Thromboelastography Applications in Feline Medicine
ACVIM 2008
Amy J. Alwood, DVM, DACVECC
Pittsburgh, PA, USA

Background

Thromboelastography (TEG) was developed in the late 1940's and early applications in human medicine centered upon "near patient" monitoring in the operating suite.1 A search of PubMed and MEDLINE databases identifies more than 2000 references for thromboelastography or thromboelastometry. In more recent years there has been expanded use for pharmaceutical monitoring and screening of patients for hemostatic disorders.

In addition to providing "point of care" testing, a major interest in use of TEG in veterinary patients has evolved because the global assessment of coagulation provides the only means to identify hypercoagulability. Dogs received more attention than cats in the earliest veterinary work with TEG. Work with TEG in dogs continues to be motivated by a desire to validate its use in identifying and studying hypercoagulable states in various naturally occurring disease states in the canine patient but has also been expanded to other applications.2-6 While thrombosis may be recognized less commonly in our feline patients, it would be anticipated that TEG could be applied to this species as well to identify hypercoagulable states and evaluate effects of antithrombotic therapies.

Initial work with TEG in cats at the University of Pennsylvania developed several years ago as part of a residency research project. The work evolved from an interest studying cardiogenic arterial thromboembolism (CATE) and a desire to evaluate the true efficacy of antithrombotic therapy (in particular, the use of heparin therapy). Ironically, this goal remains unfulfilled but the foundations have been laid with our initial work with TEG in normal cats and subsequent pilot data from cats with cardiomyopathy. A review of recently published work and a preliminary report of more recent (as yet unpublished) work will be the focus of these proceedings.

Brief Review of TEG Analysis

Standard thromboelastography can be performed with either fresh whole blood or citrated whole blood. When citrated whole blood is used (as is typically the case in veterinary medicine) calcium or a similar activator is added to the sampling cup immediately prior to initiation of analysis. The disposable sampling cup is brought into contact with a disposable pin which is connected to a torsion wire within the instrument. Once analysis is initiated the sampling cup is slowly oscillated by the instrument. As a clot begins to form the resultant tension or altered rotation is detected by the pin and torsion wire. As the clot develops this is translated via the computer connected in series to the Thromboelastograph analyzer and displays a representative tracing. The tracing begins as a straight line that then begins to separate into two parallel but divergent lines as the clot begins to form and develop. Several standard parameters have been defined based on this thromboelastogram. Principal ones utilized in the early veterinary work investigating TEG changes associated with hypercoagulable conditions include: 1) the R value or R-time, which is the time (in minutes) from initiating analysis to the beginning of clot development where the tracing just begins to split or diverge; 2) the K value or K-time, which is the time from the end of R to a standard separation (20 mm) of the arms of the tracing; 3) the alpha (α) or alpha-angle, which is the slope of the line connecting R and K; and 4) the maximum amplitude or MA, which is the height of the tracing (distance in mm between the divergent arms of the tracing). For a more detailed discussion of TEG analysis, please refer to the related Proceedings or to one of the excellent review articles.1,3

Validation of TEG in Clinically Normal Cats

While fresh whole blood can be used for TEG analysis, the need to initiate analysis within minutes and the fact that companion animals tend to initiate coagulation much more rapidly than humans, makes use of fresh whole blood samples impractical for most clinical or experimental veterinary work. Validation of TEG in healthy privately-owned cats was performed on citrated whole blood using a standardized technique that was similar to the previous validation in dogs.2-4,7 Feline blood samples (2 mL) were collected and placed into standard Vacutainer tubes containing 3.8% sodium citrate (in 9:1 ratio). Samples were held at room temperature for 30 minutes prior to analysis. Each sample was recalcified using 0.2 M calcium chloride immediately prior to analysis by the Haemoscope 5000 Thromboelastograph. Four parameters from each thromboelastogram (n=25 duplicate analyses) were recorded: R, K, α-angle, and MA or maximum amplitude.

Suggested reference ranges for each parameter were established based on the 5th and 95th percentiles for each variable and resulted in the following: R, 1.5-4.4 minutes; K, 1.0-2.8 minutes; α-angle, 59.2-79.8; MA, 46.0-69.2 mm. Compared to dogs (and humans), the normal hemostatic profile of cats was relatively hypercoagulable (*p<0.001) when each of the 4 TEG parameters was evaluated.

Table 1. Comparative TEG parameters.

TEG parameter

Feline

Canine

Human

R (min)

1.5-4.4*

5.0-7.4*

9-27

K (min)

1.0-2.8*

2.7-4.3*

2-9

α (degrees)

59.2-79.8*

44-56*

22-58

MA (mm)

46-69.2

54-61

44-64

daggerAs with any laboratory/coagulation analysis, it is advised that each institution/laboratory establish an independent reference range prior to implementing experimental or clinical use of TEG analysis.

Utility of TEG to Evaluate the Effects of Heparin Administration in an Experimental Colony

In addition to its value as a more comprehensive tool to evaluate global hemostasis [TEG analysis incorporates all phases of hemostasis from initiation to fibrinolysis and also incorporates many components of hemostasis including clotting factors, inhibitors, platelets and other cellular influences]; TEG has additional merit as it may identify hypocoagulable effects when standard coagulation times are still found to be within reference ranges. A series of experimental studies were performed administering heparin to clinically normal cats in a colony setting. Standard coagulation tests, anti-Xa analysis (as a standard measure of inhibition of Factor Xa) and TEG were performed. The initial study has been published; anti-Xa data from the companion study has been presented in abstract form.7,8

In the first study, a group of five clinically normal cats received unfractionated heparin (UFH) and the low molecular weight heparins dalteparin and enoxaparin (LMWH) serially in a cross-over study design. Heparin therapy consistently resulted in hypocoagulable TEG tracings. It should be noted, however, that at traditional doses (250 IU/kg) of UFH there is a profound effect on TEG. At peak drug effect with UFH the TEG tracing never develops the characteristic divergence and as such the R value never reaches an endpoint (our laboratory typically assigns a default R value of >30 minutes). Hence, the value of TEG analyses as a monitor of standard UFH therapy may be limited, however its use a monitoring tool for LMWH may be more applicable to clinical settings as a hypocoagulable effect was appreciated.7,8

Because LMWH dosages used in the first study did not result in significant Xa inhibition, in subsequent studies higher doses of LMWH were administered to a larger group of normal cats (also in a colony setting). Alterations of TEG were consistently seen two hours after subcutaneous administration of LMWH. Compared to a baseline R-time within reference range, R values two hours after drug administration were increased by at least 70%. In most cases the R-time was increased more than 4 times the baseline measurement. While about half of cats demonstrated their peak prolongation in R-time at 120 minutes, some cats demonstrated equal or greater prolongation of R-time at an earlier timepoint of 30 minutes. It is interesting to note that a peak hypocoagulable effect as evaluated by TEG at the 30-minute timepoint was sometimes discordant with the peak of measured anti-Xa activity (e.g., R-time was greater at 30 minutes but anti-Xa activity for the same individual was greater at the 120-minute timepoint). As anticipated, the LMWH effects on coagulation were not typically detected by standard coagulation assays.8

Previous research (both in vitro and ex vivo) in human subjects have demonstrated that TEG parameters will fall outside the reference range at concentrations of UFH and LMWH that are not detected by standard coagulation assays (i.e., aPTT analysis).9-11 In some circumstances, authors have even suggested that standard TEG may be more sensitive than anti-Xa activities.10 This relationship may, in fact, differ with heparin type and concentration. It has been suggested that paired analyses comparing standard TEG and TEG with heparinase may be enhance the sensitivity to detect effects of low concentrations of heparin however this subject requires further investigation. Certainly our experience with paired analyses using standard and heparinase cups would support a benefit to this additional analysis in both experimental and clinical cases.

In our earliest work with both UFH and LMWH in cats, significant correlation between anti-Xa activity and TEG parameters was only seen with UFH administration. With standard dosing of UFH, the R value was positively correlated (corr coef 0.577; p= 0.004) and the MA and CI were both negatively correlated (corr coef -0.791; P=0.0251 for both).7 Preliminary analyses with LMWH suggest that if a correlation exists between anti-Xa activity and TEG parameters, the correlation is weak and will likely be limited to the R-value.7,8 It is also interesting to note that despite detectable anti-Xa activity at trough in cats receiving high-doses of LMWH, the measured effect on TEG parameters does not seem to parallel that seen with measured anti-Xa activity. It is hoped that further analyses of available data and evaluation of pharmacodynamics may yet offer further clarification on the relevance of these observations.

Pilot Study: TEG in Cats with Cardiomyopathy and CATE

As TEG has offered additional insight into the pathogenesis associated with thrombosis in other species and may help identify individuals at greater risk for thrombosis, it is hoped TEG may offer similar insight in feline cardiomyopathy. With the hope of better understanding the hypercoagulable state predisposing cats with cardiomyopathy to thrombosis, we initiated a pilot study to explore global coagulation in cats with naturally occurring heart disease. Standard TEG analysis was performed in cats with cardiomyopathy (CM) with and without arterial thromboembolism (ATE). The study is a prospective observational study in a cohort of client-owned cats with CM with and without ATE; blood is collected for routine coagulation profiles and TEG analysis. As in previous work with cats, TEG analysis was performed on recalcified citrated whole blood and R, K, angle and maximum amplitude (MA) were recorded and analyzed. A significant difference is noted in affected cats compared to the reference range established based on TEG analysis in healthy cats. This study is ongoing.

Further Thoughts

Critics of TEG will likely persist and there are certainly limitations to TEG but one might argue there are limitations to standard tests of coagulation and the use of TEG in the clinical and research setting complements and expands our investigations into altered hemostasis when performed with appropriate standardization and quality control. In addition to the work discussed above, developing research interests for our group (species-specific) include the following: 1) additional work with heparinase; 2) exploring work with modified TEG analysis in cats; 3) extend analysis to include/validate fibrinolytic variables in standard or specialized TEG analysis.

References

1.  Luddington RJ. Clin Lab Haem 2005; 27:81.

2.  Otto CM, et al. JAVMA 2000; 217(10):1500.

3.  Donahue SM, et al. JVECC 2005; 15(1):9.

4.  Donahue SM, et al. JVECC 2002; 12(3):191.

5.  Wiinberg B, et al. Vet Jnl 2007; in press.

6.  Callan MB, et al. J Thromb Haemst 2006; 4:2616.

7.  Alwood AJ, et al. JVIM 2007; 21(3):378.

8.  Alwood AJ, et al. JVECC 2005; 15(3):S1.

9.  Zmuda K, et al. Am J Clin Pathol 2000: 113(5):725.

10. Coppell JA, et al. Blood Coagul Fibrinolysis 2006: 17(2):97.

11. Simons R, et al. Anesthesia 2007; 62:1175.

Speaker Information
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Amy Alwood, DVM, DACVECC
Philadelphia, PA


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