Wulf Dietrich

Whole blood multiple electrode aggregometry is a reliable point-of-care test of aspirin-induced platelet dysfunction.

BACKGROUND: Aspirin is one of the most commonly ingested over-the-counter drugs. In addition to its analgesic and antiinflammatory actions, it also potently inhibits platelet aggregation. Evaluation of aspirin-induced platelet dysfunction is relevant in various clinical situations, including during complex surgeries with high bleeding risk in individuals who have ingested aspirin. In this study, we examined the suitability of multiple electrode aggregometry (MEA) for time course assessment of the antiplatelet effects of a single oral dose of 500 mg aspirin. We also determined the applicability of this method in the point-of-care (POC) setting by comparing the results of the test after different time intervals after blood sampling. METHOD: Twenty-four adult volunteers were enrolled in the study. After blood drawing at baseline, 500 mg aspirin was administered to all volunteers. Blood samples were taken at 4, 24, 56, 80, and 124 h after aspirin ingestion. At each time point, measurements were performed immediately and 30 and 60 min after drawing blood. Whole blood MEA was performed after stimulation with thrombin receptor activating peptide (TRAPtest, 32 &mgr;M) and arachidonic acid (ASPItest, 0.5 mM). Repeated measurement analysis of variance with a Bonferroni correction for multiple comparisons was performed to detect differences between time points. Assay imprecision was determined by calculating the coefficient of variation. The level of statistical significance was set to P < 0.05. RESULTS: Platelet aggregation by ASPItest was markedly decreased 4 h after aspirin intake. From the second day after aspirin intake, ASPItest values recovered with high interindividual variability, and 5 days after aspirin intake, ASPItest values did not differ significantly from baseline. TRAP-induced platelet aggregation (TRAPtest) showed no systematic changes during the study period. The resting time of the sample did not affect TRAPtest or ASPItest values. The coefficients of variation were 10% for the ASPItest and 7% for the TRAPtest. CONCLUSIONS: MEA reliably detected the effects of aspirin. Notably, 500 mg aspirin caused complete inhibition of arachidonic acid-induced platelet aggregation for 2 days in all volunteers. Aggregation returned to baseline values with a wide interindividual variation in time course by day 5. No resting time for the blood sample was required for ASPItest or TRAPtest. These assays can be implemented as real POC tests. The reproducibility of the assays studied here is within the range of modern POC analyzers.

Tranexamic acid and aprotinin in primary cardiac operations: an analysis of 220 cardiac surgical patients treated with tranexamic acid or aprotinin.

BACKGROUND: Antifibrinolytics are widely used in cardiac surgery to reduce bleeding. Allogeneic blood transfusion, even in primary cardiac operations with low blood loss, is still high. In the present study we evaluated the impact of tranexamic acid compared to aprotinin on the transfusion incidence in cardiac surgical patients with low risk of bleeding. METHODS: This prospective, randomized, double-blind study included 220 patients undergoing primary coronary artery revascularization (coronary artery bypass grafting [CABG]) or aortic valve replacement (AVR). Randomized in blocks of 20, patients received either tranexamic acid (approximately 6 g) or full-dose aprotinin (approximately 5–6 × 106 Kallikrein Inhibiting Units). Transfusion was guided by a strict transfusion algorithm. Molecular markers of hemostasis were determined to assess differences in the mode of action of the two drugs. Primary end-points were the incidence of allogeneic red cell transfusion and 24-h postoperative blood loss. Data were analyzed according to the intention-to-treat principle and compared using the &khgr;2 and Mann-Whitney U-test. RESULTS: Two-hundred-twenty patients were enrolled (CABG: 134, AVR: 86). In the aprotinin Group 47% of patients received allogeneic blood during the hospital stay as compared to 61% in the tranexamic acid group (P = 0.036). Aprotinin conferred a 23% reduction in allogeneic transfusion risk (RR 0.77, 95% CI 0.53–0.88). Overall, no significant difference in postoperative bleeding was observed, although 24-h blood loss was reduced in aprotinin-treated CABG patients (500, 350–750 mL vs 650, 475–875 mL (median, 25th–75th percentile); P = 0.039). Despite the lower transfusion rate, the hemoglobin concentration on the first postoperative day was higher in the aprotinin group (11.3, 9.9–12.1 vs 10.6, 9.9–11.6 mg/dL; P = 0.023). The fibrinolytic activity at the end of operation determined by D-Dimer was comparable in both groups. (0.15, 0.11–0.17 mg/L [aprotinin] versus 0.18, 0.12–0.24 mg/L [tranexamic acid]). The activated partial thromboplastin time was prolonged up to 4 h postoperatively in the aprotinin group, while the heparin requirement was reduced: 19% of the patients in the aprotinin group and 45% in the tranexamic acid group received at least one additional bolus heparin during cardiopulmonary bypass (P < 0.001). Troponin T levels postoperatively and on postoperative day 1 were significantly higher in the tranexamic acid group (P = 0.017). No differences in renal, cardiac, or mortality outcomes were observed. CONCLUSION: Considering the rate of transfusion of red blood cells, tranexamic acid was slightly inferior in patients undergoing CABG, but there was no difference in patients receiving AVR. Tranexamic acid seems to be less effective in operations with increased bleeding such as CABG. Clinical benefit depends on specific patient and institution characteristics (ClinicalTrials.gov NCT00396760).

A point-of-care assessment of the effects of desmopressin on impaired platelet function using multiple electrode whole-blood aggregometry in patients after cardiac surgery.

BACKGROUND: Blood loss after cardiac surgery can be caused by acquired platelet dysfunction after cardiopulmonary bypass. Monitoring of platelet function is clinically important for the identification of patients experiencing such platelet dysfunction. 1-Deamino-8-d-arginine vasopressin (desmopressin acetate, DDAVP) has been shown to augment platelet function and to reduce blood loss in patients with platelet dysfunction. In this study, we examined the feasibility of whole blood multiple electrode aggregometry (MEA) for the detection of cardiopulmonary bypass–induced platelet dysfunction and investigated its ability to monitor DDAVP treatment. METHODS: Fifty-eight consecutive patients with blood loss exceeding 150 mL/h in the first 2 consecutive hours after cardiac surgery were screened for suspected isolated platelet dysfunction. Twenty-two patients had suspected isolated platelet dysfunction and were enrolled in the study. Platelet dysfunction was assumed if conventional coagulation analyses (platelet count, activated partial thromboplastin time, international normalized ratio, and fibrinogen) did not show abnormal values as defined for transfusion of allogenic blood products, and no surgical cause of bleeding was suspected. Eleven patients received 0.3 &mgr;g/kg DDAVP, and 11 patients received no therapy in a nonrandomized manner. MEA was performed after stimulation with thrombin receptor–activating peptide (TRAPtest, 32 &mgr;M), adenosine diphosphate (ADPtest, 6.4 &mgr;M), and arachidonic acid (ASPItest, 0.5 mM) before and 2 hours after intervention. Conventional laboratory variables were recorded. The Mann-Whitney test was used to detect differences between the groups, and the Wilcoxon test was used to detect differences before and after intervention. RESULTS: All enrolled patients showed platelet dysfunction that manifested as impaired platelet aggregation in MEA before intervention. After the intervention, platelet function improved in the DDAVP group (49 U [30/72 U], median [25th/75th percentile] postintervention vs 15 U [8/21 U] preintervention for the ASPItest [P < 0.001]; 35 U [24/54 U] vs 14 U [7/28 U] for the ADPtest [P = 0.002]; and 85 U [66/115 U] vs 64 U [26/88 U] for the TRAPtest [P = 0.007]). In contrast, MEA remained unchanged in the control group (22 U [10/50 U] postintervention vs 33 U [14/57 U] preintervention for the ASPItest [P = 0.175]; 17 U [12/20 U] vs 14 U [10/28 U] for the ADPtest [P = 0.147]; and 65 U [41/89 U] vs 57 U [30/91 U] for the TRAPtest [P = 0.123]). CONCLUSIONS: Impaired platelet function after cardiac surgery can be assessed at the bedside using MEA. The effect of DDAVP on impaired platelet function can also be detected as significant improvement in platelet aggregation to all activators. This device might be helpful for the identification of patients who may benefit from DDAVP therapy.

CU-2010--a novel small molecule protease inhibitor with antifibrinolytic and anticoagulant properties.

Background:In cardiac surgery, the contact of blood with the artificial surfaces of the cardiopulmonary bypass results in activation of coagulation, fibrinolysis, and platelets, which is recognized as reason for increased bleeding tendency. Antifibrinolytics like tranexamic acid or the broad-spectrum protease inhibitor aprotinin attenuate this response. The marketing of aprotinin has been suspended after a recent clinical trial suggested increased risks associated with aprotinin. Moreover, aprotinin is a protein of animal origin and has antigenic properties. As a result, alternative antifibrinolytic compounds are desirable. Methods:This in vitro study compared the antifibrinolytic efficacy of the synthetic small molecule CU-2010 with aprotinin and tranexamic acid. Antifibrinolytic activity in plasma and whole blood of ten healthy volunteers was examined with a turbidometric method and with tissue factor-activated thromboelastometry (ROTEM®; Pentapharm, Munich, Germany). In addition, anticoagulant effects were assessed through measurement of plasma and whole blood clotting times and thrombin generation. Results:With its high affinity for plasmin (Ki, 2 nm), CU-2010 inhibited fibrinolysis comparable to aprotinin (Ki, 4 nm) and was ten times more potent than tranexamic acid. CU-2010 also inhibited plasma kallikrein (Ki < 1 nm) and factors Xa (Ki, 45 nm) and XIa (Ki, 18 nm), which was reflected in prolongation of coagulation times and an attenuation of thrombin generation. Conclusion:These findings suggest that CU-2010 has similar antifibrinolytic potency compared to aprotinin, is more potent than tranexamic acid, and possesses some anticoagulant effects.

Effects of aprotinin dosage on renal function: an analysis of 8,548 cardiac surgical patients treated with different dosages of aprotinin.

Background:The nonspecific serine protease inhibitor aprotinin is widely used in cardiac surgery to limit bleeding. Recently, concerns about the safety of this drug were raised, especially regarding impaired renal outcome. This event rate was supposed to be dose dependent. Methods:In this observational study, the authors analyzed prospectively collected data of a single-center cardiac-anesthetic database. Adult patients treated with various dosages of aprotinin were evaluated. Logistic regression analysis identified independent predictors of renal outcome. The primary endpoint was a composite of novel postoperative renal failure requiring hemodialysis, an increased postoperative creatinine plasma concentration of 2 mg/dl or greater, or a difference between maximal postoperative and preoperative plasma creatinine of 0.7 mg/dl or greater. Results:The study analyzed 8,548 patients. Multiple logistic regression (c index = 0.861) did not show a significant association between aprotinin dosage and renal outcome, as did the bootstrap procedure (odds ratio, 0.98; confidence interval, 0.90–1.07). The most relevant predictor was a preoperatively elevated creatinine concentration (odds ratio, 11.4; confidence interval, 9.05–14.3). Patients with postoperative renal impairment or failure were at higher preoperative risk and/or underwent more complex procedures. In subgroups of patients with preoperatively impaired renal function (creatinine ≥1.5 mg/dl) (n = 1,075), complex procedures (n = 1,920), or insulin-dependent diabetes (n = 650) or patients undergoing isolated myocardial revascularization (n = 4,901), no association between aprotinin dosage and adverse renal outcome was observed. Conclusion:In the current analysis, aprotinin dosage was not associated with increased adverse renal outcome. In regard to renal outcome, this analysis did not demonstrate an essential detrimental influence of aprotinin dosage on renal function.

Tumor necrosis factor-α–863 C/A promoter polymorphism affects the inflammatory response after cardiac surgery

OBJECTIVE Cardiac surgery using cardiopulmonary bypass (CPB) initiates an inflammatory response that shows a wide inter-individual range and determines postoperative morbidity. Previous research suggests that genetic diversity contributes to individual susceptibility to perioperative trauma and stress. Nevertheless, the genetic triggering of the tumor necrosis factor-alpha (TNF-α) release remains unclear. We tested two genetic single-nucleotide polymorphisms (SNPs) from the promoter region of the TNF-α gene for associations with perioperative TNF-α level after CPB. METHODS We prospectively included 122 patients, who underwent elective coronary artery bypass grafting (CABG). Patients were genotyped for TNF-α -863 C/A (rs1800630) and TNF-α -308 G/A (rs1800629). Plasma level of TNF-α was obtained preoperatively, at the end of CPB, 6h postoperatively, and on the first postoperative day (POD). RESULTS Demographic characteristics and operative data revealed no significant differences between the different genotypes. Multiple linear regression analyses revealed significant associations for the TNF-α 863 C/A polymorphism: the major -863 CC variant was associated with higher TNF-α level preoperatively (p = 0.003), after CPB (p = 0.005), and 6h postoperatively (p = 0.010), independently from CPB time, left ventricle (LV) function and age. Contrarily, the AA allele had lower TNF-α level preoperatively (p = 0.008), after surgery (p = 0.024) and 6h postoperatively (p = 0.001). For the TNF-α 308 G/A polymorphism, only few significant associations could be observed: -308 GG carriers were associated with lower TNF-α level immediately after CPB (p = 0.020), whereas 308 AA carriers were significantly associated with elevated TNF-α level preoperatively (p = 0.032) and immediately after CPB (p = 0.05). No heterozygote variant of both SNPs revealed any significant associations with perioperative TNF-α level. CONCLUSIONS The current study suggests that the major -863 CC variant determines elevated TNF-α level preoperatively and throughout the postoperative course after CPB.

Enhanced Thrombin Generation After Cardiopulmonary Bypass Surgery

BACKGROUND:Thrombin generation has a key role in the pathophysiology of hemostasis. Research has focused on the intraoperative course of hemostasis, while little is known about postoperative hemostatic activation. Thrombin generation assays quantify the potential for thrombin generation ex vivo and may be useful for determining hypercoagulability. The thrombin dynamics test (TDT) assesses the initial kinetics of thrombin formation. We hypothesized that there would be an increase in thrombin generation as well as thrombin capacity after cardiac surgery. METHODS:Two hundred twenty patients undergoing primary coronary artery bypass grafting or aortic valve replacement (AVR) surgery were prospectively enrolled. Patients undergoing AVR received warfarin beginning on the second postoperative day. In addition to prothrombin fragment (F1+2), TDT, D-dimer, and troponin T were assessed. Blood samples were obtained preoperatively, at the end of the operation, 4 hours postoperatively, and the morning of postoperative days (PODs) 1, 3, and 5. The primary end point was the change of thrombin dynamics on POD 1. RESULTS:In all patients, F1+2 peaked at the end of the operation and remained significantly elevated until POD 5. Compared with baseline and after an initial decrease, TDT was found to be significantly elevated on POD 1. After coronary artery bypass graft, TDT remained significantly elevated, whereas in AVR patients with warfarin treatment, TDT was significantly reduced on PODs 3 and 5. CONCLUSIONS:After cardiac surgery, thrombin generation continues, accompanied by a high thrombin-generating capacity and elevated fibrinogen levels. This constellation suggests a marked procoagulopathic state in the postoperative period with the potential to aggravate the risk of thromboembolic complications. Warfarin treatment after AVR significantly reduced thrombin-generating capacity.

The Influence of Antithrombin Substitution on Heparin Sensitivity and Activation of Hemostasis During Coronary Artery Bypass Graft Surgery: A Dose-finding Study

BACKGROUND:Cardiopulmonary bypass is associated with a high degree of hemostatic system activation. Supplementation of antithrombin (AT) may attenuate this activation and increase a patient’s susceptibility to heparin. However, the appropriate dosage of AT has not been defined. We sought to determine the dosage of AT concentrate necessary to achieve >100% AT activity at the end of cardiac surgery and the influence of AT on heparin sensitivity. METHODS:Forty-one patients were included. Thirty consecutive patients undergoing primary coronary artery bypass graft surgery with cardiopulmonary bypass were assigned to 3 groups of increasing dosages of AT concentrate. Eleven additional patients served as controls. AT activity and molecular markers of thrombin generation were determined, and heparin sensitivity was calculated. RESULTS:A median amount of 36.5 U (19.0; 42.8), 47.0 U (41.3; 61.6), and 50.0 U (47.4; 66.6) AT concentrate/kilogram body weight in the low, medium, and high AT group, respectively, was administered. At the end of surgery, AT activity with substitution was 84% (77; 111), 110% (92; 120), and 104% (97; 120) (median [25th; 75th percentile]), respectively, compared with 63% (49; 79) in controls (P < 0.05 all substitution groups versus control). Heparin sensitivity increased from 1.29 (1.17; 1.66) s/U heparin/kg in the control group to 2.02 (1.43; 3.65), 2.56 (1.52; 3.64), 1.72 (1.24; 2.66) s/U heparin/kg in the groups with substitution (P < 0.05 all substitution groups versus control). Compared with preoperative values, AT activity decreased during the postoperative period in all patients with a nadir on postoperative day 3 (P < 0.05 compared with baseline except for the medium AT group). Corresponding to this decrease, an increase in prothrombin fragment 1+2 and D-dimer could be observed postoperatively. DISCUSSION:High dosages of AT were required to preserve physiologic AT activity during coronary artery bypass graft surgery and to significantly enhance heparin sensitivity, respectively. However, a significant decrease in AT activity, accompanied by high levels of thrombin generation, was encountered up to 5 days postoperatively.

Factor V Leiden Does Not Affect Bleeding in Aprotinin Recipients after Cardiopulmonary Bypass

Background:Carriers of the factor V Leiden mutation (FVL) are resistant to activated protein C proteolysis. Therefore, they are at increased risk of thromboembolic events. Aprotinin is an unspecific proteinase inhibitor frequently used during cardiac surgery procedures to reduce bleeding. However, aprotinin may cause thromboembolic complications after cardiopulmonary bypass (CPB). The primary endpoint of this study was the amount of blood loss after CPB in aprotinin recipients, and secondary endpoints were thromboembolic complications. Methods:A total of 1,447 consecutive patients who underwent cardiac surgery with CPB were prospectively enrolled. All patients were screened for FVL by a fluorescence-based polymerase chain reaction method. Linear and logistic regression analyses were performed to assess associations of FVL on bleeding and thromboembolic complications. Results:One hundred seven individuals (7.4%) were heterozygous FVL carriers. No difference was found between FVL carriers and noncarriers regarding age, sex, CPB, type of operation, EuroSCORE, antiplatelet treatment, and reoperation. FVL was not significantly associated with postoperative blood loss, whereas a significant influence was found for female sex (P < 0.0001), duration of CPB (P < 0.0001), reoperation (P = 0.001), and preoperative antiplatelet treatment (P < 0.002). Multiple linear regression analysis for total blood loss had an observed power of at least 99%. FVL carriers faced the same risk for postoperative transfusion (P = 0.391), reoperation (P = 0.675), myocardial infarction (P = 0.44), stroke (P = 0.701), and 30-day mortality (P = 0.4) as did noncarriers. Conclusions:These data suggest that FVL carriers do not have reduced blood loss compared with noncarriers. Furthermore, the combination of aprotinin and FVL does not enhance the risk for thromboembolic complications.