Monitoring activated clotting time for combined heparin and aprotinin application: in vivo evaluation of a new aprotinin-insensitive test using Sonoclot.

OBJECTIVE: Kaolin-based activated clotting time assessed by HEMOCHRON (HkACT) is a clinical standard for heparin monitoring alone and combined with aprotinin during cardiopulmonary bypass (CPB). However, aprotinin is known to prolong not only celite-based but also kaolin-based activated clotting time. Overestimation of activated clotting times implies a potential hazardous risk of subtherapeutic heparin anticoagulation. Recently, a novel 'aprotinin-insensitive' activated clotting time test has been developed for the SONOCLOT analyzer (SaiACT). The aim of our study was to evaluate SaiACT in patients undergoing CPB in presence of heparin and aprotinin. METHODS: Blood samples were taken from 44 elective cardiac surgery patients at the following measurement time points: baseline (T0); before CPB after heparinization (T1 and T2); on CPB, before administration of aprotinin (T3); 15, 30, and 60 min on CPB after administration of aprotinin (T4, T5, and T6); after protamine infusion (T7). On each measurement time point, activated clotting time was assessed with HkACT and SaiACT, both in duplicate. Furthermore, the rate of factor Xa inhibition and antithrombin concentration were measured. Statistical analysis was done using Bland and Altman analysis, Pearson's correlation, and ANOVA with post hoc Bonferroni-Dunn correction. RESULTS: Monitoring anticoagulation with SaiACT showed reliable readings. Compared to the established HkACT, SaiACT values were lower at all measurement time points. On CPB but before administration of aprotinin (T3), SaiACT values (mean+/-SD) were 44+/-118 s lower compared to HkACT. However, the difference between the two measurement techniques increased significantly on CPB after aprotinin administration (T4-T6; 89+/-152 s, P=0.032). Correlation of ACT measurements with anti-Xa activity was unchanged for SaiACT before and after aprotinin administration (r2=0.473 and 0.487, respectively; P=0.794), but was lower for HkACT after aprotinin administration (r2=0.481 and 0.361, respectively; P=0.041). On CPB after administration of aprotinin, 96% of all ACT values were classified as therapeutic by HkACT, but only 86% of all values were classified therapeutic if ACT was determined by SaiACT. Test variability was comparable for SaiACT and HkACT. CONCLUSIONS: The use of SaiACT may result in more consistent heparin management that is less affected by aprotinin and a corresponding increase in heparin administration for patients receiving aprotinin.     

Effects of heparin, haemodilution and aprotinin on kaolin-based activated clotting time: in vitro comparison of two different point of care devices

BACKGROUND: During cardiopulmonary bypass (CPB), measurement of kaolin-based activated clotting time (kACT) is a standard practice in monitoring heparin-induced anticoagulation. Despite the fact that the kACT test from the Sonoclot Analyzer (SkACT) has been commercially available for several years, no published data on the performance of SkACT are available. Thus, the aim of this in vitro study was to compare SkACT with an established kACT from Hemochron (HkACT). METHODS: Blood was withdrawn from 25 patients before elective cardiac surgery. SkACT and HkACT were measured in duplicate after in vitro administration of heparin (0, 1, 2 and 3 U/ml), calcium-free lactated Ringer's solution (25% and 50% haemodilution) and aprotinin (200 kIU/ml). RESULTS: A total of 600 duplicate kACT measurements were obtained from 25 cardiac surgery patients. Overall, mean bias +/- SD between SkACT and HkACT was 7 +/- 70 s (1.3% +/- 14.1%). Administration of heparin, haemodilution and aprotinin induced a comparable effect on both activated clotting time (ACT) tests. Mean bias ranged from -4 +/- 39 s (-1.7% +/- 12.9%) to 4 +/- 78 s (3.2% +/- 15.6%) for heparinzed blood samples after haemodilution or aprotinin application and increased after combined aprotinin administration and haemodilution. After haemodilution and administration of aprotinin, both ACT tests were less reliable for values >480 s in heparinized blood samples. CONCLUSION: Accuracy and performance of SkACT and HkACT were comparable after in vitro administration of heparin, aprotinin and haemodilution. Both ACT tests were considerably affected by aprotinin and haemodilution.     

Kaolin-based activated coagulation time measured by sonoclot in patients undergoing cardiopulmonary bypass

OBJECTIVES: In vivo data for the kaolin-based ACT test from the Sonoclot Analyzer (SkACT, Sienco Inc, Arvada, CO) are lacking. The aim of this study was to compare SkACT with an established kaolin-based ACT from Hemochron (HkACT) and anti-Xa activity in patients undergoing cardiopulmonary bypass (CPB). DESIGN: Prospective observational study. SETTING: Community hospital. PARTICIPANTS: Fifty patients scheduled for elective cardiac surgery. INTERVENTIONS: Blood samples were taken before CPB at baseline (T0) and after heparinization (T1 and T2), on CPB after administration of aprotinin (5, 15, 30, 60 minutes; T3-T6), and at the end after protamine infusion (T7). MEASUREMENTS AND MAIN RESULTS: A total of 375 blood samples were analyzed. ACT measurements were comparable for SkACT and HkACT at each measurement time point. Overall bias +/- standard deviation between SkACT and HkACT was -19 +/- 75 seconds (-2.4% +/- 11.7%). Mean bias between SkACT and HkACT at each time point ranged from -35 to 3 seconds (-4.5% to 2.6%) and showed no statistical significance over time. Heparin sensitivity of SkACT and HkACT, defined as (ACT(Tx)-ACT(T0))/(anti-Xa(Tx)-anti-Xa(T0)), significantly increased for measurements during CPB (p < 0.001) but without significant difference between the 2 methods. Test variability was comparable for both ACT measurement techniques. Overall test variability was 7.5% +/- 7.4% for SkACT and 7.8% +/- 11% for HkACT. CONCLUSIONS: Accuracy and performance of SkACT and HkACT were comparable for heparin monitoring in patients undergoing CPB for elective cardiac surgery. However, both tests were affected significantly after initiating CPB and aprotinin infusion.     

Aprotinin does not prolong the Sonoclot aprotinin-insensitive activated clotting time

STUDY OBJECTIVE: To determine whether a new Sonoclot-based, aprotinin-insensitive activated clotting time (aiACT) assay yields stable results over a broad range of aprotinin concentrations. DESIGN: Prospective trial conducted on in vitro blood samples. SETTING: Tertiary-care teaching medical center. PARTICIPANTS: 19 healthy adult volunteers. INTERVENTIONS: Whole blood samples were collected from volunteers. Heparin (2 U/mL) and escalating concentrations of aprotinin of 160 to 500 kallikrein inhibitory units (KIU)/mL were added in vitro. MEASUREMENTS AND MAIN RESULTS: Celite ACT, kaolin ACT, and aiACT assays were completed. The aiACT showed stable activated clotting time (ACT) results on heparinized, noncitrated blood with added aprotinin (P = nonsignificant). In contrast, celite ACT and kaolin ACT were greatly prolonged when aprotinin was added to heparinized, noncitrated, and citrated blood (P < 0.05). The aiACT had consistent results at all aprotinin concentrations (P = nonsignificant). CONCLUSIONS: Aprotinin (160, 320, and 500 KIU/mL) significantly prolongs the ACT value with celite and kaolin activators but not with the aprotinin-insensitive activator.     

The in vitro effects of aprotinin on twelve different ACT tests

Aprotinin is frequently used during CPB to reduce post-operative bleeding and attenuate the inflammatory response. The level of anticoagulation in these patients is monitored by using various activated clotting time (ACT) tests, which are generally accepted as being altered by the presence of aprotinin in blood. Therefore, we have investigated the effect of aprotinin on several ACT tests using whole blood from CPB patients. With IRB approval, blood samples were collected from patients undergoing CPB before and after full heparinization (300 u/kg). Each blood sample was divided into two aliquots, and aprotinin was added to one of them to yield a final calculated concentration of 300 KIU/mL. Both aliquots were used simultaneously to perform the 12 ACT tests. A paired Student's t-test was performed on the data. Overall, test results from 9 of 12 devices were significantly increased by aprotinin. Of these, four were increased only when the sample was heparinized, three were elevated by both heparinized and unheparinized blood, and two were elevated only when the sample was unheparinized. Each affected test responded uniquely to aprotinin, producing ACT test results ranging from 12 to 51% above nonaprotinized values. Several tests that were affected by aprotinin using heparinized blood samples were unaffected using unheparinized blood samples. These data emphasizes the unique manner in which individual ACT tests respond to aprotinized blood samples and should be considered when developing institutional policy for anticoagulation of aprotinized patients.     

The relationship between hirudin and activated clotting time: implications for patients with heparin-induced thrombocytopenia undergoing cardiac surgery.

Anticoagulation with recombinant hirudin (r-hirudin) (Refludan) has been suggested as an alternative to heparin for patients with heparin-induced thrombocytopenia requiring cardiac surgery. We sought to develop a modified activated coagulation time (ACT) that would allow quantification of the levels of r-hirudin required during cardiopulmonary bypass (CPB). Twenty-one patients scheduled for elective cardiac surgical procedures requiring CPB were enrolled in this IRB-approved study. R-hirudin was added to blood specimens obtained before heparin administration (before CPB) and 30 min after heparin neutralization with protamine (after CPB) to result in concentrations of 0, 2, 4, 6, 7, or 8 microg/mL. Kaolin/ACT and complete blood count measurements were assayed in native specimens (first 10 patients, Phase I) or in specimens mixed with equal volumes of commercial normal plasma (second 11 patients, Phase II). In Phase I, good (r(2) = 0.83) linear relationships between ACT values and r-hirudin concentrations (< or =4 microg/mL) were observed in specimens obtained before CPB. However, ACT values were markedly prolonged (P < 0.0001) by r-hirudin in specimens obtained after CPB, with ACT values generally exceeding the ACT's detection limit (>999 s) at hirudin concentrations >2 microg/mL. In patient specimens mixed with normal plasma (Phase II), ACT/hirudin relationships (i.e., hirudin/ACT slope values obtained with hirudin concentration < or =4 microg/mL) in the post-CPB period (0.022 +/- 0.004 microg. mL(-1). s(-1)) were similar (P = 0.47) to those (0.019 +/- 0.004 microg. mL(-1). s(-1)) obtained in the pre-CPB period. Accordingly, a significant relationship between normal plasma-supplemented ACT values and predilution hirudin concentration was obtained in the post-CPB (hirudin = 0.039ACT - 4.34, r(2) = 0.91) period. Although our data demonstrate that the ACT test cannot be used to monitor hirudin during CPB, the addition of 50% normal plasma to post-CPB hemodiluted blood specimens yields a consistent linear relationship between hirudin concentration and ACT values up to a predilution concentration of 8 microg/mL. Plasma-modified ACT may be useful in monitoring hirudin anticoagulation during CPB. Implications: A modified activated clotting time test system that may be helpful in monitoring hirudin anticoagulation in patients with heparin-induced thrombocytopenia during cardiac surgery with cardiopulmonary bypass is described.     

The in vitro effects of antithrombin III on the activated coagulation time in patients on heparin therapy

Heparin requires antithrombin III (AT) to achieve anticoagulation, and patients on continuous small-dose heparin preoperatively experience decreased levels of AT-causing heparin resistance. When this occurs, 2-4 units of fresh frozen plasma ( approximately 1000 units of AT) are often administered to increase AT levels and restore heparin responsiveness. We evaluated purified human AT concentrate (Thrombate III; Bayer, Inc., Elkhart, IN) to restore in vitro anticoagulation responses in patients receiving heparin. Blood samples were obtained from cardiac surgery patients including 22 patients receiving heparin and 21 patients not receiving heparin preoperatively. Heparin was added to blood in final concentrations of 4.1, 5.4, and 6.8 U/mL (equivalent to 300, 400, and 500 U/kg), and kaolin-activated clotting times (ACTs) were determined with and without AT at a final concentration of 0.2 units/mL to mimic fresh frozen plasma administration. The mean duration of preoperative heparin therapy was 4.0 days (range 2-10 days). AT activity was 69% +/- 9% in patients receiving heparin and 92% +/- 8% in patients not receiving heparin (P < 0.01). Heparin >4.1 U/mL failed to further increase ACT values in all patients. Attempts to increase ACT in patients receiving heparin may require supplemental AT administration. Purified AT even in small doses significantly prolongs the ACT response to heparin. Implications: In vitro addition of antithrombin III (0.2 U/mL) to heparinized blood samples (4.1-6.8 units of heparin/mL) from patients on previous heparin therapy increases sensitivity to supplemental heparin as reflected by significantly prolonged activated clotting time.     

A quick assay for monitoring recombinant hirudin during cardiopulmonary bypass in patients with heparin-induced thrombocytopenia type II : adaptation of the ecarin clotting time to the act II device

BACKGROUND: Recombinant hirudin is increasingly advocated as a promising alternative anticoagulation for patients with heparin-induced thrombocytopenia type II during cardiopulmonary bypass. This requires monitoring of the ecarin clotting time. No commercial ecarin clotting time assay is available for clinical use. We adapted the ecarin clotting time to the easy-to-handle ACT II device. METHODS: Three different concentrations of the ecarin reagent (20, 10, 5 U/mL) were investigated as preliminary studies. Standard calibration curves were constructed for concentrations of recombinant hirudin ranging from 0 to 5 microgram/mL. In vivo samples were collected from patients with heparin-induced thrombocytopenia type II who underwent cardiopulmonary bypass, and the values were compared with the values obtained by the chromogenic method. The final concentration for the assay of 5 IU/mL ecarin was further assessed in vitro for reproducibility and the influence of variations in hematocrit, platelet count, and procoagulants. RESULTS: All three concentrations of ecarin revealed linearity to 5 microgram/mL concentrations of recombinant hirudin. The ecarin concentration of 5 U/mL revealed the best correlation (0.87) to the laboratory method, was reproducible over the whole recombinant hirudin range, and was not influenced by the variations in the in vitro setup. CONCLUSIONS: The ACT II/ecarin clotting time with an ecarin concentration of 5 U/mL is a simple and reliable assay for monitoring recombinant hirudin during cardiopulmonary bypass. Use of this assay allows a wider use of recombinant hirudin in patients with heparin-induced thrombocytopenia type II during bypass and thereby may contribute to the safer management of these patients.     

Does Hemodilution by the Crystalloid Priming Solution Derange the Efficacy of Anticoagulation During Cardiopulmonary Bypass?

BACKGROUND AND AIM: Recent studies suggest the development of a procoagulant state with hemodilution. We conducted this study to investigate the effect of hemodilution, by the priming solution in a cardiopulmonary bypass (CPB) circuit, on "point of care" coagulation assays (activated clotting time [ACT] and thromboelastography [TEG]). METHODS: Twenty patients undergoing cardiac surgery with crystalloid priming of CPB circuit were evaluated. Confounding variables arising from contact activation were eliminated by minor modifications. Ten milliliter per kilogram body weight of priming solution (lactated Ringer's) was infused via the aortic cannula. ACT and TEG were performed, both prior to and immediately after hemodilution. In case of latter, four variables, reaction time (r), coagulation time (k), maximum amplitude (MA), and clot formation rate (angle alpha), were estimated and considered for the results. To see if these results are duplicated "in vitro," prebypass blood samples from eight heparinized patients, diluted (4:1) with priming solution from the venous reservoir, were also analyzed. RESULTS: Falls in ACT, from a mean of 659.7 (+/-260.6) seconds to 251.5 (+/-103.2) seconds (p < 0.01), r time (678.1 [+/-318.1] sec to 468.7 [+/-152.7] sec) (p < 0.01), and k time (211.7 [+/-161.7] sec to 123.8 [+/-32.1] sec) (p < 0.05) on TEG were noted upon hemodilution. Angle alpha and MA increased, but were not statistically significant. Results from the in vitro study closely matched the results from our in vivo analysis. CONCLUSION: The study suggests that hemodilution by crystalloid priming solution may impair the efficacy of anticoagulation during CPB. The mechanism for this phenomenon remains to be elucidated.     

Monitoring of heparin-induced anticoagulation with kaolin-activated clotting time in cardiac surgical patients treated with aprotinin.

High-dose aprotinin appears to enhance the anticoagulant effects of heparin, as documented by increases in the activated clotting times (ACTs) during cardiopulmonary bypass; hence, some authorities have advocated reducing the dose of heparin in patients treated with aprotinin. An in vitro study by our group suggested that the increase of the ACT in the presence of aprotinin and heparin may be due to the use of celite as surface activator. We compared celite and kaolin as surface activators for the measurement of the ACT in cardiac surgical patients treated with aprotinin and in patients given no aprotinin. This double-blind, randomized, placebo-controlled study included 30 patients, of whom 14 received aprotinin and 16 received a placebo. Before, during, and after cardiopulmonary bypass, the ACT was measured with two Hemochron 400 systems with 12 mg of either celite (C-ACT) or kaolin (K-ACT) used as surface activator and with one Hepcon HMS system (HR-ACT), which uses kaolin as activator. The latter also was used for measurement of the blood heparin concentration. The ACTs of blood without heparin did not differ between aprotinin and control patients. During anticoagulation with heparin and cardiopulmonary bypass, the average C-ACTs were 784 +/- 301 s (aprotinin) and 496 +/- 120 s (control) (P < .001); the K-ACTs were 502 +/- 131 s (aprotinin) and 458 +/- 101 s (control) (P > .05); the HR-ACTs were 406 +/- 87 s (aprotinin) and 423 +/- 82 s (control) (P > .05), which was consistently less than C-ACT and K-ACT.(ABSTRACT TRUNCATED AT 250 WORDS)     

A novel thrombelastograph tissue factor/kaolin assay of activated clotting times for monitoring heparin anticoagulation during cardiopulmonary bypass

We used a thrombelastograph (TEG) assay with tissue factor and kaolin (TEG TF/K) to measure activated clotting time (ACT) in 31 patients during cardiopulmonary bypass. For comparison, ACTs were also determined by a Hemochron Jr. Signature and a Hepcon HMS. The TEG TF/K correlated with both the Hepcon (r(2) = 0.789) and Hemochron (r(2) = 0.743) ACTs. The average ACT after heparin was 319 +/- 119 s (mean +/- sd) for the TEG TF/K compared with 624 +/- 118 s for the Hepcon instrument. To evaluate the effects of hemodilution on TEG TF/K and Hemochron assays, ACT assays were performed on blood diluted to 50% and titrated with heparin from 0 to 6 U/mL. Both instruments showed significant (P < 0.01) changes in the ACT-versus-heparin slope, but the 0 heparin intercept for the TEG TF/K ACTs was not significantly changed (P = 0.292), in contrast to that for the Hemochron device (P = 0.041). Both instruments also indicated the same 1.3:1 ratio of protamine to heparin for optimum heparin neutralization, with increasing ACTs at ratios >2.6:1. The TEG TF/K ACT assay rapidly monitors heparin anticoagulation, in addition to the capabilities of this instrument to monitor platelet function, clotting factors, and fibrinolysis.     

A quick anti-Xa-activity-based whole blood coagulation assay for monitoring unfractionated heparin during cardiopulmonary bypass: a pilot investigation

We developed a quick and easy method to perform anti-Xa-activity-based whole blood assay and assessed its reliability for online monitoring of unfractionated heparins (UFHs) during cardiopulmonary bypass. Seventy-five microliters of a mixture of 1:3 large- and small-range Heptest reagent were transferred into blank cartridges of the ACT II device. The plastic flags for clot detection and stirring the sample and reagent were inserted and overlaid with 75 microL of Recalmix for recalcification. One-hundred fifty microliters of citrated whole blood were added and measurements performed. In vitro, the linearity of the test over a range of 1-8 IU/mL UFH, as well as the influence of variations in hematocrit (60%, 30%, and 20%), plasma coagulation factors (50%, 30%, and 20%) and platelets (100, 50, and 20 x 10(3)/microL) on the test results were assessed. In vivo measurements performed during cardiopulmonary bypass were compared with the chromogenic assay. The test revealed linearity to concentrations of 6 IU/mL of UFH and was not significantly influenced by the variations in the in vitro set-up despite a prolongation in samples with a hematocrit of 60%. In vivo, the correlation to the chromogenic test was R: = 0.90. The ACT II anti-Xa-UFH assay performed in whole blood was reliable when used over a wide range of conditions that could be encountered clinically. Although the test is useful for point-of-care monitoring, the necessity of individual calibrations and pipetting in the operation room requires further automation before its use in clinical practice. IMPLICATIONS: The ACT II anti-Xa-unfractionated heparin assay allows for reliable monitoring of large concentrations of UFH over a wide range of hematocrit, platelet, and coagulation factor levels. Further evaluation of this point-of-care device is indicated.     

Factor Xa-activated whole blood clotting time (Xa-ACT) for bedside monitoring of dalteparin anticoagulation during haemodialysis.

BACKGROUND: Low molecular weight heparins (LMWH) like dalteparin are increasingly used for anticoagulation during haemodialysis (HD). The available laboratory tests for monitoring LMWH anticoagulation are time-consuming and expensive, and the suitability of the conventional activated clotting time (ACT) is controversial. A simple and cheap bedside test would be useful. METHODS: We studied the factor Xa-activated whole blood clotting time (Xa-ACT) in vitro and in vivo in nine patients undergoing chronic HD with i.v. dalteparin bolus anticoagulation and compared it with the conventional ACT. Plasma anti-factor Xa (antiXa) activity was determined with a chromogenic assay. Thrombin-antithrombin complexes were measured to detect coagulation activation. RESULTS: Xa-ACT and ACT were prolonged with rising dalteparin concentration. In vitro, both clotting times were strongly correlated with the antiXa levels (r = 0.94 and 0.89, respectively). Nevertheless, compared with the ACT, the Xa-ACT was considerably more sensitive to the LMWH in vitro (healthy blood: Xa-ACT 90 s/U vs ACT 26 s/U; uraemic blood: Xa-ACT 96 s/U vs ACT 31 s/U) as well as in vivo (Xa-ACT 81 s/U vs ACT 22 s/U) and reflected different intensities of anticoagulation. An initial dalteparin bolus of 80+/-11 U/kg body weight was able to prevent coagulation activation for up to 4 h of HD. CONCLUSION: For monitoring LMWH anticoagulation the Xa-ACT was superior to the conventional ACT in vitro as well as in vivo during HD. The Xa-ACT can be useful as a LMWH bedside test. The ACT was not sensitive enough to serve as a LMWH monitoring tool.     

在体外及体外循环中抑肽酶对ACT的影响

选择健康献血员及心内直视手术病，观察抑肽酶对全血活化凝血时间的影响。结果：在体外肝素剂量与ＡＣＴ有显著线性相关。抑肽酶单狡应用并不使ＡＣＴ延，但与肝素合用可协同性延长ＡＣＴ值；在体外循环中抑肽酶延长ＡＣＴ的值更为显著，一般超过８００ｓ。结论：抑肽酶可与肝素协同性延长ＡＣＴ，体外循环中应用抑肽酶时应以ＡＣＴ大于８００ｓ作为肝素抗凝标准。     

In vitro effects of aprotinin on activated clotting time measured with different activators.

The effects in vitro of aprotinin on the activated clotting time measured with both celite- and kaolin-activated tubes were investigated in 21 consecutive patients requiring cardiopulmonary bypass. Four whole-blood samples (2 ml per sample) from each patient were tested simultaneously with Hemochron automated timing systems (International Technidyne Corp., Edison, N.J.) before, during, and after cardiopulmonary bypass. One tenth milliliter of either aprotinin (at a final concentration of 80, 120, or 180 KIU/ml) or saline solution was mixed in vitro with blood samples before determination of the activated clotting time. Aprotinin had no inhibitory effect on the activated clotting times of unheparinized blood. After heparin administration, aprotinin in the above concentrations prolonged the activated clotting times measured with celite-activated tubes by 47% to 71%, as compared with the measurements of the activated clotting time without the addition of aprotinin. The activated clotting times in kaolin-activated tubes were not increased, however, by the in vitro addition of aprotinin. Our in vitro results indicate that aprotinin in concentrations from 80 to 180 KIU/ml does not significantly enhance the inhibitory effects of heparin on the intrinsic coagulation system as evaluated by measurement of the activated clotting times in kaolin-activated tubes. The anticoagulation effect of heparin in patients receiving aprotinin infusion should be monitored with kaolin-activated instead of celite-activated tubes because the celite makes the measured activated clotting time unreliable in patients receiving aprotinin therapy. These in vitro results require confirmation in vivo in patients receiving aprotinin therapy.     

The effects of aprotonin on platelets in vitro using whole blood flow cytometry

We sought to evaluate the effects of aprotinin on the number and function of the platelet glycoprotein (GP) IIb-IIIa receptor and on the expression of P-selectin in vitro in order to gain insight into the potential mechanisms involved in the platelet-protective action of aprotinin during cardiopulmonary bypass. Aprotinin at 50 to 200 kallikrein inhibiting units/mL decreased the expression of activated GP IIb-IIIa complex in response to adenosine diphosphate or thrombin receptor activator peptide 6 in a dose-dependent manner in both citrated and heparinized whole blood experiments. Aprotinin inhibited adenosine diphosphate-induced platelet aggregation, but it exhibited no effect on the expression of GP IIIa and P-selectin. These results indicate that aprotinin interferes with the platelet fibrinogen receptor function during pharmacological activation. Reduced aggregability and platelet adhesion to fibrinogen adsorbed to synthetic surfaces in the presence of aprotinin may prevent platelet consumption during clinical cardiopulmonary bypass. This in vitro study demonstrates that aprotinin decreases the agonist-induced expression of activated GP IIb-IIIa receptors that play a major role in platelet aggregation and adhesion to biomaterial surfaces. IMPLICATIONS: This in vitro study demonstrates that aprotinin decreases the agonist-induced expression of activated glycoprotein IIb-IIIa receptors that play a major role in platelet aggregation and adhesion to biomaterial surfaces.     

Reliability of the Heparin Management Test for Monitoring High Levels of Unfractionated Heparin: In Vitro Findings in Volunteers versus In Vivo Findings during Cardiopulmonary Bypass

Background: The authors assessed the heparin management test in vitro in volunteers and in vivo during cardiopulmonary bypass.

Methods: In vitro, the heparin management test was analyzed for heparin levels between 0 and 6 IU/ml using variations in hematocrit, platelets, procoagulants, and storage time. The in vivo studies consisted of two groups: In group I (cardiopulmonary bypass <= 90 min, n = 40), anticoagulation was performed according to the activated clotting time (with or without aprotinin); in group II (cardiopulmonary bypass >= 180 min, with aprotinin) included use (n = 10) and nonuse of coumadin (n = 10) and anticoagulation according to the automated heparin dose-response assay. Tests were performed in duplicate (whole blood, two heparin management test analyzers) and compared with anti-Xa activity (plasma).

Results: In vitro, the results of the heparin management test (n = 1,070) correlated well with heparin concentration (r2 = 0.98). Dilution and storage time did not affect the heparin management test; a hematocrit of 60% and reduced procoagulants (10%) prolonged clotting time. In vivo, the correlation (heparin management test vs. anti-Xa) was strong in group I (r2 = 0.97 [with aprotinin] and 0.96 [without aprotinin]; n = 960) and group II without coumadin (r2 = 0.89, n = 516). In group II with coumadin, the overall correlation was r2 = 0.87 and 0.79 (n = 484), although the range varied widely (0.57-0.94, between-analyzer differences 0-47%).     

An investigation of a new activated clotting time "MAX-ACT" in patients undergoing extracorporeal circulation

Activated clotting time (ACT) is a test used in the operating room for monitoring heparin effect. However, ACT does not correlate with heparin levels because of its lack of specificity for heparin and its variability during hypothermia and hemodilution on cardiopulmonary bypass (CPB). A modified ACT using maximal activation of Factor XII, MAX-ACT (Actalyke MAX-ACT; Array Medical, Somerville, NJ), may be less variable and more closely related to heparin levels. We compared MAX-ACT with ACT in 27 patients undergoing CPB. We measured ACT, MAX-ACT, temperature, and hematocrit at six time points: baseline; postheparin; on CPB 30, 60, and 90 min; and postprotamine. Additionally, we assessed anti-Factor Xa heparin activity and antithrombin III activity at four of these six time points. With institution of CPB and hemodilution, MAX-ACT and ACT did not change significantly but had a tendency to increase, whereas concomitant heparin levels decreased (P = 0.065). Neither test correlated with heparin levels. ACT and MAX-ACT did not differ during normothermia but did during hypothermia, and ACT was significantly longer than MAX-ACT (P = 0.009). At the postheparin time point, ACT-heparin sensitivity (defined as [ACT postheparin - ACT baseline]/[heparin concentration postheparin - heparin concentration baseline]) was greater than MAX-ACT-heparin sensitivity (analogous calculation for MAX-ACT; 520 [266 - 9366] s. U(-1). mL(-1) vs 468 [203 - 8833] s. U(-1). mL(-1); P = 0.022). IMPLICATIONS: MAX-ACT (a new activated clotting time [ACT] test) uses more maximal clotting activation in vitro and, although it is less susceptible to increase because of hypothermia and hemodilution than ACT, lack of correlation with heparin levels remains a persistent limitation.     

Platelet protection by aprotinin in cardiopulmonary bypass: electron microscopic study.

To evaluate the functional integrity of platelets in patients administered the proteinase inhibitor aprotinin during cardiopulmonary bypass, 20 patients undergoing a complicated and prolonged open heart operation were studied. They were randomized to receive either a high dose of aprotinin (total dose, 6 to 7 x 10(6) KIU) before and during cardiopulmonary bypass (10 patients) or a placebo (10 patients). Blood samples were collected preoperatively, at the termination of bypass, and 90 minutes thereafter to assess platelet count and aggregation on extracellular matrix, which was studied by scanning electron microscopy. On a scale of 1 to 4, mean preoperative platelet aggregation grades were similar in both groups (3.5 +/- 0.5). Postoperatively, at the termination of cardiopulmonary bypass and 90 minutes thereafter, all 10 patients treated with aprotinin revealed normal, unchanged platelet aggregation (grade, 3.5 +/- 0.5), whereas all placebo-treated patients showed severely disturbed aggregation (grade, 1.4 +/- 0.5) (p less than 0.001). The platelet count was similar in both groups before and after operation (preoperatively, 182 +/- 75 x 10(9)/L and 146 +/- 30 x 10(9)/L, and postoperatively, 87 +/- 13 x 10(9)/L and 80 +/- 27 x 10(9)/L for the aprotinin and placebo groups, respectively). Total 24-hour postoperative bleeding and blood requirement were significantly lower in the aprotinin group (371 +/- 84 mL and 2 +/- 0.7 units, respectively) compared with the placebo group (608 +/- 28 mL and 3.4 +/- 1.3 units, respectively) (p less than 0.01). These results demonstrate that improved postoperative hemostasis is directly related to the complete preservation of platelet function achieved by the protective properties of aprotinin.     

Influence of high-dose aprotinin treatment on blood loss and coagulation patterns in patients undergoing myocardial revascularization.

Intraoperative administration of the proteinase inhibitor aprotinin causes reduction in blood loss and homologous blood requirement in patients undergoing cardiac surgery. To ascertain the blood-saving effect of aprotinin and to obtain further information about the mode of action, 40 patients undergoing primary myocardial revascularization were randomly assigned to receive either aprotinin or placebo treatment. Aprotinin was given as a bolus of 2 x 10(6) kallikrein inactivator units (KIU) before surgery followed by a continuous infusion of 5 x 10(5) KIU/h during surgery. Additionally, 2 x 10(6) KIU were added to the pump prime. Strict criteria were used to obtain a homogeneous patient selection. Total blood loss was reduced from 1,431 +/- 760 ml in the control group to 738 +/- 411 ml in the aprotinin group (P less than 0.05) and the homologous blood requirement from 838 +/- 963 ml to 163 +/- 308 ml (P less than 0.05). In the control group, 2.3 +/- 2.2 U of homologous blood or blood products were given, and in the aprotinin group, 0.63 +/- 0.96 U were given (P less than 0.05). Twenty-five percent of patients in the control group and 63% in the aprotinin group did not receive banked blood or homologous blood products. The activated clotting time as an indicator of inhibition of the contact phase of coagulation was significantly increased before heparinization in the aprotinin group (141 +/- 13 s vs. 122 +/- 25 s) and remained significantly increased until heparin was neutralized after cardiopulmonary bypass (CPB).(ABSTRACT TRUNCATED AT 250 WORDS)