrombin complex, and prothrombin fragment 1 + 2 were assessed. A rotational thromboelastometry (ROTEM; Tem International GmbH, München, Bayern, Germany) was used to confirm the coagulation status and residual heparin after protamine administration. Anti-Xa assay showed that there is no residual heparin in the blood after 1:1 protamine administration. Nevertheless, ACT (128.89 ± 3.09 seconds before heparin administration) remained prolonged (177.14 ± 5.43 seconds at 10 minutes after protamine, 182.00 ± 5.90 seconds at 30 minutes after protamine). The blood concentrations of coagulation factors were significantly lower than those before heparin administration (p < 0.01). The low FIBTEM MCF of ROTEM (4.43 ± 0.32 mm) at 10 minutes after protamine indicated low fibrinogen concentration. Prolonged ACT after heparin neutralization by 1:1 protamine administration does not necessarily indicate residual heparin, but low blood concentrations of coagulation factors should be considered as a reason as well. Accordingly, supply of coagulation factors instead of additional protamine should be

Interactions between nattokinase and heparin/GAGs.

Science.gov (United States)

Zhang, Fuming; Zhang, Jianhua; Linhardt, Robert J

2015-12-01

Nattokinase (NK) is a serine protease extracted from a traditional Japanese food called natto. Due to its strong fibrinolytic and thrombolytic activity, NK is regarded as a valuable dietary supplement or nutraceutical for the oral thrombolytic therapy. In addition, NK has been investigated for some other medical applications including treatment of hypertension, Alzheimer's disease, and vitreoretinal disorders. The most widely used clinical anticoagulants are heparin and low molecular weight heparins. The interactions between heparin and proteins modulate diverse patho-physiological processes and heparin modifies the activity of serine proteases. Indeed, heparin plays important roles in almost all of NK's potential therapeutically applications. The current report relies on surface plasmon resonance spectroscopy to examine NK interacting with heparin as well as other glycosaminoglycans (GAGs). These studies showed that NK is a heparin binding protein with an affinity of ~250 nM. Examination with differently sized heparin oligosaccharides indicated that the interaction between NK and heparin is chain-length dependent and the minimum size for heparin binding is a hexasaccharide. Studies using chemically modified heparin showed the 6-O-sulfo as well as the N-sulfo groups but not the 2-O-sulfo groups within heparin, are essential for heparin's interaction with NK. Other GAGs (including HS, DS, and CSE) displayed modest binding affinity to NK. NK also interfered with other heparin-protein interactions, including heparin's interaction with antithrombin and fibroblast growth factors.

Furin proteolytically processes the heparin-binding region of extracellular superoxide dismutase

DEFF Research Database (Denmark)

Bowler, Russell P; Nicks, Mike; Olsen, Dorte Aa

2002-01-01

Extracellular superoxide dismutase (EC-SOD) is an antioxidant enzyme that attenuates brain and lung injury from oxidative stress. A polybasic region in the carboxyl terminus distinguishes EC-SOD from other superoxide dismutases and determines EC-SOD's tissue half-life and affinity for heparin....... There are two types of EC-SOD that differ based on the presence or absence of this heparin-binding region. It has recently been shown that proteolytic removal of the heparin-binding region is an intracellular event (Enghild, J. J., Thogersen, I. B., Oury, T. D., Valnickova, Z., Hojrup, P., and Crapo, J. D...... of intracellular proteases implicate furin as a processing protease. In vitro experiments using furin and purified EC-SOD suggest that furin proteolytically cleaves EC-SOD in the middle of the polybasic region and then requires an additional carboxypeptidase to remove the remaining lysines and arginines...

Aggregated forms of bull seminal plasma proteins and their heparin-binding activity

Czech Academy of Sciences Publication Activity Database

Jelínková, Petra; Ryšlavá, H.; Liberda, J.; Jonáková, Věra; Tichá, M.

2004-01-01

Roč. 69, - (2004), s. 616-630 ISSN 0010-0765 R&D Projects: GA ČR GA303/02/0433; GA ČR GP303/02/P069; GA MZd NJ7463 Institutional research plan: CEZ:AV0Z5052915; CEZ:MSM 113100001 Keywords : bull seminal plasma proteins * heparin-binding proteins * aggregated forms of proteins Subject RIV: CE - Biochemistry Impact factor: 1.062, year: 2004

Proteomic analysis of heparin-binding proteins from human seminal ...

Indian Academy of Sciences (India)

Prakash

(MALDI TOF/MS) for protein analysis of human HBPs. We resolved 70 ... Thus, the combined effects of seminal plasma components support the survival of ...... The BBXB motif of RANTES is the principal site for heparin binding and controls ...

Serum Heparin-binding Epidermal Growth Factor-like Growth Factor (HB-EGF) as a Biomarker for Primary Ovarian Cancer.

Science.gov (United States)

Miyata, Kohei; Yotsumoto, Fusanori; Fukagawa, Satoshi; Kiyoshima, Chihiro; Ouk, Nam Sung; Urushiyama, Daichi; Ito, Tomohiro; Katsuda, Takahiro; Kurakazu, Masamitsu; Araki, Ryota; Sanui, Ayako; Miyahara, Daisuke; Murata, Masaharu; Shirota, Kyoko; Yagi, Hiroshi; Takono, Tadao; Kato, Kiyoko; Yaegashi, Nobuo; Akazawa, Kohei; Kuroki, Masahide; Yasunaga, Shin'ichiro; Miyamoto, Shingo

2017-07-01

Ovarian cancer is the most lethal malignancy among gynaecological cancers. Although many anticancer agents have been developed for the treatment of ovarian cancer, it continues to have an extremely poor prognosis. Heparin-binding epidermal growth factor-like grown factor (HB-EGF) has been reported to be a rational therapeutic target for ovarian cancer. Here, we evaluated the clinical significance of serum HB-EGF by examining the association between prognosis and serum HB-EGF levels in patients with primary ovarian cancer. We found that high serum HB-EGF concentrations were significantly associated with poor prognosis in a combined cohort of patients with all stages of ovarian cancer, as well as in a subset of patients with advanced disease. In addition, serum HB-EGF levels increased as the cancer advanced. These data suggest that serum HB-EGF may be a target for the design of novel therapies for ovarian cancer. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Monitoring of unfractionated heparin with rotational thrombelastometry using the prothrombinase-induced clotting time reagent (PiCT®).

Science.gov (United States)

Schaden, E; Jilch, S; Hacker, S; Schober, A; Kozek-Langenecker, S

2012-12-24

To achieve sufficient and safe anticoagulation with unfractionated heparin (UFH) a close and reliable drug monitoring is necessary. In general, the activated partial thromboplastin time (APTT) is used for this purpose. In acute phase response, however, the APTT test procedure might be unreliable e.g. with false low results in the presence of elevated factor VIII. In this so called heparin resistance, measurement of anti-Xa activity is recommended over APTT to avoid potentially harmful dose escalation. A combination of anti-Xa measurement and global hemostatic testing with ROTEM® employing the anti-Xa sensitive PiCT® reagent showed high correlation with enoxaparin levels. This test modification could also be suitable for monitoring UFH. Aim of the study was to evaluate the correlation between PiCT®-ROTEM® and levels of UFH. In this in-vitro study blood samples from healthy volunteers were spiked with UFH and subjected to different ROTEM® tests. There was a linear correlation between UFH level and clotting time (CT) in the PiCT®-ROTEM® test with an excellent correlation coefficient of 0.92. Additional endpoints showed similar results (PiCT®-ROTEM® MaxVel r = -0.85 and PiCT®-ROTEM® t_MaxVel r = 0.88). As a point-of-care applicable tool ROTEM® is immediately at hand. If further clinical studies confirm sensitivity in heparin resistance, PiCT®-ROTEM® could permit rapid UFH dose adjustments especially required in critical illness with acute phase response. Copyright © 2012 Elsevier B.V. All rights reserved.

Oral direct thrombin inhibitors or oral factor Xa inhibitors for the treatment of deep vein thrombosis.

Science.gov (United States)

Robertson, Lindsay; Kesteven, Patrick; McCaslin, James E

2015-06-30

Deep vein thrombosis (DVT) is a condition in which a clot forms in the deep veins, most commonly of the leg. It occurs in approximately 1 in 1,000 people. If left untreated, the clot can travel up to the lungs and cause a potentially life-threatening pulmonary embolism (PE). Previously, a DVT was treated with the anticoagulants heparin and vitamin K antagonists. However, two forms of novel oral anticoagulants (NOACs) have been developed: oral direct thrombin inhibitors (DTI) and oral factor Xa inhibitors. The new drugs have characteristics that may be favourable over conventional treatment, including oral administration, a predictable effect, lack of frequent monitoring or re-dosing and few known drug interactions. To date, no Cochrane review has measured the effectiveness and safety of these drugs in the treatment of DVT. To assess the effectiveness of oral DTIs and oral factor Xa inhibitors for the treatment of DVT. The Cochrane Peripheral Vascular Diseases Group Trials Search Co-ordinator searched the Specialised Register (last searched January 2015) and the Cochrane Register of Studies (last searched January 2015). We searched clinical trials databases for details of ongoing or unpublished studies and the reference lists of relevant articles retrieved by electronic searches for additional citations. We included randomised controlled trials in which people with a DVT confirmed by standard imaging techniques, were allocated to receive an oral DTI or an oral factor Xa inhibitor for the treatment of DVT. Two review authors (LR, JM) independently extracted the data and assessed the risk of bias in the trials. Any disagreements were resolved by discussion with the third review author (PK). We performed meta-analyses when we considered heterogeneity low. The two primary outcomes were recurrent VTE and PE. Other outcomes included all-cause mortality and major bleeding. We calculated all outcomes using an odds ratio (OR) with a 95% confidence interval (CI). We included

Managing cancer-related venous thromboembolic disease: low-molecular-weight heparins and beyond.

Science.gov (United States)

O'Connell, Casey L; Liebman, Howard A

2008-12-01

Venous thromboembolism is a major contributor to the morbidity and mortality of patients with cancer. For patients undergoing cancer surgery, several trials support the safety and efficacy of unfractionated heparin and of low-molecular-weight heparin for the prevention of venous thromboembolism, while data regarding the efficacy and safety of these agents in the setting of medical hospitalization is less definitive and must be extracted from trials including noncancer patients with different thrombotic risk factors. Randomized clinical studies confirm that patients with cancer who develop venous thromboembolism have superior outcomes when treated with long-term low-molecular-weight heparin as compared with warfarin. Novel anticoagulants that are orally bioavailable and function by directly inhibiting factor Xa or thrombin are entering the market. To date, data regarding the efficacy and safety of these novel anticoagulants as venous thromboembolism prophylaxis and treatment in cancer patients are not available and must be extracted from larger trials with heterogeneous patient populations.

Characterization of currently marketed heparin products: key tests for quality assurance.

Science.gov (United States)

Keire, David A; Ye, Hongping; Trehy, Michael L; Ye, Wei; Kolinski, Richard E; Westenberger, Benjamin J; Buhse, Lucinda F; Nasr, Moheb; Al-Hakim, Ali

2011-01-01

During the 2007-2008 heparin crisis, it was found that the United States Pharmacopeia (USP) testing monograph for unfractionated heparin sodium (UFH) did not detect the presence of the contaminant, oversulfated chondroitin sulfate (OSCS) in heparin. In response to this concern, new tests and specifications were developed by the Food and Drug Administration (FDA) and USP and put in place to not only detect the contaminant OSCS but also to improve assurance of quality and purity of the drug product. Additional tests were also developed to monitor the heparin supply chain for other possible economically motivated additives or impurities. In 2009, a new USP monograph was put in place that includes 500 MHz (1)H NMR, SAX-HPLC, %galactosamine in total hexosamine, and anticoagulation time assays with purified factor IIa or factor Xa. These tests represent orthogonal approaches for UFH identification, measurement of bioactivity, and for detection of process impurities or contaminants in UFH. The FDA has applied these analytical approaches to the study of UFH active pharmaceutical ingredients in the marketplace. Here, we describe results from a comprehensive survey of UFH collected from seven different sources after the 2009 monograph revision and compare these data with results obtained on other heparin samples collected during the 2007-2008 crisis.

Low-molecular-weight heparins: pharmacologic profile and product differentiation.

Science.gov (United States)

Fareed, J; Jeske, W; Hoppensteadt, D; Clarizio, R; Walenga, J M

1998-09-10

The interchangeability of low-molecular-weight heparins (LMWHs) has been the subject of discussion since these products were first introduced for the prophylaxis of deep vein thrombosis. Experimental evidence now exists to show that LMWHs differ from each other in a number of characteristics. Products have been differentiated on the basis of molecular weight and biologic properties, but only limited information derived from the clinical setting is available. Potency has been described on the basis of anti-Factor Xa activity, but at equivalent anti-Xa activities, the anti-Factor IIa activity of different products shows marked variations. At the relatively small doses used for the management of postsurgical deep vein thrombosis, the effect of these interproduct differences may be relatively minor, but as LMWHs are developed for therapeutic use at much higher doses, such differences may become clinically important. Variations in safety and efficacy reported in clinical trials of LMWHs may reflect the known differences in their molecular composition and pharmacologic properties.

Daboxin P, a Major Phospholipase A2 Enzyme from the Indian Daboia russelii russelii Venom Targets Factor X and Factor Xa for Its Anticoagulant Activity.

Directory of Open Access Journals (Sweden)

Maitreyee Sharma

Full Text Available In the present study a major protein has been purified from the venom of Indian Daboia russelii russelii using gel filtration, ion exchange and Rp-HPLC techniques. The purified protein, named daboxin P accounts for ~24% of the total protein of the crude venom and has a molecular mass of 13.597 kDa. It exhibits strong anticoagulant and phospholipase A2 activity but is devoid of any cytotoxic effect on the tested normal or cancerous cell lines. Its primary structure was deduced by N-terminal sequencing and chemical cleavage using Edman degradation and tandem mass spectrometry. It is composed of 121 amino acids with 14 cysteine residues and catalytically active His48 -Asp49 pair. The secondary structure of daboxin P constitutes 42.73% of α-helix and 12.36% of β-sheet. It is found to be stable at acidic (pH 3.0 and neutral pH (pH 7.0 and has a Tm value of 71.59 ± 0.46°C. Daboxin P exhibits anticoagulant effect under in-vitro and in-vivo conditions. It does not inhibit the catalytic activity of the serine proteases but inhibits the activation of factor X to factor Xa by the tenase complexes both in the presence and absence of phospholipids. It also inhibits the tenase complexes when active site residue (His48 was alkylated suggesting its non-enzymatic mode of anticoagulant activity. Moreover, it also inhibits prothrombinase complex when pre-incubated with factor Xa prior to factor Va addition. Fluorescence emission spectroscopy and affinity chromatography suggest the probable interaction of daboxin P with factor X and factor Xa. Molecular docking analysis reveals the interaction of the Ca+2 binding loop; helix C; anticoagulant region and C-terminal region of daboxin P with the heavy chain of factor Xa. This is the first report of a phospholipase A2 enzyme from Indian viper venom which targets both factor X and factor Xa for its anticoagulant activity.

Sequence similarity between the erythrocyte binding domain of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals a functional heparin binding motif involved in binding to the Duffy antigen receptor for chemokines

Directory of Open Access Journals (Sweden)

Bolton Michael J

2011-11-01

Full Text Available Abstract Background The HIV surface glycoprotein gp120 (SU, gp120 and the Plasmodium vivax Duffy binding protein (PvDBP bind to chemokine receptors during infection and have a site of amino acid sequence similarity in their binding domains that often includes a heparin binding motif (HBM. Infection by either pathogen has been found to be inhibited by polyanions. Results Specific polyanions that inhibit HIV infection and bind to the V3 loop of X4 strains also inhibited DBP-mediated infection of erythrocytes and DBP binding to the Duffy Antigen Receptor for Chemokines (DARC. A peptide including the HBM of PvDBP had similar affinity for heparin as RANTES and V3 loop peptides, and could be specifically inhibited from heparin binding by the same polyanions that inhibit DBP binding to DARC. However, some V3 peptides can competitively inhibit RANTES binding to heparin, but not the PvDBP HBM peptide. Three other members of the DBP family have an HBM sequence that is necessary for erythrocyte binding, however only the protein which binds to DARC, the P. knowlesi alpha protein, is inhibited by heparin from binding to erythrocytes. Heparitinase digestion does not affect the binding of DBP to erythrocytes. Conclusion The HBMs of DBPs that bind to DARC have similar heparin binding affinities as some V3 loop peptides and chemokines, are responsible for specific sulfated polysaccharide inhibition of parasite binding and invasion of red blood cells, and are more likely to bind to negative charges on the receptor than cell surface glycosaminoglycans.

The heparin-binding site in tetranectin is located in the N-terminal region and binding does not involve the carbohydrate recognition domain.

Science.gov (United States)

Lorentsen, R H; Graversen, J H; Caterer, N R; Thogersen, H C; Etzerodt, M

2000-04-01

Tetranectin is a homotrimeric plasma and extracellular-matrix protein that binds plasminogen and complex sulphated polysaccharides including heparin. In terms of primary and tertiary structure, tetranectin is related to the collectin family of Ca(2+)-binding C-type lectins. Tetranectin is encoded in three exons. Exon 3 encodes the carbohydrate recognition domain, which binds to kringle 4 in plasminogen at low levels of Ca(2+). Exon 2 encodes an alpha-helix, which is necessary and sufficient to govern the trimerization of tetranectin by assembling into a triple-helical coiled-coil structural element. Here we show that the heparin-binding site in tetranectin resides not in the carbohydrate recognition domain but within the N-terminal region, comprising the 16 amino acid residues encoded by exon 1. In particular, the lysine residues in the decapeptide segment KPKKIVNAKK (tetranectin residues 6-15) are shown to be of primary importance in heparin binding.

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.

Oral direct thrombin inhibitors or oral factor Xa inhibitors for the treatment of pulmonary embolism.

Science.gov (United States)

Robertson, Lindsay; Kesteven, Patrick; McCaslin, James E

2015-12-04

Pulmonary embolism is a potentially life-threatening condition in which a clot can travel from the deep veins, most commonly in the leg, up to the lungs. Previously, a pulmonary embolism was treated with the anticoagulants heparin and vitamin K antagonists. Recently, however, two forms of direct oral anticoagulants (DOACs) have been developed: oral direct thrombin inhibitors (DTI) and oral factor Xa inhibitors. The new drugs have characteristics that may be favourable over conventional treatment, including oral administration, a predictable effect, lack of frequent monitoring or re-dosing and few known drug interactions. To date, no Cochrane review has measured the effectiveness and safety of these drugs in the long-term treatment (minimum duration of three months) of pulmonary embolism. To assess the effectiveness of oral DTIs and oral factor Xa inhibitors for the long-term treatment of pulmonary embolism. The Cochrane Vascular Trials Search Co-ordinator searched the Specialised Register (last searched January 2015) and the Cochrane Register of Studies (last searched January 2015). Clinical trials databases were also searched for details of ongoing or unpublished studies. We searched the reference lists of relevant articles retrieved by electronic searches for additional citations. We included randomised controlled trials in which patients with a pulmonary embolism confirmed by standard imaging techniques were allocated to receive an oral DTI or an oral factor Xa inhibitor for the long-term (minimum duration three months) treatment of pulmonary embolism. Two review authors (LR, JM) independently extracted the data and assessed the risk of bias in the trials. Any disagreements were resolved by discussion with the third author (PK). We used meta-analyses when we considered heterogeneity low. The two primary outcomes were recurrent venous thromboembolism and pulmonary embolism. Other outcomes included all-cause mortality and major bleeding. We calculated all outcomes

The cell-penetrating peptide domain from human heparin-binding epidermal growth factor-like growth factor (HB-EGF) has anti-inflammatory activity in vitro and in vivo

International Nuclear Information System (INIS)

Lee, Jue-Yeon; Seo, Yoo-Na; Park, Hyun-Jung; Park, Yoon-Jeong; Chung, Chong-Pyoung

2012-01-01

Highlights: ► HBP sequence identified from HB-EGF has cell penetration activity. ► HBP inhibits the NF-κB dependent inflammatory responses. ► HBP directly blocks phosphorylation and degradation of IκBα. ► HBP inhibits nuclear translocation of NF-κB p65 subunit. -- Abstract: A heparin-binding peptide (HBP) sequence from human heparin-binding epidermal growth factor-like growth factor (HB-EGF) was identified and was shown to exhibit cell penetration activity. This cell penetration induced an anti-inflammatory reaction in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages. HBP penetrated the cell membrane during the 10 min treatment and reduced the LPS-induced production of nitric oxide (NO), inducible nitric oxide synthase (iNOS), and cytokines (TNF-α and IL-6) in a concentration-dependent manner. Additionally, HBP inhibited the LPS-induced upregulation of cytokines, including TNF-α and IL-6, and decreased the interstitial infiltration of polymorphonuclear leukocytes in a lung inflammation model. HBP inhibited NF-κB-dependent inflammatory responses by directly blocking the phosphorylation and degradation of IκBα and by subsequently inhibiting the nuclear translocation of the p65 subunit of NF-κB. Taken together, this novel HBP may be potentially useful candidate for anti-inflammatory treatments and can be combined with other drugs of interest to transport attached molecules into cells.

Characterization of currently marketed heparin products: key tests for LMWH quality assurance.

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Ye, Hongping; Toby, Timothy K; Sommers, Cynthia D; Ghasriani, Houman; Trehy, Michael L; Ye, Wei; Kolinski, Richard E; Buhse, Lucinda F; Al-Hakim, Ali; Keire, David A

2013-11-01

During the 2007-2008 heparin crisis it was found that the United States Pharmacopeia (USP) testing monograph for heparin sodium or low molecular weight heparins did not detect the presence of the contaminant, oversulfated chondroitin sulfate (OSCS). In response to this concern, new tests and specifications were developed by the Food and Drug Administration (FDA) and USP and put in place to detect not only the contaminant OSCS, but also to improve assurance of quality and purity of these drug products. The USP monographs for the low molecular weight heparins (LMWHs) approved for use in the United States (dalteparin, tinzaparin and enoxaparin) are also undergoing revision to include many of the same tests used for heparin sodium, including; one-dimensional (1D) 500 MHz (1)H NMR, SAX-HPLC, percent galactosamine in total hexosamine and anticoagulation time assays with purified Factor IIa or Factor Xa. These tests represent orthogonal approaches for heparin identification, measurement of bioactivity and for detection of process impurities or contaminants in these drug products. Here we describe results from a survey of multiple lots from three types of LMWHs in the US market which were collected after the 2009 heparin sodium monograph revision. In addition, innovator and generic versions of formulated enoxaparin products purchased in 2011 are compared using these tests and found to be highly similar within the discriminating power of the assays applied. Published by Elsevier B.V.

Heparin-Binding EGF-like Growth Factor (HB-EGF) Therapy for Intestinal Injury: Application and Future Prospects

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Yang, Jixin; Su, Yanwei; Zhou, Yu; Besner, Gail E.

2014-01-01

Throughout the past 20 years, we have been investigating the potential therapeutic roles of heparin-binding EGF-like growth factor (HB-EGF), a member of the epidermal growth factor family, in various models of intestinal injury including necrotizing enterocolitis (NEC), intestinal ischemia/reperfusion (I/R) injury, and hemorrhagic shock and resuscitation (HS/R). Our studies have demonstrated that HB-EGF acts as an effective mitogen, a restitution-inducing reagent, a cellular trophic factor, an anti-apoptotic protein and a vasodilator, via its effects on various cell types in the intestine. In the current paper, we have reviewed the application and therapeutic effects of HB-EGF in three classic animal models of intestinal injury, with particular emphasis on its protection of the intestines from NEC. Additionally, we have summarized the protective functions of HB-EGF on various target cells in the intestine. Lastly, we have provided a brief discussion focusing on the future development of HB-EGF clinical applications for the treatment of various forms of intestinal injury including NEC. PMID:24345808

Heparin binding sites on Ross River virus revealed by electron cryo-microscopy

International Nuclear Information System (INIS)

Zhang Wei; Heil, Marintha; Kuhn, Richard J.; Baker, Timothy S.

2005-01-01

Cell surface glycosaminoglycans play important roles in cell adhesion and viral entry. Laboratory strains of two alphaviruses, Sindbis and Semliki Forest virus, have been shown to utilize heparan sulfate as an attachment receptor, whereas Ross River virus (RRV) does not significantly interact with it. However, a single amino acid substitution at residue 218 in the RRV E2 glycoprotein adapts the virus to heparan sulfate binding and expands the host range of the virus into chicken embryo fibroblasts. Structures of the RRV mutant, E2 N218R, and its complex with heparin were determined through the use of electron cryo-microscopy and image reconstruction methods. Heparin was found to bind at the distal end of the RRV spikes, in a region of the E2 glycoprotein that has been previously implicated in cell-receptor recognition and antibody binding

Thermodynamic parameters associated with the binding of adrenaline and norephedrine to heparin

International Nuclear Information System (INIS)

Ali-Ali, A.K.; Buchanan, J.D.; Power, D.M.; Butler, J.

1983-01-01

Pulse radiolysis was used to determine the thermodynamic parameters (ΔG', ΔH' and ΔS') governing the binding of adrenalin and norephedrine to heparin. The complexes were completely dissociated by increasing concentrations of inorganic salts. Lower concentrations of divalent cations (e.g. Ca 2+ ) were more necessary to affect dissociation than those of monovalent cations (e.g. Na + ). For each interaction, an increase in drug binding occurred as the temperature was increased from ambient. However, a transition temperature was observed (48 degC) above which the drug was progressively released as temperature was increased. These observations probably reflect conformational changes induced in the heparin below and above its melting temperature. (author)

Contemporary developments in the discovery of selective factor Xa inhibitors: A review.

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Patel, Nirav R; Patel, Dushyant V; Murumkar, Prashant R; Yadav, Mange Ram

2016-10-04

Thrombosis is a leading cause of death in cardiovascular diseases such as myocardial infarction (MI), unstable angina and acute coronary syndrome (ACS) in the industrialized world. Venous thromboembolism is observed in about 1 million people every year in United States causing significant morbidity and mortality. Conventional antithrombotic therapy has been reported to have several disadvantages and limitations like inconvenience in oral administration, bleeding risks (heparin analogs), narrow therapeutic window and undesirable interactions with food and drugs (vitamin K antagonist-warfarin). The unmet medical demand for orally active safe anticoagulants has generated widespread interest among the medicinal chemists engaged in this field. To modulate blood coagulation, various enzymes involved in the coagulation process have received great attention as potential targets by various research groups for the development of oral anticoagulants. Among these enzymes, factor Xa (FXa) has remained the centre of attention in the last decade. Intensive research efforts have been made by various research groups for the development of small, safe and orally bioavailable FXa inhibitors. This review is an attempt to compile the research work of various researchers in the direction of development of FXa inhibitors reported since 2010 onward. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Identification of potent orally active factor Xa inhibitors based on conjugation strategy and application of predictable fragment recommender system.

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Ishihara, Tsukasa; Koga, Yuji; Iwatsuki, Yoshiyuki; Hirayama, Fukushi

2015-01-15

Anticoagulant agents have emerged as a promising class of therapeutic drugs for the treatment and prevention of arterial and venous thrombosis. We investigated a series of novel orally active factor Xa inhibitors designed using our previously reported conjugation strategy to boost oral anticoagulant effect. Structural optimization of anthranilamide derivative 3 as a lead compound with installation of phenolic hydroxyl group and extensive exploration of the P1 binding element led to the identification of 5-chloro-N-(5-chloro-2-pyridyl)-3-hydroxy-2-{[4-(4-methyl-1,4-diazepan-1-yl)benzoyl]amino}benzamide (33, AS1468240) as a potent factor Xa inhibitor with significant oral anticoagulant activity. We also reported a newly developed Free-Wilson-like fragment recommender system based on the integration of R-group decomposition with collaborative filtering for the structural optimization process. Copyright © 2014 Elsevier Ltd. All rights reserved.

Activation of M1 macrophages in sepsis-induced acute kidney injury in response to heparin-binding protein.

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Li Xing

Full Text Available In the early stage of sepsis, M1 macrophages result in the production of inflammatory mediators and AKI. Heparin-binding protein (HBP have been shown to play important roles in sepsis-induced AKI. In this study, we investigate the association of HBP with M1 macrophages in sepsis-induced AKI.Male C57BL6 mice were subjected to cecal ligation and puncture (CLP or sham surgery. Biochemical and histological renal damage was assessed. Macrophage infiltration was assessed by immunohistochemistry. RT-PCR was used to investigate the expression of heparin-binding protein (HBP, the inducible nitric oxide synthase (iNOS and arginase 1 (Arg-1 mRNAs. Western blots were performed to assay the tissue levels of HBP, tumor necrosis factor alpha (TNF-α and interleukin-6 (IL-6.High levels of HBP were obviously detected 24 h after sepsis-induced AKI. Heparin inhibited HBP expression during sepsis-induced AKI. The suppression of HBP expression by heparin injection after the establishment of sepsis-induced AKI resulted in a reduction in renal injury severity accompanied with a significant repression of M1 macrophage activation and expression of TNF-α and IL-6.HBP plays an important role in the initial inflammatory reaction associated with sepsis-induced AKI, presumably by activating M1 macrophages and suppressing TNF-α and IL-6 secretion.

Neutralisation of the anti-coagulant effects of heparin by histones in blood plasma and purified systems.

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Longstaff, Colin; Hogwood, John; Gray, Elaine; Komorowicz, Erzsebet; Varjú, Imre; Varga, Zoltán; Kolev, Krasimir

2016-03-01

Neutrophil extracellular traps (NETs) composed primarily of DNA and histones are a link between infection, inflammation and coagulation. NETs promote coagulation and approaches to destabilise NETs have been explored to reduce thrombosis and treat sepsis. Heparinoids bind histones and we report quantitative studies in plasma and purified systems to better understand physiological consequences. Unfractionated heparin (UFH) was investigated by activated partial thromboplastin time (APTT) and alongside low-molecular-weight heparins (LMWH) in purified systems with thrombin or factor Xa (FXa) and antithrombin (AT) to measure the sensitivity of UFH or LMWH to histones. A method was developed to assess the effectiveness of DNA and non-anticoagulant heparinoids as anti-histones. Histones effectively neutralised UFH, the IC50 value for neutralisation of 0.2 IU/ml UFH was 1.8 µg/ml histones in APTT and 4.6 µg/ml against 0.6 IU/ml UFH in a purified system. Histones also inhibited the activities of LMWHs with thrombin (IC50 6.1 and 11.0 µg/ml histones, for different LMWHs) or FXa (IC50 7.8 and 7.0 µg/ml histones). Direct interactions of UFH and LMWH with DNA and histones were explored by surface plasmon resonance, while rheology studies showed complex effects of histones, UFH and LMWH on clot resilience. A conclusion from these studies is that anticoagulation by UFH and LMWH will be compromised by high affinity binding to circulating histones even in the presence of DNA. A complete understanding of the effects of histones, DNA and heparins on the haemostatic system must include an appreciation of direct effects on fibrin and clot structure.

Enhancement of trophoblast differentiation and survival by low molecular weight heparin requires heparin-binding EGF-like growth factor.

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Bolnick, Alan D; Bolnick, Jay M; Kohan-Ghadr, Hamid-Reza; Kilburn, Brian A; Pasalodos, Omar J; Singhal, Pankaj K; Dai, Jing; Diamond, Michael P; Armant, D Randall; Drewlo, Sascha

2017-06-01

Does low molecular weight heparin (LMWH) require heparin-binding epidermal growth factor (EGF)-like growth factor (HBEGF) signaling to induce extravillous trophoblast differentiation and decrease apoptosis during oxidative stress? LMWH increased HBEGF expression and secretion, and HBEGF signaling was required to stimulate trophoblast extravillous differentiation, increase invasion in vitro and reduce trophoblast apoptosis during oxidative stress. Abnormal trophoblast differentiation and survival contribute to placental insufficiency syndromes, including preeclampsia and intrauterine growth restriction. Preeclampsia often manifests as a pro-thrombotic state, with unsuccessful transformation of the spiral arteries that reduces oxygen supply and can produce placental infarction. LMWH improves placental function by increasing blood flow. Recent data suggest that the actions of LMWH transcend its anti-coagulative properties, but the molecular mechanism is unknown. There is evidence that LMWH alters the expression of human HBEGF in trophoblast cells, which regulates human trophoblast pathophysiology. HBEGF, itself, is capable of increasing trophoblast survival and invasiveness. First-trimester placental explants and the HTR-8/SVneo cell line, established using extravillous trophoblast outgrowths from first-trimester villous explants, were treated in vitro with LMWH to examine the effects on HBEGF signaling and trophoblast function under normal physiological and pathological conditions. A highly specific antagonist of HBEGF and other inhibitors of HBEGF downstream signaling were used to determine the relationship between LMWH treatment and HBEGF. Placental tissues (n = 5) were obtained with IRB approval and patient consent from first-trimester terminations. Placental explants and HTR-8/SVneo cells were cultured on plastic or Matrigel™ and treated with a therapeutic dose of LMWH (Enoxaparin; 10 IU/ml), with or without CRM197, pan Erb-B2 Receptor Tyrosine Kinase (ERBB

Overexpression of Heparin-Binding Epidermal Growth Factor-Like Growth Factor Mediates Liver Fibrosis in Transgenic Mice.

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Guo, Yongze; Ding, Qian; Chen, Lei; Ji, Chenguang; Hao, Huiyao; Wang, Jia; Qi, Wei; Xie, Xiaoli; Ma, Junji; Li, Aidi; Jiang, Xiaoyu; Li, Xiaotian; Jiang, Huiqing

2017-08-01

The role of heparin-binding epidermal growth factor-like growth factor (HB-EGF) in liver fibrosis is not clear and is sometimes even contradictory. To clarify this role, a HB-EGF transgenic (Tg) mouse model was, for the first time, used to evaluate the functions of HB-EGF in liver fibrosis. For the in vivo study, carbon tetrachloride injection and bile duct ligation treatment were used to induce liver fibrosis in HB-EGF Tg mice and wild-type (WT) mice, respectively. Primary hepatic satellite cells (HSCs) were isolated from HB-EGF Tg and WT mice for the in vitro study. Compared with the WT mice, HB-EGF Tg mice were shown to develop more severe liver fibrosis when treated with carbon tetrachloride or bile duct ligation, with increased matrix metalloproteinases 13 activity and enhanced expression of fibrogenic genes including α-smooth muscle actin and collagen I. HB-EGF gene transfer led to an increase in proliferation and a decrease in apoptosis in primary HSCs. The ERK signaling pathway was more highly activated in primary HSCs from HB-EGF Tg mice than in those from WT mice. Our investigation confirmed the profibrotic effect of HB-EGF on the liver using a Tg mouse model. This result may contribute to the elucidation of HB-EGF as a therapeutic target in liver fibrosis. Copyright © 2017 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.

Simple and Efficient Purification of Recombinant Proteins Using the Heparin-Binding Affinity Tag.

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Jayanthi, Srinivas; Gundampati, Ravi Kumar; Kumar, Thallapuranam Krishnaswamy Suresh

2017-11-01

Heparin, a member of the glycosaminoglycan family, is known to interact with more than 400 different types of proteins. For the past few decades, significant progress has been made to understand the molecular details involved in heparin-protein interactions. Based on the structural knowledge available from the FGF1-heparin interaction studies, we have designed a novel heparin-binding peptide (HBP) affinity tag that can be used for the simple, efficient, and cost-effective purification of recombinant proteins of interest. HBP-tagged fusion proteins can be purified by heparin Sepharose affinity chromatography using a simple sodium chloride gradient to elute the bound fusion protein. In addition, owing to the high density of positive charges on the HBP tag, recombinant target proteins are preferably expressed in their soluble forms. The purification of HBP-fusion proteins can also be achieved in the presence of chemical denaturants, including urea. Additionally, polyclonal antibodies raised against the affinity tag can be used to detect HBP-fused target proteins with high sensitivity. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

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

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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.

The differences in heparin binding for the C-terminal basic-sequence-rich peptides of HPV-16 and HPV-18 capsid protein L1

International Nuclear Information System (INIS)

Sun Jian; Yu Jisheng; Yu Zhiwu; Zha Xiao; Wu Yuqing

2012-01-01

Graphial abstract: The differences in heparin binding for the C-terminal basic-sequence-rich peptides of HPV-16 and HPV-18 capsid protein L1. Highlights: ► Several driving forces contribute to the interaction between heparin and peptides. ► C-terminal of HPV L1 is a potential candidate for the attachment to host cells. ► The C-terminal peptides of HPV-16 and -18 L1 have different heparin-binding. ► The different heparin-binding provides an explanation for the distinct prevalences. - Abstract: The high-risk types of human papillomaviruses (HPV) HPV-16 and -18 are the predominant types associated with cervical cancer. HPV-16 and -18 account for about 50% and 20%, respectively, of cervical cancers worldwide. While the reason and molecular mechanism of the distinct prevalence and distributions between them remain poorly understood, the binding affinity of cell surface receptor with capsid proteins, especially L1, may be involved. We examined heparin binding with two synthetic peptides corresponding to the 14 amino acid C-terminal peptides of HPV-16 and -18 L1 with the goal of comparing the equivalent residues in different HPV types. Using isothermal titration calorimetry (ITC) and static right-angle light scattering (SLS), we determined the binding constant K, reaction enthalpy ΔH, and other thermodynamic parameters in the interaction. Especially, we assessed the role of specific residues in binding with heparin by comparing the NMR spectra of free and heparin-bound peptides.

Identification of anthranilamide derivatives as potential factor Xa inhibitors: drug design, synthesis and biological evaluation.

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Xing, Junhao; Yang, Lingyun; Li, Hui; Li, Qing; Zhao, Leilei; Wang, Xinning; Zhang, Yuan; Zhou, Muxing; Zhou, Jinpei; Zhang, Huibin

2015-05-05

The coagulation enzyme factor Xa (fXa) plays a crucial role in the blood coagulation cascade. In this study, three-dimensional fragment based drug design (FBDD) combined with structure-based pharmacophore (SBP) model and structural consensus docking were employed to identify novel fXa inhibitors. After a multi-stage virtual screening (VS) workflow, two hit compounds 3780 and 319 having persistent high performance were identified. Then, these two hit compounds and several analogs were synthesized and screened for in-vitro inhibition of fXa. The experimental data showed that most of the designed compounds displayed significant in vitro potency against fXa. Among them, compound 9b displayed the greatest in vitro potency against fXa with the IC50 value of 23 nM and excellent selectivity versus thrombin (IC50 = 40 μM). Moreover, the prolongation of the prothrombin time (PT) was measured for compound 9b to evaluate its in vitro anticoagulant activity. As a result, compound 9b exhibited pronounced anticoagulant activity with the 2 × PT value of 8.7 μM. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Subcutaneous Administration of Low-Molecular-Weight Heparin to Horses Inhibits Ex Vivo Equine Herpesvirus Type 1-Induced Platelet Activation

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Tracy Stokol

2018-05-01

Full Text Available Equine herpesvirus type 1 (EHV-1 is a major cause of infectious respiratory disease, abortion and neurologic disease. Thrombosis in placental and spinal vessels and subsequent ischemic injury in EHV-1-infected horses manifests clinically as abortion and myeloencephalopathy. We have previously shown that addition of heparin anticoagulants to equine platelet-rich plasma (PRP can abolish ex vivo EHV-1-induced platelet activation. The goal of this study was to test whether platelets isolated from horses treated with unfractionated heparin (UFH or low-molecular-weight heparin (LMWH were resistant to ex vivo EHV-1-induced activation. In a masked, block-randomized placebo-controlled cross-over trial, 9 healthy adult horses received 4 subcutaneous injections at q. 12 h intervals of one of the following treatments: UFH (100 U/kg loading dose, 3 maintenance doses of 80 U/kg, 2 doses of LMWH (enoxaparin 80 U/kg 24 h apart with saline at the intervening 12 h intervals, or 4 doses of saline. Blood samples were collected before treatment and after 36 h, 40 h (4 h after the last injection and 60 h (24 h after the last injection. Two strains of EHV-1, Ab4 and RacL11, were added to PRP ex vivo and platelet membrane expression of P selectin was measured as a marker of platelet activation. Drug concentrations were monitored in a Factor Xa inhibition (anti-Xa bioassay. We found that LMWH, but not UFH, inhibited platelet activation induced by low concentrations (1 × 106 plaque forming units/mL of both EHV-1 strains at 40 h. At this time point, all horses had anti-Xa activities above 0.1 U/ml (range 0.15–0.48 U/ml with LMWH, but not UFH. By 60 h, a platelet inhibitory effect was no longer detected and anti-Xa activity had decreased (range 0.03 to 0.07 U/ml in LMWH-treated horses. Neither heparin inhibited platelet activation induced by high concentrations (5 × 106 plaque forming units/mL of the RacL11 strain. We found substantial between horse

Expressional and Biochemical Characterization of Rice Disease Resistance Gene Xa3/Xa26 Family

Institute of Scientific and Technical Information of China (English)

Songjie Xu; Yinglong Cao; Xianghua Li; Shiping Wang

2007-01-01

The rice (Oryza sativa L.) Xa3/Xa26 gene, conferring race-specific resistance to bacterial blight disease and encoding a leucine-rich repeat (LRR) receptor kinase-like protein, belongs to a multigene family consisting of tandem clustered homologous genes, colocalizing with several uncharacterized genes for resistance to bacterial blight or fungal blast. To provide more information on the expressional and biochemical characteristics of the Xa3/Xa26 family, we analyzed the family members. Four Xa3/Xa26 family members in the indica rice variety Teqing, which carries a bacterial blight resistance gene with a chromosomal location tightly linked to Xa3/Xa26, and five Xa3/Xa26 family members in the japonica rice variety Nipponbare, which carries at least one uncharacterized blast resistance gene, were constitutively expressed in leaf tissue. The result suggests that some of the family members may be candidates of these uncharacterized resistance genes. At least five putative N-glycosylation sites in the LRR domain of XA3/XA26 protein are not glycosylated. The XA3/XA26 and its family members MRKa and MRKc all possess the consensus sequences of paired cysteines, which putatively function in dimerization of the receptor proteins for signal transduction, immediately before the first LRR and immediately after the last LRR. However, no homo-dimer between the XA3/XA26 molecules or hetero-dimer between XA3/XA26 and MRKa or MRKc were formed, indicating that XA3/XA26 protein might function either as a monomer or a hetero-dimer formed with other protein outside of the XA3/XA26 family. These results provide valuable information for further extensive investigation into this multiple protein family.

Preparation and in vitro evaluation of heparin-loaded polymeric nanoparticles.

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Jiao, Y Y; Ubrich, N; Marchand-Arvier, M; Vigneron, C; Hoffman, M; Maincent, P

2001-01-01

Nanoparticles of a highly soluble macromolecular drug, heparin, were formulated with two biodegradable polymers (poly-E-caprolactone [PCL] and poly (D, L-lactic-co-glycolic-acid) 50/50 [PLAGA]) and two nonbiodegradable positively charged polymers (Eudragit RS and RL) by the double emulsion and solvent evaporation method, using a high-pressure homogenization device. The encapsulation efficiency and heparin release profiles were studied as a function of the type of polymers employed (alone or in combination) and the concentration of heparin. Optimal encapsulation efficiency was observed when 5000 IU of heparin were incorporated in the first emulsion. High drug entrapment efficiency was observed in both Eudragit RS and RL nanoparticles (60% and 98%, respectively), compared with PLAGA and PCL nanoparticles (PLAGA increased the encapsulation efficiency compared with these two biodegradable polymers used alone; however, the in vitro drug release was not modified and remained low. On the other hand, the addition of esterase to the dissolution medium resulted in a significant increase in heparin release. The in vitro biological activity of released heparin, evaluated by measuring the anti-Xa activity by a colorimetric assay, was conserved after the encapsulation process.

Interactions of oversulfated chondroitin sulfate (OSCS) from different sources with unfractionated heparin.

Science.gov (United States)

Gray, Angel; Litinas, Evangelos; Jeske, Walter; Fareed, Jawed; Hoppensteadt, Debra

2012-01-01

In 2008, oversulfated chondroitin sulfate (OSCS) was identified as the main contaminant in recalled heparin. Oversulfated chondroitin sulfate can be prepared from bovine (B), porcine (P), shark (Sh), or skate (S) origin and may produce changes in the antithrombotic, bleeding, and hemodynamic profile of heparins. This study examines the interactions of various OSCSs on heparin in animal models of thrombosis and bleeding, as well as on the anticoagulant and antiprotease effects in in vitro assays. Mixtures of 70% unfractionated heparin (UFH) with 30% OSCS from different sources were tested. In the in vitro activated partial thromboplastin time (aPTT) assay, all contaminant mixtures showed a decrease in clotting times. In addition, a significant increase in bleeding time compared to the control (UFH/saline) was observed. In the thrombosis model, no significant differences were observed. The OSCSs significantly increased anti-Xa activity in ex vivo blood samples. These results indicate that various sources of OSCS affect the hemostatic properties of heparin.

Targeting Heparin to Collagen within Extracellular Matrix Significantly Reduces Thrombogenicity and Improves Endothelialization of Decellularized Tissues.

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Jiang, Bin; Suen, Rachel; Wertheim, Jason A; Ameer, Guillermo A

2016-12-12

Thrombosis within small-diameter vascular grafts limits the development of bioartificial, engineered vascular conduits, especially those derived from extracellular matrix (ECM). Here we describe an easy-to-implement strategy to chemically modify vascular ECM by covalently linking a collagen binding peptide (CBP) to heparin to form a heparin derivative (CBP-heparin) that selectively binds a subset of collagens. Modification of ECM with CBP-heparin leads to increased deposition of functional heparin (by ∼7.2-fold measured by glycosaminoglycan composition) and a corresponding reduction in platelet binding (>70%) and whole blood clotting (>80%) onto the ECM. Furthermore, addition of CBP-heparin to the ECM stabilizes long-term endothelial cell attachment to the lumen of ECM-derived vascular conduits, potentially through recruitment of heparin-binding growth factors that ultimately improve the durability of endothelialization in vitro. Overall, our findings provide a simple yet effective method to increase deposition of functional heparin on the surface of ECM-based vascular grafts and thereby minimize thrombogenicity of decellularized tissue, overcoming a significant challenge in tissue engineering of bioartificial vessels and vascularized organs.

A triad of lys12, lys41, arg78 spatial domain, a novel identified heparin binding site on tat protein, facilitates tat-driven cell adhesion.

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Jing Ai

Full Text Available Tat protein, released by HIV-infected cells, has a battery of important biological effects leading to distinct AIDS-associated pathologies. Cell surface heparan sulfate protoglycans (HSPGs have been accepted as endogenous Tat receptors, and the Tat basic domain has been identified as the heparin binding site. However, findings that deletion or substitution of the basic domain inhibits but does not completely eliminate Tat-heparin interactions suggest that the basic domain is not the sole Tat heparin binding site. In the current study, an approach integrating computational modeling, mutagenesis, biophysical and cell-based assays was used to elucidate a novel, high affinity heparin-binding site: a Lys12, Lys41, Arg78 (KKR spatial domain. This domain was also found to facilitate Tat-driven β1 integrin activation, producing subsequent SLK cell adhesion in an HSPG-dependent manner, but was not involved in Tat internalization. The identification of this new heparin binding site may foster further insight into the nature of Tat-heparin interactions and subsequent biological functions, facilitating the rational design of new therapeutics against Tat-mediated pathological events.

A ΩXaV motif in the Rift Valley fever virus NSs protein is essential for degrading p62, forming nuclear filaments and virulence.

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Cyr, Normand; de la Fuente, Cynthia; Lecoq, Lauriane; Guendel, Irene; Chabot, Philippe R; Kehn-Hall, Kylene; Omichinski, James G

2015-05-12

Rift Valley fever virus (RVFV) is a single-stranded RNA virus capable of inducing fatal hemorrhagic fever in humans. A key component of RVFV virulence is its ability to form nuclear filaments through interactions between the viral nonstructural protein NSs and the host general transcription factor TFIIH. Here, we identify an interaction between a ΩXaV motif in NSs and the p62 subunit of TFIIH. This motif in NSs is similar to ΩXaV motifs found in nucleotide excision repair (NER) factors and transcription factors known to interact with p62. Structural and biophysical studies demonstrate that NSs binds to p62 in a similar manner as these other factors. Functional studies in RVFV-infected cells show that the ΩXaV motif is required for both nuclear filament formation and degradation of p62. Consistent with the fact that the RVFV can be distinguished from other Bunyaviridae-family viruses due to its ability to form nuclear filaments in infected cells, the motif is absent in the NSs proteins of other Bunyaviridae-family viruses. Taken together, our studies demonstrate that p62 binding to NSs through the ΩXaV motif is essential for degrading p62, forming nuclear filaments and enhancing RVFV virulence. In addition, these results show how the RVFV incorporates a simple motif into the NSs protein that enables it to functionally mimic host cell proteins that bind the p62 subunit of TFIIH.

Ortholog Alleles at Xa3/Xa26 Locus Confer Conserved Race-Specific Resistance against Xanthomonas oryzae in Rice

Institute of Scientific and Technical Information of China (English)

Hong-Jing Li; Xiang-Hua Li; Jing-Hua Xiao; Rod A. Wing; Shi-Ping Wang

2012-01-01

The rice disease resistance (R) gene Xa3/Xa26 (having also been named Xa3 and Xa26) against Xanthomonas oryzae pv.oryzae (Xoo),which causes bacterial blight disease,belongs to a multiple gene family clustered in chromosome 11 and is from an AA genome rice cultivar (Oryza sativa L.).This family encodes leucine-rich repeat (LRR) receptor kinasetype proteins.Here,we show that the orthologs (alleles) of Xa3/Xa26,Xa3/Xa26-2,and Xa3/Xa26-3,from wild Oryza species O.officinalis (CC genome) and O.minuta (BBCC genome),respectively,were also R genes against Xoo.Xa3/Xa26-2 and Xa3/Xa26-3 conferred resistance to 16 of the 18 Xoo strains examined.Comparative sequence analysis of the Xa3/Xa26 families in the two wild Oryza species showed that Xa3/Xa26-3 appeared to have originated from the CC genome of O.minuta.The predicted proteins encoded by Xa3/Xa26,Xa3/Xa26-2,and Xa3/Xa26-3 share 91-99％ sequence identity and 94-99％ sequence similarity.Transgenic plants carrying a single copy of Xa3/Xa26,Xa3/Xa26-2,or Xa3/Xa26-3,in the same genetic background,showed a similar resistance spectrum to a set of Xoo strains,although plants carrying Xa3/Xa26-2 or Xa3/Xa26-3 showed lower resistance levels than the plants carrying Xa3/Xa26.These results suggest that the Xa3/Xa26 locus predates the speciation of A and C genome,which is approximately 7.5 million years ago.Thus,the resistance specificity of this locus has been conserved for a long time.

Induction of Xa10-like Genes in Rice Cultivar Nipponbare Confers Disease Resistance to Rice Bacterial Blight.

Science.gov (United States)

Wang, Jun; Tian, Dongsheng; Gu, Keyu; Yang, Xiaobei; Wang, Lanlan; Zeng, Xuan; Yin, Zhongchao

2017-06-01

Bacterial blight of rice, caused by Xanthomonas oryzae pv. oryzae, is one of the most destructive bacterial diseases throughout the major rice-growing regions in the world. The rice disease resistance (R) gene Xa10 confers race-specific disease resistance to X. oryzae pv. oryzae strains that deliver the corresponding transcription activator-like (TAL) effector AvrXa10. Upon bacterial infection, AvrXa10 binds specifically to the effector binding element in the promoter of the R gene and activates its expression. Xa10 encodes an executor R protein that triggers hypersensitive response and activates disease resistance. 'Nipponbare' rice carries two Xa10-like genes in its genome, of which one is the susceptible allele of the Xa23 gene, a Xa10-like TAL effector-dependent executor R gene isolated recently from 'CBB23' rice. However, the function of the two Xa10-like genes in disease resistance to X. oryzae pv. oryzae strains has not been investigated. Here, we designated the two Xa10-like genes as Xa10-Ni and Xa23-Ni and characterized their function for disease resistance to rice bacterial blight. Both Xa10-Ni and Xa23-Ni provided disease resistance to X. oryzae pv. oryzae strains that deliver the matching artificially designed TAL effectors (dTALE). Transgenic rice plants containing Xa10-Ni and Xa23-Ni under the Xa10 promoter provided specific disease resistance to X. oryzae pv. oryzae strains that deliver AvrXa10. Xa10-Ni and Xa23-Ni knock-out mutants abolished dTALE-dependent disease resistance to X. oryzae pv. oryzae. Heterologous expression of Xa10-Ni and Xa23-Ni in Nicotiana benthamiana triggered cell death. The 19-amino-acid residues at the N-terminal regions of XA10 or XA10-Ni are dispensable for their function in inducing cell death in N. benthamiana and the C-terminal regions of XA10, XA10-Ni, and XA23-Ni are interchangeable among each other without affecting their function. Like XA10, both XA10-Ni and XA23-Ni locate to the endoplasmic reticulum (ER) membrane

Profiling Heparin-Chemokine Interactions Using Synthetic Tools

Science.gov (United States)

de Paz, Jose L.; Moseman, E. Ashley; Noti, Christian; Polito, Laura; von Andrian, Ulrich H.; Seeberger, Peter H.

2009-01-01

Glycosaminoglycans (GAGs), such as heparin or heparan sulfate, are required for the in vivo function of chemokines. Chemokines play a crucial role in the recruitment of leukocyte subsets to sites of inflammation and lymphocytes trafficking. GAG-chemokine interactions mediate cell migration and determine which leukocyte subsets enter tissues. Identifying the exact GAC sequences that bind to particular chemokines is key to understand chemokine function at the molecular level and develop strategies to interfere with chemokine-mediated processes. Here, we characterize the heparin binding profiles of eight chemokines (CCL21, IL-8, CXCL12, CXCL13, CCL19, CCL25, CCL28, and CXCL16) by employing heparin microarrays containing a small library of synthetic heparin oligosaccharides. The chemokines differ significantly in their interactions with heparin oligosaccharides: While some chemokines, (e.g., CCL21) strongly bind to a hexasaccharide containing the GlcNSO3(6-OSO3)-IdoA(2-OSO3) repeating unit, CCL19 does not bind and CXCL12 binds only weakly. The carbohydrate microarray binding results were validated by surface plasmon resonance experiments. In vitro chemotaxis assays revealed that dendrimers coated with the fully sulfated heparin hexasaccharide inhibit lymphocyte migration toward CCL21. Migration toward CXCL12 or CCL19 was not affected. These in vitro homing assays indicate that multivalent synthetic heparin dendrimers inhibit the migration of lymphocytes toward certain chemokine gradients by blocking the formation of a chemokine concentration gradient on GAG endothelial chains. These findings are in agreement with preliminary in vivo measurements of circulating lymphocytes. The results presented here contribute to the understanding of GAG-chemokine interactions, a first step toward the design of novel drugs that modulate chemokine activity. PMID:18030990

Heparin-Functionalized chitosan/ κ-carrageenan complexes as potential scaffolds for tissue engineering

International Nuclear Information System (INIS)

Dofeliz, Joni L.; Rojas, Nina Rosario L.

2015-01-01

Cell-based approaches to tissue regeneration are playing an increasingly important role in bone and cartilage repair. This is made possible through the use of growth factors, which are signaling molecules that induce a number of effects such as cell proliferation, migration and differentiation. But problems arise as these growth factors tend to be expensive, short-lived and slow moving through the extracellular matrix, making them very inefficient in their current form of introduction. One such growth factor is basic fibroblast growth factor (bFGF). The general objective of this study is to construct heparin-functionalized chitosan/κ-carrageenan scaffolds, which when bound to basic fibroblast growth factor (bFGF), may be used in the culture of mesenchymal stem cells for differentiation into cartilage. In this study, gels of semi-interpenetrating networks (semi-IPN) of chitosan and κ-carrageenan (2:4:1 blend ration) were prepared using calcium chloride as a cross-linker. The gels were then functionalized with heparin, which is known for its growth factor binding ability, using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) as cross-linker. The functionalized gels were then reshaped into scaffold on the wells of 48-well plates through freeze-dry method. The scaffolds were found to be realatively porous with pore sizes larger than 100 μm which satisfies the requirements for cell culture. Subsequent thermogravimetric analysis shows that the scaffolds were relatively stable with a degradation temperature of 277°C. Additionally, there was no observable separation of the individual degradation temperatures of chitosan and κ-carrageenan, confirming that a single miscible phase in the form of a semi-IPN structure was created. Results of scanning electron microscopy also showed that the scaffolds were stable under 2 hours of UV sterilization. The FTIR spectra of the heparinized PEC showed characteristic absorption bands (S=O asymetric stretch) in the area of 1160

Analyses of Interactions Between Heparin and the Apical Surface Proteins of Plasmodium falciparum

Science.gov (United States)

Kobayashi, Kyousuke; Takano, Ryo; Takemae, Hitoshi; Sugi, Tatsuki; Ishiwa, Akiko; Gong, Haiyan; Recuenco, Frances C.; Iwanaga, Tatsuya; Horimoto, Taisuke; Akashi, Hiroomi; Kato, Kentaro

2013-11-01

Heparin, a sulfated glycoconjugate, reportedly inhibits the blood-stage growth of the malaria parasite Plasmodium falciparum. Elucidation of the inhibitory mechanism is valuable for developing novel invasion-blocking treatments based on heparin. Merozoite surface protein 1 has been reported as a candidate target of heparin; however, to better understand the molecular mechanisms involved, we characterized the molecules that bind to heparin during merozoite invasion. Here, we show that heparin binds only at the apical tip of the merozoite surface and that multiple heparin-binding proteins localize preferentially in the apical organelles. To identify heparin-binding proteins, parasite proteins were fractionated by means of heparin affinity chromatography and subjected to immunoblot analysis with ligand-specific antibodies. All tested members of the Duffy and reticulocyte binding-like families bound to heparin with diverse affinities. These findings suggest that heparin masks the apical surface of merozoites and blocks interaction with the erythrocyte membrane after initial attachment.

The N-terminal of a heparin-binding sperm membrane mitogen possess lectin-like sequence

International Nuclear Information System (INIS)

Mor, Visesato; Chatterjee, Tapati

2007-01-01

Glycosaminoglycans like heparin and heparin sulfate in follicular fluid induce changes in the intracellular environment during the spermatozoal functional maturation. We previously reported the isolation, purification and partial characterization of a heparin binding sperm membrane protein (HBSM). In the present study, the amino acids analysis provided evidence of a single sequence, which suggest the homogeneity of the purified HBSM. Fourteen amino acids- 1 A D T I V A V E L D T Y P N 14 -correspond to the amino terminal sequence of Concanavalin A (Con A) and contain 45.2% carbohydrate by weight. HBSM possess mitogenic property on lymphocytes with comparable magnitude to the well-known mitogen; Con A, inducing 83% radiolabel thymidine incorporation in growing lymphocytes. Unlike Con A, there was no agglutination of cell by HBSM upto 5 ng/ml concentration. Interestingly, we found that heparin and chondroitin sulfate-conjugated HBSM inhibit the proliferative activity. Similar effect was also found with an in-house isolate sulfated glycans; G-I (28% sulfate). In contrast, there was no inhibition by the desulfated form; G-ID. Altogether, our data suggest that the mechanism of cell proliferative pathway may be different for HBSM and Con A

Octasaccharide is the minimal length unit required for efficient binding of cyclophilin B to heparin and cell surface heparan sulphate.

Science.gov (United States)

Vanpouille, Christophe; Denys, Agnès; Carpentier, Mathieu; Pakula, Rachel; Mazurier, Joël; Allain, Fabrice

2004-09-01

Cyclophilin B (CyPB) is a heparin-binding protein first identified as a receptor for cyclosporin A. In previous studies, we reported that CyPB triggers chemotaxis and integrin-mediated adhesion of T-lymphocytes by way of interaction with two types of binding sites. The first site corresponds to a signalling receptor; the second site has been identified as heparan sulphate (HS) and appears crucial to induce cell adhesion. Characterization of the HS-binding unit is critical to understand the requirement of HS in pro-adhesive activity of CyPB. By using a strategy based on gel mobility shift assays with fluorophore-labelled oligosaccharides, we demonstrated that the minimal heparin unit required for efficient binding of CyPB is an octasaccharide. The mutants CyPB(KKK-) [where KKK- refers to the substitutions K3A(Lys3-->Ala)/K4A/K5A] and CyPB(DeltaYFD) (where Tyr14-Phe-Asp16 has been deleted) failed to interact with octasaccharides, confirming that the Y14FD16 and K3KK5 clusters are required for CyPB binding. Molecular modelling revealed that both clusters are spatially arranged so that they may act synergistically to form a binding site for the octasaccharide. We then demonstrated that heparin-derived octasaccharides and higher degree of polymerization oligosaccharides inhibited the interaction between CyPB and fluorophore-labelled HS chains purified from T-lymphocytes, and strongly reduced the HS-dependent pro-adhesive activity of CyPB. However, oligosaccharides or heparin were unable to restore adhesion of heparinase-treated T-lymphocytes, indicating that HS has to be present on the cell membrane to support the pro-adhesive activity of CyPB. Altogether, these results demonstrate that the octasaccharide is likely to be the minimal length unit required for efficient binding of CyPB to cell surface HS and consequent HS-dependent cell responses.

Hydrolysis and Sulfation Pattern Effects on Release of Bioactive Bone Morphogenetic Protein-2 from Heparin-Based Microparticles.

Science.gov (United States)

Tellier, Liane E; Miller, Tobias; McDevitt, Todd C; Temenoff, Johnna S

2015-10-28

Glycosaminoglycans (GAGs) such as heparin are promising materials for growth factor delivery due to their ability to efficiently bind positively charged growth factors including bone morphogenetic protein-2 (BMP-2) through their negatively charged sulfate groups. Therefore, the goal of this study was to examine BMP-2 release from heparin-based microparticles (MPs) after first, incorporating a hydrolytically degradable crosslinker and varying heparin content within MPs to alter MP degradation and second, altering the sulfation pattern of heparin within MPs to vary BMP-2 binding and release. Using varied MP formulations, it was found that the time course of MP degradation for 1 wt% heparin MPs was ~4 days slower than 10 wt% heparin MPs, indicating that MP degradation was dependent on heparin content. After incubating 100 ng BMP-2 with 0.1 mg MPs, most MP formulations loaded BMP-2 with ~50% efficiency and significantly more BMP-2 release (60% of loaded BMP-2) was observed from more sulfated heparin MPs (MPs with ~100% and 80% of native sulfation). Similarly, BMP-2 bioactivity in more sulfated heparin MP groups was at least four-fold higher than soluble BMP-2 and less sulfated heparin MP groups, as determined by an established C2C12 cell alkaline phosphatase (ALP) assay. Ultimately, the two most sulfated 10 wt% heparin MP formulations were able to efficiently load and release BMP-2 while enhancing BMP-2 bioactivity, making them promising candidates for future growth factor delivery applications.

Developing an Anti-Xa-Based Anticoagulation Protocol for Patients with Percutaneous Ventricular Assist Devices.

Science.gov (United States)

Sieg, Adam; Mardis, B Andrew; Mardis, Caitlin R; Huber, Michelle R; New, James P; Meadows, Holly B; Cook, Jennifer L; Toole, J Matthew; Uber, Walter E

2015-01-01

Because of the complexities associated with anticoagulation in temporary percutaneous ventricular assist device (pVAD) recipients, a lack of standardization exists in their management. This retrospective analysis evaluates current anticoagulation practices at a single center with the aim of identifying an optimal anticoagulation strategy and protocol. Patients were divided into two cohorts based on pVAD implanted (CentriMag (Thoratec; Pleasanton, CA) / TandemHeart (CardiacAssist; Pittsburgh, PA) or Impella (Abiomed, Danvers, MA)), with each group individually analyzed for bleeding and thrombotic complications. Patients in the CentriMag/TandemHeart cohort were subdivided based on the anticoagulation monitoring strategy (activated partial thromboplastin time (aPTT) or antifactor Xa unfractionated heparin (anti-Xa) values). In the CentriMag/TandemHeart cohort, there were five patients with anticoagulation titrated based on anti-Xa values; one patient developed a device thrombosis and a major bleed, whereas another patient experienced major bleeding. Eight patients received an Impella pVAD. Seven total major bleeds in three patients and no thrombotic events were detected. Based on distinct differences between the devices, anti-Xa values, and outcomes, two protocols were created to guide anticoagulation adjustments. However, anticoagulation in patients who require pVAD support is complex with constantly evolving anticoagulation goals. The ideal level of anticoagulation should be individually determined using several coagulation laboratory parameters in concert with hemodynamic changes in the patient's clinical status, the device, and the device cannulation.

Exploiting VM/XA

International Nuclear Information System (INIS)

Boeheim, C.

1990-03-01

The Stanford Linear Accelerator Center has recently completed a conversion to IBM's VM/XA SP Release 2 operating system. The primary physics application had been constrained by the previous 16 megabyte memory limit. Work is underway to enable this application to exploit the new features of VM/XA. This paper presents a brief tutorial on how to convert an application to exploit VM/XA and discusses some of the SLAC experiences in doing so. 13 figs

Isolation of non-heprin-binding and heparin-binding proteins of boar prostate

Czech Academy of Sciences Publication Activity Database

Maňásková, Pavla; Liberda, J.; Tichá, M.; Jonáková, Věra

2002-01-01

Roč. 770, - (2002), s. 137-143 ISSN 1570-0232. [International Symposium /2./ - Separation in the BioSciences. Praha, 17.09.2001-20.09.2001] R&D Projects: GA ČR GA303/99/0357; GA ČR GV524/96/K162 Institutional research plan: CEZ:AV0Z5052915 Keywords : isolation * prostatic proteins * heparin Subject RIV: CE - Biochemistry Impact factor: 1.913, year: 2002

Enantiomeric and Diastereomeric Self-Assembled Multivalent (SAMul) Nanostructures - Understanding the Effects of Chirality on Binding to Polyanionic Heparin and DNA.

Science.gov (United States)

Thornalley, Kiri; Laurini, Erik; Pricl, Sabrina; Smith, David K

2018-05-15

A family of four self-assembling lipopeptides containing Ala-Lys peptides attached to a C16 aliphatic chain was synthesised. These compounds form two enantiomeric pairs that bear a diastereomeric relationship to one another (C16-L-Ala-L-Lys/C16-D-Ala-D-Lys) and (C16-D-Ala-L-Lys/C16-L-Ala-D-Lys). These diastereomeric pairs have very different critical micelle concentrations (CMCs), with LL/DD < DL/LD suggesting more effective assembly of the former. The self-assembled multivalent (SAMul) systems bind biological polyanions as result of the cationic lysine groups on their surfaces. Polyanion binding was investigated using dye displacement assays and isothermal calorimetry (ITC). On heparin binding, there was no significant enantioselectivity, but there was a binding preference for the diastereomeric assemblies with lower CMCs. Conversely, on binding DNA, there was a significant enantioselective preference for systems displaying D-lysine ligands, with a further slight preference for attachment to L-alanine, with the CMC being irrelevant. Binding to adaptive, ill-defined heparin has a large favourable entropic term, suggesting it depends primarily on the cationic SAMul nanostructure maximising surface contact with heparin, which can adapt, displacing solvent and other ions. Conversely, binding to well-defined, shape-persistent DNA has a larger favourable enthalpic term, and combined with the enantioselectivity, this allows us to suggest that its SAMul binding is based on optimised individual electrostatic interactions at the molecular level, with a preference for binding to D-lysine. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

In vitro effects of heparin and tissue factor pathway inhibitor on factor VII assays. possible implications for measurements in vivo after heparin therapy

DEFF Research Database (Denmark)

Bladbjerg, E-M; Larsen, L F; Ostergaard, P

2000-01-01

The coagulant activity of blood coagulation factor VII (FVII:C) can be lowered by changes in lifestyle and by therapeutic intervention, e.g. heparin infusion. The question is, however, whether FVII:C determined ex vivo is a valid measure of the FVII activity in vivo. We measured plasma FVII......:C, activated FVII (FVIIa), FVII protein (FVII:Ag), tissue factor pathway inhibitor (TFPI), triglycerides, and free fatty acids (FFA) before and 15 min after infusion of a bolus of unfractionated heparin (50 IU/kg body weight) in 12 healthy subjects. Additionally, we conducted in vitro experiments...

XA23 is an executor R protein and confers broad-spectrum disease resistance in rice.

Science.gov (United States)

Wang, Chunlian; Zhang, Xiaoping; Fan, Yinglun; Gao, Ying; Zhu, Qinlong; Zheng, Chongke; Qin, Tengfei; Li, Yanqiang; Che, Jinying; Zhang, Mingwei; Yang, Bing; Liu, Yaoguang; Zhao, Kaijun

2014-11-09

The majority of plant disease resistance (R) genes encode proteins that share common structural features. However, the transcription activator-like effector (TALE) associated executor type R genes show no considerable sequence homology to any known R genes. We adopted a map-based cloning approach and TALE-based technology to isolate and characterize Xa23, a new executor R gene derived from the wild rice (Oryza rufipogon) that confers an extremely broad spectrum of resistance to bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo). Xa23 encodes a 113-amino acid protein that shares 50% identity to the known executor R protein XA10. The predicted transmembrane helices in XA23 also overlap with those of XA10. Unlike Xa10, however, Xa23 transcription is specifically activated by AvrXa23, a TALE present in all examined Xoo field isolates. Moreover, the susceptible xa23 allele has an identical open reading frame of Xa23, but differs in promoter region by lacking the TALE binding-element (EBE) for AvrXa23. XA23 can trigger strong hypersensitive response in rice, tobacco and tomato. Our results provide the first evidence that plant genomes have an executor R gene family in which members execute their function and spectrum of disease resistance by recognizing the cognate TALEs in pathogen. © The Author 2014. Published by the Molecular Plant Shanghai Editorial Office in association with Oxford University Press on behalf of CSPB and IPPE, SIBS, CAS.

Thermodynamic parameters associated with the binding of adrenalin and norephedrine to heparin

International Nuclear Information System (INIS)

Al-Ali, A.K.; Buchanan, J.D.; Power, D.M.; Butler, J.

1983-01-01

Pulse radiolysis has been used to determine the thermodynamic parameters (ΔG', ΔH' and ΔS') governing the binding of adrenalin and norephedrine to heparin. These complexes were completely dissociated by increasing concentrations of inorganic salts. Lower concentrations of divalent cations (e.g. Ca 2+ ) than of monovalent cations (e.g. Na + ) were necessary to effect dissociation of the complex. For each interaction an increase in drug binding occurred as the temperature was increased from ambient. However, a transition temperature was observed (48 0 C) above which the drug was progressively released as the temperature was increased. These observations are discussed in terms of conformational changes induced in the polymer below and above its melting temperature. (author)

Update on the clinical use of the low-molecular-weight heparin, parnaparin

Directory of Open Access Journals (Sweden)

Giuseppe Camporese

2009-09-01

Full Text Available Giuseppe Camporese1, Enrico Bernardi2, Franco Noventa31Unit of Angiology and 3Department of Clinical and Experimental Medicine, Clinical Epidemiology Group, University Hospital of Padua, Italy; 2Department of Emergency and Accident Medicine, Hospital of Conegliano Veneto, ItalyAbstract: Parnaparin is a low-molecular-weight heparin that has widely shown its efficacy and safety in prevention of venous thromboembolism, in the treatment of chronic venous disorders, and in the treatment of venous and arterial (stable and unstable angina, acute ST-segment elevation myocardial infarction thrombosis. Parnaparin at the respective dosages of 3200, 4250, 6400, or 12800 IUaXa for a period ranging from 3 to 5 days to 6 months, is usually administered subcutaneously by means of once-daily regimen and is better tolerated than unfractionated heparin at the injection site. In the variety of commercially available low-molecular-weight heparins, parnaparin represents a useful therapeutic option, even though little evidence is available comparing the superiority or the equivalent efficacy and safety of parnaparin to that of the unfractionated heparin or placebo. This review summarizes the available literature on the use of parnaparin in different settings of cardiovascular diseases, including papers published during the past year and ongoing studies.Keywords: low-molecular-weight heparin, heparin, parnaparin, acute coronary syndromes, venous thromboembolism

Human trophoblast survival at low oxygen concentrations requires metalloproteinase-mediated shedding of heparin-binding EGF-like growth factor.

Science.gov (United States)

Armant, D Randall; Kilburn, Brian A; Petkova, Anelia; Edwin, Samuel S; Duniec-Dmuchowski, Zophia M; Edwards, Holly J; Romero, Roberto; Leach, Richard E

2006-02-01

Heparin-binding EGF-like growth factor (HBEGF), which is expressed in the placenta during normal pregnancy, is down regulated in pre-eclampsia, a human pregnancy disorder associated with poor trophoblast differentiation and survival. This growth factor protects against apoptosis during stress, suggesting a role in trophoblast survival in the relatively low O(2) ( approximately 2%) environment of the first trimester conceptus. Using a well-characterized human first trimester cytotrophoblast cell line, we found that a 4-hour exposure to 2% O(2) upregulates HBEGF synthesis and secretion independently of an increase in its mRNA. Five other expressed members of the EGF family are largely unaffected. At 2% O(2), signaling via HER1 or HER4, known HBEGF receptors, is required for both HBEGF upregulation and protection against apoptosis. This positive-feedback loop is dependent on metalloproteinase-mediated cleavage and shedding of the HBEGF ectodomain. The restoration of trophoblast survival by the addition of soluble HBEGF in cultures exposed to low O(2) and metalloproteinase inhibitor suggests that the effects of HBEGF are mediated by autocrine/paracrine, rather than juxtacrine, signaling. Our results provide evidence that a post-transcriptional mechanism induced in trophoblasts by low O(2) rapidly amplifies HBEGF signaling to inhibit apoptosis. These findings have a high clinical significance, as the downregulation of HBEGF in pre-eclampsia is likely to be a contributing factor leading to the demise of trophoblasts.

Mr 25,000 heparin-binding protein from guinea pig brain is a high molecular weight form of basic fibroblast growth factor.

OpenAIRE

Moscatelli, D; Joseph-Silverstein, J; Manejias, R; Rifkin, D B

1987-01-01

A Mr 25,000 form of basic fibroblast growth factor (bFGF) has been isolated from guinea pig brain along with the typical Mr 18,000 form. Both forms were purified to homogeneity by a combination of heparin-affinity chromatography and ion-exchange chromatography on an FPLC Mono S column. The Mr 25,000 form, like the Mr 18,000 form, was not eluted from the heparin-affinity column with 0.95 M NaCl, but was eluted with 2 M NaCl. The Mr 25,000 guinea pig protein stimulated plasminogen activator pro...

M/sub r/ 25,000 heparin-binding protein from guinea pig brain is a high molecular weight form of basic fibroblast growth factor

International Nuclear Information System (INIS)

Moscatelli, D.; Joseph-Silverstein, J.; Manejias, R.; Rifkin, D.B.

1987-01-01

A M/sub r/ 25,000 form of basic fibroblast growth factor (bFGF) has been isolated from guinea pig grain along with the typical M/sub r/ 18,000 form. Both forms were purified to homogeneity by a combination of heparin-affinity chromatography and ion-exchange chromatography on an FPLC Mono S column. The M/sub r/ 25,000 form, like the M/sub r/ 18,000 form was not eluted from the heparin-affinity column with 0.95 M NaCl, but was eluted with 2 M NaCl. The M/sub r/ 25,000 guinea pig protein stimulated plasminogen activator production by cultured bovine capillary endothelial cells in a dose-dependent manner at concentration of 0.1-10 ngml, the same range that was effective for guinea pig and human M/sub r/ 18,000 bFGFs. The binding of human 125 I-labeled bFGF to baby hamster kidney cells is inhibited equally by the M/sub r/ 25,000 guinea pig protein and the M/sub r/ 18,000 guinea pig and human bFGFs. Polyclonal antibodies raised against human bFGF recognize both the M/sub r/ 25,000 and 18,000 guinea pig proteins in an immunoblot analysis. In a radioimmunoassay, both the M/sub r/ 25,000 and M/sub r/ 18,000 guinea pig proteins compete equally well with iodinated human bFGF for binding to the anti-human bFGF antibodies. When treated with low concentrations of trypsin, the M/sub r/ 25,000 guinea pig bFGF was converted to a M/sub r/ 18,000 protein. These results show that the two molecules are closely related and suggest that the M/sub r/ 25,000 protein shares substantial homology with the M/sub r/ 18,000 bFGF

Phosphatidylserine and Phosphatidylethanolamine Bind to Protein Z Cooperatively and with Equal Affinity.

Directory of Open Access Journals (Sweden)

Tanusree Sengupta

Full Text Available Protein Z (PZ is an anticoagulant that binds with high affinity to Protein Z-dependent protease inhibitor (ZPI and accelerates the rate of ZPI-mediated inhibition of factor Xa (fXa by more than 1000-fold in the presence of Ca2+ and phospholipids. PZ promotion of the ZPI-fXa interaction results from the anchoring of the Gla domain of PZ onto phospholipid surfaces and positioning the bound ZPI in close proximity to the Gla-anchored fXa, forming a ternary complex of PZ/ZPI/fXa. Although interaction of PZ with phospholipid membrane appears to be absolutely crucial for its cofactor activity, little is known about the binding of different phospholipids to PZ. The present study was conceived to understand the interaction of different phospholipids with PZ. Experiments with both soluble lipids and model membranes revealed that PZ binds to phosphatidylserine (PS and phosphatidylethanolamine (PE with equal affinity (Kd~48 μM; further, PS and PE bound to PZ synergistically. Equilibrium dialysis experiments revealed two lipid-binding sites for both PS and PE. PZ binds with weaker affinity to other phospholipids, e.g., phosphatidic acid, phosphatidylglycerol, phosphatidylcholine and binding of these lipids is not synergistic with respect to PS. Both PS and PE -containing membranes supported the formation of a fXa-PZ complex. PZ protection of fXa from antithrombin inhibition were also shown to be comparable in presence of both PS: PC and PE: PC membranes. These findings are particularly important and intriguing since they suggest a special affinity of PZ, in vivo, towards activated platelets, the primary membrane involved in blood coagulation process.

Targeting hepatic heparin-binding EGF-like growth factor (HB-EGF) induces anti-hyperlipidemia leading to reduction of angiotensin II-induced aneurysm development.

Science.gov (United States)

Kim, Seonwook; Yang, Lihua; Kim, Seongu; Lee, Richard G; Graham, Mark J; Berliner, Judith A; Lusis, Aldons J; Cai, Lei; Temel, Ryan E; Rateri, Debra L; Lee, Sangderk

2017-01-01

The upregulated expression of heparin binding EGF-like growth factor (HB-EGF) in the vessel and circulation is associated with risk of cardiovascular disease. In this study, we tested the effects of HB-EGF targeting using HB-EGF-specific antisense oligonucleotide (ASO) on the development of aortic aneurysm in a mouse aneurysm model. Low-density lipoprotein receptor (LDLR) deficient mice (male, 16 weeks of age) were injected with control and HB-EGF ASOs for 10 weeks. To induce aneurysm, the mice were fed a high fat diet (22% fat, 0.2% cholesterol; w/w) at 5 week point of ASO administration and infused with angiotensin II (AngII, 1,000ng/kg/min) for the last 4 weeks of ASO administration. We confirmed that the HB-EGF ASO administration significantly downregulated HB-EGF expression in multiple tissues including the liver. Importantly, the HB-EGF ASO administration significantly suppressed development of aortic aneurysms including thoracic and abdominal types. Interestingly, the HB-EGF ASO administration induced a remarkable anti-hyperlipidemic effect by suppressing very low density lipoprotein (VLDL) level in the blood. Mechanistically, the HB-EGF targeting suppressed hepatic VLDL secretion rate without changing heparin-releasable plasma triglyceride (TG) hydrolytic activity or fecal neutral cholesterol excretion rate. This result suggested that the HB-EGF targeting induced protection against aneurysm development through anti-hyperlipidemic effects. Suppression of hepatic VLDL production process appears to be a key mechanism for the anti-hyperlipidemic effects by the HB-EGF targeting.

Pharmacokinetics of heparin and related polysaccharides

International Nuclear Information System (INIS)

Boneu, B.; Dol, F.; Caranobe, C.; Sie, P.; Houin, G.

1989-01-01

The pharmacodynamic profile of standard heparin (SH), a low molecular weight derivative (CY 216) and of dermatan sulfate (DS), a new potential antithrombotic drug, was investigated in the rabbit over a large range of doses. After bolus i.v. injection of low doses, the biological activity of SH disappeared exponentially; however, its half-life was prolonged when the dose injected increased, and over 158 micrograms/kg (100 anti-factor Xa U/kg) the biological activity disappeared as a concave-convex curve. CY 216 disappeared more slowly than SH at low doses but faster than SH at higher doses. More than 90% of the DS biological activity present 1 minute after the i.v. injection disappeared exponentially without dose-dependent effects. Increasing doses of the three drugs were then delivered for 5 h under continuous infusions. Below 500 micrograms/kg/h the DS and CY 216 plateau concentrations were higher than that of SH while above this dose the SH concentration was higher than that of DS and CY 216. These observations may be explained by the results of pharmacokinetics experiments where 125 I-labeled compounds were delivered by bolus i.v. injection in association with increasing doses of their unlabeled counterparts. For SH there was a 10-fold difference between the half-life of the lower dose (32 micrograms/kg or 5 anti-factor Xa U/kg) and that of the higher dose (3200 micrograms/kg); it was demonstrated that the half-life of SH continuously shortened as its plasma concentration decreased. In contrast the CY 216 and DS half-lives were very close, independent of the dose delivered, and therefore longer than that of SH at low doses and shorter than that of SH at higher doses

Heparin: The Silver Bullet of Aneurysmal Subarachnoid Hemorrhage?

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Nicolas K. Khattar

2018-03-01

Full Text Available Various neurological diseases have recently been associated with neuroinflammation and worsening outcomes. Subarachnoid hemorrhage has been shown to generate a potent neuroinflammatory response. Heparin is a potential effective anti-inflammatory agent to prevent initial injury as well as delayed neurological decline. Different mechanisms of action for heparin have been proposed including, but not limited to the binding and neutralization of oxyhemoglobin, decreased transcription and signal transduction of endothelin-1, inhibition of binding to vessel wall selectins and vascular leakage into the subarachnoid space as well as direct binding and neutralization of inflammatory molecules. With a reasonably safe side-effect profile, heparin has shown significant promise in small series in human studies of aneurysmal subarachnoid hemorrhage in decreasing both initial and delayed neurological injury. Further studies are needed to validate various neuroprotective features of heparin in subarachnoid hemorrhage as well as other disease states.

Low-molecular weight heparin increases circulating sFlt-1 levels and enhances urinary elimination.

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Henning Hagmann

Full Text Available RATIONALE: Preeclampsia is a devastating medical complication of pregnancy which leads to maternal and fetal morbidity and mortality. While the etiology of preeclampsia is unclear, human and animal studies suggest that excessive circulating levels of soluble fms-like tyrosine-kinase-1 (sFlt-1, an alternatively spliced variant of VEGF-receptor1, contribute to the signs and symptoms of preeclampsia. Since sFlt-1 binds to heparin and heparan sulfate proteoglycans, we hypothesized that the anticoagulant heparin, which is often used in pregnancy, may interfere with the levels, distribution and elimination of sFlt-1 in vivo. OBJECTIVE: We systematically determined serum and urine levels of angiogenic factors in preeclamptic women before and after administration of low molecular weight heparin and further characterized the interaction with heparin in biochemical studies. METHODS AND RESULTS: Serum and urine samples were used to measure sFlt-1 levels before and after heparin administration. Serum levels of sFlt-1 increased by 25% after heparin administration in pregnant women. The magnitude of the increase in circulating sFlt-1 correlated with initial sFlt-1 serum levels. Urinary sFlt-1 levels were also elevated following heparin administration and levels of elimination were dependent on the underlying integrity of the glomerular filtration barrier. Biochemical binding studies employing cation exchange chromatography revealed that heparin bound sFlt-1 had decreased affinity to negatively charged surfaces when compared to sFlt-1 alone. CONCLUSION: Low molecular weight heparin administration increased circulating sFlt1 levels and enhanced renal elimination. We provide evidence that both effects may be due to heparin binding to sFlt1 and masking the positive charges on sFlt1 protein.

Marker-aided Incorporation of Xa38, a Novel Bacterial Blight Resistance Gene, in PB1121 and Comparison of its Resistance Spectrum with xa13 + Xa21.

Science.gov (United States)

Ellur, Ranjith K; Khanna, Apurva; S, Gopala Krishnan; Bhowmick, Prolay K; Vinod, K K; Nagarajan, M; Mondal, Kalyan K; Singh, Nagendra K; Singh, Kuldeep; Prabhu, Kumble Vinod; Singh, Ashok K

2016-07-11

Basmati rice is preferred internationally because of its appealing taste, mouth feel and aroma. Pusa Basmati 1121 (PB1121) is a widely grown variety known for its excellent grain and cooking quality in the international and domestic market. It contributes approximately USD 3 billion to India's forex earning annually by being the most traded variety. However, PB1121 is highly susceptible to bacterial blight (BB) disease. A novel BB resistance gene Xa38 was incorporated in PB1121 from donor parent PR114-Xa38 using a modified marker-assisted backcross breeding (MABB) scheme. Phenotypic selection prior to background selection was instrumental in identifying the novel recombinants with maximum recovery of recurrent parent phenome. The strategy was effective in delimiting the linkage drag to <0.5 mb upstream and <1.9 mb downstream of Xa38 with recurrent parent genome recovery upto 96.9% in the developed NILs. The NILs of PB1121 carrying Xa38 were compared with PB1121 NILs carrying xa13 + Xa21 (developed earlier in our lab) for their resistance to BB. Both NILs showed resistance against the Xoo races 1, 2, 3 and 6. Additionally, Xa38 also resisted Xoo race 5 to which xa13 + Xa21 was susceptible. The PB1121 NILs carrying Xa38 gene will provide effective control of BB in the Basmati growing region.

Anti-tumor activity of a novel HS-mimetic-vascular endothelial growth factor binding small molecule.

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Kazuyuki Sugahara

Full Text Available The angiogenic process is controlled by variety of factors of which the vascular endothelial growth factor (VEGF pathway plays a major role. A series of heparan sulfate mimetic small molecules targeting VEGF/VEGFR pathway has been synthesized. Among them, compound 8 (2-butyl-5-chloro-3-(4-nitro-benzyl-3H-imidazole-4-carbaldehyde was identified as a significant binding molecule for the heparin-binding domain of VEGF, determined by high-throughput-surface plasmon resonance assay. The data predicted strong binding of compound 8 with VEGF which may prevent the binding of VEGF to its receptor. We compared the structure of compound 8 with heparan sulfate (HS, which have in common the functional ionic groups such as sulfate, nitro and carbaldehyde that can be located in similar positions of the disaccharide structure of HS. Molecular docking studies predicted that compound 8 binds at the heparin binding domain of VEGF through strong hydrogen bonding with Lys-30 and Gln-20 amino acid residues, and consistent with the prediction, compound 8 inhibited binding of VEGF to immobilized heparin. In vitro studies showed that compound 8 inhibits the VEGF-induced proliferation migration and tube formation of mouse vascular endothelial cells, and finally the invasion of a murine osteosarcoma cell line (LM8G7 which secrets high levels of VEGF. In vivo, these effects produce significant decrease of tumor burden in an experimental model of liver metastasis. Collectively, these data indicate that compound 8 may prevent tumor growth through a direct effect on tumor cell proliferation and by inhibition of endothelial cell migration and angiogenesis mediated by VEGF. In conclusion, compound 8 may normalize the tumor vasculature and microenvironment in tumors probably by inhibiting the binding of VEGF to its receptor.

Platelet-derived growth factor inhibits platelet activation in heparinized whole blood.

Science.gov (United States)

Selheim, F; Holmsen, H; Vassbotn, F S

1999-08-15

We previously have demonstrated that human platelets have functionally active platelet-derived growth factor alpha-receptors. Studies with gel-filtered platelets showed that an autocrine inhibition pathway is transduced through this tyrosine kinase receptor during platelet activation. The physiological significance of this inhibitory effect of platelet-derived growth factor on gel-filtered platelets activation is, however, not known. In the present study, we investigated whether platelet-derived growth factor inhibits platelet activation under more physiological conditions in heparinized whole blood, which represents a more physiological condition than gel-filtered platelets. Using flow cytometric assays, we demonstrate here that platelet-derived growth factor inhibits thrombin-, thrombin receptor agonist peptide SFLLRN-, and collagen-induced platelet aggregation and shedding of platelet-derived microparticles from the platelet plasma membrane during platelet aggregation in stirred heparinized whole blood. The inhibitory effect of platelet-derived growth factor was dose dependent. However, under nonaggregating conditions (no stirring), we could not demonstrate any significant effect of platelet-derived growth factor on thrombin- and thrombin receptor agonist peptide-induced platelet surface expression of P-selectin. Our results demonstrate that platelet-derived growth factor appears to be a true antithrombotic agent only under aggregating conditions in heparinized whole blood.

Marker-aided Incorporation of Xa38, a Novel Bacterial Blight Resistance Gene, in PB1121 and Comparison of its Resistance Spectrum with xa13 + Xa21

Science.gov (United States)

Ellur, Ranjith K.; Khanna, Apurva; S, Gopala Krishnan.; Bhowmick, Prolay K.; Vinod, K. K.; Nagarajan, M.; Mondal, Kalyan K.; Singh, Nagendra K.; Singh, Kuldeep; Prabhu, Kumble Vinod; Singh, Ashok K.

2016-01-01

Basmati rice is preferred internationally because of its appealing taste, mouth feel and aroma. Pusa Basmati 1121 (PB1121) is a widely grown variety known for its excellent grain and cooking quality in the international and domestic market. It contributes approximately USD 3 billion to India’s forex earning annually by being the most traded variety. However, PB1121 is highly susceptible to bacterial blight (BB) disease. A novel BB resistance gene Xa38 was incorporated in PB1121 from donor parent PR114-Xa38 using a modified marker-assisted backcross breeding (MABB) scheme. Phenotypic selection prior to background selection was instrumental in identifying the novel recombinants with maximum recovery of recurrent parent phenome. The strategy was effective in delimiting the linkage drag to <0.5 mb upstream and <1.9 mb downstream of Xa38 with recurrent parent genome recovery upto 96.9% in the developed NILs. The NILs of PB1121 carrying Xa38 were compared with PB1121 NILs carrying xa13 + Xa21 (developed earlier in our lab) for their resistance to BB. Both NILs showed resistance against the Xoo races 1, 2, 3 and 6. Additionally, Xa38 also resisted Xoo race 5 to which xa13 + Xa21 was susceptible. The PB1121 NILs carrying Xa38 gene will provide effective control of BB in the Basmati growing region. PMID:27403778

AVE5026, a new hemisynthetic ultra-low-molecular-weight heparin for the prevention of venous thromboembolism in patients after total knee replacement surgery

DEFF Research Database (Denmark)

Lassen, Michael Rud; Dahl, O E; Mismetti, P

2009-01-01

BACKGROUND: AVE5026 is a new hemisynthetic ultra-low-molecular-weight heparin, with a novel anti-thrombotic profile resulting from high anti-factor (F)Xa activity and residual anti-FIIa activity. AVE5026 is in clinical development for venous thromboembolism (VTE) prevention, a frequent complication....... The primary safety outcome was the incidence of major bleeding. RESULTS: The primary efficacy outcome was assessed in 464 patients. There was a significant dose-response across the five AVE5026 groups for VTE prevention (Pincidence of VTE ranging from 5.3% to 44.1% compared with 35...

A mutation in the heparin-binding site of noggin as a novel mechanism of proximal symphalangism and conductive hearing loss.

Science.gov (United States)

Masuda, Sawako; Namba, Kazunori; Mutai, Hideki; Usui, Satoko; Miyanaga, Yuko; Kaneko, Hiroki; Matsunaga, Tatsuo

2014-05-09

The access of bone morphogenetic protein (BMP) to the BMP receptors on the cell surface is regulated by its antagonist noggin, which binds to heparan-sulfate proteoglycans on the cell surface. Noggin is encoded by NOG and mutations in the gene are associated with aberrant skeletal formation, such as in the autosomal dominant disorders proximal symphalangism (SYM1), multiple synostoses syndrome, Teunissen-Cremers syndrome, and tarsal-carpal coalition syndrome. NOG mutations affecting a specific function may produce a distinct phenotype. In this study, we investigated a Japanese pedigree with SYM1 and conductive hearing loss and found that it carried a novel heterozygous missense mutation of NOG (c.406C>T; p.R136C) affecting the heparin-binding site of noggin. As no mutations of the heparin-binding site of noggin have previously been reported, we investigated the crystal structure of wild-type noggin to investigate molecular mechanism of the p.R136C mutation. We found that the positively charged arginine at position 136 was predicted to be important for binding to the negatively charged heparan-sulfate proteoglycan (HSPG). An in silico docking analysis showed that one of the salt bridges between noggin and heparin disappeared following the replacement of the arginine with a non-charged cysteine. We propose that the decreased binding affinity of NOG with the p.R136C mutation to HSPG leads to an excess of BMP signaling and underlies the SYM1 and conductive hearing loss phenotype of carriers. Copyright © 2014 Elsevier Inc. All rights reserved.

Structure-Activity Relationships of Bioengineered Heparin/Heparan Sulfates Produced in Different Bioreactors

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Ha Na Kim

2017-05-01

Full Text Available Heparin and heparan sulfate are structurally-related carbohydrates with therapeutic applications in anticoagulation, drug delivery, and regenerative medicine. This study explored the effect of different bioreactor conditions on the production of heparin/heparan sulfate chains via the recombinant expression of serglycin in mammalian cells. Tissue culture flasks and continuously-stirred tank reactors promoted the production of serglycin decorated with heparin/heparan sulfate, as well as chondroitin sulfate, while the serglycin secreted by cells in the tissue culture flasks produced more highly-sulfated heparin/heparan sulfate chains. The serglycin produced in tissue culture flasks was effective in binding and signaling fibroblast growth factor 2, indicating the utility of this molecule in drug delivery and regenerative medicine applications in addition to its well-known anticoagulant activity.

Apixaban versus enoxaparin for thromboprophylaxis after hip replacement

DEFF Research Database (Denmark)

Lassen, Michael Rud; Gallus, Alexander; Raskob, Gary E

2010-01-01

There are various regimens for thromboprophylaxis after hip replacement. Low-molecular-weight heparins such as enoxaparin predominantly inhibit factor Xa but also inhibit thrombin to some degree. Orally active, specific factor Xa inhibitors such as apixaban may provide effective thromboprophylaxis...

Sequence similarity between the erythrocyte binding domain of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals a functional heparin binding motif involved in binding to the Duffy antigen receptor for chemokines

OpenAIRE

Bolton, Michael J; Garry, Robert F

2011-01-01

Abstract Background The HIV surface glycoprotein gp120 (SU, gp120) and the Plasmodium vivax Duffy binding protein (PvDBP) bind to chemokine receptors during infection and have a site of amino acid sequence similarity in their binding domains that often includes a heparin binding motif (HBM). Infection by either pathogen has been found to be inhibited by polyanions. Results Specific polyanions that inhibit HIV infection and bind to the V3 loop of X4 strains also inhibited DBP-mediated infectio...

Cleavage of spike protein of SARS coronavirus by protease factor Xa is associated with viral infectivity

International Nuclear Information System (INIS)

Du, Lanying; Kao, Richard Y.; Zhou, Yusen; He, Yuxian; Zhao, Guangyu; Wong, Charlotte; Jiang, Shibo; Yuen, Kwok-Yung; Jin, Dong-Yan; Zheng, Bo-Jian

2007-01-01

The spike (S) protein of SARS coronavirus (SARS-CoV) has been known to recognize and bind to host receptors, whose conformational changes then facilitate fusion between the viral envelope and host cell membrane, leading to viral entry into target cells. However, other functions of SARS-CoV S protein such as proteolytic cleavage and its implications to viral infection are incompletely understood. In this study, we demonstrated that the infection of SARS-CoV and a pseudovirus bearing the S protein of SARS-CoV was inhibited by a protease inhibitor Ben-HCl. Also, the protease Factor Xa, a target of Ben-HCl abundantly expressed in infected cells, was able to cleave the recombinant and pseudoviral S protein into S1 and S2 subunits, and the cleavage was inhibited by Ben-HCl. Furthermore, this cleavage correlated with the infectivity of the pseudovirus. Taken together, our study suggests a plausible mechanism by which SARS-CoV cleaves its S protein to facilitate viral infection

Nerve growth factor loaded heparin/chitosan scaffolds for accelerating peripheral nerve regeneration.

Science.gov (United States)

Li, Guicai; Xiao, Qinzhi; Zhang, Luzhong; Zhao, Yahong; Yang, Yumin

2017-09-01

Artificial chitosan scaffolds have been widely investigated for peripheral nerve regeneration. However, the effect was not as good as that of autologous grafts and therefore could not meet the clinical requirement. In the present study, the nerve growth factor (NGF) loaded heparin/chitosan scaffolds were fabricated via electrostatic interaction for further improving nerve regeneration. The physicochemical properties including morphology, wettability and composition were measured. The heparin immobilization, NGF loading and release were quantitatively and qualitatively characterized, respectively. The effect of NGF loaded heparin/chitosan scaffolds on nerve regeneration was evaluated by Schwann cells culture for different periods. The results showed that the heparin immobilization and NGF loading did not cause the change of bulk properties of chitosan scaffolds except for morphology and wettability. The pre-immobilization of heparin in chitosan scaffolds could enhance the stability of subsequently loaded NGF. The NGF loaded heparin/chitosan scaffolds could obviously improve the attachment and proliferation of Schwann cells in vitro. More importantly, the NGF loaded heparin/chitosan scaffolds could effectively promote the morphology development of Schwann cells. The study may provide a useful experimental basis to design and develop artificial implants for peripheral nerve regeneration and other tissue regeneration. Copyright © 2017 Elsevier Ltd. All rights reserved.

Cell adhesion to fibrillin-1: identification of an Arg-Gly-Asp-dependent synergy region and a heparin-binding site that regulates focal adhesion formation

DEFF Research Database (Denmark)

Bax, Daniel V; Mahalingam, Yashithra; Cain, Stuart

2007-01-01

We have defined the molecular basis of cell adhesion to fibrillin-1, the major structural component of extracellular microfibrils that are associated with elastic fibres. Using human dermal fibroblasts, and recombinant domain swap fragments containing the Arg-Gly-Asp motif, we have demonstrated...... a requirement for upstream domains for integrin-alpha(5)beta(1)-mediated cell adhesion and migration. An adjacent heparin-binding site, which supports focal adhesion formation, was mapped to the fibrillin-1 TB5 motif. Site-directed mutagenesis revealed two arginine residues that are crucial for heparin binding...

The clinical significance and risk factors of anti-platelet factor 4/heparin antibody on maintenance hemodialysis patients: a two-year prospective follow-up.

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Delong Zhao

Full Text Available BACKGROUND: Heparin-induced thrombocytopenia is an immune response mediated by anti-PF4/heparin antibody, which is clinically characterized by thrombocytopenia and thromboembolic events. In this study, a prospective and multi-center clinical investigation 1 determined the positive rate of anti-PF4/heparin antibody in maintenance hemodialysis patients in China, 2 identified the related risk factors, and 3 further explored the effect of the anti-PF4/heparin antibody on bleeding, thromboembolic events, and risk of death in the patients. METHODS: The serum anti-PF4/heparin antibody was measured in 661 patients from nine hemodialysis centers, detected by IgG-specific ELISA and followed by confirmation with excess heparin. Risk factors of these patients were analyzed. Based on a two-year follow-up, the association between the anti-PF4/heparin antibody and bleeding, thromboembolic events, and risk of death in the patients was investigated. RESULTS: 1 The positivity rate of the anti-PF4/heparin antibody in maintenance hemodialysis patients was 5.6%. With diabetes as an independent risk factor, the positivity rate of the anti-PF4/heparin antibody decreased in the patients undergoing weekly dialyses ≥3 times. 2 The positivity rate of the anti-PF4/heparin antibody was not related to the occurrence of clinical thromboembolic events and was not a risk factor for death within two years in maintenance hemodialysis patients. 3 Negativity for the anti-PF4/heparin antibody combined with a reduction of the platelet count or combined with the administration of antiplatelet drugs yielded a significant increase in bleeding events. However, the composite determination of the anti-PF4/heparin antibody and thrombocytopenia, as well as the administration of antiplatelet drugs, was not predictive for the risk of thromboembolic events in the maintenance hemodialysis patients. CONCLUSIONS: A single detection of the anti-PF4/heparin antibody did not predict the occurrence

Anti-Platelet Factor 4/Heparin Antibody Formation Occurs Endogenously and at Unexpected High Frequency in Polycythemia Vera

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Sara C. Meyer

2017-01-01

Full Text Available Background. Myeloproliferative neoplasms (MPN encounter thromboses due to multiple known risk factors. Heparin-induced thrombocytopenia (HIT is a thrombotic syndrome mediated by anti-platelet factor 4 (PF4/heparin antibodies with undetermined significance for thrombosis in MPN. We hypothesized that anti-PF4/heparin Ab might occur in MPN and promote thrombosis. Methods. Anti-PF4/heparin antibodies were analyzed in 127 MPN patients including 76 PV and 51 ET. Screening, validation testing, and isotype testing of anti-PF4/heparin Ab were correlated with disease characteristics. Results. Anti-PF4/heparin antibodies were detected in 21% of PV and 12% of ET versus 0.3–3% in heparin-exposed patients. Validation testing confirmed anti-PF4/heparin immunoglobulins in 15% of PV and 10% of ET. Isotype testing detected 9.2% IgG and 5.3% IgM in PV and exclusively IgM in ET. IgG-positive PV patients encountered thromboses in 57.1% suggesting anti-PF4/heparin IgG may contribute to higher risk for thrombosis in MPN. Overall, 45% of PV patients experienced thromboses with 11.8% positive for anti-PF4/heparin IgG versus 7.1% in PV without thrombosis. Conclusion. Anti-PF4/heparin antibodies occur endogenously and more frequently in MPN than upon heparin exposure. Thrombotic risk increases in anti-PF4/heparin IgG-positive PV reflecting potential implications and calling for larger, confirmatory cohorts. Anti-PF4/heparin IgG should be assessed upon thrombosis in PV to facilitate avoidance of heparin in anti-PF4/heparin IgG-positive PV.

The heparin-binding domain of HB-EGF mediates localization to sites of cell-cell contact and prevents HB-EGF proteolytic release

Energy Technology Data Exchange (ETDEWEB)

Prince, Robin N.; Schreiter, Eric R.; Zou, Peng; Wiley, H. S.; Ting, Alice Y.; Lee, Richard T.; Lauffenburger, Douglas A.

2010-07-01

Heparin-binding EGF-like growth factor (HB-EGF) is a ligand for EGF receptor (EGFR) and possesses the ability to signal in juxtacrine, autocrine and/or paracrine mode, with these alternatives being governed by the degree of proteolytic release of the ligand. Although the spatial range of diffusion of released HB-EGF is restricted by binding heparan-sulfate proteoglycans (HSPGs) in the extracellular matrix and/or cellular glycocalyx, ascertaining mechanisms governing non-released HB-EGF localization is also important for understanding its effects. We have employed a new method for independently tracking the localization of the extracellular EGFlike domain of HB-EGF and the cytoplasmic C-terminus. A striking observation was the absence of the HB-EGF transmembrane proform from the leading edge of COS-7 cells in a wound-closure assay; instead, this protein localized in regions of cell-cell contact. A battery of detailed experiments found that this localization derives from a trans interaction between extracellular HSPGs and the HBEGF heparin-binding domain, and that disruption of this interaction leads to increased release of soluble ligand and a switch in cell phenotype from juxtacrine-induced growth inhibition to autocrine-induced proliferation. Our results indicate that extracellular HSPGs serve to sequester the transmembrane pro-form of HB-EGF at the point of cell-cell contact, and that this plays a role in governing the balance between juxtacrine versus autocrine and paracrine signaling.

Selective interaction of heparin with the variable region 3 within surface glycoprotein of laboratory-adapted feline immunodeficiency virus.

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Qiong-Ying Hu

Full Text Available Heparan sulfate proteoglycans (HSPG can act as binding receptors for certain laboratory-adapted (TCA strains of feline immunodeficiency virus (FIV and human immunodeficiency virus (HIV. Heparin, a soluble heparin sulfate (HS, can inhibit TCA HIV and FIV entry mediated by HSPG interaction in vitro. In the present study, we further determined the selective interaction of heparin with the V3 loop of TCA of FIV. Our current results indicate that heparin selectively inhibits infection by TCA strains, but not for field isolates (FS. Heparin also specifically interferes with TCA surface glycoprotein (SU binding to CXCR4, by interactions with HSPG binding sites on the V3 loop of the FIV envelope protein. Peptides representing either the N- or C-terminal side of the V3 loop and containing HSPG binding sites were able to compete away the heparin block of TCA SU binding to CXCR4. Heparin does not interfere with the interaction of SU with anti-V3 antibodies that target the CXCR4 binding region or with the interaction between FS FIV and anti-V3 antibodies since FS SU has no HSPG binding sites within the HSPG binding region. Our data show that heparin blocks TCA FIV infection or entry not only through its competition of HSPG on the cell surface interaction with SU, but also by its interference with CXCR4 binding to SU. These studies aid in the design and development of heparin derivatives or analogues that can inhibit steps in virus infection and are informative regarding the HSPG/SU interaction.

AVE5026, a new hemisynthetic ultra-low-molecular-weight heparin for the prevention of venous thromboembolism in patients after total knee replacement surgery--TREK: a dose-ranging study

DEFF Research Database (Denmark)

Lassen, M R; Dahl, O E; Mismetti, P

2009-01-01

BACKGROUND: AVE5026 is a new hemisynthetic ultra-low-molecular-weight heparin, with a novel anti-thrombotic profile resulting from high anti-factor (F)Xa activity and residual anti-FIIa activity. AVE5026 is in clinical development for venous thromboembolism (VTE) prevention, a frequent complication....... The primary safety outcome was the incidence of major bleeding. RESULTS: The primary efficacy outcome was assessed in 464 patients. There was a significant dose-response across the five AVE5026 groups for VTE prevention (Pincidence of VTE ranging from 5.3% to 44.1% compared with 35...

D-fructose-binding proteins in bull seminal plasma: Isolation and characterization

Czech Academy of Sciences Publication Activity Database

Liberda, J.; Kraus, Marek; Ryšlavá, H.; Vlasáková, M.; Jonáková, Věra; Tichá, M.

2001-01-01

Roč. 47, č. 4 (2001), s. 113-119 ISSN 0015-5500 R&D Projects: GA ČR GA303/99/0357; GA ČR GV524/96/K162 Institutional research plan: CEZ:AV0Z5052915 Keywords : bull seminal plasma * non-heparin-binding and heparin-binding proteins * D-fructose-binding proteins Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.519, year: 2001

FGF growth factor analogs

Science.gov (United States)

Zamora, Paul O [Gaithersburg, MD; Pena, Louis A [Poquott, NY; Lin, Xinhua [Plainview, NY; Takahashi, Kazuyuki [Germantown, MD

2012-07-24

The present invention provides a fibroblast growth factor heparin-binding analog of the formula: ##STR00001## where R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5, X, Y and Z are as defined, pharmaceutical compositions, coating compositions and medical devices including the fibroblast growth factor heparin-binding analog of the foregoing formula, and methods and uses thereof.

Influence of heparin on radioimmunological assay of ACTH

International Nuclear Information System (INIS)

Dupouy, J.P.; Godaut, M.; Chatelain, A.

1986-01-01

1 - Heparin traps plasma ACTH, promoting the formation of aggregates with apparent high molecular weight as shown by chromatography on Sephadex G 50 fine columns. The percentage of 125 I-ACTH which appeared in the void volume of the column, increased linearly with the log. dose of heparin. 2 - Heparin at concentrations of up to 100 IU/ml does not impair ACTH adsorption on either silicic acid or Quso G 32 as well as further elution by acetic acid/acetone/water (I: 40: 59; V/V) or HCl O.I N. Silicic acid traps selectively ACTH but not heparin. 3 - Heparin interferes with direct RIA-ACTH in the plasma by decreasing 125 I-ACTH binding to the antibodies and modifying the slope of the standard curve. Unsuitable artefacts induced by heparin, as overestimation or underestimation of plasma ACTH levels by RIA, can be avoid by previous hormone extraction from heparinized plasmas. Such results emphasized the importance of the sample preparation in order to obtain consistent results [fr

A need for evidence-based clinical practice guidelines for the use of heparins in the elderly

Directory of Open Access Journals (Sweden)

Isabelle Gouin-Thibault

2010-04-01

Full Text Available Isabelle Gouin-Thibault1,2, Virginie Siguret1,2, Eric Pautas2,31Assistance Publique Hôpitaux de Paris, Laboratoire d’Hématologie, Hôpital Charles Foix, Paris, France; 2Université Paris Descartes, INSERM U, Paris, France; 3Assistance Publique Hôpitaux de Paris, Unité de Gériatrie Aiguë, Hôpital Charles Foix, Paris, FranceAbstract: Low-molecular-weight heparins (LMWHs have been widely studied in pivotal clinical trials or in several meta-analyses. However, the safety and optimal use of LMWHs in high-risk patients such as the very elderly remains uncertain since these patients are usually excluded from clinical trials. In terms of LMWHs in the elderly, the main concerns are renal failure and the risk of accumulation. A clinical approach consisting of a LMWH dose reduction in the elderly should be considered with great caution in terms of efficacy, since it has been tested neither in the treatment of VTE nor in VTE prophylaxis. If monitoring is considered in patients receiving therapeutic dose LMWHs, appropriate target ranges for peak anti-Xa activity levels should be used and so far, no anti-Xa activity-based guidelines have been issued. Moreover, no data support any laboratory monitoring in elderly patients treated with prophylactic dose LMWHs.Keywords: elderly patients, low-molecular-weight heparin, renal insufficiency, evidence-based medicine

Structural characterization of pharmaceutical heparins prepared from different animal tissues.

Science.gov (United States)

Fu, Li; Li, Guoyun; Yang, Bo; Onishi, Akihiro; Li, Lingyun; Sun, Peilong; Zhang, Fuming; Linhardt, Robert J

2013-05-01

Although most pharmaceutical heparin used today is obtained from porcine intestine, heparin has historically been prepared from bovine lung and ovine intestine. There is some regulatory concern about establishing the species origin of heparin. This concern began with the outbreak of mad cow disease in the 1990s and was exacerbated during the heparin shortage in the 2000s and the heparin contamination crisis of 2007-2008. Three heparins from porcine, ovine, and bovine were characterized through state-of-the-art carbohydrate analysis methods with a view profiling their physicochemical properties. Differences in molecular weight, monosaccharide and disaccharide composition, oligosaccharide sequence, and antithrombin III-binding affinity were observed. These data provide some insight into the variability of heparins obtained from these three species and suggest some analytical approaches that may be useful in confirming the species origin of a heparin active pharmaceutical ingredient. Copyright © 2013 Wiley Periodicals, Inc.

Heparin conjugated quantum dots for in vitro imaging applications.

Science.gov (United States)

Maguire, Ciaran Manus; Mahfoud, Omar Kazem; Rakovich, Tatsiana; Gerard, Valerie Anne; Prina-Mello, Adriele; Gun'ko, Yurii; Volkov, Yuri

2014-11-01

In this work heparin-gelatine multi-layered cadmium telluride quantum dots (QDgel/hep) were synthesised using a novel 'one-pot' method. The QDs produced were characterised using various spectroscopic and physiochemical techniques. Suitable QDs were then selected and compared to thioglycolic acid stabilised quantum dots (QDTGA) and gelatine coated quantum dots (QDgel) for utilisation in in vitro imaging experiments on live and fixed permeabilised THP-1, A549 and Caco-2 cell lines. Exposure of live THP-1 cells to QDgel/hep resulted in localisation of the QDs to the nucleus of the cells. QDgel/hep show affinity for the nuclear compartment of fixed permeabilised THP-1 and A549 cells but remain confined to cytoplasm of fixed permeabilised Caco-2 cells. It is postulated that heparin binding to the CD11b receptor facilitates the internalisation of the QDs into the nucleus of THP-1 cells. In addition, the heparin layer may reduce the unfavourable thrombogenic nature of quantum dots observed in vivo. In this study, heparin conjugated quantum dots were found to have superior imaging properties compared to its native counterparts. The authors postulate that heparin binding to the CD11b receptor facilitates QD internalization to the nucleus, and the heparin layer may reduce the in vivo thrombogenic properties of quantum dots. Copyright © 2014 Elsevier Inc. All rights reserved.

Conditional loss of heparin-binding EGF-like growth factor results in enhanced liver fibrosis after bile duct ligation in mice

Energy Technology Data Exchange (ETDEWEB)

Takemura, Takayo; Yoshida, Yuichi [Department of Gastroenterology and Hepatology, Osaka University, Graduate School of Medicine, Osaka (Japan); Kiso, Shinichi, E-mail: kiso@gh.med.osaka-u.ac.jp [Department of Gastroenterology and Hepatology, Osaka University, Graduate School of Medicine, Osaka (Japan); Kizu, Takashi; Furuta, Kunimaro; Ezaki, Hisao; Hamano, Mina; Egawa, Mayumi; Chatani, Norihiro; Kamada, Yoshihiro [Department of Gastroenterology and Hepatology, Osaka University, Graduate School of Medicine, Osaka (Japan); Imai, Yasuharu [Department of Gastroenterology, Ikeda Municipal Hospital, Ikeda, Osaka (Japan); Higashiyama, Shigeki [Department of Biochemistry and Molecular Genetics, Ehime University, Graduate School of Medicine and Department of Cell Growth and Tumor Regulation, Proteo-Medicine Research Center (ProMRes), Ehime University, Shitsukawa, Toon, Ehime (Japan); Iwamoto, Ryo; Mekada, Eisuke [Department of Cell Biology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka (Japan); Takehara, Tetsuo [Department of Gastroenterology and Hepatology, Osaka University, Graduate School of Medicine, Osaka (Japan)

2013-07-26

Highlights: •HB-EGF expression was increased during the development of liver fibrosis. •Conditional HB-EGF knockout mouse showed enhanced experimental liver fibrosis. •HB-EGF antagonized TGF-β-induced activation of hepatic stellate cells. •We report a possible protective role of HB-EGF in cholestatic liver fibrosis. -- Abstract: Our aims were to evaluate the involvement of heparin-binding EGF-like growth factor (HB-EGF) in liver fibrogenesis of humans and mice and to elucidate the effect of HB-EGF deficiency on cholestatic liver fibrosis using conditional HB-EGF knockout (KO) mice. We first demonstrated that gene expression of HB-EGF had a positive significant correlation with that of collagen in human fibrotic livers, and was increased in bile duct ligation (BDL)-induced fibrotic livers in mouse. We then generated conditional HB-EGF knockout (KO) mice using the interferon inducible Mx-1 promoter driven Cre recombinase transgene and wild type (WT) and KO mice were subjected to BDL. After BDL, KO mice exhibited enhanced liver fibrosis with increased expression of collagen, compared with WT mice. Finally, we used mouse hepatic stellate cells (HSCs) to examine the role of HB-EGF in the activation of these cells and showed that HB-EGF antagonized TGF-β-induced gene expression of collagen in mouse primary HSCs. Interestingly, HB-EGF did not prevent the TGF-β-induced nuclear accumulation of Smad3, but did lead to stabilization of the Smad transcriptional co-repressor TG-interacting factor. In conclusion, our data suggest a possible protective role of HB-EGF in cholestatic liver fibrosis.

Conditional loss of heparin-binding EGF-like growth factor results in enhanced liver fibrosis after bile duct ligation in mice

International Nuclear Information System (INIS)

Takemura, Takayo; Yoshida, Yuichi; Kiso, Shinichi; Kizu, Takashi; Furuta, Kunimaro; Ezaki, Hisao; Hamano, Mina; Egawa, Mayumi; Chatani, Norihiro; Kamada, Yoshihiro; Imai, Yasuharu; Higashiyama, Shigeki; Iwamoto, Ryo; Mekada, Eisuke; Takehara, Tetsuo

2013-01-01

Highlights: •HB-EGF expression was increased during the development of liver fibrosis. •Conditional HB-EGF knockout mouse showed enhanced experimental liver fibrosis. •HB-EGF antagonized TGF-β-induced activation of hepatic stellate cells. •We report a possible protective role of HB-EGF in cholestatic liver fibrosis. -- Abstract: Our aims were to evaluate the involvement of heparin-binding EGF-like growth factor (HB-EGF) in liver fibrogenesis of humans and mice and to elucidate the effect of HB-EGF deficiency on cholestatic liver fibrosis using conditional HB-EGF knockout (KO) mice. We first demonstrated that gene expression of HB-EGF had a positive significant correlation with that of collagen in human fibrotic livers, and was increased in bile duct ligation (BDL)-induced fibrotic livers in mouse. We then generated conditional HB-EGF knockout (KO) mice using the interferon inducible Mx-1 promoter driven Cre recombinase transgene and wild type (WT) and KO mice were subjected to BDL. After BDL, KO mice exhibited enhanced liver fibrosis with increased expression of collagen, compared with WT mice. Finally, we used mouse hepatic stellate cells (HSCs) to examine the role of HB-EGF in the activation of these cells and showed that HB-EGF antagonized TGF-β-induced gene expression of collagen in mouse primary HSCs. Interestingly, HB-EGF did not prevent the TGF-β-induced nuclear accumulation of Smad3, but did lead to stabilization of the Smad transcriptional co-repressor TG-interacting factor. In conclusion, our data suggest a possible protective role of HB-EGF in cholestatic liver fibrosis

Heparin (GAG-hed) inhibits LCR activity of Human Papillomavirus type 18 by decreasing AP1 binding

International Nuclear Information System (INIS)

Villanueva, Rita; Morales-Peza, Néstor; Castelán-Sánchez, Irma; García-Villa, Enrique; Tapia, Rocio; Cid-Arregui, Ángel; García-Carrancá, Alejandro; López-Bayghen, Esther; Gariglio, Patricio

2006-01-01

High risk HPVs are causative agents of anogenital cancers. Viral E6 and E7 genes are continuously expressed and are largely responsible for the oncogenic activity of these viruses. Transcription of the E6 and E7 genes is controlled by the viral Long Control Region (LCR), plus several cellular transcription factors including AP1 and the viral protein E2. Within the LCR, the binding and activity of the transcription factor AP1 represents a key regulatory event in maintaining E6/E7 gene expression and uncontrolled cell proliferation. Glycosaminoglycans (GAGs), such as heparin, can inhibit tumour growth; they have also shown antiviral effects and inhibition of AP1 transcriptional activity. The purpose of this study was to test the heparinoid GAG-hed, as a possible antiviral and antitumoral agent in an HPV18 positive HeLa cell line. Using in vivo and in vitro approaches we tested GAG-hed effects on HeLa tumour cell growth, cell proliferation and on the expression of HPV18 E6/E7 oncogenes. GAG-hed effects on AP1 binding to HPV18-LCR-DNA were tested by EMSA. We were able to record the antitumoral effect of GAG-hed in vivo by using as a model tumours induced by injection of HeLa cells into athymic female mice. The antiviral effect of GAG-hed resulted in the inhibition of LCR activity and, consequently, the inhibition of E6 and E7 transcription. A specific diminishing of cell proliferation rates was observed in HeLa but not in HPV-free colorectal adenocarcinoma cells. Treated HeLa cells did not undergo apoptosis but the percentage of cells in G 2 /M phase of the cell cycle was increased. We also detected that GAG-hed prevents the binding of the transcription factor AP1 to the LCR. Direct interaction of GAG-hed with the components of the AP1 complex and subsequent interference with its ability to correctly bind specific sites within the viral LCR may contribute to the inhibition of E6/E7 transcription and cell proliferation. Our data suggest that GAG-hed could have

Identifying the functional part of heparin-binding protein (HBP) as a monocyte stimulator and the novel role of monocytes as HBP producers

DEFF Research Database (Denmark)

Schou, Morten; Djurup, René; Norris, Kjeld

2011-01-01

Heparin-binding protein (HBP), an evolutionary ancient and biologically highly important molecule in inflammation, is an inactive serine protease due to mutations in the catalytic triad. The histidine (position 41) in the conserved sequence TAAHC is mutated to serine and this sequence (TAASC) pla...

A spectroscopic study of interaction of cationic dyes with heparin

Directory of Open Access Journals (Sweden)

R. Nandini

2010-01-01

Full Text Available The interaction of two cationic dyes namely, acridine orange and pinacyanol chloride with an anionic polyelectrolyte, heparin, has been investigated by spectrophotometric method.The polymer induced metachromasy in the dyes resulting in the shift of the absorption maxima of the dyes towards shorter wavelengths. The stability of the complexes formed between acridine orange and heparin was found to be lesser than that formed between pinacyanol chloride and heparin. This fact was further confirmed by reversal studies using alcohols, urea and surfactants. The interaction of acridine orange with heparin has also been investigated fluorimetrically.The interaction parameters revealed that binding between acridine orange and heparin arises due to electrostatic interaction while that between pinacyanol chloride and heparin is found to involve both electrostatic and hydrophobic forces. The effect of the structure of the dye in inducing metachromasy has also been discussed.

Pharmacological inhibition of heparin-binding EGF-like growth factor promotes peritoneal angiogenesis in a peritoneal dialysis rat model.

Science.gov (United States)

Li, Zhenyuan; Yan, Hao; Yuan, Jiangzi; Cao, Liou; Lin, Aiwu; Dai, Huili; Ni, Zhaohui; Qian, Jiaqi; Fang, Wei

2018-04-01

Molecular mechanisms of peritoneal dialysis (PD) ultrafiltration failure, peritoneal neo-angiogenesis, and fibrosis remain to be determined. We aimed to determine the role of heparin-binding EGF-like growth factor (HB-EGF) inhibition on angiogenesis of peritoneal membrane in a PD rat model. 32 male Wistar rats were assigned into (1) control group; (2) uremic non-PD group: subtotal nephrectomy-induced uremic rats without PD; (3) uremic rats subjected to PD: uremic rats that were dialyzed with Dianeal ® for 4 weeks; (4) CRM 197 group: dialyzed uremic rats were supplemented with CRM197, a specific HB-EGF inhibitor. Peritoneal transport function was examined by peritoneal equilibration test. Expression of HB-EGF and EGFR in peritoneal samples were examined by real-time PCR, immunohistochemical staining, and western blot. Progressive angiogenesis and fibrosis were observed in uremic PD rats, and there were associated with decreased net ultrafiltration (nUF), increased permeability of peritoneal membrane, and reduced expression of HB-EGF and EGFR protein and mRNA in uremic PD rats compared to uremic non-PD or control groups (both p CRM197 significantly induced peritoneal membrane permeability, decreased nUF, increased higher vessel density, and reduced pericyte count compared to that of uremic PD rats. The levels of HB-EGF and EGFR expression negatively correlated with vessel density in peritoneal membrane (both p < 0.001). PD therapy was associated with peritoneal angiogenesis, functional deterioration, and downregulation of HB-EGF/EGFR. Pharmacological inhibition of HB-EGF promoted PD-induced peritoneal angiogenesis and fibrosis and ultrafiltration decline, suggesting that HB-EGF downregulation contributes to peritoneal functional deterioration in the uremic PD rat model.

The rice XA21 ectodomain fused to the Arabidopsis EFR cytoplasmic domain confers resistance to Xanthomonas oryzae pv. oryzae.

Science.gov (United States)

Thomas, Nicholas C; Oksenberg, Nir; Liu, Furong; Caddell, Daniel; Nalyvayko, Alina; Nguyen, Yen; Schwessinger, Benjamin; Ronald, Pamela C

2018-01-01

Rice ( Oryza sativa ) plants expressing the XA21 cell-surface receptor kinase are resistant to Xanthomonas oryzae pv. oryzae (Xoo) infection. We previously demonstrated that expressing a chimeric protein containing the ELONGATION FACTOR Tu RECEPTOR (EFR) ectodomain and the XA21 endodomain (EFR:XA21) in rice does not confer robust resistance to Xoo . To test if the XA21 ectodomain is required for Xoo resistance, we produced transgenic rice lines expressing a chimeric protein consisting of the XA21 ectodomain and EFR endodomain (XA21:EFR) and inoculated these lines with Xoo . We also tested if the XA21:EFR rice plants respond to a synthetic sulfated 21 amino acid derivative (RaxX21-sY) of the activator of XA21-mediated immunity, RaxX. We found that five independently transformed XA21:EFR rice lines displayed resistance to Xoo as measured by lesion length analysis, and showed that five lines share characteristic markers of the XA21 defense response (generation of reactive oxygen species and defense response gene expression) after treatment with RaxX21-sY. Our results indicate that expression of the XA21:EFR chimeric receptor in rice confers resistance to Xoo . These results suggest that the endodomain of the EFR and XA21 immune receptors are interchangeable and the XA21 ectodomain is the key determinant conferring robust resistance to Xoo .

Cerebral Venous Sinus Thrombosis in a Patient with Undiagnosed Factor VII Deficiency.

Science.gov (United States)

Qadir, Hira; Rashid, Anila; Adil, Salman Naseem

2017-09-01

Factor VII (FVII) deficiency is one of the rare inherited bleeding disorders. Thrombosis has been occasionally described in inherited FVII deficiency. Here, we report a young female with undiagnosed FVII deficiency who presented with cerebral venous sinus thrombosis (CVST). Oral contraceptive pill was found to be prothrombotic risk factor. The CVSToccurred in spite of the congenital FVII deficiency indicating that no definitive antithrombotic protection is assured by this defect. Low molecular weight heparin and anti-Xa assay were found to be safe choice of anticoagulation and monitoring, respectively, in this patient.

Voltammetric determination of heparin based on its interaction with malachite green

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Xueliang Niu

2008-08-01

Full Text Available In this paper malachite green (MG was used as a bioprobe to determine heparin concentration by linear sweep voltammetry on the dropping mercury working electrode (DME. In Britton-Robinson (B-R buffer solution of pH 1.5, MG had a well-defined second order derivative linear sweep voltammetric reductive peak at –0.618 V (vs. SCE. After the addition of heparin into the MG solution, the reductive peak current decreased apparently without the movement of peak potential. Based on the difference of the peak current, a new voltammetric method for the determination of heparin was established. The conditions for the binding reaction and the electrochemical detection were optimized. Under the selected experimental conditions the difference of peak current was directly proportional to the concentration of heparin in the range from 0.3 to 10.0 mg/L with the linear regression equation as ∆ip″ (nA = 360.19 C (mg/L + 178.88 (n = 15, γ = 0.998 and the detection limit as 0.28 mg/L (3σ. The effects of coexisting substances such as metal ions, amino acids on the determination of heparin were investigated and the results showed that this method had good selectivity. This method was further applied to determine the heparin content in heparin sodium injection samples with satisfactory results and good recovery. The stoichiometry of the biocomplex was calculated by the electrochemical method and the binding mechanism was further discussed.

Intracellular protein delivery activity of peptides derived from insulin-like growth factor binding proteins 3 and 5

International Nuclear Information System (INIS)

Goda, Natsuko; Tenno, Takeshi; Inomata, Kosuke; Shirakawa, Masahiro; Tanaka, Toshiki; Hiroaki, Hidekazu

2008-01-01

Insulin-like growth factor binding proteins (IGFBPs) have various IGF-independent cellular activities, including receptor-independent cellular uptake followed by transcriptional regulation, although mechanisms of cellular entry remain unclear. Herein, we focused on their receptor-independent cellular entry mechanism in terms of protein transduction domain (PTD) activity, which is an emerging technique useful for clinical applications. The peptides of 18 amino acid residues derived from IGFBP-3 and IGFBP-5, which involve heparin-binding regions, mediated cellular delivery of an exogenous protein into NIH3T3 and HeLa cells. Relative protein delivery activities of IGFBP-3/5-derived peptides were approximately 20-150% compared to that of the HIV-Tat peptide, a potent PTD. Heparin inhibited the uptake of the fusion proteins with IGFBP-3 and IGFBP-5, indicating that the delivery pathway is heparin-dependent endocytosis, similar to that of HIV-Tat. The delivery of GST fused to HIV-Tat was competed by either IGFBP-3 or IGFBP-5-derived synthetic peptides. Therefore, the entry pathways of the three PTDs are shared. Our data has shown a new approach for designing protein delivery systems using IGFBP-3/5 derived peptides based on the molecular mechanisms of IGF-independent activities of IGFBPs

Heparin-binding peptide as a novel affinity tag for purification of recombinant proteins.

Science.gov (United States)

Morris, Jacqueline; Jayanthi, Srinivas; Langston, Rebekah; Daily, Anna; Kight, Alicia; McNabb, David S; Henry, Ralph; Kumar, Thallapuranam Krishnaswamy Suresh

2016-10-01

Purification of recombinant proteins constitutes a significant part of the downstream processing in biopharmaceutical industries. Major costs involved in the production of bio-therapeutics mainly depend on the number of purification steps used during the downstream process. Affinity chromatography is a widely used method for the purification of recombinant proteins expressed in different expression host platforms. Recombinant protein purification is achieved by fusing appropriate affinity tags to either N- or C- terminus of the target recombinant proteins. Currently available protein/peptide affinity tags have proved quite useful in the purification of recombinant proteins. However, these affinity tags suffer from specific limitations in their use under different conditions of purification. In this study, we have designed a novel 34-amino acid heparin-binding affinity tag (HB-tag) for the purification of recombinant proteins expressed in Escherichia coli (E. coli) cells. HB-tag fused recombinant proteins were overexpressed in E. coli in high yields. A one-step heparin-Sepharose-based affinity chromatography protocol was developed to purify HB-fused recombinant proteins to homogeneity using a simple sodium chloride step gradient elution. The HB-tag has also been shown to facilitate the purification of target recombinant proteins from their 8 M urea denatured state(s). The HB-tag has been demonstrated to be successfully released from the fusion protein by an appropriate protease treatment to obtain the recombinant target protein(s) in high yields. Results of the two-dimensional NMR spectroscopy experiments indicate that the purified recombinant target protein(s) exist in the native conformation. Polyclonal antibodies raised against the HB-peptide sequence, exhibited high binding specificity and sensitivity to the HB-fused recombinant proteins (∼10 ng) in different crude cell extracts obtained from diverse expression hosts. In our opinion, the HB-tag provides a

Activated factor X signaling via protease-activated receptor 2 suppresses pro-inflammatory cytokine production from LPS-stimulated myeloid cells.

LENUS (Irish Health Repository)

Gleeson, Eimear M

2013-07-19

Vitamin K-dependent proteases generated in response to vascular injury and infection enable fibrin clot formation, but also trigger distinct immuno-regulatory signaling pathways on myeloid cells. Factor Xa, a protease crucial for blood coagulation, also induces protease-activated receptor-dependent cell signaling. Factor Xa can bind both monocytes and macrophages, but whether factor Xa-dependent signaling stimulates or suppresses myeloid cell cytokine production in response to Toll-like receptor activation is not known. In this study, exposure to factor Xa significantly impaired pro-inflammatory cytokine production from lipopolysaccharide-treated peripheral blood mononuclear cells, THP-1 monocytic cells and murine macrophages. Furthermore, factor Xa inhibited nuclear factor-kappa B activation in THP-1 reporter cells, requiring phosphatidylinositide 3-kinase activity for its anti-inflammatory effect. Active-site blockade, γ-carboxyglutamic acid domain truncation and a peptide mimic of the factor Xa inter-epidermal growth factor-like region prevented factor Xa inhibition of lipopolysaccharide-induced tumour necrosis factor-α release. In addition, factor Xa anti-inflammatory activity was markedly attenuated by the presence of an antagonist of protease-activated receptor 2, but not protease-activated receptor 1. The key role of protease-activated receptor 2 in eliciting factor Xa-dependent anti-inflammatory signaling on macrophages was further underscored by the inability of factor Xa to mediate inhibition of tumour necrosis factor-α and interleukin-6 release from murine bone marrow-derived protease-activated receptor 2-deficient macrophages. We also show for the first time that, in addition to protease-activated receptor 2, factor Xa requires a receptor-associated protein-sensitive low-density lipoprotein receptor to inhibit lipopolysaccharide-induced cytokine production. Collectively, this study supports a novel function for factor Xa as an endogenous, receptor

Heparin Interaction with the Primed Polymorphonuclear Leukocyte CD11b Induces Apoptosis and Prevents Cell Activation

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Meital Cohen-Mazor

2015-01-01

Full Text Available Heparin is known to have anti-inflammatory effects, yet the mechanisms are not completely understood. In this study, we tested the hypothesis that heparin has a direct effect on activated polymorphonuclear leukocytes (PMNLs, changing their activation state, and can explain its anti-inflammatory effect. To test our hypothesis, we designed both in vitro and ex vivo studies to elucidate the mechanism by which heparin modulates PMNL functions and therefore the inflammatory response. We specifically tested the hypothesis that priming of PMNLs renders them more susceptible to heparin. Amplified levels of CD11b and increased rate of superoxide release manifested PMNL priming. Increase in cell priming resulted in a dose-dependent increase in heparin binding to PMNLs followed by augmented apoptosis. Blocking antibodies to CD11b inhibited heparin binding and abolished the apoptotic response. Moreover, heparin caused a significant dose-dependent decrease in the rate of superoxide release from PMNLs, which was blunted by blocking antibodies to CD11b. Altogether, this study shows that the interaction of heparin with the PMNL CD11b results in cell apoptosis and explains heparin’s anti-inflammatory effects.

RNA binding specificity of Ebola virus transcription factor VP30.

Science.gov (United States)

Schlereth, Julia; Grünweller, Arnold; Biedenkopf, Nadine; Becker, Stephan; Hartmann, Roland K

2016-09-01

The transcription factor VP30 of the non-segmented RNA negative strand Ebola virus balances viral transcription and replication. Here, we comprehensively studied RNA binding by VP30. Using a novel VP30:RNA electrophoretic mobility shift assay, we tested truncated variants of 2 potential natural RNA substrates of VP30 - the genomic Ebola viral 3'-leader region and its complementary antigenomic counterpart (each ∼155 nt in length) - and a series of other non-viral RNAs. Based on oligonucleotide interference, the major VP30 binding region on the genomic 3'-leader substrate was assigned to the internal expanded single-stranded region (∼ nt 125-80). Best binding to VP30 was obtained with ssRNAs of optimally ∼ 40 nt and mixed base composition; underrepresentation of purines or pyrimidines was tolerated, but homopolymeric sequences impaired binding. A stem-loop structure, particularly at the 3'-end or positioned internally, supports stable binding to VP30. In contrast, dsRNA or RNAs exposing large internal loops flanked by entirely helical arms on both sides are not bound. Introduction of a 5´-Cap(0) structure impaired VP30 binding. Also, ssDNAs bind substantially weaker than isosequential ssRNAs and heparin competes with RNA for binding to VP30, indicating that ribose 2'-hydroxyls and electrostatic contacts of the phosphate groups contribute to the formation of VP30:RNA complexes. Our results indicate a rather relaxed RNA binding specificity of filoviral VP30, which largely differs from that of the functionally related transcription factor of the Paramyxoviridae which binds to ssRNAs as short as 13 nt with a preference for oligo(A) sequences.

Molecular screening of rice ( Oryza sativa L.) germplasm for Xa4 ...

African Journals Online (AJOL)

DNA fingerprinting results indicated the presence of Xa4 gene in 41 entries, while 14 lines were positive for xa5 gene. Only one local line was carrying Xa21 gene along with Xa4. Thus, the present study will not only be helpful for rice breeders to develop new rice varieties carrying disease resistant genes, but will also ...

Bioinspired Heparin Nanosponge Prepared by Photo-crosslinking for Controlled Release of Growth Factors

DEFF Research Database (Denmark)

Choi, Won Il; Sahu, Abhishek; Vilos, Cristian

2017-01-01

to overcome these limitations. Herein, we have developed a thermosensitive heparin nanosponge (Hep-NS) by a one step photopolymerization reaction between diacrylated pluronic and thiolated heparin molecules. The amount of heparin in Hep-NS was precisely controlled by varying the heparin amount in the reaction...

The ligand-binding profile of HARE: hyaluronan and chondroitin sulfates A, C, and D bind to overlapping sites distinct from the sites for heparin, acetylated low-density lipoprotein, dermatan sulfate, and CS-E.

Science.gov (United States)

Harris, Edward N; Weigel, Paul H

2008-08-01

The hyaluronic acid receptor for endocytosis (HARE)/ Stabilin-2 is the primary systemic scavenger receptor for hyaluronan (HA), the chondroitin sulfates (CS), dermatan sulfate (DS), and nonglycosaminoglycan (GAG) ligands such as acetylated low-density lipoprotein (AcLDL), pro-collagen propeptides, and advanced glycation end products. We recently discovered that HARE is also a systemic scavenger receptor for heparin (Hep) (Harris EN, Weigel JA, Weigel PH. 2008. The human hyaluronan receptor for endocytosis [HARE/Stabilin-2] is a systemic clearance receptor for heparin. J Biol Chem. 283:17341-17350). Our goal was to map the binding sites of eight different ligands within HARE. We used biotinylated GAGs and radio-iodinated streptavidin or AcLDL to assess the binding activities of ligands directly or indirectly (by competition with unlabeled ligands) in endocytosis assays using stable cell lines expressing the 315 or 190 kDa HA receptor for endocytosis (315- or 190-HARE) isoforms, and ELISA-like assays, with purified recombinant soluble 190-HARE ecto-domain. For example, Hep binding to HARE was competed by DS, CS-E, AcLDL, and dextran sulfate, but not by other CS types, HA, dextran, or heparosan. (125)I-AcLDL binding to HARE was partially competed by Hep and dextran sulfate, but not competed by HA. Two ligands, DS and CS-E, competed with both Hep and HA to some degree. Hep and HA binding or endocytosis is mutually inclusive; binding of these two GAGs occurs with functionally separate, noncompetitive, and apparently noninteracting domains. Thus, HARE binds to HA and Hep simultaneously. Although the domain(s) responsible for Hep binding remains unknown, the Link domain was required for HARE binding to HA, CS-A, CS-C, and CS-D. These results enable us to outline, for the first time, a binding activity map for multiple ligands of HARE.

Heparin molecularly imprinted polymer thin flm on gold electrode by plasma-induced graft polymerization for label-free biosensor.

Science.gov (United States)

Orihara, Kouhei; Hikichi, Atsushi; Arita, Tomohiko; Muguruma, Hitoshi; Yoshimi, Yasuo

2018-03-20

Heparin, a highly sulfated glycosaminoglycan, is an important biomaterial having biological and therapeutic functionalities such as anticoagulation, regeneration, and protein stabilization. This study addresses a label-free quartz crystal microbalance (QCM) biosensor for heparin detection based on a macromolecularly imprinted polymer (MIP) as an artificial recognition element. We demonstrate the novel strategy for MIP in the form of thin film on a gold (Au) electrode with the plasma-induced graft polymerization (PIP) technique. The procedure of PIP is as follows: (i) Hexamethyldisiloxane plasma-polymerized thin film (PPF) as a pre-coating scaffold of active species for PIP (post-polymerization) is deposited on an Au electrode. (ii) The PPF/Au electrode is soaked in an water solution containing heparin (template), (2-(methacryloxy)-ethyl)trimethylammonium chloride acrylamide (functional monomer), acrylamide, and N,N-methylenebisacrylamide (crosslinker). Double bonds of monomer and crosslinker attacked by residually active species in pre-coating PPF cause radical chain reaction. Consequently, a growing polymer network of 20 nm thickness of PIP-MIP thin film is formed and grafted on the PPF/Au surface. (iii) The PIP-MIP/PPF/Au is washed by sodium chloride solution so as to remove the template. Non-imprinted polymer (NIP) is carried out like the same procedure without a template. The AFM, XPS, and QCM measurements show that the PIP process facilitates macromolecularly surface imprinting of template heparin where the template is easily removed and is rapidly rebound to PIP-MIP without a diffusional barrier. The heparin-PIP-MIP specifically binds to heparin compared with heparin analog chondroitin sulfate C (selective factor: 4.0) and a detectable range of heparin in the presence of CS (0.1 wt%) was 0.001-0.1 wt%. The PIP-NIP does not show selectivity between them. The evaluated binding kinetics are association (k a  = 350 ± 100 M -1  s -1

Biomedical Application of Low Molecular Weight Heparin/Protamine Nano/Micro Particles as Cell- and Growth Factor-Carriers and Coating Matrix

Directory of Open Access Journals (Sweden)

Masayuki Ishihara

2015-05-01

Full Text Available Low molecular weight heparin (LMWH/protamine (P nano/micro particles (N/MPs (LMWH/P N/MPs were applied as carriers for heparin-binding growth factors (GFs and for adhesive cells including adipose-derived stromal cells (ADSCs and bone marrow-derived mesenchymal stem cells (BMSCs. A mixture of LMWH and P yields a dispersion of N/MPs (100 nm–3 μm in diameter. LMWH/P N/MPs can be immobilized onto cell surfaces or extracellular matrix, control the release, activate GFs and protect various GFs. Furthermore, LMWH/P N/MPs can also bind to adhesive cell surfaces, inducing cells and LMWH/P N/MPs-aggregate formation. Those aggregates substantially promoted cellular viability, and induced vascularization and fibrous tissue formation in vivo. The LMWH/P N/MPs, in combination with ADSCs or BMSCs, are effective cell-carriers and are potential promising novel therapeutic agents for inducing vascularization and fibrous tissue formation in ischemic disease by transplantation of the ADSCs and LMWH/P N/MPs-aggregates. LMWH/P N/MPs can also bind to tissue culture plates and adsorb exogenous GFs or GFs from those cells. The LMWH/P N/MPs-coated matrix in the presence of GFs may provide novel biomaterials that can control cellular activity such as growth and differentiation. Furthermore, three-dimensional (3D cultures of cells including ADSCs and BMSCs using plasma-medium gel with LMWH/P N/MPs exhibited efficient cell proliferation. Thus, LMWH/P N/MPs are an adequate carrier both for GFs and for stromal cells such as ADSCs and BMSCs, and are a functional coating matrix for their cultures.

Interaction of blood coagulation factor Va with phospholipid vesicles examined by using lipophilic photoreagents

International Nuclear Information System (INIS)

Krieg, U.C.; Isaacs, B.S.; Yemul, S.S.; Esmon, C.T.; Bayley, H.; Johnson, A.E.

1987-01-01

Two different lipophilic photoreagents, [ 3 H]adamantane diazirine and 3-(trifluoromethyl)-3-(m-[ 125 I]iodophenyl)diazirine (TID), have been utilized to examine the interactions of blood coagulation factor Va with calcium, prothrombin, factor Xa, and, in particular, phospholipid vesicles. With each of these structurally dissimilar reagents, the extent of photolabeling of factor Va was greater when the protein was bound to a membrane surface than when it was free in solution. Specifically, the covalent photoreaction with Vl, the smaller subunit of factor Va, was 2-fold higher in the presence of phosphatidylcholine/phosphatidylserine (PC/PS, 3:1) vesicles, to which factor Va binds, than in the presence of 100% PC vesicles, to which the protein does not bind. However, the magnitude of the PC/PS-dependent photolabeling was much less than has been observed previously with integral membrane proteins. It therefore appears that the binding of factor Va to the membrane surface exposes Vl to the lipid core of the bilayer, but that only a small portion of the Vl polypeptide is exposed to, or embedded in, the bilayer core. Addition of either prothrombin or active-site-blocked factor Xa to PC/PS-bound factor Va had little effect on the photolabeling of Vl with TID, but reduced substantially the covalent labeling of Vh, the larger subunit of factor Va. This indicates that prothrombin and factor Xa each cover nonpolar surfaces on Vh when the macromolecules associate on the PC/PS surface. It therefore seems likely that the formation of the prothrombinase complex involves a direct interaction between Vh and factor Xa and between Vh and prothrombin.(ABSTRACT TRUNCATED AT 250 WORDS)

Maximize x(a - x)

Science.gov (United States)

Lange, L. H.

1974-01-01

Five different methods for determining the maximizing condition for x(a - x) are presented. Included is the ancient Greek version and a method attributed to Fermat. None of the proofs use calculus. (LS)

Heparin kinetics

International Nuclear Information System (INIS)

Swart, C.A.M. de.

1983-01-01

The author has studied the kinetics of heparin and heparin fractions after intravenous administration in humans and in this thesis the results of this study are reported. Basic knowledge about the physico-chemical properties of heparin and its interactions with proteins resulting in anticoagulant and lipolytic effects are discussed in a review (chapter II), which also comprises some clinical aspects of heparin therapy. In chapter III the kinetics of the anticoagulant effect are described after intravenous administration of five commercial heparin preparations. A mathematical model is presented that fits best to these kinetics. The kinetics of the anticoagulant and lipolytic effects after intravenous injection of various 35 S-radiolabelled heparin fractions and their relationship with the disappearance of the radiolabel are described in chapter IV. Chapter V gives a description of the kinetics of two radiolabels after injection of in vitro formed complexes consisting of purified, 125 I-radiolabelled antithrombin III and various 35 S-radiolabelled heparin fractions. (Auth.)

Sensitive detection of oversulfated chondroitin sulfate in heparin sodium or crude heparin with a colorimetric microplate based assay.

Science.gov (United States)

Sommers, Cynthia D; Mans, Daniel J; Mecker, Laura C; Keire, David A

2011-05-01

In this work we describe a 96-well microplate assay for oversulfated chondroitin sulfate A (OSCS) in heparin, based on a water-soluble cationic polythiophene polymer (3-(2-(N-(N'-methylimidazole))ethoxy)-4-methylthiophene (LPTP)) and heparinase digestion of heparin. The assay takes advantage of several unique properties of heparin, OSCS, and LPTP, including OSCS inhibition of heparinase I and II activity, the molecular weight dependence of heparin-LPTP spectral shifts, and the distinct association of heparin fragments and OSCS to LPTP. These factors combine to enable detection of the presence of 0.003% w/w spiked OSCS in 10 μg of heparin sodium active pharmaceutical ingredient (API) using a plate reader and with visual detection to 0.1% levels. The same detection limit for OSCS was observed in the presence of 10% levels of dermatan sulfate (DS) or chondroitin sulfate A (CSA) impurities. In addition, we surveyed a selection of crude heparin samples received by the agency in 2008 and 2009 to determine average and extreme DS, CSA, and galactosamine weight percent levels. In the presence of these impurities and the variable heparin content in the crude heparin samples, spiked OSCS was reliably detected to the 0.1% w/w level using a plate reader. Finally, authentically OSCS contaminated heparin sodium API and crude samples were distinguished visually by color from control samples using the LPTP/heparinase test.

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.

N-acetyl-heparin attenuates acute lung injury caused by acid aspiration mainly by antagonizing histones in mice.

Science.gov (United States)

Zhang, Yanlin; Zhao, Zanmei; Guan, Li; Mao, Lijun; Li, Shuqiang; Guan, Xiaoxu; Chen, Ming; Guo, Lixia; Ding, Lihua; Cong, Cuicui; Wen, Tao; Zhao, Jinyuan

2014-01-01

Acute lung injury (ALI) is the leading cause of death in intensive care units. Extracellular histones have recently been recognized to be pivotal inflammatory mediators. Heparin and its derivatives can bind histones through electrostatic interaction. The purpose of this study was to investigate 1) the role of extracellular histones in the pathogenesis of ALI caused by acid aspiration and 2) whether N-acetyl-heparin (NAH) provides more protection than heparin against histones at the high dose. ALI was induced in mice via intratracheal instillation of hydrochloric acid (HCl). Lethality rate, blood gas, myeloperoxidase (MPO) activity, lung edema and pathological changes were used to evaluate the degree of ALI. Heparin/NAH was administered intraperitoneally, twice a day, for 3 days or until death. Acid aspiration caused an obvious increase in extracellular histones. A significant correlation existed between the concentration of HCl aspirated and the circulating histones. Heparin/NAH (10 mg/kg) improved the lethality rate, blood gas, MPO activity, lung edema and pathological score. At a dose of 20 mg/kg, NAH still provided protection, however heparin tended to aggravate the injury due to hemorrhagic complications. The specific interaction between heparin and histones was verified by the binding assay. In summary, high levels of extracellular histones can be pathogenic in ALI caused by acid aspiration. By neutralizing extracellular histones, heparin/NAH can offer similar protection at the moderate doses. At the high dose, NAH provides better protection than heparin.

Unfractionated heparin versus low molecular weight heparins for avoiding heparin-induced thrombocytopenia in postoperative patients.

Science.gov (United States)

Junqueira, Daniela R; Zorzela, Liliane M; Perini, Edson

2017-04-21

Heparin-induced thrombocytopenia (HIT) is an adverse drug reaction presenting as a prothrombotic disorder related to antibody-mediated platelet activation. It is a paradoxical immune reaction resulting in thrombin generation in vivo, which leads to a hypercoagulable state and the potential to initiate venous or arterial thrombosis. A number of factors are thought to influence the incidence of HIT including the type and preparation of heparin (unfractionated heparin (UFH) or low molecular weight heparin (LMWH)) and the heparin-exposed patient population, with the postoperative patient population at higher risk.Although LMWH has largely replaced UFH as a front-line therapy, there is evidence supporting a lack of superiority of LMWH compared with UFH regarding prevention of deep vein thrombosis and pulmonary embolism following surgery, and similar frequencies of bleeding have been described with LMWH and UFH. The decision as to which of these two preparations of heparin to use may thus be influenced by harmful effects such as HIT. We therefore sought to determine the relative impact of UFH and LMWH on HIT in postoperative patients receiving thromboembolism prophylaxis. This is an update of a review first published in 2012. The objective of this review was to compare the incidence of heparin-induced thrombocytopenia (HIT) and HIT complicated by venous thromboembolism in postoperative patients exposed to unfractionated heparin (UFH) versus low molecular weight heparin (LMWH). For this update, the Cochrane Vascular Information Specialist searched the Specialised Register (May 2016), CENTRAL (2016, Issue 4) and trials registries. The authors searched Lilacs (June 2016) and additional trials were sought from reference lists of relevant publications. We included randomised controlled trials (RCTs) in which participants were postoperative patients allocated to receive prophylaxis with UFH or LMWH, in a blinded or unblinded fashion. Studies were excluded if they did not use

Review of fondaparinux sodium injection for the prevention of venous thromboembolism in patients undergoing surgery

Directory of Open Access Journals (Sweden)

David Bergqvist

2006-12-01

Full Text Available David BergqvistDepartment of Surgery, University Hospital, Uppsala, SwedenAbstract: The antithrombin binding sequence of heparin, a pentasaccharide, has been synthesized as fondaparinux, an indirect, selective, and reversible factor Xa inhibitor. It can be administered subcutaneously, is well absorbed, and has a half-life of c. 17 hours permitting once-daily injection. It has been evaluated in an extensive study program in major orthopedic surgery, including hip fracture, and in major abdominal surgery with a large proportion of surgery for cancer. The effect is at least as effective as for low-molecular-weight heparins and it has also been shown effective for extended prophylaxis in hip fracture patients. Several thousands of patients have been studied and the substance is safe, although a slightly higher frequency of bleedings is found than in patients on low-molecular-weight heparins. There is no specific antidote but if necessary, recombinant activated factor VII can be used. Other side-effects are rare. Fondaparinux is cost saving and sometimes cost neutral when compared with enoxaparin. Keywords: fondaparinux, venous thrombosis, venous thromboembolism, surgery, orthopedic surgery, major abdominal surgery, bleeding complications

Immunomodulating effects of heparin on human B cell proliferation

International Nuclear Information System (INIS)

Wasik, Maria; Stepien-Sopniewska, Barbara; Gorski, Andrzej

1993-01-01

Recent data indicate that heparin may act as an immunomodulator. In this paper we have analyzed the effect of this agent on human B cell proliferation ''in vitro'' induced by ''S. aureus'' Cowan. The action of heparin is complex, but there was a trend for inhibition of B cell responses obtained from defibrinated but not heparinized blood samples. This suggest that heparin interacts with platelet products (growth factors, cytokines) and the results of such interactions determine the final effect. (author). 6 refs, 4 figs

Parent heparin and daughter LMW heparin correlation analysis using LC-MS and NMR

Energy Technology Data Exchange (ETDEWEB)

Liu, Xinyue, E-mail: liux22@rpi.edu [National Glycoengineering Research Center, Shandong Provincial Key Laboratory of Carbohydrate Chemistry and Glycobiology, State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong, 250100 (China); Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180 (United States); St Ange, Kalib, E-mail: stangk2@rpi.edu [Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180 (United States); Wang, Xiaohua, E-mail: wangx35@rpi.edu [Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180 (United States); School of Computer and Information, Hefei University of Technology, Hefei (China); Lin, Lei, E-mail: Linl5@rpi.edu [Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180 (United States); Zhang, Fuming, E-mail: zhangf2@rpi.edu [Department of Chemistry and Chemical Biology, Department of Chemical and Biological Engineering, Department of Biology, Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, 12180 (United States); and others

2017-04-08

Heparin is a structurally complex, polysaccharide anticoagulant derived from livestock, primarily porcine intestinal tissues. Low molecular weight (LMW) heparins are derived through the controlled partial depolymerization of heparin. Increased manufacturing and regulatory concerns have provided the motivation for the development of more sophisticated analytical methods for determining both their structure and pedigree. A strategy, for the comprehensive comparison of parent heparins and their LMW heparin daughters, is described that relies on the analysis of monosaccharide composition, disaccharide composition, and oligosaccharide composition. Liquid chromatography-mass spectrometry is rapid, robust, and amenable to automated processing and interpretation of both top-down and bottom-up analyses. Nuclear magnetic resonance spectroscopy provides complementary top-down information on the chirality of the uronic acid residues and glucosamine substitution. Principal component analysis (PCA) was applied to the normalized abundance of oligosaccharides, calculated in the bottom-up analysis, to show parent and daughter correlation in oligosaccharide composition. Using these approaches, six pairs of parent heparins and their daughter generic enoxaparins from two different manufacturers were comprehensively analyzed. Enoxaparin is the most widely used LMW heparin and is prepared through controlled chemical β-eliminative cleavage of porcine intestinal heparin. Lovenox{sup ®}, the innovator version of enoxaparin marketed in the US, was analyzed as a reference for the daughter LMW heparins. The results, show similarities between LMW heparins from two different manufacturers with Lovenox{sup ®}, excellent lot-to-lot consistency of products from each manufacturer, and detects a correlation between each parent heparin and daughter LMW heparin. - Highlights: • Low molecular weight heparins prepared from different heparin parents were analyzed. • An integrated analytical

Parent heparin and daughter LMW heparin correlation analysis using LC-MS and NMR

International Nuclear Information System (INIS)

Liu, Xinyue; St Ange, Kalib; Wang, Xiaohua; Lin, Lei; Zhang, Fuming

2017-01-01

Heparin is a structurally complex, polysaccharide anticoagulant derived from livestock, primarily porcine intestinal tissues. Low molecular weight (LMW) heparins are derived through the controlled partial depolymerization of heparin. Increased manufacturing and regulatory concerns have provided the motivation for the development of more sophisticated analytical methods for determining both their structure and pedigree. A strategy, for the comprehensive comparison of parent heparins and their LMW heparin daughters, is described that relies on the analysis of monosaccharide composition, disaccharide composition, and oligosaccharide composition. Liquid chromatography-mass spectrometry is rapid, robust, and amenable to automated processing and interpretation of both top-down and bottom-up analyses. Nuclear magnetic resonance spectroscopy provides complementary top-down information on the chirality of the uronic acid residues and glucosamine substitution. Principal component analysis (PCA) was applied to the normalized abundance of oligosaccharides, calculated in the bottom-up analysis, to show parent and daughter correlation in oligosaccharide composition. Using these approaches, six pairs of parent heparins and their daughter generic enoxaparins from two different manufacturers were comprehensively analyzed. Enoxaparin is the most widely used LMW heparin and is prepared through controlled chemical β-eliminative cleavage of porcine intestinal heparin. Lovenox"®, the innovator version of enoxaparin marketed in the US, was analyzed as a reference for the daughter LMW heparins. The results, show similarities between LMW heparins from two different manufacturers with Lovenox"®, excellent lot-to-lot consistency of products from each manufacturer, and detects a correlation between each parent heparin and daughter LMW heparin. - Highlights: • Low molecular weight heparins prepared from different heparin parents were analyzed. • An integrated analytical approach relied

Apixaban or enoxaparin for thromboprophylaxis after knee replacement

DEFF Research Database (Denmark)

Lassen, Michael Rud; Raskob, Gary E; Gallus, Alexander

2009-01-01

BACKGROUND: The optimal strategy for thromboprophylaxis after major joint replacement has not been established. Low-molecular-weight heparins such as enoxaparin predominantly target factor Xa but to some extent also inhibit thrombin. Apixaban, a specific factor Xa inhibitor, may provide effective...... (P=0.03). CONCLUSIONS: As compared with enoxaparin for efficacy of thromboprophylaxis after knee replacement, apixaban did not meet the prespecified statistical criteria for noninferiority, but its use was associated with lower rates of clinically relevant bleeding and it had a similar adverse...

RUBY-1: a randomized, double-blind, placebo-controlled trial of the safety and tolerability of the novel oral factor Xa inhibitor darexaban (YM150) following acute coronary syndrome

DEFF Research Database (Denmark)

Steg, Ph Gabriel; Mehta, Shamir R; Jukema, J Wouter

2011-01-01

To establish the safety, tolerability and most promising regimen of darexaban (YM150), a novel, oral, direct factor Xa inhibitor, for prevention of ischaemic events in acute coronary syndrome (ACS)....

Angiogenic Factor Profiles in Pregnant Women With a History of Early-Onset Severe Preeclampsia Receiving Low-Molecular-Weight Heparin Prophylaxis.

Science.gov (United States)

Lecarpentier, Edouard; Gris, Jean Christophe; Cochery-Nouvellon, Eva; Mercier, Erick; Touboul, Cyril; Thadhani, Ravi; Karumanchi, S Ananth; Haddad, Bassam

2018-01-01

To evaluate whether daily low-molecular-weight (LMW) heparin prophylaxis during pregnancy alters profile of circulating angiogenic factors that have been linked with the pathogenesis of preeclampsia and fetal growth restriction. This is a planned ancillary study of the Heparin-Preeclampsia trial, a randomized trial in pregnant women with a history of severe early-onset preeclampsia (less than 34 weeks of gestation). In the parent study, all women were treated with aspirin and then randomized to receive LMW heparin or aspirin alone. In this study, we measured serum levels of circulating angiogenic factors (soluble fms-like tyrosine kinase-1, placental growth factor, and soluble endoglin by immunoassay) at the following gestational windows: 10-13 6/7 weeks, 14-17 6/7 weeks, 18-21 6/7 weeks, 22-25 6/7 weeks, 26-29 6/7 weeks, 30-33 6/7 weeks, and 34-37 6/7 weeks. Samples were available from 185 patients: LMW heparin+aspirin (n=92) and aspirin alone (n=93). The two groups had comparable baseline characteristics and had similar adverse composite outcomes (35/92 [38.0%] compared with 36/93 [38.7%]; P=.92). There were no significant differences in serum levels of soluble fms-like tyrosine kinase-1, placental growth factor, and soluble endoglin in the participants who received LMW heparin and aspirin compared with those who received aspirin alone regardless of gestational age period. Finally, women who developed an adverse composite outcome at less than 34 weeks of gestation demonstrated significant alterations in serum angiogenic profile as early as 10-13 6/7 weeks that was most dramatic 6-8 weeks preceding delivery. Prophylactic LMW heparin therapy when beginning from before 14 weeks of gestation with aspirin during pregnancy is not associated with an improved angiogenic profile. This may provide a molecular explanation for the lack of clinical benefit noted in recent trials. ClinicalTrials.gov, NCT00986765.

Heparin-Binding EGF-like Growth Factor (HB-EGF) stimulates the proliferation of Müller glia-derived progenitor cells in avian and murine retinas

Science.gov (United States)

Todd, Levi; Volkov, Leo I.; Zelinka, Chris; Squires, Natalie; Fischer, Andy J.

2015-01-01

Müller glia can be stimulated to de-differentiate, proliferate and form Müller glia-derived progenitor cells (MGPCs) that regenerate retinal neurons. In the zebrafish retina, Heparin-Binding EGF-like Growth Factor (HB-EGF) may be one of the key factors that stimulate the formation of proliferating MGPCs. Currently nothing is known about the influence of HB-EGF on the proliferative potential of Müller glia in retinas of birds and rodents. In the chick retina, we found that levels of both hb-egf and egf-receptor are rapidly and transiently up-regulated following NMDA-induced damage. Although intraocular injections of HB-EGF failed to stimulate cell-signaling or proliferation of Müller glia in normal retinas, HB-EGF stimulated proliferation of MGPCs in damaged retinas. By comparison, inhibition of the EGF-receptor (EGFR) decreased the proliferation of MGPCs in damaged retinas. HB-EGF failed to act synergistically with FGF2 to stimulate the formation of MGPCs in the undamaged retina and inhibition of EGF-receptor did not suppress FGF2-mediated formation of MGPCs. In the mouse retina, HB-EGF stimulated the proliferation of Müller glia following NMDA-induced damage. Furthermore, HB-EGF stimulated not only MAPK-signaling in Müller glia/MGPCs, but also activated mTor- and Jak/Stat-signaling. We propose that levels of expression of EGFR are rate-limiting to the responses of Müller glia to HB-EGF and the expression of EGFR can be induced by retinal damage, but not by FGF2-treatment. We conclude that HB-EGF is mitogenic to Müller glia in both chick and mouse retinas, and HB-EGF is an important player in the formation of MGPCs in damaged retinas. PMID:26500021

Heparin binding chitosan derivatives for production of pro-angiogenic hydrogels for promoting tissue healing

Energy Technology Data Exchange (ETDEWEB)

Yar, Muhammad, E-mail: drmyar@ciitlahore.edu.pk [Interdisciplinary Research Center in Biomedical Materials, COMSATS Institute of Information Technology, Lahore 54000 (Pakistan); Shahzad, Sohail [Interdisciplinary Research Center in Biomedical Materials, COMSATS Institute of Information Technology, Lahore 54000 (Pakistan); Department of Chemistry, The Islamia University of Bahawalpur, Bahawalpur 63100 (Pakistan); Shahzadi, Lubna [Interdisciplinary Research Center in Biomedical Materials, COMSATS Institute of Information Technology, Lahore 54000 (Pakistan); Shahzad, Sohail Anjum [Department of Chemistry, COMSATS Institute of Information Technology, Abbottabad 22060 (Pakistan); Mahmood, Nasir [Department of Allied Health Sciences and Chemical Pathology, University of Health Sciences, Lahore (Pakistan); Department of Human Genetics and Molecular Biology, University of Health Sciences, Lahore (Pakistan); Chaudhry, Aqif Anwar [Interdisciplinary Research Center in Biomedical Materials, COMSATS Institute of Information Technology, Lahore 54000 (Pakistan); Rehman, Ihtesham ur [Interdisciplinary Research Center in Biomedical Materials, COMSATS Institute of Information Technology, Lahore 54000 (Pakistan); Materials Science and Engineering, North Campus, University of Sheffield, Broad Lane, Sheffield S3 7HQ (United Kingdom); MacNeil, Sheila, E-mail: s.macneil@sheffield.ac.uk [Materials Science and Engineering, North Campus, University of Sheffield, Broad Lane, Sheffield S3 7HQ (United Kingdom)

2017-05-01

Our aim was to develop a biocompatible hydrogel that could be soaked in heparin and placed on wound beds to improve the vasculature of poorly vascularized wound beds. In the current study, a methodology was developed for the synthesis of a new chitosan derivative (CSD-1). Hydrogels were synthesized by blending CSD-1 for either 4 or 24 h with polyvinyl alcohol (PVA). The physical/chemical interactions and the presence of specific functional groups were confirmed by Fourier transform infrared (FT-IR) spectroscopy and proton nuclear magnetic resonance ({sup 1}H NMR). The porous nature of the hydrogels was confirmed by scanning electron microscopy (SEM). Thermal gravimetric analysis (TGA) showed that these hydrogels have good thermal stability which was slightly increased as the blending time was increased. Hydrogels produced with 24 h of blending supported cell attachment more and could be loaded with heparin to induce new blood vessel formation in a chick chorionic allantoic membrane assay. - Highlights: • Chitosan based hydrogels were designed to stimulate angiogenesis. • Two new derivatives of chitosan were produced using a Mannich type reaction. • Blending a chitosan derivative with PVA gave a porous biocompatible hydrogel. • Heparin bound to the hydrogel on immersion changing its morphology. • Heparin loaded hydrogel stimulated blood vessel formation in a chick model.

N-terminal and C-terminal heparin-binding domain polypeptides derived from fibronectin reduce adhesion and invasion of liver cancer cells

International Nuclear Information System (INIS)

Tang, Nan-Hong; Chen, Yan-Lin; Wang, Xiao-Qian; Li, Xiu-Jin; Wu, Yong; Zou, Qi-Lian; Chen, Yuan-Zhong

2010-01-01

Fibronectin (FN) is known to be a large multifunction glycoprotein with binding sites for many substances, including N-terminal and C-terminal heparin-binding domains. We investigated the effects of highly purified rhFNHN29 and rhFNHC36 polypeptides originally cloned from the two heparin-binding domains on the adhesion and invasion of highly metastatic human hepatocellular carcinoma cells (MHCC97H) and analyzed the underlying mechanism involved. The MHCC97H cells that adhered to FN in the presence of various concentrations of rhFNHN29 and rhFNHC36 polypeptides were stained with crystal violet and measured, and the effects of rhFNHN29 and rhFNHC36 on the invasion of the MHCC97H cells were then detected using the Matrigel invasion assay as well as a lung-metastasis mouse model. The expression level of integrins and focal adhesion kinase (FAK) phosphotyrosyl protein was examined by Western blot, and the activity of matrix metalloproteinases (MMPs) and activator protein 1 (AP-1) was analyzed by gelatin zymography and the electrophoretic mobility band-shift assay (EMSA), respectively. Both of the polypeptides rhFNHN29 and rhFNHC36 inhibited adhesion and invasion of MHCC97H cells; however, rhFNHC36 exhibited inhibition at a lower dose than rhFNHN29. These inhibitory effects were mediated by integrin αvβ3 and reversed by a protein tyrosine phosphatase inhibitor. Polypeptides rhFNHN29 and rhFNHC36 abrogated the tyrosine phosphorylation of focal adhesion kinase (p-FAK) and activation of activator protein 1 (AP-1), resulting in the decrease of integrin αv, β3 and β1 expression as well as the reduction of MMP-9 activity. Polypeptides rhFNHN29 and rhFNHC36 could potentially be applicable to human liver cancer as anti-adhesive and anti-invasive agents

Cellular adhesion responses to the heparin-binding (HepII) domain of fibronectin require heparan sulfate with specific properties

DEFF Research Database (Denmark)

Mahalingam, Yashithra; Gallagher, John T; Couchman, John R

2006-01-01

of fibronectin (HepII domain) through its HS chains. The fine structure of HS is critical to growth factor responses, and whether this extends to matrix ligands is unknown but is suggested from in vitro experiments. Cell attachment to HepII showed that heparin oligosaccharides of >or=14 sugar residues were...

Cutaneous reactions to heparin therapy: when are they caused by heparin allergy?

Directory of Open Access Journals (Sweden)

Giuliana Zisa

2013-03-01

Full Text Available Introduction: Little is known about the incidence and causes of heparin-induced skin lesions. The most commonly reported causes are delayed-type hypersensitivity reactions. We describe 3 patients who were referred to our staff between March and October 2009 for suspected heparin allergies. All were scheduled to undergo major surgery (cardiovascular or orthopedic. Materials and methods: All 3 patients reported the development of itchy, erythematous rashes a few days after the subcutaneous administration of heparin (nadroparin calcium in cases 1 and 2, unspecified in case 3. Each of them underwent a diagnostic work-up for heparin allergy, which included prick and intradermal tests with commonly used heparins and patch testing with undiluted heparins and disinfectants. Results: Patch tests with disinfectants were negative in all 3 cases. In case 2, all allergological tests were negative. In cases 1 and 3, delayed positivity emerged for nadroparin calcium and at least one other heparin tested. Intravenous and/or subcutaneous provocation testing was done with an alternative heparin which produced negative results in skin tests (heparin sodium in case 1, pentasaccharide fondaparinux in case 3. In both cases the alternative drug was tolerated. After our evaluation, all 3 patients underwent surgery with no heparin-related complications. Discussion: The presenting clinical features in these 3 cases provided no information on which reactions were likely to be allergic: all 3 patients presented with similar local delayed reaction. The allergic reactions were identified only after cutaneous testing.

Theoretical calculation of shakeup intensities using Xa--SW wave functions

International Nuclear Information System (INIS)

Tse, J.S.; Loubriel, G.

1981-01-01

The ground and 1s core hole state molecular wave functions of CH 4 , NH 3 , H 2 O, and HF obtained from Xa--SW calculations using the touching spheres (TS) and overlapping spheres (OS) approximations are used to calculate the intensity of shakeup satellites observed in their ls core level photoelectron spectra. The sudden approximation was assumed in the calculation. In case of TS Xa--SW wave functions, the one electron overlap integral inside the intersphere was calculated via Green's theorem. For OS Xa--SW wave functions, the integration over the awkwardly shaped intersphere region was circumvented by distributing the intersphere charge into the atomic spheres according to the charge partition scheme suggested by Case and Karplus. Our results show that there are no significant differences between the shakeup energies calculated from the TS and OS approximations. However, shakeup intensities calculated from TS Xa--SW wave functions are more reliable and in better numerical agreement with experiment

Effect of MCM09, an active site-directed inhibitor of factor Xa, on B16-BL6 melanoma lung colonies in mice.

Science.gov (United States)

Rossi, C; Hess, S; Eckl, R W; di Lena, A; Bruno, A; Thomas, O; Poggi, A

2006-03-01

Treatment with anticoagulant drugs has shown potential inhibitory effect on tumor invasion, although the relationship with clotting inhibition was not clear. The aim of our study was to evaluate the potential antitumor activity of MCM09, a newly developed, active site-directed, small molecule inhibitor of factor Xa (FXa) [WO0216312], and to relate the findings to anticlotting potency. MCM09 (0.1-10 mg kg(-1)) or heparin (H; 10 mg kg(-1)) was injected intravenously (i.v.), with 5 x 10(4) B16-BL6 melanoma cells, in C57BL/6 mice. Mice were killed after 18 days, to count lung colonies. Ex vivo anticoagulant activity was measured by activated partial thromboplastin time (APTT) on mouse plasma. MCM09, a selective inhibitor of FXa (IC-50 = 2.4 nm against human FXa), inhibited in a dose-dependent manner B16-BL6 melanoma lung colonies in mice. Mean lung metastasis number was 20.9 +/- 4.8 in controls (n = 10), 1.2 +/- 0.4 in mice treated with H, 10 mg kg(-1) i.v. (P < 0.01), 0.9 +/- 0.3, 9.2 +/- 2.2 and 15.5 +/- 2.6 in mice treated with MCM09, at 10 (P < 0.01), 1 (P < 0.05) and 0.1 mg kg(-1) i.v. (ns), respectively. MCM09 (10 mg kg(-1) i.v.) significantly prolonged APTT (57.1 +/- 10.2 s) 30 min after i.v. injection when compared with controls (25.3 +/- 1.6 s; P < 0.05). Lung colonies were 74.2-72.6% reduced by MCM09 (10 mg kg(-1)) given 60 or 120 min before cells, but not by MCM09 given 60 min thereafter, suggesting a direct cell interaction as a mechanism underlying antitumor activity.

Heparin for assisted reproduction.

Science.gov (United States)

Akhtar, Muhammad A; Sur, Shyamaly; Raine-Fenning, Nick; Jayaprakasan, Kannamannadiar; Thornton, Jim G; Quenby, Siobhan

2013-08-17

Heparin as an adjunct in assisted reproduction (peri-implantation heparin) is given at or after egg collection or at embryo transfer during assisted reproduction. Heparin has been advocated to improve embryo implantation and clinical outcomes.  It has been proposed that heparin enhances the intra-uterine environment by improving decidualisation with an associated activation of growth factors and a cytokine expression profile in the endometrium that is favourable to pregnancy. To investigate whether the administration of heparin around the time of implantation (peri-implantation heparin) improves clinical outcomes in subfertile women undergoing assisted reproduction. A comprehensive and exhaustive search strategy was developed in consultation with the Trials Search Co-ordinator of the Cochrane Menstrual Disorders and Subfertility Group (MDSG). The strategy was used in an attempt to identify all relevant studies regardless of language or publication status (published, unpublished, in press, and in progress). Relevant trials were identified from both electronic databases and other resources (last search 6 May 2013). All randomised controlled trials (RCTs) were included where peri-implantation heparin was given during assisted reproduction. Peri-implantation low molecular weight heparin (LMWH) during IVF/ICSI was given at or after egg collection or at embryo transfer in the included studies. Live birth rate was the primary outcome. Two review authors independently assessed the eligibility and quality of trials and extracted relevant data. The quality of the evidence was evaluated using GRADE methods. Three RCTs (involving 386 women) were included in the review.Peri-implantation LMWH administration during assisted reproduction was associated with a significant improvement in live birth rate compared with placebo or no LMWH (odds ratio (OR) 1.77, 95% confidence interval (CI) 1.07 to 2.90, three studies, 386 women, I(2) = 51%, very low quality evidence with high

Sequence similarity between the erythrocyte binding domain 1 of the Plasmodium vivax Duffy binding protein and the V3 loop of HIV-1 strain MN reveals binding residues for the Duffy Antigen Receptor for Chemokines

Directory of Open Access Journals (Sweden)

Garry Robert F

2011-01-01

Full Text Available Abstract Background The surface glycoprotein (SU, gp120 of the human immunodeficiency virus (HIV must bind to a chemokine receptor, CCR5 or CXCR4, to invade CD4+ cells. Plasmodium vivax uses the Duffy Binding Protein (DBP to bind the Duffy Antigen Receptor for Chemokines (DARC and invade reticulocytes. Results Variable loop 3 (V3 of HIV-1 SU and domain 1 of the Plasmodium vivax DBP share a sequence similarity. The site of amino acid sequence similarity was necessary, but not sufficient, for DARC binding and contained a consensus heparin binding site essential for DARC binding. Both HIV-1 and P. vivax can be blocked from binding to their chemokine receptors by the chemokine, RANTES and its analog AOP-RANTES. Site directed mutagenesis of the heparin binding motif in members of the DBP family, the P. knowlesi alpha, beta and gamma proteins abrogated their binding to erythrocytes. Positively charged residues within domain 1 are required for binding of P. vivax and P. knowlesi erythrocyte binding proteins. Conclusion A heparin binding site motif in members of the DBP family may form part of a conserved erythrocyte receptor binding pocket.

Cultured human foreskin fibroblasts produce a factor that stimulates their growth with properties similar to basic fibroblast growth factor

International Nuclear Information System (INIS)

Story, M.T.

1989-01-01

To determine if fibroblasts could be a source of fibroblast growth factor (FGF) in tissue, cells were initiated in culture from newborn human foreskin. Fibroblast cell lysates promoted radiolabeled thymidine uptake by cultured quiescent fibroblasts. Seventy-nine percent of the growth-promoting activity of lysates was recovered from heparin-Sepharose. The heparin-binding growth factor reacted on immunoblots with antiserum to human placenta-derived basic FGF and competed with iodinated basic FGF for binding to antiserum to (1-24)bFGF synthetic peptide. To confirm that fibroblasts were the source of the growth factor, cell lysates were prepared from cells incubated with radiolabeled methionine. Heparin affinity purified material was immunoprecipitated with basic FGF antiserum and electrophoresed. Radiolabeled material was detected on gel autoradiographs in the same molecular weight region as authentic iodinated basic FGF. The findings are consistant with the notion that cultured fibroblasts express basic FGF. As these cells also respond to the mitogen, it is possible that the regulation of their growth is under autocrine control. Fibroblasts may be an important source of the growth factor in tissue

The regulatory role of heparin on c-Met signaling in hepatocellular carcinoma cells.

Science.gov (United States)

İşcan, Evin; Güneş, Aysim; Korhan, Peyda; Yılmaz, Yeliz; Erdal, Esra; Atabey, Neşe

2017-06-01

The role of heparin as an anticoagulant is well defined; however, its role in tumorigenesis and tumor progression is not clear yet. Some studies have shown that anticoagulant treatment in cancer patients improve overall survival, however, recent clinical trials have not shown a survival benefit in cancer patients receiving heparin treatment. In our previous studies we have shown the inhibitory effects of heparin on Hepatocyte Growth Factor (HGF)-induced invasion and migration in hepatocellular carcinoma (HCC) cells. In this study, we showed the differential effects of heparin on the behaviors of HCC cells based on the presence or absence of HGF. In the absence of HGF, heparin activated HGF/c-Met signaling and promoted motility and invasion in HCC cells. Heparin treatment led to c-Met receptor dimerization and activated c-Met signaling in an HGF independent manner. Heparin-induced c-Met activation increased migration and invasion through ERK1/2, early growth response factor 1 (EGR1) and Matrix Metalloproteinases (MMP) axis. Interestingly, heparin modestly decreased the proliferation of HCC cells by inhibiting activatory phosphorylation of Akt. The inhibition of c-Met signaling reversed heparin-induced increase in motility and invasion and, proliferation inhibition. Our study provides a new perspective into the role of heparin on c-Met signaling in HCC.

Surface biomimetic modification with laminin-loaded heparin/poly-L-lysine nanoparticles for improving the biocompatibility

Energy Technology Data Exchange (ETDEWEB)

Liu, Tao, E-mail: 11140021@hyit.edu.cn [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an (China); Hu, Youdong [Department of Geriatrics, The Affiliated Huai' an Hospital of Xuzhou Medical College, Huai' an (China); Tan, Jianying [Key Lab. of Advanced Technology for Materials of Chinese Education Ministry, School of Materials Science and Engineering, Southwest Jiaotong University, Chengdu (China); Liu, Shihui [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an (China); Chen, Junying [Key Lab. of Advanced Technology for Materials of Chinese Education Ministry, School of Materials Science and Engineering, Southwest Jiaotong University, Chengdu (China); Guo, Xin; Pan, Changjiang [Jiangsu Provincial Key Laboratory for Interventional Medical Devices, Huaiyin Institute of Technology, Huai' an (China); Li, Xia, E-mail: xial_li@qq.com [Department of Geriatrics, The Affiliated Huai' an Hospital of Xuzhou Medical College, Huai' an (China)

2017-02-01

Late thrombus and restenosis caused by delayed endothelialization and insufficient biocompatibility of polymer coating continue to be the greatest limitations of drug-eluting stents. In this study, based on the specific structure of vascular basement membrane, a novel biomimetic nano-coating was constructed by incorporating laminin into electrostatic-assembled heparin/poly-L-lysine nanoparticles. Alteration of heparin and poly-L-lysine concentration ratio in a certain range has no significantly influence nanoparticle size, uniformity and stability, but may affect the chemical property and subsequently the binding efficiency to dopamine-coated titanium surface. By use of this feature, four different nanoparticles were synthesized and immobilized on titanium surface for creating gradient nanoparticle binding density. According to in vitro biocompatibility evaluation, the nanoparticle modified surfaces were found to effectively block coagulation pathway and reduce thrombosis formation. Moreover, NP10L and NP15L modified surface with relatively low heparin exposing density (4.9 to 7.1 μg/cm2) showed beneficial effect in selective promoting EPCs and ECs proliferation, as well as stimulating cell migration and NO synthesis. - Highlights: • A novel laminin-loaded anticoagulant nanoparticle was prepared and used for titanium surface modification. • The nanoparticle binding density was adjustable by alteration the concentration ratio of heparin and poly-L-lysine. • In a certain range of NPs density, the surface was found to selectively direct platelet and vascular cells behavior.

Surface biomimetic modification with laminin-loaded heparin/poly-L-lysine nanoparticles for improving the biocompatibility

International Nuclear Information System (INIS)

Liu, Tao; Hu, Youdong; Tan, Jianying; Liu, Shihui; Chen, Junying; Guo, Xin; Pan, Changjiang; Li, Xia

2017-01-01

Late thrombus and restenosis caused by delayed endothelialization and insufficient biocompatibility of polymer coating continue to be the greatest limitations of drug-eluting stents. In this study, based on the specific structure of vascular basement membrane, a novel biomimetic nano-coating was constructed by incorporating laminin into electrostatic-assembled heparin/poly-L-lysine nanoparticles. Alteration of heparin and poly-L-lysine concentration ratio in a certain range has no significantly influence nanoparticle size, uniformity and stability, but may affect the chemical property and subsequently the binding efficiency to dopamine-coated titanium surface. By use of this feature, four different nanoparticles were synthesized and immobilized on titanium surface for creating gradient nanoparticle binding density. According to in vitro biocompatibility evaluation, the nanoparticle modified surfaces were found to effectively block coagulation pathway and reduce thrombosis formation. Moreover, NP10L and NP15L modified surface with relatively low heparin exposing density (4.9 to 7.1 μg/cm2) showed beneficial effect in selective promoting EPCs and ECs proliferation, as well as stimulating cell migration and NO synthesis. - Highlights: • A novel laminin-loaded anticoagulant nanoparticle was prepared and used for titanium surface modification. • The nanoparticle binding density was adjustable by alteration the concentration ratio of heparin and poly-L-lysine. • In a certain range of NPs density, the surface was found to selectively direct platelet and vascular cells behavior.

Ontogeny of basic fibroblast growth factor binding sites in mouse ocular tissues

International Nuclear Information System (INIS)

Fayein, N.A.; Courtois, Y.; Jeanny, J.C.

1990-01-01

Basic fibroblast growth factor (bFGF) binding to ocular tissues has been studied by autoradiographical and biochemical approaches directly performed on sections during mouse embryonic and postnatal development. Frozen sections of embryos (9 to 18 days), newborns, and adults (1 day to 6 months) were incubated with iodinated bFGF. One specific FGF binding site (KD = 2.5 nM) is colocalized with heparan sulfate proteoglycans of the basement membranes and is heparitinase sensitive. It first appears at Day 9 around the neural tube, the optic vesicles, and below the head ectoderm and by Day 14 of embryonic development is found in all basement membranes of the eye. At Day 16, very intensely labeled patches appear, corresponding to mast cells which have been characterized by metachromatic staining of their heparin-rich granulations with toluidine blue. In addition to the latter binding, we have also observed a general diffuse distribution of silver grains on all tissues and preferentially in the ecto- and neuroectodermic tissues. From Days 17-18, there is heterogeneous labeling inside the retina, localized in the pigmented epithelium and in three different layers colocalized with the inner and outer plexiform layers and with the inner segments of the photoreceptors. This binding is heparitinase resistant but N-glycanase sensitive and may represent a second specific binding site corresponding to cellular FGF receptors (KD = 280 pM). Both types of binding patterns observed suggest a significant role for bFGF in eye development and physiology

Role of the A+ helix in heparin binding to protein C inhibitor

NARCIS (Netherlands)

Elisen, M. G.; Maseland, M. H.; Church, F. C.; Bouma, B. N.; Meijers, J. C.

1996-01-01

Interactions between proteins and heparin(-like) structures involve electrostatic forces and structural features. Based on charge distributions in the linear sequence of protein C inhibitor (PCI), two positively charged regions of PCI were proposed as possible candidates for this interaction. The

Apixaban versus enoxaparin for thromboprophylaxis after knee replacement (ADVANCE-2): a randomised double-blind trial

DEFF Research Database (Denmark)

Lassen, Michael Rud; Raskob, Gary E; Gallus, Alexander

2010-01-01

BACKGROUND: Low-molecular-weight heparins such as enoxaparin are preferred for prevention of venous thromboembolism after major joint replacement. Apixaban, an orally active factor Xa inhibitor, might be as effective, have lower bleeding risk, and be easier to use than is enoxaparin. We assessed ...

Heparin modulates the endopeptidase activity of Leishmania mexicana cysteine protease cathepsin L-Like rCPB2.8.

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Wagner A S Judice

Full Text Available Cysteine protease B is considered crucial for the survival and infectivity of the Leishmania in its human host. Several microorganism pathogens bind to the heparin-like glycosaminoglycans chains of proteoglycans at host-cell surface to promote their attachment and internalization. Here, we have investigated the influence of heparin upon Leishmania mexicana cysteine protease rCPB2.8 activity.THE DATA ANALYSIS REVEALED THAT THE PRESENCE OF HEPARIN AFFECTS ALL STEPS OF THE ENZYME REACTION: (i it decreases 3.5-fold the k 1 and 4.0-fold the k -1, (ii it affects the acyl-enzyme accumulation with pronounced decrease in k 2 (2.7-fold, and also decrease in k 3 (3.5-fold. The large values of ΔG  =  12 kJ/mol for the association and dissociation steps indicate substantial structural strains linked to the formation/dissociation of the ES complex in the presence of heparin, which underscore a conformational change that prevents the diffusion of substrate in the rCPB2.8 active site. Binding to heparin also significantly decreases the α-helix content of the rCPB2.8 and perturbs the intrinsic fluorescence emission of the enzyme. The data strongly suggest that heparin is altering the ionization of catalytic (Cys(25-S(-/(His(163-Im(+ H ion pair of the rCPB2.8. Moreover, the interaction of heparin with the N-terminal pro-region of rCPB2.8 significantly decreased its inhibitory activity against the mature enzyme.Taken together, depending on their concentration, heparin-like glycosaminoglycans can either stimulate or antagonize the activity of cysteine protease B enzymes during parasite infection, suggesting that this glycoconjugate can anchor parasite cysteine protease at host cell surface.

Heparin-based hydrogels with tunable sulfation & degradation for anti-inflammatory small molecule delivery.

Science.gov (United States)

Peng, Yifeng; Tellier, Liane E; Temenoff, Johnna S

2016-08-16

Sustained release of anti-inflammatory agents remains challenging for small molecule drugs due to their low molecular weight and hydrophobicity. Therefore, the goal of this study was to control the release of a small molecule anti-inflammatory agent, crystal violet (CV), from hydrogels fabricated with heparin, a highly sulfated glycosaminoglycan capable of binding positively-charged molecules such as CV. In this system, both electrostatic interactions between heparin and CV and hydrogel degradation were tuned simultaneously by varying the level of heparin sulfation and varying the amount of dithiothreitol within hydrogels, respectively. It was found that heparin sulfation significantly affected CV release, whereby more sulfated heparin hydrogels (Hep and Hep(-N)) released CV with near zero-order release kinetics (R-squared values between 0.96-0.99). Furthermore, CV was released more quickly from fast-degrading hydrogels than slow-degrading hydrogels, providing a method to tune total CV release between 5-15 days while maintaining linear release kinetics. In particular, N-desulfated heparin hydrogels exhibited efficient CV loading (∼90% of originally included CV), near zero-order CV release kinetics, and maintenance of CV bioactivity after release, making this hydrogel formulation a promising CV delivery vehicle for a wide range of inflammatory diseases.

XA readout chip characteristics and CdZnTe spectral measurements

International Nuclear Information System (INIS)

Barbier, L.M.; Birsa, F.; Odom, J.

1999-01-01

The authors report on the performance of a CdZnTe (CZT) array readout by an XA (X-ray imaging chip produced at the AMS foundry) application specific readout chip (ASIC). The array was designed and fabricated at NASA/Goddard Space Flight Center (GSFC) as a prototype for the Burst Arc-Second Imaging and Spectroscopy gamma-ray instrument. The XA ASIC was obtained from Integrated Detector and Electronics (IDE), in Norway. Performance characteristics and spectral data for 241 Am are presented both at room temperature and at -20 C. The measured noise (σ) was 2.5 keV at 60 keV at room temperature. This paper represents a progress report on work with the XA ASIC and CZT detectors. Work is continuing and in particular, larger arrays are planned for future NASA missions

Purification of foot-and-mouth disease virus by heparin as ligand for certain strains.

Science.gov (United States)

Du, Ping; Sun, Shiqi; Dong, Jinjie; Zhi, Xiaoying; Chang, Yanyan; Teng, Zhidong; Guo, Huichen; Liu, Zaixin

2017-04-01

The goal of this project was to develop an easily operable and scalable process for the recovery and purification of foot-and-mouth disease virus (FMDV) from cell culture. Heparin resins HipTrap Heparin HP and AF-Heparin HC-650 were utilized to purify FMDV O/HN/CHA/93. Results showed that the purity of AF-Heparin HC-650 was ideal. Then, the O/HN/CHA/93, O/Tibet/CHA/99, Asia I/HN/06, and A/CHA/HB/2009 strains were purified by AF-Heparin HC-650. Their affinity/virus recoveries were approximately 51.2%/45.8%, 71.5%/70.9%, 96.4%/73.5, and 59.5%/42.1%, respectively. During a stepwise elution strategy, the viral particles were mainly eluted at 300mM ionic strength peaks. The heparin affinity chromatography process removed more than 94% of cellular and medium proteins. Anion exchange resin Capto Q captured four FMD virus particles; 40% of binding proteins and 80%-90% of viral particles were eluted at 450mM NaCl. Moreover, ionic strength varied from 30 to 450mM had no effect on the immunity to FMDV. The results revealed that heparin sulfate may be the main receptor for CHA/99 strain attachment-susceptible cells. Heparin affinity chromatography can reach perfect results, especially when used as a ligand of the virus. Anion exchange is useful only as previous step for further purification. Copyright © 2016. Published by Elsevier B.V.

Sterilization of heparinized cuprophan hemodialysis membranes

OpenAIRE

ten Hoopen, Hermina W.M.; Hinrichs, W.L.J.; Hinrichs, W.L.J.; Engbers, G.H.M.; Feijen, Jan

1996-01-01

The effects of sterilization of dry heparinized Cuprophan hemodialysis membranes by means of ethylene oxide (EtO) exposure, gamma irradiation, or steam on the anticoagulant activity and chemical characteristics of immobilized heparin and the permeability of the membrane were investigated. Sterilization did not result in a release of heparin or heparin fragments from heparinized Cuprophan. Sterilization of heparinized Cuprophan by means of EtO exposure and gamma irradiation induced a slight, i...

Protein binding of glufosinate and factors affecting it revealed by an equilibrium dialysis technique.

Science.gov (United States)

Hori, Y; Koyama, K; Fujisawa, M; Nakajima, M; Shimada, K; Hirose, Y; Kohda, Y; Akuzawa, H

2001-09-01

We investigated the protein binding of glufosinate ammonium (GLF) and several factors affecting this binding using human serum albumin (HSA) and human volunteer serum under various conditions. The mean ratios of the free GLF (RFr-GLF) to 4% HSA were examined in the sera of patients described elsewhere at GLF levels from 1 microg/mL to 500 microg/mL; the range was found to be only from 0.80 to 0.88. Neither the incubation temperature nor buffers containing different chloride ion concentrations had any effect on the RFr-GLF to HSA. Moreover, the addition of heparin, glycoprotein-alpha1-acid (AAG), and sodium azide had no effect on the RFr-GLF. However, pH of the isotonic phosphate buffer and the addition of palmitic or oleic acid were seen to have an effect. In this study, the mean RFr-GLF to healthy human serum was 0.99. This high value was evidenced that GLF was rapidly excreted through the renal route.

Heparin pharmacovigilance in Brazil.

Science.gov (United States)

Junqueira, Daniela Rezende Garcia; Viana, Thércia Guedes; Peixoto, Eliane R de M; Barros, Fabiana C R de; Carvalho, Maria das Graças; Perini, Edson

2011-01-01

To investigate the biological origin of injectable unfractioned heparin available in Brazilian market by discussing the impact of the profile of commercial products and the changes in heparin monograph on the drug safety. The Anvisa data base for the Registered Products of Pharmaceutical Companies and the Dictionary of Pharmaceutical Specialties (DEF 2008/2009) were searched. A survey with industries having an active permission for marketing the drug in Brazil was conducted. Five companies were granted a permission to market unfractioned heparin in Brazil. Three of them are porcine in origin and two of them are bovine in origin, with only one explicitly showing this information in the package insert. The effectiveness and safety of heparin studied in non-Brazilian populations may not represent the Brazilian reality, since most countries no longer produce bovine heparin. The currently marketed heparin has approximately 10% less anticoagulant activity than that previously produced and this change may have clinical implications. Evidence about the lack of dose interchangeability between bovine and porcine heparins and the unique safety profile of these drugs indicates the need to follow the treatment and the patients' response. Events threatening the patient's safety must be reported to the pharmacovigilance system in each particular country.

Covalent Binding of Heparin to Functionalized PET Materials for Improved Haemocompatibility

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Metod Kolar

2015-03-01

Full Text Available The hemocompatibility of vascular grafts made from poly(ethylene terephthalate (PET is insufficient due to the rapid adhesion and activation of blood platelets that occur upon incubation with whole blood. PET polymer was treated with NHx radicals created by passing ammonia through gaseous plasma formed by a microwave discharge, which allowed for functionalization with amino groups. X-ray photoelectron spectroscopy characterization using derivatization with 4-chlorobenzaldehyde indicated that approximately 4% of the –NH2 groups were associated with the PET surface after treatment with the gaseous radicals. The functionalized polymers were coated with an ultra-thin layer of heparin and incubated with fresh blood. The free-hemoglobin technique, which is based on the haemolysis of erythrocytes, indicated improved hemocompatibility, which was confirmed by imaging the samples using confocal optical microscopy. A significant decrease in number of adhered platelets was observed on such samples. Proliferation of both human umbilical vein endothelial cells and human microvascular endothelial cells was enhanced on treated polymers, especially after a few hours of cell seeding. Thus, the technique represents a promising substitute for wet-chemical modification of PET materials prior to coating with heparin.

A genome-wide association study of heparin-induced thrombocytopenia using an electronic medical record

DEFF Research Database (Denmark)

Karnes, Jason H; Cronin, Robert M; Rollin, Jerome

2015-01-01

Heparin-induced thrombocytopenia (HIT) is an unpredictable, potentially catastrophic adverse effect of heparin treatment resulting from an immune response to platelet factor 4 (PF4)/heparin complexes. No genome-wide evaluations have been performed to identify potential genetic influences on HIT. ...

Deep vein thrombosis, ecythyma gangrenosum and heparin-induced thrombocytopenia occurring in a man with a heterozygous Factor V Leiden mutation

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Mariya Apostolova

2012-11-01

Full Text Available Skin necrosis and limb gangrene are occasional thrombotic manifestations of anticoagulation therapy. We report a man heterozygous for the Factor V Leiden (FVL mutation, and with a history of recurrent deep venous thrombosis, who initially presented with a necrotic skin lesion of the right flank while on warfarin therapy with a therapeutic international normalized ratio. Warfarin was discontinued and he received intravenous heparin. Thereafter he developed thrombocytopenia and pedal erythema and was diagnosed with heparin-induced thrombocytopenia (HIT. Heparin was replaced with argatroban. He ultimately underwent bilateral below-knee amputations for the thrombotic complications of the HIT. The initial necrotic lesion healed with antibiotics and wound care. Pathologic examination of multiple biopsy specimens revealed two separate lesions. One was necrotic tissue infiltrated with methicillin resistant Staphylococcus aureus having features of ecthyma gangrenosum. The second showed thrombotic changes consistent with HIT. The case illustrates the differential diagnosis of skin necrosis and limb gangrene in patients on warfarin and heparin, and also the clinical complexities that can occur in a FVL heterozygote.

Cationization of heparin for film applications

Czech Academy of Sciences Publication Activity Database

Šimkovic, I.; Mendichi, R.; Kelnar, Ivan; Filip, J.; Hricovíni, M.

2015-01-01

Roč. 115, 22 January (2015), s. 551-558 ISSN 0144-8617 Institutional support: RVO:61389013 Keywords : heparin * cationization * NMR Subject RIV: CD - Macromolecular Chemistry Impact factor: 4.219, year: 2015

Heparin: Past, Present, and Future.

Science.gov (United States)

Oduah, Eziafa I; Linhardt, Robert J; Sharfstein, Susan T

2016-07-04

Heparin, the most widely used anticoagulant drug in the world today, remains an animal-derived product with the attendant risks of adulteration and contamination. A contamination crisis in 2007-2008 increased the impetus to provide non-animal-derived sources of heparin, produced under cGMP conditions. In addition, recent studies suggest that heparin may have significant antineoplastic activity, separate and distinct from its anticoagulant activity, while other studies indicate a role for heparin in treating inflammation, infertility, and infectious disease. A variety of strategies have been proposed to produce a bioengineered heparin. In this review, we discuss several of these strategies including microbial production, mammalian cell production, and chemoenzymatic modification. We also propose strategies for creating "designer" heparins and heparan-sulfates with various biochemical and physiological properties.

Comparative study of heparin-binding proteins profile of Murrah buffalo (Bubalus bubalis semen

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S. S. Ramteke

2014-09-01

Full Text Available Aim: The experiment was conducted to study the total seminal plasma protein (TSPP and heparin-binding proteins (HBPs in relation to initial semen quality of buffalo bull. Materials and Methods: Semen from two Murrah buffalo bulls (bull no. 605 and 790 with mass motility of ≥3+ were used for the study and categorized into three groups (Group I- Mass motility 3+, Group II- Mass motility 4+ and Group III- Mass motility 5+. Seminal plasma from semen was separated by centrifugation. HBPs was isolated and purified from heparin-agarose affinity column by modified elution buffer. TSPP and isolated HBPs concentration was estimated by Lowry’s method. The purified HBPs were resolved on Sodium dodecyl sulfate polyacrylamide gel electrophoresis to check the protein profile of two bulls. Results: The mean values of TSPP concentrations in bull no. 605 and 790 in Group I, II and III were 30.64±0.12, 31.66±0.09, 32.53±0.19 and 28.51±0.09, 29.49±0.15, 30.45±0.17 mg/mL, respectively. The mean values of HBPs concentrations in bull no. 605 and 790 in Group I, II and III were 3.11±0.07, 3.32±0.06, 3.46±0.08 and 2.51±0.08, 2.91±0.05, 3.10±0.03 mg/mL, respectively. Both the values of TSPP and HBPs were significantly higher (p<0.01 in bull no. 605 when compared to 790 in all the three groups. 31 kDa HBP was more intensely present in bull no. 605, thus may indicate its superiority over bull no. 790 in relation to fertility potential. Conclusion: TSPP and HBPs shows variation in concentration with respect to initial semen quality. Furthermore, presence of fertility related 31 kDa HBPs in one of the bull may be an indication of high fertility of a bull. In future, in-vivo and in-vitro correlative study on larger basis is needed for the establishment of fertility-related HBPs in semen which might establish criteria for selection of buffalo bull with high fertility potential.

Activation of factor VII bound to tissue factor: A key early step in the tissue factor pathway of blood coagulation

International Nuclear Information System (INIS)

Rao, L.V.M.; Rapaport, S.I.

1988-01-01

Whether the factor VII/tissue factor complex that forms in tissue factor-dependent blood coagulation must be activated to factor VIIa/tissue factor before it can activate its substrates, factor X and IX, has been a difficult question to answer because the substrates, once activated, back-activate factor VII. The earlier studies suggested that human factor VII/tissue factor cannot activate factor IX. Studies have now been extended to the activation of factor X. Reaction mixtures were made with purified factor VII, X, and tissue factor; in some experiments antithrombin III and heparin were added to prevent back-activation of factor VII. Factor X was activated at similar rates in reaction mixtures containing either VII or factor VIIa after an initial 30-sec lag with factor VII. In reaction mixtures with factor VII a linear activation of factor X was established several minutes before cleavage of 125 I-labeled factor VII to the two-chain activated molecule was demonstrable on gel profiles. These data suggest that factor VII/tissue factor cannot activate measurable amounts of factor X over several minutes. Overall, the results support the hypothesis that a rapid preferential activation of factor VII bound to tissue factor by trace amounts of factor Xa is a key early step in tissue factor-dependent blood coagulation

Heparin induced thrombocytopenia type ii and myocardial infarction: Two case reports

Directory of Open Access Journals (Sweden)

Antonijević Nebojša

2004-01-01

Full Text Available Heparin-induced thrombocytopenia (HIT type II is an acquired thrombophylic state and life-threatening immune complication of a heparin treatment mainly clinically manifested by marked thrombocytopenia, frequently by arterial and venous thrombosis, and sometimes by skin changes. Functional assay as heparin aggregation test and 14C-serotonin release assays are used in diagnostics as well as antigen assays of which detection tests for heparin-platelet factor 4 antibodies are most frequently used. Considering the fact that there is no single reliable assays for HIT II detection available, sometimes it is necessary to combine both of the above-mentioned types of assays. We present the case of a 57-year-old patient with an acute anterior myocardial infarction with cardiac insufficiency of III and IV degree according to Killip, recurrent ventricular fibrillation and diabetes mellitus type II developing thrombocytopenia to 37x10 9/l accompanied with typical skin changes. The diagnosis was confirmed by the heparin aggregation test. The second patient aged 70 undergoing the treatment for anteroseptal myocardial infarction and reinfarction of the inferior wall complicated by a cardiogenic shock and acute right bundle branch block developed thrombocytopenia 59x10 9/I on the third day of the heparin therapy, with the remark that he had received a heparin therapy during the first infarction as well. Antibodies against heparin-platelet factor 4 were detected by particle gel ID-HPF4 immunoassay. In both patients, the disease had a lethal outcome despite all then available therapeutic measures applied. Further on we discuss advantages of certain types of tests, a therapy doctrine, need for urgent therapeutic measures, inclusive of the administration of anitithrombins, avoidance of harmful procedures like low-molecular-weight heparins administration and prophylactic platelet transfusion as well as preventive measures.

Human recombinant interleukin-1 beta- and tumor necrosis factor alpha-mediated suppression of heparin-like compounds on cultured porcine aortic endothelial cells

International Nuclear Information System (INIS)

Kobayashi, M.; Shimada, K.; Ozawa, T.

1990-01-01

Cytokines are known to tip the balance of the coagulant-anticoagulant molecules on the endothelial cell surface toward intravascular coagulation. Their effects on endothelial cell surface-associated heparin-like compounds have not been examined yet. Incorporation of [35S]sulfate into heparan sulfate on cultured porcine aortic endothelial cells was suppressed by human recombinant interleukin-1 beta (rIL-1 beta) or tumor necrosis factor alpha (rTNF alpha) in a dose- and time-dependent manner with little effect on cell number, protein content, and [3H]leucine incorporation of cells. Maximal inhibition was achieved by incubation of cells with 100 ng/ml of rIL-1 beta or 5 ng/ml of rTNF alpha for 12-24 hours, resulting in a reduction of the synthesis of heparan sulfate on the cell surface by approximately 50%. The dose dependency was consistent with that seen in the stimulation of endothelial cell procoagulant activity by each cytokine. The suppression of heparan sulfate synthesis was sustained for at least 48 hours after pretreatment of cells with cytokines and was unchanged after the addition of indomethacin or polymyxin B. The rate of degradation of prelabeled 35S-heparan sulfate on the cell surface was not altered by cytokine treatments. Neither the size, the net negative charge, nor the proportion of the molecule with high affinity for antithrombin III of endothelial cell heparan sulfate was changed by cytokines. Furthermore, specific binding of 125I-labeled antithrombin III to the endothelial cell surface was reduced to 40-60% of control by cytokines. In parallel with reduction in binding, antithrombin III cofactor activity was partially diminished in cytokine-treated endothelial cells. Thus, cytokine-mediated suppression of heparin-like substance on endothelial cells appears to be another cytokine-inducible endothelial effects affecting coagulation

Fine mapping of the rice bacterial blight resistance gene Xa-4 and its co-segregation marker

Institute of Scientific and Technical Information of China (English)

无

2000-01-01

An F2 population developed from the Xa-4 near isogenic lines,IR24 and IRBB4,was used for fine mapping of the rice bacterial blight resistance gene,Xa-4.Some restriction fragment length polymorphism (RFLP) markers on the high-density map constructed by Harushima et al.and the amplified DNA fragments homologous to the conserved domains of plant disease resistance (R) genes were used to construct the genetic linkage map around the gene Xa-4 by scoring susceptible individuals in the population.Xa-4 was mapped between the RFLP marker G181 and the polymerase chain reaction (PCR) marker M55.The R gene homologous fragment marker RS13 was found co-segregating with Xa-4 by analyzing all the plants in the population.This result opened an approach to map-based cloning of this gene,and marker RS13 can be applied to molecular marker-assisted selection of Xa-4 in rice breeding programs.

Low molecular weight heparin versus unfractionated heparin in the initial treatment of venous thromboembolism

NARCIS (Netherlands)

Hettiarachchi, R. J.; Prins, M. H.; Lensing, A. W.; Buller, H. R.

1998-01-01

In this review, we analyze data from randomized trials in which low molecular weight heparin was compared with unfractionated heparin, both to estimate the treatment effect of low molecular weight heparin in the initial treatment of venous thromboembolism and to evaluate the effect of the varied

Determination of enoxaparin with rotational thrombelastometry using the prothrombinase-induced clotting time reagent.

Science.gov (United States)

Schaden, Eva; Schober, Andreas; Hacker, Stefan; Spiss, Christian; Chiari, Astrid; Kozek-Langenecker, Sibylle

2010-04-01

Drug monitoring of low molecular weight heparin is generally not recommended, but could be reasonable in critically ill patients, whose risk for bleeding or thrombosis shows a high interpatient variability. Anti-Xa assays are not available around the clock even in central hospitals, whereas rotational thrombelastometry (ROTEM) becomes increasingly used at the bedside. Prothrombinase-induced clotting time (PiCT) reagent allows determination of factor Xa-inhibition in plasma. The aim of our study was to evaluate enoxaparin determination in whole blood with the ROTEM using specific test modifications, including PiCT. After ethics committee's approval, citrated whole blood obtained from overall 16 healthy volunteers was incubated with enoxaparin at 16 different anti-Xa concentrations. Main endpoint was the clotting time (CT) in ROTEM representing initial activation of clot formation. CT was determined in the new PiCT-ROTEM test, in a low-tissue factor-activated modification (LowTF-ROTEM) as well as in the commercially available heparin-sensitive ROTEM assays (HEPTEM and INTEM). In the absence of enoxaparin, CT values were 168.6 +/- 6.1 s (PiCT-ROTEM), 247.3 +/- 18.6 s (LowTF-ROTEM), and -6.2 +/- 7.9 s (INTEM-HEPTEM). A linear dependency (P anti-Xa concentration and CT was found for PiCT-ROTEM, LowTF-ROTEM, and for INTEM-HEPTEM with correlation coefficients of 0.93 for PiCT-ROTEM, 0.94 for LowTF-ROTEM, and 0.81 for INTEM-HEPTEM. This in-vitro experiment demonstrates a strong correlation between enoxaparin anti-Xa concentrations and specific ROTEM tests. These promising assays should be further evaluated for monitoring anticoagulation in high-risk patients in clinical studies.

Qualitative and quantitative analysis of heparin and low molecular weight heparins using size exclusion chromatography with multiple angle laser scattering/refractive index and inductively coupled plasma/mass spectrometry detectors.

Science.gov (United States)

Ouyang, Yilan; Zeng, Yangyang; Yi, Lin; Tang, Hong; Li, Duxin; Linhardt, Robert J; Zhang, Zhenqing

2017-11-03

Heparin, a highly sulfated glycosaminoglycan, has been used as a clinical anticoagulant over 80 years. Low molecular weight heparins (LMWHs), heparins partially depolymerized using different processes, are widely used as clinical anticoagulants. Qualitative molecular weight (MW) and quantitative mass content analysis are two important factors that contribute to LMWH quality control. Size exclusion chromatography (SEC), relying on multiple angle laser scattering (MALS)/refractive index (RI) detectors, has been developed for accurate analysis of heparin MW in the absence of standards. However, the cations, which ion-pair with the anionic polysaccharide chains of heparin and LMWHs, had not been considered in previous reports. In this study, SEC with MALS/RI and inductively coupled plasma/mass spectrometry detectors were used in a comprehensive analytical approach taking both anionic polysaccharide and ion-paired cations heparin products. This approach was also applied to quantitative analysis of heparin and LMWHs. Full profiles of MWs and mass recoveries for three commercial heparin/LMWH products, heparin sodium, enoxaparin sodium and nadroparin calcium, were obtained and all showed higher MWs than previously reported. This important improvement more precisely characterized the MW properties of heparin/LMWHs and potentially many other anionic polysaccharides. Copyright © 2017 Elsevier B.V. All rights reserved.

Collection of heparinized plasma by plasmapheresis

NARCIS (Netherlands)

van der Meer, P. F.; Vrielink, H.; Pietersz, R. N.; Dekker, W. J.; Reesink, H. W.

1999-01-01

BACKGROUND AND OBJECTIVES: Heparinized plasma can be used for exchange transfusions in neonates and is usually collected by drawing whole blood using heparin as anticoagulant. The heparinized red blood cells and buffy coat cannot be used and are therefore discarded. To collect heparinized plasma

Analysis and characterization of heparin impurities.

Science.gov (United States)

Beni, Szabolcs; Limtiaco, John F K; Larive, Cynthia K

2011-01-01

This review discusses recent developments in analytical methods available for the sensitive separation, detection and structural characterization of heparin contaminants. The adulteration of raw heparin with oversulfated chondroitin sulfate (OSCS) in 2007-2008 spawned a global crisis resulting in extensive revisions to the pharmacopeia monographs on heparin and prompting the FDA to recommend the development of additional physicochemical methods for the analysis of heparin purity. The analytical chemistry community quickly responded to this challenge, developing a wide variety of innovative approaches, several of which are reported in this special issue. This review provides an overview of methods of heparin isolation and digestion, discusses known heparin contaminants, including OSCS, and summarizes recent publications on heparin impurity analysis using sensors, near-IR, Raman, and NMR spectroscopy, as well as electrophoretic and chromatographic separations.

Epitope mapping of functional domains of human factor V with human and mouse monoclonal antibodies

International Nuclear Information System (INIS)

Annamalai, A.E.; Rao, A.K.; Chiu, H.C.; Wang, D.; Dutta-Roy, A.K.; Colman, R.W.

1986-01-01

The authors previously described two human monoclonal antibodies (MAbs) which inactivated factor V. The authors have now purified the predominant antibody (H2) on protein A Sepharose using a pH gradient and typed it as IgG 1 ,. Immunoprecipitation of 125 I-human factor Va with H2 demonstrated specificity for the heavy chain (D), Mr = 105,000. The authors compared using ELISA the competitive binding to factor Va, of H2, H1 and two mouse MAbs, B38 (directed to E) and B10 (to activation peptide, Cl). All four antibodies recognized distinct epitopes in factor V with steric overlap in some cases. Factor Xa showed a concentration dependent competition for binding of H1, H2 and B38 but not B10 to factor V/Va in ELISA. All MAbs bound to factor V/Va in the absence of Ca ++ . However, Ca ++ at 8 mM increased the binding of H1 and H2 to 165% and 360% and did not have any effect on the binding of either mouse MAbs. Prothrombin at a concentration of up to 400 μg/ml did not inhibit binding of any of these antibodies. Thus, both the light (E) and heavy (D) chains of factor Va but not the activation peptide (Cl) interact with factor Xa as defined by the MAbs. In addition, sites on both chains for Ca ++ are recognized by particular MAbs (H1 and H2). These studies increase their knowledge of the interactions of factor V domains in the formation of prothrombinase complex

Cellular Responses Modulated by FGF-2 Adsorbed on Albumin/Heparin Layer-by-Layer Assemblies.

Science.gov (United States)

Kumorek, Marta; Kubies, Dana; Filová, Elena; Houska, Milan; Kasoju, Naresh; Mázl Chánová, Eliška; Matějka, Roman; Krýslová, Markéta; Bačáková, Lucie; Rypáček, František

2015-01-01

In a typical cell culture system, growth factors immobilized on the cell culture surfaces can serve as a reservoir of bio-signaling molecules, without the need to supplement them additionally into the culture medium. In this paper, we report on the fabrication of albumin/heparin (Alb/Hep) assemblies for controlled binding of basic fibroblast growth factor (FGF-2). The surfaces were constructed by layer-by-layer adsorption of polyelectrolytes albumin and heparin and were subsequently stabilized by covalent crosslinking with glutaraldehyde. An analysis of the surface morphology by atomic force microscopy showed that two Alb/Hep bilayers are required to cover the surface of substrate. The formation of the Alb/Hep assemblies was monitored by the surface plasmon resonance (SPR), the infrared multiinternal reflection spectroscopy (FTIR MIRS) and UV/VIS spectroscopy. The adsorption of FGF-2 on the cross-linked Alb/Hep was followed by SPR. The results revealed that FGF-2 binds to the Alb/Hep assembly in a dose and time-dependent manner up to the surface concentration of 120 ng/cm(2). The bioactivity of the adsorbed FGF-2 was assessed in experiments in vitro, using calf pulmonary arterial endothelial cells (CPAE). CPAE cells could attach and proliferate on Alb/Hep surfaces. The adsorbed FGF-2 was bioactive and stimulated both the proliferation and the differentiation of CPAE cells. The improvement was more pronounced at a lower FGF-2 surface concentration (30 ng/cm(2)) than on surfaces with a higher concentration of FGF-2 (120 ng/cm(2)).

[Thrombocytopenia induced by type II heparin and myocardial infarct: 2 case reports].

Science.gov (United States)

Antonijević, Nabojsa; Stanojević, Milica; Perunicić, Jovan; Djokić, Milan; Miković, Danijla; Kovac, Mirjana; Miljić, Predrag; Milosević, Rajko; Terzić, Branka; Vasiljević, Zorana

2004-01-01

Heparin-induced thrombocytopenia (HIT) type II is an acquired thrombophylic state and life-threatening immune complication of a heparin treatment mainly clinically manifested by marked thrombocytopenia, frequently by arterial and venous thrombosis, and sometimes by skin changes. Functional assay as heparin aggregation test and 14C-serotonin release assays are used in diagnostics as well as antigen assays of which detection tests for heparin-platelet factor 4 antibodies are most frequently used. Considering the fact that there is no single reliable assays for HIT II detection available, sometimes it is necessary to combine both of the above-mentioned types of assays. We present the case of a 57-year-old patient with an acute anterior myocardial infarction with cardiac insufficiency of III and IV degree according to Killip, recurrent ventricular fibrillation and diabetes mellitus type II developing thrombocytopenia to 37 x 10(9)/l accompanied with typical skin changes. The diagnosis was confirmed by the heparin aggregation test. The second patient aged 70 undergoing the treatment for anteroseptal myocardial infarction and reinfarction of the inferior wall complicated by a cardiogenic shock and acute right bundle branch block developed thrombocytopenia 59 x 10(9)/l on the third day of the heparin therapy, with the remark that he had received a heparin therapy during the first infarction as well. Antibodies against heparin-platelet factor 4 were detected by particle gel ID-HPF4 immuno-assay. In both patients, the disease had a lethal outcome despite all then available therapeutic measures applied. Further on we discuss advantages of certain types of tests, a therapy doctrine, need for urgent therapeutic measures, inclusive of the administration of antithrombins, avoidance of harmful procedures like low-molecular-weight heparins administration and prophylactic platelet transfusion as well as preventive measures.

Polyguluronate sulfate and its oligosaccharides but not heparin promotes FGF19/FGFR1c signaling

Science.gov (United States)

Lan, Ying; Zeng, Xuan; Guo, Zhihua; Zeng, Pengjiao; Hao, Cui; Zhao, Xia; Yu, Guangli; Zhang, Lijuan

2017-06-01

Fibroblast growth factor 19(FGF19) functions as a hormone by affecting glucose metabolism. FGF19 improves glucose tolerance when overexpressed in mice with impaired glucose tolerance or diabetes. A functional cellular FGF19 receptor consists of FGF receptor (FGFR) and glycosaminoglycan complexed with either α Klotho or β Klotho. Interestingly, in mice with diet-induced diabetes, a single injection of FGF1 is enough to restore blood sugar levels to a healthy range. FGF1 binds heparin with high affinity whereas FGF19 does not, indicating that polysaccharides other than heparin might enhance FGF19/FGFR signaling. Using a FGFs/FGFR1c signaling-dependent BaF3 cell proliferation assay, we discovered that polyguluronate sulfate (PGS) and its oligosaccharides, PGS12 and PGS25, but not polyguluronate (PG), a natural marine polysaccharide, enhanced FGF19/FGFR1c signaling better than that of heparin based on 3H-thymidine incorporation. Interestingly, PGS6, PGS8, PGS10, PGS12, PGS25, and PGS, but not PG, had comparable FGF1/FGFR1c signal-stimulating activity compared to that of heparin. These results indicated that PGS and its oligosaccharides were excellent FGF1/FGFR1c and FGF19/FGFR1c signaling enhancers at cellular level. Since the inexpensive PGS and PGS oligosaccharides can be absorbed through oral route, these seaweed-derived compounds merit further investigation as novel agents for the treatment of type 2 diabetes through enhancing FGF1/FGFR1c and FGF19/FGFR1c signaling in future.

Heparin-induced thrombocytopenia: real-world issues.

Science.gov (United States)

Linkins, Lori-Ann; Warkentin, Theodore E

2011-09-01

Heparin-induced thrombocytopenia (HIT) is a prothrombotic drug reaction caused by platelet-activating antibodies. HIT sera often activate platelets without needing heparin-such heparin-"independent" platelet activation can be associated with HIT beginning or worsening despite stopping heparin ("delayed-onset HIT"). We address important issues in HIT diagnosis and therapy, using a recent cohort of HIT patients to illustrate influences of heparin type; triggers for HIT investigation; serological features of heparin-independent platelet activation; and treatment. In our cohort of recent HIT cases ( N = 13), low-molecular-weight heparin (dalteparin) was a common causative agent ( N = 8, 62%); most patients were diagnosed after HIT-thrombosis had occurred; and danaparoid was the most frequently selected treatment. Heparin-independent platelet activation was common (7/13 [54%]) and predicted slower platelet count recovery (>1 week) among evaluable patients (5/5 vs 1/6; P = 0.015). In our experience with argatroban-treated patients, HIT-associated consumptive coagulopathy confounds anticoagulant monitoring. Our observations provide guidance on practical aspects of HIT diagnosis and management. Thieme Medical Publishers.

Rivaroxaban-once daily, oral, direct factor Xa inhibition compared with vitamin K antagonism for prevention of stroke and Embolism Trial in Atrial Fibrillation: rationale and design of the ROCKET AF study

DEFF Research Database (Denmark)

NN, NN

2010-01-01

BACKGROUND: Atrial fibrillation (AF), the most common significant cardiac arrhythmia, increases the risk of stroke, particularly in the elderly. Warfarin is effective in reducing stroke risk but is burdensome to patients and is difficult to control. Rivaroxaban is an oral direct factor Xa inhibit...

77 FR 7584 - Draft Guidance for Industry on Heparin for Drug and Medical Device Use; Monitoring Crude Heparin...

Science.gov (United States)

2012-02-13

... strategies to ensure that the heparin supply chain is not contaminated with OSCS or any non- porcine origin... device manufacturers of finished products, and others to the potential risk of crude heparin...) among patients injected with heparin sodium in 2008, FDA identified the contaminant OSCS in heparin API...

78 FR 38058 - Guidance for Industry on Heparin for Drug and Medical Device Use: Monitoring Crude Heparin for...

Science.gov (United States)

2013-06-25

... public health. FDA developed this guidance to alert manufacturers to the risks of crude heparin contaminants and to recommend strategies to ensure that the heparin supply chain is not contaminated with OSCS... heparin sodium in 2008, FDA identified the contaminant OSCS in crude heparin sourced from China. FDA is...

Comparative Transcriptome Profiling of Rice Near-Isogenic Line Carrying Xa23 under Infection of Xanthomonas oryzae pv. oryzae.

Science.gov (United States)

Tariq, Rezwan; Wang, Chunlian; Qin, Tengfei; Xu, Feifei; Tang, Yongchao; Gao, Ying; Ji, Zhiyuan; Zhao, Kaijun

2018-03-02

Bacterial blight, caused by Xanthomonas oryzae pv. oryzae ( Xoo ), is an overwhelming disease in rice-growing regions worldwide. Our previous studies revealed that the executor R gene Xa23 confers broad-spectrum disease resistance to all naturally occurring biotypes of Xoo . In this study, comparative transcriptomic profiling of two near-isogenic lines (NILs), CBB23 (harboring Xa23 ) and JG30 (without Xa23 ), before and after infection of the Xoo strain, PXO99 A , was done by RNA sequencing, to identify genes associated with the resistance. After high throughput sequencing, 1645 differentially expressed genes (DEGs) were identified between CBB23 and JG30 at different time points. Gene Ontlogy (GO) analysis categorized the DEGs into biological process, molecular function, and cellular component. KEGG analysis categorized the DEGs into different pathways, and phenylpropanoid biosynthesis was the most prominent pathway, followed by biosynthesis of plant hormones, flavonoid biosynthesis, and glycolysis/gluconeogenesis. Further analysis led to the identification of differentially expressed transcription factors (TFs) and different kinase responsive genes in CBB23, than that in JG30. Besides TFs and kinase responsive genes, DEGs related to ethylene, jasmonic acid, and secondary metabolites were also identified in both genotypes after PXO99 A infection. The data of DEGs are a precious resource for further clarifying the network of Xa23 -mediated resistance.

Risk factors for epistaxis in patients followed in general practices in Germany.

Science.gov (United States)

Seidel, D U; Jacob, L; Kostev, K; Sesterhenn, A M

2017-12-01

The goal of the present study was to analyze the risk factors for epistaxis in patients followed in general practices in Germany. The current study sample included patients aged 18 years or older who received a first epistaxis diagnosis between January 2012 and December 2016 (index date). Epistaxis patients and controls without epistaxis were matched (1:1) on the basis of age, gender, insurance status and physician. A total of 16,801 patients with epistaxis and 16,801 control subjects were included in this study. Of the subjects, 53.2% were men, and the mean age was 59.6 years (SD=21.2 years). Epistaxis was found to be positively associated with hypertension, obesity, chronic sinusitis, other disorders of the nose and nasal sinuses, anxiety disorder, and adjustment disorder (ORs ranging from 1.13 to 1.44). Epistaxis was also associated with the prescription of vitamin K antagonists, preparations from the heparin group, platelet aggregation inhibitors excluding heparin, direct thrombin inhibitors, direct factor Xa inhibitors, other antithrombotic agents, selective serotonin reuptake inhibitors and nasal steroids (ORs ranging from 1.15 to 3.55). Overall, epistaxis risk is increased by multiple medical and psychiatric disorders. Several antithrombotic and nasal steroid therapies are also associated with this risk.

Effect of buffer at nanoscale molecular recognition interfaces - electrostatic binding of biological polyanions.

Science.gov (United States)

Rodrigo, Ana C; Laurini, Erik; Vieira, Vânia M P; Pricl, Sabrina; Smith, David K

2017-10-19

We investigate the impact of an over-looked component on molecular recognition in water-buffer. The binding of a cationic dye to biological polyanion heparin is shown by isothermal calorimetry to depend on buffer (Tris-HCl > HEPES > PBS). The heparin binding of self-assembled multivalent (SAMul) cationic micelles is even more buffer dependent. Multivalent electrostatic molecular recognition is buffer dependent as a result of competitive interactions between the cationic binding interface and anions present in the buffer.

Heparin and heparin-induced thrombocytopenia

African Journals Online (AJOL)

2007-06-15

Jun 15, 2007 ... Heparin is one of the most widely used drugs. It is used routinely for treatment and thromboprophylaxis in a broad spectrum of conditions including venous thromboembolism, atrial fibrillation, acute coronary syndromes, peripheral vascular disease and to maintain the patency of indwelling catheters and ...

Results of the HepZero study comparing heparin-grafted membrane and standard care show that heparin-grafted dialyzer is safe and easy to use for heparin-free dialysis

NARCIS (Netherlands)

Laville, Maurice; Dorval, Marc; Ros, Joan Fort; Fay, Renaud; Cridlig, Joelle; Nortier, Joelle L.; Juillard, Laurent; Debska-Slizien, Alicja; Lorente, Loreto Fernandez; Thibaudin, Damien; Franssen, Casper; Schulz, Michael; Moureau, Frederique; Loughraieb, Nathalie; Rossignol, Patrick

2014-01-01

Heparin is used to prevent clotting during hemodialysis, but heparin-free hemodialysis is sometimes needed to decrease the risk of bleeding. The HepZero study is a randomized, multicenter international controlled open-label trial comparing no-heparin hemodialysis strategies designed to assess

Coexistence of Antiphospholipid Syndrome and Heparin-Induced Thrombocytopenia in a Patient with Recurrent Venous Thromboembolism

Directory of Open Access Journals (Sweden)

Samuel Adediran

2017-01-01

Full Text Available Heparin-induced thrombocytopenia (HIT is a prothrombotic adverse drug reaction in which heparin forms complexes with platelet factor 4 forming neoantigens that are recognized by autoantibodies. Antiphospholipid syndrome (APS is similar to HIT in that it is mediated by autoantibodies that are also prothrombotic. We present a case of rare coexistence of antiphospholipid antibody syndrome and heparin-induced thrombocytopenia.

Elevated urine levels of heparin-binding protein in children with urinary tract infection.

Science.gov (United States)

Kjölvmark, Charlott; Akesson, Per; Linder, Adam

2012-08-01

Urinary tract infection (UTI) is a common infection diagnosis in children, and efficient diagnosis and treatment are important to avoid serious complications. In this study we investigated whether urinary levels of neutrophil-derived heparin-binding protein (HBP) can be used as a marker of UTI in children. These results were compared to those of dipstick analysis, interleukin-6 (IL-6) analysis in urine, and bacterial culturing. Seventy-eight children aged 0-18 years with fever and/or symptoms indicating UTI were enrolled in a prospective consecutive study. Urine samples were cultured and analyzed with dipstick, and concentrations of HBP and IL-6 were measured. Fifteen patients were classified as having UTI, 30 patients had fever but were diagnosed with a non-urinary tract infection, and 33 patients had neither UTI nor fever. Using a urine HBP (U-HBP) cut-off level of 32 ng/mL, the sensitivity and specificity for detecting UTI were 93.3 and 90.3 %, respectively. Receiver operating characteristic curves demonstrated that U-HBP levels were a higher specificity indicator of UTI than urine white blood cell counts or urine IL-6 levels; they also showed a higher sensitivity than the results of the urine nitrite test. All patients with significant growth of clinically relevant bacteria had elevated U-HBP levels. The results indicate that rapid analysis of U-HBP can provide helpful guidance in the management of children with suspected UTI.

Backbone dynamics of a biologically active human FGF-1 monomer, complexed to a hexasaccharide heparin-analogue, by 15N NMR relaxation methods

International Nuclear Information System (INIS)

Canales-Mayordomo, Angeles; Fayos, Rosa; Angulo, Jesus; Ojeda, Rafael; Martin-Pastor, Manuel; Nieto, Pedro M.; Martin-Lomas, Manuel; Lozano, Rosa; Gimenez-Gallego, Guillermo; Jimenez-Barbero, Jesus

2006-01-01

The binding site and backbone dynamics of a bioactive complex formed by the acidic fibroblast growth factor (FGF-1) and a specifically designed heparin hexasaccharide has been investigated by HSQC and relaxation NMR methods. The comparison of the relaxation data for the free and bound states has allowed showing that the complex is monomeric, and still induces mutagenesis, and that the protein backbone presents reduced motion in different timescale in its bound state, except in certain points that are involved in the interaction with the fibroblast growth factor receptor (FGFR)

Introduction of a rice blight resistance gene, Xa21, into five Chinese rice varieties through an Agrobacterium-mediated system

Institute of Scientific and Technical Information of China (English)

无

2000-01-01

A cloned gene, Xa21 was transferred into five widely-used Chinese rice varieties through an Agrobacterium-mediated system, and over 110 independent transgenic lines were obtained. PCR and Southern analysis of transgenic plants revealed the integration of the whole Xa21 gene into the host genomes. The integrated Xa21 gene was stably inherited, and segregated in a 3∶1 ratio in the selfed T1 generation when one copy of the gene was integrated in the transformants. Inoculation tests displayed that transgenic T0 plants and Xa21 PCR-positive T1 plants were highly resistant to bacterial blight disease. The selected Xa21 homozygous resistant transgenic lines with desirable qualities may be propagated as new varieties or utilized in hybrid rice breeding.

Introduction of a rice blight resistance gene, Xa21, into five Chinese rice varieties through an Agrobacterium -mediated system

Institute of Scientific and Technical Information of China (English)

翟文学; 李晓兵; 田文忠; 周永力; 潘学彪; 曹守云; 赵显峰; 赵彬; 章琦; 朱立煌

2000-01-01

A cloned gene, Xa21 was transferred into five widely-used Chinese rice varieties through an Agrobacterium-mediated system, and over 110 independent transgenic lines were obtained. PCR and Southern analysis of transgenic plants revealed the integration of the whole Xa21 gene into the host genomes. The integrated Xa21 gene was stably inherited, and segregated in a 3 : 1 ratio in the selfed T1 generation when one copy of the gene was integrated in the transfor-mants. Inoculation tests displayed that transgenic T0 plants and Xa21 PCR-positive T1 plants were highly resistant to bacterial blight disease. The selected Xa21 homozygous resistant transgenic lines with desirable qualities may be propagated as new varieties or utilized in hybrid rice breeding.

Role of Heparan Sulfate in Cellular Infection of Integrin-Binding Coxsackievirus A9 and Human Parechovirus 1 Isolates.

Directory of Open Access Journals (Sweden)

Pirjo Merilahti

Full Text Available Heparan sulfate/heparin class of proteoglycans (HSPG have been shown to function in cellular attachment and infection of numerous viruses including picornaviruses. Coxsackievirus A9 (CV-A9 and human parechovirus 1 (HPeV-1 are integrin-binding members in the family Picornaviridae. CV-A9 Griggs and HPeV-1 Harris (prototype strains have been reported not to bind to heparin, but it was recently shown that some CV-A9 isolates interact with heparin in vitro via VP1 protein with a specific T132R/K mutation. We found that the infectivity of both CV-A9 Griggs and HPeV-1 Harris was reduced by sodium chlorate and heparinase suggestive of HSPG interactions. We analyzed the T132 site in fifty-four (54 CV-A9 clinical isolates and found that only one of them possessed T132/R mutation while the other nine (9 had T132K. We then treated CV-A9 Griggs and HPeV-1 Harris and eight CV-A9 and six HPeV-1 clinical isolates with heparin and protamine. Although infectivity of Griggs strain was slightly reduced (by 25%, heparin treatment did not affect the infectivity of the CV-A9 isolates that do not possess the T132R/K mutation, which is in line with the previous findings. Some of the HPeV-1 isolates were also affected by heparin treatment, which suggested that there may be a specific heparin binding site in HPeV-1. In contrast, protamine (a specific inhibitor of heparin completely inhibited the infection of both prototypes and clinical CV-A9 and HPeV-1 isolates. We conclude that T132R/K mutation has a role in heparin binding of CV-A9, but we also show data, which suggest that there are other HSPG binding sites in CV-A9. In all, we suggest that HSPGs play a general role in both CV-A9 and HPeV-1 infections.

Heparin/heparan sulfates bind to and modulate neuronal L-type (Cav1.2) voltage-dependent Ca²⁺ channels

DEFF Research Database (Denmark)

Garau, Gianpiero; Magotti, Paola; Heine, Martin

2015-01-01

Our previous studies revealed that L-type voltage-dependent Ca2+ channels (Cav1.2 L-VDCCs) are modulated by the neural extracellular matrix backbone, polyanionic glycan hyaluronic acid. Here we used isothermal titration calorimetry and screened a set of peptides derived from the extracellular......M), integrating their enthalpic and entropic binding contributions. Interaction between heparin and recombinant as well as native full-length neuronal Cav1.2α1 channels was confirmed using the heparin–agarose pull down assay. Whole cell patch clamp recordings in HEK293 cells transfected with neuronal Cav1.......2 channels revealed that enzymatic digestion of highly sulfated heparan sulfates with heparinase 1 affects neither voltage-dependence of channel activation nor the level of steady state inactivation, but did speed up channel inactivation. Treatment of hippocampal cultures with heparinase 1 reduced the firing...

Severe heparin osteoporosis in pregnancy.

OpenAIRE

Griffiths, H. T.; Liu, D. T.

1984-01-01

A case of severe osteoporosis following administration of low dose subcutaneous heparin in pregnancy is reported. Possible reasons for the condition are suggested which caution against the indiscriminate use of subcutaneous heparin in pregnancy.

Nuclear androgen receptors in human prostatic tissue. Extraction with heparin and estimation of the number of binding sites with different methods

International Nuclear Information System (INIS)

Foekens, J.A.; Bolt-de Vries, J.; Mulder, E.; Blankenstein, M.A.; Schroeder, F.H.; Molen, H.J. van der

1981-01-01

A procedure for the estimation of nuclear androgen receptors in benign prostatic hyperplastic tissue is described, which employs extraction of receptors from nuclei with buffers containing heparin. Extraction of a nuclear pellet with a heparin-containing (1 g/l) buffer appeared to have definite advantages over 0.4 mol/l KCl extraction. Heparin appeared to be twice as efficient in extracting androgen receptors. In addition aggregated receptor proteins, formed after storage at -80 0 C, were partly deaggregated by heparin. Specific isolation of the androgen receptor was performed using either agar gel electrophoresis, protamine sulphate precipitation or LH-20 gel filtration. A comparison was made between the amounts of estimated receptors with these different techniques. Protamine sulphate precipitation resulted in the highest estimates of receptor-bound 5α-[ 3 H]dihydrotestosterone ( 3 H-DHT). Treatment of the labelled nuclear extracts with a charcoal suspension prior to the receptor assay resulted in lower amounts of estimated androgen receptors. A method for routine evaluation of nuclear androgen receptors in prostatic tissue has been evaluated, which involves extraction of nuclear pellets with a heparin-containing (1 g/l) buffer, exchange labelling of the nuclear extracts for 20 h at 10 0 C and quantification of the receptors with protamine sulphate precipitation. (Auth.)

Backbone dynamics of a biologically active human FGF-1 monomer, complexed to a hexasaccharide heparin-analogue, by {sup 15}N NMR relaxation methods

Energy Technology Data Exchange (ETDEWEB)

Canales-Mayordomo, Angeles; Fayos, Rosa [Centro de Investigaciones Biologicas, CSIC, Departamento de Estructura y Funcion de Proteinas (Spain); Angulo, Jesus; Ojeda, Rafael [Instituto de Investigaciones Quimicas, CSIC, Grupo de Carbohidratos (Spain); Martin-Pastor, Manuel [Unidad de RM y Unidad de RMN de Biomoleculas Asociada al CSIC, Laboratorio de Estructura e Estructura de Biomoleculas Jose Carracido (Spain); Nieto, Pedro M.; Martin-Lomas, Manuel [Instituto de Investigaciones Quimicas, CSIC, Grupo de Carbohidratos (Spain); Lozano, Rosa; Gimenez-Gallego, Guillermo; Jimenez-Barbero, Jesus [Centro de Investigaciones Biologicas, CSIC, Departamento de Estructura y Funcion de Proteinas (Spain)], E-mail: jjbarbero@cib.csic.es

2006-08-15

The binding site and backbone dynamics of a bioactive complex formed by the acidic fibroblast growth factor (FGF-1) and a specifically designed heparin hexasaccharide has been investigated by HSQC and relaxation NMR methods. The comparison of the relaxation data for the free and bound states has allowed showing that the complex is monomeric, and still induces mutagenesis, and that the protein backbone presents reduced motion in different timescale in its bound state, except in certain points that are involved in the interaction with the fibroblast growth factor receptor (FGFR)

Assessment of anti-factor Xa activity of enoxaparin for venous thromboembolism prophylaxis in morbidly obese surgical patients

Directory of Open Access Journals (Sweden)

Nouf Al Otaib

2017-01-01

Conclusions: Weight-based enoxaparin dose led to the anticipated peak anti-Xa levels (0.2–0.6 IU/mL in most of the morbidly obese study patients undergoing surgery without any evidence of major side effects. The weight-based dosing of enoxaparin was also effective in preventing VTE in all patients. Although these results are promising, further comparative trials are needed in the setting of morbidly obese surgical patients.

Allergic anaphylaxis due to subcutaneously injected heparin

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Anders Diana

2013-01-01

Full Text Available Abstract Heparins are one of the most used class of anticoagulants in daily clinical practice. Despite their widespread application immune-mediated hypersensitivity reactions to heparins are rare. Among these, the delayed-type reactions to s.c. injected heparins are well-known usually presenting as circumscribed eczematous plaques at the injection sites. In contrast, potentially life-threatening systemic immediate-type anaphylactic reactions to heparins are extremely rare. Recently, some cases of non-allergic anaphylaxis could be attributed to undesirable heparin contaminants. A 43-year-old patient developed severe anaphylaxis symptoms within 5–10 minutes after s.c. injection of enoxaparin. Titrated skin prick testing with wheal and flare responses up to an enoxaparin dilution of 1:10.000 indicated a probable allergic mechanism of the enoxaparin-induced anaphylaxis. The basophil activation test as an additional in-vitro test method was negative. Furthermore, skin prick testing showed rather broad cross-reactivity among different heparin preparations tested. In the presented case, history, symptoms, and results of skin testing strongly suggested an IgE-mediated allergic hypersensitivity against different heparins. Therefore, as safe alternative anticoagulants the patient could receive beneath coumarins the hirudins or direct thrombin inhibitors. Because these compounds have a completely different molecular structure compared with the heparin-polysaccharides.

Bioassay-directed fractionation of a blood coagulation factor Xa inhibitor, betulinic acid from Lycopus lucidus

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Tan Yin-Feng

2018-03-01

Full Text Available Thrombosis is a major cause of morbidity and mortality worldwide and plays a pivotal role in the pathogenesis of several cardiovascular disorders, including acute coronary syndrome, unstable angina, myocardial infarction, sudden cardiac death, peripheral arterial occlusion, ischemic stroke, deep-vein thrombosis, and pulmonary embolism. Anticoagulants, antiplatelet agents, and fibrinolytics can reduce the risks of these clinical events. Especially, the blood coagulation factor Xa (FXa inhibitor is a proven anticoagulant. Promoting blood circulation, using traditional Chinese medicine (TCM, for the treatment of these diseases has been safely used for thousands of years in clinical practice. Therefore, highly safe and effective anticoagulant ingredients, including FXa inhibitors, could be found in TCM for activating the blood circulation. One FXa inhibitor, a pentacyclic triterpene (compound 1, betulinic acid characterized by IR, MS and NMR analyses, was isolated from the ethyl acetate fraction of Lycopus lucidus by bioassay-directed fractionation. Compound 1 exhibited an inhibitory effect on FXa with IC50 25.05 μmol/L and reduced the thrombus weight in an animal model at 25-100 mg/kg. These results indicate that betulinic acid could be the potential for anticoagulant therapy.

Interference of heparin in carcinoembryonic antigen radioimmunoassays

International Nuclear Information System (INIS)

Wu, J.T.

1983-01-01

A false Roche carcinoembryonic antigen (CEA) activity could be detected in all commercial and noncommercial heparin preparations examined. The possibility of 'due to contamination' has been ruled out. Using the Roche procedure, heparin solutions, in the absence of CEA, gave positive CEA activity; on the other hand, no CEA activity was detected in solutions containing only heparin when the Abbott Kit was used. When heparin was present in specimens containing CEA, the Abbott Kit underestimated the CEA activity, whereas the Roche Kit gave false elevated values. However, the negative effect of heparin could be reduced by heat treatment in the presence of plasma proteins. (Auth.)

Degranulating mast cells in fibrotic regions of human tumors and evidence that mast cell heparin interferes with the growth of tumor cells through a mechanism involving fibroblasts

International Nuclear Information System (INIS)

Samoszuk, Michael; Kanakubo, Emi; Chan, John K

2005-01-01

The purpose of this study was to test the hypothesis that mast cells that are present in fibrotic regions of cancer can suppress the growth of tumor cells through an indirect mechanism involving peri-tumoral fibroblasts. We first immunostained a wide variety of human cancers for the presence of degranulated mast cells. In a subsequent series of controlled in vitro experiments, we then co-cultured UACC-812 human breast cancer cells with normal fibroblasts in the presence or absence of different combinations and doses of mast cell tryptase, mast cell heparin, a lysate of the human mast cell line HMC-1, and fibroblast growth factor-7 (FGF-7), a powerful, heparin-binding growth factor for breast epithelial cells. Degranulating mast cells were localized predominantly in the fibrous tissue of every case of breast cancer, head and neck cancer, lung cancer, ovarian cancer, non-Hodgkin's lymphoma, and Hodgkin's disease that we examined. Mast cell tryptase and HMC-1 lysate had no significant effect on the clonogenic growth of cancer cells co-cultured with fibroblasts. By contrast, mast cell heparin at multiple doses significantly reduced the size and number of colonies of tumor cells co-cultured with fibroblasts, especially in the presence of FGF-7. Neither heparin nor FGF-7, individually or in combination, produced any significant effect on the clonogenic growth of breast cancer cells cultured without fibroblasts. Degranulating mast cells are restricted to peri-tumoral fibrous tissue, and mast cell heparin is a powerful inhibitor of clonogenic growth of tumor cells co-cultured with fibroblasts. These results may help to explain the well-known ability of heparin to inhibit the growth of primary and metastatic tumors

Degranulating mast cells in fibrotic regions of human tumors and evidence that mast cell heparin interferes with the growth of tumor cells through a mechanism involving fibroblasts

Directory of Open Access Journals (Sweden)

Kanakubo Emi

2005-09-01

Full Text Available Abstract Background The purpose of this study was to test the hypothesis that mast cells that are present in fibrotic regions of cancer can suppress the growth of tumor cells through an indirect mechanism involving peri-tumoral fibroblasts. Methods We first immunostained a wide variety of human cancers for the presence of degranulated mast cells. In a subsequent series of controlled in vitro experiments, we then co-cultured UACC-812 human breast cancer cells with normal fibroblasts in the presence or absence of different combinations and doses of mast cell tryptase, mast cell heparin, a lysate of the human mast cell line HMC-1, and fibroblast growth factor-7 (FGF-7, a powerful, heparin-binding growth factor for breast epithelial cells. Results Degranulating mast cells were localized predominantly in the fibrous tissue of every case of breast cancer, head and neck cancer, lung cancer, ovarian cancer, non-Hodgkin's lymphoma, and Hodgkin's disease that we examined. Mast cell tryptase and HMC-1 lysate had no significant effect on the clonogenic growth of cancer cells co-cultured with fibroblasts. By contrast, mast cell heparin at multiple doses significantly reduced the size and number of colonies of tumor cells co-cultured with fibroblasts, especially in the presence of FGF-7. Neither heparin nor FGF-7, individually or in combination, produced any significant effect on the clonogenic growth of breast cancer cells cultured without fibroblasts. Conclusion Degranulating mast cells are restricted to peri-tumoral fibrous tissue, and mast cell heparin is a powerful inhibitor of clonogenic growth of tumor cells co-cultured with fibroblasts. These results may help to explain the well-known ability of heparin to inhibit the growth of primary and metastatic tumors.

Thrombin and factor Xa link the coagulation system with liver fibrosis.

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Dhar, Ameet; Sadiq, Fouzia; Anstee, Quentin M; Levene, Adam P; Goldin, Robert D; Thursz, Mark R

2018-05-08

Thrombin activates hepatic stellate cells via protease-activated receptor-1. The role of Factor Xa (FXa) in hepatic fibrosis has not been elucidated. We aimed to evaluate the impact of FXa and thrombin in vitro on stellate cells and their respective inhibition in vivo using a rodent model of hepatic fibrosis. HSC-LX2 cells were incubated with FXa and/or thrombin in cell culture, stained for αSMA and relative gene expression and gel contraction calculated. C57BL/6 J mice were administered thioacetamide (TAA) for 8 weeks with Rivaroxaban (n = 15) or Dabigatran (n = 15). Control animals received TAA alone (n = 15). Fibrosis was scored and quantified using digital image analysis and hepatic tissue hydroxyproline estimated. Stellate cells treated with FXa and thrombin demonstrated upregulation of procollagen, TGF-beta, αSMA and significant cell contraction (43.48%+/- 4.12) compared to culturing with FXa or thrombin alone (26.90%+/- 8.90, p = 0.02; 13.1%+/- 9.84, p < 0.001). Mean fibrosis score, percentage area of fibrosis and hepatic hydroxyproline content (2.46 vs 4.08, p = 0.008; 2.02% vs 3.76%, p = 0.012; 276.0 vs 651.3, p = 0.0001) were significantly reduced in mice treated with the FXa inhibitor compared to control mice. FXa inhibition was significantly more effective than thrombin inhibition in reducing percentage area of fibrosis and hepatic hydroxyproline content (2.02% vs 3.70%,p = 0.031; 276.0 vs 413.1,p = 0.001). FXa promotes stellate cell contractility and activation. Early inhibition of coagulation using a FXa inhibitor significantly reduces TAA induced murine liver fibrosis and may be a viable treatment for liver fibrosis in patients.

Interaction of the amyloid precursor protein-like protein 1 (APLP1) E2 domain with heparan sulfate involves two distinct binding modes

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Dahms, Sven O., E-mail: sdahms@fli-leibniz.de [Leibniz Institute for Age Research (FLI), Beutenbergstrasse 11, 07745 Jena (Germany); Mayer, Magnus C. [Freie Universität Berlin, Thielallee 63, 14195 Berlin (Germany); Miltenyi Biotec GmbH, Robert-Koch-Strasse 1, 17166 Teterow (Germany); Roeser, Dirk [Leibniz Institute for Age Research (FLI), Beutenbergstrasse 11, 07745 Jena (Germany); Multhaup, Gerd [McGill University Montreal, Montreal, Quebec H3G 1Y6 (Canada); Than, Manuel E., E-mail: sdahms@fli-leibniz.de [Leibniz Institute for Age Research (FLI), Beutenbergstrasse 11, 07745 Jena (Germany)

2015-03-01

Two X-ray structures of APLP1 E2 with and without a heparin dodecasaccharide are presented, revealing two distinct binding modes of the protein to heparan sulfate. The data provide a mechanistic explanation of how APP-like proteins bind to heparan sulfates and how they specifically recognize nonreducing structures of heparan sulfates. Beyond the pathology of Alzheimer’s disease, the members of the amyloid precursor protein (APP) family are essential for neuronal development and cell homeostasis in mammals. APP and its paralogues APP-like protein 1 (APLP1) and APP-like protein 2 (APLP2) contain the highly conserved heparan sulfate (HS) binding domain E2, which effects various (patho)physiological functions. Here, two crystal structures of the E2 domain of APLP1 are presented in the apo form and in complex with a heparin dodecasaccharide at 2.5 Å resolution. The apo structure of APLP1 E2 revealed an unfolded and hence flexible N-terminal helix αA. The (APLP1 E2){sub 2}–(heparin){sub 2} complex structure revealed two distinct binding modes, with APLP1 E2 explicitly recognizing the heparin terminus but also interacting with a continuous heparin chain. The latter only requires a certain register of the sugar moieties that fits to a positively charged surface patch and contributes to the general heparin-binding capability of APP-family proteins. Terminal binding of APLP1 E2 to heparin specifically involves a structure of the nonreducing end that is very similar to heparanase-processed HS chains. These data reveal a conserved mechanism for the binding of APP-family proteins to HS and imply a specific regulatory role of HS modifications in the biology of APP and APP-like proteins.

M2 macrophages induce ovarian cancer cell proliferation via a heparin binding epidermal growth factor/matrix metalloproteinase 9 intercellular feedback loop.

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Carroll, Molly J; Kapur, Arvinder; Felder, Mildred; Patankar, Manish S; Kreeger, Pamela K

2016-12-27

In ovarian cancer, a high ratio of anti-inflammatory M2 to pro-inflammatory M1 macrophages correlates with poor patient prognosis. The mechanisms driving poor tumor outcome as a result of the presence of M2 macrophages in the tumor microenvironment remain unclear and are challenging to study with current techniques. Therefore, in this study we utilized a micro-culture device previously developed by our lab to model concentrated paracrine signaling in order to address our hypothesis that interactions between M2 macrophages and ovarian cancer cells induce tumor cell proliferation. Using the micro-culture device, we determined that co-culture with M2-differentiated primary macrophages or THP-1 increased OVCA433 proliferation by 10-12%. This effect was eliminated with epidermal growth factor receptor (EGFR) or heparin-bound epidermal growth factor (HB-EGF) neutralizing antibodies and HBEGF expression in peripheral blood mononuclear cells from ovarian cancer patients was 9-fold higher than healthy individuals, suggesting a role for HB-EGF in tumor progression. However, addition of HB-EGF at levels secreted by macrophages or macrophage-conditioned media did not induce proliferation to the same extent, indicating a role for other factors in this process. Matrix metalloproteinase-9, MMP-9, which cleaves membrane-bound HB-EGF, was elevated in co-culture and its inhibition decreased proliferation. Utilizing inhibitors and siRNA against MMP9 in each population, we determined that macrophage-secreted MMP-9 released HB-EGF from macrophages, which increased MMP9 in OVCA433, resulting in a positive feedback loop to drive HB-EGF release and increase proliferation in co-culture. Identification of multi-cellular interactions such as this may provide insight into how to most effectively control ovarian cancer progression.

Tranexamic Acid Failed to Reverse the Anticoagulant Effect and Bleeding by an Oral Direct Factor Xa Inhibitor Edoxaban.

Science.gov (United States)

Honda, Yuko; Furugohri, Taketoshi; Morishima, Yoshiyuki

2018-01-01

Agents to reverse the anticoagulant effect of edoxaban, an oral direct factor Xa inhibitor, would be desirable in emergency situations. The aim of this study is to determine the effect of tranexamic acid, an antifibrinolytic agent, on the anticoagulant activity and bleeding by edoxaban in rats. A supratherapeutic dose of edoxaban (3 mg/kg) was intravenously administered to rats. Three minutes after dosing, tranexamic acid (100 mg/kg) was given intravenously. Bleeding was induced by making an incision with a blade on the planta 8 min after edoxaban injection and bleeding time was measured. Prothrombin time (PT) and clot lysis were examined. A supratherapeutic dose of edoxaban significantly prolonged PT and bleeding time. Tranexamic acid did not affect PT or bleeding time prolonged by edoxaban, although tranexamic acid significantly inhibited clot lysis in rat plasma. An antifibrinolytic agent tranexamic acid failed to reverse the anticoagulant effect and bleeding by edoxaban in rats. © 2017 S. Karger AG, Basel.

A new peptide (Ruviprase) purified from the venom of Daboia russelii russelii shows potent anticoagulant activity via non-enzymatic inhibition of thrombin and factor Xa.

Science.gov (United States)

Thakur, Rupamoni; Kumar, Ashok; Bose, Biplab; Panda, Dulal; Saikia, Debashree; Chattopadhyay, Pronobesh; Mukherjee, Ashis K

2014-10-01

Compounds showing dual inhibition of thrombin and factor Xa (FXa) are the subject of great interest owing to their broader specificity for effective anticoagulation therapy against cardiovascular disorders. This is the first report on the functional characterization and assessment of therapeutic potential of a 4423.6 Da inhibitory peptide (Ruviprase) purified from Daboia russelii russelii venom. The secondary structure of Ruviprase is composed of α-helices (61.9%) and random coils (38.1%). The partial N-terminal sequence (E(1)-V(2)-X(3)-W(4)-W(5)-W(6)-A(7)-Q(8)-L(9)-S(10)) of Ruviprase demonstrated significant similarity (80.0%) with an internal sequence of apoptosis-stimulating protein reported from the venom of Ophiophagus hannah and Python bivittatus; albeit Ruviprase did not show sequence similarity with existing thrombin/FXa inhibitors, suggesting its uniqueness. Ruviprase demonstrated a potent in vitro anticoagulant property and inhibited both thrombin and FXa following slow binding kinetics. Ruviprase inhibited thrombin by binding to its active site via an uncompetitive mechanism with a Ki value and dissociation constant (KD) of 0.42 μM and 0.46 μM, respectively. Conversely, Ruviprase demonstrated mixed inhibition (Ki = 0.16 μM) of FXa towards its physiological substrate prothrombin. Furthermore, the biological properties of Ruviprase could not be neutralized by commercial polyvalent or monovalent antivenom. Ruviprase at a dose of 2.0 mg/kg was non-toxic and showed potent in vivo anticoagulant activity after 6 h of intraperitoneal treatment in mice. Because of the potent anticoagulant property as well as non-toxic nature of Ruviprase, the possible application of the peptide as an antithrombotic agent for combating thrombosis-associated ailments appears promising. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Structural and functional characterization of the interaction between cyclophilin B and a heparin-derived oligosaccharide.

Science.gov (United States)

Hanoulle, Xavier; Melchior, Aurélie; Sibille, Nathalie; Parent, Benjamin; Denys, Agnès; Wieruszeski, Jean-Michel; Horvath, Dragos; Allain, Fabrice; Lippens, Guy; Landrieu, Isabelle

2007-11-23

The chemotaxis and integrin-mediated adhesion of T lymphocytes triggered by secreted cyclophilin B (CypB) depend on interactions with both cell surface heparan sulfate proteoglycans (HSPG) and the extracellular domain of the CD147 membrane receptor. Here, we use NMR spectroscopy to characterize the interaction of CypB with heparin-derived oligosaccharides. Chemical shift perturbation experiments allowed the precise definition of the heparan sulfate (HS) binding site of CypB. The N-terminal extremity of CypB, which contains a consensus sequence for heparin-binding proteins was modeled on the basis of our experimental NMR data. Because the HS binding site extends toward the CypB catalytic pocket, we measured its peptidyl-prolyl cis-trans isomerase (PPIase) activity in the absence or presence of a HS oligosaccharide toward a CD147-derived peptide. We report the first direct evidence that CypB is enzymatically active on CD147, as it is able to accelerate the cis/trans isomerization of the Asp(179)-Pro(180) bond in a CD147-derived peptide. However, HS binding has no significant influence on this PPIase activity. We thus conclude that the glycanic moiety of HSPG serves as anchor for CypB at the cell surface, and that the signal could be transduced by CypB via its PPIase activity toward CD147.

Preparation and characterization of microspheres of albumin-heparin conjugates

NARCIS (Netherlands)

Kwon, Glen S.; Bae, You Han; Kim, Sung Wan; Cremers, Harry; Cremers, H.F.M.; Feijen, Jan

1991-01-01

Albumin-heparin microspheres have been prepared as a new drug carrier. A soluble albumin-heparin conjugate was synthesized by forming amide bonds between human serum albumin and heparin. After purification the albumin-heparin conjugate was crosslinked in a water-in-oil emulsion to form

3D/3D registration of coronary CTA and biplane XA reconstructions for improved image guidance

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Dibildox, Gerardo, E-mail: g.dibildox@erasmusmc.nl; Baka, Nora; Walsum, Theo van [Biomedical Imaging Group Rotterdam, Departments of Radiology and Medical Informatics, Erasmus Medical Center, 3015 GE Rotterdam (Netherlands); Punt, Mark; Aben, Jean-Paul [Pie Medical Imaging, 6227 AJ Maastricht (Netherlands); Schultz, Carl [Department of Cardiology, Erasmus Medical Center, 3015 GE Rotterdam (Netherlands); Niessen, Wiro [Quantitative Imaging Group, Faculty of Applied Sciences, Delft University of Technology, 2628 CJ Delft, The Netherlands and Biomedical Imaging Group Rotterdam, Departments of Radiology and Medical Informatics, Erasmus Medical Center, 3015 GE Rotterdam (Netherlands)

2014-09-15

Purpose: The authors aim to improve image guidance during percutaneous coronary interventions of chronic total occlusions (CTO) by providing information obtained from computed tomography angiography (CTA) to the cardiac interventionist. To this end, the authors investigate a method to register a 3D CTA model to biplane reconstructions. Methods: The authors developed a method for registering preoperative coronary CTA with intraoperative biplane x-ray angiography (XA) images via 3D models of the coronary arteries. The models are extracted from the CTA and biplane XA images, and are temporally aligned based on CTA reconstruction phase and XA ECG signals. Rigid spatial alignment is achieved with a robust probabilistic point set registration approach using Gaussian mixture models (GMMs). This approach is extended by including orientation in the Gaussian mixtures and by weighting bifurcation points. The method is evaluated on retrospectively acquired coronary CTA datasets of 23 CTO patients for which biplane XA images are available. Results: The Gaussian mixture model approach achieved a median registration accuracy of 1.7 mm. The extended GMM approach including orientation was not significantly different (P > 0.1) but did improve robustness with regards to the initialization of the 3D models. Conclusions: The authors demonstrated that the GMM approach can effectively be applied to register CTA to biplane XA images for the purpose of improving image guidance in percutaneous coronary interventions.

Prospective multicentre cohort study of heparin-induced thrombocytopenia in acute ischaemic stroke patients

Science.gov (United States)

Kawano, Hiroyuki; Yamamoto, Haruko; Miyata, Shigeki; Izumi, Manabu; Hirano, Teruyuki; Toratani, Naomi; Kakutani, Isami; Sheppard, Jo-Ann I; Warkentin, Theodore E; Kada, Akiko; Sato, Shoichiro; Okamoto, Sadahisa; Nagatsuka, Kazuyuki; Naritomi, Hiroaki; Toyoda, Kazunori; Uchino, Makoto; Minematsu, Kazuo

2011-01-01

Acute ischaemic stroke patients sometimes receive heparin for treatment and/or prophylaxis of thromboembolic complications. This study was designed to elucidate the incidence and clinical features of heparin-induced thrombocytopenia (HIT) in acute stroke patients treated with heparin. We conducted a prospective multicentre cohort study of 267 patients who were admitted to three stroke centres within 7 d after stroke onset. We examined clinical data until discharge and collected blood samples on days 1 and 14 of hospitalization to test anti-platelet factor 4/heparin antibodies (anti-PF4/H Abs) using an enzyme-linked immunosorbent assay (ELISA); platelet-activating antibodies were identified by serotonin-release assay (SRA). Patients with a 4Ts score ≥4 points, positive-ELISA, and positive-SRA were diagnosed as definite HIT. Heparin was administered to 172 patients (64·4%: heparin group). Anti-PF4/H Abs were detected by ELISA in 22 cases (12·8%) in the heparin group. Seven patients had 4Ts ≥ 4 points. Among them, three patients (1·7% overall) were also positive by both ELISA and SRA. National Institutes of Health Stroke Scale score on admission was high (range, 16–23) and in-hospital mortality was very high (66·7%) in definite HIT patients. In this study, the incidence of definite HIT in acute ischaemic stroke patients treated with heparin was 1·7% (95% confidence interval: 0·4–5·0). The clinical severity and outcome of definite HIT were unfavourable. PMID:21671895

How to give a heparin shot

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... you put the injection. Storing Your Heparin and Supplies Ask your pharmacist how to store your heparin ... M. is also a founding member of Hi-Ethics and subscribes to the principles of the Health ...

Laboratory tests for identification or exclusion of heparin induced thrombocytopenia: HIT or miss?

Science.gov (United States)

Favaloro, Emmanuel J

2018-02-01

Heparin induced thrombocytopenia (HIT) is a potentially fatal condition that arises subsequent to formation of antibodies against complexes containing heparin, usually platelet-factor 4-heparin ("anti-PF4-heparin"). Assessment for HIT involves both clinical evaluation and, if indicated, laboratory testing for confirmation or exclusion, typically using an initial immunological assay ("screening"), and only if positive, a secondary functional assay for confirmation. Many different immunological and functional assays have been developed. The most common contemporary immunological assays comprise enzyme-linked immunosorbent assay [ELISA], chemiluminescence, lateral flow, and particle gel techniques. The most common functional assays measure platelet aggregation or platelet activation events (e.g., serotonin release assay; heparin-induced platelet activation (HIPA); flow cytometry). All assays have some sensitivity and specificity to HIT antibodies, but differ in terms of relative sensitivity and specificity for pathological HIT, as well as false negative and false positive error rate. This brief article overviews the different available laboratory methods, as well as providing a suggested approach to diagnosis or exclusion of HIT. © 2017 Wiley Periodicals, Inc.

Acid-citrate-dextrose compared with heparin in the preparation of in vivo/in vitro technetium-99m red blood cells

International Nuclear Information System (INIS)

Porter, W.C.; Dees, S.M.; Freitas, J.E.; Dworkin, H.J.

1983-01-01

Red blood cells labeled in vivo/in vitro with Tc-99m (Tc-99m RBC) were prepared in a series of 21 patients and two normal volunteers. In each subject both heparin and acid-citrate-dextrose (ACD) solutions were used to label tandem blood samples. The immediate preinjection binding efficiency (BE) was then determined. In each of the 23 studies, the ACD preparation yielded superior BE. The average BE was 93.47% (+/- 3.78) with ACD and 87.23% (+/- 4.29) with heparin. With the ACD method the effect of carrier Tc-99 may be as great as a 24% reduction in BE observed when initial eluates from long-ingrowth-time generators were used. Improved image quality with minimal renal and urinary-bladder activity results with ACD labeling. It is concluded that the use of ACD results in superior RBC labeling with less nontarget activity relative to heparin and is preferred over heparin for preparing in vivo/in vitro Tc-99m RBC

Endogenous heparin levels in the controlled asthmatic patient ...

African Journals Online (AJOL)

Background. Since heparin possesses anti-inflammatory properties, it is hypothesised that asthmatic patients have decreased levels of circulating heparin compared with healthy individuals. Design. We compared endogenous heparin levels in controlled asthmatic patients (53 adults) from the Asthma Clinic at ...

[Heparin-induced thrombocytopenia developed during the acute phase after left upper lobectomy for lung cancer].

Science.gov (United States)

Mitomo, Hideki; Miyamoto, Akira; Tabata, Toshiharu; Sugawara, Takafumi; Yabuki, Hiroshi; Fujimura, Shigefumi

2014-12-01

Heparin-induced thrombocytopenia (HIT) is a serious adverse effect of heparin administration. This must not be rarely encountered but is not often reported in Japan compared to Western countries. A 68-year-old woman underwent left upper lobectomy for lung cancer. Low-dose unfractionated heparin was administrated to prevent thromboembolism after the operation. Two days later, sudden dyspnea appeared and ultracardiosonography showing an extensive thromboembolus from the main trunk to both main branches of pulmonary artery indicated pulmonary embolization. After the establishment of percutaneous cardiopulmonary support (PCPS) support, the embolus was removed by emergent open heart surgery. However, despite further unfractionated heparin administration following embolization surgery, other thrombus was identified in both the bi-lateral internal jagular veins and inferior vena cava by ultrasonography and contrast computed tomography( CT). Her platelet count was decreased gradually despite platelet transfusion. Plate factor 4( PF4) antibody against heparin in her blood examination was found, and HIT II was diagnosed. Discontinuation of unfractionated heparin and administration of antithrombin agent improved platelet count, and no additional embolization was identified.

Heparin increases food intake through AgRP neurons

Science.gov (United States)

Although the widely used anticoagulant drug heparin has been shown to have many other biological functions independent of its anticoagulant role, its effects on energy homeostasis are unknown. Here, we demonstrate that heparin level is negatively associated with nutritional states and that heparin t...

Protein interactions with quaternized chitosan/heparin multilayers

Czech Academy of Sciences Publication Activity Database

Kumorek, Marta M.; Kubies, Dana; Riedel, Tomáš

2016-01-01

Roč. 65, Suppl. 2 (2016), S253-S261 ISSN 0862-8408 R&D Projects: GA MŠk(CZ) LQ1604 Institutional support: RVO:61389013 Keywords : heparin * chitosan * protein Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.461, year: 2016 http://www.biomed.cas.cz/physiolres/pdf/65%20Suppl%202/65_S253.pdf

Bilateral adrenal haemorrhage associated with heparin-induced thrombocytopaenia during treatment of Fournier gangrene.

Science.gov (United States)

Tattersall, Timothy Lee; Thangasamy, Isaac A; Reynolds, Jamie

2014-10-14

We present a case of bilateral adrenal haemorrhage (BAH) associated with heparin-induced thrombocytopaenia (HIT) in a 61-year-old man admitted to hospital for the treatment of Fournier's gangrene. He presented to hospital with scrotal swelling and fever, and developed spreading erythaema and a gangrenous scrotum. His scrotum was surgically debrided and intravenous broad-spectrum antibiotics were administered. Unfractionated heparin was given postoperatively for venous thromboembolism prophylaxis. The patient deteriorated clinically 8-11 days postoperatively with delirium, chest pain and severe hypertension followed by hypotension and thrombocytopaenia. Abdominal CT scan revealed bilateral adrenal haemorrhage. Antibodies to the heparin-platelet factor 4 complex were present. HIT-associated BAH was diagnosed and heparin was discontinued. Intravenous bivalirudin and hydrocortisone were started, with rapid improvement in clinical status. BAH is a rare complication of HIT and should be considered in the postoperative patient with unexplained clinical deterioration. 2014 BMJ Publishing Group Ltd.

Sterilization of heparinized cuprophan hemodialysis membranes

NARCIS (Netherlands)

ten Hoopen, Hermina W.M.; Hinrichs, W.L.J.; Hinrichs, W.L.J.; Engbers, G.H.M.; Feijen, Jan

1996-01-01

The effects of sterilization of dry heparinized Cuprophan hemodialysis membranes by means of ethylene oxide (EtO) exposure, gamma irradiation, or steam on the anticoagulant activity and chemical characteristics of immobilized heparin and the permeability of the membrane were investigated.

Eldecalcitol (ED-71), an analog of 1α,25(OH)2D3, inhibits the growth of squamous cell carcinoma (SCC) cells in vitro and in vivo by down-regulating expression of heparin-binding protein 17/fibroblast growth factor-binding protein-1 (HBp17/FGFBP-1) and FGF-2.

Science.gov (United States)

Shintani, T; Takatsu, F; Rosli, S N Z; Usui, E; Hamada, A; Sumi, K; Hayashido, Y; Toratani, S; Okamoto, Tetsuji

2017-10-01

Heparin-binding protein 17 (HBp17)/fibroblast growth factor-binding protein-1 (FGFBP-1) was first purified from medium conditioned by A431 cells for its capacity to bind to fibroblast growth factors 1 and 2 (FGF-1 and -2). Among FGF family members, FGF-2 is a potent mitogen for various cell types, including vascular endothelial cells, fibroblasts, and cancer cells such as oral squamous cell carcinoma (OSCC) cells. Besides being well known in bone metabolism, the active form of vitamin D 3 , i.e., 1α,25(OH) 2 D 3 (1,25D 3 ), was reported to have protective effects for heart disease and cancer. Previously, we reported that 1,25D 3 inhibited HBp17/FGFBP-1 expression in OSCC cell lines through NF-κB inhibition (IκBα activation) and resulted in the inactivation of FGF-2. In this study, we examined the potential anti-tumor effect of ED-71, an analog of 1α,25(OH) 2 D 3 , for squamous cell carcinoma cells in vitro and in vivo. The cell lines used were OSCC cell lines (NA-HO-1-n-1 and UE-HO-1-u-1), established from oral cancer patients in our laboratory, and an epidermoid carcinoma/SCC cell line (A431). The growth assay in serum-free culture revealed that ED-71 inhibited the growth of the cancer cell lines in a dose-dependent manner. In addition, ED-71 suppressed HBp17/FGFBP-1 expression by inhibiting the NF-κB pathway as did 1,25D 3 . Furthermore, a luciferase reporter assay revealed that the promoter activity of HBp17/FGFBP-1 (region between -217 and +61) was down-regulated by ED-71. Oral administration of ED-71 significantly inhibited the growth of A431-derived tumors in athymic nude mice. Immunohistochemical analysis revealed that the expression of HBp17/FGFBP-1, FGF-2, CD31, and Ki-67 in the tumors of ED71-treated group was down-regulated in comparison to control. These results suggest that ED-71 possesses potential anti-tumor activity for SCCs both in vitro and in vivo. This compound may act directly on the tumor cells or on endothelial cells by modulating the

Novel direct factor Xa inhibitory compounds from Tenebrio molitor with anti-platelet aggregation activity.

Science.gov (United States)

Lee, Wonhwa; Kim, Mi-Ae; Park, InWha; Hwang, Jae Sam; Na, MinKyun; Bae, Jong-Sup

2017-11-01

Tenebrio molitor is an edible insect that has antimicrobial, anticancer, and antihypertensive effects. The aim of this study was to identify the unreported bioactive compounds from T. molitor larvae with inhibitory activities against factor Xa (FXa) and platelet aggregation. Isolated compounds were evaluated for their anti-FXa and anti-platelet aggregation properties by monitoring clotting time, platelet aggregation, FXa activity, and thrombus formation. A diketopiperazine (1, cyclo( L -Pro- L -Tyr)) and a phenylethanoid (2, N-acetyltyramine) were isolated and inhibited the catalytic activity of FXa in a mixed inhibition model and inhibited platelet aggregation induced by adenosine diphosphate (ADP) and U46619. They inhibited ADP- and U46619-induced phosphorylation of myristoylated alanine-rich C kinase substrate (MARCKS) and the expression of P-selectin and PAC-1 in platelets. They also improved the production of nitric oxide and inhibited the oversecretion of endothelin-1 compared to that of the ADP- or U46619-treated group. In an animal model of arterial and pulmonary thrombosis, the isolated compounds showed enhanced antithrombotic effects. They also elicited anticoagulant effects in mice. Compounds 1-2 inhibited ADP-, collagen-, or U46619-induced platelet aggregation and showed similar anti-thrombotic efficacy to rivaroxaban, a positive control. Therefore, 1-2 could serve as candidates and provide scaffolds for the development of new anti-FXa and anti-platelet drugs. Copyright © 2017 Elsevier Ltd. All rights reserved.

Does ′heparin-induced thrombocytopenia′ hit our minds?

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Arun R Thangavel

2016-01-01

Full Text Available Unfractionated heparin is a widely used drug to prevent deep vein thrombosis and pulmonary emboli in patients at risk. With the advent of newer anticoagulants having lesser side effects, its use has diminished but not out of service. Here, we report a case of deep venous thrombosis, in a patient on prophylactic dose of heparin, which was later found to be a manifestation of heparin-induced thrombocytopenia (HIT. Thrombosis in the presence of heparin prophylaxis should be considered as HIT rather than a failure of anticoagulation.

Heparin octasaccharide decoy liposomes inhibit replication of multiple viruses

Science.gov (United States)

Hendricks, Gabriel L.; Velazquez, Lourdes; Pham, Serena; Qaisar, Natasha; Delaney, James C.; Viswanathan, Karthik; Albers, Leila; Comolli, James C.; Shriver, Zachary; Knipe, David M.; Kurt-Jones, Evelyn A.; Fygenson, Deborah K.; Trevejo, Jose M.

2016-01-01

Heparan sulfate (HS) is a ubiquitous glycosaminoglycan that serves as a cellular attachment site for a number of significant human pathogens, including respiratory syncytial virus (RSV), human parainfluenza virus 3 (hPIV3), and herpes simplex virus (HSV). Decoy receptors can target pathogens by binding to the receptor pocket on viral attachment proteins, acting as ‘molecular sinks’ and preventing the pathogen from binding to susceptible host cells. Decoy receptors functionalized with HS could bind to pathogens and prevent infection, so we generated decoy liposomes displaying HS-octasaccharide (HS-octa). These decoy liposomes significantly inhibited RSV, hPIV3, and HSV infectivity in vitro to a greater degree than the original HS-octa building block. The degree of inhibition correlated with the density of HS-octa displayed on the liposome surface. Decoy liposomes with HS-octa inhibited infection of viruses to a greater extent than either full-length heparin or HS-octa alone. Decoy liposomes were effective when added prior to infection or following the initial infection of cells in vitro. By targeting the well-conserved receptor-binding sites of HS-binding viruses, decoy liposomes functionalized with HS-octa are a promising therapeutic antiviral agent and illustrate the utility of the liposome delivery platform. PMID:25637710

Heparin- induced thrombocytopenia (HIT: a case report of CABG patient

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Alireza Jahangirifard

2016-08-01

Full Text Available Heparin- induced thrombocytopenia (HIT is an antibody mediated adverse effect of heparin therapy which is classified into two subtypes, HITI which is non-immune, spontaneously reversible thrombocytopenia and; HITII which is an autoimmune-mediated adverse effect of heparin therapy. In this case report, we described a 65-year old male patient with HITII after coronary artery bypass grafting.Key words: Heparin- induced thrombocytopenia, Heparin- induced thrombosis, coronary artery bypass grafting.

RNA-Seq analysis reveals insight into enhanced rice Xa7-mediated bacterial blight resistance at high temperature.

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Stephen P Cohen

Full Text Available Plant disease is a major challenge to agriculture worldwide, and it is exacerbated by abiotic environmental factors. During some plant-pathogen interactions, heat stress allows pathogens to overcome host resistance, a phenomenon which could severely impact crop productivity considering the global warming trends associated with climate change. Despite the importance of this phenomenon, little is known about the underlying molecular mechanisms. To better understand host plant responses during simultaneous heat and pathogen stress, we conducted a transcriptomics experiment for rice plants (cultivar IRBB61 containing Xa7, a bacterial blight disease resistance (R gene, that were infected with Xanthomonas oryzae, the bacterial blight pathogen of rice, during high temperature stress. Xa7-mediated resistance is unusual relative to resistance mediated by other R genes in that it functions better at high temperatures. Using RNA-Seq technology, we identified 8,499 differentially expressed genes as temperature responsive in rice cultivar IRBB61 experiencing susceptible and resistant interactions across three time points. Notably, genes in the plant hormone abscisic acid biosynthesis and response pathways were up-regulated by high temperature in both mock-treated plants and plants experiencing a susceptible interaction and were suppressed by high temperature in plants exhibiting Xa7-mediated resistance. Genes responsive to salicylic acid, an important plant hormone for disease resistance, were down-regulated by high temperature during both the susceptible and resistant interactions, suggesting that enhanced Xa7-mediated resistance at high temperature is not dependent on salicylic acid signaling. A DNA sequence motif similar to known abscisic acid-responsive cis-regulatory elements was identified in the promoter region upstream of genes up-regulated in susceptible but down-regulated in resistant interactions. The results of our study suggest that the plant

21 CFR 864.7525 - Heparin assay.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification. A...

Investigation of a Potential Protective Mechanism Against Heparin-Induced Thrombocytopenia in Patients on Chronic Intermittent Hemodialysis

Science.gov (United States)

Tanhehco, Yvette C.; Cuker, Adam; Rudnick, Michael; Sachais, Bruce S.

2015-01-01

BACKGROUND Heparin-induced thrombocytopenia (HIT) develops as a result of platelet (PLT) activation by anti-platelet factor 4 (PF4)/heparin complex antibodies. Despite repeated exposure to heparin, patients undergoing chronic intermittent hemodialysis (HD) rarely develop HIT. We investigated the possibility that HD decreases/removes PF4 from PLT surfaces and/or plasma, thereby disfavoring immune complex formation as a mechanism of protection against HIT. MATERIALS AND METHODS We enrolled 20 patients undergoing chronic HD at the Penn Presbyterian Medical Center. Blood samples were drawn before, during and after treatment in the presence and absence of heparin. PF4, PF4/heparin antibody, heparin, and P-selectin levels were measured. RESULTS No patients demonstrated clinical symptoms of HIT. PLT surface PF4 levels decreased and plasma PF4 levels increased concurrently with increase in plasma heparin concentration. In the absence of heparin, PLT surface and plasma PF4 levels were unchanged. Anti-PF4/heparin antibodies, which were non-functional by the serotonin release assay, were detectable in 8 patients. PLT surface P-selectin levels did not change during treatment. CONCLUSIONS Removal of PLT surface and/or plasma PF4 as a mechanism of protection against HIT in patients undergoing HD is not supported by the results of our study, although the transient decrease in PLT surface PF4 in the presence of large amounts of heparin remains a candidate mechanism. The small sample size, single type of dialyzer membrane, and early sampling time points may have led to the inability to detect changes in PF4 levels. Future studies should explore other potential protective mechanisms. PMID:23305841

Proteolytic activation transforms heparin cofactor II into a host defense molecule.

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Kalle, Martina; Papareddy, Praveen; Kasetty, Gopinath; Tollefsen, Douglas M; Malmsten, Martin; Mörgelin, Matthias; Schmidtchen, Artur

2013-06-15

The abundant serine proteinase inhibitor heparin cofactor II (HCII) has been proposed to inhibit extravascular thrombin. However, the exact physiological role of this plasma protein remains enigmatic. In this study, we demonstrate a previously unknown role for HCII in host defense. Proteolytic cleavage of the molecule induced a conformational change, thereby inducing endotoxin-binding and antimicrobial properties. Analyses employing representative peptide epitopes mapped these effects to helices A and D. Mice deficient in HCII showed increased susceptibility to invasive infection by Pseudomonas aeruginosa, along with a significantly increased cytokine response. Correspondingly, decreased levels of HCII were observed in wild-type animals challenged with bacteria or endotoxin. In humans, proteolytically cleaved HCII forms were detected during wounding and in association with bacteria. Thus, the protease-induced uncovering of cryptic epitopes in HCII, which transforms the molecule into a host defense factor, represents a previously unknown regulatory mechanism in HCII biology and innate immunity.

Advanced nanocarriers based on heparin and its derivatives for cancer management.

Science.gov (United States)

Yang, Xiaoye; Du, Hongliang; Liu, Jiyong; Zhai, Guangxi

2015-02-09

To obtain a satisfying anticancer effect, rationally designed nanocarriers are intensively studied. In this field, heparin and its derivatives have been widely attempted recently as potential component of nanocarriers due to their unique biological and physiochemical features, especially the anticancer activity. This review focuses on state-of-the-art nanocarriers with heparin/heparin derivatives as backbone or coating material. At the beginning, the unique advantages of heparin used in cancer nanotechnology are discussed. After that, different strategies of heparin chemical modification are reviewed, laying the foundation of developing various nanocarriers. Then a systematic summary of diverse nanoparticles with heparin as component is exhibited, involving heparin-drug conjugate, polymeric nanoparticles, nanogels, polyelectrolyte complex nanoparticles, and heparin-coated organic and inorganic nanoparticles. The application of these nanoparticles in various novel cancer therapy (containing targeted therapy, magnetic therapy, photodynamic therapy, and gene therapy) will be highlighted. Finally, future challenges and opportunities of heparin-based biomaterials in cancer nanotechnology are discussed.

Characterization of heparin aerosols generated in jet and ultrasonic nebulizers

DEFF Research Database (Denmark)

Bendstrup, K.E.; Newhouse, M.T.; Pedersen, Ole Finn

1999-01-01

Inhaled heparin has been used for asthma treatment, but results have been inconsistent, probably due to highly varying lung doses. We determined the output per unit time and the particle size distributions of sodium heparin, calcium heparin, and low molecular weight (LMW) heparin formulations in ...... on the exhalation filter, and 15,000 IU was captured on the inhalation filter (inhaled mass). This corresponds to a respirable mass of 10,000 IU of heparin with a high probability of reaching the lower respiratory tract in normal healthy adults....

Citrate Anticoagulation for CRRT in Children: Comparison with Heparin

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Sara Nicole Fernández

2014-01-01

Full Text Available Regional anticoagulation with citrate is an alternative to heparin in continuous renal replacement therapies, which may prolong circuit lifetime and decrease hemorrhagic complications. A retrospective comparative cohort study based on a prospective observational registry was conducted including critically ill children undergoing CRRT. Efficacy, measured as circuit survival, and secondary effects of heparin and citrate were compared. 12 patients on CRRT with citrate anticoagulation and 24 patients with heparin anticoagulation were analyzed. Median citrate dose was 2.6 mmol/L. Median calcium dose was 0.16 mEq/kg/h. Median heparin dose was 15 UI/kg/h. Median circuit survival was 48 hours with citrate and 31 hours with heparin (P=0.028. 66.6% of patients treated with citrate developed mild metabolic alkalosis, which was directly related to citrate dose. There were no cases of citrate intoxication: median total calcium/ionic calcium index (CaT/I of 2.16 and a maximum CaT/I of 2.33, without metabolic acidosis. In the citrate group, 45.5% of patients developed hypochloremia and 27.3% hypomagnesemia. In the heparin group, 27.8% developed hypophosphatemia. Three patients were moved from heparin to citrate to control postoperatory bleeding. In conclusion citrate is a safe and effective anticoagulation method for CRRT in children and it achieves longer circuit survival than heparin.

Structure of rat acidic fibroblast growth factor at 1.4 A resolution

DEFF Research Database (Denmark)

Kulahin, Nikolaj; Kiselyov, Vladislav; Kochoyan, Artur

2007-01-01

Fibroblast growth factors (FGFs) constitute a family of 22 structurally related heparin-binding polypeptides that are involved in the regulation of cell growth, survival, differentiation and migration. Here, a 1.4 A resolution X-ray structure of rat FGF1 is presented. Two molecules are present...

Electrophoresis for the analysis of heparin purity and quality.

Science.gov (United States)

Volpi, Nicola; Maccari, Francesca; Suwan, Jiraporn; Linhardt, Robert J

2012-06-01

The adulteration of raw heparin with oversulfated chondroitin sulfate (OSCS) in 2007-2008 produced a global crisis resulting in extensive revisions to the pharmacopeia monographs and prompting the FDA to recommend the development of additional methods for the analysis of heparin purity. As a consequence, a wide variety of innovative analytical approaches have been developed for the quality assurance and purity of unfractionated and low-molecular-weight heparins. This review discusses recent developments in electrophoresis techniques available for the sensitive separation, detection, and partial structural characterization of heparin contaminants. In particular, this review summarizes recent publications on heparin quality and related impurity analysis using electrophoretic separations such as capillary electrophoresis (CE) of intact polysaccharides and hexosamines derived from their acidic hydrolysis, and polyacrylamide gel electrophoresis (PAGE) for the separation of heparin samples without and in the presence of its relatively specific depolymerization process with nitrous acid treatment. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biological distribution of 51Cr-heparin

International Nuclear Information System (INIS)

Almeida, M.A.T.M. de.

1979-01-01

The kinetics of heparin in normal Wistar rats using the radioactive tracer 51 Cr, has been studied. The labeled and purified 51 Cr-heparin was injected into rats intravenously and by intraperitoneal injection. In measuring the radioactivity of organs it was possible to conclude that the tissues rich in mast cells, liver and spleen, were found to take up the greater amounts of heparin. The curve that represents the logarithm of the concentration of heparin versus time is biexponential. The half-lives of the two exponential were determined. The volume of distribution, the rate constant and the renal clearance were determined by the values of the plasma levels and urinary excretions. The biological half-time, the turnover rate and the turnover time were determined by measuring the residual radioactivity of the total body and urinary excretions. With the data obtained from the mentioned experiments a compartmental model was performed in which the plasma is the central compartment for the distribution of the drug, exchanging with another extraplasmatic compartment and finally the drug being stored in reticulo endothelial system cells. (Author) [pt

Influence of spacer length on heparin coupling efficiency and fibrinogen adsorption of modified titanium surfaces

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Gbureck Uwe

2007-07-01

Full Text Available Abstract Background Chemical bonding of the drug onto surfaces by means of spacer molecules is accompanied with a reduction of the biological activity of the drug due to a constricted mobility since normally only short spacer molecule like aminopropyltrimethoxysilane (APMS are used for drug coupling. This work aimed to study covalent attachment of heparin to titanium(oxide surfaces by varying the length of the silane coupling agent, which should affect the biological potency of the drug due to a higher mobility with longer spacer chains. Methods Covalent attachment of heparin to titanium metal and TiO2 powder was carried out using the coupling agents 3-(Trimethoxysilyl-propylamine (APMS, N- [3-(Trimethoxysilylpropyl]ethylenediamine (Diamino-APMS and N1- [3-(Trimethoxy-silyl-propyl]diethylenetriamine (Triamino-APMS. The amount of bound coupling agent and heparin was quantified photometrically by the ninhydrin reaction and the tolidine-blue test. The biological potency of heparin was determined photometrically by the chromogenic substrate Chromozym TH and fibrinogen adsorption to the modified surfaces was researched using the QCM-D (Quartz Crystal Microbalance with Dissipation Monitoring technique. Results Zeta-potential measurements confirmed the successful coupling reaction; the potential of the unmodified anatase surface (approx. -26 mV shifted into the positive range (> + 40 mV after silanisation. Binding of heparin results in a strongly negatively charged surface with zeta-potentials of approx. -39 mV. The retaining biological activity of heparin was highest for the spacer molecule Triamino-APMS. QCM-D measurements showed a lower viscosity for adsorbed fibrinogen films on heparinised surfaces by means of Triamino-APMS. Conclusion The remaining activity of heparin was found to be highest for the covalent attachment with Triamino-APMS as coupling agent due to the long chain of this spacer molecule and therefore the highest mobility of the drug

Heparin defends against the toxicity of circulating histones in sepsis.

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Wang, Feifei; Zhang, Naipu; Li, Biru; Liu, Lanbo; Ding, Lei; Wang, Ying; Zhu, Yimin; Mo, Xi; Cao, Qing

2015-06-01

Although circulating histones were demonstrated as major mediators of death in septic mice models, their roles in septic patients are not clarified. The present study sought to evaluate the clinical relevance of the circulating histone levels in septic children, and the antagonizing effects of heparin on circulating histones. Histone levels in the plasma of septic children were significantly higher than healthy controls, and positively correlated with disease severity. Histone treatment could activate NF-κB pathway of the endothelial cells and induce the secretion of large amount of cytokines that further amplify inflammation, subsequently leading to organ damage. Co-injection of low dose heparin with lethal dose histones could protect mouse from organ damage and death by antagonizing circulating histones, and similar effects were also observed in other septic models. Collectively, these findings indicated that circulating histones might serve as key factors in the pathogenesis of sepsis and their levels in plasma might be a marker for disease progression and prognosis. Furthermore, low dose heparin might be an effective therapy to hamper sepsis progression and reduce the mortality.

Heparin and Heparin-Derivatives in Post-Subarachnoid Hemorrhage Brain Injury: A Multimodal Therapy for a Multimodal Disease

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Erik G. Hayman

2017-05-01

Full Text Available Pharmacologic efforts to improve outcomes following aneurysmal subarachnoid hemorrhage (aSAH remain disappointing, likely owing to the complex nature of post-hemorrhage brain injury. Previous work suggests that heparin, due to the multimodal nature of its actions, reduces the incidence of clinical vasospasm and delayed cerebral ischemia that accompany the disease. This narrative review examines how heparin may mitigate the non-vasospastic pathological aspects of aSAH, particularly those related to neuroinflammation. Following a brief review of early brain injury in aSAH and heparin’s general pharmacology, we discuss potential mechanistic roles of heparin therapy in treating post-aSAH inflammatory injury. These roles include reducing ischemia-reperfusion injury, preventing leukocyte extravasation, modulating phagocyte activation, countering oxidative stress, and correcting blood-brain barrier dysfunction. Following a discussion of evidence to support these mechanistic roles, we provide a brief discussion of potential complications of heparin usage in aSAH. Our review suggests that heparin’s use in aSAH is not only safe, but effectively addresses a number of pathologies initiated by aSAH.

Efficacy and safety of once daily low molecular weight heparin (tinzaparin sodium) in high risk pregnancy.

LENUS (Irish Health Repository)

Ní Ainle, Fionnuala

2008-10-01

Low molecular weight heparin (LMWH) is widely regarded as the anticoagulant treatment of choice for the prevention and treatment of venous thromboembolism during pregnancy. However, previous studies have demonstrated that the pharmacokinetic profiles of LMWH vary significantly with increasing gestation. Consequently, it remains unclear whether LMWH regimens recommended for use in nonpregnant individuals can be safely extrapolated to pregnant women. The aims of this study were to assess the safety and the efficacy of tinzaparin sodium (Innohep) administered only once daily during pregnancy. A systematic retrospective review identified a cohort of 37 high-risk pregnancies which had been managed using tinzaparin 175 IU\\/kg once daily. In 26 cases, the index pregnancy had been complicated by development of an acute venous thromboembolism (17 deep vein thrombosis and nine pulmonary embolism). For each individual, case notes were examined and data extracted using a predetermined questionnaire. No episodes of recurrent venous thromboembolism were identified amongst this cohort of pregnancies managed using once daily LMWH administration. However, two unusual thrombotic complications were observed, including a parietal infarct in one patient, and a postpartum cerebral venous thrombosis in another. Once daily tinzaparin was well tolerated, with no cases of heparin-induced thrombocytopaenia, symptomatic osteoporosis, or foetal malformations. Tinzaparin dose modification based upon peak anti-Xa levels occurred in 45% of the cases examined. The present study is the largest study to have examined the clinical efficacy of once daily LMWH for use in pregnant women at high risk of venous thromboembolism. Our data support the safety and efficacy of antenatal tinzaparin at a dose of 175 IU\\/kg. In order to determine whether this once daily regimen provides equivalent (or indeed greater) thromboprophylaxis to twice daily LMWH regimens during pregnancy will require highly powered

Adaptive evolution of transcription factor binding sites

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Berg Johannes

2004-10-01

Full Text Available Abstract Background The regulation of a gene depends on the binding of transcription factors to specific sites located in the regulatory region of the gene. The generation of these binding sites and of cooperativity between them are essential building blocks in the evolution of complex regulatory networks. We study a theoretical model for the sequence evolution of binding sites by point mutations. The approach is based on biophysical models for the binding of transcription factors to DNA. Hence we derive empirically grounded fitness landscapes, which enter a population genetics model including mutations, genetic drift, and selection. Results We show that the selection for factor binding generically leads to specific correlations between nucleotide frequencies at different positions of a binding site. We demonstrate the possibility of rapid adaptive evolution generating a new binding site for a given transcription factor by point mutations. The evolutionary time required is estimated in terms of the neutral (background mutation rate, the selection coefficient, and the effective population size. Conclusions The efficiency of binding site formation is seen to depend on two joint conditions: the binding site motif must be short enough and the promoter region must be long enough. These constraints on promoter architecture are indeed seen in eukaryotic systems. Furthermore, we analyse the adaptive evolution of genetic switches and of signal integration through binding cooperativity between different sites. Experimental tests of this picture involving the statistics of polymorphisms and phylogenies of sites are discussed.

Heparin-Induced Thrombocytopenia

Science.gov (United States)

... HIT information card. Early identification of HIT and avoidance of inappropriate heparin therapy can help promote a ... Heart Association is a qualified 501(c)(3) tax-exempt organization. *Red Dress™ DHHS, Go Red™ AHA; ...

Identification of bacterial blight resistance genes Xa4 in Pakistani ...

African Journals Online (AJOL)

SERVER

2008-03-04

Mar 4, 2008 ... Bacterial blight (BB) caused by Xanthomonas oryzae pv oryzae (Xoo) is a major biotic constraint in the irrigated rice belts. Genetic resistance is the most effective and economical control for bacterial blight. Molecular survey was conducted to identify the rice germplasm/lines for the presence of Xa4, a.

Increased synthesis of heparin affin regulatory peptide in the perforant path lesioned mouse hippocampal formation

DEFF Research Database (Denmark)

Poulsen, F R; Lagord, C; Courty, J

2000-01-01

Heparin affin regulatory peptide (HARP), also known as pleiotrophin or heparin-binding growth-associated molecule, is a developmentally regulated extracellular matrix protein that induces cell proliferation and promotes neurite outgrowth in vitro as well as pre- and postsynaptic developmental...... differentiation in vivo. Here we have investigated the expression of HARP mRNA and protein in the perforant path lesioned C57B1/6 mouse hippocampal formation from 1 to 35 days after surgery. This type of lesion induces a dense anterograde and terminal axonal degeneration, activation of glial cells, and reactive...... axonal sprouting within the perforant path zones of the fascia dentata and hippocampus as well as axotomy-induced retrograde neuronal degeneration in the entorhinal cortex. Analysis of sham- and unoperated control mice showed that HARP mRNA is expressed in neurons and white and gray matter glial cells...

Identification of a novel structure in heparin generated by potassium permanganate oxidation

Science.gov (United States)

Beccati, Daniela; Roy, Sucharita; Yu, Fei; Gunay, Nur Sibel; Capila, Ishan; Lech, Miroslaw; Linhardt, Robert J.; Venkataraman, Ganesh

2012-01-01

The worldwide heparin contamination crisis in 2008 led health authorities to take fundamental steps to better control heparin manufacture, including implementing appropriate analytical and bio-analytical methods to ensure production and release of high quality heparin sodium product. Consequently, there is an increased interest in the identification and structural elucidation of unusually modified structures that may be present in heparin. Our study focuses on the structural elucidation of species that give rise to a signal observed at 2.10 ppm in the N-acetyl region of the 1H NMR spectrum of some pharmaceutical grade heparin preparations. Structural elucidation experiments were carried out using homonuclear (COSY, TOSCY and NOESY) and heteronuclear (HSQC, HSQC-DEPT, HMQC-COSY, HSQC-TOCSY, and HMBC) 2D NMR spectroscopy on both heparin as well as heparin-like model compounds. Our results identify a novel type of oxidative modification of the heparin chain that results from a specific step in the manufacturing process used to prepare heparin. PMID:25147414

Anticoagulant activity in salivary glands of the insect vector Culicoides variipennis sonorensis by an inhibitor of factor Xa.

Science.gov (United States)

Pérez de León, A A; Valenzuela, J G; Tabachnick, W J

1998-02-01

Blood feeding by the insect vector Culicoides variipennis sonorensis involves laceration of superficial host tissues, an injury that would be expected to trigger the coagulation cascade. Accordingly, the salivary glands of C.v. sonorensis were examined for the presence of an antihemostatic that prevents blood coagulation. Assays using salivary gland extracts showed a delay in the recalcification time of plasma devoid of platelets, indicating the presence of anticoagulant activity. Retardation in the formation of a fibrin clot was also observed after the addition of tissue factor to plasma that was preincubated with salivary gland extracts. Similarly, an inhibitory effect by salivary gland extracts was detected in assays that included factors of the intrinsic pathway. Inhibition of the catalytic activity of purified factor Xa toward its chromogenic substrate suggested that it was the target of the salivary anticoagulant of C.v. sonorensis. This was corroborated by the coincidence of anticoagulant and anti-FXa activities obtained by reverse-phase HPLC. The depletion of anti-FXa activity from salivary glands during blood feeding suggests that the FXa inhibitor functions as anticoagulant. Molecular sieving HPLC yielded an apparent molecular mass of 28 kDa for the salivary FXa inhibitor of C.v. sonorensis. Preventing the formation of thrombin through the inhibition of FXa likely facilitates blood feeding by maintaining the pool of blood fluid at the feeding site. The salivary FXa inhibitor of C.v. sonorensis could impair the network of host-defense mechanisms in the skin microenvironment by avoiding blood coagulation at the site of feeding.

Construction of mussel-inspired coating via the direct reaction of catechol and polyethyleneimine for efficient heparin immobilization

Energy Technology Data Exchange (ETDEWEB)

Liu, Yujie [School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); The Institute of Biomaterials and Surface Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Luo, Rifang, E-mail: lrifang@126.com [School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); The Institute of Biomaterials and Surface Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Shen, Fangyu; Tang, Linlin [School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); The Institute of Biomaterials and Surface Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Wang, Jin, E-mail: jinxxwang@263.net [School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); The Institute of Biomaterials and Surface Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Huang, Nan [School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); The Institute of Biomaterials and Surface Engineering, Southwest Jiaotong University, Chengdu 610031 (China)

2015-02-15

Highlights: • Catechol (CA) and PEI copolymerization was a mimetic and dopamine-like coating method. • CA/PEI film provided amine groups and was effective in heparin immobilization. • CA/PEI coating could inhibit smooth muscle cell proliferation. • CA/PEI coating did not show any significant cytotoxicity to endothelial cell. - Abstract: Dopamine could self-polymerize to form the coating on various substrates and the co-existence of catechols and amines was crucial in performing such polymerization process. In this work, a mimetic approach of coating formation was carried out based on the co-polymerization of catechol (CA) and polyethyleneimine (PEI). Mussel-inspired CA/PEI coating was deposited on 316L stainless steel (SS). Fourier transform infrared spectra (FTIR) and X-ray photoelectron spectroscopy (XPS) demonstrated the successful coating formation. QCM measurement showed good affinity of heparin immobilization on CA/PEI coating surface ascribed to the amine groups. Herein, vascular cell-material interactions like endothelial cells (ECs) and smooth muscle cells (SMCs) were also investigated. Interestingly, CA/PEI and heparin modified coatings presented no cytotoxicity to ECs, however to a certain extent, decreased SMCs proliferation. Moreover, heparin-binding surface presented significant anti-platelet adhesion and activation properties. These results effectively suggested that the mussel-inspired CA/PEI coating might be promising when served as a platform for biomolecule immobilization.

Cyclic Voltammetry of Biopolymer Heparin at PVC Plasticized Liquid Membrane

Czech Academy of Sciences Publication Activity Database

Samec, Zdeněk; Trojánek, Antonín; Langmaier, Jan; Samcová, E.

2003-01-01

Roč. 5, - (2003), s. 867-870 ISSN 1388-2481 R&D Projects: GA ČR GA203/04/0424 Institutional research plan: CEZ:AV0Z4040901 Keywords : cyclic voltammetry * PVC plasticized liquit membrane * heparin Subject RIV: CG - Electrochemistry Impact factor: 2.300, year: 2003

The Use of Heparin during Endovascular Peripheral Arterial Interventions: A Synopsis

Directory of Open Access Journals (Sweden)

Arno M. Wiersema

2016-01-01

Full Text Available A large variety exists for many aspects of the use of heparin as periprocedural prophylactic antithrombotics (PPAT during peripheral arterial interventions (PAI. This variation is present, not only within countries, but also between them. Due to a lack of (robust data, no systematic review on the use of heparin during PAI could be justified. A synopsis of all available literature on heparin during PAI describes that heparin is used on technical equipment to reduce the thrombogenicity and in the flushing solution with saline. Heparin could have a cumulative anticoagulant effect when used in combination with ionic contrast medium. No level-1 evidence exists on the use of heparin. A measurement of actual anticoagulation status by means of an activated clotting time should be mandatory.

Factor XI and XII as antithrombotic targets.

Science.gov (United States)

Müller, Felicitas; Gailani, David; Renné, Thomas

2011-09-01

Arterial and venous thrombosis are major causes of morbidity and mortality, and the incidence of thromboembolic diseases increases as a population ages. Thrombi are formed by activated platelets and fibrin. The latter is a product of the plasma coagulation system. Currently available anticoagulants such as heparins, vitamin K antagonists and inhibitors of thrombin or factor Xa target enzymes of the coagulation cascade that are critical for fibrin formation. However, fibrin is also necessary for terminating blood loss at sites of vascular injury. As a result, anticoagulants currently in clinical use increase the risk of bleeding, partially offsetting the benefits of reduced thrombosis. This review focuses on new targets for anticoagulation that are associated with minimal or no therapy-associated increased bleeding. Data from experimental models using mice and clinical studies of patients with hereditary deficiencies of coagulation factors XI or XII have shown that both of these clotting factors are important for thrombosis, while having minor or no apparent roles in processes that terminate blood loss (hemostasis). Hereditary deficiency of factor XII (Hageman factor) or factor XI, plasma proteases that initiate the intrinsic pathway of coagulation, impairs thrombus formation and provides protection from vascular occlusive events, while having a minimal impact on hemostasis. As the factor XII-factor XI pathway contributes to thrombus formation to a greater extent than to normal hemostasis, pharmacological inhibition of these coagulation factors may offer the exciting possibility of anticoagulation therapies with minimal or no bleeding risk.

Use of heparin in the investigation of obscure gastrointestinal bleeding

International Nuclear Information System (INIS)

Mernagh, J.R.; O'Donovan, N.; Somers, S.; Gill, G.; Sridhar, S.

2001-01-01

To determine if the administration of heparin improves the predictive value of angiography in the investigation of obscure gastrointestinal (GI) bleeding. 18 patients with a history of chronic GI bleeding were investigated with angiography. For 6 patients, the cause of GI bleeding was established with angiography; the 12 patients who had negative results were given heparin for 24 h and were reassessed with angiography. After heparin administration, the source of GI bleeding was determined with angiography for 6 of the remaining 12 patients. Thus, heparinization increased diagnostic yield from 33% (6 of 18) to 67% (12 of 18). No significant complications, such as uncontrolled GI bleeding, occurred. Heparinization improves the diagnostic yield of angiography when obscure GI bleeding is being investigated. (author)

Use of Enoxaparin in Obese Adolescents During Bariatric Surgery--a Pilot Study.

Science.gov (United States)

Mushtaq, Alvina; Vaughns, Janelle D; Ziesenitz, Victoria C; Nadler, Evan P; van den Anker, John N

2015-10-01

Obese patients have a higher risk of venous thromboembolism when immobilized due to surgery. The objective of this study was to assess anti-factor Xa activity in adolescent bariatric surgical patients receiving prophylactic enoxaparin. Four morbidly obese adolescents undergoing laparoscopic sleeve gastrectomy were enrolled. Enoxaparin was administered (40 mg subcutaneous (SC) if BMI ≤50 kg/m(2) or 60 mg SC if BMI >50 kg/m(2)) for prevention of venous thromboembolism every 12 h starting after induction of anesthesia until discharge. Plasma anti-factor Xa activity was assessed over 12 h after the first dose and used as a surrogate marker for enoxaparin levels. Non-compartmental analysis of anti-factor Xa activity levels was performed and compared with previously published studies. Patients recruited were 16 to 18 years of age with a mean BMI of 52.6 ± 5.8 kg/m(2) (>99th BMI percentile). Peak anti-factor Xa activity ranged from 0.20 to 0.23 IU/mL in our study population, compared to 0.38 to 0.53 IU/mL in the cited lean comparator groups. Our current dosing practice of 40 mg SC for individuals with a BMI ≤50 kg/m(2) and 60 mg for individuals with a BMI ≥50 kg/m(2) resulted in anti-factor Xa activity that was sufficient for adequate thromboprophylaxis in adolescent bariatric surgical patients. Our data also demonstrates lower drug exposures in the obese when compared to lean patients. Therefore, randomized controlled efficacy and safety studies are urgently needed to guide the use of low-molecular-weight heparins in the pediatric and adolescent obese population.

The effect of a new direct Factor Xa inhibitor on human osteoblasts: an in-vitro study comparing the effect of rivaroxaban with enoxaparin

LENUS (Irish Health Repository)

Solayar, Gandhi N

2011-10-28

Abstract Background Current treatments for the prevention of thromboembolism include heparin and low-molecular weight heparins (LMWHs). A number of studies have suggested that long term administration of these drugs may adversely affect osteoblasts and therefore, bone metabolism. Xarelto™ (Rivaroxaban) is a new anti-thrombotic drug for the prevention of venous thromboembolism in adult patients undergoing elective hip and knee replacement surgery. The aim of this in vitro study was to investigate the possible effects of rivaroxaban on osteoblast viability, function and gene expression compared to enoxaparin, a commonly used LMWH. Methods Primary human osteoblast cultures were treated with varying concentrations of rivaroxaban (0.013, 0.13, 1.3 and 13 μg\\/ml) or enoxaparin (1, 10 and 100 μg\\/ml). The effect of each drug on osteoblast function was evaluated by measuring alkaline phosphatase activity. The MTS assay was used to assess the effect of drug treatments on cell proliferation. Changes in osteocalcin, Runx2 and BMP-2 messenger RNA (mRNA) expression following drug treatments were measured by real-time polymerase chain reaction (PCR). Results Rivaroxaban and enoxaparin treatment did not adversely affect osteoblast viability. However, both drugs caused a significant reduction in osteoblast function, as measured by alkaline phosphatase activity. This reduction in osteoblast function was associated with a reduction in the mRNA expression of the bone marker, osteocalcin, the transcription factor, Runx2, and the osteogenic factor, BMP-2. Conclusions These data show that rivaroxaban treatment may negatively affect bone through a reduction in osteoblast function.

Novel Drosophila receptor that binds multiple growth factors

International Nuclear Information System (INIS)

Rosner, M.R.; Thompson, K.L.; Garcia, V.; Decker, S.J.

1986-01-01

The authors have recently reported the identification of a novel growth factor receptor from Drosophila cell cultures that has dual binding specificity for both insulin and epidermal growth factor (EGF). This 100 kDa protein is also antigenically related to the cytoplasmic region of the mammalian EGF receptor-tyrosine kinase. They now report that this protein binds to mammalian nerve growth factor and human transforming growth factor alpha as well as insulin and EGF with apparent dissociation constants ranging from 10 -6 to 10 -8 M. The 100 kDa protein can be affinity-labeled with these 125 I-labeled growth factors after immunoprecipitation with anti-EGF receptor antiserum. These four growth factors appear to share a common binding site, as evidenced by their ability to block affinity labelling by 125 I-insulin. No significant binding to the 100 kDa protein was observed with platelet-derived growth factor, transforming growth factor beta, or glucagon. The 100 kDa Drosophila protein has a unique ligand-binding spectrum with no direct counterpart in mammalian cells and may represent an evolutionary precursor of the mammalian receptors for these growth factors

Pathogenic Leptospira species acquire factor H and vitronectin via the surface protein LcpA.

Science.gov (United States)

da Silva, Ludmila Bezerra; Miragaia, Lidia Dos Santos; Breda, Leandro Carvalho Dantas; Abe, Cecilia Mari; Schmidt, Mariana Costa Braga; Moro, Ana Maria; Monaris, Denize; Conde, Jonas Nascimento; Józsi, Mihály; Isaac, Lourdes; Abreu, Patrícia Antônia Estima; Barbosa, Angela Silva

2015-03-01

Upon infection, pathogenic Leptospira species bind several complement regulators in order to overcome host innate immunity. We previously characterized a 20-kDa leptospiral surface protein which interacts with C4b binding protein (C4BP): leptospiral complement regulator-acquiring protein A (LcpA). Here we show that LcpA also interacts with human factor H (FH), which remains functionally active once bound to the protein. Antibodies directed against short consensus repeat 20 (SCR20) inhibited binding of FH to LcpA by approximately 90%, thus confirming that this particular domain is involved in the interaction. We have also shown for the first time that leptospires bind human vitronectin and that the interaction is mediated by LcpA. Coincubation with heparin blocked LcpA-vitronectin interaction in a dose-dependent manner, strongly suggesting that binding may occur through the heparin binding domains of vitronectin. LcpA also bound to the terminal pathway component C9 and inhibited Zn(2+)-induced polymerization and membrane attack complex (MAC) formation. Competitive binding assays indicated that LcpA interacts with C4BP, FH, and vitronectin through distinct sites. Taken together, our findings indicate that LcpA may play a role in leptospiral immune evasion. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Investigation of the heparin-thrombin interaction by dynamic force spectroscopy.

Science.gov (United States)

Wang, Congzhou; Jin, Yingzi; Desai, Umesh R; Yadavalli, Vamsi K

2015-06-01

The interaction between heparin and thrombin is a vital step in the blood (anti)coagulation process. Unraveling the molecular basis of the interactions is therefore extremely important in understanding the mechanisms of this complex biological process. In this study, we use a combination of an efficient thiolation chemistry of heparin, a self-assembled monolayer-based single molecule platform, and a dynamic force spectroscopy to provide new insights into the heparin-thrombin interaction from an energy viewpoint at the molecular scale. Well-separated single molecules of heparin covalently attached to mixed self-assembled monolayers are demonstrated, whereby interaction forces with thrombin can be measured via atomic force microscopy-based spectroscopy. Further these interactions are studied at different loading rates and salt concentrations to directly obtain kinetic parameters. An increase in the loading rate shows a higher interaction force between the heparin and thrombin, which can be directly linked to the kinetic dissociation rate constant (koff). The stability of the heparin/thrombin complex decreased with increasing NaCl concentration such that the off-rate was found to be driven primarily by non-ionic forces. These results contribute to understanding the role of specific and nonspecific forces that drive heparin-thrombin interactions under applied force or flow conditions. Copyright © 2015 Elsevier B.V. All rights reserved.

Artemin Crystal Structure Reveals Insights into Heparan Sulfate Binding

Energy Technology Data Exchange (ETDEWEB)

Silvian,L.; Jin, P.; Carmillo, P.; Boriack-Sjodin, P.; Pelletier, C.; Rushe, M.; Gong, B.; Sah, D.; Pepinsky, B.; Rossomando, A.

2006-01-01

Artemin (ART) promotes the growth of developing peripheral neurons by signaling through a multicomponent receptor complex comprised of a transmembrane tyrosine kinase receptor (cRET) and a specific glycosylphosphatidylinositol-linked co-receptor (GFR{alpha}3). Glial cell line-derived neurotrophic factor (GDNF) signals through a similar ternary complex but requires heparan sulfate proteoglycans (HSPGs) for full activity. HSPG has not been demonstrated as a requirement for ART signaling. We crystallized ART in the presence of sulfate and solved its structure by isomorphous replacement. The structure reveals ordered sulfate anions bound to arginine residues in the pre-helix and amino-terminal regions that were organized in a triad arrangement characteristic of heparan sulfate. Three residues in the pre-helix were singly or triply substituted with glutamic acid, and the resulting proteins were shown to have reduced heparin-binding affinity that is partly reflected in their ability to activate cRET. This study suggests that ART binds HSPGs and identifies residues that may be involved in HSPG binding.

Current Trends in Heparin Use During Arterial Vascular Interventional Radiology

International Nuclear Information System (INIS)

Durran, Alexandra C.; Watts, Christopher

2012-01-01

Purpose: This study was designed to assess the current use of heparinized saline and bolus doses of heparin in non-neurological interventional radiology and to determine whether consensus could be reached to produce guidance for heparin use during arterial vascular intervention. Methods: An interactive electronic questionnaire was distributed to members of the British Society of Interventional Radiology regarding their current practice in the use, dosage, and timing of heparin boluses and heparinized flushing solutions.ResultsA total of 108 completed questionnaires were received. More than 80% of respondents used heparinized saline with varying concentrations; the most prevalent was 1,000 IU/l (international units of heparin per liter) and 5,000 IU/l. Fifty-one percent of interventionalists use 3,000 IU as their standard bolus dose; however, the respondents were split regarding the timing of bolus dose with ∼60% administering it after arterial access is obtained and 40% after crossing the lesion. There was no consensus on altering dose according to body weight, and only 4% monitored clotting parameters. Conclusions: There seems to be some coherence among practicing interventionalists regarding heparin administration. We hypothesize that heparinized saline should be used at a recognized standard concentration of 1,000 IU/l as a flushing concentration in all arterial vascular interventions and that 3,000 IU bolus is considered the standard dose for straightforward therapeutic procedures and 5000 IU for complex, crural, and endovascular aneurysm repair work. The bolus should be given after arterial access is obtained to allow time for optimal anticoagulation to be achieved by the time of active intervention and stenting. Further research into clotting abnormalities following such interventional procedures would be an interesting quantifiable follow-up to this initial survey of opinions and practice.

The intracellular uptake and protracted release of exogenous heparins by cultured endothelial cells

International Nuclear Information System (INIS)

Hiebert, L.M.; McDuffie, N.M.

1989-01-01

Heparins from bovine or porcine sources were fed in media for 48 hrs to cultured porcine aortic and human umbilical vein endothelial cells. Heparin was found in pericellular and cellular fractions after extraction by chemical methods and 125 I radiolabelled heparins were recovered when radiolabelled heparin was included in the feed. Even after washing and media changes heparin was detected in media and cell fractions up to 6 days post feeding. Metachromatic vacuoles within cells were demonstrated histologically up to 7 days post feeding after staining with toluidine blue. This is the first report of protracted internalization of exogenous heparin by cultured endothelial cells with concurrent prolonged release of the heparin to the media. This clearly demonstrates that the endothelium plays an important role in the distribution and metabolism of heparin

Community nurse resource implications for a change in heparin prophylaxis policy

Directory of Open Access Journals (Sweden)

Parker Martyn J.

2015-01-01

Full Text Available Introduction: A review was undertaken for a consecutive series of hip fracture patients for the year before and then after a change in low dose heparin prophylaxis policy. Patients and methods: For the first year heparin was administered in hospital for a maximum of 14 days only. Patients sent home before this time were not discharged taking heparin. For the second year heparin was administered as recommended by NICE guidelines for 28 days from admission regardless of whether the patient was discharged. Results: For the first year 486 patients were treated with a mean of 10.4 doses of heparin per patient. For the second year 465 patients were treated with a mean of 24.3 doses per patient. In total an extra 6,464 doses of heparin were administered. 33.8% of patients were unable to administer their heparin at home therefore a district nurse administered 2,284 of these doses of subcutaneous heparin at the patient’s home. The increased cost associated with the change in policy was estimated to be £161 per patient, with over 90% of this increase being incurred by the district nurse expense. If applied nationally for the England, using extended heparin prophylaxis for hip fracture patients would cost in excess of 12 million pounds each year. Conclusion: Whilst the necessity for and duration of thromboembolic prophylaxis for these patients remains undetermined, there is a need to re-evaluate the cost effectiveness of the current recommendations for hip fracture patients.

Determination of rivaroxaban in patient’s plasma samples by anti-Xa chromogenic test associated to High Performance Liquid Chromatography tandem Mass Spectrometry (HPLC-MS/MS)

Science.gov (United States)

Derogis, Priscilla Bento Matos; Sanches, Livia Rentas; de Aranda, Valdir Fernandes; Colombini, Marjorie Paris; Mangueira, Cristóvão Luis Pitangueira; Katz, Marcelo; Faulhaber, Adriana Caschera Leme; Mendes, Claudio Ernesto Albers; Ferreira, Carlos Eduardo dos Santos; França, Carolina Nunes; Guerra, João Carlos de Campos

2017-01-01

Rivaroxaban is an oral direct factor Xa inhibitor, therapeutically indicated in the treatment of thromboembolic diseases. As other new oral anticoagulants, routine monitoring of rivaroxaban is not necessary, but important in some clinical circumstances. In our study a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was validated to measure rivaroxaban plasmatic concentration. Our method used a simple sample preparation, protein precipitation, and a fast chromatographic run. It was developed a precise and accurate method, with a linear range from 2 to 500 ng/mL, and a lower limit of quantification of 4 pg on column. The new method was compared to a reference method (anti-factor Xa activity) and both presented a good correlation (r = 0.98, p highly correlated and should be used as clinical tools for drug monitoring. The method was applied successfully in a group of 49 real-life patients, which allowed an accurate determination of rivaroxaban in peak and trough levels. PMID:28170419

Chitosan-capped gold nanoparticles for selective and colorimetric sensing of heparin

International Nuclear Information System (INIS)

Chen, Zhanguang; Wang, Zhen; Chen, Xi; Xu, Haixiong; Liu, Jinbin

2013-01-01

In this contribution, novel chitosan-stabilized gold nanoparticles (AuNPs) were prepared by mixing chitosan with citrate-reductive AuNPs under appropriate conditions. The as-prepared chitosan-stabilized AuNPs were positively charged and highly stably dispersed in aqueous solution. They exhibited weak resonance light scattering (RLS) intensity and a wine red color. In addition, the chitosan-stabilized AuNPs were successfully utilized as novel sensitive probes for the detection of heparin for the first time. It was found that the addition of heparin induced a strong increase of RLS intensity for AuNPs and the color change from red to blue. The increase in RLS intensity and the color change of chitosan-stabilized AuNPs caused by heparin allowed the sensitive detection of heparin in the range of 0.2–60 μM (∼6.7 U/mL). The detection limit for heparin is 0.8 μM at a signal-to-noise ratio of 3. The present sensor for heparin detection possessed a low detection limit and wide linear range. Additionally, the proposed method was also applied to the detection of heparin in biological media with satisfactory results

Chitosan-capped gold nanoparticles for selective and colorimetric sensing of heparin

Energy Technology Data Exchange (ETDEWEB)

Chen, Zhanguang, E-mail: kqlu@stu.edu.cn; Wang, Zhen; Chen, Xi [Shantou University, Department of Chemistry (China); Xu, Haixiong [Shantou Central Hospital, Affiliated Shantou Hospital of SUN YAT-SEN University (China); Liu, Jinbin [University of Texasat Dallas, Department of Chemistry (United States)

2013-09-15

In this contribution, novel chitosan-stabilized gold nanoparticles (AuNPs) were prepared by mixing chitosan with citrate-reductive AuNPs under appropriate conditions. The as-prepared chitosan-stabilized AuNPs were positively charged and highly stably dispersed in aqueous solution. They exhibited weak resonance light scattering (RLS) intensity and a wine red color. In addition, the chitosan-stabilized AuNPs were successfully utilized as novel sensitive probes for the detection of heparin for the first time. It was found that the addition of heparin induced a strong increase of RLS intensity for AuNPs and the color change from red to blue. The increase in RLS intensity and the color change of chitosan-stabilized AuNPs caused by heparin allowed the sensitive detection of heparin in the range of 0.2-60 {mu}M ({approx}6.7 U/mL). The detection limit for heparin is 0.8 {mu}M at a signal-to-noise ratio of 3. The present sensor for heparin detection possessed a low detection limit and wide linear range. Additionally, the proposed method was also applied to the detection of heparin in biological media with satisfactory results.

Potentiation of C1-esterase inhibitor by heparin and interactions with C1s protease as assessed by surface plasmon resonance.

Science.gov (United States)

Rajabi, Mohsen; Struble, Evi; Zhou, Zhaohua; Karnaukhova, Elena

2012-01-01

Human C1-esterase inhibitor (C1-INH) is a multifunctional plasma protein with a wide range of inhibitory and non-inhibitory properties, mainly recognized as a key down-regulator of the complement and contact cascades. The potentiation of C1-INH by heparin and other glycosaminoglycans (GAGs) regulates a broad spectrum of C1-INH activities in vivo both in normal and disease states. SCOPE OF RESEARCH: We have studied the potentiation of human C1-INH by heparin using Surface Plasmon Resonance (SPR), circular dichroism (CD) and a functional assay. To advance a SPR for multiple-unit interaction studies of C1-INH we have developed a novel (consecutive double capture) approach exploring different immobilization and layout. Our SPR experiments conducted in three different design versions showed marked acceleration in C1-INH interactions with complement protease C1s as a result of potentiation of C1-INH by heparin (from 5- to 11-fold increase of the association rate). Far-UV CD studies suggested that heparin binding did not alter C1-INH secondary structure. Functional assay using chromogenic substrate confirmed that heparin does not affect the amidolytic activity of C1s, but does accelerate its consumption due to C1-INH potentiation. This is the first report that directly demonstrates a significant acceleration of the C1-INH interactions with C1s due to heparin by using a consecutive double capture SPR approach. The results of this study may be useful for further C-INH therapeutic development, ultimately for the enhancement of current C1-INH replacement therapies. Published by Elsevier B.V.

Sensitive and selective turn off-on fluorescence detection of heparin based on the energy transfer platform using the BSA-stabilized Au nanoclusters/amino-functionalized graphene oxide hybrids.

Science.gov (United States)

Lan, Jing; Zou, Hong Yan; Wang, Qiang; Zeng, Ping; Li, Yuan Fang; Huang, Cheng Zhi

2016-12-01

An ultra-sensitive and selective turn off-on fluorescence detection of heparin based on the energy transfer in the BSA-stabilized gold nanoclusters/amino-functionalized graphene oxide (BSA-AuNCs/NH 2 -GO) hybrids was successfully realized. The BSA-AuNCs containing amounts of carboxyl groups could be absorbed on the surface of NH 2 -GO through the electrostatic interaction, which resulted in the fluorescence quenching of BSA-AuNCs with high efficiency. However, heparin, possessing high density of negative charge, could compete with BSA-AuNCs to bind NH 2 -GO and block the energy transfer from BSA-AuNCs to NH 2 -GO. The fluorescence recovery of BSA-AuNCs was closely related to the amount of heparin and there was a good linear relationship between fluorescence recovery of BSA-AuNCs and heparin over the range of 100ng/mL to 30μg/mL with a detection limit of 40ng/mL. What's more, the fluorescence assay was successfully applied for heparin sensing in human serums and intracellular imaging. Copyright © 2016 Elsevier B.V. All rights reserved.

Prevention of fatal postoperative pulmonary embolism by low doses of heparin. An international multicentre trial.

Science.gov (United States)

1975-07-12

The efficacy of low-dose heparin in preventing fatal postoperative pulmonary embolism has been investigated in a multicentre prospective randomised trial. 4121 patients over the age of forty years undergoing a variety of elective major surgical procedures were included in the trial; 2076 of these were in the control group and 2045 patients received heparin. The two groups were well matched for age, sex, weight, blood-group, and other factors which could predispose to the development of venous thromboembolism. 180 (4-4 %) patients died during the postoperative period, 100 in the control and 80 in the heparin group: 72% of deaths in the control and 66% in the heparin group had necropsy examination. 16 patients in the control group and 2 in the heparin group were found at necropsy to have died due to acute massive pulmonary embolism (P smaller than 0-005). In addition, emboli found at necropsy in 6 patients in the control group and 3 in the heparin group were considered either contributory to death or an incidental finding since death in these patients was attributed to other causes. Taking all pulmonary emboli together, the findings were again significant (P smaller than 0-005). Of 1292 patients in whom the 125-I-fibrinogen test was performed to detect deep-vein thrombosis (D.V.T.) 667 were in the control group and 625 in the heparin group. The frequency of isotopic D.V.T. was reduced from 24-6% in the control group 7-7% in the heparin group (P smaller 0-005). In 30 patients D.V.T. was detected at necropsy; 24 in the control and 6 in the heparin group (P smaller 0-005). 32 patients in the control group and 11 in the heparin group developed clinically diagnosed D.V.T. which was confirmed by venography (P smaller than 0-005). In addition, 24 patients in the control and 8 in the heparin group were treated for clinically suspected pulmonary emoblism. The difference in the number of patients requiring treatment for D.V.T. and/or pulmonary embolism in the two groups was

DEGRADATION AND INTRAHEPATIC COMPATIBILITY OF ALBUMIN-HEPARIN CONJUGATE MICROSPHERES

NARCIS (Netherlands)

CREMERS, HFM; WOLF, RFE; BLAAUW, EH; SCHAKENRAAD, JM; LAM, KH; NIEUWENHUIS, P; VERRIJK, R; KWON, G; BAE, YH; KIM, SW; FEIJEN, J

The in vitro degradation properties of glutaraldehyde cross-linked albumin and albumin-heparin conjugate microspheres (AMS and AHCMS respectively) were evaluated using light microscopy, turbidity measurements and heparin release determinations, showing that the microspheres are degraded by

The heparin-binding domain of HB-EGF as an efficient cell-penetrating peptide for drug delivery.

Science.gov (United States)

Luo, Zhao; Cao, Xue-Wei; Li, Chen; Wu, Miao-Dan; Yang, Xu-Zhong; Zhao, Jian; Wang, Fu-Jun

2016-11-01

Cell-penetrating peptides (CPPs) have been shown to be potential drug carriers for cancer therapy. The inherently low immunogenicity and cytotoxicity of human-derived CPPs make them more suitable for intracellular drug delivery compared to other delivery vehicles. In this work, the protein transduction ability of a novel CPP (termed HBP) derived from the heparin-binding domain of HB-EGF was evaluated. Our data shows, for the first time, that HBP possesses similar properties to typical CPPs and is a potent drug delivery vector for improving the antitumor activity of impermeable MAP30. The intrinsic bioactivities of recombinant MAP30-HBP were well preserved compared to those of free MAP30. Furthermore, HBP conjugated to the C-terminus of MAP30 promoted the cellular uptake of recombinant MAP30-HBP. Moreover, the fusion of HBP to MAP30 gave rise to significantly enhanced cytotoxic effects in all of the tumor cell lines tested. In HeLa cells, this cytotoxicity was mainly caused by the induction of cell apoptosis. Further investigation revealed that HBP enhanced MAP30-induced apoptosis through the activation of the mitochondrial- and death receptor-mediated signaling pathways. In addition, the MAP30-HBP fusion protein caused more HeLa cells to become arrested in S phase compared to MAP30 alone. These results highlight the MAP30-HBP fusion protein as a promising drug candidate for cancer therapy and demonstrate HBP, a novel CPP derived from human HB-EGF, as a new potential vector for antitumor drug delivery. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

Increased accuracy in heparin and protamine administration decreases bleeding

DEFF Research Database (Denmark)

Runge, Marx; Møller, Christian H; Steinbrüchel, Daniel A

2009-01-01

Three to 5 percent of the patients undergoing cardiac surgery are reoperated because of bleeding. When a surgical cause can be excluded, heparin/protamine mismatch may be considered. Insufficient reversal of heparin and overdosing of protamine may cause postoperative bleeding. The purpose......). A reduced number of patients needed blood transfusions in the RxDx group, although this was not statistically significant (19% vs. 38%, respectively; p = .13). Initial heparin dose was significantly reduced in the RxDx group (250 mg; range, 100-375 mg) compared with the control group (300 mg; range, 200...

Prophylaxis of postoperative thromboembolism with low molecular weight heparins

DEFF Research Database (Denmark)

Jørgensen, L N; Wille-Jørgensen, P; Hauch, O

1993-01-01

To evaluate the thromboprophylactic use of low molecular weight heparins (LMWHs), publications from 27 orthopaedic trials and 35 studies of patients undergoing general or gynaecological surgery were scrutinized and subjected to a partial meta-analysis. In orthopaedic surgery, LMWHs were superior...... to placebo or dextran and at least as efficient as unfractionated heparin in the prevention of deep vein thrombosis (DVT). Compared with unfractionated heparin, one of the LMWH preparations significantly reduced the total incidence of DVT. The rate of non-fatal pulmonary embolism was 0.49 per cent...

Transfer of heparin polyion across a polarized water/ionic liquid membrane interface

Czech Academy of Sciences Publication Activity Database

Langmaier, Jan; Samec, Zdeněk; Samcová, E.; Tůma, P.

2012-01-01

Roč. 24, OCT 2012 (2012), s. 25-27 ISSN 1388-2481 R&D Projects: GA ČR GAP206/11/0707 Institutional support: RVO:61388955 Keywords : heparin polyion * ionic liquid membrane * amperometric detection Subject RIV: CG - Electrochemistry Impact factor: 4.425, year: 2012

Determination of rivaroxaban in patient's plasma samples by anti-Xa chromogenic test associated to High Performance Liquid Chromatography tandem Mass Spectrometry (HPLC-MS/MS).

Science.gov (United States)

Derogis, Priscilla Bento Matos; Sanches, Livia Rentas; de Aranda, Valdir Fernandes; Colombini, Marjorie Paris; Mangueira, Cristóvão Luis Pitangueira; Katz, Marcelo; Faulhaber, Adriana Caschera Leme; Mendes, Claudio Ernesto Albers; Ferreira, Carlos Eduardo Dos Santos; França, Carolina Nunes; Guerra, João Carlos de Campos

2017-01-01

Rivaroxaban is an oral direct factor Xa inhibitor, therapeutically indicated in the treatment of thromboembolic diseases. As other new oral anticoagulants, routine monitoring of rivaroxaban is not necessary, but important in some clinical circumstances. In our study a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was validated to measure rivaroxaban plasmatic concentration. Our method used a simple sample preparation, protein precipitation, and a fast chromatographic run. It was developed a precise and accurate method, with a linear range from 2 to 500 ng/mL, and a lower limit of quantification of 4 pg on column. The new method was compared to a reference method (anti-factor Xa activity) and both presented a good correlation (r = 0.98, p plasma samples for rivaroxaban measurement by HPLC-MS/MS without interferences. The chromogenic and HPLC-MS/MS methods were highly correlated and should be used as clinical tools for drug monitoring. The method was applied successfully in a group of 49 real-life patients, which allowed an accurate determination of rivaroxaban in peak and trough levels.

Heparin release from thermosensitive polymer coatings: in vivo studies

NARCIS (Netherlands)

Gutowska, Anna; Bae, You Han; Jacobs, Harvey; Mohammad, Fazal; Mix, Donald; Feijen, Jan; Kim, Sung Wan

1995-01-01

Biomer/poly(N-isopropylacrylamide)/[poly(NiPAAm)] thermosensitive polymer blends were prepared and their application as heparin-releasing polymer coatings for the prevention of surface-induced thrombosis was examined. The advantage of using poly(NiPAAm)-based coatings as heparin-releasing polymers

Poly(vinyl alcohol)-heparin hydrogels as sensor catheter membranes

NARCIS (Netherlands)

Brinkman, E.; van der Does, L.; Bantjes, A.

1991-01-01

Poly(vinyl alcohol)-heparin hydrogels with varying water content were synthesized for use as sensor catheter membranes. Films were cast from aqueous mixtures of poly(viny) alcohol) (PVA), a photosensitive cross-linker p-diazonium diphenyl amine polymer (PA), glutaraldehyde (GA) and heparin. After

From Farm to Pharma: An Overview of Industrial Heparin Manufacturing Methods.

Science.gov (United States)

van der Meer, Jan-Ytzen; Kellenbach, Edwin; van den Bos, Leendert J

2017-06-21

The purification of heparin from offal is an old industrial process for which commercial recipes date back to 1922. Although chemical, chemoenzymatic, and biotechnological alternatives for this production method have been published in the academic literature, animal-tissue is still the sole source for commercial heparin production in industry. Heparin purification methods are closely guarded industrial secrets which are not available to the general (scientific) public. However by reviewing the academic and patent literature, we aim to provide a comprehensive overview of the general methods used in industry for the extraction of heparin from animal tissue.

Apixaban versus enoxaparin for thromboprophylaxis after knee replacement (ADVANCE-2): a randomised double-blind trial

DEFF Research Database (Denmark)

Lassen, Michael Rud; Raskob, Gary E; Gallus, Alexander

2010-01-01

efficacy and safety of these drugs after elective total knee replacement. METHODS: In ADVANCE-2, a multicentre, randomised, double-blind phase 3 study, patients undergoing elective unilateral or bilateral total knee replacement were randomly allocated through an interactive central telephone system......BACKGROUND: Low-molecular-weight heparins such as enoxaparin are preferred for prevention of venous thromboembolism after major joint replacement. Apixaban, an orally active factor Xa inhibitor, might be as effective, have lower bleeding risk, and be easier to use than is enoxaparin. We assessed...

Low-thrombogenic fibrin-heparin coating promotes in vitro endothelialization

Czech Academy of Sciences Publication Activity Database

Kaplan, Ondřej; Hierlemann, T.; Krajewski, S.; Kurz, J.; Nevoralová, Martina; Houska, Milan; Riedel, Tomáš; Riedelová, Zuzana; Zárubová, Jana; Wendel, H. P.; Brynda, Eduard

2017-01-01

Roč. 105, č. 11 (2017), s. 2995-3005 ISSN 1549-3296 R&D Projects: GA MZd(CZ) NV15-29153A Institutional support: RVO:61389013 ; RVO:67985823 Keywords : fibrin-heparin coating * hemocompatibility * endothelialization Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery; FA - Cardiovascular Diseases incl. Cardiotharic Surgery (FGU-C) OBOR OECD: Cardiac and Cardiovascular systems; Cardiac and Cardiovascular systems (FGU-C) Impact factor: 3.076, year: 2016

P-selectin- and heparanase-dependent antimetastatic activity of non-anticoagulant heparins.

Science.gov (United States)

Hostettler, Nina; Naggi, Annamaria; Torri, Giangiacomo; Ishai-Michaeli, Riva; Casu, Benito; Vlodavsky, Israel; Borsig, Lubor

2007-11-01

Vascular cell adhesion molecules, P- and L-selectins, facilitate metastasis of cancer cells in mice by mediating interactions with platelets, endothelium, and leukocytes. Heparanase is an endoglycosidase that degrades heparan sulfate of extracellular matrix, thereby promoting tumor invasion and metastasis. Heparin is known to efficiently attenuate metastasis in different tumor models. Here we identified modified, nonanticoagulant species of heparin that specifically inhibit selectin-mediated cell-cell interactions, heparanase enzymatic activity, or both. We show that selective inhibition of selectin interactions or heparanase with specific heparin derivatives in mouse models of MC-38 colon carcinoma and B16-BL6 melanoma attenuates metastasis. Selectin-specific heparin derivatives attenuated metastasis of MC-38 carcinoma, but heparanase-specific derivatives had no effect, in accordance with the virtual absence of heparanase activity in these cells. Heparin derivatives had no further effect on metastasis in mice deficient in P- and L-selectin, indicating that selectins are the primary targets of heparin antimetastatic activity. Selectin-specific and heparanase-specific derivatives attenuated metastasis of B16-BL6 melanomas to a similar extent. When mice were injected with a derivative containing both heparanase and selectin inhibitory activity, no additional attenuation of metastasis could be observed. Thus, selectin-specific heparin derivatives efficiently attenuated metastasis of both tumor cell types whereas inhibition of heparanase led to reduction of metastasis only in tumor cells producing heparanase.

Amblyomma americanum tick calreticulin binds C1q but does not inhibit activation of the classical complement cascade.

Science.gov (United States)

Kim, Tae Kwon; Ibelli, Adriana Mércia Guaratini; Mulenga, Albert

2015-02-01

In this study we characterized Amblyomma americanum (Aam) tick calreticulin (CRT) homolog in tick feeding physiology. In nature, different tick species can be found feeding on the same animal host. This suggests that different tick species found feeding on the same host can modulate the same host anti-tick defense pathways to successfully feed. From this perspective it's plausible that different tick species can utilize universally conserved proteins such as CRT to regulate and facilitate feeding. CRT is a multi-functional protein found in most taxa that is injected into the vertebrate host during tick feeding. Apart from it's current use as a biomarker for human tick bites, role(s) of this protein in tick feeding physiology have not been elucidated. Here we show that annotated functional CRT amino acid motifs are well conserved in tick CRT. However our data show that despite high amino acid identity levels to functionally characterized CRT homologs in other organisms, AamCRT is apparently functionally different. Pichia pastoris expressed recombinant (r) AamCRT bound C1q, the first component of the classical complement system, but it did not inhibit activation of this pathway. This contrast with reports of other parasite CRT that inhibited activation of the classical complement pathway through sequestration of C1q. Furthermore rAamCRT did not bind factor Xa in contrast to reports of parasite CRT binding factor Xa, an important protease in the blood clotting system. Consistent with this observation, rAamCRT did not affect plasma clotting or platelet aggregation. We discuss our findings in the context of tick feeding physiology.

Heparin modulates human intestinal smooth muscle (HISM) cell proliferation and matrix production

International Nuclear Information System (INIS)

Graham, M.; Perr, H.; Drucker, D.E.; Diegelmann, R.F.

1986-01-01

(HISM) cell proliferation and collagen production may play a role in the pathogenesis of intestinal stricture in Crohn's disease. The present studies were performed to evaluate the effects of heparin, a known modulator of vascular smooth muscle cells, on HISM cell proliferation and collagen production. Heparin (100 μg/ml) was added daily to HISM cell cultures for cell proliferation studies and for 24 hours at various time points during culture for collagen synthesis studies. Collagen synthesis was determined by the uptake of 3 H proline into collagenase-sensitive protein. Heparin completely inhibited cell proliferation for 7 days, after which cell numbers increased but at a slower rate than controls. Cells released from heparin inhibition demonstrated catch-up growth to control levels. Collagen production was significantly inhibited by 24 hours exposure to heparin but only at those times during culture when collagen synthesis was maximal (8 to 12 days). Non-collagen protein synthesis was inhibited by heparin at all time points during culture. Heparin through its modulation of HISM cells may play an important role in the control of the extracellular matrix of the intestinal wall

Obstacles in the diagnostics and therapy of heparin-induced thrombocytopenia.

Science.gov (United States)

Antonijević, Nebojsa M; Radovanović, Nebojsa; Obradović, Slobodan; Vucelić, Dragica; Stojanović, Bojan; Miković, Danijela; Kovac, Mirjana; Kocica, Tina; Tadić, Svetlana; Antonijević, Irina; Drasković, Snezana; Djordjević, Valentina; Calija, Branko; Perunicić, Jovan; Vasiljević, Zorana

2010-01-01

An immune-mediated, severe, acquired prothrombotic disorder, heparin-induced thrombocytopenia type II (HIT II) occurs in 0.5-5% of patients exposed to unfractionated heparin longer than 5-7 days. Arterial and venous thromboses are induced by HIT II in about 35-50% of patients. Typical death rate for HIT is about 29%, while 21% of HIT patients result in amputation of a limb. The trend towards the occurrence of HIT due to the administration of low molecular weight heparins (LMWH) taking ever conspicuous place in the standard venous thromboembolism (VTE) prophylaxis has been more frequently observed recently. It is considered that LMWH may cause HIT II in about 0.25-1%. The need for further modification of HIPA assays with LMWH has been imposed in the HIT laboratory diagnostics, heretofore overburdened with complexity. There are several constantly opposing problems arising in HIT laboratory diagnostics, one of which is that in a certain number of patients immunologic assays detect nonpathogenic antibodies (mainly IgM or IgA heparin-PF4 antibodies) while, on the other hand, the occurrence of HIT pathogenetically mediated by minor antigens (neutrophil-activating peptide 2 or interleukin 8) may be neglected in certain cases. The following factors play an important role in the interpretation of each laboratory HIT assays performed: 1. correlation with HIT clinical probability test, the best known of which is 4T'score, 2. the interpretation of the laboratory findings dependent on the time of the thrombocytopenia onset, as well as 3. the sensitivity and specificity of each test respectively. The HIT diagnostics in the presence of other comorbid states which may also induce thrombocytopenia, more precisely known as pseudo HIT (cancer, sepsis, disseminated intravascular coagulation, pulmonary embolism, antiphospholipid syndrome, etc), represents a specific clinical problem.

Heparin-Based Nanoparticles: An Overview of Their Applications

Directory of Open Access Journals (Sweden)

Maria del Pilar Rodriguez-Torres

2018-01-01

Full Text Available This review deals with nanoparticles synthesized using heparin. Such nanoparticles have been widely studied since a long time ago, obtaining satisfactory outcomes. An outstanding aspect of these nanoparticles is that they possess good biocompatible characteristics, and since heparin is produced in the human body within the mast cells, this makes these nanoparticles useful for future applications like imaging, disease and cancer treatment, and antibacterial activity. They can also be used for applications that are not oriented directly to the medical and biological areas such as in the case of analyte detection in aqueous solution, although such studies are very few. These nanoparticles synthesis is mainly through wet chemistry methods, using heparin that could have been modified or not.

ABFs, a family of ABA-responsive element binding factors.

Science.gov (United States)

Choi, H; Hong, J; Ha, J; Kang, J; Kim, S Y

2000-01-21

Abscisic acid (ABA) plays an important role in environmental stress responses of higher plants during vegetative growth. One of the ABA-mediated responses is the induced expression of a large number of genes, which is mediated by cis-regulatory elements known as abscisic acid-responsive elements (ABREs). Although a number of ABRE binding transcription factors have been known, they are not specifically from vegetative tissues under induced conditions. Considering the tissue specificity of ABA signaling pathways, factors mediating ABA-dependent stress responses during vegetative growth phase may thus have been unidentified so far. Here, we report a family of ABRE binding factors isolated from young Arabidopsis plants under stress conditions. The factors, isolated by a yeast one-hybrid system using a prototypical ABRE and named as ABFs (ABRE binding factors) belong to a distinct subfamily of bZIP proteins. Binding site selection assay performed with one ABF showed that its preferred binding site is the strong ABRE, CACGTGGC. ABFs can transactivate an ABRE-containing reporter gene in yeast. Expression of ABFs is induced by ABA and various stress treatments, whereas their induction patterns are different from one another. Thus, a new family of ABRE binding factors indeed exists that have the potential to activate a large number of ABA/stress-responsive genes in Arabidopsis.

New strategies for the treatment of acute venous thromboembolism.

Science.gov (United States)

Prandoni, Paolo; Lensing, Anthonie W A; Pesavento, Raffaele

2006-11-01

In recent years, new opportunities have emerged that have the potential to change rapidly the therapeutic scenario of patients with acute venous thromboembolism (VTE). Selected patients with deep vein thrombosis (DVT) can be treated effectively and safely at home with fixed doses of low molecular weight heparins. The prompt administration of compression elastic stockings in addition to anticoagulant drugs in patients with acute DVT has the potential to halve the rate of late postthrombotic sequelae. The long-term use of low molecular weight heparins is likely to be more effective than oral anticoagulants for the secondary prevention of VTE in patients with advanced malignancy. Patients with pulmonary embolism and right ventricular dysfunction might benefit from the early administration of thrombolytic drugs in combination with heparin to a greater extent than from heparin alone. Despite an impressive amount of clinical information on the proper duration of oral anticoagulants in patients with unprovoked VTE, the optimal long-term treatment of these patients remains undefined. Finally, new categories of drugs are emerging that have the potential to replace conventional anticoagulants in the near future. They include compounds that inhibit factor Xa or thrombin.

Surface Modification of Titanium with BMP-2/GDF-5 by a Heparin Linker and Its Efficacy as a Dental Implant

Directory of Open Access Journals (Sweden)

Dae Hyeok Yang

2017-01-01

Full Text Available In this study, we prepared human bone morphogenetic protein-2 (hBMP-2/human growth and differentiation factor-5 (hGDF-5-coated titanium (Ti disc and screw types for controlled release of the growth factors (GFs. The two growth factors were coated onto Ti with a smooth surface using their specific interaction with heparin, because they have heparin binding sites in their molecular structures. Efficacy of the two growth factor-coated Ti for enhancement of bone formation and osseointegration was compared to pristine Ti, and hBMP-2- and hGDF-5-coated Ti in vivo. The surface chemical composition, surface morphology, and wettability characteristics of the metal samples were determined by X-ray photoelectron spectroscopy (XPS, scanning electron microscopy (SEM, and contact angle measurement, respectively. The initial burst of hBMP-2, hGDF-5, and their combination, occurred within one day of the release study, resulting in 12.5%, 4.5%, and 13.5%/3.2%, and then there was a sustained, even release of these two growth factors from the coated metal for 30 days. In vitro tests revealed that MC3T3-E1 cells cultured on the two growth factor-coated Ti had a higher proliferation rate and a higher activity for alkaline phosphatase (ALP, which led to a larger amount of calcium deposition and larger expressions of type I collagen (COL 1, ALP, and osteocalcin (OCN mRNAs. In vivo animal tests using ten white New Zealand rabbits showed that the two growth factor-coated Ti enhanced bone formation and osseointegration at the interface between the implants and host bone. In addition, histological evaluation showed that bone remodeling, including bone formation by osteoblasts and bone resorption by osteoclasts, actively occurred between the two growth factor-coated Ti and host bone. Consequently, it is suggested that Ti surface modification with the combination of hBMP-2 and hGDF-5 for the two growth factor-coated Ti implants can improve the clinical properties of

Anticoagulant effect of low molecular weight heparin on central ...

African Journals Online (AJOL)

Purpose: To analyse the effect of low molecular weight heparin on venous catheters in haemodialysis patients. Methods: This study included 140 eligible patients who were randomly and evenly divided into two groups, viz, a study group that received low molecular weight heparin and a control group that received ...

Modulation of DNA binding by gene-specific transcription factors.

Science.gov (United States)

Schleif, Robert F

2013-10-01

The transcription of many genes, particularly in prokaryotes, is controlled by transcription factors whose activity can be modulated by controlling their DNA binding affinity. Understanding the molecular mechanisms by which DNA binding affinity is regulated is important, but because forming definitive conclusions usually requires detailed structural information in combination with data from extensive biophysical, biochemical, and sometimes genetic experiments, little is truly understood about this topic. This review describes the biological requirements placed upon DNA binding transcription factors and their consequent properties, particularly the ways that DNA binding affinity can be modulated and methods for its study. What is known and not known about the mechanisms modulating the DNA binding affinity of a number of prokaryotic transcription factors, including CAP and lac repressor, is provided.

New Functional Tools for Antithrombogenic Activity Assessment of Live Surface Glycocalyx.

Science.gov (United States)

Dimitrievska, Sashka; Gui, Liqiong; Weyers, Amanda; Lin, Tylee; Cai, Chao; Wu, Wei; Tuggle, Charles T; Sundaram, Sumati; Balestrini, Jenna L; Slattery, David; Tchouta, Lise; Kyriakides, Themis R; Tarbell, John M; Linhardt, Robert J; Niklason, Laura E

2016-09-01

It is widely accepted that the presence of a glycosaminoglycan-rich glycocalyx is essential for endothelialized vasculature health; in fact, a damaged or impaired glycocalyx has been demonstrated in many vascular diseases. Currently, there are no methods that characterize glycocalyx functionality, thus limiting investigators' ability to assess the role of the glycocalyx in vascular health. We have developed novel, easy-to-use, in vitro assays that directly quantify live endothelialized surface's functional heparin weights and their anticoagulant capacity to inactivate Factor Xa and thrombin. Using our assays, we characterized 2 commonly used vascular models: native rat aorta and cultured human umbilical vein endothelial cell monolayer. We determined heparin contents to be ≈10 000 ng/cm(2) on the native aorta and ≈10-fold lower on cultured human umbilical vein endothelial cells. Interestingly, human umbilical vein endothelial cells demonstrated a 5-fold lower anticoagulation capacity in inactivating both Factor Xa and thrombin relative to native aortas. We verified the validity and accuracy of the novel assays developed in this work using liquid chromatography-mass spectrometry analysis. Our assays are of high relevance in the vascular community because they can be used to establish the antithrombogenic capacity of many different types of surfaces such as vascular grafts and transplants. This work will also advance the capacity for glycocalyx-targeting therapeutics development to treat damaged vasculatures. © 2016 American Heart Association, Inc.

Naked eye detection of infertility based on sperm protamine-induced aggregation of heparin gold nanoparticles.

Science.gov (United States)

Vidya, Raj; Saji, Alex

2018-05-01

The development of an easy to use, one-pot, environmentally friendly, non-invasive and label-free colorimetric probe for the determination of semen protamines, the biochemical marker of male fertility, using heparin gold nanoparticles (HAuNPs) is presented. The affinity of HAuNPs for protamines was due to the electrostatic interactions between polycationic protamine and polyanionic heparin. The binding of HAuNPs to protamine was characterized by variation in the plasmon absorption spectra followed by a visibly observable colour change of the solution from red to blue. We observed a red shift in the plasmon peak and the method exhibited linearity in the range of 10-70 ng/mL with a detection limit of 5 ng/mL, which is much lower than that reported for colorimetric sensors of protamine. The colour change and the variation in the absorbance of HAuNPs were highly specific for protamines in the presence of different interfering compounds and the method was successfully applied for determining protamine in real samples of semen and serum. Rather than a quantitative estimation, it seems that the method provides a quick screening between a large array of positive and negative samples and, moreover, it maintains the privacy of the user. The method appears to be simple and would be very useful in third-world countries where high-tech diagnostic aids are inaccessible to the majority of the population. Graphical Abstract Heparin gold nanoparticles aided visual detection of infertility.

Species of /sup 67/Ga-binding acid mucopolysaccharide in liver

Energy Technology Data Exchange (ETDEWEB)

Ando, A.; Ando, I.

1985-01-01

It was determined from measuring neutral saccharide in the structure that the principal /sup 67/Ga-binding acid mucopolysaccharide in liver was keratan sulfate and/or keratan polysulfate. On the other hand, it was clarified from the results of mucopolysaccharase treatment that the main /sup 67/Ga-binding acid mucopolysaccharide in liver was neither keratan sulfate, heparan sulfate, heparin, nor chondroitin sulfate A, B and C. Based on the present results, it was deduced that the main /sup 67/Ga-binding acid mucopolysaccharide in liver was keratan polysulfate.

Distribution of heparin-/sup 67/Ga in tumor-bearing rats

Energy Technology Data Exchange (ETDEWEB)

Hiraki, T; Ando, A; Sanada, S [Kanazawa Univ. (Japan). School of Paramedicine; Ando, I; Hisada, K

1976-06-01

Heparin is a kind of acidic mucopolysaccharide. The distribution of heparin-/sup 67/Ga complex in tumor-bearing rats was investigated by administering it to rats into which Yoshida sarcoma had been transplanted subcutaneously. These rats were sacrificed at 10 minutes, 30 minutes, 1 hour and 3 hours after injection. Radioactivity of the tumor, blood, muscle, liver, kidney, spleen and urine were measured with a well-type scintillation counter. Retention values in these organs and excretion rates in the urine were calculated. Excretion rates (%/dose) of heparin-/sup 67/Ga in 10 min., 30 min., 1 hour, and 3 hours were 38.2%, 67.5%, 79.5% and 78.0%, respectively. From these facts, it was thought that heparin-/sup 67/Ga complex was not suitable for tumor scanning, but that this compound might be a suitable agent for the renal function test.

Involvement of three meningococcal surface-exposed proteins, the heparin-binding protein NhbA, the α-peptide of IgA protease and the autotransporter protease NalP, in initiation of biofilm formation

KAUST Repository

Arenas, Jesús

2012-12-04

Neisseria meningitidis is a common and usually harmless inhabitant of the mucosa of the human nasopharynx, which, in rare cases, can cross the epithelial barrier and cause meningitis and sepsis. Biofilm formation favours the colonization of the host and the subsequent carrier state. Two different strategies of biofilm formation, either dependent or independent on extracellular DNA (eDNA), have been described for meningococcal strains. Here, we demonstrate that the autotransporter protease NalP, the expression of which is phase variable, affects eDNA-dependent biofilm formation in N.meningitidis. The effect of NalP was found in biofilm formation under static and flow conditions and was dependent on its protease activity. Cleavage of the heparin-binding antigen NhbA and the α-peptide of IgA protease, resulting in the release of positively charged polypeptides from the cell surface, was responsible for the reduction in biofilm formation when NalP is expressed. Both NhbA and the α-peptide of IgA protease were shown to bind DNA. We conclude that NhbA and the α-peptide of IgA protease are implicated in biofilm formation by binding eDNA and that NalP is an important regulator of this process through the proteolysis of these surface-exposed proteins. © 2012 Blackwell Publishing Ltd.

Involvement of three meningococcal surface-exposed proteins, the heparin-binding protein NhbA, the α-peptide of IgA protease and the autotransporter protease NalP, in initiation of biofilm formation

KAUST Repository

Arenas, Jesú s; Nijland, Reindert; Rodriguez, Francisco J.; Bosma, Tom N. P.; Tommassen, Jan

2012-01-01

Neisseria meningitidis is a common and usually harmless inhabitant of the mucosa of the human nasopharynx, which, in rare cases, can cross the epithelial barrier and cause meningitis and sepsis. Biofilm formation favours the colonization of the host and the subsequent carrier state. Two different strategies of biofilm formation, either dependent or independent on extracellular DNA (eDNA), have been described for meningococcal strains. Here, we demonstrate that the autotransporter protease NalP, the expression of which is phase variable, affects eDNA-dependent biofilm formation in N.meningitidis. The effect of NalP was found in biofilm formation under static and flow conditions and was dependent on its protease activity. Cleavage of the heparin-binding antigen NhbA and the α-peptide of IgA protease, resulting in the release of positively charged polypeptides from the cell surface, was responsible for the reduction in biofilm formation when NalP is expressed. Both NhbA and the α-peptide of IgA protease were shown to bind DNA. We conclude that NhbA and the α-peptide of IgA protease are implicated in biofilm formation by binding eDNA and that NalP is an important regulator of this process through the proteolysis of these surface-exposed proteins. © 2012 Blackwell Publishing Ltd.

Optical sensing of sulfate by polymethinium salt receptors: colorimetric sensor for heparin

Czech Academy of Sciences Publication Activity Database

Bříza, T.; Kejík, Z.; Císařová, I.; Králová, Jarmila; Martásek, P.; Král, V.

2008-01-01

Roč. 16, - (2008), s. 1901-1903 ISSN 1359-7345 R&D Projects: GA AV ČR KAN200200651; GA ČR(CZ) GA203/06/1038 Institutional research plan: CEZ:AV0Z50520514 Keywords : colorimetric sensor * heparin * polymethinium salt Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.340, year: 2008

Computational investigation of potential dosing schedules for a switch of medication from warfarin to rivaroxaban-an oral, direct Factor Xa inhibitor.

Science.gov (United States)

Burghaus, Rolf; Coboeken, Katrin; Gaub, Thomas; Niederalt, Christoph; Sensse, Anke; Siegmund, Hans-Ulrich; Weiss, Wolfgang; Mueck, Wolfgang; Tanigawa, Takahiko; Lippert, Jörg

2014-01-01

The long-lasting anticoagulant effect of vitamin K antagonists can be problematic in cases of adverse drug reactions or when patients are switched to another anticoagulant therapy. The objective of this study was to examine in silico the anticoagulant effect of rivaroxaban, an oral, direct Factor Xa inhibitor, combined with the residual effect of discontinued warfarin. Our simulations were based on the recommended anticoagulant dosing regimen for stroke prevention in patients with atrial fibrillation. The effects of the combination of discontinued warfarin plus rivaroxaban were simulated using an extended version of a previously validated blood coagulation computer model. A strong synergistic effect of the two distinct mechanisms of action was observed in the first 2-3 days after warfarin discontinuation; thereafter, the effect was close to additive. Nomograms for the introduction of rivaroxaban therapy after warfarin discontinuation were derived for Caucasian and Japanese patients using safety and efficacy criteria described previously, together with the coagulation model. The findings of our study provide a mechanistic pharmacologic rationale for dosing schedules during the therapy switch from warfarin to rivaroxaban and support the switching strategies as outlined in the Summary of Product Characteristics and Prescribing Information for rivaroxaban.

Computational investigation of potential dosing schedules for a switch of medication from warfarin to rivaroxaban—an oral, direct Factor Xa inhibitor

Science.gov (United States)

Burghaus, Rolf; Coboeken, Katrin; Gaub, Thomas; Niederalt, Christoph; Sensse, Anke; Siegmund, Hans-Ulrich; Weiss, Wolfgang; Mueck, Wolfgang; Tanigawa, Takahiko; Lippert, Jörg

2014-01-01

The long-lasting anticoagulant effect of vitamin K antagonists can be problematic in cases of adverse drug reactions or when patients are switched to another anticoagulant therapy. The objective of this study was to examine in silico the anticoagulant effect of rivaroxaban, an oral, direct Factor Xa inhibitor, combined with the residual effect of discontinued warfarin. Our simulations were based on the recommended anticoagulant dosing regimen for stroke prevention in patients with atrial fibrillation. The effects of the combination of discontinued warfarin plus rivaroxaban were simulated using an extended version of a previously validated blood coagulation computer model. A strong synergistic effect of the two distinct mechanisms of action was observed in the first 2–3 days after warfarin discontinuation; thereafter, the effect was close to additive. Nomograms for the introduction of rivaroxaban therapy after warfarin discontinuation were derived for Caucasian and Japanese patients using safety and efficacy criteria described previously, together with the coagulation model. The findings of our study provide a mechanistic pharmacologic rationale for dosing schedules during the therapy switch from warfarin to rivaroxaban and support the switching strategies as outlined in the Summary of Product Characteristics and Prescribing Information for rivaroxaban. PMID:25426077

Quantitation of heparosan with heparin lyase III and spectrophotometry.

Science.gov (United States)

Huang, Haichan; Zhao, Yingying; Lv, Shencong; Zhong, Weihong; Zhang, Fuming; Linhardt, Robert J

2014-02-15

Heparosan is Escherichia coli K5 capsule polysaccharide, which is the key precursor for preparing bioengineered heparin. A rapid and effective quantitative method for detecting heparosan is important in the large-scale production of heparosan. Heparin lyase III (Hep III) effectively catalyzes the heparosan depolymerization, forming unsaturated disaccharides that are measurable using a spectrophotometer at 232 nm. We report a new method for the quantitative detection of heparosan with heparin lyase III and spectrophotometry that is safer and more specific than the traditional carbazole assay. In an optimized detection system, heparosan at a minimum concentration of 0.60 g/L in fermentation broth can be detected. Copyright © 2013 Elsevier Inc. All rights reserved.

Quantitative determination of heparin levels in serum with microtiter plate-format optode

International Nuclear Information System (INIS)

Kim, Sung Bae; Kang, Tae Young; Cha, Geun Sig; Nam, Hakhyun

2006-01-01

A new assay method has been developed for the quantitative determination of heparin in serum using a microtiter plate-format optode (MPO). Heparin and proton in physiological sample are favorably co-extracted into the solvent polymeric optode membrane containing both cationic lipophilic additive, tridodecylmethyl ammonium chloride (TDMAC), and proton-selective ionophore, 3-hydroxy-4-(4-nitrophenylazo)-phenyloctadecanoate (ETH 2412), resulting in the absorbance change of the membrane to varying heparin levels. The optimized MPO composition contains low polymer-to-plasticizer ratio compared to those of conventional ion-selective optodes or electrodes, i.e., poly(vinyl chloride) (20.0)/dioctylsebacate (76.3)/ETH 2412 (1.7)/TDMAC (1.0) (wt.%): it resulted in a quantitative response to heparin from 0 to 15 unit/mL in serum with high sensitivity. The heparin-protamine titration on the MPO could provide rapid and precise determination of heparin. It was shown that the heparin levels in serum sample could be determined from the rate of absorbance change over time (ΔA/Δt); this method was more effective than the direct absorbance measurement in minimizing the interferences from color and turbidity of serum samples. MPO has been developed as a high throughput and convenient disposable sensing device, and may find a wide application in the determination of polyions and charged macromolecules

Cancer Cell Adhesion and Metastasis: Selectins, Integrins, and the Inhibitory Potential of Heparins

Directory of Open Access Journals (Sweden)

Gerd Bendas

2012-01-01

Full Text Available Cell adhesion molecules play a significant role in cancer progression and metastasis. Cell-cell interactions of cancer cells with endothelium determine the metastatic spread. In addition, direct tumor cell interactions with platelets, leukocytes, and soluble components significantly contribute to cancer cell adhesion, extravasation, and the establishment of metastatic lesions. Clinical evidence indicates that heparin, commonly used for treatment of thromboembolic events in cancer patients, is beneficial for their survival. Preclinical studies confirm that heparin possesses antimetastatic activities that lead to attenuation of metastasis in various animal models. Heparin contains several biological activities that may affect several steps in metastatic cascade. Here we focus on the role of cellular adhesion receptors in the metastatic cascade and discuss evidence for heparin as an inhibitor of cell adhesion. While P- and L-selectin facilitation of cellular contacts during hematogenous metastasis is being accepted as a potential target of heparin, here we propose that heparin may also interfere with integrin activity and thereby affect cancer progression. This review summarizes recent findings about potential mechanisms of tumor cell interactions in the vasculature and antimetastatic activities of heparin.

99m Tc-labeled heparin test in orthopaedic surgery

International Nuclear Information System (INIS)

Bouvier, J.F.; Lafon, J.C.; Colin, M.; Chatelut, J.; Beaubatie, F.

1983-01-01

99m Tc-labeled heparin test was performed for early detection of phlebitis or pulmonary embolism after orthopaedic prothesis. Heparinic treatment and surgery per se were demonstrated to have no effect on the results. If this test demonstrates a statistical difference for pathologic patients, it is of greater value to consider ratio between rates before and after intervention [fr

Evidence-based algorithm for heparin dosing before cardiopulmonary bypass. Part 1: Development of the algorithm.

Science.gov (United States)

McKinney, Mark C; Riley, Jeffrey B

2007-12-01

The incidence of heparin resistance during adult cardiac surgery with cardiopulmonary bypass has been reported at 15%-20%. The consistent use of a clinical decision-making algorithm may increase the consistency of patient care and likely reduce the total required heparin dose and other problems associated with heparin dosing. After a directed survey of practicing perfusionists regarding treatment of heparin resistance and a literature search for high-level evidence regarding the diagnosis and treatment of heparin resistance, an evidence-based decision-making algorithm was constructed. The face validity of the algorithm decisive steps and logic was confirmed by a second survey of practicing perfusionists. The algorithm begins with review of the patient history to identify predictors for heparin resistance. The definition for heparin resistance contained in the algorithm is an activated clotting time 450 IU/kg heparin loading dose. Based on the literature, the treatment for heparin resistance used in the algorithm is anti-thrombin III supplement. The algorithm seems to be valid and is supported by high-level evidence and clinician opinion. The next step is a human randomized clinical trial to test the clinical procedure guideline algorithm vs. current standard clinical practice.

Biomimetic synthesis and biocompatibility evaluation of carbonated apatites template-mediated by heparin

Energy Technology Data Exchange (ETDEWEB)

Deng, Yi [Department of Oral and Maxillofacial Surgery, Laboratory of Interdisciplinary Studies, School and Hospital of Stomatology, Peking University, Beijing 100081 (China); Center for Biomedical Materials and Tissue Engineering, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871 (China); Sun, Yuhua [Department of Oral and Maxillofacial Surgery, Laboratory of Interdisciplinary Studies, School and Hospital of Stomatology, Peking University, Beijing 100081 (China); Chen, Xiaofang [Center for Biomedical Materials and Tissue Engineering, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871 (China); Zhu, Peizhi, E-mail: pzzhu@umich.edu [Center for Biomedical Materials and Tissue Engineering, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871 (China); Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055 (United States); Wei, Shicheng, E-mail: sc-wei@pku.edu.cn [Department of Oral and Maxillofacial Surgery, Laboratory of Interdisciplinary Studies, School and Hospital of Stomatology, Peking University, Beijing 100081 (China); Center for Biomedical Materials and Tissue Engineering, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing 100871 (China)

2013-07-01

Biomimetic synthesis of carbonated apatites with good biocompatibility is a promising strategy for the broadening application of apatites for bone tissue engineering. Most researchers were interested in collagen or gelatin-based templates for synthesis of apatite minerals. Inspired by recent findings about the important role of polysaccharides in bone biomineralization, here we reported that heparin, a mucopolysaccharide, was used to synthesize carbonated apatites in vitro. The results indicated that the Ca/P ratio, carbon content, crystallinity and morphology of the apatites varied depending on the heparin concentration and the initial pH value. The morphology of apatite changed from flake-shaped to needle-shaped, and the degree of crystallinity decreased with the increasing of heparin concentration. Biocompatibility of the apatites was tested by proliferation and alkaline phosphatase activity of MC3T3-E1 cells. The results suggested that carbonated apatites synthesized in the presence of heparin were more favorable to the proliferation and differentiation of MC3T3-E1 cells compared with traditional method. In summary, the heparin concentration and the initial pH value play a key role in the chemical constitution and morphology, as well as biological properties of apatites. These biocompatible nano-apatite crystals hold great potential to be applied as bioactive materials for bone tissue engineering. - Highlights: • Heparin was used as a template to synthesize needle-shaped nano-apatite. • Changing the pH value and concentration led to different properties of apatite. • Apatite prepared by heparin was more favorable to the osteogenic differentiation. • Possible synthesis mechanism of apatite templated by heparin was described.

Detection and properties of A-factor-binding protein from Streptomyces griseus

International Nuclear Information System (INIS)

Miyake, K.; Horinouchi, S.; Yoshida, M.; Chiba, N.; Mori, K.; Nogawa, N.; Morikawa, N.; Beppu, T.

1989-01-01

The optically active form of tritium-labeled A-factor (2-isocapryloyl-3R-hydroxymethyl-gamma-butyrolactone), a pleiotropic autoregulator responsible for streptomycin production, streptomycin resistance, and sporulation in Streptomyces griseus, was chemically synthesized. By using the radioactive A-factor, a binding protein for A-factor was detected in the cytoplasmic fraction of this organism. The binding protein had an apparent molecular weight of approximately 26,000, as determined by gel filtration. Scatchard analysis suggested that A-factor bound the protein in the molar ratio of 1:1 with a binding constant, Kd, of 0.7 nM. The number of the binding protein was roughly estimated to be 37 per genome. The inducing material virginiae butanolide C (VB-C), which has a structure very similar to that of A-factor and is essential for virginiamycin production in Streptomyces virginiae, did not inhibit binding. In addition, no protein capable of specifically binding 3 H-labeled VB-C was found in S. griseus. Together with the observation that VB-C had almost no biological activity on the restoration of streptomycin production or sporulation in an A-factor-deficient mutant of S. griseus, these results indicated that the binding protein had a strict ligand specificity. Examination for an A-factor-binding protein in Streptomyces coelicolor A3(2) and Streptomyces lividans showed the absence of any specifically binding protein

Genetics Home Reference: core binding factor acute myeloid leukemia

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... binding factor acute myeloid leukemia Core binding factor acute myeloid leukemia Printable PDF Open All Close All Enable Javascript ... on PubMed (1 link) PubMed OMIM (1 link) LEUKEMIA, ACUTE MYELOID Sources for This Page Goyama S, Mulloy JC. Molecular ...

In vitro evidence of a tissue factor-independent mode of action of recombinant factor VIIa in hemophilia.

Science.gov (United States)

Augustsson, Cecilia; Persson, Egon

2014-11-13

Successful competition of activated factor VII (FVIIa) with zymogen factor VII (FVII) for tissue factor (TF) and loading of the platelet surface with FVIIa are plausible driving forces behind the pharmacological effect of recombinant FVIIa (rFVIIa) in hemophilia patients. Thrombin generation measurements in platelet-rich hemophilia A plasma revealed competition for TF, which potentially could reduce the effective (r)FVIIa:TF complex concentration and thereby attenuate factor Xa production. However, (auto)activation of FVII apparently counteracted the negative effect of zymogen binding; a small impact was observed at endogenous concentrations of FVII and FVIIa but was virtually absent at pharmacological amounts of rFVIIa. Moreover, corrections of the propagation phase in hemophilia A required rFVIIa concentrations above the range where a physiological level of FVII was capable to downregulate thrombin generation. These data strongly suggest that rFVIIa acts independently of TF in hemophilia therapy and that FVII displacement by rFVIIa is a negligible mechanistic component. © 2014 by The American Society of Hematology.

L2₁ and XA Ordering Competition in Hafnium-Based Full-Heusler Alloys Hf₂VZ (Z = Al, Ga, In, Tl, Si, Ge, Sn, Pb).

Science.gov (United States)

Wang, Xiaotian; Cheng, Zhenxiang; Wang, Wenhong

2017-10-20

For theoretical designing of full-Heusler based spintroinc materials, people have long believed in the so-called Site Preference Rule (SPR). Very recently, according to the SPR, there are several studies on XA-type Hafnium-based Heusler alloys X₂YZ, i.e., Hf₂VAl, Hf₂CoZ (Z = Ga, In) and Hf₂CrZ (Z = Al, Ga, In). In this work, a series of Hf₂-based Heusler alloys, Hf₂VZ (Z = Al, Ga, In, Tl, Si, Ge, Sn, Pb), were selected as targets to study the site preferences of their atoms by first-principle calculations. It has been found that all of them are likely to exhibit the L2₁-type structure instead of the XA one. Furthermore, we reveal that the high values of spin-polarization of XA-type Hf₂VZ (Z = Al, Ga, In, Tl, Si, Ge, Sn, Pb) alloys have dropped dramatically when they form the L2₁-type structure. Also, we prove that the electronic, magnetic, and physics nature of these alloys are quite different, depending on the L2₁-type or XA-type structures.

Marker-Assisted Selection of Xa21 Conferring Resistance to Bacterial Leaf Blight in indica Rice Cultivar LT2

Institute of Scientific and Technical Information of China (English)

Hue Thi NGUYEN; Trung Nguyen DINH; Nakano TOSHITSUGU; Liet Van VU; Quang Hong VU; Tan Van MAI; Thu Thi NGUYEN; Lam Duc VU; Tung Thanh NGUYEN; Long Viet NGUYEN; Hien Thu Thi VU; Hue Thi NONG

2018-01-01

Bacterial leaf blight of rice (BLB), caused by Xanthomonas oryzae pv. oryzae, is one of the most destructive diseases in Asian rice fields. A high-quality rice variety, LT2, was used as the recipient parent. IRBB21, which carries the Xa21 gene, was used as the donor parent. The resistance gene Xa21 was introduced into LT2 by marker-assisted backcrossing. Three Xoo races were used to inoculate the improved lines following the clipping method. Eleven BC3F3lines carrying Xa21 were obtained based on molecular markers and agronomic performance. The 11 lines were then inoculated with the three Xoo races. All the 11 improved lines showed better resistance to BLB than the recipient parent LT2. Based on the level of resistance to BLB and their agronomic performance, five lines (BC3F35.1.5.1, BC3F35.1.5.12, BC3F38.5.6.44, BC3F3 9.5.4.1 and BC3F39.5.4.23) were selected as the most promising for commercial release. These improved lines could contribute to rice production in terms of food security.

Transcription factor binding sites prediction based on modified nucleosomes.

Directory of Open Access Journals (Sweden)

Mohammad Talebzadeh

Full Text Available In computational methods, position weight matrices (PWMs are commonly applied for transcription factor binding site (TFBS prediction. Although these matrices are more accurate than simple consensus sequences to predict actual binding sites, they usually produce a large number of false positive (FP predictions and so are impoverished sources of information. Several studies have employed additional sources of information such as sequence conservation or the vicinity to transcription start sites to distinguish true binding regions from random ones. Recently, the spatial distribution of modified nucleosomes has been shown to be associated with different promoter architectures. These aligned patterns can facilitate DNA accessibility for transcription factors. We hypothesize that using data from these aligned and periodic patterns can improve the performance of binding region prediction. In this study, we propose two effective features, "modified nucleosomes neighboring" and "modified nucleosomes occupancy", to decrease FP in binding site discovery. Based on these features, we designed a logistic regression classifier which estimates the probability of a region as a TFBS. Our model learned each feature based on Sp1 binding sites on Chromosome 1 and was tested on the other chromosomes in human CD4+T cells. In this work, we investigated 21 histone modifications and found that only 8 out of 21 marks are strongly correlated with transcription factor binding regions. To prove that these features are not specific to Sp1, we combined the logistic regression classifier with the PWM, and created a new model to search TFBSs on the genome. We tested the model using transcription factors MAZ, PU.1 and ELF1 and compared the results to those using only the PWM. The results show that our model can predict Transcription factor binding regions more successfully. The relative simplicity of the model and capability of integrating other features make it a superior method

Amperometric Sensor for Heparin: Sensing Mechanism and Application in Human Blood Plasma Analysis

Czech Academy of Sciences Publication Activity Database

Langmaier, Jan; Olšák, J.; Samcová, E.; Samec, Zdeněk; Trojánek, Antonín

2006-01-01

Roč. 18, 13-14 (2006), s. 1329-1338 ISSN 1040-0397 R&D Projects: GA ČR GA203/04/0424 Institutional research plan: CEZ:AV0Z40400503 Keywords : heparin * amperometry * PVC membrane electrode * sensing mechanism * human blood plasma Subject RIV: CG - Electrochemistry Impact factor: 2.444, year: 2006

Fibroblast growth factor regulates insulin-like growth factor-binding protein production by vascular smooth muscle cells.

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Ververis, J; Ku, L; Delafontaine, P

1994-02-01

Insulin-like growth factor I is an important mitogen for vascular smooth muscle cells, and its effects are regulated by several binding proteins. Western ligand blotting of conditioned medium from rat aortic smooth muscle cells detected a 24 kDa binding protein and a 28 kDa glycosylated variant of this protein, consistent with insulin-like growth factor binding protein-4 by size. Low amounts of a glycosylated 38 to 42 kDa doublet (consistent with binding protein-3) and a 31 kDa non-glycosylated protein also were present. Basic fibroblast growth factor markedly increased secretion of the 24 kDa binding protein and its 28 kDa glycosylated variant. This effect was dose- and time-dependent and was inhibited by co-incubation with cycloheximide. Crosslinking of [125I]-insulin-like growth factor I to cell monolayers revealed no surface-associated binding proteins, either basally or after agonist treatment. Induction of binding protein production by fibroblast growth factor at sites of vascular injury may be important in vascular proliferative responses in vivo.

The US regulatory and pharmacopeia response to the global heparin contamination crisis.

Science.gov (United States)

Szajek, Anita Y; Chess, Edward; Johansen, Kristian; Gratzl, Gyöngyi; Gray, Elaine; Keire, David; Linhardt, Robert J; Liu, Jian; Morris, Tina; Mulloy, Barbara; Nasr, Moheb; Shriver, Zachary; Torralba, Pearle; Viskov, Christian; Williams, Roger; Woodcock, Janet; Workman, Wesley; Al-Hakim, Ali

2016-06-09

The contamination of the widely used lifesaving anticoagulant drug heparin in 2007 has drawn renewed attention to the challenges that are associated with the characterization, quality control and standardization of complex biological medicines from natural sources. Heparin is a linear, highly sulfated polysaccharide consisting of alternating glucosamine and uronic acid monosaccharide residues. Heparin has been used successfully as an injectable antithrombotic medicine since the 1930s, and its isolation from animal sources (primarily porcine intestine) as well as its manufacturing processes have not changed substantially since its introduction. The 2007 heparin contamination crisis resulted in several deaths in the United States and hundreds of adverse reactions worldwide, revealing the vulnerability of a complex global supply chain to sophisticated adulteration. This Perspective discusses how the US Food and Drug Administration (FDA), the United States Pharmacopeial Convention (USP) and international stakeholders collaborated to redefine quality expectations for heparin, thus making an important natural product better controlled and less susceptible to economically motivated adulteration.

Veno-venous bypass without systemic heparinization using a centrifugal pump: a blind comparison of a heparin bonded circuit versus a non heparin bonded circuit

NARCIS (Netherlands)

van der Hulst, V. P.; Henny, C. P.; Moulijn, A. C.; Engbers, G.; ten Cate, H.; Gründeman, P. F.; Klopper, P. J.

1989-01-01

Veno-venous bypass without the use of systemic heparinization has recently become of increasing interest for application during liver transplantation and surgery on the large abdominal veins. However, possible adverse effects on blood components as demonstrated by means of hematologic and hemostatic

Structural Fingerprints of Transcription Factor Binding Site Regions

Directory of Open Access Journals (Sweden)

Peter Willett

2009-03-01

Full Text Available Fourier transforms are a powerful tool in the prediction of DNA sequence properties, such as the presence/absence of codons. We have previously compiled a database of the structural properties of all 32,896 unique DNA octamers. In this work we apply Fourier techniques to the analysis of the structural properties of human chromosomes 21 and 22 and also to three sets of transcription factor binding sites within these chromosomes. We find that, for a given structural property, the structural property power spectra of chromosomes 21 and 22 are strikingly similar. We find common peaks in their power spectra for both Sp1 and p53 transcription factor binding sites. We use the power spectra as a structural fingerprint and perform similarity searching in order to find transcription factor binding site regions. This approach provides a new strategy for searching the genome data for information. Although it is difficult to understand the relationship between specific functional properties and the set of structural parameters in our database, our structural fingerprints nevertheless provide a useful tool for searching for function information in sequence data. The power spectrum fingerprints provide a simple, fast method for comparing a set of functional sequences, in this case transcription factor binding site regions, with the sequences of whole chromosomes. On its own, the power spectrum fingerprint does not find all transcription factor binding sites in a chromosome, but the results presented here show that in combination with other approaches, this technique will improve the chances of identifying functional sequences hidden in genomic data.

Intra-articular TSG-6 delivery from heparin-based microparticles reduces cartilage damage in a rat model of osteoarthritis.

Science.gov (United States)

Tellier, Liane E; Treviño, Elda A; Brimeyer, Alexandra L; Reece, David S; Willett, Nick J; Guldberg, Robert E; Temenoff, Johnna S

2018-05-01

As a potential treatment for osteoarthritis (OA), we have developed injectable and hydrolytically degradable heparin-based biomaterials with tunable sulfation for the intra-articular delivery of tumor necrosis factor-alpha stimulated gene-6 (TSG-6), a protein known to inhibit plasmin which may degrade extracellular matrix within OA joints. We first assessed the effect of heparin sulfation on TSG-6 anti-plasmin activity and found that while fully sulfated (Hep) and heparin desulfated at only the N position (Hep-N) significantly enhanced TSG-6 bioactivity in vitro, fully desulfated heparin (Hep-) had no effect, indicating that heparin sulfation plays a significant role in modulating TSG-6 bioactivity. Next, TSG-6 loaded, degradable 10 wt% Hep-N microparticles (MPs) were delivered via intra-articular injection into the knee at 1, 7, and 15 days following medial meniscal transection (MMT) injury in a rat model. After 21 days, cartilage thickness, volume, and attenuation were significantly increased with soluble TSG-6, indicating degenerative changes. In contrast, no significant differences were observed with TSG-6 loaded MP treatment, demonstrating that TSG-6 loaded MPs reduced cartilage damage following MMT injury. Ultimately, our results indicate that Hep-N can enhance TSG-6 anti-plasmin activity and that Hep-N-based biomaterials may be an effective method for TSG-6 delivery to treat OA.

Heparin concentration is critical for cell culture with human platelet lysate.

Science.gov (United States)

Hemeda, Hatim; Kalz, Jana; Walenda, Gudrun; Lohmann, Michael; Wagner, Wolfgang

2013-09-01

Culture media for mesenchymal stromal cells (MSCs) are generally supplemented with fetal bovine serum. Human platelet lysate (hPL) has been proven to be a very effective alternative without the risk of xenogeneic infections or immune reactions. In contrast to fetal bovine serum, hPL comprises plasma, and anticoagulants-usually unfractionated heparin (UFH)-need to be added to prevent gel formation. Cultures of MSCs in hPL media with various concentrations of UFH and enoxaparin, a low-molecular-weight heparin (LMWH), were systematically compared with regard to proliferation, fibroblastoid colony-forming unit frequency, immunophenotype and in vitro differentiation. At least 0.61 IU/mL UFH or 0.024 mg/mL LMWH was necessary for reliable prevention of coagulation of hPL pools used in this study. Higher concentrations impaired cellular proliferation in a dose-dependent manner even without benzyl alcohol, which is commonly added to heparins as a bacteriostatic agent. Colony-forming unit frequency was also reduced at higher heparin concentrations, particularly with LMWH, whereas no significant effect was observed on cellular morphology or immunophenotype. High concentrations of heparins reduced the in vitro differentiation toward adipogenic and osteogenic lineages. Heparin concentration is critical for culture of MSCs in hPL media; this is of particular relevance for cellular therapy where cell culture procedures need to be optimized and standardized. Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Isolation of a cDNA for a Growth Factor of Vascular Endothelial Cells from Human Lung Cancer Cells: Its Identity with Insulin‐like Growth Factor II

Science.gov (United States)

Hagiwara, Koichi; Kobayashi, Tatsuo; Tobita, Masato; Kikyo, Nobuaki; Yazaki, Yoshio

1995-01-01

We have found growth‐promoting activity for vascular endothelial cells in the conditioned medium of a human lung cancer cell line, T3M‐11. Purification and characterization of the growth‐promoting activity have been carried out using ammonium sulfate precipitation and gel‐exclusion chromatography. The activity migrated as a single peak just after ribonuclease. It did not bind to a heparin affinity column. These results suggest that the activity is not a heparin‐binding growth factor (including fibroblast growth factors) or a vascular endothelial growth factor. To identify the molecule exhibiting the growth‐promoting activity, a cDNA encoding the growth factor was isolated through functional expression cloning in COS‐1 cells from a cDNA library prepared from T3M‐11 cells. The nucleotide sequence encoded by the cDNA proved to be identical with that of insulin‐like growth factor II. PMID:7730145

The effect of different forms of heparin on point-of-care blood gas ...

African Journals Online (AJOL)

and heparin vacutainers on blood gas and electrolyte analysis and ... This prospective, cross-sectional study took place in the ED of a ... the effect of two concentrations of liquid heparin and the use of heparin vacutainers on the reliability of blood gas ... Germany) and (iv) a 2 mL plastic syringe (BD) washed with 5 000 IU/.

Nycthemeral variations of 99Tcsup(m)-labelled heparin pharmacokinetic parameters

International Nuclear Information System (INIS)

Decousus, M.; Gremillet, E.; Decousus, H.; Champailler, A.; Houzard, C.; Perpoint, B.; Jaubert, J.

1985-01-01

Six healthy volunteers received four i.v.boluses of 99 Tcsup(m)-heparin at 8.00, 14.00, 20.00 and 02.00 hours at seven-day intervals. Nine blood samples were taken covering a period of 2 h after administration. Simultaneously urine was collected and diuresis not noted. Plasma and urinary radioactivity were measured and standard pharmacokinetic parameters were calculated. Nycthemeral variations of these kinetic parameters were detected by means of distribution-free tests. Circadian rhythms were analysed by means of the cosinor method and the Gauss-Marquardt method. The mean raw value of the following parameters: apparent volume of distribution, plasmatic clearance and extra-renal metabolic clearance, increased significantly between 8.00 and 14.00 and decreased between 14.00 and 20.00. A circadian rhythm was found for the plasmatic clearance only. On the other hand the elimination half-lives and the renal clearance were unaffected by the time of the injections. These results obtained for low doses of 99 Tcsup(m)-heparin suggest a circadian rhythm of the bio-availability of heparin in man. This fact should be taken into account for the use of 99 Tcsup(m)-heparin in the diagnosis of deep-vein thrombosis and for the safe adjustment of the heparin dosages in the treatment of severe thromboembolism. (author)

Highly sensitive ratiometric detection of heparin and its oversulfated chondroitin sulfate contaminant by fluorescent peptidyl probe.

Science.gov (United States)

Mehta, Pramod Kumar; Lee, Hyeri; Lee, Keun-Hyeung

2017-05-15

The selective and sensitive detection of heparin, an anticoagulant in clinics as well as its contaminant oversulfated chondroitin sulfate (OSCS) is of great importance. We first reported a ratiometric sensing method for heparin as well as OSCS contaminants in heparin using a fluorescent peptidyl probe (Pep1, pyrene-GSRKR) and heparin-digestive enzyme. Pep1 exhibited a highly sensitive ratiometric response to nanomolar concentration of heparin in aqueous solution over a wide pH range (2~11) and showed highly selective ratiometric response to heparin among biological competitors such as hyaluronic acid and chondroitin sulfate. Pep1 showed a linear ratiometric response to nanomolar concentrations of heparin in aqueous solutions and in human serum samples. The detection limit for heparin was calculated to be 2.46nM (R 2 =0.99) in aqueous solutions, 2.98nM (R 2 =0.98) in 1% serum samples, and 3.43nM (R 2 =0.99) in 5% serum samples. Pep1 was applied to detect the contaminated OSCS in heparin with heparinase I, II, and III, respectively. The ratiometric sensing method using Pep1 and heparinase II was highly sensitive, fast, and efficient for the detection of OSCS contaminant in heparin. Pep1 with heparinase II could detect as low as 0.0001% (w/w) of OSCS in heparin by a ratiometric response. Copyright © 2017 Elsevier B.V. All rights reserved.

Heparin induced alterations in clearance and distribution of blood-borne microparticles following operative trauma.

Science.gov (United States)

Saba, T M; Antikatzides, T G

1979-04-01

The influence of systemic heparin administration on the vascular clearance and tissue distribution of blood-borne microparticles was evaluated in normal rats and rats after operation (laparotomy plus intestinal manipulation) utilizing an (131)I- colloid which is phagocytized by the reticuloendothelial system (RES). Intravenous heparin administration (100 USP/100g body weight) into normal animals three minutes prior to colloid injection (50 mg/lOOg) induced a significant increase in pulmonary localization of the microparticles as compared to nonheparinized control rats, while hepatic and splenic uptake were decreased. Surgical trauma decreased hepatic RE uptake and increased pulmonary localization of the microparticles when injected systemically at 60 minutes postsurgery. Heparin administration 60 minutes after surgery and three minutes prior to colloid injection, magnified the increased pulmonary localization response with an associated further depression of the RES. The ability of heparin to alter both RE clearance function and lung localization of microparticles was dose dependent and a function of the interval between heparin administration and systemic particulate infusion. Thus, low dose heparin administration was capable of stimulating RE activity while heparin in doses of excess of 50 USP units/lOOg body weight decreased RE function. These findings suggest that the functional state of the hepatic RE system can be greatly affected in a dose-dependent manner by systemic heparin administration which may influence distribution of blood-borne microparticles.

12500 E heparin and 12500 E of a semisynthetic heparin analogue (SSHA) in preventing thrombosis during radiotherapy of gynaecological carcinomas

International Nuclear Information System (INIS)

Hilscher, T.M.

1983-01-01

The effects of 12500 E calcium heparin given once daily were contrasted with those seen under daily treatment with 12500 E of a semisynthetic heparin analogue (SSHA) and evaluated using iodine-125-labelled fibrinogen. The study included 80 patients, who were randomly assigned to the two treatment groups on a 1:1 basis. The findings revealed here led to the conclusion that both drugs, administered once daily by the subcutaneous route, were effective in preventing the occurrence of thrombosis during radiation treatment of gynaecological tumours. (orig./MG) [de

Heparin and glutathione II: correlation between decondensation of bull sperm cells and its nucleons.

Science.gov (United States)

Delgado, N M; Flores-Alonso, J C; Rodríguez-Hernández, H M; Merchant-Larios, H; Reyes, R

2001-01-01

The correlation between the kinetics of bull sperm nuclear and nucleon decondensation induced by the action of physiological concentrations of heparin/GSH was studied. Sperm and nucleon suspensions were incubated at 37 degrees C in salt medium, at a constant concentration of either heparin or GSH and increasing concentrations of the other reagent. Even though nucleons are pretreated with DTT/CTAB, when they are incubated alone with GSH for 96 h, they remain intact, no matter which concentration is employed, and it was impossible to observe the slightest sign of nuclei decondensation. Therefore, rupture of disulfide bridges is not the main mechanism to induce nuclei decondensation and perhaps the GSH role resides in potentate the heparin effect by increasing its negative charge. Nevertheless, nucleons reach 95% of chromatin decondensation in the presence of heparin plus GSH or heparin alone. The fact that the correlation between heparin and GSH concentrations needed to induce sperm nuclei decondensation was 3- to 4-fold greater that in nucleons might be due to the complete lack of nucleon membranes. Heparin/GSH seem to induce nuclei decondensation by an ionic chromatin charge neutralization mechanism.

Photochemically synthesized heparin-based silver nanoparticles: an antimicrobial activity study

Science.gov (United States)

Rodriguez-Torres, Maria del Pilar; Acosta-Torres, Laura Susana; Díaz-Torres, Luis Armando

2017-08-01

The antimicrobial activity of silver nanoparticles has been extensively studied in the last years. Such nanoparticles constitute a potential and promising approach for the development of new antimicrobial systems especially due to the fact that several microorganisms are developing resistance to some already existing antimicrobial agents, therefore making antibacterial and antimicrobial studies on alternative materials necessary to overcome this issue. Silver nanoparticle concentration and size are determining factors on the antimicrobial activity of these nano systems. Heparin is a polysaccharide that belongs to the glycosaminoglycans (GAGs) family, molecules formed by a base disaccharide whose components are joined by a glycosidic linkage that is a repeating unit along their structure. It is highly sulfated making it a negatively charged material that is also widely used as an anticoagulant in Medicine because its biocompatibility besides it is also produced within the human body, specifically in the mast cells. Heparin alone possesses antimicrobial activity although it has not been studied very much in detail, it only has been demonstrated that it inhibits E. coli, P. aeruginosa, S. aureus and S. epidermidis, so taking this into account, this study is dedicated to assess UV photochemically-synthesized (λ=254 nm) heparin-based silver nanoparticles antimicrobial activity using the agar disk diffusion method complemented by the broth microdilution method to estimate de minimum inhibitory concentration (MIC), that is the lowest concentration at which an antimicrobial will inhibit visible growth of a microorganism. The strains used were the ones aforementioned to assess the antimicrobial activity degree these heparinbased nanoparticles exhibit.

Multifunctional silk-heparin biomaterials for vascular tissue engineering applications

Science.gov (United States)

Seib, F. Philipp; Herklotz, Manuela; Burke, Kelly A.; Maitz, Manfred F.; Werner, Carsten; Kaplan, David L.

2013-01-01

Over the past 30 years, silk has been proposed for numerous biomedical applications that go beyond its traditional use as a suture material. Silk sutures are well tolerated in humans, but the use of silk for vascular engineering applications still requires extensive biocompatibility testing. Some studies have indicated a need to modify silk to yield a hemocompatible surface. This study examined the potential of low molecular weight heparin as a material for refining silk properties by acting as a carrier for vascular endothelial growth factor (VEGF) and improving silk hemocompatibility. Heparinized silk showed a controlled VEGF release over 6 days; the released VEGF was bioactive and supported the growth of human endothelial cells. Silk samples were then assessed using a humanized hemocompatibility system that employs whole blood and endothelial cells. The overall thrombogenic response for silk was very low and similar to the clinical reference material polytetrafluoroethylene. Despite an initial inflammatory response to silk, apparent as complement and leukocyte activation, the endothelium was maintained in a resting, anticoagulant state. The low thrombogenic response and the ability to control VEGF release support the further development of silk for vascular applications. PMID:24099708

SPECIFIC ASPECTS OF INTERACTION OF PLATELETS WITH THE HEPARINIZED MATERIALS

Directory of Open Access Journals (Sweden)

E.A. Nemets

2012-01-01

Full Text Available Comparative analysis of anticoagulant nature on medical materials testing was done. It was found that change of citrate by heparin is accompanied by significant changes in platelet adhesion and activation. This results allowed us to arrive at a conclusion about reasonability of heparin usage as anticoagulant in in vitro testing. 

Safety of low-molecular-weight heparin in pregnancy: a systematic review

NARCIS (Netherlands)

Sanson, B. J.; Lensing, A. W.; Prins, M. H.; Ginsberg, J. S.; Barkagan, Z. S.; Lavenne-Pardonge, E.; Brenner, B.; Dulitzky, M.; Nielsen, J. D.; Boda, Z.; Turi, S.; Mac Gillavry, M. R.; Hamulyák, K.; Theunissen, I. M.; Hunt, B. J.; Büller, H. R.

1999-01-01

Unfractionated heparin (UFH) remains the anticoagulant of choice during pregnancy. Low-molecular-weight heparins (LMWH) are an attractive alternative to UFH due to their logistic advantages and their association with a lower incidence of osteoporosis and HIT. We reviewed all published clinical

Qualitative and Quantitative Analysis of Heparin during Precipitation by Near-Infrared Spectroscopy

OpenAIRE

Lian Li; Jinfeng Wang; Hengchang Zang; Hui Zhang; Wei Jiang; Shang Chen; Fengshan Wang

2016-01-01

Heparin is a glycosaminoglycan (GAG) that plays an important role in the blood coagulation system. Its quality is of great importance, so it is necessary to develop a fast analytical method during the manufacture process to analyse the quality of heparin produced. In this study, the heparin contents of 80 samples collected from five batches during the precipitation process were analysed using nearinfrared (NIR) spectroscopy and a chemometrics approach. This was done in order to improve the ef...

Rat embryo fibroblasts require both the cell-binding and the heparin-binding domains of fibronectin for survival

DEFF Research Database (Denmark)

Jeong, J; Han, I; Lim, Y

2001-01-01

of the cell-binding domain of FN with integrin is sufficient to rescue rat embryo fibroblasts (REFs) from detachment-induced apoptosis. REFs attached and spread normally after plating on substrates coated with either intact FN or a FN fragment, FN120, that contains the cell-binding domain but lacks the C...

The role of heparin in sepsis: much more than just an anticoagulant.

Science.gov (United States)

Li, Xu; Ma, Xiaochun

2017-11-01

Despite progress in antibiotic treatment, mechanical ventilation, fluid resuscitation and blood glucose maintenance, sepsis remains a cause of high mortality in the intensive care unit to date, there are no proven treatment strategies for the routine management of septic patients. The extensive interaction between inflammation and coagulation contributes to the basic pathophysiology of sepsis. Thus, the agents that attenuate the activation of both inflammation and coagulation may improve the outcome in sepsis. Apart from the well-known anticoagulant effects of heparin, it also possesses various immunomodulatory properties and protects glycocalyx from shedding. Hence, heparin seems to be such an agent. Immunothrombosis plays an important role in early host defence against bacterial dissemination, thus the proper timing for anticoagulant therapy should be determined. We review the available experimental and clinical data supporting the use of heparin in sepsis. At this time the use of heparin in the treatment of sepsis is conflicting. Future trials of heparin therapy for sepsis should concentrate on the very severely ill patients, in whom benefit is most likely to be demonstrated. © 2017 John Wiley & Sons Ltd.

Extracellular matrix inspired surface functionalization with heparin, fibronectin and VEGF provides an anticoagulant and endothelialization supporting microenvironment

International Nuclear Information System (INIS)

Wang, Xue; Liu, Tao; Chen, Yuan; Zhang, Kun; Maitz, Manfred F.; Pan, Changjiang; Chen, Junying; Huang, Nan

2014-01-01

Highlights: • Surface modification with fibronectin, heparin and VEGF could selectively anticoagulant and promote endothelialization. • The bioactivity of biomolecules was more efficiently maintained via specific intermolecular interaction. • Poly-l-lysine interlayer was more feasible and the degradation product had no harm to human body. - Abstract: The biocompatibility of currently used coronary artery stent is still far from perfect, which closely related to insufficient endothelialization and thrombus formation. In this study, heparin, fibronectin and VEGF were immobilized on Ti surface to construct a multifunctional microenvironment with favorable properties to inhibit thrombosis formation and promote endothelialization simultaneously. The microenvironment on Ti surface was characterized in detail and demonstrated that the Hep/Fn/VEGF biofunctional coating was constructed successfully on Ti surface. The influence of surface properties such as chemical composition, roughness, hydrophilicity, and binding density of biomolecules on the performances of hemocompatibility and cytocompatibility was evaluated and discussed. Modified surface significantly enhanced the AT III binding density and prolonged the clotting time. In vitro platelet adhesion and activation assays further proved that the modified surface presented favorable anti-coagulant property. In addition, the proliferation of endothelial progenitor cells (EPCs) and endothelial cells (ECs) on the Hep/Fn/VEGF biofunctional coating was significantly promoted. In conclusion, the Hep/Fn/VEGF biofunctional coating was successfully constructed with desirable anticoagulant and endothelialization supporting properties. This work may provide a promising approach for biofunctional surface modification of coronary artery stent to acquire a desired multifunctional microenvironment

Using TESS to predict transcription factor binding sites in DNA sequence.

Science.gov (United States)

Schug, Jonathan

2008-03-01

This unit describes how to use the Transcription Element Search System (TESS). This Web site predicts transcription factor binding sites (TFBS) in DNA sequence using two different kinds of models of sites, strings and positional weight matrices. The binding of transcription factors to DNA is a major part of the control of gene expression. Transcription factors exhibit sequence-specific binding; they form stronger bonds to some DNA sequences than to others. Identification of a good binding site in the promoter for a gene suggests the possibility that the corresponding factor may play a role in the regulation of that gene. However, the sequences transcription factors recognize are typically short and allow for some amount of mismatch. Because of this, binding sites for a factor can typically be found at random every few hundred to a thousand base pairs. TESS has features to help sort through and evaluate the significance of predicted sites.

Covalently bound conjugates of albumin and heparin: Synthesis, fractionation and characterization

NARCIS (Netherlands)

Hennink, Wim E.; Feijen, Jan; Ebert, Charles D.; Kim, Sung Wan

1983-01-01

Covalently bound conjugates of human serum albumin and heparin were prepared as compounds which could improve the blood-compatibility of polymer surfaces either by preadsorption or by covalent coupling of the conjugates onto blood contacting surfaces. The conjugates (10–16 weight % of heparin) were

Meningococcal factor H-binding protein vaccines with decreased binding to human complement factor H have enhanced immunogenicity in human factor H transgenic mice.

Science.gov (United States)

Rossi, Raffaella; Granoff, Dan M; Beernink, Peter T

2013-11-04

Factor H-binding protein (fHbp) is a component of a meningococcal vaccine recently licensed in Europe for prevention of serogroup B disease, and a second vaccine in clinical development. The protein specifically binds human factor H (fH), which down-regulates complement activation and enhances resistance to bactericidal activity. There are conflicting data from studies in human fH transgenic mice on whether binding of human fH to fHbp vaccines decreases immunogenicity, and whether mutant fHbp vaccines with decreased fH binding have enhanced immunogenicity. fHbp can be classified into two sub-families based on sequence divergence and immunologic cross-reactivity. Previous studies of mutant fHbp vaccines with low fH binding were from sub-family B, which account for approximately 60% of serogroup B case isolates. In the present study, we evaluated the immunogenicity of two mutant sub-family A fHbp vaccines containing single substitutions, T221A or D211A, which resulted in 15- or 30-fold lower affinity for human fH, respectively, than the corresponding control wild-type fHbp vaccine. In transgenic mice with high serum concentrations of human fH, both mutant vaccines elicited significantly higher IgG titers and higher serum bactericidal antibody responses than the control fHbp vaccine that bound human fH. Thus, mutations introduced into a sub-family A fHbp antigen to decrease fH binding can increase protective antibody responses in human fH transgenic mice. Collectively the data suggest that mutant fHbp antigens with decreased fH binding will result in superior vaccines in humans. Copyright © 2013 Elsevier Ltd. All rights reserved.

Marker-Assisted Selection of Xa21 Conferring Resistance to Bacterial Leaf Blight in indica Rice Cultivar LT2

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Hue Thi Nguyen

2018-01-01

Full Text Available Bacterial leaf blight of rice (BLB, caused by Xanthomonas oryzae pv. oryzae, is one of the most destructive diseases in Asian rice fields. A high-quality rice variety, LT2, was used as the recipient parent. IRBB21, which carries the Xa21 gene, was used as the donor parent. The resistance gene Xa21 was introduced into LT2 by marker-assisted backcrossing. Three Xoo races were used to inoculate the improved lines following the clipping method. Eleven BC3F3 lines carrying Xa21 were obtained based on molecular markers and agronomic performance. The 11 lines were then inoculated with the three Xoo races. All the 11 improved lines showed better resistance to BLB than the recipient parent LT2. Based on the level of resistance to BLB and their agronomic performance, five lines (BC3F3 5.1.5.1, BC3F3 5.1.5.12, BC3F3 8.5.6.44, BC3F3 9.5.4.1 and BC3F3 9.5.4.23 were selected as the most promising for commercial release. These improved lines could contribute to rice production in terms of food security.

Factor VIIa binding and internalization in hepatocytes

DEFF Research Database (Denmark)

Hjortoe, G; Sorensen, B B; Petersen, L C

2005-01-01

The liver is believed to be the primary clearance organ for coagulation proteases, including factor VIIa (FVIIa). However, at present, clearance mechanisms for FVIIa in liver are unknown. To obtain information on the FVIIa clearance mechanism, we investigated the binding and internalization...... no effect. HEPG2 cells internalized FVIIa with a rate of 10 fmol 10(-5) cells h(-1). In contrast to HEPG2 cells, FVIIa binding to primary rat hepatocytes was completely independent of TF, and excess unlabeled FVIIa partly reduced the binding of 125I-FVIIa to rat hepatocytes. Further, compared with HEPG2...... cells, three- to fourfold more FVIIa bound to rat primary hepatocytes, and the bound FVIIa was internalized at a faster rate. Similar FVIIa binding and internalization profiles were observed in primary human hepatocytes. Plasma inhibitors had no effect on FVIIa binding and internalization in hepatocytes...

Heparin-mimicking multilayer coating on polymeric membrane via LbL assembly of cyclodextrin-based supramolecules.

Science.gov (United States)

Deng, Jie; Liu, Xinyue; Ma, Lang; Cheng, Chong; Shi, Wenbin; Nie, Chuanxiong; Zhao, Changsheng

2014-12-10

In this study, multifunctional and heparin-mimicking star-shaped supramolecules-deposited 3D porous multilayer films with improved biocompatibility were fabricated via a layer-by-layer (LbL) self-assembly method on polymeric membrane substrates. Star-shaped heparin-mimicking polyanions (including poly(styrenesulfonate-co-sodium acrylate; Star-PSS-AANa) and poly(styrenesulfonate-co-poly(ethylene glycol)methyl ether methacrylate; Star-PSS-EGMA)) and polycations (poly(methyl chloride-quaternized 2-(dimethylamino)ethyl methacrylate; Star-PMeDMA) were first synthesized by atom transfer radical polymerization (ATRP) from β-cyclodextrin (β-CD) based cores. Then assembly of 3D porous multilayers onto polymeric membrane surfaces was carried out by alternating deposition of the polyanions and polycations via electrostatic interaction. The surface morphology and composition, water contact angle, blood activation, and thrombotic potential as well as cell viability for the coated heparin-mimicking films were systematically investigated. The results of surface ATR-FTIR spectra and XPS spectra verified successful deposition of the star-shaped supramolecules onto the biomedical membrane surfaces; scanning electron microscopy (SEM) and atomic force microscopy (AFM) observations revealed that the modified substrate had 3D porous surface morphology, which might have a great biological influence on the biointerface. Furthermore, systematic in vitro investigation of protein adsorption, platelet adhesion, human platelet factor 4 (PF4, indicates platelet activation), activate partial thromboplastin time (APTT), thrombin time (TT), coagulation activation (thrombin-antithrombin III complex (TAT, indicates blood coagulant)), and blood-related complement activation (C3a and C5a, indicates inflammation potential) confirmed that the heparin-mimicking multilayer coated membranes exhibited ultralow blood component activations and excellent hemocompatibility. Meanwhile, after surface coating

Factor VII and protein C are phosphatidic acid-binding proteins.

Science.gov (United States)

Tavoosi, Narjes; Smith, Stephanie A; Davis-Harrison, Rebecca L; Morrissey, James H

2013-08-20

Seven proteins in the human blood clotting cascade bind, via their GLA (γ-carboxyglutamate-rich) domains, to membranes containing exposed phosphatidylserine (PS), although with membrane binding affinities that vary by 3 orders of magnitude. Here we employed nanodiscs of defined phospholipid composition to quantify the phospholipid binding specificities of these seven clotting proteins. All bound preferentially to nanobilayers in which PS headgroups contained l-serine versus d-serine. Surprisingly, however, nanobilayers containing phosphatidic acid (PA) bound substantially more of two of these proteins, factor VIIa and activated protein C, than did equivalent bilayers containing PS. Consistent with this finding, liposomes containing PA supported higher proteolytic activity by factor VIIa and activated protein C toward their natural substrates (factors X and Va, respectively) than did PS-containing liposomes. Moreover, treating activated human platelets with phospholipase D enhanced the rates of factor X activation by factor VIIa in the presence of soluble tissue factor. We hypothesize that factor VII and protein C bind preferentially to the monoester phosphate of PA because of its accessibility and higher negative charge compared with the diester phosphates of most other phospholipids. We further found that phosphatidylinositol 4-phosphate, which contains a monoester phosphate attached to its myo-inositol headgroup, also supported enhanced enzymatic activity of factor VIIa and activated protein C. We conclude that factor VII and protein C bind preferentially to monoester phosphates, which may have implications for the function of these proteases in vivo.

Effect of crosslinking chemistry of albumin/heparin multilayers on FGF-2 adsorption and endothelial cell behavior

Czech Academy of Sciences Publication Activity Database

Kumorek, Marta M.; Janoušková, Olga; Höcherl, Anita; Houska, Milan; Mázl Chánová, Eliška; Kasoju, Naresh; Cuchalová, Lucie; Matějka, R.; Kubies, Dana

2017-01-01

Roč. 411, 31 July (2017), s. 240-250 ISSN 0169-4332 R&D Projects: GA MŠk(CZ) LQ1604; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:61389013 Keywords : basic fibroblast growth factor * heparin * cross-linking Subject RIV: CD - Macromolecular Chemistry OBOR OECD: Polymer science Impact factor: 3.387, year: 2016

Deciphering the Role of Sulfonated Unit in Heparin-Mimicking Polymer to Promote Neural Differentiation of Embryonic Stem Cells.

Science.gov (United States)

Lei, Jiehua; Yuan, Yuqi; Lyu, Zhonglin; Wang, Mengmeng; Liu, Qi; Wang, Hongwei; Yuan, Lin; Chen, Hong

2017-08-30

Glycosaminoglycans (GAGs), especially heparin and heparan sulfate (HS), hold great potential for inducing the neural differentiation of embryonic stem cells (ESCs) and have brought new hope for the treatment of neurological diseases. However, the disadvantages of natural heparin/HS, such as difficulty in isolating them with a sufficient amount, highly heterogeneous structure, and the risk of immune responses, have limited their further therapeutic applications. Thus, there is a great demand for stable, controllable, and well-defined synthetic alternatives of heparin/HS with more effective biological functions. In this study, based upon a previously proposed unit-recombination strategy, several heparin-mimicking polymers were synthesized by integrating glucosamine-like 2-methacrylamido glucopyranose monomers (MAG) with three sulfonated units in different structural forms, and their effects on cell proliferation, the pluripotency, and the differentiation of ESCs were carefully studied. The results showed that all the copolymers had good cytocompatibility and displayed much better bioactivity in promoting the neural differentiation of ESCs as compared to natural heparin; copolymers with different sulfonated units exhibited different levels of promoting ability; among them, copolymer with 3-sulfopropyl acrylate (SPA) as a sulfonated unit was the most potent in promoting the neural differentiation of ESCs; the promoting effect is dependent on the molecular weight and concentration of P(MAG-co-SPA), with the highest levels occurring at the intermediate molecular weight and concentration. These results clearly demonstrated that the sulfonated unit in the copolymers played an important role in determining the promoting effect on ESCs' neural differentiation; SPA was identified as the most potent sulfonated unit for copolymer with the strongest promoting ability. The possible reason for sulfonated unit structure as a vital factor influencing the ability of the copolymers

Direct oral anticoagulants and venous thromboembolism

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Massimo Franchini

2016-09-01

Full Text Available Venous thromboembolism (VTE, consisting of deep vein thrombosis and pulmonary embolism, is a major clinical concern associated with significant morbidity and mortality. The cornerstone of management of VTE is anticoagulation, and traditional anticoagulants include parenteral heparins and oral vitamin K antagonists. Recently, new oral anticoagulant drugs have been developed and licensed, including direct factor Xa inhibitors (e.g. rivaroxaban, apixaban and edoxaban and thrombin inhibitors (e.g. dabigatran etexilate. This narrative review focusses on the characteristics of these direct anticoagulants and the main results of published clinical studies on their use in the prevention and treatment of VTE.

Examination of the ligand-binding and enzymatic properties of a bilin-binding protein from the poisonous caterpillar Lonomia obliqua.

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Ana B G Veiga

Full Text Available The bilin-binding proteins (BBP from lepidopteran insects are members of the lipocalin family of proteins and play a special role in pigmentation through the binding of biliverdin IXγ. Lopap, a BBP-like protein from the venom of the toxic caterpillar Lonomia obliqua has been reported to act as a serine protease that activates the coagulation proenzyme prothrombin. Here we show that BBPLo, a variant of lopap from the same organism binds biliverdin IXγ, forming a complex that is spectrally identical with previously described BBP proteins. Although BBPLo is nearly identical in sequence to lopap, no prothrombinase activity was detected in our recombinant preparations using reconstituted systems containing coagulation factors Xa and Va, as well as anionic phospholipids. In addition to biliverdin, BBPLo was found to form a 1:1 complex with heme prompting us to examine whether the unusual biliverdin IXγ ligand of BBPs forms as a result of oxidation of bound heme in situ rather than by a conventional heme oxygenase. Using ascorbate or a NADPH(+-ferredoxin reductase-ferredoxin system as a source of reducing equivalents, spectral changes are seen that suggest an initial reduction of heme to the Fe(II state and formation of an oxyferrous complex. The complex then disappears and a product identified as a 5-coordinate carbonyl complex of verdoheme, an intermediate in the biosynthesis of biliverdin, is formed. However, further reaction to form biliverdin was not observed, making it unlikely that biliverdin IXγ is formed by this pathway.

Morbidity associated with heparin therapy in spinal surgery patients with cardiovascular diseases

International Nuclear Information System (INIS)

Sawakami, Kimihiko; Ishikawa, Seiichi; Ito, Takui

2011-01-01

The objectives of this study were to investigate morbidity associated with heparin therapy in spinal surgery patients. The management of patients on anticoagulant therapy who undergo spinal surgery is becoming a common clinical problem. Although guidelines for the management of gastrointestinal endoscopy patients on heparin therapy have been published, spinal surgery may lead to specific complications, especially because of heparin therapy. However, only few studies have examined the clinical significance of heparin therapy in spinal surgery patients. The subjects of this study were 116 consecutive patients who were on anticoagulant or antiplatelet therapy. This says that all of the patients were receiving heparin or another anticoagunt. The patients were divided into 2 groups: a group that received heparin therapy before and after surgery (H group, n=25) and a group that did not receive heparin therapy (NH group, n=91). The results of clinical examinations and magnetic resonance imaging (MRI) in the 2 groups were compared. There were no significant differences between the 2 groups in baseline data. Comorbidities in both groups included valvular heart disease, atrial fibrillation, angina pectoris/myocardial infarction, and cerebral infarction. Mean intraoperative and postoperative blood loss in the H group were 324 ml and 536 ml, respectively, and the corresponding values in the NH group were 431 ml and 449 ml, respectively. MRI of all patients was performed within 10 days after surgery and T2-weighted images in the axial plane were examined for evidence of an epidural hematoma. Although the proportion of patients with an epidural hematoma, detected by MRI was higher in the H group than in the NH group (71% vs. 64%), none of the patients in either group required revision surgery because of intolerable pain or muscle weakness. Thrombocytopenia and skin necrosis were observed as complications of the heparin therapy in 1 patient in the H group (4%). The rate of

How much heparin do we really need to go on pump? A rethink of current practices.

LENUS (Irish Health Repository)

Shuhaibar, M N

2012-02-03

OBJECTIVES: Patients undergoing myocardial revascularisation using extracorporeal circulation require heparin anticoagulation. We aimed to evaluate the effect of reducing heparin dosage on target activated clotting time (ACT) and postoperative blood loss. METHODS: In a prospective randomised trial, 195 patients undergoing isolated primary CABG were randomised into four groups A, B, C, and D receiving an initial heparin dosage of 100, 200, 250 and 300 iu\\/kg, respectively. Extra incremental heparin (50 iu\\/kg) was added if required to achieve a target ACT of 480 s before initiating cardiopulmonary bypass. Postoperative blood loss was measured from the time of heparin reversal to drain removal 24h later. RESULTS: Target ACT was achieved in 0, 63, 68.3 and 82.4% of patients in groups A, B, C and D, respectively, after the initial dose of heparin. In group B, of those not achieving target act a single increment of heparin was sufficient to achieve target ACT in further 18.6%. The mean ACT after the initial dose in groups B, C and D was 482.9, 519 and 588 s, respectively (P<0.05). Postoperative blood loss in millilitre per kilogram was directly proportional to preoperative heparin dose. CONCLUSIONS: Patients receiving lower dose of heparin has lower postoperative blood loss. Of those achieving the target ACT, group B was significantly the closest to the target ACT. A starting dose of 200 iu\\/kg of heparin and if necessary one 50 iu\\/kg increment achieved target ACT in 81.5% of patients. The added benefit of significant drop in postoperative blood loss is evident.

The Effect of Low Molecular Weight Heparins on Fracture Healing.

Science.gov (United States)

Kapetanakis, Stylianos; Nastoulis, Evangelos; Demesticha, Theano; Demetriou, Thespis

2015-01-01

Venous Thromboembolism is a serious complication in the trauma patient. The most commonly studied and used anticoagulant treatment in prophylaxis of thrombosis is heparin. The prolonged use of unfractionated heparin has been connected with increased incidence of osteoporotic fractures. Low molecular-weight-heparins (LMWHs) have been the golden rule in antithrombotic therapy during the previous two decades as a way to overcome the major drawbacks of unfractioned heparin. However there are few studies reporting the effects of LMWHs on bone repair after fractures. This review presents the studies about the effects of LMWHs on bone biology (bone cells and bone metabolism) and underlying the mechanisms by which LMWHs may impair fracture healing process. The authors' research based on literature concluded that there are no facts and statistics for the role of LMWHs on fracture healing process in humans and the main body of evidence of their role comes from in vitro and animal studies. Further large clinical studies designed to compare different types of LMWHs, in different dosages and in different patient or animal models are needed for exploring the effects of LMWHs on fracture healing process.

Heparin-Induced Cardiac Tamponade and Life-Threatening Hyperkalemia in a Patient with Chronic Hemodialysis

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Ho-Ming Su

2005-03-01

Full Text Available Heparin, a commonly used anticoagulant agent, is frequently used in patients undergoing hemodialysis. As with most medications, heparin has a significant side effect profile. Two of its most important side effects, major bleeding and hyperkalemia, may be devastating without immediate diagnosis and treatment. Major bleeding such as gastrointestinal, genitourinary or intracranial bleeding is occasionally encountered and rarely neglected. However, heparin-induced cardiac tamponade is rarely encountered and may be easily overlooked. Another side effect, heparin-induced hyperkalemia, an unusual but well-described side effect, is frequently forgotten until life-threatening arrhythmia has occurred. We report a case involving a 40-year-old male patient with uremia, who had received heparin for 10 days for deep vein thrombosis in the left lower extremity. Hemopericardium with cardiac tamponade and life-threatening hyperkalemia were both noted in this patient.

Heparinization of alimentation solutions administered through peripheral veins in premature infants: a controlled study.

Science.gov (United States)

Alpan, G; Eyal, F; Springer, C; Glick, B; Goder, K; Armon, J

1984-09-01

A randomized controlled study was done to determine whether the addition of heparin (1 U/mL) to peripheral intravenous alimentation solutions would affect the incidence of phlebitis and duration of patency of intravenous catheters in premature infants. Twenty-two-gauge Teflon catheters were uniformly used. One hundred five catheters infused with heparin were placed in 13 infants, and 122 catheters were placed in the control group of 13 infants. The time, nature, and incidence of complications were noted for each infusion site. Infusion of heparin was found to double the duration of patency of intravenous catheters and to reduce significantly the incidence of phlebitis. No complications related to the administration of heparin were noted. Heparinization of intravenous alimentation solutions should therefore be considered in premature infants as a means of reducing the work load and incidence of complications associated with peripheral lines.

Effects of heparin on platelet aggregation and release and thromboxane A2 production

International Nuclear Information System (INIS)

Mohammad, S.F.; Anderson, W.H.; Smith, J.B.; Chuang, H.Y.; Mason, R.G.

1981-01-01

Heparin, when added to citrated platelet-rich plasma (PRP), caused potentiation of platelet aggregation and the release reaction induced by the aggregating agents adenosine diphosphate (ADP), arachidonic acid, collagen, and epinephrine. At low concentrations (4.7 x 10(-5) M) arachidonic acid failed to cause aggregation of platelets in citrated PRP. However, in the presence of heparin, the same concentration of arachidonic acid caused aggregation. Examination of PRP for the presence of thromboxane A2 (TxA2) by use of a bioassay revealed that heparin also stimulated release of TxA2. This finding indicated that platelets released more TxA2 when they were challenged by low concentrations of arachidonic acid in the presence of heparin than in its absence. Platelets were labeled with 3 H-arachidonic acid and 14 C-serotonin, and attempts were made to determine whether heparin stimulated the platelet release reaction first with subsequent increased production of TxA2, or alternatively, whether heparin stimulated TxA2 production first with subsequent enhancement of the release reaction. In view of the demonstrated simultaneous release of 14 C-serotonin and 3 H-arachidonic acid metabolites, it appeared that either release of 14 C and 3 H occurs concurrently or, even if one of these events is dependent on the other, both events take place in rapid succession. Timed sequential studies revealed that in the presence of arachidonic acid, the addition of heparin hastened the apparently simultaneous release of both 14 C and 3 H

Increased accuracy in heparin and protamine administration decreases bleeding: a pilot study

DEFF Research Database (Denmark)

Runge, Marx; Møller, Christian H; Steinbrüchel, Daniel A

2009-01-01

Three to 5 percent of the patients undergoing cardiac surgery are reoperated because of bleeding. When a surgical cause can be excluded, heparin/protamine mismatch may be considered. Insufficient reversal of heparin and overdosing of protamine may cause postoperative bleeding. The purpose......). A reduced number of patients needed blood transfusions in the RxDx group, although this was not statistically significant (19% vs. 38%, respectively; p = .13). Initial heparin dose was significantly reduced in the RxDx group (250 mg; range, 100-375 mg) compared with the control group (300 mg; range, 200...

Prevention of adverse events of interferon γ gene therapy by gene delivery of interferon γ-heparin-binding domain fusion protein in mice

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Mitsuru Ando

2014-01-01

Full Text Available Sustained gene delivery of interferon (IFN γ can be an effective treatment, but our previous study showed high levels of IFNγ-induced adverse events, including the loss of body weight. These unwanted events could be reduced by target-specific delivery of IFNγ after in vivo gene transfer. To achieve this, we selected the heparin-binding domain (HBD of extracellular superoxide dismutase as a molecule to anchor IFNγ to the cell surface. We designed three IFNγ derivatives, IFNγ-HBD1, IFNγ-HBD2, and IFNγ-HBD3, each of which had 1, 2, or 3 HBDs, respectively. Each plasmid-encoding fusion proteins was delivered to the liver, a model target in this study, by hydrodynamic tail vein injection. The serum concentration of IFNγ-HBD2 and IFNγ-HBD3 after gene delivery was lower than that of IFNγ or IFNγ-HBD1. Gene delivery of IFNγ-HBD2, but not of IFNγ-HBD3, effectively increased the mRNA expression of IFNγ-inducible genes in the liver, suggesting liver-specific distribution of IFNγ-HBD2. Gene delivery of IFNγ-HBD2-suppressed tumor growth in the liver as efficiently as that of IFNγ with much less symptoms of adverse effects. These results indicate that the adverse events of IFNγ gene transfer can be prevented by gene delivery of IFNγ-HBD2, a fusion protein with high cell surface affinity.

Tissue factor pathway inhibitor (TFPI) release after heparin stimulation is increased in Type 1 diabetic patients with albuminuria

NARCIS (Netherlands)

Leurs, PB; van Oerle, R; Hamulyak, K; Wolffenbuttel, BHR

Aims To study heparin-stimulated TFPI release in relation to complications in Type 1 diabetic patients. Subjects and methods Nineteen uncomplicated Type 1 diabetic patients (group I) were compared with 18 patients with retinopathy (group II), and nine patients with retinopathy and albuminuria (group

Complement-mediated bactericidal activity of anti-factor H binding protein monoclonal antibodies against the meningococcus relies upon blocking factor H binding.

Science.gov (United States)

Giuntini, Serena; Reason, Donald C; Granoff, Dan M

2011-09-01

Binding of the complement-downregulating protein factor H (fH) to the surface of the meningococcus is important for survival of the organism in human serum. The meningococcal vaccine candidate factor H binding protein (fHbp) is an important ligand for human fH. While some fHbp-specific monoclonal antibodies (MAbs) block binding of fH to fHbp, the stoichiometry of blocking in the presence of high serum concentrations of fH and its effect on complement-mediated bactericidal activity are unknown. To investigate this question, we constructed chimeric antibodies in which the human IgG1 constant region was paired with three murine fHbp-specific binding domains designated JAR 3, JAR 5, and MAb502. By surface plasmon resonance, the association rates for binding of all three MAbs to immobilized fHbp were >50-fold higher than that for binding of fH to fHbp, and the MAb dissociation rates were >500-fold lower than that for fH. While all three MAbs elicited similar C1q-dependent C4b deposition on live bacteria (classical complement pathway), only those antibodies that inhibited binding of fH to fHbp (JAR 3 and JAR 5) had bactericidal activity with human complement. MAb502, which did not inhibit fH binding, had complement-mediated bactericidal activity only when tested with fH-depleted human complement. When an IgG1 anti-fHbp MAb binds to sparsely exposed fHbp on the bacterial surface, there appears to be insufficient complement activation for bacteriolysis unless fH binding also is inhibited. The ability of fHbp vaccines to elicit protective antibodies, therefore, is likely to be enhanced if the antibody repertoire is of high avidity and includes fH-blocking activity.

In vitro binding of 67Ga to Ehrlich ascites tumor cells

International Nuclear Information System (INIS)

Kojima, S.; Kubodera, A.

1984-01-01

The binding of 67 Ga to Ehrlich ascites tumor cells (ETC) was studied in vitro. Acid mucopolysaccharide (AMPS) present at the cell surface of ETC was identified as heparan sulfate (HS). The extent of 67 Ga binding to ETC reached a plateau (ca. 10% of the added dose) at 1-2 h after the start of incubation. The binding was higher under neutral or alkaline conditions than under acidic conditions. Heparin and heparitinase treatment both significantly decreased the extent of 67 Ga binding to ETC. Mild treatment with protease, including trypsin or papain, also decreased the binding. On the contrary, the treatment with trypsin under severe conditions markedly increased the extent of 67 Ga binding to ETC. These results support the hypothesis that HS plays an important role as a 67 Ga receptor in the mechanism of gallium binding to ETC. (orig.)

Functional interaction of the DNA-binding transcription factor Sp1 through its DNA-binding domain with the histone chaperone TAF-I.

Science.gov (United States)

Suzuki, Toru; Muto, Shinsuke; Miyamoto, Saku; Aizawa, Kenichi; Horikoshi, Masami; Nagai, Ryozo

2003-08-01

Transcription involves molecular interactions between general and regulatory transcription factors with further regulation by protein-protein interactions (e.g. transcriptional cofactors). Here we describe functional interaction between DNA-binding transcription factor and histone chaperone. Affinity purification of factors interacting with the DNA-binding domain of the transcription factor Sp1 showed Sp1 to interact with the histone chaperone TAF-I, both alpha and beta isoforms. This interaction was specific as Sp1 did not interact with another histone chaperone CIA nor did other tested DNA-binding regulatory factors (MyoD, NFkappaB, p53) interact with TAF-I. Interaction of Sp1 and TAF-I occurs both in vitro and in vivo. Interaction with TAF-I results in inhibition of DNA-binding, and also likely as a result of such, inhibition of promoter activation by Sp1. Collectively, we describe interaction between DNA-binding transcription factor and histone chaperone which results in negative regulation of the former. This novel regulatory interaction advances our understanding of the mechanisms of eukaryotic transcription through DNA-binding regulatory transcription factors by protein-protein interactions, and also shows the DNA-binding domain to mediate important regulatory interactions.

Influence of Human Leukocyte Antigen (HLA) Alleles and Killer Cell Immunoglobulin-Like Receptors (KIR) Types on Heparin-Induced Thrombocytopenia (HIT).

Science.gov (United States)

Karnes, Jason H; Shaffer, Christian M; Cronin, Robert; Bastarache, Lisa; Gaudieri, Silvana; James, Ian; Pavlos, Rebecca; Steiner, Heidi E; Mosley, Jonathan D; Mallal, Simon; Denny, Joshua C; Phillips, Elizabeth J; Roden, Dan M

2017-09-01

Heparin-induced thrombocytopenia (HIT) is an unpredictable, life-threatening, immune-mediated reaction to heparin. Variation in human leukocyte antigen (HLA) genes is now used to prevent immune-mediated adverse drug reactions. Combinations of HLA alleles and killer cell immunoglobulin-like receptors (KIR) are associated with multiple autoimmune diseases and infections. The objective of this study is to evaluate the association of HLA alleles and KIR types, alone or in the presence of different HLA ligands, with HIT. HIT cases and heparin-exposed controls were identified in BioVU, an electronic health record coupled to a DNA biobank. HLA sequencing and KIR type imputation using Illumina OMNI-Quad data were performed. Odds ratios for HLA alleles and KIR types and HLA*KIR interactions using conditional logistic regressions were determined in the overall population and by race/ethnicity. Analysis was restricted to KIR types and HLA alleles with a frequency greater than 0.01. The p values for HLA and KIR association were corrected by using a false discovery rate qHIT cases and 350 matched controls were identified. No statistical differences in baseline characteristics were observed between cases and controls. The HLA-DRB3*01:01 allele was significantly associated with HIT in the overall population (odds ratio 2.81 [1.57-5.02], p=2.1×10 -4 , q=0.02) and in individuals with European ancestry, independent of other alleles. No KIR types were associated with HIT, although a significant interaction was observed between KIR2DS5 and the HLA-C1 KIR binding group (p=0.03). The HLA-DRB3*01:01 allele was identified as a potential risk factor for HIT. This class II HLA gene and allele represent biologically plausible candidates for influencing HIT pathogenesis. We found limited evidence of the role of KIR types in HIT pathogenesis. Replication and further study of the HLA-DRB3*01:01 association is necessary. © 2017 Pharmacotherapy Publications, Inc.

Elimination of heparin interference during microarray processing of fresh and biobank-archived blood samples.

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Hebels, Dennie G A J; van Herwijnen, Marcel H M; Brauers, Karen J J; de Kok, Theo M C M; Chalkiadaki, Georgia; Kyrtopoulos, Soterios A; Kleinjans, Jos C S

2014-07-01

In the context of environmental health research, biobank blood samples have recently been identified as suitable for high-throughput omics analyses enabling the identification of new biomarkers of exposure and disease. However, blood samples containing the anti-coagulant heparin could complicate transcriptomic analysis because heparin may inhibit RNA polymerase causing inefficient cRNA synthesis and fluorophore labelling. We investigated the inhibitory effect of heparin and the influence of storage conditions (0 or 3 hr bench times, storage at room temperature or -80°C) on fluorophore labelling in heparinized fresh human buffy coat and whole blood biobank samples during the mRNA work-up protocol for microarray analysis. Subsequently, we removed heparin by lithium chloride (LiCl) treatment and performed a quality control analysis of LiCl-treated biobank sample microarrays to prove their suitability for downstream data analysis. Both fresh and biobank samples experienced varying degrees of heparin-induced inhibition of fluorophore labelling, making most samples unusable for microarray analysis. RNA derived from EDTA and citrate blood was not inhibited. No effect of bench time was observed but room temperature storage gave slightly better results. Strong correlations were observed between original blood sample RNA yield and the amount of synthesized cRNA. LiCl treatment restored sample quality to normal standards in both fresh and biobank samples and the previously identified correlations disappeared. Microarrays hybridized with LiCl-treated biobank samples were of excellent quality with no identifiable influence of heparin. We conclude that, to obtain high quality results, in most cases heparin removal is essential in blood-derived RNA samples intended for microarray analysis. Copyright © 2014 Wiley Periodicals, Inc.

Transgenic expression of the rice Xa21 pattern-recognition receptor in banana (Musa sp.) confers resistance to Xanthomonas campestris pv. musacearum.

Science.gov (United States)

Tripathi, Jaindra N; Lorenzen, Jim; Bahar, Ofir; Ronald, Pamela; Tripathi, Leena

2014-08-01

Banana Xanthomonas wilt (BXW), caused by the bacterium Xanthomonas campestris pv. musacearum (Xcm), is the most devastating disease of banana in east and central Africa. The spread of BXW threatens the livelihood of millions of African farmers who depend on banana for food security and income. There are no commercial chemicals, biocontrol agents or resistant cultivars available to control BXW. Here, we take advantage of the robust resistance conferred by the rice pattern-recognition receptor (PRR), XA21, to the rice pathogen Xanthomonas oryzae pv. oryzae (Xoo). We identified a set of genes required for activation of Xa21-mediated immunity (rax) that were conserved in both Xoo and Xcm. Based on the conservation, we hypothesized that intergeneric transfer of Xa21 would confer resistance to Xcm. We evaluated 25 transgenic lines of the banana cultivar 'Gonja manjaya' (AAB) using a rapid bioassay and 12 transgenic lines in the glasshouse for resistance against Xcm. About 50% of the transgenic lines showed complete resistance to Xcm in both assays. In contrast, all of the nontransgenic control plants showed severe symptoms that progressed to complete wilting. These results indicate that the constitutive expression of the rice Xa21 gene in banana results in enhanced resistance against Xcm. Furthermore, this work demonstrates the feasibility of PRR gene transfer between monocotyledonous species and provides a valuable new tool for controlling the BXW pandemic of banana, a staple food for 100 million people in east Africa. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Preparation and characterization of chitosan-heparin composite matrices for blood contacting tissue engineering

International Nuclear Information System (INIS)

He Qing; Gong Kai; Gong Yandao; Zhang Xiufang; Ao Qiang; Zhang Lihai; Hu Min

2010-01-01

Chitosan has been widely used for biomaterial scaffolds in tissue engineering because of its good mechanical properties and cytocompatibility. However, the poor blood compatibility of chitosan has greatly limited its biomedical utilization, especially for blood contacting tissue engineering. In this study, we exploited a polymer blending procedure to heparinize the chitosan material under simple and mild conditions to improve its antithrombogenic property. By an optimized procedure, a macroscopically homogeneous chitosan-heparin (Chi-Hep) blended suspension was obtained, with which Chi-Hep composite films and porous scaffolds were fabricated. X-ray photoelectron spectroscopy and sulfur elemental analysis confirmed the successful immobilization of heparin in the composite matrices (i.e. films and porous scaffolds). Toluidine blue staining indicated that heparin was distributed homogeneously in the composite matrices. Only a small amount of heparin was released from the matrices during incubation in normal saline for 10 days. The composite matrices showed improved blood compatibility, as well as good mechanical properties and endothelial cell compatibility. These results suggest that the Chi-Hep composite matrices are promising candidates for blood contacting tissue engineering.

Heparin-Induced Thrombocytopenia in a Patient with Essential Thrombocythemia: A Case Based Update

Directory of Open Access Journals (Sweden)

Edva Noel

2015-01-01

Full Text Available Vascular thrombosis is a common clinical feature of both essential thrombocythemia (ET and heparin-induced thrombocytopenia (HIT. The development of HIT in a patient with ET is rare and underrecognized. We report the case of a 77-year-old woman with preexisting ET, who was admitted with acute coronary syndrome, and IV heparin was started. She was exposed to unfractionated heparin (UFH 5 days prior to this admission. Decrease in platelet count was noted, and HIT panel was sent. Heparin was discontinued. Patient developed atrial fibrillation, and Dabigatran was started. On day three, patient also developed multiple tiny cerebral infarctions and acute right popliteal DVT. On day ten of admission, HIT panel was positive, and Dabigatran was changed to Lepirudin. Two days later, Lepirudin was also discontinued because patient developed pseudoaneurysm on the right common femoral artery at the site of cardiac catheterization access. A progressive increase in the platelet count was noted after discontinuing heparin. Physicians should be aware of the coexistence of HIT and ET, accompanied challenges of the prompt diagnosis, and initiation of appropriate treatment.

Covalent co-immobilization of heparin/laminin complex that with different concentration ratio on titanium surface for selectively direction of platelets and vascular cells behavior

International Nuclear Information System (INIS)

Wang, Jian; Chen, Yuan; Liu, Tao; Wang, Xue; Liu, Yang; Wang, Yuan; Chen, Junying; Huang, Nan

2014-01-01

Highlights: • Extracellular matrix inspired surface modification with fibronectin, heparin and VEGF to construct a favorable microenvironment for selectively anticoagulant and promote endothelialization. • Take the advantage of specific intermolecular interaction, the bioactivity of above biomolecules was more efficiently maintained in compared with the common used covalent immobilization method. • Poly-l-lysine was used as a novel interlayer for surface amination, and in comparison, PLL coating was more feasible and the degradation product had no harm to human body. - Abstract: Surface biofunctional modification of coronary artery stent to improve the hemocompatibility and selectively accelerate endothelium regeneration but prevent restenosis have been become a new hotspot. For this, a novel method was developed in this work by co-immobilization of Ln and heparin complex on poly-L-lysine modified Ti surface. Take the advantage of the specific interaction between Ln and heparin, Ln and heparin complexes with different concentration ratios were set up for creating different exposure density of these two types of biomolecules. According to biocompatibility evaluation results, the Hep/Ln complexes modified surface displayed less platelet adhesion and activation. Especially, on L(150)H and L(200)H surface, the AT III binding quantity, APTT value and anti-coagulation property of modified surface were significantly promoted. Furthermore, the adherent density and proliferation activity of ECs and EPCs were positively correlated with Ln concentration. Notably, the proliferation of both ECs and EPCs on L(100)H, L(150)H and L(200)H surface were greatly promoted. Another hand, the proliferation activity of SMCs was significantly inhibited on Hep/Ln modified surfaces, which was considered mainly due to the inhibitory effect of heparin to SMCs. According to the existing results, this study demonstrated that in a certain range of heparin and laminin concentration ratio

Comparative study on collagen-binding enzyme-linked immunosorbent assay and ristocetin cofactor activity assays for detection of functional activity of von Willebrand factor.

Science.gov (United States)

Turecek, Peter L; Siekmann, Jürgen; Schwarz, Hans Peter

2002-04-01

For more than two decades, the ristocetin cofactor (RCo) assay, which measures the von Willebrand factor (vWF)-mediated agglutination of platelets in the presence of the antibiotic ristocetin, has been the most common method for measuring the functional activity of vWF. There is, however, general agreement among clinical analysts that this method has major practical disadvantages in performance and reproducibility. Today, collagen-binding assays (CBA) based on the enzyme-linked immunosorbent assay (ELISA) technique that measure the interaction of vWF and collagen are an alternative analytic procedure based on a more physiological function than that of the RCo procedure. We used both assay systems in a comparative study to assess the functional activity of vWF in plasma as well as in therapeutic preparations. We measured RCo activities of plasma from healthy donors and patients with different types of von Willebrand disease (vWD) and of vWF as a drug substance in factor (F) VIII/vWF concentrates using both the aggregometric and the macroscopic methods. In addition, we measured collagen-binding activity (vWF:CB) using a recently developed commercially available CBA system. To investigate the relation between the structure and the functional activity of vWF, we isolated vWF species with different numbers of multimers from FVIII/vWF concentrates by affinity chromatography on immobilized heparin. The vWF:RCo and vWF:CB of the different fractions were measured, and the multimeric structure of vWF was analyzed by sodium dodecyl sulfate (SDS) agarose gel electrophoresis. (vWF:CB and vWF:RCo are part of the nomenclature proposed by the International Society on Thrombosis and Hemostasis Scientific and Standardization Committee [ISTH SSC] subcommittee on von Willebrand factor, in Maastricht, Germany, June 16, 2000.) Measurement of functional vWF activity by CBA can be carried out with substantially higher interassay reproducibility than can measurement of RCo. Both assay

Heparin interferes with the radioenzymatic and homogeneous enzyme immunoassays for aminoglycosides

International Nuclear Information System (INIS)

Krogstad, D.J.; Granich, G.G.; Murray, P.R.; Pfaller, M.A.; Valdes, R.

1981-01-01

Heparin interferes with measurement of aminoglycosides in serum by biological, radioenzymatic, and homogeneous enzyme immunoassay techniques, but not with radioimmunoassay. At concentrations greater than or equal to 10 5 and greater than or equal to 3 X 10 6 USP units/L, respectively, it interferes with the radioenzymatic assay by inhibiting the gentamicin 3-acetyltransferase and kanamycin 6'-acetyltransferase enzymes used in the assay. It interferes with the homogeneous enzyme immunoassays for gentamicin and tobramycin (at concentrations greater than or equal to 10 5 and greater than or equal to10 4 USP units/L, respectively), but not with the commercially available homogeneous enzyme immunoassays for other drugs. Heparin interference with the homogeneous enzyme immunoassay for aminoglycosides requires both the heparin polyanion and glucose-6-phosphate dehydrogenase bound to a cationic aminoglycoside. This interference can be reproduced with dextran sulfate (but not dextran), and does not occur with free enzyme (glucose-6-phosphate dehydrogenase) alone. Heparin interference with these two assays and at concentrations that may be present in intravenous infusions or in seriously underfilled blood-collection tubes is described

Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

Energy Technology Data Exchange (ETDEWEB)

Loegel, Thomas N.; Trombley, John D.; Taylor, Richard T. [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States); Danielson, Neil D., E-mail: danielnd@muohio.edu [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States)

2012-11-13

Highlights: Black-Right-Pointing-Pointer Ethylenediamine is likely acting as an ion-pairing agent. Black-Right-Pointing-Pointer Oversulfated chondroitin sulfate is last peak instead of first peak. Black-Right-Pointing-Pointer There is about a factor of five improved detectability with a 12.5 min analysis time. Black-Right-Pointing-Pointer Use of a 50 {mu}m ID capillary is possible. - Abstract: This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with -14 V applied across a 50 {mu}m ID Multiplication-Sign 24.5 cm fused silica capillary at 15 Degree-Sign C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

International Nuclear Information System (INIS)

Loegel, Thomas N.; Trombley, John D.; Taylor, Richard T.; Danielson, Neil D.

2012-01-01

Highlights: ► Ethylenediamine is likely acting as an ion-pairing agent. ► Oversulfated chondroitin sulfate is last peak instead of first peak. ► There is about a factor of five improved detectability with a 12.5 min analysis time. ► Use of a 50 μm ID capillary is possible. - Abstract: This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with −14 V applied across a 50 μm ID × 24.5 cm fused silica capillary at 15 °C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

78 FR 24677 - Safety Zone; XA The Experimental Agency Fireworks, Pier 34, East River, NY

Science.gov (United States)

2013-04-26

... Final Rule This rule establishes a temporary safety zone on the navigable waters of the East River, in... Fireworks, Pier 34, East River, NY. (a) Regulated Area. The following area is a temporary safety zone: all... 1625-AA00 Safety Zone; XA The Experimental Agency Fireworks, Pier 34, East River, NY AGENCY: Coast...

Prevention of equine herpesvirus myeloencephalopathy - Is heparin a novel option? A case report.

Science.gov (United States)

Walter, Jasmin; Seeh, Christoph; Fey, Kerstin; Bleul, Ulrich; Osterrieder, Nikolaus

2016-10-12

Equine herpesvirus myeloencephalopathy (EHM) is a severe manifestation of equine herpesvirus 1 (EHV-1) infection. Prevention and treatment of EHM during EHV-1 outbreaks is critical, but no reliable and tested specific medication is available. Due to the thromboischemic nature of EHM and due to the fact that EHV-1 entry in cells is blocked by heparin, it was hypothesized that this compound may be useful in reduction of EHM incidence and severity. Therefore, during an acute EHV-1 outbreak with the neuropathogenic G 2254 /D 752 Pol variant, metaphylactic treatment with heparin to prevent EHM was initiated. Clinical signs were present in 61 horses (fever n = 55; EHM n = 8; abortion n = 6). Heparin (25000 IU subcutaneously twice daily for 3 days) was given to 31 febrile horses from day 10 of the outbreak, while the first 30 horses exhibiting fever remained untreated. Treatment outcome was analyzed retrospectively. Heparin-treated horses showed a lower EHM incidence (1/31; 3.2%) than untreated horses (7/30; 23.3%; p = 0.03). Results indicate that heparin may be useful for prevention of EHM during an EHV-1 outbreak. These promising data highlight the need for randomized and possibly blinded studies for the use of heparin in EHV-1 outbreaks.

Preparation of Low Molecular Weight Heparin by Microwave Discharge Electrodeless Lamp/TiO2 Photo-Catalytic Reaction.

Science.gov (United States)

Lee, Do-Jin; Kim, Byung Hoon; Kim, Sun-Jae; Kim, Jung-Sik; Lee, Heon; Jung, Sang-Chul

2015-01-01

An MDEL/TiO2 photo-catalyst hybrid system was applied, for the first time, for the production of low molecular weight heparin. The molecular weight of produed heparin decreased with increasing microwave intensity and treatment time. The abscission of the chemical bonds between the constituents of heparin by photo-catalytic reaction did not alter the characteristics of heparin. Formation of by-products due to side reaction was not observed. It is suggested that heparin was depolymerized by active oxygen radicals produced during the MDEL/TiO2 photo-chemical reaction.

Cost-utility of enoxaparin compared with unfractionated heparin in unstable coronary artery disease

Directory of Open Access Journals (Sweden)

Milne Ruairidh

2001-10-01

Full Text Available Abstract Background Low molecular weight heparins hold several advantages over unfractionated heparin including convenience of administration. Enoxaparin is one such heparin licensed in the UK for use in unstable coronary artery disease (unstable stable angina and non-Q wave myocardial infarction. In these patients, two large randomised controlled trials and their meta-analysis showed small benefits for enoxaparin over unfractionated heparin at 30–43 days and potentially at one year. We found no relevant published full economic evaluations, only cost studies, one of which was conducted in the UK. The other studies, from the US, Canada and France, are difficult to interpret since their resource use and costs may not reflect UK practice. Methods We aimed to compare the benefits and costs of short-term treatment (two to eight days with enoxaparin and unfractionated heparin in unstable coronary artery disease. We used published data sources to estimate the incremental cost per quality adjusted life year (QALY, adopting a NHS perspective and using 1998 prices. Results The base case was a 0.013 QALY gain and net cost saving of £317 per person treated with enoxaparin instead of unfractionated heparin. All but one sensitivity analysis showed net savings and QALY gains, the exception (the worst case being a cost per QALY of £3,305. Best cases were a £495 saving and 0.013 QALY gain, or a £317 saving and 0.014 QALY gain per person. Conclusions Enoxaparin appears cost saving compared with unfractionated heparin in patients with unstable coronary artery disease. However, cost implications depend on local revascularisation practice.

Two-step purification of His-tagged Nef protein in native condition using heparin and immobilized metal ion affinity chromatographies.

Science.gov (United States)

Finzi, Andrés; Cloutier, Jonathan; Cohen, Eric A

2003-07-01

The Nef protein encoded by human immunodeficiency virus type 1 (HIV-1) has been shown to be an important factor of progression of viral growth and pathogenesis in both in vitro and in vivo. The lack of a simple procedure to purify Nef in its native conformation has limited molecular studies on Nef function. A two-step procedure that includes heparin and immobilized metal ion affinity chromatographies (IMACs) was developed to purify His-tagged Nef (His(6)-Nef) expressed in bacteria in native condition. During the elaboration of this purification procedure, we identified two closely SDS-PAGE-migrating contaminating bacterial proteins, SlyD and GCHI, that co-eluted with His(6)-Nef in IMAC in denaturing condition and developed purification steps to eliminate these contaminants in native condition. Overall, this study describes a protocol that allows rapid purification of His(6)-Nef protein expressed in bacteria in native condition and that removes metal affinity resin-binding bacterial proteins that can contaminate recombinant His-tagged protein preparation.

Heparan sulfate C5-epimerase is essential for heparin biosynthesis in mast cells.

Science.gov (United States)

Feyerabend, Thorsten B; Li, Jin-Ping; Lindahl, Ulf; Rodewald, Hans-Reimer

2006-04-01

Biosynthesis of heparin, a mast cell-derived glycosaminoglycan with widespread importance in medicine, has not been fully elucidated. In biosynthesis of heparan sulfate (HS), a structurally related polysaccharide, HS glucuronyl C5-epimerase (Hsepi) converts D-glucuronic acid (GlcA) to L-iduronic acid (IdoA) residues. We have generated Hsepi-null mouse mutant mast cells, and we show that the same enzyme catalyzes the generation of IdoA in heparin and that 'heparin' lacking IdoA shows a distorted O-sulfation pattern.

Heparin-induced increase in serum levels of aminotranferases. A controlled clinical trial.

Science.gov (United States)

Nielsen, H K; Husted, S E; Koopmann, H D; Fasting, H; Simonsen, O; Andersen, K; Husegaard, H C; Petersen, T K

1984-01-01

Sixty-four patients over the age of 40 years, undergoing elective surgery of at least one hour's duration, were randomized to treatment with either a thromboembolic deterrent ( TED ) stocking (Kendall Co.) or subcutaneous low-dose heparin 5 000 IU every 12 hours. Serum levels of alanine aminotransferase (S-ALAT), aspartate aminotransferase (S-ASAT), gamma-glutamyl transpeptidase (S-gamma-GT) and alkaline phosphatase (S-ALP) were measured. S-ALAT increased significantly on the 5th and 10th postoperative day, from 27 +/- 2 (x +/- SE) to 40 +/- 4 (p less than 0.01) and 55 +/- 7 U/l (p less than 0.001), respectively, in the heparin group and was significantly higher in the heparin than in the TED group both on the 5th (p less than 0.01) and 10th (p less than 0.05) postoperative day. S-ASAT and S-gamma-GT increased significantly during heparin treatment, but did not differ significantly from the values of the TED group. No change in S-ALP was registered in either group. It is concluded that prophylactic treatment with low-dose heparin induces a significant increase in S-aminotransferase levels, especially in S-ALAT. The phenomenon has profound differential diagnostic implications in conditions such as pulmonary embolism and acute myocardial infarction.

Enhanced effect of BCG vaccine against pulmonary Mycobacterium tuberculosis infection in mice with lung Th17 response to mycobacterial heparin-binding hemagglutinin adhesin antigen.

Science.gov (United States)

Fukui, Masayuki; Shinjo, Kikuko; Umemura, Masayuki; Shigeno, Satoko; Harakuni, Tetsuya; Arakawa, Takeshi; Matsuzaki, Goro

2015-12-01

Although the BCG vaccine can prevent tuberculosis (TB) in infants, its ability to prevent adult pulmonary TB is reportedly limited. Therefore, development of a novel effective vaccine against pulmonary TB has become an international research priority. We have previously reported that intranasal vaccination of mice with a mycobacterial heparin-binding hemagglutinin adhesin (HBHA) plus mucosal adjuvant cholera toxin (CT) enhances production of IFN-γ and anti-HBHA antibody and suppresses extrapulmonary bacterial dissemination after intranasal infection with BCG. In the present study, the effects of intranasal HBHA + CT vaccine on murine pulmonary Mycobacterium tuberculosis (Mtb) infection were examined. Intranasal HBHA + CT vaccination alone failed to reduce the bacterial burden in the infected lung. However, a combination vaccine consisting of s.c. BCG priming and an intranasal HBHA + CT booster significantly enhanced protective immunity against pulmonary Mtb infection on day 14 compared with BCG vaccine alone. Further, it was found that intranasal HBHA + CT vaccine enhanced not only IFN-γ but also IL-17A production by HBHA-specific T cells in the lung after pulmonary Mtb infection. Therefore, this combination vaccine may be a good candidate for a new vaccine strategy against pulmonary TB. © 2015 The Societies and Wiley Publishing Asia Pty Ltd.

Alternative method for determination of contaminated heparin using chiral recognition.

Science.gov (United States)

Szekely, J; Collins, M; Currie, C A

2014-05-15

Since 2008 a significant amount of work has focused on the development of methods to analyze contaminated heparin. This work focuses on utilizing heparin's ability to serve as a chiral selector as a means for determining contamination. Specifically, the effect of contamination on the separation of pheniramine and chloroquine enantiomers was explored. Separations were conducted using heparin contaminated with chondroitin sulfate at varying levels. For each pair of enantiomers, electrophoretic mobility and resolution were calculated. For pheniramine enantiomers, an increase in contamination leads to a decrease in the electrophoretic mobility and resolution. A linear relationship between contamination level and electrophoretic mobility of the pheniramine enantiomers was observed for the entire contamination range. A linear relationship was also found between contamination level and resolution of the enantiomers between 0 and 70 percent contamination. For the separation of chloroquine enantiomers, it was found that at low levels of contamination, the resolution of enantiomers was increased due to the secondary interaction between the chloroquine enantiomers and the chondroitin sulfate. Results of this study illustrate the potential of using chiral recognition as a means to determine heparin contamination as well as the improvement of the chiral resolution of chloroquine with the additional of low levels of chondroitin sulfate A. Copyright © 2014 Elsevier B.V. All rights reserved.

Comprehensive human transcription factor binding site map for combinatory binding motifs discovery.

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Arnoldo J Müller-Molina

Full Text Available To know the map between transcription factors (TFs and their binding sites is essential to reverse engineer the regulation process. Only about 10%-20% of the transcription factor binding motifs (TFBMs have been reported. This lack of data hinders understanding gene regulation. To address this drawback, we propose a computational method that exploits never used TF properties to discover the missing TFBMs and their sites in all human gene promoters. The method starts by predicting a dictionary of regulatory "DNA words." From this dictionary, it distills 4098 novel predictions. To disclose the crosstalk between motifs, an additional algorithm extracts TF combinatorial binding patterns creating a collection of TF regulatory syntactic rules. Using these rules, we narrowed down a list of 504 novel motifs that appear frequently in syntax patterns. We tested the predictions against 509 known motifs confirming that our system can reliably predict ab initio motifs with an accuracy of 81%-far higher than previous approaches. We found that on average, 90% of the discovered combinatorial binding patterns target at least 10 genes, suggesting that to control in an independent manner smaller gene sets, supplementary regulatory mechanisms are required. Additionally, we discovered that the new TFBMs and their combinatorial patterns convey biological meaning, targeting TFs and genes related to developmental functions. Thus, among all the possible available targets in the genome, the TFs tend to regulate other TFs and genes involved in developmental functions. We provide a comprehensive resource for regulation analysis that includes a dictionary of "DNA words," newly predicted motifs and their corresponding combinatorial patterns. Combinatorial patterns are a useful filter to discover TFBMs that play a major role in orchestrating other factors and thus, are likely to lock/unlock cellular functional clusters.

Preparation of tin -heparin complex to be applied for myocardial infarct diagnosis

International Nuclear Information System (INIS)

Badi, J. M.; Al-Azzawi, H. A.; Resen, H. M.; Abed, I. G.; Owiad, H.; Manji, A. N.

2012-12-01

Tin-heparin complex has been prepared (liquid form) to be labeled with technetium-99 can be applied for diagnosis of myocardial infarcts vascular diseases and deep vein thrombosis. The preparation contents are 0.1mg tin chloride dehydrate and 1250 1.U of heparin. The results of the pH effect on the labeling yield indicated that high percentage of labeling yield (96.1%) was obtained in the optimal pH (5.50). The obtained results showed that the quantity of reducing agent (tin chloride dehydrate) and chelating agent (heparin) has no effect on the labeling yield. Results of radio analytical studies by paper chromatography technique wear confirmed by data obtained by Gel chromatography column scanning techniques. These techniques showed the high labeling yield of the tin-heparin complex. The persistence of high labeling yield for 8 hours is a good indication for its stability and efficiency for radio diagnosis examination in nuclear medicine centers. (Author)

Human corpus luteum: presence of epidermal growth factor receptors and binding characteristics

International Nuclear Information System (INIS)

Ayyagari, R.R.; Khan-Dawood, F.S.

1987-01-01

Epidermal growth factor receptors are present in many reproductive tissues but have not been demonstrated in the human corpus luteum. To determine the presence of epidermal growth factor receptors and its binding characteristics, we carried out studies on the plasma cell membrane fraction of seven human corpora lutea (days 16 to 25) of the menstrual cycle. Specific epidermal growth factor receptors were present in human corpus luteum. Insulin, nerve growth factor, and human chorionic gonadotropin did not competitively displace epidermal growth factor binding. The optimal conditions for corpus luteum-epidermal growth factor receptor binding were found to be incubation for 2 hours at 4 degrees C with 500 micrograms plasma membrane protein and 140 femtomol 125 I-epidermal growth factor per incubate. The number (mean +/- SEM) of epidermal growth factor binding sites was 12.34 +/- 2.99 X 10(-19) mol/micrograms protein; the dissociation constant was 2.26 +/- 0.56 X 10(-9) mol/L; the association constant was 0.59 +/- 0.12 X 10(9) L/mol. In two regressing corpora lutea obtained on days 2 and 3 of the menstrual cycle, there was no detectable specific epidermal growth factor receptor binding activity. Similarly no epidermal growth factor receptor binding activity could be detected in ovarian stromal tissue. Our findings demonstrate that specific receptors for epidermal growth factor are present in the human corpus luteum. The physiologic significance of epidermal growth factor receptors in human corpus luteum is unknown, but epidermal growth factor may be involved in intragonadal regulation of luteal function

Effects of chronic intraventricular infusion of heparin glycosaminoglycan on learning and brain acetylcholine parameters in aged rats

Czech Academy of Sciences Publication Activity Database

Ježek, Karel; Schulz, D.; De Souza Silva, M. A.; Müller, H. W.; Huston, J. P.; Hasenöhrl, R. U.

2003-01-01

Roč. 147, 1-2 (2003), s. 115-123 ISSN 0166-4328 Grant - others:Deutsche Forschungsgemeinschaft(DE) HU 306/11-3 Institutional research plan: CEZ:AV0Z5011922 Keywords : heparin * memory * ageing Subject RIV: FH - Neurology Impact factor: 2.817, year: 2003

High-throughput differentiation of heparin from other glycosaminoglycans by pyrolysis mass spectrometry.

Science.gov (United States)

Nemes, Peter; Hoover, William J; Keire, David A

2013-08-06

Sensors with high chemical specificity and enhanced sample throughput are vital to screening food products and medical devices for chemical or biochemical contaminants that may pose a threat to public health. For example, the rapid detection of oversulfated chondroitin sulfate (OSCS) in heparin could prevent reoccurrence of heparin adulteration that caused hundreds of severe adverse events including deaths worldwide in 2007-2008. Here, rapid pyrolysis is integrated with direct analysis in real time (DART) mass spectrometry to rapidly screen major glycosaminoglycans, including heparin, chondroitin sulfate A, dermatan sulfate, and OSCS. The results demonstrate that, compared to traditional liquid chromatography-based analyses, pyrolysis mass spectrometry achieved at least 250-fold higher sample throughput and was compatible with samples volume-limited to about 300 nL. Pyrolysis yielded an abundance of fragment ions (e.g., 150 different m/z species), many of which were specific to the parent compound. Using multivariate and statistical data analysis models, these data enabled facile differentiation of the glycosaminoglycans with high throughput. After method development was completed, authentically contaminated samples obtained during the heparin crisis by the FDA were analyzed in a blinded manner for OSCS contamination. The lower limit of differentiation and detection were 0.1% (w/w) OSCS in heparin and 100 ng/μL (20 ng) OSCS in water, respectively. For quantitative purposes the linear dynamic range spanned approximately 3 orders of magnitude. Moreover, this chemical readout was successfully employed to find clues in the manufacturing history of the heparin samples that can be used for surveillance purposes. The presented technology and data analysis protocols are anticipated to be readily adaptable to other chemical and biochemical agents and volume-limited samples.

The Factor of Structure and Mechanical Properties in the Production of Critical Fixing Hardware 38XA

Directory of Open Access Journals (Sweden)

G.V. Pachurin

2016-09-01

Full Text Available Fixing hardware made of carbon, high-carbon, and alloyed wires is one of the widespread critical parts in mechanical engineering. The characteristics of fixtures and fasteners and their performance figures are assessed at all stages of steel-making operation, from the choice of burden stock for metal smelting to the method of preparing calibrated rolled metal parts and upsetting end products. Material used for producing long high-duty bolts must be both sufficiently strong and ductile, have homogeneous mechanical properties and chemistry, and no inner or surface defects. When manufacturing fasteners, hot-rolled metal is often plastically deformed by drafting before cold upsetting, and all unacceptable defects are removed by the expensive procedure of lathe-turning. Moreover, this technology of metal processing entails losses of up to 5.5 % in chips. This paper suggests a resource-efficient and environmentally friendlier fabrication method for calibrated rolled metal items made of steel 38XA, 9.65 mm diameter, for cold die-forging of high-duty bolts used in automotive engines, which helps spare expensive lathe-machining processes. Moreover, rolled steel produced using this technology is characterized by high resistance to plastic deformation when cold, which leads to increase n wear-resistance of tools in cold die-forging of bolts.

[Obstetrical APS: Is there a place for additional treatment to aspirin-heparin combination?

Science.gov (United States)

Mekinian, A; Kayem, G; Cohen, J; Carbillon, L; Abisror, N; Josselin-Mahr, L; Bornes, M; Fain, O

2017-01-01

Obstetrical APS is defined by thrombosis and/or obstetrical morbidity associated with persistent antiphospholipid antibodies. The aspirin and low molecular weighted heparin combination dramatically improved obstetrical outcome in APS patients. Several factors could be associated with obstetrical prognosis, as previous history of thrombosis, associated SLE, the presence of lupus anticoagulant and triple positivity of antiphospholipid antibodies. Obstetrical APS with isolated recurrent miscarriages is mostly associated with isolated anticardiolipids antibodies and have better obstetrical outcome. The pregnancy loss despite aspirin and heparin combination define the refractory obstetrical APS, and the prevalence could be estimated to 20-39%. Several other treatments have been used in small and open labeled studies, as steroids, intravenous immunoglobulins, plasma exchanges and hydroxychloroquine to improve the obstetrical outcome. Some other drugs as eculizumab and statins could also have physiopathological rational, but studies are necessary to define the place of these various drugs. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

A radioisotope dilution assay for unlabelled vitamin B12-intrinsic factor complex employing the binding intrinsic factor antibody: probable evidence for two types of binding antibody

International Nuclear Information System (INIS)

Jacob, E.; O'Brien, H.A.W.; Mollin, D.L.

1977-01-01

A new radioisotope dilution assay for vitamin B 12 -intrinsic factor complex is described. The method is based on the use of the binding type intrinsic antibody (the binding reagent), which when combined with the intrinsic factor-vitamin B 12 complex (labelled ligand), is quantitatively adsorbed onto zirconium phosphate gel pH 6.25. The new assay has been shown to provide a measure of intrinsic factor comparable with other intrinsic factor assays, but it has the important advantage of being able to measure the unlabelled vitamin B 12 -intrinsic factor complex (unlabelled ligand), and will, therefore, be valuable in the study of physiological events in the gastrointestinal tract. During the study, it was found that there is some evidence for at least two types of binding intrinsic factor antibody: One which combines preferentially with the intrinsic factor-vitamin B 12 complex and one which combines equally well with this complex or with free intrinsic factor. (author)