Anesth Analg 2004;99:120-123
© 2004 International Anesthesia Research Society
doi: 10.1213/01.ANE.0000123012.24871.62
TECHNOLOGY, COMPUTING, AND SIMULATION
Steven J. Barker Section Editor
The Impact of Factor XIII on Coagulation Kinetics and Clot Strength Determined by Thrombelastography
Vance G. Nielsen, MD,
William Q. Gurley, Jr, MD, and
Thomas M. Burch, MD
Department of Anesthesiology, The University of Alabama at Birmingham
Address correspondence and reprint requests to Vance G. Nielsen, MD, Department of Anesthesiology, The University of Alabama at Birmingham, 619 S. 19th St., Birmingham, AL 35249-6810. Address e-mail to vance.nielsen{at}ccc.uab.edu
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Abstract
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Fibrinogen has been shown to be responsible for most protein-mediated clot strength via thrombelastography. However, factor XIII (FXIII) activity also plays a prominent role in the development of clot strength. Thus, we hypothesized that changes in FXIII activity would significantly increase clot strength. FXIII (0%, 1%, 6.25%, 12.5%, 25%, 50%, and 100% normal activity) was placed in a fixed volume of citrated FXIII-deficient plasma with 1% tissue factor and calcium chloride and underwent thrombelastography for 10 min. We measured the variables reaction time (R; a measurement of clot initiation), 
(a measure of the rate of clot formation), amplitude (A; a measure of clot strength), and shear elastic modulus (G; a measure of clot strength). FXIII activity significantly decreased R in a pattern of exponential decay (R2 = 0.77; P < 0.001). FXIII activity significantly increased , following a sigmoidal pattern (R2 = 0.88; P < 0.001). Finally, increases in FXIII activity significantly increased A and G in a sigmoidal pattern (R2 = 0.89; P < 0.001). We concluded that FXIII significantly affects R, alpha, A, and G. Thus, transfusion decision making with protein-mediated thrombelastographic patterns must account for the contribution of both fibrinogen and FXIII.
IMPLICATIONS: Changes in fibrinogen concentration have been implicated as the primary determinant of protein-mediated clot strength via thrombelastography. We determined that factor XIII (FXIII) activity significantly enhanced clot strength and speed of clot initiation and formation. Transfusion decision making with thrombelastography must account for the contribution of both fibrinogen and FXIII.
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Introduction
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Thrombelastography (TEG®) is used to assess coagulopathy in the settings of cardiac surgery (1,2), liver transplantation (3–5), and other procedures (6). Modifications of TEG® methodology can query specific components of the coagulation system. In particular, either the separation of platelets from plasma (7) or the inhibition of platelet effects via glycoprotein IIb/IIIa blockade (1,8) has been performed to assess coagulation protein-mediated phenomena via TEG®. A linear relationship between clot strength and fibrinogen concentration in platelet-inhibited whole blood (R2 = 0.80) (1) or purified fibrinogen solutions (R2 = 0.94) (8) has been demonstrated. Given these relationships, perhaps platelet-inhibited TEG® could serve as a surrogate for fibrinogen determinations.
Although fibrinogen concentration has been correlated with clot strength via TEG®, factor XIII (FXIII) also modulates clot strength (9–11). Further, fibrinogen concentration and FXIII activity, but not thrombin activity, significantly increase fibrin sealant strength in vitro (12). FXIII is a 320-kd tetramer, and when activated by thrombin, it covalently cross-links the glutamine and lysine residues of forming fibrin strands and thus increases clot strength and stability (13). FXIII activities as low as 1%–2% of normal can be sufficient for hemostasis in patients with congenital FXIII deficiency (13), a disorder associated with hemorrhagic complications such as intracranial hematoma. Thus, perhaps the amount of FXIII activity present could influence TEG® variables in human plasma.
The purpose of this study was to characterize the effect of FXIII on coagulation kinetics and clot strength via TEG®. This goal was achieved by the addition of various activities of purified FXIII to FXIII-deficient plasma in vitro.
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Methods
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Serial dilutions of FXIII obtained from one lot (Enzyme Research Laboratories, South Bend, IN) in 50 mM Tris buffer (pH 7.4) were made so that a final concentration of 0%, 1%, 6.25%, 12.5%, 25%, 50%, and 100% activity was present when 20 µL of FXIII solution was added to 340 µL of other constituents in the TEG® reaction solution (n = 8 per concentration). Citrated, FXIII-deficient plasma (<1% activity; George King Bio-Medical, Overland Park, KS) was obtained from one lot procured from one donor. This plasma was considered to have an FXIII activity of 0%. With the exception of FXIII, all other procoagulant and anticoagulant activities were within the normal range. Plasma (300 µL) was placed with 10 µL of tissue factor (final concentration, 1% of rabbit brain tissue factor; international sensitivity index, 1.07; Trinity Biotech, Ventura, CA), 20 µL of FXIII solution, and 30 µL of 200 mM CaCl2 into a disposable cup in a computer-controlled TEG® (Model 5000; Haemoscope Corp., Niles, IL). Control plasmas obtained from Haemoscope were used daily to verify TEG® function. The following variables were determined at 37°C: reaction time (R; minutes), a measure of clot initiation; angle (; degrees), a measure of the speed of clot formation; amplitude (A; millimeters), a measure of clot strength; and shear elastic modulus (G; dynes per square centimeter). G is a measure of clot strength calculated from A as follows:
The TEG® sample data were recorded for 10 min. A detailed description of the methodology of TEG® has been previously described (7). Finally, prothrombin time, activated partial thromboplastin time, and fibrinogen concentration of FXIII plasma were determined in duplicate with a coagulation analyzer (ACL 100/200; Instrumentation Laboratory, Lexington, MA).
Variables are expressed as mean ± SD. Analyses of the effects of different doses of FXIII on TEG® variables were conducted with one-way analysis of variance with the Holm-Sidak post hoc test for multiple comparisons. The correlation of changes in FXIII activity and TEG® variables was calculated with commercially available software (Origin 7.0; OriginLab Corp., Northampton, MA). A P value of <0.05 was considered significant.
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Results
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FXIII significantly enhanced R, , A, and G in an activity-dependent fashion (Table 1). With regard to R, the addition of FXIII significantly decreased time to R until activities were 25% or more. In contrast, increasing FXIII activity resulted in , A, and G values that were significantly different from one another throughout the dose range.
Changes in FXIII activity were significantly related in nonlinear fashions to R, , A, and G values (Fig. 1). Increasing FXIII activity resulted in an exponential decay of R values, whereas , A, and G values demonstrated a sigmoidal increase. The correlation coefficients and equations describing these relationships were significant (Table 2). Finally, the FXIII-deficient plasma had a prothrombin time of 12.3 ± 0.0 s (international normalized ratio of 1.01), an activated partial thromboplastin time of 33.3 ± 0.0 s (control, 27.9 s), and a fibrinogen concentration of 248 ± 6 mg/dL (reference range, 170–410 mg/dL).
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Figure 1. Correlations of factor XIII (FXIII) activity and thrombelastographic variables. R = reaction time, a measure of clot initiation; alpha = a measure of the speed of clot formation; amplitude = a measure of clot strength; G = shear elastic modulus, a measure of clot strength derived from amplitude.
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Discussion
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We found that in a human plasma system, wherein thrombin activity and fibrinogen concentration are a constant, FXIII activity significantly affects R, , A, and G, as measured by TEG®. Whereas R appeared to be maximally decreased after 25% of normal FXIII activity was present, all remaining variables continued to be significantly increased throughout the dose range of FXIII used. Further, increases in A observed in blood obtained from FXIII-deficient mice administered human FXIII (11) were consistent with the A and G dose-response relationship to FXIII activity in this study. Given that all TEG® variables are derived from an increase in viscoelastic resistance mediated by fibrin mesh formation, FXIII-mediated increases in fibrin cross-linking should significantly affect R, , A, and G.
Although this study demonstrates that FXIII activity significantly affects TEG® variables, caution should be observed in extrapolating our data to clinical situations. An important limitation of this study is that it was plasma based and devoid of fibrin-platelet interactions. Further, in the setting of massive intravascular volume resuscitation, the inevitable decrease of fibrinogen and thrombin in conjunction with a decrease in FXIII activity could have even more profound effects on TEG® variables than our data suggest. Despite these limitations, this study demonstrated that FXIII activity significantly affects all TEG® variables.
In conclusion, FXIII activity significantly affects not only A and G, but also the speed of R and . In clinical situations involving significant blood loss, changes in protein-mediated thrombelastographic patterns may be mediated by fluctuations of both fibrinogen concentration and FXIII activity. Thus, transfusion decision making (e.g., the administration of cryoprecipitate versus fresh frozen plasma) involving protein-mediated TEG® must account for the contribution of both fibrinogen and FXIII.
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Acknowledgments
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This investigation was supported by the Department of Anesthesiology.
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References
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Accepted for publication January 30, 2004.
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Abstract
Introduction
