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INTRODUCTION: Thrombin generation assay (TGA) is a laboratory method that provides the global evaluation of hemostasis. The association between thrombin generation and all-cause mortality is poorly investigated and results are contradictory. This study evaluated whether TGA parameters are associated with all-cause mortality in a prospective cohort. METHODS: This study was conducted in 2,588 participants enrolled at baseline of the Brazilian Longitudinal Study of Adult Health (ELSA-Brasil). TGA was performed using the Calibrated Automated Thrombogram (CAT) method, and its parameters lagtime, time-to-peak, peak, Endogenous Thrombin Potential (ETP) and normalized ETP (nETP) were evaluated according to the reference interval (RI). The association between TGA parameters and all-cause mortality was estimated by Cox regression and adjusted for confounders. RESULTS: The mean follow-up time was 6.6 ± 2.7 years and 85 deaths occurred. After adjustment, time-to-peak values above the RI at low and high tissue factor (TF) concentrations were associated with higher risk of death [HR = 2.45 (95 % CI: 1.17-5.13) and HR = 2.24 (95 % CI: 1.02-4.93), respectively] and nETP and peak values below RI at high TF concentration were associated with higher risk of death [HR = 3.85 (95 % CI: 1.39-10.68) and HR = 2.56 (95 % CI: 1.17-5.61), respectively]. CONCLUSIONS: Delayed thrombin generation was associated with higher risk of all-cause mortality.
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Trombina , Adulto , Humanos , Testes de Coagulação Sanguínea , Brasil , Estudos Prospectivos , Estudos LongitudinaisRESUMO
Rapidly reversible anticoagulant agents have great clinical potential. Oligonucleotide-based anticoagulant agents are uniquely positioned to fill this clinical niche, as they are able to be deactivated through the introduction of the reverse complement oligo. Once the therapeutic and the antidote oligos meet in solution, they are able to undergo isothermal reassociation to form short, inactive, duplexes that are rapidly secreted via filtration by the kidneys. The formation of the duplexes interrupts the structure of the anticoagulant oligo, allowing normal coagulation to be restored. To effectively assess these new anticoagulants, a variety of methods may be employed. The measurement of thrombin generation (TG) reflects the overall capacity of plasma to produce active thrombin and provides a strong contribution to identifying new anticoagulant drugs, including DNA/RNA thrombin binding aptamer carrying fibers which are used through this chapter as an example. Here we describe the TG assessed by Calibrated Automated Thrombogram (CAT) assay in a fully automated system. This method is based on the detection of TG in plasma samples by measuring fluorescent signals released from a quenched fluorogenic thrombin substrate and the subsequent conversion of these signals in TG curves.
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Nanopartículas , Ácidos Nucleicos , Trombina/metabolismo , Anticoagulantes/farmacologia , Plasma/metabolismo , Corantes Fluorescentes , Testes de Coagulação Sanguínea/métodosRESUMO
Background: Twenty-minute whole blood clotting test (20WBCT) and Modified Lee and White (MLW) method are the most routinely employed bedside tests for detecting coagulopathic snake envenomation. Our study compared the diagnostic utility of MLW and 20WBCT for snakebite victims at a tertiary care hospital in Central Kerala, South India. Methods: This single-center study recruited 267 patients admitted with snake bites. 20WBCT and MLW were performed simultaneously at admission along with the measurement of Prothrombin Time (PT). The diagnostic utility of 20WBCT and MLW was determined by comparing the sensitivity (Sn), specificity (Sp), positive and negative predictive values, likelihood ratios, and accuracy at admission with an INR value > 1.4. Results: Out of 267 patients, 20 (7.5%) patients had VICC. Amongst those who had venom-induced consumption coagulopathy (VICC), MLW was prolonged for 17 patients, (Sn 85% 95% confidence interval [CI]: 61.1-96.0) whereas 20WBCT was abnormal for 11 patients (Sn 55%, 95% CI: 32.04-76.17). MLW and 20WBCT were falsely positive for the same patient (Sp 99.6%, 95% CI: 97.4-99.9%). Conclusion: MLW is more sensitive than 20WBCT to detect coagulopathy at the bedside amongst snakebite victims. However, further studies are necessary for standardizing bedside coagulation tests in snakebite cases.
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Resumen: Introducción: Las pruebas de coagulación carecen de valor para determinar el riesgo de sangrado perioperatorio. Material y métodos: Se realizó un estudio observacional, descriptivo, y transversal en 2,114 pacientes en la consulta de Anestesiología del Hospital Universitario «Dr. Celestino Hernández Robau¼, los resultados se evaluaron mediante estadística descriptiva. Resultados: El tiempo de coagulación y sangrado se realizó en 100% de los casos y el conteo de plaquetas en 93.99%, mientras que el tiempo de protrombina y tiempo de tromboplastina parcial activado se efectuó en 66.27 y 55.62% de los casos respectivamente. De 8.834 exámenes realizados se encontraron 49 alterados en 0.55%. Los pacientes con exámenes alterados fueron 33 en 1.56%, los enfermos en riesgo de sangrado por exámenes de coagulación fueron 30 en 1.42% y los pacientes en riesgo sin antecedentes de sangrados detectados por exámenes de coagulación fueron tres en 0.14%. Se reportó sangrado perioperatorio en 16 pacientes en 0.76%, siete pacientes con interrogatorio positivo y exámenes normales y nueve pacientes con interrogatorio negativo y exámenes normales. Conclusiones: La historia clínica y el examen físico del paciente son las mejores herramientas para predecir el riesgo de sangrado quirúrgico y los exámenes aislados de coagulación no constituyen un buen predictor del sangrado perioperatorio.
Abstract: Introduction: Coagulation tests are no value to determine the risk of perioperative bleeding. Material and methods: An observational descriptive cross-sectional study was carried out in 2,114 patients in the anesthesiology consultation of the University Hospital «Dr. Celestino Hernández Robau¼. Results: The clotting and bleeding time was performed in 100% of cases, the platels count in 93.99%. While the prothrombin time and activated partial tromboplastin time were performed in 66.27 and 55.62% respectively. Of 8,834 tests carried out, 49 were found to be altered for 0.55%. Patients with altered tests were for 1.56%, patients at risk of bleeding from coagulation tests were 30 for 1.42% and patients at risk with no history of bleeding detected by coagulation tests were three for 0.14%. Perioperative bleeding was reported in 16 patients for 0.76%, seven patients with positive questioning and normal tests and nine patients with negative questioning and normal tests. Conclusions: The patient's medical history and physical examination are the best tools to predict the risk of surgical bleeding and isolated coagulation tests do not constitute a good predictor of perioperative bleeding.
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There is a trend towards increased perioperative bleeding in patients with plasma fibrinogen levels < 200 mg/dL-1. This study aimed to assess whether there is an association between preoperative fibrinogen levels and perioperative blood-product transfusion up to 48 h after major orthopedic surgery. This cohort study included 195 patients who underwent primary or revision hip arthroplasty for nontraumatic etiologies. Plasma fibrinogen, blood count, coagulation tests, and platelet count were measured preoperatively. A plasma fibrinogen level of 200 mg/dL-1 was the cutoff value used to predict blood transfusion. The mean (SD) plasma fibrinogen level was 325 (83) mg/dL-1. Only thirteen patients had levels < 200 mg/dL-1, and only one of them received a blood transfusion, with an absolute risk of 7.69% (1/13; 95%CI: 1.37-33.31%). Preoperative plasma fibrinogen levels were not associated with the need for blood transfusion (p = 0.745). The sensitivity and the positive predictive value of plasma fibrinogen < 200 mg/dL-1 as a predictor of blood transfusion were 4.17% (95%CI: 0.11-21.12%) and 7.69% (95%CI: 1.12-37.99%), respectively. Test accuracy was 82.05% (95%CI: 75.93-87.17%), but positive and negative likelihood ratios were poor. Therefore, preoperative plasma fibrinogen level in hip-arthroplasty patients was not associated with the need for blood-product transfusion.
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Abstract Background: Twenty-minute whole blood clotting test (20WBCT) and Modified Lee and White (MLW) method are the most routinely employed bedside tests for detecting coagulopathic snake envenomation. Our study compared the diagnostic utility of MLW and 20WBCT for snakebite victims at a tertiary care hospital in Central Kerala, South India. Methods: This single-center study recruited 267 patients admitted with snake bites. 20WBCT and MLW were performed simultaneously at admission along with the measurement of Prothrombin Time (PT). The diagnostic utility of 20WBCT and MLW was determined by comparing the sensitivity (Sn), specificity (Sp), positive and negative predictive values, likelihood ratios, and accuracy at admission with an INR value > 1.4. Results: Out of 267 patients, 20 (7.5%) patients had VICC. Amongst those who had venom-induced consumption coagulopathy (VICC), MLW was prolonged for 17 patients, (Sn 85% 95% confidence interval [CI]: 61.1-96.0) whereas 20WBCT was abnormal for 11 patients (Sn 55%, 95% CI: 32.04-76.17). MLW and 20WBCT were falsely positive for the same patient (Sp 99.6%, 95% CI: 97.4-99.9%). Conclusion: MLW is more sensitive than 20WBCT to detect coagulopathy at the bedside amongst snakebite victims. However, further studies are necessary for standardizing bedside coagulation tests in snakebite cases.
Assuntos
Tempo de Protrombina/métodos , Mordeduras de Serpentes/diagnóstico , Transtornos da Coagulação Sanguínea/diagnóstico , Fatores de Coagulação Sanguínea/análiseRESUMO
Resumen El objetivo de este trabajo fue comparar los niveles de fibrinógeno (FBG) obtenidos por el método de Clauss con los obtenidos por el método de fibrinógeno derivado del tiempo de protrombina (FBG PT-d), con dos tromboplastinas, en pacientes anticoagulados con distintas drogas. Se estudiaron pacientes anticoagulados consecutivos: 105 con antagonistas de la vitamina K (AVK), 55 con heparina no fraccionada (HNF), 58 con heparina de bajo peso molecular (HBPM), 60 con rivaroxabán, 45 con apixabán, 60 con dabigatrán y 100 controles normales (CN). El FBG se determinó por el método de Clauss y FBG PT-d utilizando tromboplastina de cerebro de conejo o recombinante humana; los niveles de heparina, rivaroxabán y apixabán por método cromogénico anti Xa; el dabigatrán con el ensayo de tiempo de trombina diluido. Existió un sesgo positivo (p<0,001) al comparar el FBG PT-d vs. FBG por Clauss: CN: 13,7%, AVK: 31,8%, rivaroxabán: 34,8% y apixabán: 20,0% cuando se utilizó tromboplastina de conejo. En el caso de las muestras que contenían HBPM se observó este desvío con ambas tromboplastinas. El sesgo porcentual en presencia de dabigatrán y heparina no fraccionada no fue estadísticamente distinto del obtenido en el grupo control. El ensayo de FBG PT-d no debe utilizarse en pacientes anticoagulados con rivaroxabán, apixabán, HBPM o AVK, ya que sobreestima los niveles de FBG. El porcentaje de sesgo depende del tipo de tromboplastina utilizado y fue mayor con la de cerebro de conejo en el sistema de detección utilizado.
Abstract The aim of this study was to compare fibrinogen (FBG) results obtained by Clauss method (FBG-C) and by the prothrombin time-derived fibrinogen assay (FBG PT-d) with two thromboplastins in patients under anticoagulation. Consecutive anticoagulated patients were studied: 105 vitamin-K antagonist (VKA), 55 unfractioned heparin, 58 LMWH, 60 rivaroxaban, 45 apixaban and 60 dabigatran, and 100 healthy controls (NC). FBG-C was performed by Clauss and FIB PT-d with rabbit brain and human recombinant thromboplastins, respectively. Heparins, rivaroxaban and apixaban levels were measured by antiXa; dabigatran by thrombin diluted assay. A positive bias of FBG PT-d vs. FBG-C with both thromboplastins were seen in NC (13.7 and 19.0 % for HS and RP, respectively), but bias with HS in rivaroxaban, apixaban and VKA patients were significantly higher compared to NC: 34.8%, 20.0 % and 31.8 %, respectively. LMWH presented higher BIAS compared to NC with both thromboplastins. Samples with unfraction heparin and dabigatran presented similar bias to NC. FBG PT-d should not be used in patients under anticoagulant treatment because of an important overestimation of FBG could be obtained in these patients. The percentage of bias depends on the type of thromboplastin used; it was higher with rabbit brain thromboplastin in the detection system used.
Resumo O objetivo deste trabalho foi comparar os níveis de fibrinogênio (FBG) obtidos pelo método de Clauss com aqueles obtidos pelo método do fibrinogênio derivado do tempo de protrombina (FBG PT-d), com duas tromboplastinas, em pacientes anticoagulados com diferentes drogas. Pacientes anticoagulados consecutivos foram estudados: 105 com antagonista da vitamina K (AVK); 55 com heparina não fracionada (UFH); 58 com heparina de baixo peso molecular (HBPM), 60 com rivaroxabana, 45 com apixabana, 60 com dabigatrana e 100 controles normais (CN). FBG foi determinado pelo método de Clauss e FBG PT-d usando tromboplastina de cérebro de coelho ou tromboplastina humana recombinante; níveis de heparina, rivaroxabana e apixabana pelo método cromogênico anti-Xa; dabigatrana com ensaio de tempo de trombina diluída. Há um viés positivo (p<0,001) ao comparar o FBG PT-d vs FBG de Clauss: CN: 13,7%; AVK: 31,8%, rivaroxabana: 34,8% e apixabana 20,0% quando foi utilizada tromboplastina de coelho. No caso das amostras contendo HBPM, esse desvio foi observado com ambas as tromboplastinas. O viés percentual na presença de dabigatrana e heparina não fracionada não foi estatisticamente diferente daquela obtida no grupo controle. O ensaio de FBG PT-d não deve ser usado em pacientes anticoagulados com rivaroxabana, apixabana, LMWH ou VKA, pois superestima os níveis de FBG. A porcentagem de viés depende do tipo de tromboplastina utilizado e foi maior com a de cérebro de coelho, no sistema de detecção utilizado.
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Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Fibrinogênio/análise , Protrombina/administração & dosagem , Coagulação Sanguínea , Tromboplastina , Preparações Farmacêuticas/administração & dosagemRESUMO
BACKGROUND AND OBJECTIVES: Neuraxial hematoma is a rare complication of the epidural technique which is commonly used for high quality postoperative pain relief. In case of urgent initiation of multiple antithrombotic therapy, the optimal timing of epidural catheter removal and need for treatment modification may be quite challenging. There are no specific guidelines and published reports are scarce. CASE REPORT: We present the uneventful removal of an indwelling epidural catheter in a patient who was put on emergency triple antithrombotic treatment with Low Molecular Weight Heparin (LMWH), aspirin and clopidogrel in the immediate postoperative period, due to acute coronary syndrome. In order to define the optimal conditions and timing for catheter removal, so as to reduce the risk of complications, various laboratory tests were conducted 3 hours after aspirin/clopidogrel intake. Standard coagulation tests revealed normal platelet count, normal prothrombin time and normal activated partial thromboplastin time, while Platelet Function Analysis (PFA-200) revealed abnormal values (increased COL/EPI and COL/ADP values, both indicating inhibition of platelet function). The anti-Xa level, estimated 4 hours after LMWH administration, was within therapeutic range. At the same time, Rotational Thromboelastometry (ROTEM) showed a relatively satisfactory coagulation status overall. The epidural catheter was removed 26 hours after the last dual antiplatelet dose and the next dose was given 2 hours after removal. Enoxaparin was withheld for 24 hours and was resumed after 6 hours. Neurologic checks were performed regularly for alarming signs and symptoms suggesting development of an epidural hematoma. No complications occurred. CONCLUSION: Point-of-care coagulation and platelet function monitoring may provide a helpful guidance in order to define the optimal timing for catheter removal, so as to reduce the risk of complications. A case-specific management plan based on a multidisciplinary approach is also important.
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Síndrome Coronariana Aguda , Fibrinolíticos , Síndrome Coronariana Aguda/terapia , Catéteres , Fibrinolíticos/uso terapêutico , Heparina de Baixo Peso Molecular , Humanos , Período Pós-OperatórioRESUMO
Abstract Introduction: The efficacy of 20-minute whole blood clotting (WBCT20) and the Lee-White clotting time (LWCT) tests in diagnosing coagulation alterations from snakebites were compared. Methods: We evaluated 89 snakebite cases treated at the Hospital Regional do Juruá em Cruzeiro do Sul, Acre, Brazil. Results: WBCT20 results were normal in 33.7% and unclottable in 66.3% of cases, while LWCT results were normal in 23.6% and altered (prolonged or unclottable) in 76.4% of cases, with no significant differences. Conclusions: The WBCT20 is important for rapidly diagnosing coagulation alterations from snakebites. Furthermore, it is efficient, inexpensive, and can be deployed in isolated hospitals.
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Humanos , Mordeduras de Serpentes/diagnóstico , Coagulação Sanguínea , Brasil , HospitaisRESUMO
OBJECTIVES: Despite more than 40 years of experience performing the Bethesda assay (BA), poor intra- and interlaboratory precision remains the biggest laboratory challenge to date. METHODS: The BA procedure was modeled using stochastic simulation techniques to determine the precision of the BA up to dilutions of 1:4,096, to estimate the minimum significant relative change at various inhibitor titers, and to understand the laboratory procedural variables that could significantly affect the performance of the BA at high dilutions. RESULTS: Selecting the lowest dilution tube with a residual activity closest to 25% for calculating the reported Bethesda titer (BT), using a factor activity assay with a coefficient of variation less than or equal toâ 7.5% in the range of 15% to 50% factor activity level, performing the factor activity measurement in replicates, and minimizing pipette volumetric error resulted in the lowest imprecision in the reported BT. The factor neutralization kinetics of the inhibitor appear to have little impact on the precision of the assay if the incubation time is greater than 90 minutes. CONCLUSIONS: This in silico model will assist future laboratory efforts in standardizing the quantification of specific coagulation factor inhibitors and improving the precision of the reported results.
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Testes de Coagulação Sanguínea/normas , Simulação por Computador , Testes de Coagulação Sanguínea/métodos , Humanos , Sensibilidade e EspecificidadeRESUMO
Abstract Background: Heparin decreases the risks of thrombotic phenomena in extracorporeal circulation. However, it must present a robust safety profile itself, especially for bleeding. Contamination of porcine heparin demands an alternative source and consequent assessment of safety. Objective: To evaluate the safety of unfractionated bovine heparin during on-pump cardiac surgery. Methods: Descriptive, retrospective study, evaluating medical records from all patients who had on-pump cardiac surgery over four years. We observed the occurrence of bleeding, thrombocytopenia, postoperative vasoplegia, activated clotting time values and any other coagulation phenomena as safety profile parameters. Results: We evaluated 204 medical records reporting the use of unfractionated bovine heparin. 66.18% of the patients presented thrombocytopenia, 1.04% presented bleeding of more than 2000 mL in the first 24 hours of the postoperative period. One patient presented clots in the surgical field. Median activated clotting time was 137 seconds at baseline, 803 seconds after the first dose of heparin and, after protamine, it returns to similar baseline values, that is, 149.5 seconds. Conclusion: Unfractionated bovine heparin did not present unusual adverse effects and can be considered safe for on-pump cardiac surgery.
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Humanos , Animais , Masculino , Feminino , Adolescente , Adulto , Pessoa de Meia-Idade , Idoso , Idoso de 80 Anos ou mais , Bovinos , Adulto Jovem , Heparina/uso terapêutico , Circulação Extracorpórea , Procedimentos Cirúrgicos Cardíacos , Período Pós-Operatório , Trombocitopenia , Testes de Coagulação Sanguínea , Heparina/efeitos adversos , Epidemiologia Descritiva , Estudos Retrospectivos , Hemorragia Pós-Operatória , VasoplegiaRESUMO
BACKGROUND/AIM: There are few studies in the literature evaluating possible alterations in laboratory tests in patients with maxillofacial fractures. The aim of this study was to analyze the changes in admission laboratory tests of patients with maxillofacial fractures with indications for surgical treatment, including the influence of dento-alveolar trauma. MATERIAL AND METHODS: Data from complete blood counts, blood coagulation tests, blood chemistries, and urinalysis results were obtained. The occurrence of concomitant dento-alveolar trauma was noted. The medical records were also re-evaluated later to verify the treatment outcome and the occurrence of complications. Statistical analyses were performed using the likelihood-ratio test to verify significant changes in the evaluated parameters (P ≤ .050). RESULTS: There was a prevalence of males (78%) with a mean age of 35.9 years. Lower erythrocyte counts, hemoglobin levels, and/or hematocrit were observed in males with associated fractures and with simultaneous dento-alveolar fractures. Higher mean neutrophil, segmented neutrophil, and lymphocyte counts were observed in patients with simultaneous dento-alveolar trauma. A higher mean activated partial thromboplastin time (aPTT) ratio was also observed. Lower potassium levels were observed for patients in the fourth decade of life. Higher leukocyte counts not associated with trauma were observed in the urinalysis results of females and in the group of patients aged 20 or younger. Verification of treatment outcome showed two cases of infections and two cases that needed re-operation after mandible fractures. These four cases showed no significant changes in laboratory tests regarding the predisposition for complications. CONCLUSION: Patients with maxillofacial fractures had neutrophilia, increased aPTT, and non-traumatic leukocyturia. There was an influence of associated fractures and dento-alveolar trauma on the decrease in red blood cell parameters, neutrophilia, and lymphocytosis and of age on hypokalemia.
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Traumatismos Maxilofaciais , Fraturas Cranianas , Traumatismos Dentários , Adulto , Ossos Faciais/lesões , Feminino , Hospitalização , Humanos , Masculino , Estudos Retrospectivos , Adulto JovemRESUMO
Background: It is necessary to establish biological parameters for each population. Objective: To establish reference values for prothrombin time (PT), activated partial thromboplastin time (PTT) and fibrinogen in healthy population and to determine intra- and inter-assay concordance. Methods: Cross-sectional study that included 204 women and 202 men from the Blood Donor service. Coagulation tests were carried out in order to obtain reference ranges. All procedures were made according to the Clinical & Laboratory Standards Institute guidelines. Results: Mean PT, PTT and fibrinogen were 14.1 s, 28.8 s and 381 mg/dL in men, and 15.1 s, 29.0 s and 381 mg/dL in women. The proposed PT, PTT and fibrinogen reference ranges for men were 12.7 to 16.3 s, 24.2 to 36.3 s and 239 to 276 mg/dL, respectively; for women, 12.7 to 16.6 s, 23.5 to 35.4 s and 276 to 598 mg/dL. The latter was statistically significant (p ≤ 0.001). Conclusions: Reference values for blood coagulation tests were determined. This is of great importance for fast medical diagnosis and treatment. The results from this study can be adopted by other clinic laboratories after appropriate validation procedures.
Introducción: Es necesario establecer valores de referencia biológicos para cada población. Objetivo: Establecer los límites de referencia de tiempo de protrombina (TP), tiempo parcial de tromboplastina (TTP) y fibrinógeno en población mestizo-mexicana sana, así como la correlación y la concordancia en la determinación de estas pruebas con los dos equipos utilizados. Métodos: Estudio transversal en 204 mujeres y 202 hombres que acudieron al servicio de donadores y se les determinó TP, TTP y fibrinógeno para obtener los límites de referencia. Los procedimientos se realizaron de acuerdo con las guías del Instituto de Estándares de Laboratorio y Clínicos (CLSI C28-A3). Resultados: La media de TP, TTP y fibrinógeno en hombres fue de 14.1 s, 28.8 s y 381 mg/dL, y en mujeres de 15.1 s, 29.0 s y 381 mg/dL, respectivamente. Los límites de referencia para hombres en TP, TTP y fibrinógeno fueron de 12.7 a 16.3 s, de 24.2 a 36.3 s y de 239 a 276 mg/dL; para mujeres de 12.7 a 16.6 s, de 23.5 a 35.4 s y de 276 a 598 mg/dL, respectivamente. Este último fue estadísticamente significativo (p ≤ 0.001). Conclusiones: Se determinaron los límites de referencia para las pruebas de coagulación. Los resultados obtenidos en este estudio pueden ser adoptados por otros laboratorios clínicos, después de su apropiada validación.
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Fibrinogênio/metabolismo , Tempo de Tromboplastina Parcial , Tempo de Protrombina , Adolescente , Adulto , Idoso , Biomarcadores/sangue , Estudos Transversais , Feminino , Humanos , Masculino , México , Pessoa de Meia-Idade , Valores de Referência , Adulto JovemRESUMO
Essentials Bovine (HBI) and porcine (HPI) heparins differ in structure and anticoagulant activity. Protamine-neutralization was evaluated on a variety of physical-chemical methods. HBI requires more protamine than HPI to fully neutralize its anticoagulant activity. Protamine preferentially removes higher-sulfated chains of HBI while HPI is evenly precipitated. SUMMARY: Background Protamine neutralization is an essential step for the safe use and inactivation of the unfractionated heparin (UFH) that is widely employed in surgical and non-surgical procedures involving extracorporeal circulation. Objective To compare protamine neutralization of different pharmaceutical-grade UFHs prepared from porcine or bovine intestine (HPI and HBI, respectively). HBI has approximately half the anticoagulant potency of HPI, mostly as consequence of its fraction enriched with N-sulfated α-glucosamine disaccharides. Methods Protamine neutralization of HPI and HBI was evaluated with in vitro, ex vivo and in vivo assays. We also performed in-depth assessments of the complexation of protamine with these distinct UFHs by using nuclear magnetic resonance and mass spectroscopy. Results HPI and HBI interact similarly with protamine on a mass/mass basis; however, HBI requires more protamine than HPI to have its anticoagulant activity fully neutralized, because of its lower potency, which entails the use of higher doses. Nuclear magnetic resonance spectra revealed that HPI precipitates homogeneously with protamine. On the other hand, the low-sulfated fraction of HBI, enriched with N-sulfated α-glucosamine, precipitates at higher concentrations of protamine than the fraction more like HPI, with a preponderance of N,6-disulfated α-glucosamine disaccharides. Finally, mass spectroscopy spectra showed that some of the different peptide components of protamine interact preferentially with the heparins, irrespective of their animal origin. Conclusion Our results have important medical implications, indicating that protamine neutralization of HBI, determined exclusively by point-of-care coagulation assessments, must fail because of its lower-sulfated fraction with reduced anticoagulant activity that could remain in the circulation after the neutralization procedure.
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Anticoagulantes/farmacologia , Antagonistas de Heparina/farmacologia , Heparina/farmacologia , Protaminas/farmacologia , Animais , Anticoagulantes/química , Anticoagulantes/isolamento & purificação , Bioensaio , Bovinos , Precipitação Química , Cromatografia de Afinidade , Dissacarídeos/química , Relação Dose-Resposta a Droga , Heparina/química , Heparina/isolamento & purificação , Mucosa Intestinal/química , Espectrometria de Massas , Ressonância Magnética Nuclear Biomolecular , Tempo de Tromboplastina Parcial , Protaminas/química , Ratos , Especificidade da Espécie , Enxofre/análise , SuínosRESUMO
Essentials Fibrinogen prothrombin time-derived (FIBPT-d) behavior in anticoagulated patients is under studied. FIBPT-d method overestimates fibrinogen in rivaroxaban and low molecular weight heparin samples. Unfractionated heparin and dabigatran samples showed similar bias to the control group. Rabbit brain and human recombinant thromboplastin behavior was different in rivaroxaban samples. SUMMARY: Background The fibrinogen prothrombin time-derived (FIBPT-d) method with photo-optical coagulometers is easy and economical. However, there are few reports on the behavior of this test on samples from patients anticoagulated with direct oral anticoagulants or low molecular weight heparin (LMWH). Objective To compare fibrinogen results obtained with the Clauss (FIB C) method and the FIBPT-d method with two thromboplastins in anticoagulated patients. Population The study population comprised 295 consecutive anticoagulated patients: 99 treated with vitamin K antagonists (VKAs), 49 treated with unfractionated heparin (UFH), 47 treated with LMWH, 50 treated with rivaroxaban, 50 treated with dabigatran, and 100 normal controls (NCs). Methods Dabigatran samples were analyzed by the use of FIB C with HemosIL Fibrinogen C or 100 NHI thrombin units mL-1 reagents; rabbit brain and human recombinant thromboplastins with HemosIL PTFibrinogen HS plus (HS) and Recombiplastin 2G (RP) were used for FIBPT-d method. Heparin and rivaroxaban levels were assessed with HemosIL Liq antiXa with specific calibrators; dabigatran levels were determined with the HemosIL Direct Thrombin Inhibitor Assay. All assays were performed on the ACL TOP platform in two laboratories. Percentage biases for the FIBPT-d method versus the FIB C method were calculated by the use of Bland-Altman plots. Results Positive biases of the FIBPT-d method versus the FIB C method with both thromboplastins were seen in NC samples (13.7% and 18.9% for HS and RP, respectively), but biases with HS in rivaroxaban and VKA patient samples were higher than that in NC samples, at 31.9% and 34.0%, respectively. LMWH patient samples showed higher bias than NC samples: 26.5% and 29.3.0% with HS and RP, respectively. UFH and dabigatran patient samples showed similar bias as NC samples. Conclusion The FIBPT-d method should not be used in anticoagulated patients, because the FIBPT-d mathematical algorithm has been validated only in normal subjects, so overestimation could occur in these patients.
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Background: Compared to standard coagulation essays (SCE), such as international normalized ratio (INR), prothrombin activated partial thromboplastin time (aPTT), or platelet count, thromboelastograhy (TEG) offers precise and real-time information about hemostasis. TEG tests both platelet function and coagulation by assaying several parameters of clot formation dynamically in whole blood. Aim: To evaluate hemostasis in cirrhotic patients undergoing liver transplantation and determine the positive and negative predictive values of SCE for alterations of TEG. Material and Methods: Preoperative SCE and TEG were prospectively analyzed in 25 patients. Results were categorized as normal, laboratory alteration or clinical alteration. SCE results were compared with TEG parameters to determine positive (PPV) and negative predictive values (NPV). Results: Hemostasis was abnormal and laboratory abnormalities were observed in all patients. One patient had clinical signs of excessive bleeding. SCE were abnormal in all patients and TEG was normal in nine patients. The most common alteration in TEG was hypocoagulability, in some cases associated with hypercoagulability and hyperfibrinolysis. Two patients had solely hypercoagulability. PPV of INR, aPTT, platelet count and fibrinogenemia were 0, 0, 0.5 and 0.17 respectively. NPV of the same tests were 1, 1, 0.34 and 1 respectively. Conclusions: Hypocoagulability was the most common laboratory alteration, however, clinical signs of coagulopathy were rarely present. SCE had a poor predictive value to diagnose o discard hemostatic abnormalities.
Assuntos
Humanos , Masculino , Feminino , Pessoa de Meia-Idade , Tromboelastografia , Testes de Coagulação Sanguínea , Transplante de Fígado , Período Pré-Operatório , Estudos ProspectivosRESUMO
The biotechnological value of macroalgae for screening assays of thrombin generation-TG using sulfated polysaccharides-SPs as substitutes to heparin has been poorly explored. Five Brazilian species of macroalgae (Gracilaria birdiae, Acanthophora muscoides, Halymenia sp., Caulerpa cupressoides and C. racemosa) wereanalyzed and compared for their abundance, physical-chemical characteristics and in vitro anticoagulant assays of activated partial thromboplastin time-APTT, prothrombin time-PT and TG. Papain extraction yielded (p 100 kDa. These procedures,combined with the use of Stains-All, also indicated nonSPs. APTTs ranged from 2.81 (A. muscoides) to 21.30 IU(Halymenia sp.) vs. heparin (193 IU), and were dependent on sulfation of the crude SPs. PT was not altered. Withrespect to TG assay, crude SPs modified concentration-dependent and independently from molecular mass TGby both intrinsic/extrinsic pathways in 60-fold diluted human plasma, with total intrinsic inactivation using crudeSPs from A. muscoides in parallel to heparin (p < 0.05). Thrombosis in vitro is differentially modulated by distinctcrude SPs from Brazilian seaweeds.
O valor biotecnológico das macroalgas para ensaios de varredura de geração de trombina-GT pouco tem sido explorado usando polissacarídeos sulfatados-PSs como substitutos à heparina. Foramanalisadas e comparadas cinco espécies brasileiras de macroalgas (Gracilaria birdiae, Acanthophora muscoides, Halymenia sp., Caulerpa cupressoides e C. racemosa) quanto à abundância, às característicasfísico-químicas e os ensaios anticoagulantes in vitro de tempo de tromboplastina parcial ativada-TTPA, aotempo de protrombina-TP e a GT. A extração com papaína rendeu (p 100 kDa. Esses procedimentos,combinados ao uso de azul de toluidina/Stains-All, indicaram também polissacarídeos-não sulfatados. OsTTPAs foram dependentes da sulfatação dos PSs brutos e variaram de 2,81 (A. muscoides) a 21,30 UI (Halymenia sp.) vs. heparina (193 UI). O TP não foi alterado. Com respeito ao ensaio de GT, os PSs brutos modificaram, dependente de concentração e independentemente de massa molecular, GT pelas viasintrínseca/extrínseca no plasma humano diluído 60 vezes, com inativação intrínseca total usando PSs brutosde A. muscoides em paralelo à heparina (p < 0,05). A trombose in vitro é modulada diferencialmente porPSs brutos distintos de algas marinhas brasileiras.
Assuntos
Alga Marinha/enzimologia , Alga Marinha/química , Trombina/análiseRESUMO
The biotechnological value of macroalgae for screening assays of thrombin generation-TG using sulfated polysaccharides-SPs as substitutes to heparin has been poorly explored. Five Brazilian species of macroalgae (Gracilaria birdiae, Acanthophora muscoides, Halymenia sp., Caulerpa cupressoides and C. racemosa) wereanalyzed and compared for their abundance, physical-chemical characteristics and in vitro anticoagulant assays of activated partial thromboplastin time-APTT, prothrombin time-PT and TG. Papain extraction yielded (p < 0.001) from 0.66±0.03% (C. racemosa) to 41.60±1.10% (Halymenia sp.) of crude SPs varying sulfate (8.41-42.60%) and total sugars (47.80-70.53%). Crude SPs showed difference in mobility and resolution pattern by agarose electrophoresis, while polyacrylamide analysis revealed SPs of > 100 kDa. These procedures,combined with the use of Stains-All, also indicated nonSPs. APTTs ranged from 2.81 (A. muscoides) to 21.30 IU(Halymenia sp.) vs. heparin (193 IU), and were dependent on sulfation of the crude SPs. PT was not altered. Withrespect to TG assay, crude SPs modified concentration-dependent and independently from molecular mass TGby both intrinsic/extrinsic pathways in 60-fold diluted human plasma, with total intrinsic inactivation using crudeSPs from A. muscoides in parallel to heparin (p < 0.05). Thrombosis in vitro is differentially modulated by distinctcrude SPs from Brazilian seaweeds.(AU)
O valor biotecnológico das macroalgas para ensaios de varredura de geração de trombina-GT pouco tem sido explorado usando polissacarídeos sulfatados-PSs como substitutos à heparina. Foramanalisadas e comparadas cinco espécies brasileiras de macroalgas (Gracilaria birdiae, Acanthophora muscoides, Halymenia sp., Caulerpa cupressoides e C. racemosa) quanto à abundância, às característicasfísico-químicas e os ensaios anticoagulantes in vitro de tempo de tromboplastina parcial ativada-TTPA, aotempo de protrombina-TP e a GT. A extração com papaína rendeu (p < 0,001) de 0,66±0,03%(C. racemosa) a 41,60±1,10% (Halymenia sp.) de PSs brutos variando sulfato (8,41-42,60%) e açúcarestotais (47,80-70,53%). Os PSs brutos mostraram diferenças na mobilidade e resolução por eletroforese emagarose, enquanto pela análise em poliacrilamida revelou PSs brutos de >100 kDa. Esses procedimentos,combinados ao uso de azul de toluidina/Stains-All, indicaram também polissacarídeos-não sulfatados. OsTTPAs foram dependentes da sulfatação dos PSs brutos e variaram de 2,81 (A. muscoides) a 21,30 UI (Halymenia sp.) vs. heparina (193 UI). O TP não foi alterado. Com respeito ao ensaio de GT, os PSs brutos modificaram, dependente de concentração e independentemente de massa molecular, GT pelas viasintrínseca/extrínseca no plasma humano diluído 60 vezes, com inativação intrínseca total usando PSs brutosde A. muscoides em paralelo à heparina (p < 0,05). A trombose in vitro é modulada diferencialmente porPSs brutos distintos de algas marinhas brasileiras.(AU)
Assuntos
Alga Marinha/química , Alga Marinha/enzimologia , Trombina/análiseRESUMO
AIM: The aim of this study was to assess the effects of the injectable depot-medroxyprogesterone acetate (DMPA) contraceptive on selected blood coagulation parameters in young, healthy new users. METHODS: The prospective study included 39 healthy women aged 20-39 years, with a body mass index (BMI; kg/m2 ) < 30, who were never users of DMPA, and who opted to use DMPA (21 women) or a copper intrauterine device (IUD; 18 women). The women in the two groups were matched for age (±1 year) and BMI (±1). Blood samples were obtained from all participants at baseline and at 6 and 12 months. Activated partial thromboplastin time, D-dimer, protein C, antithrombin, protein S, and thrombin generation test (lag time, endogenous thrombin potential, time to peak, and velocity index of thrombin generation) were analyzed. Repeated-measures analysis of variance was used to compare the groups. RESULTS: There were no significant differences between the groups at baseline with respect to any of the parameters evaluated; however, in the DMPA group, D-dimer levels were lower and the time to peak thrombin generation was longer than in the IUD group at 12 months of evaluation. CONCLUSION: Lower D-dimer and longer time to peak thrombin generation in new users of DMPA suggest a positive profile against hypercoagulability.
Assuntos
Coagulação Sanguínea/efeitos dos fármacos , Anticoncepcionais Femininos/efeitos adversos , Acetato de Medroxiprogesterona/efeitos adversos , Adulto , Anticoncepcionais Femininos/administração & dosagem , Preparações de Ação Retardada , Feminino , Humanos , Acetato de Medroxiprogesterona/administração & dosagem , Estudos Prospectivos , Tromboembolia Venosa/induzido quimicamente , Adulto JovemRESUMO
BACKGROUND: Liver failure patients have traditionally been empirically transfused prior to invasive procedures. Blood transfusion is associated with immunologic and nonimmunologic reactions, increased risk of adverse outcomes and high costs. Scientific evidence supporting empirical transfusion is lacking, and the best approach for blood transfusion prior to invasive procedures in cirrhotic patients has not been established so far. The aim of this study is to compare three transfusion strategies (routine coagulation test-guided - ordinary or restrictive, or thromboelastometry-guided) prior to central venous catheterization in critically ill patients with cirrhosis. METHODS/DESIGN: Design and setting: a double-blinded, parallel-group, single-center, randomized controlled clinical trial in a tertiary private hospital in São Paulo, Brazil. INCLUSION CRITERIA: adults (aged 18 years or older) admitted to the intensive care unit with cirrhosis and an indication for central venous line insertion. Patients will be randomly assigned to three groups for blood transfusion strategy prior to central venous catheterization: standard coagulation tests-based, thromboelastometry-based, or restrictive. The primary efficacy endpoint will be the proportion of patients transfused with any blood product prior to central venous catheterization. The primary safety endpoint will be the incidence of major bleeding. Secondary endpoints will be the proportion of transfusion of fresh frozen plasma, platelets and cryoprecipitate; infused volume of blood products; hemoglobin and hematocrit before and after the procedure; intensive care unit and hospital length of stay; 28-day and hospital mortality; incidence of minor bleeding; transfusion-related adverse reactions; and cost analysis. DISCUSSION: This study will evaluate three strategies to guide blood transfusion prior to central venous line placement in severely ill patients with cirrhosis. We hypothesized that thromboelastometry-based and/or restrictive protocols are safe and would significantly reduce transfusion of blood products in this population, leading to a reduction in costs and transfusion-related adverse reactions. In this manner, this trial will add evidence in favor of reducing empirical transfusion in severely ill patients with coagulopathy. TRIAL REGISTRATION: ClinicalTrials.gov, identifier: NCT02311985 . Retrospectively registered on 3 December 2014.