enhanced thrombin generation after cardiopulmonary …...after prosthetic avr, all patients received...

Enhanced Thrombin Generation After CardiopulmonaryBypass SurgerySusanne Lison, MD,* Wulf Dietrich, MD, PhD,* Siegmund Braun, MD,† Johannes Boehm, MD,‡Tibor Schuster, MS,§ Anna Englhard, MD,* Anna Perchuc, PhD,� Michael Spannagl, MD, PhD,*and Raimund Busley, MD#

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 aboutpostoperative hemostatic activation. Thrombin generation assays quantify the potential forthrombin generation ex vivo and may be useful for determining hypercoagulability. The thrombindynamics test (TDT) assesses the initial kinetics of thrombin formation. We hypothesized thatthere would be an increase in thrombin generation as well as thrombin capacity after cardiacsurgery.METHODS: Two hundred twenty patients undergoing primary coronary artery bypass grafting oraortic valve replacement (AVR) surgery were prospectively enrolled. Patients undergoing AVRreceived warfarin beginning on the second postoperative day. In addition to prothrombinfragment (F1�2), TDT, D-dimer, and troponin T were assessed. Blood samples were obtainedpreoperatively, at the end of the operation, 4 hours postoperatively, and the morning ofpostoperative days (PODs) 1, 3, and 5. The primary end point was the change of thrombindynamics on POD 1.RESULTS: In all patients, F1�2 peaked at the end of the operation and remained significantlyelevated until POD 5. Compared with baseline and after an initial decrease, TDT was found to besignificantly elevated on POD 1. After coronary artery bypass graft, TDT remained significantlyelevated, whereas in AVR patients with warfarin treatment, TDT was significantly reduced onPODs 3 and 5.CONCLUSIONS: After cardiac surgery, thrombin generation continues, accompanied by a highthrombin-generating capacity and elevated fibrinogen levels. This constellation suggests amarked procoagulopathic state in the postoperative period with the potential to aggravate therisk of thromboembolic complications. Warfarin treatment after AVR significantly reducedthrombin-generating capacity. (Anesth Analg 2011;112:37–45)

Thrombin formation is the key regulatory step thatmaintains intravascular hemostasis in patients un-dergoing cardiac surgery.1–4 Once formed, thrombin

converts fibrinogen into fibrin and is the most potentphysiological platelet activator, leading to platelet aggrega-tion. Thrombin also activates factor XIII and terminates itsown formation through the thrombomodulin–protein Cpathway.1,2 Its actions, however, are not restricted to thecoagulation system; thrombin can also induce a variety ofcellular responses involved in inflammation and tissueregeneration5,6 in what is characterized as “crosstalk be-tween coagulation and inflammation.”7,8 To suppressthrombin generation during cardiopulmonary bypass

(CPB), large amounts of unfractionated heparin are used.Nevertheless, systemic thrombin generation is still beingstimulated during and after CPB, as demonstrated byelevated markers of hemostatic activation.3,9

Studies on the activation of hemostasis in patientsundergoing cardiac surgery have focused almost exclu-sively on the intraoperative period,9–15 whereas only a fewstudies have evaluated the postoperative period.16–18 Theseinvestigations revealed a pronounced hypercoagulablestate after cardiac operations. Additionally, failure tosuppress uncontrolled thrombin formation resulted indisordered hemostasis in the perioperative period.19,20

Furthermore, previous investigations concentratedmainly on bleeding and transfusion13,14,21,22 but not theconsequences of hypercoagulability.

Increased thrombin generation results in poorer clinicaloutcomes in patients with acute coronary syndrome.23–25

Likewise, most of the adverse events during and aftercardiac surgery, such as myocardial infarction, stroke, andvenous thromboembolism, may be at least partly linked toa prothrombotic state.26–28

In contrast to prothrombin fragment (F1�2) as a markerof thrombin activation and the thrombin-antithrombincomplex as a marker of thrombin activity, thrombin gen-eration tests reflect the potential for thrombin generation invitro.29 Thus, assessment of thrombin generation has beenproposed to be an indicator of the functionality of thecoagulation system and a “tool” to assess increased “clot-ting ability.” It is usually determined by measuring the

Authors’ affiliations are listed at the end of the article.

Accepted for publication August 30, 2010.

Supported solely by the Medical Faculty of the Technische UniversitaetMuenchen and the Deutsches Herzzentrum Muenchen, Munich, Germany.

Clinical trial registration: ClinicalTrials.gov NCT00396760 (http://www.clinicaltrials.gov).

Conflicts of Interest: See Disclosures at the end of the article.

The authors WD and RB formerly held positions at the German Heart CenterMunich. AP is currently affiliated with August-Lenz-Stiftung am Institutfuer Prophylaxe und Epidemiologie der Kreislaufkrankheiten, LudwigMaximilians Universitaet, Munich, Germany.

Address correspondence and reprint requests to Wulf Dietrich, MD, PhD,Working Group of Perioperative Hemostasis, Ludwig Maximilians Uni-versitaet, Winthirstr. 4, 80639 Munich, Germany. Address e-mail [email protected].

Copyright © 2010 International Anesthesia Research SocietyDOI: 10.1213/ANE.0b013e3181fc6df0

January 2011 • Volume 112 • Number 1 www.anesthesia-analgesia.org 37

formation and inhibition of thrombin using “slow” throm-bin substrates over a period of up to 1 hour.30–32 Thethrombin dynamics test (TDT)33,34 uses a “fast” chromo-genic substrate (i.e., a substrate that is rapidly converted bythrombin) to detect initial thrombin-formation kinetics.This test was applied in the present investigation to de-scribe the dynamics of thrombin generation during andafter cardiac surgery.

The aim of this study was to evaluate the activation ofplasmatic coagulation in cardiac surgery with special em-phasis on the postoperative period. We hypothesized thatcardiac surgery leads to a prothrombotic state postopera-tively that may be partly independent of the degree ofintraoperative hemostatic activation.

METHODSAfter IRB approval (Ethical Committee, Medical Faculty ofthe Technische Universitaet Muenchen, Munich, Germany)and written informed consent, 220 patients older than 18years and scheduled for primary coronary artery bypassgraft (CABG) or aortic valve replacement (AVR) surgerywere prospectively enrolled in this study. Exclusion criteriawere emergency operation, known previous exposure toaprotinin, warfarin treatment within 5 days of the opera-tion, or refusal of allogeneic blood transfusion. Aspirin orthienopyridine therapy did not lead to exclusion. Demo-graphic data, medical and surgical history, medications,and outcomes were recorded by trained research staff usingdefined protocols in a purpose-built case report form. Thisnonindustry sponsored study was supported by the De-partment of Anaesthesiology of the Deutsches Herzzen-trum Muenchen, Munich, Germany. Quantitative dataabout blood loss and transfusion requirements are pub-lished elsewhere.35

In all patients, standard anesthesia was performed usingsufentanil and midazolam supplemented with sevofluraneinhalation; neuromuscular blockade was achieved andmaintained by either pancuronium bromide or rocuro-nium. During CPB, nonpulsatile pump flow was used withmoderate systemic hypothermia of 32°C. CPB was per-formed using a membrane oxygenator, an open cardiotomyreservoir, and uncoated tubing systems. The oxygenatorwas primed with a total volume of 1500 mL crystalloidsolution. Myocardial protection consisted of administrationof either cold crystalloid or blood cardioplegia. Intraopera-tive cell salvage of shed blood was used for all patients.

The patients were randomly assigned to receive eitheraprotinin (Trasylol Bayer AG, Leverkusen, Germany) ortranexamic acid (Cyklokapron; Pfizer Pharma GmbH, Ber-lin, Germany) during surgery. The anesthesiologist wasblinded to group allocation. Patients received either 2 � 106

kIU aprotinin over 10 minutes, followed by a continuousinfusion of 5 � 105 kIU aprotinin/h for the duration ofsurgery with an additional bolus of 2 � 106 kIU aprotinin tothe CPB priming, or 2 g tranexamic acid followed by acontinuous infusion of 1 g/h and an additional bolus of 2 gto the CPB circuit.

For CPB, the patients received anticoagulation withporcine heparin 375 U/kg; 10,000 U of heparin was addedto the priming solution. The degree of heparinization wasdetermined by measuring the kaolin activated clotting time

(Hemochron 800; Intern Technidyne Corp., Edison, NJ)every 30 minutes, with a target activated clotting time of480 seconds. After termination of CPB, heparin was re-versed by protamine chloride in a ratio of 1:1 of the initialheparin dosage.

The trigger for transfusion of allogeneic blood was ahematocrit �18% during CPB or �21%–24% postopera-tively or if physiological signs (tachycardia �100/min withadequate volume load and pain care and/or tachypneawith �25 breaths per minute and/or a decrease in centralvenous oxygen saturation below 65%) of the patient indi-cated a need for improved oxygen supply. Postoperatively,patients’ lungs were ventilated in the intensive care unituntil warmed to 37°C, oxygenation and hemodynamicswere sufficient, and blood loss was �100 mL/h. A standard12-lead electrocardiography was applied postoperativelyon the morning of the first, third, and fifth postoperativedays (PODs).

After prosthetic AVR, all patients received oral warfarintreatment beginning the second day after the operation.

Blood SamplingBlood samples were obtained before induction of anesthe-sia, at the end of the operation, 4 hours postoperatively,and the morning of PODs 1, 3, and 5. After discarding thefirst 10 mL, blood was collected in citrated plastic tubes(sodium citrate 0.106 mol/L; Sarstedt, Numbrecht, Ger-many) without stasis from an arterial catheter or a centralvenous catheter. Blood samples were centrifuged immedi-ately (10 minutes, 3000g); the plasma was frozen in multiplealiquots of platelet-poor plasma and stored at �80°C. Allmeasurements were performed directly after thawing theplasma.

Thrombin Dynamics TestThe TDT (Pefakit, in-TDT; Pentapharm, Basel, Switzer-land) allows for the rapid detection of thrombin-formation kinetics in platelet-poor plasma by activationof the intrinsic coagulation pathway and optical detec-tion of clots in standard coagulation analyzers. The assayis commercially available from Pentapharm, Basel, Swit-zerland using intrinsic coagulation activation, i.e., viafactors XII, XI, IX, VIII, X, V, and ultimately thrombin.Analyses were performed on the Behring CoagulationSystem Analyzer (BCS; Siemens, Marburg, Germany).Briefly, coagulation is triggered by the addition of 60 �Lactivator reagent to 60 �L platelet-poor plasma (contactphase activator based on ellagic acid and phospholipids),diluted 1:3 in 0.9% saline. After incubation for 180seconds at 37°C, 60 �L of reagent containing CaCl2, fibrinpolymerization inhibitor (H-Gly-Pro-Arg-Pro-OH) thatallows for undisturbed optical detection,36,37 and a chro-mogenic substrate (H-D-CHG-Ala-Arg-pNa�2AcOH) isadded. Thrombin formation is detected with a “fast” chro-mogenic substrate, i.e., a substrate that is readily convertedby thrombin and releases p-nitroaniline; color intensity iscontinuously recorded at 405 nm over 4 to 10 minutes. Thepeak value of the first derivative representing the maxi-mum velocity of the conversion of the thrombin substrate isdetermined. This parameter characterizes the dynamics of

Thrombin Generation and Cardiac Surgery

38 www.anesthesia-analgesia.org ANESTHESIA & ANALGESIA

thrombin formation and is expressed as a percentage ofthrombin formation of a normal control population.

Further laboratory tests determined activated partialthromboplastin time (aPTT) (Pathromtin SL; Dade Behring,Marburg, Germany) (reference range, 25–37 seconds);prothrombin time (PT) (%) (Innovin, Dade Behring) (refer-ence range PT, 70%–120%); antithrombin (Berichrom Anti-thrombin, Dade Behring) (reference range, 80%–120%);d-dimer (d-dimer PLUS immunoturbidimetric Assay, DadeBehring) (reference range, 0.06–0.25 mg/dL); cardiac tro-ponin T (on Elecsys 2010 analyzer with an enzyme immu-noassay based on electrochemiluminescence; Roche Diag-nostics, Rotkreuz, Switzerland) (lower detection limit 0.01ng/mL); fibrinogen with a modification of the Claussmethod (Multifibren, Dade Behring) (reference range,180–350 mg/dL); and thrombin activation, determined byformation of prothrombin fragment F1�2 using a microtiterplate–based sandwich immunoassay (Enzygnost monoclo-nal, Dade Behring) (reference range, 69–229 pmol/L).

Myocardial infarction was defined as the new occur-rence of Q waves on electrocardiogram or an increase introponin T with a postoperative cutoff �1.58 ng/mL38;ischemic stroke was confirmed by computed tomographyafter clinical findings.

Sample Size and Power ConsiderationsStudy design and calculation of sample size were initiallybased on 24-hour blood loss as reported previously byDietrich et al.35 A sample size of 220 was deemed sufficientfor detecting changes in TDT values of at least 2.5% frombaseline to POD 1, with 80% power, assuming a commonstandard deviation of 6%.

Statistical AnalysisThe primary end point of the investigation was the changein thrombin dynamics on the first POD as compared withpreoperative values. Secondary end points were changes inthrombin dynamics and prothrombin fragment F1�2 levelsup to the fifth POD. Additionally, we analyzed the influ-ence of 2 different intraoperative antifibrinolytic treatmentsas well as postoperative warfarin treatment on these pa-rameters. Distributions of quantitative data are described

by reporting the median with 25th to 75th percentiles.Categorical variables are reported as percentages.

The Friedman test was used to evaluate longitudinalchanges in quantitative measurements over time. In thecase of a statistically significant Friedman test, the nonpara-metric post hoc multiple comparison analysis by Daniel39

was applied to assess the significance of changes in eachtime point compared with baseline values. In addition,analysis of patients divided in subpopulations (CABG andAVR) or subgroups (aprotinin and tranexamic acid), re-spectively, was performed in the same manner. To assessintergroup differences at single predetermined time points,the Mann-Whitney U test was used. In all statistical com-parisons, a P value �0.05 was considered statisticallysignificant. The Pearson correlation coefficient (r) was usedto quantify bivariate correlation of quantitative measure-ments at distinct time points.

RESULTSTwo hundred twenty patients were enrolled in this study.CABG was performed in 134 patients; 86 patients hadAVRs. All patients underwent first-time cardiac surgery.Demographic and procedural data are summarized inTables 1 and 2.

In all patients, thrombin dynamics (median; 25th–75thpercentile) were significantly reduced postoperatively(83%; 77%–87%, P � 0.05) compared with baseline (97%;93%–101%) (Table 3; Figs. 1 and 2). On POD 1, a significantincrease in thrombin dynamics was found (100%;96%–103%, P � 0.05) (Table 3; Figs. 1 and 2). Separateanalysis of the aprotinin and tranexamic subgroups onPOD 1 revealed a decreased TDT in the aprotinin subgroupcompared with the tranexamic acid subgroup (aprotininsubgroup 99%; 94%–102% vs tranexamic subgroup 101%;96%–105%, P � 0.05) (Fig. 3).

All patients with prosthetic AVR began oral warfarinanticoagulation on POD 2 and demonstrated significantlyattenuated thrombin dynamics on the third and fifth PODscompared with baseline (P � 0.05) and compared withpatients without warfarin treatment after CABG (P � 0.05)

Table 1. Demographic DataAll patients(n � 220)

Aprotinin(n � 110)

TA(n � 110)

CABG(n � 134)

AVR(n � 86)

Gender (female) 66 (30.1) 28 (25.5) 38 (34.5) 22 (16.4) 44 (51.2)Age (y) 70 (64–76) 69 (63–75) 71 (64–77) 70 (64–75) 64 (71–46)BMI (kg/m2) 26 (24–29) 26 (25–29) 26 (24–29) 26 (24–29) 26 (25–29)Previous MI 62 (28.2) 28 (25.5) 34 (30.9) 52 (38.8) 10 (11.6)Previous stroke 3 (1.4) 3 (2.7) 0 (0) 2 (1.5) 1 (1.2)Aspirin 60 (27.3) 33 (30) 27 (24.5) 44 (32.8) 16 (18.6)Clopidogrel 27 (12.3) 15 (13.6) 12 (10.9) 24 (17.9) 3 (3.5)Hemoglobin preoperatively (g/dL) 13.5 (12.4–14.6) 13.6 (12.5–14.6) 13.2 (12.3–14.7) 13,6 (12,5–15,0) 13.1 (12.4–13.9)EF (%) 60 (48–70) 58 (43–68) 62 (49–70) 60 (47–70) 61 (50–70)EuroScore 2.0 (1.0–3.0) 2.0 (1.0–3.0) 2.0 (1.0–3.0) 2.0 (1.0–3.0) 2.0 (1.0–3.0)Preoperative risk score 2.0 (1.0–3.0) 2.0 (1.0–3.0) 2.0 (1.0–3.0) 2.0 (1.0–3.0) 2.0 (1.0–3.0)NYHA II 75 (34.1) 36 (32.7) 39 (35.5) 45 (33.6) 30 (34.9)NYHA III 141 (64.1) 70 (63.6) 71 (64.5) 86 (64.2) 55 (64)NYHA IV 4 (1.8) 4 (3.6) 0 (0) 3 (2.2) 1 (1.2)

Values are n (%) or median (25th–75th percentiles). Preoperative risk score was calculated according to the Cleveland Clinic Risk Score.TA � tranexamic acid; CABG � coronary artery bypass graft; AVR � aortic valve replacement; BMI � body mass index; MI � myocardial infarction; EF �left-ventricular ejection fraction; NYHA � New York Heart Association.


(Fig. 4). In addition to these findings, in the AVR sub-population, the PT was significantly decreased comparedwith baseline (97%; 92%–103%) on POD 3 (60%;33%–94%, P � 0.05) and POD 5 (31%; 23%– 45%, P � 0.05)as well as in comparison to patients after CABG on PODs3 and 5 (P � 0.05).

Molecular markers of hemostasis are summarized inTable 3 and Figure 5. Prothrombin fragment F1�2, a param-eter of prothrombin cleavage, peaked immediately aftersurgery and remained significantly elevated throughoutthe postoperative period (P � 0.05). After CABG, signifi-cantly elevated prothrombin fragment F1�2 levels werefound on POD 5 compared with warfarin-treated patientsafter AVR (after CABG 301 pmol/L; 222–389 pmol/L vsafter AVR 198 pmol/L; 159–273 pmol/L, P � 0.05). Nosignificant difference in postoperative prothrombin frag-ment F1�2 levels was found between patients treated withaprotinin and those treated with tranexamic acid. Levels offibrinogen were significantly decreased at the end of sur-gery and 4 hours postoperatively but later increased sig-nificantly above baseline on POD 1 (P � 0.05). Levelsincreased further during the residual observation periodand reached a median of 693 mg/dL (584–778 mg/dL) on

POD 5 (P � 0.05 vs baseline). Plasmin-dependent fibrindegradation as measured by d-dimer levels showed aminimal increase at the end of surgery and increasedfurther in the postoperative period. Antithrombin wassignificantly reduced at the end of surgery and 4 hourspostoperatively (P � 0.05).

For the first POD, a positive but weak correlation wasfound between thrombin dynamics and prothrombin frag-ment F1�2 levels (r � 0.18).

In all patients, troponin T values (Table 3) were signifi-cantly increased in the postoperative course compared withbaseline and peaked 4 hours after the operation (P � 0.05).Nine patients showed an increase above the defined cutoffof 1.58 ng/mL postoperatively; 5 of these patients hadelevated values above 1.58 ng/mL on POD 1. No correla-tion was found between troponin T and TDT. No Q wavemyocardial infarction was observed.

Three patients (1.4%) died either in the hospital orwithin 30 days after the operation. One patient died ofischemic stroke on POD 5. The other patient died ofmultiorgan failure on POD 42. The third patient experi-enced sudden cardiac death after discharge on POD 14(Table 2).

Table 2. Procedural DataAll patients(n � 220)

Aprotinin(n � 110)

TA(n � 110)

CABG(n � 134)

AVR(n � 86)

CABG 134 (60.9) 66 (60.0) 68 (61.8)AVR 86 (39.1) 44 (40.0) 42 (38.2)OP duration (min) 200 (165–235) 195 (165–235) 200 (170–234) 210 (185–250) 175 (146–210)CPB duration (min) 85 (72–101) 88 (73–102) 83 (71–98) 83 (71–99) 88 (74–102)Time on ventilator (min) 520 (380–715) 548 (406–724) 480 (373–689) 435 (345–675) 530 (409–706)ICU stay (d) 1.0 (1.0–1.0) 1.0 (1.0–1.0) 1.0 (1.0–1.8) 1.0 (1.0–1.8) 1 (1.0–1.0)Hospital stay (d) 9 (8–12) 9 (8–12) 9 (8–12) 9 (8–11) 10 (8–13)Hospital mortality�Mortality within 30 d

3 (1.4) 2 (1.8) 1 (0.9) 2 (1.5) 1 (1.2)

Rethoracotomy for bleeding 5 (2.3) 2 (1.8) 3 (2.7) 2 (1.5) 3 (3.5)Blood loss 24 h (mL) 500 (300–713) 500 (300–700) 565 (300–800) 600 (400–800) 340 (250–588)Packet cells, allogenic (yes) 119 (54) 52 (47.3) 67 (60.9) 67 (50) 52 (60.5)Platelets total (yes) 4 (1.8) 0 4 (3.6) 3 (2.2) 1 (1.2)Fresh frozen plasma total (yes) 20 (9.1) 10 (9.1) 10 (9.1) 17 (12.7) 3 (3.5)

Values are n (%) or SD or median (25th–75th percentiles).TA � tranexamic acid; CABG � coronary artery bypass graft; AVR � aortic valve replacement; OP � operation; CPB � cardiopulmonary bypass; ICU � intensivecare unit.

Table 3. Laboratory Data (All Patients)Hemoglobin

(g/dL)Fibrinogen(mg/dL)

D-dimer(mg/dL)

Troponin(ng/mL)

Preop 13.5 (12.4–14.6) 328 (249–418) 0.13 (0.09–0.21) 0.01 (0.01–0.01)Opend 10.5 (9.6–11.4) 213 (181–338) 0.17 (0.12–0.23)4 h opend 247 (215–338) 0.14 (0.10–0.21) 0.25 (0.13–0.43)POD 1 10.8 (9.9–11.8) 411 (328–479) 0.15 (0.11–0.21) 0.23 (0.13–0.44)POD 3 692 (574–778) 0.20 (0.15–0.25) 0.19 (0.09–0.39)POD 5 10.7 (9.8–11.6) 693 (584–778) 0.26 (0.21–0.34) 0.16 (0.06–0.36)

F1�2 (pmol/L) TDT (%) PT (%) aPTT (s) AT (%)Preop 160 (119–220) 97 (93–101) 98 (92–104) 32.7 (30.3–35.5) 74 (68–80)Opend 922 (652–1194) 83 (77–88) 70 (62–81) 47.8 (38.2–58.9) 58 (51–63)4 h opend 512 (349–800) 96 (91–100) 92 (82–100) 34.7 (31.3–40.1) 65 (58–72)POD 1 222 (149–333) 100 (96–103) 95 (86–103) 34.5 (31.9–38.2) 74 (65–81)POD 3 246 (190–306) 98 (92–102) 93 (55–107) 37.0 (32.6–43.5) 77 (66–86)POD 5 256 (184–361) 98 (85–104) 74 (30–108) 34.3 (29.8–43.5) 86 (75–96)

Values are median (25th–75th percentiles).Preop � before the operation; opend � at the end of the operation; POD � postoperative day; F1�2 � prothrombin fragment; TDT � thrombin dynamics test; PT �prothrombin time; aPTT � activated partial thromboplastin time; AT � antithrombin.



DISCUSSIONOur study revealed a pronounced hypercoagulable stateafter cardiac surgery as demonstrated by increased pro-thrombin fragment F1�2, accompanied by a high thrombin-generating capacity and increased fibrinogen levels. The

majority of studies evaluating hemostasis and cardiac sur-gery focus on hemostatic changes during the course of CPBand the immediate postoperative period.40 Hemostaticmarkers of the activation and activity of prothrombin(prothrombin fragment F1�2 and thrombin-antithrombincomplex, respectively) increase during surgery and peakafter protamine administration.10,11The current study sup-ports previous research41 demonstrating alterations in he-mostasis up to 30 days after surgery. Prothrombin fragmentF1�2, as an index of continuing thrombin generation,peaked immediately after surgery9 and remained signifi-cantly elevated until POD 5. On the contrary, TDT, as amarker of thrombin-generating capacity, decreased imme-diately after the operation but increased again up to POD 1in all patients. Fibrinogen showed a similar course withsignificantly reduced values postoperatively and a consid-erable and significant increase starting on POD 1 andcontinuing until POD 5. d-dimer values showed only aminimal increase at the end of the operation as well as 4hours after the operation. However, one must consider thatall patients were treated with high-dose antifibrinolytics.

When thrombin is finally formed, prothrombin frag-ment F1�2 is cleaved from the prothrombin molecule.Therefore, during and immediately after surgery, pro-thrombin fragment F1�2 should be enhanced. Thrombingeneration capacity reflects the potential amount of throm-bin that is reduced during and early after surgery becauseof the consumption of clotting factors and hemodilution.This situation contributes to the possible risk of the widelydocumented13,14,21,22 increased bleeding tendency in theperioperative period.

In patients with acute coronary syndrome, but withoutsurgical intervention, high levels of thrombin generationare predictors of an increased risk of an unfavorable

Figure 1. First derivative of the optical density curve of a fastchromogenic substrate converted by thrombin preoperatively, at theend of the operation, and on the first postoperative day. The thickline represents results in all patients; the dotted line represents thenormal control plasma; and the dashed line represents the patho-logical control plasma.

Figure 2. Course of thrombin dynamics in all pa-tients before the operation (1), after the operation(2), 4 hours after the operation (3), and on postop-erative day (POD) 1 (4), POD 3 (5), and POD 5 (6).Immediately after the operation, a decrease fol-lowed by an increase in thrombin dynamics wasnoted. Values are presented as median values(25th–75th percentiles, whiskers 10%/90%) andthe asterisk denotes a significant difference invalues at the primary end point POD 1 comparedwith baseline values (P � 0.05).


outcome.24,25 Activation of hemostasis during cardiac sur-gery may present a comparable “hemostatic constellation.”CPB results in a significant enhancement of thrombingeneration and activation of fibrinolysis, despite adequateheparinization.3 In addition to an increased perioperativebleeding tendency, impaired hemostasis after CPB also

leads to prothrombotic effects.6,21,22,42 Increased levels ofhemostatic activation are associated with evidence of post-operative organ dysfunction after cardiac surgery.43 Ourresults demonstrate continuing thrombin activity until thefifth POD after cardiac surgery, data that may explain theclinical findings of postoperative ischemic events such asmyocardial infarction, graft occlusions, ischemic stroke, orvenous thromboembolism. However, the number of pa-tients in the present study was too small to demonstrate anassociation between hemostatic activation and adverseoutcome.

The kinetics of thrombin formation are of fundamentalimportance for the hemostatic process in vivo and are notreflected by routine coagulation tests such as aPTT or PT.44

Thrombin generation capacity is a general “function test”providing a global picture of plasmatic hemostasis byshowing the potential rate of thrombin production.31,45

Hemker and Beguin44 once compared prothrombin frag-ment F1�2 to a smoke detector that reports a fire, whereasthe thrombin generation test would depict the risk of fire.Thrombin generation tests have been used to assesshypocoagulable states such as clotting factor deficiencies(e.g., hemophilia),46,47 hypercoagulable states such asthrombosis29,48 and coronary disease,49 and it is alsoused to monitor different antithrombotic therapies.31,50

TDT is a fairly new modification of the thrombin-generating assay described by Hemker et al.45 In contrastto the original assay, TDT focuses on the physiologicallyrelevant initial phase of thrombin generation. Thus, a“fast” thrombin substrate is used, which increases thespeed of the test.34 Additionally, it is an easily applicablemethod that is performed on standard automated hemo-stasis instrumentation.

Only a few studies examined TDT,33,34,51 and no data arefound concerning TDT after cardiac surgery. Similar to ourresults, Tanaka et al.30 demonstrated a reduced thrombingeneration compared with prebypass values by using the

Figure 3. Separate analysis of thrombin dynamicsin the aprotinin group and the tranexamic groupbefore the operation (1), after the operation (2), 4hours after the operation (3), and on postoperativeday (POD) 1 (4), POD 3 (5), and POD 5 (6). Afteraprotinin administration, reduced TDT values wereobserved after the operation and on POD 1. Valuesare median values (25th–75th percentiles, whis-kers 10%/90%).

Figure 4. To test the influence of warfarin treatment, patients with(after aortic valve replacement [AVR]) or without (after coronary arterybypass graft [CABG]) postoperative warfarin treatment were analyzedseparately. Thrombin dynamics are shown before the operation (1),after the operation (2), 4 hours after the operation (3), and onpostoperative day (POD) 1 (4), POD 3 (5), and POD 5 (6). Warfarinuse decreased thrombin dynamics test values in the postoperativeperiod. Values are presented as median values (25th–75th percen-tiles, whiskers 10%/90%).



method described by Hemker et al.,45 which was also con-firmed by others.32,52 The postoperative period has not yetbeen investigated. In our patients, high TDT values and aweak but positive correlation between prothrombin fragmentF1�2 levels and TDT on the first POD suggest that althoughongoing thrombin generation is observed, the capacity of theplasma to generate thrombin remains. The high postoperativefibrinogen levels are attributable to the acute phase responseand contribute to further hypercoagulability.

Before again increasing, a decrease in TDT was observedin our study at the end of the operation. This indicates theconsumption of clotting factors during and immediatelyafter the operation as well as the hemodilution effect ofCPB. The marked reincrease in TDT values from measure-ments immediately postoperatively to measurements onPOD 1 highlights that the prothrombotic state is at leastpartly independent of CPB. Thus, these results raise thequestion of whether CPB is really the culprit of hemostaticdisturbances after open heart surgery.15,16,53 This hypoth-esis is supported by the results of Parolari et al.18 who

found an increased prothrombotic state (as measured byprothrombin fragment F1�2) after on-pump as well asoff-pump coronary surgery, which lasted as long as 1month after the operation.

Interestingly, when started on POD 2, warfarin de-creased TDT as well as PT and increased aPTT on PODs 3and 5.54,55 PT and aPTT only detect the onset time ofthrombin formation, whereas thrombin generation tests ingeneral quantify the thrombin generation potential andtherefore provide information in addition to PT in warfarintherapy.55,56 In the CABG subpopulation, TDT values werehigher in comparison to the warfarin-treated patients.Particularly, in regard to increased prothrombin fragmentF1�2 levels, this result reveals a pronounced hypercoagu-lable state. Currently, postoperative antithrombotic pro-phylaxis is often disregarded in cardiac surgery. Futurestudies must evaluate whether more aggressive antithrom-botic therapy may reduce thromboembolic events in car-diac surgery.

After aprotinin administration, less pronounced TDTvalues were observed at the end of surgery and on POD 1in the aprotinin group compared with the tranexamic acidgroup. Aprotinin has anticoagulant properties that mightpreserve hemostasis by inhibiting the contact pathwayduring and directly after surgery.20,57

This study has several limitations. First, because of thesmall number of patients and the low incidence of compli-cations, our study could not demonstrate an associationbetween adverse events or troponin T increases with el-evated TDT values. Second, at the time of the study, only anintrinsic activator of TDT was commercially available formeasurements. Extrinsic activation, indicating factor VIIand factor X activity and mainly activated by tissue factor,might provide additional information about a procoagula-tory state. Third, blood samples were obtained discontinu-ously with a total of 6 samples during a study period of 5days. Therefore, we might have missed peaks of prothrom-bin fragment F1�2 levels or TDT during the study period.Fourth, even though TDT values were significantly in-creased on POD 1 in all patients and stayed increased onPODs 3 and 5 in patients without warfarin treatment, itshould be noted that TDT values stayed within the range ofnormal during the whole observational period.

In conclusion, this study demonstrated enhanced throm-bin generation and a high thrombin capacity after cardiacsurgery. These observations confirm the hypothesis thathemostatic alterations after cardiac surgery result in aprothrombotic state, which potentially increases the risk ofthrombotic complications. This hypercoagulable milieumight be partly independent of CPB and partially related tosurgical stress.

AUTHOR AFFILIATIONSFrom the *Working Group of Perioperative Hemostasis,Ludwig Maximilians Universitaet, Munich; †Department ofClinical Chemistry, and ‡Department of Cardiac Surgery,Deutsches Herzzentrum Muenchen, Munich; §Department ofMedical Statistics and Epidemiology, Klinikum Rechts der Isar,Technische Universitaet Muenchen, Munich, Germany; �De-partment for Research & Development, Pentapharm Ltd.,

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Figure 5. The course of prothrombin fragment F1�2 (top panel),fibrinogen (middle panel), and D-dimer (bottom panel) before theoperation (1), after the operation (2), 4 hours after the operation (3),and on postoperative day (POD) 1 (4), POD 3 (5), and POD 5 (6). Thecourse of these molecular markers demonstrated an intermittenthypercoagulable state in the postoperative period. Values are pre-sented as median values (25th–75th percentiles).


Basel, Switzerland; and #Department of Anesthesiology, Be-handlungszentrum Vogtareuth, Vogtareuth, Germany.

DISCLOSURESSL, WD, and MS received speaker honoraria from Bayer Corp.WD was a paid consultant at the FDA advisory committeemeetings in September 2006 and September 2007 in WashingtonDC. RB was a member of an advisory board for aprotinin. APworked in the Department of Research & Development at Pen-tapharm Ltd., Basel, Switzerland in the course of her dissertationat the University of Basel, Switzerland. No other potential con-flicts of interest relevant to this article are reported.

ACKNOWLEDGMENTSWe thank Mrs. Seggebrock for assistance in data collection andreview of patients’ records and Mrs. Kramer for technicalassistance in collecting and analyzing blood samples. Drugpreparation and randomization were performed by the phar-macist, Mrs. Graf.

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