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Pesquisa : D08.811.520.241.700.512 [Categoria DeCS]
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[PMID]:28465034
[Au] Autor:Antia IU; Yagnik DR; Pantoja Munoz L; Shah AJ; Hills FA
[Ad] Endereço:Glycan Research Group, Department of Natural Sciences, Faculty of Science and Technology, Middlesex University, The Burroughs, London NW4 4BT, UK. Electronic address: i.antia@mdx.ac.uk.
[Ti] Título:Heparan sulfate disaccharide measurement from biological samples using pre-column derivatization, UPLC-MS and single ion monitoring.
[So] Source:Anal Biochem;530:17-30, 2017 08 01.
[Is] ISSN:1096-0309
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Glycosaminoglycans are a heterogeneous family of linear polysaccharides comprised of repeating disaccharide subunits that mediate many effects at the cellular level. There is increasing evidence that the nature of these effects is determined by differences in disaccharide composition. However, the determination of GAG disaccharide composition in biological samples remains challenging and time-consuming. We have developed a method that uses derivatization and selected ion recording and RP-UPLCMS resulting in rapid separation and quantification of twelve heparin/heparin sulfate disaccharides from 5 µg GAG. Limits of detection and quantitation were 0.02-0.15 and 0.07-0.31 µg/ml respectively. We have applied this method to the novel analysis of disaccharide levels extracted from heparan sulfate and human cancer cell lines. Heparan sulfate disaccharides extracted from biological samples following actinase and heparinase incubation and derivatized using reductive amination with 2-aminoacridone. Derivatized disaccharides were analyzed used UPLC-MS with single ion monitoring. Eight HS disaccharide subunits were separated and quantified from HS and cell lines in eleven minutes per sample. In all samples the most abundant subunits present were the unsulfated ΔUA-GlcNAc, ΔUA-GlcNAc,6S and ΔUA,2S-GlcNS,6S. There was considerable variation in the proportions and concentrations of disaccharides between different cell lines. Further studies are needed to examine the significance of these differences.
[Mh] Termos MeSH primário: Cromatografia Líquida de Alta Pressão/métodos
Dissacarídeos/análise
Heparina/análogos & derivados
Heparitina Sulfato/análise
Espectrometria de Massas/métodos
Neoplasias/metabolismo
[Mh] Termos MeSH secundário: Aminoacridinas/química
Dissacarídeos/química
Dissacarídeos/isolamento & purificação
Heparina/análise
Heparina/química
Heparina/isolamento & purificação
Heparina Liase/metabolismo
Heparitina Sulfato/química
Heparitina Sulfato/isolamento & purificação
Seres Humanos
Células Tumorais Cultivadas
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Aminoacridines); 0 (Disaccharides); 0 (heparin disaccharide); 27918-14-5 (2-aminoacridone); 9005-49-6 (Heparin); 9050-30-0 (Heparitin Sulfate); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1708
[Cu] Atualização por classe:180207
[Lr] Data última revisão:
180207
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170504
[St] Status:MEDLINE


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[PMID]:28867189
[Au] Autor:Li S; Zhang F; Cui Y; Wu M; Lee C; Song J; Cao C; Chen H
[Ad] Endereço:Department of Cardiology, Peking University People's Hospital, Beijing, 100044, China; Beijing Key Laboratory of Early Prediction and Intervention of Acute Myocardial Infarction, Peking University People's Hospital, Beijing, 100044, China; Center for Cardiovascular Translational Research, Peking Uni
[Ti] Título:Modified high-throughput quantification of plasma microRNAs in heparinized patients with coronary artery disease using heparinase.
[So] Source:Biochem Biophys Res Commun;493(1):556-561, 2017 Nov 04.
[Is] ISSN:1090-2104
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:Heparin, a widely used anticoagulant in cardiovascular diseases, is notorious for its inhibitory effect on qRT-PCR-based detection. Heparinase I could degrade heparin in RNA. qRT-PCR-based TaqMan Low Density Array (TLDA) technology is commonly used for circulating microRNAs (miRNAs) profiling analysis. However, the effect of heparin contamination on inhibition of miRNAs TLDA amplification, as well as the method for removing heparin during this process, are not yet well investigated. We obtained the plasma RNA samples from patients undergoing percutaneous coronary intervention (PCI) before and after heparinization (n = 26). We found that heparin suppressed the miRNAs amplification by ∼8 cycles in the TLDA assay, which was absolutely reversed after treating the RNA samples with heparinase I using the components from TLDA reverse transcription system. We further observed that heparin inhibited the miRNAs amplification by ∼4 cycles in the qRT-PCR assay, which was also reversed by heparinase I using the similar method. Furthermore, we demonstrated that plasma miR-92a and miR-155 were differentially expressed in the patients undergoing PCI tested by TLDA assay, which was validated by qRT-PCR. In conclusion, we present a simple method for the removal of heparin with heparinase I, and for the subsequent successful miRNAs TLDA or RT-qPCR amplification.
[Mh] Termos MeSH primário: Doença da Artéria Coronariana/sangue
Heparina Liase/sangue
Heparina/sangue
Ensaios de Triagem em Larga Escala/métodos
MicroRNAs/sangue
Reação em Cadeia da Polimerase/métodos
[Mh] Termos MeSH secundário: Artefatos
Biomarcadores/sangue
Análise Química do Sangue/métodos
Seres Humanos
Reprodutibilidade dos Testes
Sensibilidade e Especificidade
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Biomarkers); 0 (MicroRNAs); 9005-49-6 (Heparin); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1710
[Cu] Atualização por classe:171023
[Lr] Data última revisão:
171023
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170905
[St] Status:MEDLINE


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[PMID]:28581441
[Au] Autor:Putz EM; Mayfosh AJ; Kos K; Barkauskas DS; Nakamura K; Town L; Goodall KJ; Yee DY; Poon IK; Baschuk N; Souza-Fonseca-Guimaraes F; Hulett MD; Smyth MJ
[Ad] Endereço:Immunology in Cancer and Infection Laboratory, QIMR Berghofer Medical Research Institute, Herston, Queensland, Australia.
[Ti] Título:NK cell heparanase controls tumor invasion and immune surveillance.
[So] Source:J Clin Invest;127(7):2777-2788, 2017 Jun 30.
[Is] ISSN:1558-8238
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:NK cells are highly efficient at preventing cancer metastasis but are infrequently found in the core of primary tumors. Here, have we demonstrated that freshly isolated mouse and human NK cells express low levels of the endo-ß-D-glucuronidase heparanase that increase upon NK cell activation. Heparanase deficiency did not affect development, differentiation, or tissue localization of NK cells under steady-state conditions. However, mice lacking heparanase specifically in NK cells (Hpsefl/fl NKp46-iCre mice) were highly tumor prone when challenged with the carcinogen methylcholanthrene (MCA). Hpsefl/fl NKp46-iCre mice were also more susceptible to tumor growth than were their littermate controls when challenged with the established mouse lymphoma cell line RMA-S-RAE-1ß, which overexpresses the NK cell group 2D (NKG2D) ligand RAE-1ß, or when inoculated with metastatic melanoma, prostate carcinoma, or mammary carcinoma cell lines. NK cell invasion of primary tumors and recruitment to the site of metastasis were strictly dependent on the presence of heparanase. Cytokine and immune checkpoint blockade immunotherapy for metastases was compromised when NK cells lacked heparanase. Our data suggest that heparanase plays a critical role in NK cell invasion into tumors and thereby tumor progression and metastases. This should be considered when systemically treating cancer patients with heparanase inhibitors, since the potential adverse effect on NK cell infiltration might limit the antitumor activity of the inhibitors.
[Mh] Termos MeSH primário: Heparina Liase/imunologia
Vigilância Imunológica
Células Matadoras Naturais/imunologia
Neoplasias Experimentais/imunologia
[Mh] Termos MeSH secundário: Animais
Linhagem Celular Tumoral
Citocinas/genética
Citocinas/imunologia
Feminino
Heparina Liase/genética
Seres Humanos
Células Matadoras Naturais/patologia
Masculino
Camundongos
Camundongos Knockout
Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética
Subfamília K de Receptores Semelhantes a Lectina de Células NK/imunologia
Invasividade Neoplásica/genética
Invasividade Neoplásica/imunologia
Metástase Neoplásica
Neoplasias Experimentais/genética
Neoplasias Experimentais/patologia
Proteínas Associadas à Matriz Nuclear/genética
Proteínas Associadas à Matriz Nuclear/imunologia
Proteínas de Transporte Nucleocitoplasmático/genética
Proteínas de Transporte Nucleocitoplasmático/imunologia
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Cytokines); 0 (KLRK1 protein, human); 0 (NK Cell Lectin-Like Receptor Subfamily K); 0 (Nuclear Matrix-Associated Proteins); 0 (Nucleocytoplasmic Transport Proteins); 0 (RAE1 protein, human); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1709
[Cu] Atualização por classe:170930
[Lr] Data última revisão:
170930
[Sb] Subgrupo de revista:AIM; IM
[Da] Data de entrada para processamento:170606
[St] Status:MEDLINE


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[PMID]:28300590
[Au] Autor:Bhushan I; Alabbas A; Sistla JC; Saraswat R; Desai UR; Gupta RB
[Ad] Endereço:Department of Biotechnology, Shri Mata Vaishno Devi University, Katra, J&K 182320, India; Department of Chemical and Life Science Engineering, Virginia Commonwealth University, Richmond, VA 23284-3068, USA; Institute for Structural Biology, Drug Discovery and Development Virginia Commonwealth Un
[Ti] Título:Heparin depolymerization by immobilized heparinase: A review.
[So] Source:Int J Biol Macromol;99:721-730, 2017 Jun.
[Is] ISSN:1879-0003
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:Heparin is a member of the glycosaminoglycan (GAG) family composed of glucosamine and uronic acid units containing O-sulfo, N-acetyl and N-sulfo groups, which are alternating in the chain and linked by 1→4 manner. It is a naturally occurring anticoagulant that prevents the formation of clots and their growth within blood. Certain low molecular weight heparins (LMWHs) are considered as better therapeutic agents than natural heparin because of the reduced side effects and smaller risk of bleeding. LMWHs can be produced from heparin by chemical or enzymatic depolymerizations. Heparinases catalyze the cleavage of glycosidic linkage between amino sugars and uronic acids in heparin. There are three kinds of heparinases which are frequently used for depolymerization of heparin. Despite wide range of applications of heparinases in health care, their use still has been hampered due to poor stability and high cost. To overcome this problem heparinases are recommended for immobilization to reduce the cost of product and enhance stability. Heparinases have been successfully immobilized using various methods and supports, mostly for deheparinization of blood through extracorporeal devices. The focus of this review is to present the current status of heparinase immobilization including various supports and methods used, stability and applications.
[Mh] Termos MeSH primário: Enzimas Imobilizadas/química
Enzimas Imobilizadas/metabolismo
Heparina Liase/química
Heparina Liase/metabolismo
Heparina/química
Polimerização
[Mh] Termos MeSH secundário: Animais
Assistência à Saúde
Seres Humanos
Peso Molecular
[Pt] Tipo de publicação:JOURNAL ARTICLE; REVIEW
[Nm] Nome de substância:
0 (Enzymes, Immobilized); 9005-49-6 (Heparin); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1705
[Cu] Atualização por classe:170509
[Lr] Data última revisão:
170509
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170317
[St] Status:MEDLINE


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[PMID]:28086125
[Au] Autor:Mehta PK; Lee H; Lee KH
[Ad] Endereço:Center for Design and Applications of Molecular Catalysts, Department of Chemistry and Chemical Engineering, Inha University, Incheon 402-751, South Korea.
[Ti] Título:Highly sensitive ratiometric detection of heparin and its oversulfated chondroitin sulfate contaminant by fluorescent peptidyl probe.
[So] Source:Biosens Bioelectron;91:545-552, 2017 May 15.
[Is] ISSN:1873-4235
[Cp] País de publicação:England
[La] Idioma:eng
[Ab] Resumo: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 =0.99) in aqueous solutions, 2.98nM (R =0.98) in 1% serum samples, and 3.43nM (R =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.
[Mh] Termos MeSH primário: Anticoagulantes/sangue
Sulfatos de Condroitina/sangue
Corantes Fluorescentes/química
Heparina/sangue
Peptídeos/química
Espectrometria de Fluorescência/métodos
[Mh] Termos MeSH secundário: Anticoagulantes/análise
Anticoagulantes/metabolismo
Técnicas Biossensoriais/métodos
Sulfatos de Condroitina/análise
Sulfatos de Condroitina/metabolismo
Contaminação de Medicamentos
Corantes Fluorescentes/metabolismo
Heparina/análise
Heparina/metabolismo
Heparina Liase/metabolismo
Seres Humanos
Limite de Detecção
Peptídeos/metabolismo
Polissacarídeo-Liase/metabolismo
Pirenos/química
Pirenos/metabolismo
[Pt] Tipo de publicação:EVALUATION STUDIES; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Anticoagulants); 0 (Fluorescent Dyes); 0 (Peptides); 0 (Pyrenes); 9005-49-6 (Heparin); 9007-28-7 (Chondroitin Sulfates); 9E0T7WFW93 (pyrene); EC 4.2.2.- (Polysaccharide-Lyases); EC 4.2.2.- (heparinase II); EC 4.2.2.7 (Heparin Lyase); EC 4.2.2.8 (heparitinsulfate lyase)
[Em] Mês de entrada:1703
[Cu] Atualização por classe:170531
[Lr] Data última revisão:
170531
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:170114
[St] Status:MEDLINE


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[PMID]:28002635
[Au] Autor:Cuq B; Dunn ME; Bédard C
[Ad] Endereço:Department of Clinical Sciences, Faculty of Veterinary Medicine, Université de Montréal, St Hyacinthe, QC, Canada.
[Ti] Título:Heparinase-modified thromboelastography in cats.
[So] Source:J Vet Emerg Crit Care (San Antonio);27(1):127-130, 2017 Jan.
[Is] ISSN:1476-4431
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:BACKGROUND: Evaluation of underlying hemostatic function is challenging when feline patients are receiving an anticoagulant medication. Discontinuing the anticoagulant to obtain accurate results for hemostatic testing may lead to thrombotic complications. The addition of heparinase to blood samples may mitigate the effects of exogenous heparin and allow hemostatic testing. METHODS: Tissue factor (TF)-activated thromboelastography (TEG) was performed using citrated whole blood from 19 cats. Assays were performed using citrated whole blood, with and without addition of unfractionated heparin to a concentration of 0.2 U/mL. For each blood sample, TEG assays were performed using a standard cup and a heparinase-coated cup. KEY FINDINGS: For TEG variables R, k, α-angle, and MA, mean values were not statistically different when citrated blood was used with standard or heparinase-coated cups. Heparinized blood assayed in standard cups displayed a significantly increased R and k, and significantly decreased α-angle and MA when compared to heparinized blood assayed in heparinase-coated cups. TEG variables for heparinized blood assayed in heparinase cups was not statistically different from those of the citrated whole blood without added heparin. SIGNIFICANCE: Heparinase modified, TF-activated, TEG reverses heparin effects in feline-citrated blood.
[Mh] Termos MeSH primário: Anticoagulantes/farmacologia
Coagulação Sanguínea/efeitos dos fármacos
Heparina Liase/farmacologia
Tromboelastografia/veterinária
[Mh] Termos MeSH secundário: Animais
Anticoagulantes/administração & dosagem
Testes de Coagulação Sanguínea/veterinária
Gatos
Feminino
Heparina Liase/administração & dosagem
Masculino
[Pt] Tipo de publicação:EVALUATION STUDIES; JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Anticoagulants); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1704
[Cu] Atualização por classe:170817
[Lr] Data última revisão:
170817
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161222
[St] Status:MEDLINE
[do] DOI:10.1111/vec.12559


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[PMID]:27923477
[Au] Autor:Sun X; Guo Z; Yu M; Lin C; Sheng A; Wang Z; Linhardt RJ; Chi L
[Ad] Endereço: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 Biologi
[Ti] Título:Hydrophilic interaction chromatography-multiple reaction monitoring mass spectrometry method for basic building block analysis of low molecular weight heparins prepared through nitrous acid depolymerization.
[So] Source:J Chromatogr A;1479:121-128, 2017 Jan 06.
[Is] ISSN:1873-3778
[Cp] País de publicação:Netherlands
[La] Idioma:eng
[Ab] Resumo:Low molecular weight heparins (LMWHs) are important anticoagulant drugs that are prepared through depolymerization of unfractionated heparin. Based on the types of processing reactions and the structures of the products, LMWHs can be divided into different classifications. Enoxaparin is prepared by benzyl esterification and alkaline depolymerization, while dalteparin and nadroparin are prepared through nitrous acid depolymerization followed by borohydride reduction. Compositional analysis of their basic building blocks is an effective way to provide structural information on heparin and LMWHs. However, most current compositional analysis methods have been limited to heparin and enoxaparin. A sensitive and comprehensive approach is needed for detailed investigation of the structure of LMWHs prepared through nitrous acid depolymerization, especially their characteristic saturated non-reducing end (NRE) and 2,5-anhydro-d-mannitol reducing end (RE). A maltose modified hydrophilic interaction column offers improved separation of complicated mixtures of acidic disaccharides and oligosaccharides. A total of 36 basic building blocks were unambiguously identified by high-resolution tandem mass spectrometry (MS). Multiple reaction monitoring (MRM) MS/MS quantification was developed and validated in the analysis of dalteparin and nadroparin samples. Each group of building blocks revealed different aspects of the properties of LMWHs, such as functional motifs required for anticoagulant activity, the structure of heparin starting materials, cleavage sites in the depolymerization reaction, and undesired structural modifications resulting from side reactions.
[Mh] Termos MeSH primário: Heparina de Baixo Peso Molecular/química
Heparina/química
Ácido Nitroso/química
[Mh] Termos MeSH secundário: Cromatografia Líquida
Heparina/metabolismo
Heparina Liase/metabolismo
Heparina de Baixo Peso Molecular/isolamento & purificação
Interações Hidrofóbicas e Hidrofílicas
Nadroparina/análise
Espectrometria de Massas em Tandem
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Heparin, Low-Molecular-Weight); 0 (Nadroparin); 9005-49-6 (Heparin); EC 4.2.2.7 (Heparin Lyase); T2I5UM75DN (Nitrous Acid)
[Em] Mês de entrada:1703
[Cu] Atualização por classe:170817
[Lr] Data última revisão:
170817
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161208
[St] Status:MEDLINE


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[PMID]:27725938
[Au] Autor:Wu CS; Lin FC; Chen SJ; Chen YL; Chung WJ; Cheng CI
[Ad] Endereço:Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan.
[Ti] Título:Optimized Collection Protocol for Plasma MicroRNA Measurement in Patients with Cardiovascular Disease.
[So] Source:Biomed Res Int;2016:2901938, 2016.
[Is] ISSN:2314-6141
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:. Various microRNAs (miRNAs) are used as markers of acute coronary syndrome, in which heparinization is considered mandatory therapy. Nevertheless, a standard method of handling plasma samples has not been proposed, and the effects of heparin treatment on miRNA detection are rarely discussed. . This study used quantitative polymerase chain reaction (qPCR) analysis to investigate how storage temperature, standby time, hemolysis, and heparin treatment affect miRNA measurement in plasma samples from 25 patients undergoing cardiac catheterization. . For most miRNAs, the qPCR results remained consistent during the first 2 hours. The miRNA signals did not significantly differ between samples stored at 4°C before processing and samples stored at room temperature (RT) before processing. miR-451a/miR-23a ratio < 60 indicated < 0.12% hemolysis with 100% sensitivity and 100% specificity. Pretreatment with 0.25 U heparinase I recovered qPCR signals that were reduced by heparinization. . For miRNA measurement, blood samples stored at RT should be processed into plasma within 2 hours after withdrawal and should be pretreated with 0.25 U heparinase I to overcome heparin-attenuated miRNA signals. The miR-451a/miR-23a ratio is a reliable indicator of significant hemolysis.
[Mh] Termos MeSH primário: Coleta de Amostras Sanguíneas/métodos
Doenças Cardiovasculares/sangue
Doenças Cardiovasculares/genética
MicroRNAs/sangue
MicroRNAs/genética
[Mh] Termos MeSH secundário: Adulto
Idoso
Primers do DNA/metabolismo
Sondas de DNA/metabolismo
Demografia
Feminino
Regulação da Expressão Gênica
Hemólise
Heparina Liase/metabolismo
Seres Humanos
Masculino
Meia-Idade
Reação em Cadeia da Polimerase
Preservação Biológica
Temperatura Ambiente
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (DNA Primers); 0 (DNA Probes); 0 (MicroRNAs); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1702
[Cu] Atualização por classe:170224
[Lr] Data última revisão:
170224
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:161012
[St] Status:MEDLINE


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[PMID]:27598820
[Au] Autor:Roest HP; Verhoeven CJ; de Haan JE; de Jonge J; IJzermans JN; van der Laan LJ
[Ad] Endereço:Department of Surgery, Erasmus MC-University Medical Center, Rotterdam, the Netherlands. Electronic address: h.roest@erasmusmc.nl.
[Ti] Título:Improving Accuracy of Urinary miRNA Quantification in Heparinized Patients Using Heparinase I Digestion.
[So] Source:J Mol Diagn;18(6):825-833, 2016 Nov.
[Is] ISSN:1943-7811
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:miRNAs have emerged as promising biomarkers because of their association with cell stress and diseases and their easy detection and stability in many body fluids. Because of the sensitivity, the method of choice to detect miRNAs is quantitative RT-PCR (RT-qPCR). Therapeutics, in particular circulating anticoagulants, are notorious for their inhibitory effect on RT-qPCR-based measurements. The effect of heparin contamination on inhibition of RT-qPCR from miRNAs isolated from urine has, however, never been investigated. We obtained urine samples from healthy controls and from heparinized patients undergoing major surgery (live kidney donation or liver transplantation) (n = 27). Samples were spiked with synthetic miRNAs to monitor RNA loss during workup, and levels of endogenous and spiked-in miRNAs were quantified by RT-qPCR. Endogenous miRNAs in urine were protected from degradation, but levels differed substantially within surgery groups. Variability in detection levels of spiked-in miRNAs was low in nonhospitalized controls, but was high in both surgery groups, and the difference in miRNA levels correlated well with the heparin concentration in urinary samples. Treatment of urinary RNA with heparinase I during RT-qPCR strongly reduced this variation in a dose-dependent manner. Heparinase I should therefore be considered as standard step for detection of miRNA in urine from hospitalized individuals.
[Mh] Termos MeSH primário: Heparina Liase
Heparina
MicroRNAs/genética
Reação em Cadeia da Polimerase em Tempo Real
[Mh] Termos MeSH secundário: Adulto
Anticoagulantes/efeitos adversos
Biomarcadores
Estudos de Casos e Controles
Feminino
Heparina/efeitos adversos
Seres Humanos
Masculino
MicroRNAs/urina
Meia-Idade
Proteólise
Estabilidade de RNA
Reação em Cadeia da Polimerase em Tempo Real/métodos
Reação em Cadeia da Polimerase em Tempo Real/normas
Reprodutibilidade dos Testes
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Anticoagulants); 0 (Biomarkers); 0 (MicroRNAs); 9005-49-6 (Heparin); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1706
[Cu] Atualização por classe:170615
[Lr] Data última revisão:
170615
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160907
[St] Status:MEDLINE


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[PMID]:27461795
[Au] Autor:Willems A; Savan V; Faraoni D; De Ville A; Rozen L; Demulder A; Van der Linden P
[Ad] Endereço:Pediatric Intensive Care Unit, Department of Pediatrics, Queen Fabiola University Children's Hospital, Brussels, Belgium. Electronic address: ariane.willems@huderf.be.
[Ti] Título:Heparin Reversal After Cardiopulmonary Bypass: Are Point-of-Care Coagulation Tests Interchangeable?
[So] Source:J Cardiothorac Vasc Anesth;30(5):1184-9, 2016 Oct.
[Is] ISSN:1532-8422
[Cp] País de publicação:United States
[La] Idioma:eng
[Ab] Resumo:OBJECTIVE: Protamine is used to neutralize heparin after patient separation from cardiopulmonary bypass (CPB). Different bedside tests are used to monitor the adequacy of heparin neutralization. For this study, the interchangeability of the activated coagulation time (ACT) and thromboelastometry (ROTEM; Tem Innovations GmbH, Basel, Switzerland) clotting time (CT) ratios in children undergoing cardiac surgery was assessed. DESIGN: Single-center, retrospective, cohort study between September 2010 and January 2012. SETTING: University children's hospital. PARTICIPANTS: The study comprised children 0 to 16 years old undergoing elective cardiac surgery with CPB. Exclusion criteria were preoperative coagulopathy, Jehovah's witnesses, and children in a moribund condition (American Society of Anesthesiologists score 5). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: After heparin neutralization with protamine, the ratio between ACT, with and without heparinase, and the CT measured with INTEM/HEPTEM (intrinsic test activated with ellagic acid was performed without heparinase [INTEM] and with heparinase [HEPTEM]) using tests of ROTEM were calculated. Agreement was evaluated using Cohen's kappa statistics, Passing-Bablok regression, and Bland-Altman analysis. Among the 173 patients included for analysis, agreement between both tests showed a Cohen's kappa statistic of 0.06 (95% CI: -0.02 to 0.14; p = 0.22). Bland-Altman analysis showed a bias of 0.01, with a standard deviation of 0.13, and limits of agreement between -0.24 and 0.26. Passing-Bablok regression showed a systematic difference of 0.40 (95% CI: 0.16-0.59) and a proportional difference of 0.61 (95% CI: 0.42-0.86). The residual standard deviation was 0.11 (95% CI: -0.22 to 0.22), and the test for linearity showed p = 0.10. CONCLUSION: ACT, with or without heparinase, and the INTEM/HEPTEM CT ratios are not interchangeable to evaluate heparin reversal after pediatric patient separation from CPB. Therefore, the results of these tests should be corroborated with the absence/presence of bleeding and integrated into center-specific treatment algorithms.
[Mh] Termos MeSH primário: Coagulação Sanguínea/efeitos dos fármacos
Ponte Cardiopulmonar
Antagonistas de Heparina/uso terapêutico
Sistemas Automatizados de Assistência Junto ao Leito
Cuidados Pós-Operatórios/métodos
[Mh] Termos MeSH secundário: Adolescente
Testes de Coagulação Sanguínea/métodos
Criança
Pré-Escolar
Heparina Liase/uso terapêutico
Seres Humanos
Lactente
Masculino
Protaminas/uso terapêutico
Estudos Retrospectivos
Tromboelastografia/efeitos dos fármacos
Tempo de Coagulação do Sangue Total
[Pt] Tipo de publicação:JOURNAL ARTICLE
[Nm] Nome de substância:
0 (Heparin Antagonists); 0 (Protamines); EC 4.2.2.7 (Heparin Lyase)
[Em] Mês de entrada:1708
[Cu] Atualização por classe:170817
[Lr] Data última revisão:
170817
[Sb] Subgrupo de revista:IM
[Da] Data de entrada para processamento:160728
[St] Status:MEDLINE



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