EXTEM Assays Research Articles

Whole blood thromboelastometry profiles in women with preeclampsia.

Preeclampsia (PE) is a pregnancy complication characterized by high blood pressure and significant amounts of protein in the urine. Various coagulation abnormalities have been described in pregnant women with PE. The aim of the present case-control study was to evaluate whole blood thromboelastometry profiles, performed by ROTEM(®), in women with PE in order to better characterize the PE-related discoagulopathy. Standard ROTEM(®) (Tem International GmbH, Munich, Germany) parameters evaluating clot initiation [clotting time (CT)], propagation [clot formation time (CFT); α-angle], stability [maximum clot firmness (MCF)] and lysis [maximum lysis (ML)] in INTEM, EXTEM, NATEM, and FIBTEM assays were performed in 30 consecutive pregnant women with PE at diagnosis. Sixty (1:2 ratio with cases) healthy pregnant women, matched for gestational age (± 2 weeks) with the cases, acted as controls. Platelet count, prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen, antithrombin and D-Dimer were also evaluated. Preeclamptic women showed a significantly more rapid propagation phase in EXTEM assay than controls (CFT 62 ± 15 vs. 75 ± 15 s and α-angle 78 ± 4 vs. 75 ± 4°, p<0.01 in both cases). Moreover, MCF was significantly higher and ML significantly lower in women with PE than in healthy pregnant women (p=0.001 for all comparisons). ROTEM(®) profiles in women with PE were characterized by an increased tissue factor driven clot propagation capability. In addition, higher clot stability due both to the increase in clot firmness and the decrease in blood fibrinolysis was observed. Larger studies are needed to identify the clinical relevance of ROTEM(®) alterations in women with PE.

Single vs double antiplatelet therapy in acute coronary syndrome: Predictors of bleeding after coronary artery bypass grafting.

To investigate the contribution of anti-platelet therapy and derangements of pre-operative classical coagulation and thromboelastometry parameters to major bleeding post-coronary artery bypass grafting (CABG). Two groups of CABG patients were studied: Group A, treated with aspirin alone (n = 50), and Group B treated with aspirin and clopidogrel (n = 50). Both had similar preoperative, clinical, biologic characteristics and operative management. Classic coagulation parameters and rotational thromboelastometry (ROTEM) profiles were determined preoperatively for both groups and the same heparin treatment was administered. ROTEM profiles (INTEM and EXTEM assays) were analyzed, both for traditional parameters, and thrombin generation potential, expressed by area-under-curve (AUC). There was no significant difference between rates of major bleeding between patients treated with aspirin alone, compared with those treated with aspirin and clopidogrel (12% vs 16%, P = 0.77). In the 14 cases of major bleeding, pre-operative classic coagulation and traditional ROTEM parameters were comparable. Conversely we observed that the AUC in the EXTEM test was significantly lower in bleeders (5030 ± 1115 Ohm*min) than non-bleeders (6568 ± 548 Ohm*min) (P < 0.0001). We observed that patients with a low AUC value were at a significantly higher risk of bleeding compared to patients with higher AUC, regardless of antiplatelet treatment. This suggests that thrombin generation potential, irrespective of the degree of platelet inhibition, correlates with surgical bleeding.

Aspects of bleeding complications and hemostasis at central line insertion and mild induced hypothermia

Bleeding complications range from 0.5 to 1.6% in connection with central venous catheter insertions but are more frequent in patients with bone marrow failure and severe thrombocytopenia. Although supportive evidence is scarce, prophylactic platelet transfusion is sometimes performed in these patients before catheter insertion. Furthermore, the ideal threshold platelet count and timing of transfusion remain controversial among clinical studies. Hypothermia is generally considered to reduce coagulation and platelet function. However, studies performed in animals, healthy volunteers, and patients have shown conflicting results. Paper I, a retrospective study in non-intensive care unit patients, showed that serious bleeding complications in association with central line insertions were uncommon and that insertion of a large bore catheter may be an independent risk factor for mild bleeding complications in this population. Paper II, a prospective observational study, evaluated the efficacy of prophylactic platelet transfusions in thrombocytopenic patients with bone marrow failure. Transfusion improved hemostatic parameters in ROTEM and Multiplate tests by increasing the number of platelets and not by enhancing platelet function. Improved clotting parameters persisted between 1 and 4 hours after transfusion. Paper III demonstrated increased platelet aggregation and strengthened clot formation over time in out-of-hospital cardiac arrest patients treated with hypothermia. In patients with dual platelet inhibition, including ticagrelor and aspirin, this effect was offset by powerful P2Y12 blockade, confirmed by analysis of vasodilator-stimulated phosphoprotein. The effect of ticagrelor was delayed in survivors of cardiac arrest, probably due to slow gastric emptying. Paper IV demonstrated prolonged clot initiation and decreased clot propagation in ROTEM testing (EXTEM, FIBTEM, and APTEM assays) in whole blood from out-of-hospital cardiac arrest patients treated with mild induced hypothermia. Paper V investigated whole blood from acute coronary syndrome patients treated with ticagrelor and aspirin. In vitro applied hypothermia to 33 degrees C markedly increased platelet activity measured by flow cytometry, whereas a viscoelastic coagulation test (Sonoclot) revealed a hypocoagulative response.

Laboratory Assessment of Anti-Coagulant Properties of a Von Willebrand Factor Targeted Aptamer

Introduction: Many cardiovascular procedures including percutaneous coronary intervention, coronary bypass surgery, and peripheral vascular surgery require antithrombotic therapy during the procedure. Heparin is frequently used, due to low cost, ease of monitoring and reversibility with protamine. Antiplatelet agents such as aspirin and clopidogrel are also frequently used but these agents are not reversible. Alternative reversible antithrombotic strategies that interfere with platelet-mediated thrombosis are needed. We are investigating the use of a reversible agent that is specific for von Willebrand factor (VWF) for use during cardiovascular procedures.We previously developed an antidote-controlled 80 oligonucleotide RNA aptamer Ch-9.14-T10 (T10). (Nimjee, S.M., Lohrmann, Wang, H, et al. Molecular Therapy 2012; 2:391-397). T10 bound to VWF with high affinity and inhibited thrombosis in a murine carotid artery damage model. The aptamer also greatly increased bleeding in mice that were surgically challenged by tail transection and blood loss decreased when complementary antidote oligonucleotides were administered. In addition, T10 inhibited primary hemostasis in the PFA-100 assay and this inhibition was completely reversed by the antidote. This study further characterizes the T10 aptamer, as well as a truncated version of the T10 aptamer, using clinical laboratory assays that would be used to monitor antithrombotic therapies during cardiovascular procedures.Methods: We initiated our study with T10 and then developed a truncated version, T25, a 60 oligonucleotide with which we performed additional investigations. We recruited normal donors for analysis of aptamer and antidotes. All participants had normal platelet counts, VWF activity and antigen levels, and were not taking antiplatelet agents. Laboratory tests included: ristocetin induced platelet agglutination (RIPA), VWF activity, ristocetin cofactor activity, cone and plate(let) analyzer, thromboelastography (TEG) and thromboelastometry (ROTEM).Results: We first determined the effects of the VWF aptamers on RIPA, VWF activity, and ristocetin cofactor activity to confirm functional inhibition of VWF. We found that T10 (500-750 nM) and T25 (250-750nM) completely inhibited RIPA depending on the donor. T25 also inhibited VWF activity with a Ki of 160-320 nM and ristocetin cofactor activity with an IC50 of 3000 nM. Employing a cone and plate(let) analyzer we found that T25 (750 nm) reduced platelet adhesion by 33-49% on a polystyrene surface under arterial flow conditions (1800 s-1, for 2 minutes). The aggregate size was also reduced by 30%.As ACT and TEG are frequently used during cardiovascular procedures, we performed several measures with our VWF aptamer. T10 had no effect on the ACT (730-2000nM), TEG (1600 nM), or ROTEM (750 nM). Standard methods were used for the TEG (Kaolin activation followed by CaCl2) and ROTEM (EXTEM assay) Topf et al. reported a modified ROTEM protocol where they found that incubating citrated whole blood with ristocetin decreased the clot strength as measured by the maximum amplitude (Thromb.Haemost 2011; 105:1091). We reasoned that an aptamer that bound to VWF would inhibit the effect of ristocetin on the maximum amplitude. We found that T25 did inhibit the effect of ristocetin and titration studies showed that 200-400 nM T25 was needed to restore the maximum amplitude to at least 80% of its original value. An example of the ROTEM tracings is shown in figure 1.Conclusion: The VWF-specific aptamer T25 can be detected by several clinical assays, including the RIPA, VWF activity assays, and the cone and plate(let) analyzer. It is not detectable by an ACT or standard TEG or ROTEM, however. We are able to detect the aptamer using a modified ROTEM and are further defining the adaptability of this assay to facilitate monitoring during cardiovascular procedures.Citrated whole blood from was incubated with buffer or ristocetin (0.7 mg/ml) or ristocetin and T25 (300 nM) as indicated. Clotting was initiated with EXTEM (tissue factor pathway activation) and CaCl2. [Display omitted] DisclosuresBecker:Regado Biosciences: Membership on an entity’s Board of Directors or advisory committees.

The Evaluation of Hydroxyethyl Starch (6% HES 130/0.4) Solution's Potential Preventive Effects on Coagulation Status in Women with Gynecologic Malignancies Using Rotation Thromboelastography.

Objective: The aim of this study was to determine the effects of in vitro hemodilution with 6% hydroxyethyl starch (HES) 130/0.4 solution on the coagulation status of women with gynecologic malignancies by using rotation thromboelastogram (ROTEM®). Materials and Methods: Twenty-two patients with gynecological tumors scheduled for anesthesia were enrolled. Blood samples were diluted by 20% with 6% HES (130/0.4) solution.Results: In the INTEM assay, clotting time (CT) (p<0.01) and clot formation time (CFT) (p<0.001) were significantly increased and maximum maximum clot formation (MCF) (p< 0.001) was significantly decreased in HES hemodilution compared with the undiluted control samples. In the EXTEM assay, there was a similar significant increase in increase in CFT (p<0.01) and a decrease in maximum a decrease in MCF (p<0.01) in HES hemodilution when compared with control samples. Conclusion: HES 130/0.4 solution causes significant hypocoagulable changes in the thromboelastographic profile of gynecologic cancer patients in vitro.

Assessment of early thromboelastometric variables from extrinsically activated assays with and without aprotinin for rapid detection of fibrinolysis.

Although thromboelastometry (ROTEM®) and thrombelastography can be used for bedside diagnosis of fibrinolysis, the time needed for detection is often prolonged. Since untreated fibrinolysis can result in consumption of coagulation factors and bleeding, early diagnosis and decision making are desirable. Accordingly, we assessed ROTEM variables from extrinsically activated assays with (APTEM) and without (EXTEM) addition of aprotinin for their ability to rapidly identify fibrinolysis. Specifically, we tested the hypotheses that prolonged clotting time, clot formation time, low clot firmness (at 5, 10, 15, and 20 minutes, designated A5, A10, A15, and A20, respectively), low maximum clot firmness (MCF) in EXTEM assays, and differences in these variables from parallel APTEM and EXTEM assays (designated as Δvariables) predict fibrinolysis. Data from 411 thromboelastometric measurements (obtained from 352 patients) with fibrinolysis and from 2537 measurements without fibrinolysis (obtained from 1605 patients) were assessed and analyzed using receiver operating characteristics. Data were analyzed as a pooled fibrinolysis cohort, and subanalyses were performed from sets assigned to categories of fibrinolysis related to the timing of thrombus lysis (i.e., a decrease of clot firmness to <15% of MCF within 30, 45, and 60 minutes, respectively). A lower 95% confidence limit of the area under the receiver operating characteristic curve (AUC [SE] <0.6) was considered a failure to substantially improve detection of increased fibrinolysis. AUCs were compared to identify the variable providing the best predictive association with fibrinolysis. As a secondary end point, optimum cutoff values at the point estimate corresponding to the greatest Youden index were calculated along with the respective sensitivities and specificities. In the pooled cohort, clot formation time (AUC: 0.652 [0.016]), α-angle (AUC: 0.675 [0.015]), A5 (AUC: 0.718 [0.013]), A10 (AUC: 0.734 [0.0.13]), A15 (AUC: 0.752 [0.013]), A20 (AUC: 0.771 [0.013]), and MCF (AUC: 0.799 [0.012]) predicted fibrinolysis. Fibrinolysis was also predicted by ΔA15 (AUC: 0.675 [0.016]), ΔA20 (AUC: 0.719 [0.015]), and ΔMCF (AUC: 0.812 [0.013]). AUCs increased in a time-related fashion. The ability to predict subsequent fibrinolysis based on thromboelastometry was higher when it occurred early rather than later during testing. However, for prediction of late fibrinolysis, only MCF (AUC: 0.655 [0.025]) appears to be potentially clinically useful. Low early values of clot firmness in extrinsically activated thromboelastometric assays are associated with fibrinolysis and improve its early detection. Additional assays with aprotinin fail to improve the early diagnosis of fibrinolysis compared with assays without aprotinin.

Wide temperature range testing with ROTEM coagulation analyses.

Mild induced hypothermia is used for neuroprotection in patients successfully resuscitated after cardiac arrest. Temperature-dependent effects on rotational thromboelastometry (ROTEM(®)) assays with EXTEM(®), FIBTEM(®), or APTEM(®) in cardiac arrest patients have not previously been studied. Ten patients with out-of-hospital cardiac arrest who underwent induced hypothermia were studied during stable hypothermia at 33°C. ROTEM temperature effects on EXTEM, FIBTEM, and APTEM assays were studied at temperatures set between 30°C and 42°C. Citrated whole blood test tubes were incubated in temperature-adjusted heating blocks and then investigated at respective temperature in the temperature-adjusted ROTEM. The following variables were determined: clotting time (CT), clot formation time (CFT), α-angle, and maximum clot firmness (MCF). The results from hypo- and hyperthermia samples were compared with the samples incubated at 37°C using the Wilcoxon matched-pairs signed-rank test. A p-value of <0.05 was considered significant. CT-EXTEM(®) and CT-APTEM(®) were prolonged by hypothermia at 30°C (p<0.01 for both) and 33°C (p<0.05 for both). Hyperthermia at 42°C shortened CT-EXTEM (p<0.05) and CT-APTEM (p<0.01). CFT-EXTEM(®) and CFT-APTEM(®) were markedly prolonged by hypothermia at 30°C, 33°C, and 35°C (p<0.01 for all except CFT-EXTEM, 35°C [p<0.05]). The α-angle-EXTEM was markedly decreased at 30°C, 33°C, and 35°C (p<0.01) but increased at 40°C (p<0.05) and 42°C (p<0.01); α-angle-APTEM showed similar results. MCF was unchanged at different temperatures for all tests. ROTEM (EXTEM, FIBTEM, and APTEM assays) revealed a hypocoagulative response to in vitro-applied hypothermia in the blood of cardiac arrest patients reflected in the prolonged clot initiation and decreased clot propagation. Hyperthermia showed the opposite effects. Clot firmness was not affected by temperature.

Ex Vivo Prothrombin Complex Concentrates and a Specific Antidote Are Effective In Reversing Dabigatran-Induced Coagulopathy In Pigs

IntroductionNew oral anticoagulants are effective alternatives to warfarin; however, no specific reversal agents are currently available for life-threatening bleeding or emergency surgery. This study measured the effects of prothrombin complex concentrate (PCC), activated PCC (aPCC) and recombinant factor VIIa (rFVIIa) and compared this to reversal with the specific antibody fragment for dabigatran after high-dose dabigatran etexilate (DE) treatment in a porcine model of trauma, to assess reversal of anticoagulation. MethodsStudies were performed in 5 male pigs after ethical approval. DE was given orally for 3 days (30 mg/kg bid) and, on the 4th day, dabigatran was infused at 0.77mg/kg/h for 30 min followed by 0.52 mg/kg/min for 60 min. The infusion was administered immediately before trauma (blunt liver insult). Blood samples were taken before dabigatran infusion and 60 min post-injury. Two doses of PCC (30 and 60 IU/kg), aPCC (30 and 60 IU/kg) and rVIIa (90 and 180 μg/kg) and two doses anti-Dabi Fab (30 and 60 mg/kg) were added to blood samples ex vivo. Coagulation was assessed by thromboelastometry (ExTEM assay) and PT and dabigatran levels with diluted TT. Data were analyzed by ANOVA (± SEM). ResultsOral DE prolonged clot formation (CT: 389 ± 128 sec; CFT: 159 ± 39 sec) and PT (27 ± 9 sec), but coagulopathy was more severe after achieving high dabigatran plasma levels (943 ± 147 ng/ml) and inducing trauma (CT: 2378 ± 480 sec; CFT: 3374 ± 625 sec; PT: 167 ± 40 sec). At this time point, 30 IU/kg PCC and 30 IU/kg aPCC significantly accelerated clot formation (PCC: CT 416 ± 70 sec, CFT 250 ± 91 sec; aPCC: CT 525 ± 126 sec, CFT 309 ± 60 sec; p<0.05 vs. untreated samples). These interventions also shortened PT (PCC: 53 ± 14 sec; aPCC: 66 ± 16 sec; p<0.05 vs. untreated samples). Higher doses of PCC and aPCC had no further effect on hemostatic parameters. rFVIIa had no impact on coagulopathy at any time point. Addition of the specific antidote to dabigatran completely normalized all coagulation parameters to baseline levels. ConclusionThe ex vivo addition of either PCC or aPCC is effective in reducing coagulopathy in a porcine model of high-dose dabigatran therapy and trauma. However the specific aDabi-Fab is the most promising agent to reverse the anticoagulant effects of dabigatran. The potential for reducing blood loss remains to be investigated. Disclosures:van Ryn:Boehringer Ingelheim Pharma GmbH & Co. KG: Employment. Spronk:Boehringer Ingelheim Pharma GmbH & Co. KG: Research Funding. Rossaint:Boehringer Ingelheim: Research Funding. Grottke:Boehringer Ingelheim: Research Funding; CSL Behring: Consultancy.

A calcium-containing electrolyte-balanced hydroxyethyl starch (HES) solution is associated with higher factor VIII activity than is a non-balanced HES solution, but does not affect von Willebrand factor function or thromboelastometric measurements--results of a model of in vitro haemodilution.

Hydroxyethyl starch (HES) is known to impair blood coagulation. The impact of calcium-containing, balanced carrier solutions of HES on coagulation is controversial. We investigated the effects of increasing degrees of haemodilution with modern 6%, electrolyte-balanced HES vs non-balanced HES on coagulation in vitro, and compared the balanced HES to a balanced crystalloid solution for an internal control. Blood samples from ten healthy volunteers were diluted in vitro by 20%, 40% and 60% with either calcium-containing balanced 130/0.42 HES, non-balanced 130/0.4 HES or balanced crystalloid. In all samples, blood counts, prothrombin time ratio, activated partial thromboplastin time, ionized calcium, factor VIII activity, von Willebrand factor antigen, von Willebrand factor collagen binding activity, and von Willebrand factor activity were determined, and activated rotational thromboelastometry (EXTEM and FIBTEM assays) was performed. Haemodilution impaired coagulation in a dilution-dependent manner as determined by both conventional laboratory assays and thromboelastometry. Ionized calcium increased with balanced HES (p≤0.004), but decreased with non-balanced HES (p≤0.004). Prothrombin time ratio (p≤0.002) and factor VIII levels (p=0.001) were better preserved with balanced HES than with non-balanced HES in dilutions ≥40%. Thromboelastometry showed no differences between values in blood diluted with the balanced or non-balanced HES. In vitro, a balanced calcium-containing carrier solution of 6% HES 130/0.42 preserved coagulation better than did non-balanced HES 130/0.4 as quantified by conventional coagulation assays, but not in activated thromboelastometry. One explanation could be the increased ionized calcium levels after dilution with calcium-containing carrier solutions.

The role of hemostatic mechanisms in the development of thrombosis in Behcet's disease: an analysis by modified rotation thromboelastogram (ROTEM)

Behcet's disease (BD) is a multisystemic disorder characterised by recurrent oral and genital ulcers. Vasculitis and thrombotic events are the most important causes of mortality. Thrombosis is the major clinical finding in patients with BD, but the cause of the thrombosis is still unclear. Thromboelastography is an alternative method to evaluate almost all steps of the hemostatic system. Today, the modified rotation thromboelastogram (ROTEM) is a newer coagulation test and a more powerful technique. Our aim in this study was to investigate whether hemostatic mechanisms play a role in the development of thrombosis in BD patients by using ROTEM. Thirty BD patients, 20 ankylosing spondylitis patients, and 14 healthy controls who are not taking anti-aggregant or anti-coagulant therapy were included in the study. Whole blood count, protrombin time, activated protrombin time, fibrinogen, D-dimer levels, and ROTEM parameters (clotting time, clot formation time (CFT), and maximum clot formation(MCF)) were determined by INTEM and EXTEM analysis. Of the 30 patients with BD, 19 were women and 11 were men, and mean age was 40.6 ± 11.2. Two of the BD patients had vascular involvement, but none of them were in active phase of the disease during the study. In INTEM assay, MCF (p < 0,001) value was significantly increased, and CFT (p>0.05) value was decreased in BD patients compared with the control group. In the EXTEM assay, there was a similar significant increase in MCF (p=0.002) value and a decrease in CFT (p=0.002) value in BD patients compared with the control group. The results of our study indicated that primary hemostatic mechanisms which can be detected by ROTEM may play a role in the development of thrombosis in patients with BD.

Changes in fibrinogen availability and utilization in an animal model of traumatic coagulopathy

BackgroundImpaired haemostasis following shock and tissue trauma is frequently detected in the trauma setting. These changes occur early, and are associated with increased mortality. The mechanism behind trauma-induced coagulopathy (TIC) is not clear. Several studies highlight the crucial role of fibrinogen in posttraumatic haemorrhage. This study explores the coagulation changes in a swine model of early TIC, with emphasis on fibrinogen levels and utilization of fibrinogen.MethodsA total of 18 landrace pigs were anaesthetized and divided into four groups. The Trauma-Shock group (TS) were inflicted bilateral blast femoral fractures with concomitant soft tissue injury by a high-energy rifle shot to both hind legs, followed by controlled exsanguination. The Shock group (S) was exposed to shock by exsanguination, whereas a third group was exposed to trauma only (T). A fourth group (C) served as control. Physiological data, haematological measurements, blood gas analyses and conventional coagulation assays were recorded at baseline and repeatedly over 60 minutes. Thrombelastometry were performed by means of the tissue factor activated ExTEM assay and the platelet inhibiting FibTEM assay. Data were statistically analysed by repeated measurements analyses method.ResultsA significant reduction of fibrinogen concentration was observed in both the TS and S groups. INR increased significantly in the S group and differed significantly from the TS group. Maximum clot firmness (MCF) of the ExTEM assay was significantly reduced over time in both TS and S groups. In the FibTEM assay a significant shortening of the clotting time and an increase in MCF was observed in the TS group compared to the S group.ConclusionDespite a reduction in clotting capability measured by ExTEM MCF and a reduced fibrinogen concentration, extensive tissue trauma may induce an increased fibrin based clotting activity that attenuates the hypocoagulable tendency in exsanguinated animals.

Whole blood rotation thromboelastometry (ROTEM®) profiles in subjects with non-neoplastic portal vein thrombosis

The coagulation pattern and the determinants of portal vein thrombosis (PVT), both in patients with and without cirrhosis, are still largely unknown. The aim of this study was to evaluate whole blood thromboelastometry profile, performed by ROTEM®, of both cirrhotic and non-cirrhotic subjects with PVT. Two different groups were considered: i) 14 non-cirrhotic PVT patients, ii) 35 cirrhotic patients with PVT. Controls were sex- and age-matched healthy volunteers and cirrhotic subjects without PVT, respectively. ROTEM® assays (i.e. INTEM, EXTEM, NATEM, and FIBTEM) and traditional coagulative parameters (i.e. platelet count, PT/INR, aPTT, and fibrinogen) were performed on blood samples from each subject. There were no significant differences in ROTEM® profile, as for INTEM, EXTEM, and NATEM assays, and in traditional coagulative parameters, between PVT patients, both with and without cirrhosis, and control groups. Interestingly, Maximum Clot Firmness (MCF) in FIBTEM was significantly higher in non-cirrhotic PVT patients (19 mm) than in healthy volunteers (11 mm, p<0.05). The amplitude of MCF in FIBTEM revealed to be a useful tool to discriminate non-cirrhotic subjects with PVT from those without thrombotic events. Larger prospective studies are needed to evaluate the relevance of the association between the alterations of ROTEM® profiles and PVT in cirrhotic patients.

FRI0238 Can thromboloelastography be a new tool to asses the risk of thrombosis in behcet’s disease?

BackgroundBehcet’s Disease (BD) is a multisystemic disorder characterised by recurrent oral and genital ulcers (1). Vasculitis and thrombotic events are the most important causes of mortality (2). Thrombosis is the...

Thrombelastographic evaluation of the influence of 2-RBC apheresis on donor’s coagulation system

Rotation thrombelastogram (ROTEM®/TEG®) assays allow rapid global assessment of hemostatic function using whole blood. Since published data about the effects of automated red cell collection on coagulation system are scarce, we aimed to investigate the effects of 2-RBC apheresis on donor’s coagulation system using ROTEM® assays.In INTEM assay, CFT was significantly shortened 24h after apheresis compared with baseline value (p<0.05) and MCF was significantly prolonged immediately after apheresis and 24h after apheresis compared with baseline value (p<0.05 and p<0.01, respectively). In EXTEM assay, CFT was significantly prolonged immediately after apheresis and 24h after apheresis compared with baseline value (p=0.001 and p<0.001, respectively) and MCF was significantly prolonged 24h after apheresis compared with baseline value (p<0,001).Our results demonstrate thromboelastographic signs of hypercoagulability in donors undergoing 2-RBC apheresis.

Rotational thromboelastometry for diagnosing sudden hyperfibrinolysis immediately after cardiopulmonary bypass during cardiac surgery

Rotational thromboelastometry for diagnosing sudden hyperfibrinolysis immediately after cardiopulmonary bypass during cardiac surgery

In-vitro comparison of free oscillation rheometry (ReoRox) and rotational thromboelastometry (ROTEM) in trauma patients upon hospital admission

Plasma-based assays do not provide accurate information on haemostatic resuscitation hence viscoelastic point-of-care haemostatic assays such as rotational thromboelastometry (ROTEM Delta, Pentapharm) are used to monitor coagulopathy in trauma patients. Free oscillation rheometry (FOR) is a new whole blood haemostatic assay that measures not only the clot-forming process but also the initial viscous phase; this could potentially be of value when assessing traumatic coagulopathy. A comparative analysis between FOR and ROTEM was therefore performed. This is a prospective observational study of 40 adult trauma patients admitted to a level 1 trauma centre. Citrated whole blood was analysed with ROTEM EXTEM and FIBTEM assays and FOR Fibscreen1 and Fibscreen2 assays. Predefined variables of ROTEM and FOR were compared using Spearman's ρ. ROTEM maximum clot function (MCF) in both EXTEM and FIBTEM correlated (P < 0.0001 for both) with FOR maximum elasticity Fibscreen1 and Fibscreen2, respectively. Interestingly, ROTEM EXTEM clotting time did not correlate with any of the FOR clot initiation parameters COT1, COT2 or COT2-1 of Fibscreen1. A correlation between ROTEM EXTEM and FIBTEM and FOR Fibscreen1 and Fibscreen2 clot formation and clot strength was found as was a significant correlation between lysis index after 60 min and ClotSR30. ROTEM EXTEM did not correlate with COT1, COT2 or COT2-1 of Fibscreen1 and this warrants further investigation.

Alterations in rotation thromboelastometry (ROTEM®) parameters: point-of-care testing vs analysis after pneumatic tube system transport

Thromboelastometry as point-of-care (POC) testing enables the analysis of the clotting process at the bedside, providing rapid results to guide haemostatic therapy. However, POC testing utilizes medical staff who are managing critically ill patients, as non-laboratory personnel may not be sufficiently trained to run the devices. To resolve these problems, thromboelastometry can be performed in the central laboratory and rapid transport of samples can be accomplished via a pneumatic tube system (PTS). This study compares thromboelastometry parameters of blood samples analysed immediately with those analysed after PTS transport. In patients with normal haemostasis, two arterial blood samples were collected from each patient (n=92) in citrated plastic tubes to investigate the assays INTEM (n=35), EXTEM (n=27), and FIBTEM (n=30). One blood sample was analysed immediately, the other sample after PTS transport. Thromboelastometry was performed using a single ROTEM(®) device. The mean clot firmness values were significantly lower for PTS samples in both the INTEM (-0.7 mm cf. -1.1 mm) and EXTEM (-1.4 cf. -1.7 mm) assays. INTEM coagulation time (CT) was significantly lower in PTS samples with a mean difference of -13 s. EXTEM CT was significantly higher in PTS samples with a mean difference of +3.9 s. Thromboelastometry parameters of blood samples analysed after PTS transport are significantly altered compared with those analysed immediately. However, in patients with normal haemostasis, the alterations were small and without clinical consequence, implying that analysis after PTS transport is an acceptable alternative to prompt analysis at the bedside. Further studies should focus on patients with impaired haemostasis.

Evaluating coagulation disorders in the use of bevacizumab for metastatic colorectal cancer by thrombelastography

The aim of this study was to evaluate coagulation disorders in patients with metastatic colorectal cancer treated with bevacizumab by using rotation thrombelastogram (ROTEM(®)) and correlate ROTEM(®) parameters with routine coagulation tests. A total of 18 colorectal cancer patients who received bevacizumab combined with chemotherapy were included. There was no statistically significant difference between results of platelet count, prothrombin time (PT) and activated partial thromboplastin time (APTT), fibrinogen, and D-Dimer obtained at baseline and on day 1 of chemotherapy cycles 4, 8, and 12. CFT value was significantly increased on day 1 of cycle 12 compared with baseline value by both INTEM and EXTEM assays while CT and MCF showed no significant difference. Correlation analysis revealed significant correlation between laboratory parameters and ROTEM(®) parameters. Platelet count showed a positive correlation with MCF in INTEM (r = 0.627) and EXTEM (r = 0.699) assays while showed a negative correlation with CFT in EXTEM (r = -603). There was a significant negative correlation between fibrinogen levels and CFT in INTEM (r = -0.617) and EXTEM (r = -0.512). Our data demonstrated the value of TEG over conventional coagulation tests in evaluating antiangiogenesis agents-induced coagulation disorders.

The effects of pneumatic tube system transport on ROTEM analysis and contact activation assessed by thrombin generation test

Thromboelastometry (ROTEM) is a popular point-of-care test. It generates results quickly and may benefit individualised guided haemostatic therapy. However, processing of specimens by non-technicians might decrease the quality and reproducibility of results. Centralised laboratory equipment receiving specimens through a pneumatic tube system (PTS) could avoid this. This study aimed to evaluate the influence of PTS transport on ROTEM results and its contribution to contact activation assessed by thrombin generation (TG). MethodsSpecimens from 44 patients were drawn immediately after arterial puncture. Two were anticoagulated by citrate and two by citrate/corn trypsin inhibitor, a Factor XIIa pathway inhibitor. Both types of samples were transported by walking and PTS. Subsequently, analysis was performed: ROTEM on citrated blood, and TG on citrated and corn trypsin inhibitor (CTI) blood using either 0 or 1 pM tissue factor (TF). ResultsIn ROTEM analysis the NATEM assay showed significant differences. The EXTEM assay revealed small significant differences for clot formation time: 65seconds (SD±20) versus 67seconds (SD±17), and alpha angle 79° (SD±3) versus 77° (SD±3). The results remained within reference range. TG was not significantly affected by the type of tube transport, independent of the amount of TF. ConclusionPTS for ROTEM analysis is feasible except for NATEM assays. The amount of contact activation via Factor XIIa in terms of TG is independent of transport type. However, due to the different characteristics of pneumatic systems, hospitals should check its impact on the results before introducing this route of transport.

Similarities in Thromboelastometric (ROTEM®) Findings between Humans and Baboons

IntroductionInterest in visco-elastic testing in different clinical scenarios has increased but few data are available on thromboelastometric findings in primates. Materials and MethodsBlood cell count (hemoglobin, hematocrit, platelet count), coagulation parameters (prothrombin time, International Normalized Ratio, fibrinogen), and ROTEM® (Tem International GmbH, Munich, Germany) variables were analyzed using blood from 25 anesthetized male baboons and 21 non-anesthetized healthy volunteers. The platelet component of the clot was calculated as the difference in maximum clot elasticity (MCE) between the whole blood clot (EXTEM test) and the fibrin-based clot (FIBTEM test). In subgroups of each species, 10μg abciximab was added to the regular FIBTEM reagent (cytochalasin D) for additional platelet inhibition. ResultsBlood cell count was comparable between humans and primates. Both fibrinogen concentration (p<0.0001) and maximum clot firmness (MCF) in FIBTEM assays were significantly lower in baboons (p>0.0001, and p=0.006, respectively). PT, INR, and clotting time in NATEM assays were significantly prolonged in humans compared with baboons. MCF in NATEM, EXTEM and INTEM assays was not different between baboons and humans. Clot lysis in NATEM, EXTEM and INTEM assays was significantly higher in humans (p<0.0001). In contrast FIBTEM clot lysis was significantly higher in baboons (p=0.01). Addition of abciximab into the FIBTEM assay resulted in a significant reduction in MCF and MCE (p<0.001) and, consequently, the platelet component increased similar in both humans and baboons (p<0.001). ConclusionActivated ROTEM® tests revealed broad similarities between humans and baboons. ROTEM® assays developed for use in humans can also be used in baboons.