Comparison of three methods for measuring PEG-hirudin in blood.

Three methods for measuring pegylated hirudin (PEG-hirudin), a new antithrombotic agent, in blood were compared using clinical samples. The ecarin clotting time (ECT) was performed in whole blood using a point-of-care device (TAS analyzer). The ECT was also performed in plasma, using a clotting assay in a conventional automated coagulation analyzer. Finally, a chromogenic method was used, based on thrombin inhibition and the substrate S-2238. Both clotting assays showed a linear relationship between the ECT and the PEG-hirudin concentration up to 3.0 microg/ml. The chromogenic substrate method was linear only between 0.1 and 1.0 microg/ml PEG-hirudin. The intra-assay coefficient of variation was 3.0% for the automated ECT method, 6.4% for the point-of-care ECT method and 3.4% for the chromogenic method. The inter-assay coefficient of variation was approximately 10% for both clotting methods and 3.2% for the chromogenic method. There was a high correlation (r = 0.954) in PEG-hirudin concentration between both ECT methods over the entire measuring range. The correlation of the chromogenic method with any ECT was significantly less (r < 0.89), even if only PEG-hirudin concentrations < 1.0 microg/ml were taken into account (r < 0.92). Although we clearly prefer the conventional ECT, any of the other methods may be used for monitoring PEG-hirudin in patients treated with this drug, depending on the specific application and local circumstances.     

Antithrombotic Effect of LB-30057 (CI-1028), a New Synthetic Thrombin Inhibitor,in a Rabbit Model of Thrombosis: Comparison with Inogatran

LB-30057 (CI-1028) is a novel, orally bioavailable, direct thrombin inhibitor with a Ki of 0.38[emsp4 ]nM against human thrombin. The effects of LB-30057 on thrombus formation and hemostasis were evaluated in a veno-venous shunt model of thrombosis in rabbits, and compared with inogatran, another direct inhibitor of thrombin. Each compound was studied at 5 or 6 different doses with 5 or 6 rabbits in each group. After administration as a bolus i.v. injection followed by continuous infusion, both LB-30057 and inogatran dose-dependently inhibited thrombus formation, which was measured as an increase in time to occlusion (TTO) and a decrease in thrombus weight. Both compounds also improved vena caval blood flow and reduced the overall incidence of thrombotic occlusion. LB-30057 significantly prolonged TTO from 23±4[emsp4 ]min (before dose) to 110±10[emsp4 ]min at the highest dose (0.7[emsp4 ]mg/kg+47[emsp4 ]g/kg/min) (p<0.001), and reduced thrombus weight from 57±2[emsp4 ]mg to 15±5[emsp4 ]mg (p<0.001). Occlusive thrombus formed in only one of six rabbits that received the highest dose of LB-30057 (vs. 13/13 in the control group, p<0.01). At the dose that produced the maximum antithrombotic effect (0.7[emsp4 ]mg/kg+47[emsp4 ]g/kg/min), LB-30057 increased aPTT and bleeding time approximately 2-and 2.5-fold above baseline, respectively. On a gravimetric basis, LB-30057 and inogatran displayed comparable in vivo antithrombotic efficacy. When compared to equally effective anti thrombotic doses of inogatran, LB-30057 caused less prolongation in aPTT, had no effect on PT, and tended to have less of effect on bleeding time. These results indicate that LB-30057 is an effective antithrombotic compound and it appears to have a better benefit/risk profile than inogatran in this experimental model.     

Methods for the monitoring of direct thrombin inhibitors

Direct thrombin inhibitors are available for prophylactic as well as therapeutic purposes. Application of hirudin in therapeutic doses has been shown to require drug monitoring. Currently, most experience is available for recombinant hirudin, but the principle aspects of drug monitoring are the same for all direct thrombin inhibitors. Most frequently, activated partial thromboplastin time (aPTT) and modifications of the activated clotting time (ACT) have been used for the monitoring of hirudin therapy. However, these methods are insensitive at plasma levels higher than 0.6 mg/L of hirudin, so that overdoses may be missed despite monitoring. Correlations between ecarin clotting time (ECT), enzyme immunoassays, and chromogenic substrate assays on one side and global tests on the other side are poor. Fully automated chromogenic substrate-based assays, also available as point-of-care tests (POCT), are more precise and sensitive and are not disturbed by interferents such as heparin and antithrombin. Good correlations can be observed between chromogenic assays and the ECT performed in plasma or whole blood samples. ECT can also be determined with POCT systems. Test characteristics such as imprecision and measuring range are comparable to those of the chromogenic assays. In conclusion, therapy with direct thrombin inhibitors should be monitored with chromogenic assays or ECT.     

Management of lepirudin therapy for a patient with antiphospholipid antibody syndrome using the whole blood ecarin clot time and activated partial thromboplastin time.

A patient with antiphospholipid antibody syndrome (APS) and a history of heparin-induced thrombocytopenia required lepirudin therapy. The patient had an abnormal baseline activated partial thromboplastin time (aPTT), complicating management of his therapy. We investigated whether an alternative monitoring system, using a dry reagent technology [Thrombolytic Assessment System (TAS)], could be used to monitor the patient's whole blood ecarin clot time (ECT) and aPTT. Baseline values for the ECT and aPTT were normal with this system. During a continuous infusion of lepirudin, the patient's whole blood ECT was maintained between a desired range of 150-200 s for 73% of the time. Similarly, his whole blood aPTT was maintained between 60 and 80 s for 80% of the time. In contrast, the patient's plasma-based aPTT by standard methods was consistently > 150 s. The patient underwent surgical procedures without complications. To further investigate the finding that the patient's antibody did not affect the aPTT with this system, we performed the ECT and the aPTT assays on the TAS Analyzer with plasma samples from 10 patients with APS and abnormal aPTTs. All 10 samples had plasma ECT values within the normal range. Four patients had normalization of the aPTT, suggesting that a subset of patients with APS may benefit from the TAS aPTT assay when monitoring heparin or other anticoagulation therapy.     

Comparison of GP IIB/IIIA Inhibitors and Their Activity as Measured by Aggregometry, Flow Cytometry, Single Platelet Counting, and the Rapid Platelet Function Analyzer

Background: GP IIb/IIIa inhibitors have primarily been used short-term e.g., during PTCA. They failed to show clinical benefit during long-term therapy. One reason might be the absence of a method to monitor inhibitor activity. This study compared platelet aggregometry, the rapid platelet function analyzer (RPFA) test, single platelet counting, and flow cytometric determination of receptor occupancy to measure GP IIb/IIIa-receptor inhibitor activity. Methods: Increasing doses of abciximab, tirofiban, and eptifibatide were added to whole blood in vitro. Whole blood was used for the RPFA, for single platelet counting and flow cytometry. Platelet rich plasma was prepared for aggregometry. Results: The correlation between aggregometry and RPFA results was linear for abciximab and eptifibatide. Tirofiban was a stronger inhibitor with the RPFA (IC₅₀ 7.7[emsp4 ]nM) than with aggregometry (IC₅₀ 19.6[emsp4 ]nM). The single platelet counting technique showed that even supratherapeutic concentrations of all three inhibitors could not completely suppress microaggregation. Abciximab concentrations that were equipotent to tirofiban with aggregometry were less potent with regards to the inhibition of microaggregation. This difference was more pronounced with TRAP induced microaggregation than with ADP. The flow cytometric receptor occupancy test showed that occupancy was 95[emsp4 ]% with 5[emsp4 ]g/ml abciximab and almost 97[emsp4 ]% with 10[emsp4 ]g/ml. Tirofiban reached a maximum receptor occupancy of 56[emsp4 ]%, eptifibatide 64[emsp4 ]%. Conclusions: While aggregometry is time consuming the RPFA provides results fast and with little variability. There is still a discrepancy between aggregometry and RPFA results for tirofiban. The single platelet counting technique detects the inhibition of microaggregation the relevance of which for the clinical outcome is not known. The flow cytometric receptor occupancy assay is best suited for abciximab.     

The Hemostatic Effects of Warfarin Titration in Post CABG Patients in Comparison to Placebo Treatment

Background: Since coronary artery bypass graft patients remain at risk of coronary artery and bypass graft occlusion after successful surgery, adjunct treatment regimens are under investigation. In a study of the patients of the multicenter Post Coronary Artery Bypass Graft (Post CABG) Trial, 1[emsp4 ]mg warfarin was found to have no important effect on coagulation parameters. Study design: The effects of 1, 2 and 3[emsp4 ]mg warfarin were evaluated at six-week intervals in 20 Post CABG Trial patients receiving titrated dose increases in comparison to 20 patients of similar age, gender and time from CABG treated with placebo. Results: International normalized ratio (INR) values increased with warfarin dose increments for 1, 2, and 3[emsp4 ]mg, respectively (0.95±0.16, 1.08±0.19, and 1.34±0.39) and in comparison to placebo treated patients (dose×treatment p<0.001). Factor VII coagulant activity decreased with warfarin titration (1[emsp4 ]mg, 119.0±18.3%%; 2[emsp4 ]mg, 100.6±32.8%%; 3[emsp4 ]mg, 95.0±27.8%%) and in comparison to placebo (dose×treatment p=0.008). Levels of prothrombin fragment F1.2, tissue plasminogen activator, fibrinogen and von Willebrand factor were unchanged with warfarin dose increments and in comparison to placebo. Conclusions: At doses up to 3[emsp4 ]mg, warfarin acts on the INR through a reduction of factor VII with no effect on the fibrinolytic system, fibrinogen or von Willebrand factor. At these doses F1.2 did not document reduced coagulation activity. The observations of this study were consistent with the decision in the Post CABG Trial to increase the warfarin dose above 1[emsp4 ]mg to achieve a distinct effect of warfarin that was less than full anticoagulation.     

Comparison of Two Different Ecarin Clotting Time Methods

Background: Ecarin Clotting Time (ECT) assay specifically determines the inhibition of meizothrombin by direct thrombin inhibitors (DTI). Blood coagulation factor levels lowered by vitamin K antagonists (VKA) may prolong ECT. Concomitant treatment of VKA with DTI may influence differently the two published ECT methods.Methods: Lepirudin (100–3000 ng/ml), argatroban (300–3000 ng/ml) and melagatran (30–1000 ng/ml) were added to normal plasma (NP; n = 12) samples and to plasma of patients on stable vitamin K antagonist therapy with warfarin (VKAP; n = 12). ECT assays were performed according to [5] (method 1) and according to [6] (method 2). Data were subjected to multifactorial variance analysis.Results: Normal ranges were 35.5 ± 2.8 s in NP versus 31.8 ± 1.2 s in VKAP with method 1 (p < 0.001) and 44.3 ± 3.9 s in NP vs. 51.4 ± 8.3 s in VKAP with method 2 (p < 0.004). Besides the inhibitors (p < 0.0001), the method used (p < 0.0001) and the group (NP vs. VKAP, p = 0.003) had an influence on the ECT. Inhibitors (p < 0.02) or method used (p < 0.03) and the group (NP vs. VKAP, p = 0.0001) influenced also the ECT ratio.Discussion: Both ECT methods are suitable for monitoring different DTIs over a large linear range with both methods during concomitant treatments with vitamin K antagonists. The ECT ratio improves but not abolishes the differences between the methods. Additive effects of vitamin K antagonists on ECT methods have to be taken into consideration in clinical routine.     

Clinical monitoring of hirudin and direct thrombin inhibitors.

In addition to heparin, the standard medication for prophylaxis and therapy of thromboembolism, several other substances have been developed and tested for clinical use with the aim of decreasing or eliminating side effects. Most of all, hirudin, a direct antithrombin (AT), has proved to be effective. To define the therapeutic range and to avoid underdosage or overdosage, clinical monitoring is necessary. For monitoring of hirudin, thrombin time (TT) is not suited because of the missing linearity of the standard curve. Activated partial thromboplastin time (aPTT) can be used only in the lower hirudin level range, where the standard curve is quite linear. However, high and toxic hirudin levels cannot be determined using aPTT. Another drawback is a high variation in single measurements and in the normal value of patients. Methods using chromogenic substrates are suited for determination of hirudin in plasma but cannot be used at bedside. Especially for monitoring of hirudin, the ecarin clotting time (ECT) was developed. The standard curve is linear over the entire concentration range. There are no influences by other coagulation parameters or anticoagulants. For both acute clinical situations and long-term monitoring, this method capable of point-of-care therapy (POCT) will be the method of choice.     

Cardiac Swelling Associated with Linear Radiofrequency Ablation in the Atrium

Objectives: To characterize myocardial swelling in response to application of endocardial radio- frequency ablation lesions. Background: In individual patients, we have observed that ablation in the posterior right atrium was associated with echocardiographic evidence of atrial and contiguous right pulmonary vein wall swelling. Methods: 1. Human Subjects: linear ablation was performed in the posterior right atrium in 10 subjects; a portion of the ablation lesion was contiguous to the right pulmonary vein; this area was defined as the contiguity zone. In the contiguity zone, right atrial wall thickness and pulmonary vein lumen diameter were measured utilizing intracardiac echocardiography. Measurements were made just prior to (baseline) and immediately after ablation.2. Porcine Subjects: linear ablation was performed in the posterior right atrium of 14 pigs. In the contiguity zone, atrial wall thickness, interstitial space thickness, right pulmonary vein wall thickness and lumen diameter were measured using intracardiac echocardiography. Measurements were made at baseline, immediately after ablation, and at 1, 4, 8 or 12 weeks after ablation (followup). Post-mortem pathologic evaluation of the contiguity zone was performed. Results: 1. Human Subjects: Immediately after ablation, relative to baseline right atrial wall thickness was significantly increased (9.4±3.1[emsp4 ]mm versus 5.4±1.5[emsp4 ]mm) and right pulmonary vein lumen diameter was significantly decreased (6.2±2.9[emsp4 ]mm versus 8.1±2.9[emsp4 ]mm).2. Porcine Subjects: Immediately after ablation, right atrial wall thickness (4.1±1.2[emsp4 ]mm), interstitial space thickness (1.9±1.1[emsp4 ]mm), and right pulmonary vein wall thickness (1.2±0.4[emsp4 ]mm) were each significantly increased relative to baseline (1.0±0.3[emsp4 ]mm, 0±0[emsp4 ]mm, and 0.7±0.2[emsp4 ]mm, respectively) and pulmonary vein lumen diameter was significantly decreased (5.0±1.4[emsp4 ]mm versus 6.9±2.2[emsp4 ]mm). Similar findings were made at the 1 week followup interval. At 4, 8 and 12 week followup intervals, thicknesses and lumen diameter were not significantly different from baseline. At post-mortem examination, direct measurements of wall thickness were significantly correlated with echocardiographic measurements. Histologic analysis demonstrated edema to be the cause of the early wall thickness and lumen diameter changes. Ablation lesions were transmural in the right atria of all animals; in some animals, lesion formation was also observed in the pulmonary vein wall. Conclusions: Cardiac edema resulting from right atrial linear ablation results in swelling of atrial and contiguous right pulmonary vein walls, as well as the interposed extracardiac interstitial space. These changes are associated with a decrease in pulmonary vein lumen diameter. Swelling evolves rapidly and resolves within 4 weeks.     

Determining the Optimal Pacing Intervention Rate for Vasovagal Syncope

Introduction: In this study, patients with rate hysteresis pacemakers implanted for vasovagal syncope were re-studied using serial tilt testing to determine whether, once triggered, pacing was more effective if the intervention rate was higher than the standard rate. Methods and Results: Twenty patients (mean age 55.4 years, range 23–81, 14 male) were studied, with randomisation to either initial standard rate (80–90[emsp4 ]beats/min) intervention, or to initial high rate (120[emsp4 ]beats/min) intervention. Although 18 of the 20 reported complete abolition of syncope since pacing, only 8 patients could be objectively assessed. The respective mean time to tilt down after symptom onset with standard and high rate intervention was 193±234[emsp4 ]s and 185±143[emsp4 ]s, (P >0.05). Conclusion: Repeat tilt testing was only of limited value in assessing the benefit of pacing. There was no advantage with high rate intervention in delaying the loss of consciousness (or intolerable symptoms) after the initial onset of symptoms.     

Prevention of Short Term Reversible Chronic Atrial Fibrillation by Permanent Pacing at the Triangle of Koch

Objectives: The purpose of this study was to investigate if single lead interatrial septum pacing could be effective in maintaining sinus rhythm in patients in whom restoration of sinus rhythm was only possible for a period of 2–24 hours after one or more previous electrical cardioversions, and in whom a sinus bradycardia was documented before arrhythmia restarted. The two hours limit was chosen because it was considered a sufficient time to implant a dual chamber pacemaker. Background: Alternative atrial pacing techniques have been demonstrated to be successful in preventing recurrences of atrial fibrillation (AF) in patients with sinus bradycardia. Excluding the AF occurring after only a few sinus beats, at 24 hours from electrical cardioversion an early restart of chronic AF has been reported in 12[emsp4 ]% to 17[emsp4 ]% of the patients. Methods: After sinus rhythm was restored by internal electrical cardioversion, 17 patients, 7 ablated at the AV junction, underwent a dual chamber rate response (DDDR) pacemaker implantation with a screw-in atrial lead placed in the interatrial septum. Results: After a follow-up period of 17±5 months (range 12 to 27 months) persistence of sinus rhythm was observed in 11 patients (65[emsp4 ]%). Six patients (35[emsp4 ]%) had recurrences of paroxysmal attacks, while five (30[emsp4 ]%) were totally free of AF. Recurrence of chronic AF was observed in six cases (35[emsp4 ]%) after 2 days–12 months from implantation. No dislodgements of the atrial lead and no complications were observed at implantation and during follow-up. Conclusions: Interatrial septum pacing is a safe and feasible technique with a satisfying success rate (65[emsp4 ]%) in long-term maintaining sinus rhythm in previously unsuccessfully cardioverted patients.     

Effects of Sodium Channel Blockade on Ibutilide Induced Prolongation of Repolarization in the Isolated Rabbit Ventricle

Introduction: Ibutilide fumarate is indicated for the termination of atrial fibrillation and atrial flutter. It's mechanism of action is unclear but may involve activation of a late inward Na⁺ current. Methods and Results: Twenty seven experiments were performed using an isolated perfused rabbit right ventricle preparation. In each experiment effective refractory periods (ERP) and transmembrane 90% action potential durations (APD) were measured. In 8 experiments ERP and APD were measured at baseline, in the presence of ibutilide (0.1[emsp4 ]uM), and in the presence of both ibutilide and tetrodotoxin (TTX, 2[emsp4 ]uM). In 8 experiments lidocaine (10[emsp4 ]uM) was used in place of TTX. Measures were made at 200, 400, and 800[emsp4 ]msec paced cycle lengths under each condition. The baseline values for APD at 200, 400 and 800[emsp4 ]msec cycle lengths for the experiments treated with ibutilide and TTX were 111±8, 140±14 and 159±22[emsp4 ]msec, respectively. In the presence of ibutilide, APD increased to 130±19, 192±26 and 217±35[emsp4 ]msec at 200, 400 and 800[emsp4 ]msec cycle lengths, respectively (all p0.03). After the addition of TTX there was no shortening of APD or ERP compared to treatment with ibutilide alone at any cycle length (all p0.062). Similarly, in the presence of ibutilide and lidocaine there were no changes in APD or ERP compared to treatment with ibutilide alone (all p0.41). In 11 control experiments, there were no changes in APD or ERP on serial measures after placebo and TTX or lidocaine. Conclusion: Ibutilide induced prolongation of ventricular repolarization is not affected by Na⁺ channel blockade with lidocaine or TTX in the isolated rabbit heart. These findings suggest that the effects of ibutilide are not mediated by a Na⁺ channel dependent late current or that this mechanism contributes minimally to its action in this model.     

Recombinant hirudin (HBW 023) produces stable anticoagulation unaffected by circadian variation in patients with thrombolysis for acute myocardial infarction

BACKGROUND: Circadian variations have been described for a number of haemostaticand physiological factors, all of which might predispose towardsclotting in the late morning. The anticoagulation effect ofheparin has been shown to respond in a circadian manner, resultingin minimal prolongation of the activated partial thromboplastintime (aPTT) in the morning. METHODS: Recombinant hirudin (HBW 023) given as a bolus of 0.07, 0.1,0.2 or 0.4 mg.kg^–1 followed by an infusion of 0.05, 0.06,0.1 or 0.15 mg. kg^–1 over 48 h was used as conjunctivetherapy to thrombolysis with frontloaded recombinant tissue-typeplasminogen activator (100 mg . 90 min^–1) in 40 patientswith acute myocardial infarction. APTT, activated clotting timeand free hirudin plasma levels were determined at baseline andat 8, 12, 16, 20, 24, 32, 40 and 48 h. RESULTS: The prolongation of aPTT and activated clotting time was dose-dependentand stable. In 82.5% of the patients, aPTT values were rangedbetween the highest and the lowest aPTT of <30 s. When theresults were divided into four time intervals (0000–0600,0600–1200, 1200–1800, 1800–2400) neither inthe individual patients nor in the mean values of the four differentdose groups was any significant circadian variation in aPTTor activated clotting time prolongation observed. The pharmacokineticstudies of free hirudin plasma levels revealed no circadianrhythm either. All but one patient (97.5%) had a patent vessel(TIMI grade 2/3) at the end of the hirudin infusion. CONCLUSIONS: Recombinant hirudin, in contrast to heparin, does not show anycircadian variation in its anticoagulation effect. This might,in part, explain the more stable and predictable anticoagulationachieved by hirudin, which is associated with a reduced rateof reocclusions after thrombolysis.     

The Determinants of Activated Partial Thromboplastin Time,Relation of Activated Partial Thromboplastin Time to Clinical Outcomes,and Optimal Dosing Regimens for Heparin Treated Patients with Acute Coronary Syndromes: A Review of GUSTO-IIb

Context: Unfractionated heparin remains widely utilized in the treatment of acute coronary syndromes (ACS). However, limited data exist on optimal dosing and range of activated partial thromboplastin time (aPTT) in this setting. A large trial of thrombolysis for acute myocardial infarction has reported an association between longer aPTTs and adverse outcomes. Objectives: Estimate the optimal heparin-dosing regimen in achieving early therapeutic aPTTs (50 to 75 seconds) and determine the association of aPTT and death, reinfarction, and bleeding in population with ACS. Design: Subgroup analysis within a randomized, controlled trial of 5861 patients given unfractionated heparin who had aPTTs at 6, 12, or 24 hours, with outcome analyses by weight categories. Setting: In 373 hospitals in 13 countries from May 1994 to October 1995. Patients: A total of 12,142 patients admitted for ACS, stratified by the presence (n = 4131) or absence (n = 8011) of ST-segment elevation, and randomized to 72 hours of unfractionated heparin. Results: In a simulated weight-adjusted model, based on retrospective grouping by weight, a simulated dose of 60-U/kg bolus and 12-U/kg/h infusion resulted in the highest proportion of therapeutic aPTTs. After adjustment for baseline variables, longer 12-hour aPTT was associated with the composite of 30-day death or reinfarction in patients not treated with thrombolytic therapy (odds ratio, 1.10; 95% CI, 1.00 to 1.22; P = 0.047). Longer aPTT at 6 hours was associated with increased moderate or severe bleeding for the entire cohort. There was also a significant, nonlinear correlation of the 12-hour aPTT with moderate or severe bleeding in thrombolysis-treated patients. Conclusions: For ACS patients who are treated with heparin, aPTT is highly associated with body weight. Longer aPTT within the first 12 hours is associated with adverse outcomes in ACS. Heparin dosing for ACS should be weight based.     

Endocardial Stimulation of Efferent Parasympathetic Nerves to the Atrioventricular Node in Humans: Optimal Stimulation Sites and the Effects of Digoxin

The purposes of this study were to identify optimal sites of stimulation of efferent parasympathetic nerve fibers to the human atrioventricular node via an endocardial catheter and to investigate the interaction between digoxin and vagal activation at the end organ. Methods: The ventricular rate was measured during atrial fibrillation, prior to and during parasympathetic nerve stimulation, in 8 patients taking digoxin and in 10 controls. High frequency electrical stimuli were delivered via an hexapolar or quadripolar electrode catheter, placed at the posteroseptal right atrium near the atrioventricular node (n=18 patients) or in the coronary sinus (n=12 of 18 patients). In 4 patients, stimulation was repeated after intravenous administration of 1 to 2[emsp4 ]mg of atropine. Results: Nerve stimulation prolonged the R-R interval in all patients. Stimulation close to the posteroseptal right atrium led to maximal atrioventricular nodal slowing. The mean R-R intervals at baseline and during parasympathetic nerve stimulation (60[emsp4 ]mA) from the posteroseptal right atrium and the proximal coronary sinus were 581±79[emsp4 ]ms, 2440±466, and 900±228[emsp4 ]ms respectively (p=0.0001). The response to nerve stimulation was greater in patients taking digoxin than in patients not taking the drug (p=0.02). Junctional rhythm occurred during nerve stimulation in 8/8 patients taking digoxin and 0/10 not taking the drug (p=0.0001). The response to stimulation was eliminated after atropine (p=0.01). Conclusions: Parasympathetic nerves to the atrioventricular node were stimulated from the proximal coronary sinus as well as the posteroseptal right atrium. Stimulation at the posteroseptal right atrium resulted in the greatest response, and digoxin enhanced this response. The augmented response suggests that an interaction may exist between parasympathetic stimulation and digoxin at the end organ.     

Electroanatomical Mapping (CARTO®) of Ectopic Atrial Tachycardia: Impact of Bipolar and Unipolar Local Electrogram Annotation for Localization the Focal Origin

Electroanatomical Mapping (CARTO®) allows a tridimensional localization of ectopic atrial tachycardia (EAT). No standardized recommendation exists for annotation the local activation time in EAT using this new technology. In the present study bipolar local electrogram were used for CARTO® guided RF ablation of EAT. In comparison the same maps were retrospectively analyzed by annotation the unipolar local electrogram. Methods: In 15 consecutive patients (6[emsp4 ]m, 51[emsp4 ]±[emsp4 ]14 y) with EAT CARTO® mapping was guided by annotation the earliest onset of the bipolar local electrogram. Following successful RF ablation the obtained EAT maps were subsequently evaluated by annotation of the earliest steepest negative intrinsic deflection of the unipolar local electrogram. Both CARTO® maps were compared with regard to the region of focal EAT origin. Results: RF ablation of all 15 EAT foci guided by annotation the bipolar local electrogram with CARTO® was successful with a median of 3 [1–18] pulses and a median fluoroscopy time of 10[emsp4 ]min [4–25]. All but one focus was located in the right atrium: posterior to posteroinferior region of the terminal crest in 6, septal region in 5, anterior superior region in 3 cases. One left sided EAT was located at the septum. The bipolar CARTO® map demonstrated a small territory location of earliest activation (extension of the focus 0.4[emsp4 ]cm²) in 14 out of 15 patients. In a single patient the bipolar map showed several sites of earliest local activation (extension >0.4[emsp4 ]cm²). On the other side the retrospectively achieved unipolar maps demonstrated an extended region of earliest local activation in 6 out of 15 patients (>0.4[emsp4 ]cm²). Conclusions: CARTO® maps of EAT by annotation the earliest onset of the bipolar local electrogram provide an efficacious guide for location the focal origin. Extended regions of earliest local activation in EAT might be rather determined by annotation the unipolar in comparison to the bipolar local electrogram.     

Echocardiographic Transponder-Guided Catheter Ablation Feasibility and Accuracy

Background: The utility of echocardiography for catheter guidance during percutaneous endocardial ablation is increasingly apparent. However, the technique is currently imperfect due to limitations in discerning the ablation electrode from other parts of the catheter shaft. Purpose: To examine the feasibility and accuracy of echocardiography-guided ablation using commercial ablation catheters fitted with a transponder to improve localization of the ablation electrode. Methods: Fifteen healthy pigs and five pigs with chronic anterior myocardial infarction were studied. In healthy animals, echocardiographically distinct endocardial sites in right and left cardiac chambers were targeted for ablation. In infarcted animals, the left ventricular infarction border zone was targeted. Both intracardiac (ICE; 12.5[emsp4 ]megahertz and 5[emsp4 ]megahertz) and transesophageal echocardiographic (TEE) techniques were utilized. Results: In healthy animals, transponder-guided ablation was feasible with each of the echocardiographic techniques. Accuracy was 82 % (45 of 55 lesions) with ICE-12.5[emsp4 ]MHz, 87 % (27 of 31 lesions) with ICE-5[emsp4 ]MHz, and 81 % (22 of 27 lesions) with TEE. In infarcted animals, the accuracy was 38 % (3 of 8 lesions) for ICE-5[emsp4 ]MHz and 38 % (3 of 8 lesions) for TEE. Errant lesions in healthy animals were observed in earlier experiments, due to operator misinterpretation of the plane of imaging. Errant lesions in infarcted animals were observed throughout the experimental series, and may have been due to a variable relationship between echocardiographic and histologic infarction border zones. Conclusions: Echocardiographic transponder-guided catheter ablation is feasible. Accuracy for normal endocardial targets was excellent, less so for chronic infarction border.     

Monitoring of anticoagulant effects of direct thrombin inhibitors

Monitoring of direct thrombin inhibitors with the activated partial thromboplastin time (aPTT) is limited by poor linearity and reproducibility. Recently, direct prothrombin activation methods have been developed for coagulation analysis: ecarin clotting time (ECT) and prothrombinase-induced clotting time (PiCT). Laboratory monitoring of the direct thrombin inhibitors lepirudin, argatroban, and melagatran was analyzed and compared with monitoring unfractionated heparin (UFH). Plasma samples of six healthy donors were spiked with lepirudin and argatroban extending to 3000 ng/mL, melagatran extending to 1000 ng/mL, and UFH up to 0.48 IU/mL. Clotting times of aPTT (two reagents), ECT, PiCT, and prothrombin time were determined in a KC 10, a micro instrument. At 3000 ng/mL ECT values were 339.1 +/- 25.0 seconds with lepirudin and 457.5 +/- 29.5 seconds with argatroban. ECT was 586.0 +/- 38.2 seconds with 1000 ng/mL melagatran. The PiCT method provided clotting times of 137.0 +/- 8.4 seconds with UFH, 128.0 +/- 23.4 seconds with lepirudin, and 151 +/- 23.9 seconds with argatroban, and 153.5 +/- 9.9 seconds with melagatran, with the concentrations mentioned. ECT is more sensitive to therapeutic drug concentration ranges than aPTT (prolongations of 3-7 versus 2-3). PiCT yields comparable results with direct thrombin inhibitors and UFH. This method could therefore be suitable for monitoring both drug groups.     

Differences in angiotensin convertase enzyme (ACE) activity and expression may contribute to shorter event free period after coronary artery bypass graft surgery

Aims: The goal of this study was to investigate the importance of the vascular angiotensin convertase enzyme (ACE) in coronary artery bypass graft surgery (CABG) patients. Methods: Vascular tissue (distal saphenous vein [n= 163] and/or radial artery [n= 120] segments) and blood samples were collected from CABG patients (n= 81). We studied (i) the potency of angiotensin I (AngI) and angiotensin II (AngII) to evoke vascular contractions; (ii) vascular and plasma ACE concentrations; and (iii) ACE genotype of the patients enrolled. Results: The ratio of the potencies (EC₅₀) of AngII and AngI was significantly lower in radial artery compared to the saphenous vein (0.17 ± 0.03 nM and 0.51 ± 0.14 nM, respectively, P= 0.003), suggesting a 3‐fold more effective AngI conversion in saphenous vein samples. Angiotensin constrictions were inhibited with telmisartan and captopril in both saphenous veins and radial arteries. Vascular ACE expression was significantly higher in saphenous vein compared to radial artery (9.7 ± 1.0 ng/mg and 5.3 ± 0.7 ng/mg, respectively, P= 0.01). Serum but no tissue ACE concentration was determined by ACE insertion/deletion polymorphism. Accordingly, no relation was found between serum and tissue ACE expression. Conclusion: ACE‐inhibitor therapy targeting tissue located ACE may be beneficial to patients with saphenous vein grafts after CABG surgery.     

Cytochrome P450 2B6 516G→T is associated with plasma concentrations of nevirapine at both 200 mg twice daily and 400 mg once daily in an ethnically diverse population

Objectives

The aim of the study was to characterize the impact of the cytochrome P450 2B6 (CYP2B6), CYP3A4, CYP3A5 and ATP‐binding cassette sub‐family B member 1 (ABCB1) polymorphisms on nevirapine plasma concentrations.

Methods

A total of 104 patients (82% male; 26% non‐Caucasian) were genotyped for eight single nucleotide polymorphisms at four loci (CYP2B6, CYP3A4, CYP3A5 and MDR1). Nevirapine plasma concentrations were determined using high‐performance liquid chromatography.

Results

Non‐Caucasian ethnicity [5609 ng/mL (n=27) for non‐Caucasians vs. 3771 ng/mL (n=77) for Caucasians; P<0.0001] and CYP2B6 516G→T [GG, 3574 ng/mL (n=50); GT, 4634 ng/mL (n=50); TT, 8170 ng/mL (n=4); P_{analysis of variance (anova)} =0.001] were significantly associated with a higher nevirapine trough concentration (C_trough). The latter association was maintained with both 200 mg twice daily (bid) and 400 mg once daily (qd) dosing [GG, 3527 ng/mL (n=30); GT, 4525 ng/mL (n=32); TT, 7020 ng/mL (n=2); P_anova =0.05 and GG, 3645 ng/mL (n=20); GT, 4861 ng/mL (n=17); TT, 9508 ng/mL (n=2); P_anova =0.01, respectively]. In a multivariable analysis, CYP2B6 516G→T and non‐Caucasian ethnicity remained significant predictors of nevirapine C_trough but CYP2B6 516G→T homozygosity had the greatest effect (108% higher, 46% higher). No associations were found between nevirapine C_trough and the remaining polymorphisms.

Conclusion

In this population, both non‐Caucasian ethnicity and carriage of the variant allele of CYP2B6 516G→T were significant predictors of nevirapine C_trough. The association between CYP2B6 516G→T and higher plasma nevirapine exposure was maintained at both bid and qd dosing.