Oliver Grottke

Increasing concentrations of prothrombin complex concentrate induce disseminated intravascular coagulation in a pig model of coagulopathy with blunt liver injury

Despite increasing use of prothrombin complex concentrate (PCC) to treat hemorrhage-associated coagulopathy, few studies have investigated PCC in trauma, and there is a particular lack of safety data. This study was performed to evaluate PCC therapy in a porcine model of coagulopathy with blunt liver injury. Coagulopathy was induced in 27 anesthetized pigs by replacing approximately 70% blood volume with hydroxyethyl starch 130/0.4 and Ringers lactate solution; erythrocytes were collected and retransfused. Ten minutes after trauma, animals randomly received PCC (35 or 50 IU/kg) or saline. Coagulation parameters including thromboelastometry, thrombin generation, and blood loss were monitored for 2 hours. Internal organs were examined macroscopically and histologically to determine the presence of emboli and assess liver injury. Total blood loss was significantly lower and survival was higher in both PCC groups versus the control group (P < .05). These outcomes appeared to be dose-independent. Thromboembolism was found in all animals treated with 50 IU/kg PCC; 44% also showed signs of disseminated intravascular coagulation. Liver injury was similar in all animals. In conclusion, 35 IU/kg PCC safely improved coagulation and attenuated blood loss. However, the higher dose of PCC (50 IU/kg) appeared to increase the risk of thromboembolism and disseminated intravascular coagulation.

Argon: neuroprotection in in vitro models of cerebral ischemia and traumatic brain injury.

IntroductionRecently, it has been shown in several experimental settings that the noble gases xenon and helium have neuroprotective properties. In this study we tested the hypothesis that the noble gas argon has a neuroprotective potential as well. Since traumatic brain injury and stroke are widespread and generate an enormous economic and social burden, we investigated the possible neuroprotective effect in in vitro models of traumatic brain injury and cerebral ischemia.MethodsOrganotypic hippocampal slice cultures from mice pups were subjected to either oxygen-glucose deprivation or to a focal mechanical trauma and subsequently treated with three different concentrations (25, 50 and 74%) of argon immediately after trauma or with a two-or-three-hour delay. After 72 hours of incubation tissue injury assessment was performed using propidium iodide, a staining agent that becomes fluorescent when it diffuses into damaged cells via disintegrated cell membranes.ResultsWe could show argons neuroprotective effects at different concentrations when applied directly after oxygen-glucose deprivation or trauma. Even three hours after application, argon was still neuroprotective.ConclusionsArgon showed a neuroprotective effect in both in vitro models of oxygen-glucose deprivation and traumatic brain injury. Our promising results justify further in vivo animal research.

Reversal of dabigatran anticoagulation ex vivo: Porcine study comparing prothrombin complex concentrates and idarucizumab

Urgent surgery or life-threatening bleeding requires prompt reversal of the anticoagulant effects of dabigatran. This study assessed the ability of three- and four-factor prothrombin complex concentrate (PCC) and idarucizumab (specific antidote for dabigatran) to reverse the anticoagulant effects of dabigatran in a porcine model of trauma. Twelve animals were given dabigatran etexilate (DE) orally and dabigatran intravenously, before infliction of trauma. Six animals received tranexamic acid plus fibrinogen concentrate 12 minutes post-injury. Six PCCs (each 30 and 60 U/kg) and idarucizumab (30 and 60 mg/kg) were added to blood samples ex vivo. Coagulation was assessed by several coagulation assays. All coagulation parameters were altered after dabigatran infusion (plasma level: 442 ± 138 ng/ml). Both three- and four-factor PCCs mostly or completely reversed the effects of dabigatran on thromboelastometry variables and PT but not on aPTT. Idarucizumab neutralised plasma concentrations of dabigatran, and reversed the effects of the drug on coagulation variables. Thrombin generation showed dose-dependent over-correction following the addition of PCC, implying that elevated levels of thrombin are required to overcome dabigatran-induced coagulopathy. In contrast, treatment with idarucizumab returned thrombin generation to baseline levels. Following trauma, therapy with tranexamic acid plus fibrinogen improved correction of coagulation parameters by PCC, and thromboelastometry parameters by idarucizumab. All investigated PCCs improved dabigatran- and trauma-induced coagulopathy to a similar degree. In conclusion, this study shows that three- and four-factor PCCs are similarly effective for dabigatran reversal. Idarucizumab also reversed the effects of dabigatran and, unlike PCCs, was not associated with over-correction of thrombin generation.

Prevention and treatment of coagulopathy in patients receiving massive transfusions

Patients undergoing massive transfusions frequently develop a coagulopathy, which is already present in a considerable percentage of patients upon admission to the emergency room. This derangement of coagulation may aggravate the bleeding tendency and is associated with significant morbidity and mortality. Existing guidelines for optimal transfusion therapy in massively bleeding patients advocate early administration of crystalloid or colloid fluids in conjunction with transfusion of red cells. And, according to the guidelines, fresh frozen plasma (FFP) and platelets should only be administered when a whole blood volume or more has been replaced and then only in patients with excessive or microvascular bleeding and, at best, according to conventional laboratory coagulation analysis. However, this approach may cause dilution coagulopathy and a further impairment of hemostasis due to direct effects of plasma replacement treatment on platelet-vessel wall interaction and thus compromise haemostatic ability further in severely bleeding patients. In recent years, there has been increasing evidence, although mainly coming from non-randomized studies, that early and more intense replacement of coagulation factors and platelets may improve the outcome in patients undergoing massive transfusion. The recommendations in the existing guidelines are based on the results of conventional coagulation assays such as the activated partial thromboplastin time. However, these assays poorly correlate with clinically relevant coagulopathies [1, 2]. Cell-based whole blood viscoelastical assays such as thromboelastography (TEG) provide quantitative information of the haemostatic process and thus give a profile of the haemostatic changes that occur during clotting. Such tests may provide a better guide for blood component therapy for patients with massive bleeding, although also for these tests the clinical relevance has never been adequately validated [3–5]. It seemed of interest to obtain information on these issues by sending the following questions to experts in the field. Question 1: What is your definition of ‘massive blood transfusion’? Question 2: When treating a patient with massive bleeding, do you still follow the official guidelines i.e. restoration of blood volume initially with cristalloids or colloids followed by packed red cells and subsequently the use of FFP, platelets, cryoprecipitate, and other coagulation concentrates depending on the results of coagulation tests and platelet counts? If no: please explain. Question 3: Or do you follow a more aggressive regimen administering FFP and platelets as part of the standard transfusion program? If so, which FFP:RBC ratio do you apply? Please describe your transfusion policy in detail. Question 4: If you use coagulation parameters in your setting, which tests do you apply? Do you think that the conventional tests are satisfactory for this purpose? If not, please explain why. What would be an acceptable turn-around time for any test? Question 5: Have you evidence that a more aggressive regimen with regard to FFP and platelet transfusions improves outcome? Or do you think FFP and platelet transfusion may be harmful? We received 12 contributions to this Forum. Many of the answers are extensive and contain much detailed information. It is impossible to include all this information in an editorial. The reader is therefore strongly advised to read the answers. Although some participants still use the standard definition of massive transfusion, i.e. 10 units of RBC within 24 h, most now use a different definition. Most

Effects of different fibrinogen concentrations on blood loss and coagulation parameters in a pig model of coagulopathy with blunt liver injury

IntroductionThe early application of fibrinogen could potentially reverse haemodilution-induced coagulopathy, although the impact of varying concentrations of fibrinogen to reverse dilutional coagulopathy has not been studied in vivo. We postulated that fibrinogen concentration is correlated with blood loss in a pig model of coagulopathy with blunt liver injury.MethodsCoagulopathy was induced in 18 anaesthetized pigs (32 ± 1.6 kg body weight) by replacing 80% of blood volume with hydroxyethylstarch 130/0.4 and Ringers lactated solution, and re-transfusion of erythrocytes. Animals were randomly assigned to receive either 70 mg kg-1 (F-70) or 200 mg kg-1 (F-200) fibrinogen or placebo before inducing blunt liver injury using a force of 225 ± 26 Newton. Haemodynamics, coagulation parameters and blood loss were monitored for 2 hours. After death, histological examination of internal organs was performed to assess the presence of emboli and the equality of liver injury.ResultsPlasma dilution caused severe coagulopathy. Measured by thromboelastography fibrinogen restored coagulation dose-dependently. Total blood loss was significantly lower and survival better in both fibrinogen groups as compared to controls (P < 0.05). Between the F-70 (1317 ± 113 ml) and the F-200 group (1155 ± 232 ml) no significant difference in total blood loss could be observed, despite improved coagulation parameters in the F-200 group (P < 0.05). Microscopy revealed even injury pattern and no (micro) thrombi for either group.ConclusionsRestoring fibrinogen with 70 or 200 mg kg-1 after severe dilutional coagulopathy safely improved coagulation and attenuated blood loss after experimental blunt liver trauma. The higher dosage of fibrinogen was not associated with a further reduction in blood loss.

Virtual reality-based simulator for training in regional anaesthesia

BACKGROUND The safe performance of regional anaesthesia (RA) requires theoretical knowledge and good manual skills. Virtual reality (VR)-based simulators may offer trainees a safe environment to learn and practice different techniques. However, currently available VR simulators do not consider individual anatomy, which limits their use for realistic training. We have developed a VR-based simulator that can be used for individual anatomy and for different anatomical regions. METHODS Individual data were obtained from magnetic resonance imaging (MRI) and magnetic resonance angiography (MRA) without contrast agent to represent morphology and the vascular system, respectively. For data handling, registration, and segmentation, an application based on the Medical Imaging Interaction Toolkit was developed. Suitable segmentation algorithms such as the fuzzy c-means clustering approach were integrated, and a hierarchical tree data structure was created to model the flexible anatomical structures of peripheral nerve cords. The simulator was implemented in the VR toolkit ViSTA using modules for collision detection, virtual humanoids, interaction, and visualization. A novel algorithm for electric impulse transmission is the core of the simulation. RESULTS In a feasibility study, MRI morphology and MRA were acquired from five subjects for the inguinal region. From these sources, three-dimensional anatomical data sets were created and nerves modelled. The resolution obtained from both MRI and MRA was sufficient for realistic simulations. Our high-fidelity simulator application allows trainees to perform virtual peripheral nerve blocks based on these data sets and models. CONCLUSIONS Subject-specific training of RA is supported in a virtual environment. We have adapted segmentation algorithms and developed a VR-based simulator for the inguinal region for use in training for different peripheral nerve blocks. In contrast to available VR-based simulators, our simulation offers anatomical variety.

Prothrombin complex concentrates in trauma and perioperative bleeding.

923 April 2015 P rothrombin complex concentrates (PCCs) are recommended in preference to other treatments such as therapeutic plasma for urgent reversal of vitamin K antagonists.1,2 PCCs contain either three or four coagulation factors (factors ii, iX, and X, with or without factor Vii) and, depending on formulation, low doses of coagulation inhibitors such as protein C, protein S, and heparin (table 1). There are considerable variations among countries in the availability and licensing status of PCCs. For example, four-factor PCCs have been used for many years in Europe, where their license is not restricted to vitamin K antagonist reversal—they have broad approval for “treatment and prophylaxis of bleeding in acquired deficiency of the prothrombin complex coagulation factors.” in the United States, however, the first four-factor PCC was only recently approved, specifically for urgent reversal of vitamin K antagonist therapy. The mechanism of action of PCCs is important for understanding their therapeutic applications. Vitamin K antagonists such as warfarin function by reducing levels of four coagulation factors: ii, Vii, iX, and X, with the aim of preventing thromboembolism. For patients with life-threatening bleeding, rapid replacement of these coagulation factors is required, and PCCs serve as a concentrated source of the required coagulation factors. Three-factor as well as four-factor PCCs have been explored for vitamin K antagonist reversal. however, due to the absence of factor Vii, it appears that three-factor PCCs are less suitable than four-factor PCCs for patients with an international normalized ratio (inr) greater than 3.7.3 in trauma and perioperative bleeding, patients present with a variety of coagulopathies. PCCs increase thrombin generation potential by ensuring adequate levels of the key coagulation factors—notably factor ii (prothrombin), whose conversion to thrombin is facilitated by activated factor X and activated factor V. treatment with PCCs may potentially be effective in facilitating hemostasis in trauma and perioperative bleeding. however, the potential role of PCCs in these settings must be considered in the context of other treatment options and the patient’s overall coagulation status. Fibrinogen is generally the first coagulation factor to decrease below critical levels during bleeding.4 The physiological response to trauma includes an increase in thrombin generation,5 whereas fibrinogen levels are typically reduced in trauma patients upon admission to hospital.6 in cardiovascular surgery, it has been shown that fibrin formation is impaired to a greater extent than thrombin generation after cardiopulmonary bypass.7 Coagulation management algorithms based on coagulation factor concentrates have been published in relation to cardiovascular surgery8,9 and trauma (fig. 1).10 Some authors advocate the use of PCC in accordance with implementing such algorithms although it should be noted that, after antifibrinolytic medication, fibrinogen supplementation is recommended as first-line hemostatic treatment.8,10,11 PCCs may subsequently be considered for patients with ongoing bleeding despite restoration of fibrinogen levels. Throughout the management of coagulopathic bleeding, therapy should be tailored to the patient’s coagulation status. Point-of-care assessment, for example using rotEm® (tem international Gmbh, Germany) or tEG® (haemonetics Corp., USA), can help determine which of the available hemostatic therapies (e.g., therapeutic plasma, platelets, coagulation factor concentrates, or cryoprecipitate) should be administered. A deficiency in thrombin generation is more likely to arise later during the course of surgery. The potential risk of thromboembolic complications necessitates a cautious approach when using PCCs in trauma and perioperative bleeding. in these settings— unlike vitamin K antagonist reversal—levels of coagulation

Intraoperative Wake-up Test and Postoperative Emergence in Patients Undergoing Spinal Surgery: A Comparison of Intravenous and Inhaled Anesthetic Techniques Using Short-acting Anesthetics

Surgical procedures on the vertebral column may result in spinal cord damage, leading to neurological deficits that demand immediate therapeutical intervention. We designed this study to determine which anesthetic regimen allows a rapid wake-up test during and after surgery to detect neurological deficits. Fifty-four patients were randomly allocated to the following groups: group PR (propofol/remifentanil): target-controlled infusion with propofol (plasma concentration, 2–4 &mgr;g/mL) and remifentanil 0.2–0.5 &mgr;g · kg−1 · min−1; group PS (propofol/sufentanil): propofol (2–4 &mgr;g/mL) and repetitive boluses of 0.1–0.2 &mgr;g/kg of sufentanil adjusted to patients requirements; and group DR (desflurane/remifentanil): desflurane/air 3.0–4.0 vol% combined with remifentanil 0.2–0.5 &mgr;g · kg−1 · min−1. Group PS required significantly longer times for the onset of breathing (8.9 ± 1.6 min), elevation of the head (17.0 ± 3.8 min), and motion of the feet (17.0 ± 7.4 min) than group PR (6.9 ± 2.6 min, 9.3 ± 2.2 min, and 9.4 ± 2.4 min, respectively) or group DR (5.4 ± 0.8 min, 6.1 ± 1.0 min, and 6.2 ± 1.0 min, respectively). The anesthetic regimen with desflurane and remifentanil allowed faster awakening during and after surgery that permitted immediate neurological examination after spinal surgery compared with propofol/remifentanil.

Thrombin Generation Capacity of Prothrombin Complex Concentrate in an In Vitro Dilutional Model

Background The use of PCC for the treatment of trauma-induced coagulopathy potentially increase the risk of thromboembolism and disseminated intravascular coagulation, which is addressed to an imbalance of both pro- and anticoagulants. As PCCs differ in composition, we used an in vitro dilutional approach to assess the overall thrombin generation of five different PCCs through various laboratory assays. Methods The vitamin K-dependent coagulation factors, heparin, and antithrombin were assessed in five commercially available PCCs. The procoagulant potential of the PCCs was assessed in plasma and whole blood from 4 healthy donors by means of classical coagulation assays, thrombin generation assay and thromboelastometry. In order to reflect coagulopathy, whole blood was diluted to 80, 60, 40, and 20% with Ringer’s lactate solution. Results The five different PCCs were characterised by comparable levels of factors II, VII, IX and X (all around 20–30 IU/mL), whereas the heparin (0 to 17.6 IU/mL) and antithrombin (0.06 to 1.29 IU/mL) levels were remarkably different between manufactures. In vitro dilution of blood induced a prolongation of the PT and aPTT, and attenuation of thrombin generation and ExTem induced thromboelastometry. Overall, non- or low-heparin containing PCCs restored the in vitro dilutional coagulopathy, whereas PCCs containing heparin have an anticoagulant effect. The thrombin generation assay showed to be the most sensitive method for assessment of PCC effects. Conclusions This study shows that most available PCCs are not balanced regarding their pro- and anticoagulants. The effect of measured differences in thrombin generation among different PCCs requires further investigations to elaborate the clinical meaning of this finding in the treatment of trauma induced coagulopathy.

Prothrombin complex concentrate reduces blood loss and enhances thrombin generation in a pig model with blunt liver injury under severe hypothermia

Although prothrombin complex concentrate (PCC) is increasingly used for the treatment of trauma-induced coagulopathy, few studies have investigated the impact and safety of PCC for this indication. The present study was performed to assess PCC for treatment of coagulopathy after blunt liver injury under severe hypothermia. Coagulopathy in 14 anaesthetised pigs was induced by haemodilution. Subsequently, standardised blunt liver injury was induced under severe hypothermia (32.8-33.2°C). Animals were randomised to receive either PCC (35 IU kg⁻¹) or saline (control). Coagulation was assessed over the following 2 hours by thromboelastometry and thrombin generation. Internal organs were examined to determine presence of emboli. The administration of PCC showed a significant reduction in blood loss (p=0.002 vs. controls) and a significant increase in the rate of survival (p=0.022 vs. controls). Plasma thrombin generation in the PCC group increased considerably above baseline levels, with significant increases in peak thrombin levels and endogenous thrombin potential versus controls throughout the follow-up period. In addition, PT decreased significantly in the PCC group versus the control group. However, only slight improvements in thromboelastometry variables were observed. Histology showed an equal degree of liver injury in both groups, and no thromboembolism. In severely hypothermic pigs, the application of PCC corrected trauma-induced coagulopathy and reduced blood loss. Thus, the infusion of PCC might be a reasonable approach to reduce the need for blood cell transfusion in trauma. Furthermore, the impact and safety of PCC application can be monitored through thrombin generation and thromboelastometry under hypothermia.