By C. Guillermo Couto, DVM, Dipl. ACVIM

Sighthounds in general, and greyhounds in particular, have evolved over the past 6,000 to 7,000 years to follow their prey by sight. Hence, they have developed numerous physiologic and hematologic adaptations specific to the breeds.¹

In retired racing greyhounds (RRGs), the packed cell volume (PCV), hemoglobin concentration, red blood cell (RBC) count, and whole blood viscosity are higher, while the white blood cell count, neutrophil count, and platelet count are lower than in other dogs (reviewed in 2). The serum total protein, globulin, alpha-globulin, and beta-globulin concentrations are also lower than in non-greyhound dogs.^3-5 Interestingly, platelet aggregation under high shear, as determined with the platelet function analyzer 100 (PFA-100), is significantly higher in RRGs than in non-greyhound dogs.⁶ The latter is particularly noteworthy, since lower platelet counts typically lead to prolonged closure times when using PFA.⁷

There was anecdotal information suggesting the hemostatic system in greyhounds is also different. After losing a greyhound patient due to unexplained bleeding in early 2003, I embarked in a saga to determine why greyhounds bleed spontaneously. Shortly thereafter, I also discovered that some actually clot excessively. This review article summarizes my clinical and research experience during the past 15 years trying to decipher hemostasis in greyhounds.

Hemostasis for clinicians

Under normal conditions, injury to a blood vessel leads to immediate vascular changes (e.g. vasoconstriction) and rapid activation of the hemostatic system. Changes in axial blood flow lead to exposure of circulating blood to subendothelial collagen, resulting in rapid adhesion of platelets to the affected area. The adhesion of platelets to the subendothelium is mediated by adhesive proteins, such as von Willebrand factor (vWF) and fibrinogen, among others. After adhering to the area of endothelial damage, platelets aggregate and form the primary hemostatic plug, which is short-lived and unstable. The primary hemostatic plug serves as a framework in which secondary hemostasis occurs because most of the clotting factors "assemble" the thrombus or clot on the platelet plug (reviewed in 8).

Although the intrinsic, extrinsic, and common coagulation pathways have been well characterized and are still used to teach physiology of hemostasis, coagulation in vivo does not necessarily follow these distinct pathways. For example, factors XII and XI do not appear to be necessary for the initiation of coagulation (e.g. dogs and cats with factor XII deficiency do not have spontaneous bleeding tendencies). It is generally accepted the physiologic mechanism responsible for clotting in vivo is primarily tissue factor (TF) activation of factor VII. TF is ubiquitous, and it is present in most every cell surface, with the exception of resting endothelium. Disruption or "damage" to the endothelial cells causes exposure of circulating blood (containing factor VII) to TF, and rapid activation of the coagulation cascade via the traditional extrinsic system. Thrombin plays a pivotal role in both the activation and inactivation (through thrombomodulin) of the hemostatic mechanisms (reviewed in 9). In the past two decades, the traditional coagulation cascade has been thought of as a common pathway from early in the process; the traditional intrinsic, extrinsic, and common pathways are now known to be interrelated.^9-11

The stimuli that activate coagulation also trigger the fibrinolytic and kinin pathways. Fibrinolysis is an extremely important safeguard mechanism because it prevents excessive clot or thrombus formation. When plasmin lyses fibrinogen and fibrin, it generates fibrin degradation products (FDPs), which impair additional platelet adhesion and aggregation in the site of injury. Once fibrin has been stabilized by complexing factor XIII, plasmin biodegradation generates D-dimers instead. The activation of plasminogen into plasmin results in the destruction (lysis) of an existing clot (or thrombus) and interferes with the normal clotting mechanisms (inhibition of platelet aggregation and clotting factor activation in the affected area). Therefore, excessive fibrinolysis usually leads to spontaneous bleeding. Two main molecules stimulate plasminogen activation into plasmin: tissue plasminogen activator (tPA) and urokinase-type plasminogen activator. Three plasminogen activator inhibitors (PAI) termed PAI-1, -2, and -3 inhibit fibrinolysis, leading to thrombosis (reviewed in 8).

Other systems opposing blood coagulation also become operational once intravascular clotting has occurred. The best characterized ones include antithrombin (AT), a protein synthesized by hepatocytes that is a cofactor for heparin and inhibits the activation of factors IX, X, and thrombin. AT also inhibits tPA. Proteins C and S are two vitamin K-dependent anticoagulants also produced by hepatocytes. These three factors are some of the natural anticoagulants that prevent excessive clot formation (reviewed in 8).

The thrombelastograph (TEG) is a whole blood coagulation analyser that evaluates cell/protein interaction; it allows for a global analysis of the hemostatic system, including primary and secondary hemostasis, and the fibrinolytic system.^12,13 The TEG parameters reported by the instrument are as follows: The "R-time" is the time from addition of the agonist (CaCl2 or TF) to the citrated whole blood in the cup, until the clot formation reaches detectable levels and represents the enzymatic portion of coagulation. The "K-time" is the time from detection of the endpoint "R-time" until the clot reaches a determined firmness, a measure of the speed to reach a certain level of clot strength, representing the clot kinetics. The angle ("alpha") is related to the fibrinogen concentration (and function) and the rapidity of fibrin formation and cross-linking, also related to the kinetics of clot formation. The "MA" is the maximum amplitude or ultimate strength of the fibrin clot and represents primarily the contribution of platelet aggregation to clot formation. "G" provides a measure of clot strength viscoelastographically and is calculated from the MA. Finally, LY60 represents the percent or proportion of clot lysis (i.e. clot retraction and fibrinolysis) or decrease in amplitude from the MA at 60 minutes, under the area of the tracing.^12,13 In a recent study, we demonstrated thromboelastographic features in RRGs are different from those in other dogs; mainly the maximum amplitude and clot strength are significantly lower than in other breeds.¹³

Surgery and hemostasis

Surgery typically induces a hypercoagulable state.¹⁴ A recent study in humans demonstrated a continuous increase in clot firmness, as determined by TEG, two to six days after surgery; however, there were no changes in one-stage prothrombin time (OSPT) or activated partial thromboplastin time (APTT) that suggested hypercoagulability.¹⁴ The proposed mechanism of this hypercoagulability is associated with local tissue trauma, release of tissue factor from damaged vessels, decreased blood flow, activation of inflammation, and compromised fibrinolysis.^14,15

Intraoperative and immediate postoperative bleeding can be due to local surgical technique (i.e. surgical bleeding) or to systemic abnormalities (i.e. nonsurgical bleeding).¹⁶ The latter includes primary hemostatic defects, such as thrombocytopenia, platelet dysfunction, or von Willebrand disease (vWD), as well as secondary hemostatic defects, including hypofibrinogenemia, hypoprothrombinemia, hemophilia A or B, factor VII deficiency, or combined clotting factor deficiencies.¹⁶ Coagulopathy due to trauma must also be considered in such patients.¹⁷ Finally, systemic endothelial damage or dysfunction after postoperative septic complications or hypertensive crises can result in thrombocytopenia and generalized bleeding, as described in women with hemolysis, elevated liver enzymes, low platelet count (HELLP) syndrome associated with preeclampsia and in children with hemolytic-uremic syndrome.¹⁸ Delayed postoperative bleeding is more likely due to abnormal fibrin stabilization, factor XI deficiency, or enhanced fibrinolysis.¹⁹