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HEMOSTASIS
Signs of abnormalities seen in animals with bleeding disorders include:
II PHYSIOLOGY
The inner surface of normal vasculature is smooth and coated with anticoagulant factors. When the endothelium is damaged, collagen fibers are exposed and thromboplastin and vonWillebrand factor leak from the injured cells. Activation occurs due to the contact of platelets with these collagen fibers and the released vonWillebrand factor. The coagulation system is activated when blood proteins contact tromboplastin leaking from damaged tissue cells.
The function of the platelet is primary hemostasis—the formation of a platelet plug. This is initiated by adhesion. The platelets adhere to the site of injury, in response to contact with collagen and vonWillebrand factor. These activated platelets aggregate, attracted to each other and sticking together. The cell membranes dissolve and viscous metamorphosis occurs, as a fragile jellylike plug is formed (imagine sneezing right after a nosebleed stops—it’s the platelet plug that comes out). This platelet mass is the substrate on which the coagulation system reacts to form a fibrin clot (scab). The coagulation system is a series of enzymatic reactions that results in the formation of an insoluble fibrin mesh around the platelets. This series of reactions is called the coagulation cascade. Most of the coagulation factors are protein-based enzymes that circulate in inactive form. The coagulation factors were numbers by order of discovery, not by their place in the coagulation cascade. Don’t learn the names of these factors, but for information, they are:
Factor I fibrinogen vonWillebrand factor (factor VIIIa) is part of factor VIII, but functions independently of the coagulation cascade. It is an adhesive protein produced by the endothelial cells and the megakaryocytes. It facilitates the attachments of platelets to collagen fibers. There are three coagulation pathways, two that initiate coagulation (the intrinsic and extrinsic pathways) and the common pathway that completes the fibrin clot formation. The intrinsic pathway is initiated by the contact of blood components with abnormal endothelial surfaces (i.e. collagen fibers). This is a slow process and generally takes several minutes to complete. The extrinsic pathway is very rapid, completing its action within seconds. This occurs when shed blood contact thromboplastin in the tissues surrounding the damaged blood vessel. The intrinsic and extrinsic pathways merge to form the common pathway. This part of the cascade converts fibrinogen to fibrin, forming a protective scab. Coagulation must be followed by fibrinolysis—the breakdown of fibrin—or we (and our animals) would be giant walking scabs. The fibrinolytic system replaces fibrin with collagen, re-establishing the normal channel. As fibrinolysis occur, fibrin degradation products are formed, coating platelets and interfering with further clotting.
III LABORATORY EVALUATION OF BLEEDING DISORDERS
Sodium citrate is the anticoagulant of choice. It has a bright blue top. Plastic tubes are preferred over glass tubes, because plastic inhibits the coagulation cascade. Plasma should be harvested within thirty minutes of collection and, if shipped, should be with dry ice. Inadequate platelet numbers (thrombocytopenia) is the most common acquired coagulopathy in small animal practice, and it is important to be able to accurately assess platelets. The reference range for platelets in all species is 200,000 – 500,000/ul. It is easy to estimate platelet numbers and the results are fairly accurate an animal’s platelet count is relatively normal. All animals should have three to thirty platelets per oil immersion field. A manual platelet count can be obtained by using Unopette® 5855. The accuracy and precision are only fair, however. Generally, automatic blood cell counters are a more accurate method to enumerate platelets, but you should check your machine. Bleeding time is a method that evaluates both the quantity and function of platelets. The protocol requires the incision of the buccal mucosa. The incision should be blotted every 30 – 60 seconds without actually touching the skin. If there are sufficient platelets that are capable of forming a platelet plug, clotting should occur in one to five minutes. Some automatic blood cell counters can determine the mean platelet volume (MPV). Like the MCV, this measures the volume of the average platelet. Platelets that have recently been released from the bone marrow are larger than other platelets. Some machines will also calculate the platelet distribution width (PDW). A decrease in the number of functional platelets is called thrombocytopenia. Hemorrhage is usually not associated with thrombocytopenia, especially if the condition is chronic. Clinical signs may occur if the platelet count drops below 30,000/ul. Bone marrow hypoplasia may result in a thrombocytopenia. Immune-mediated thrombocytopenia (IMTP) may occur; it is rarely the primary problem. It is often idiopathic, but it may also be associated with infections organisms such as rickettsias, immune-mediate hemolytic anemia, drug-induced (many drugs have been implicated, including prednisolone), and viral infections (such as parvovirus and the retroviruses). Clinical signs of IMTP are usually minor, although lethargy, anorexia, depression and petechia can occur. Diagnosis is usually due to exclusion of all other possibilities. Transfusion of platelets is ineffective for IMTP, as the transfused platelets are destroyed within three or four hours. Thrombocytopathia occurs when there are sufficient numbers of platelets but they are unable to function properly. These often result in bleeding and are frequently associated with vonWillebrand factor deficiency or viral infections. Transfusion of platelets is generally the best treatment. There are several tests that can be performed to evaluate the coagulation system. Regardless of the test selected, controls should always be run and used to evaluate the results. The intrinsic coagulation pathway can be evaluated in the clinic with three tests. The whole blood clotting time (WBCT) is an insensitive (thrombocytopenia can affect the results) but simple screening test. Blood is placed in a test tube and gently rocked every 30 – 60 seconds. Dog blood should clot within 2 – 10 minutes and horse blood clots in 4 – 15 minutes. The activated coagulation time (ACT®) test is a modification of the WBCT using silica in a blood collection tube. This is a simple and rapid test. Like the WBCT, it is affected by thrombocytopenia and it is not diagnostic if the blood clots WNL (you must perform the following test). The activated partial thromboplastin time (APTT) is the most sensitive and specific in-clinic test for the intrinsic coagulation system. Unlike the WBCT and ACT tests, it is independent of thrombocytopenia. The patient results are compared to the control; clotting time more than 25% lover than the control is considered abnormal. The APTT in the dog is 17 – 35 seconds. The most common in-clinic test for the extrinsic coagulation pathway is prothrombin time (PT). Like the APTT, it is independent of thrombocytopenia and the results should be within 25% of the control results. The dog’s APTT averages 7 – 10 second. Commercial assays exist for all of the factors and fibrin degradation products (FDP). REVIEW QUESTIONS: 1. Define
hemostasis and list the four components. |