Gerhardus J.A.J.M. Kuiper

Rapid and correct prediction of thrombocytopenia and hypofibrinogenemia with rotational thromboelastometry in cardiac surgery.

OBJECTIVES In the present study, the authors have investigated whether rotational thromboelastometry (ROTEM) could predict thrombocytopenia and hypofibrinogenemia in cardiac surgery using the clot amplitude after 5 minutes (A5). Another parameter, PLTEM, in which the contribution of fibrinogen is eliminated by subtracting a fibrin-specific ROTEM test (FIBTEM) from an extrinsically-activated ROTEM test (EXTEM), was investigated. Furthermore, the turnaround time of ROTEM was compared to conventional laboratory tests. DESIGN Prospective cohort study. SETTING Single academic medical center. PARTICIPANTS Ninety-seven patients undergoing cardiac surgery between July 2011 until August 2012. INTERVENTIONS The correlations between EXTEM/FIBTEM A5, A10, and maximal clot formation (MCF), EXTEM/PLTEM (A5/A10, and MCF) and platelet count, and FIBTEM (A5/A10, and MCF) and fibrinogen were evaluated using the Pearsons correlation coefficient and receiver-operating characteristic curves. Turnaround times of ROTEM tests and conventional laboratory tests were assessed in the central laboratory. MEASUREMENTS AND MAIN RESULTS EXTEM A5 and FIBTEM A5 showed an excellent correlation with A10 (R:0.99/1.00) and MCF (R:0.97/0.99). The correlation between EXTEM A5 and platelet count (R:0.74) was comparable with the correlation of A10 (R:0.73) and MCF (R:0.70) with platelet count. FIBTEM A5 predicted fibrinogen levels (R:0.87) as well as A10 (R:0.86) and MCF (R:0.87). PLTEM A5 (R:0.85) correlated better with platelet count than EXTEM A5 (R:0.74; p = 0.04) and showed significantly better area under the curve values than EXTEM for predicting thrombocytopenia (A5 p = 0.012, A10 p = 0.019). Turnaround time for ROTEM tests, 12 minutes, was comparable with emergency requests for platelet count, 13 minutes, and shorter than emergency requests for fibrinogen levels, 37 minutes. CONCLUSIONS Implementation of PLTEM and FIBTEM A5 in ROTEM-guided transfusion protocols may improve transfusion management.

Perioperative dilutional coagulopathy treated with fresh frozen plasma and fibrinogen concentrate: a prospective randomized intervention trial

Background and objectives  Treatment of dilutional coagulopathy by transfusing fresh frozen plasma (FFP) remains sub‐optimal. We hypothesized that partial replacement of transfused FFP by fibrinogen concentrate results in improved coagulant activity and haemostasis. This was tested in a controlled clinical intervention trial with patients experiencing massive bleeding during major surgery.

Additive roles of platelets and fibrinogen in whole-blood fibrin clot formation upon dilution as assessed by thromboelastometry

Blood dilution after transfusion fluids leads to diminished coagulant activity monitored by rotational thromboelastometry, assessing elastic fibrin clot formation, or by thrombin generation testing. We aimed to determine the contributions of blood cells (platelets, red blood cells) and plasma factors (fibrinogen, prothrombin complex concentrate) to fibrin clot formation under conditions of haemodilution in vitro or in vivo.Whole blood or plasma diluted in vitro was supplemented with platelets, red cells, fibrinogen or prothrombin complex concentrate (PCC). Thromboelastometry was measured in whole blood as well as plasma; thrombin generation was determined in parallel. Similar tests were performed with blood from 48 patients, obtained before and after massive fluid infusion during cardiothoracic surgery.Addition of platelets or fibrinogen, in additive and independent ways, reversed the impaired fibrin clot formation (thromboelastometry) in diluted whole blood. In contrast, supplementation of red blood cells or prothrombin complex concentrate was ineffective. Platelets and fibrinogen independently restored clot formation in diluted plasma, resulting in thromboelastometry curves approaching those in whole blood. In whole blood from patients undergoing dilution during surgery, elastic clot formation was determined by both the platelet count and the fibrinogen level. Thrombin generation in diluted (patient) plasma was not changed by fibrinogen, but improved markedly by prothrombin complex concentrate. In conclusion, in dilutional coagulopathy, platelets and fibrinogen, but not red blood cells or vitamin K-dependent coagulation factors, independently determine thromboelastometry parameters measured in whole blood and plasma. Clinical decisions for transfusion based on thromboelastometry should take into account the platelet concentration.

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

UNLABELLED 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). METHODS Specimens 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). RESULTS In ROTEM analysis the NATEM assay showed significant differences. The EXTEM assay revealed small significant differences for clot formation time: 65 seconds (SD ± 20) versus 67 seconds (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. CONCLUSION PTS 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.

The procoagulant effects of fine particulate matter in vivo

Inhalation of fine particulate matter (<2.5 μm; fine PM) has been shown to increase the risk for cardiovascular events. In this letter, we reappraise the role of tissue factor (TF) antigen and we also summarize changes in measured coagulation proteins in humans and rodents by other studies with fine PM. By considering all studies including ours, we conclude that monitoring the overall coagulation state by measuring capacity assays such as thrombin generation, and quantification of TF activity would be more suitable than determining single coagulation proteins (such as TF antigen) in order to better assess the systemic prothrombotic effects of fine PM.

Validation of a modified thromboelastometry approach to detect changes in fibrinolytic activity

BackgroundThus far, validated whole blood assays used in in vitro fibrinolysis experiments using thromboelastometry (ROTEM) are lacking or have yet to be tested in humans.The objective was first, to establish a standardized modified ROTEM approach to detect both hypo- and hyperfibrinolysis. And second, to perform a technical and clinical validation of the assay.MethodsBlood was used of healthy volunteers, patients with sepsis, patients after cardiothoracic surgery, pregnant women, and cirrhotic liver disease patients. A whole blood tissue factor (TF) activated ROTEM assay with and without the addition of recombinant tissue plasminogen activator (rTPA) was developed. Plasma fibrinolysis determinants were measured in all volunteers and patients.ResultsThirty five pM TF and additions of 125 and 175 ng/ml rTPA resulted in full lysis within 60 min in healthy volunteers. Coefficients of variation were below 10 % without and below 20 % with rTPA addition. In sepsis the hypofibrinolytic ROTEM profiles with 175 ng/ml rTPA were in line with the plasma determinants (high PAI-1, high fibrinogen, low tPA activity, and high d-dimers). After cardiothoracic surgery, reduced fibrinogen and platelet levels accounted for the reduced maximum clot firmness. The hypofibrinolytic profile is attributed to tranexamic acid use and elevated PAI-1 levels. The lowest rTPA concentration in cirrhosis resulted in hyperfibrinolysis in only few of the patients. In pregnancy normal profiles were found.DiscussionOur high rTPA concentration demonstrates hypofibrinolytic profiles adequately in sepsis and after cardiothoracic surgery. Our low rTPA concentration of 125 ng/ml seems too high for demonstrating hyperfibrinolysis in cirrhotic liver disease.ConclusionsWe were able to present a validated whole blood ROTEM approach to fibrinolysis testing using added rTPA, which can be of added value next to classical plasma based fibrinolysis assays.

The use of regression analysis in determining reference intervals for low hematocrit and thrombocyte count in multiple electrode aggregometry and platelet function analyzer 100 testing of platelet function

Abstract Low platelet counts and hematocrit levels hinder whole blood point-of-care testing of platelet function. Thus far, no reference ranges for MEA (multiple electrode aggregometry) and PFA-100 (platelet function analyzer 100) devices exist for low ranges. Through dilution methods of volunteer whole blood, platelet function at low ranges of platelet count and hematocrit levels was assessed on MEA for four agonists and for PFA-100 in two cartridges. Using (multiple) regression analysis, 95% reference intervals were computed for these low ranges. Low platelet counts affected MEA in a positive correlation (all agonists showed r2 ≥ 0.75) and PFA-100 in an inverse correlation (closure times were prolonged with lower platelet counts). Lowered hematocrit did not affect MEA testing, except for arachidonic acid activation (ASPI), which showed a weak positive correlation (r2 = 0.14). Closure time on PFA-100 testing was inversely correlated with hematocrit for both cartridges. Regression analysis revealed different 95% reference intervals in comparison with originally established intervals for both MEA and PFA-100 in low platelet or hematocrit conditions. Multiple regression analysis of ASPI and both tests on the PFA-100 for combined low platelet and hematocrit conditions revealed that only PFA-100 testing should be adjusted for both thrombocytopenia and anemia. 95% reference intervals were calculated using multiple regression analysis. However, coefficients of determination of PFA-100 were poor, and some variance remained unexplained. Thus, in this pilot study using (multiple) regression analysis, we could establish reference intervals of platelet function in anemia and thrombocytopenia conditions on PFA-100 and in thrombocytopenia conditions on MEA.

Early platelet recovery following cardiac surgery with cardiopulmonary bypass

Abstract Coronary artery bypass grafting (CABG) with cardiopulmonary bypass (CPB) is frequently associated with low platelet count (PC) and disturbed platelet function (PF). While PC is easy to measure, PF is more difficult to assess. Moreover, the time-related platelet dysfunction and recovery after CPB is not fully elucidated. Platelet dysfunction could lead to bleeding but also to coronary graft failure. Laboratory tests could provide more insights into PF after CABG. The aim of the current study was to investigate the time-related PF induced by CPB. Blood samples of 20 patients with a preoperative PC of more than 250 × 109/L were collected before incision, after weaning from CPB, and 24 h postoperative. Platelet contribution to coagulation was quantified by PLTEM (calculated by means of EXTEM and FIBTEM results). PF was assessed by multiple electrode impedance aggregometry (MEIA) in whole blood and by light transmission aggregometry (LTA) in platelet-rich plasma after stimulation with arachidonic acid (AA), adenosine diphosphate, collagen, and thrombin-receptor-activating peptide. LTA and MEIA analysis demonstrated significant platelet dysfunction after CPB, with partial recovery within 24 h after surgery. AA-induced platelet aggregation increased to higher levels within 24 h after surgery compared to baseline values as measured by LTA. PLTEM maximum clot firmness remained unchanged throughout the study. Correlation analyses revealed that MEIA and rotational thromboelastometry (ROTEM), but not LTA, were dependent on PC and hematocrit. No correlations were found between LTA, MEIA, ROTEM, PC, and clinical outcome parameters. Our results demonstrate a reversible platelet dysfunction recovering within 24 h after CPB. Interestingly, AA-induced platelet aggregation increases to higher levels during the first 24 h postoperatively, which might be important for early initiation of antiplatelet therapy after CABG. MEIA as POC test is able to detect platelet dysfunction during cardiac surgery with a PC of ≥150 × 109/L.

Platelet monitoring follow-up in a pregnant patient with HELLP syndrome

Monitoring the course of platelet function in HELLP (haemolysis, elevated liver-enzymes and low platelets) syndrome is important for clinical decision-making. We present a primigravid woman developing HELLP syndrome at 29 weeks and 6 days. Platelet function was monitored by multiple electrode aggregometry (MEA), platelet function analyzer (PFA-100®), platelet count and mean platelet volume (MPV) over an 11-day period. MPV and PFA-100® seem better predictors for platelet function than platelet levels.

Routine haemostasis testing before transplanted kidney biopsy: a cohort study

Kidney biopsy can result in bleeding complications. Prebiopsy testing using bleeding time (BT) is controversial. New whole blood haemostasis tests, such as platelet function analyser‐100 (PFA‐100) and multiple electrode aggregometry (MEA), might perform better. We postulated that PFA‐100 would be suitable to replace BT prebiopsy. In 154 patients, transplanted kidney biopsies were performed after measurement of bleeding time, PFA‐100, MEA and mean platelet volume (MPV). Bleeding outcome (haemoglobin (Hb) drop, haematuria (±bladder catheterization), ultrasound finding of a bleeding, need for (non)surgical intervention and/or transfusion) after the biopsy was correlated to each test. Male–female ratio was 2:1. 50% had a surveillance biopsy at either three or 12 months. Around 17% (had) used acetylsalicylic acid (ASA) prebiopsy. Of 17 bleeding events, one subject needed a transfusion. Most bleeding events were Hb reductions over 1 mmol/l and all resolved uneventful. BT, PFA‐100, MEA and MPV did not predict a bleeding outcome; prior ASA use however could (odds ratio 3.19; 95%‐CI 1.06 to 9.61). Diagnostic performance data and Bland–Altman analysis showed that BT could not be substituted by PFA‐100. ASA use was the best determinant of bleeding after kidney biopsy. Routine haemostasis testing prebiopsy has no added value.