"What I like about the LaserCyte® is that I can tell if a patient has regenerative or nonregenerative anemia right away, so I don't have to send out a test and wait for that information. We get everything we need at once, and it's very easy to use. It's foolproof!"

Dr. Nikki Kelly
Animal Hospital of Polaris
Lewis Center, Ohio

Thrombocytopenia: What is it and why is it important?*

Thrombocytopenia (decreased platelet count) is the most common cause of a bleeding disorder in veterinary medicine. It may be an isolated finding or may accompany other hematologic abnormalities, such as anemia and neutropenia because of marrow disease. Preanesthetic evaluation of platelet numbers is very helpful for the surgeon because the surgical hemorrhage secondary to thrombocytopenia can be life-threatening.

Most cases of thrombocytopenia are spurious (particularly in cats), however, platelets are essential for hemostasis and evaluation of platelets is very important. Examination of a complete hemogram is essential to establish whether thrombocytopenia is an isolated finding or associated with anemia or leucopenia.

Clinical Signs of Thrombocytopenia
The clinical hallmark of thrombocytopenia is the occurrence of petechiae. Petechiae reflect capillary or postcapillary venule bleeding and usually occur at sites of increased intravascular pressure, including the lower abdomen, oral or genital mucosae, and axillary or inguinal friction sites. Large coalescing petechiae are called purpura. Purpura are not usually palpable (skin texture, thickness are normal). Palpable purpura suggest an underlying systemic vasculitis such as ecchymosis, or bruising, and is also associated with primary hemostatic defects.

Clinical signs of bleeding can be seen in any thrombocytopenic patient. The rapidity with which thrombocytopenia occurs affects the platelet number at which clinical signs occur. Rapid platelet destruction leads to hemorrhagic tendencies at much higher platelet counts. If platelets are also dysfunctional, clinical signs occur with only modest reductions in numbers, such as often occurs with rickettsial diseases. Patients with concomitant illnesses and thrombocytopenia bleed more.

Causes of Thrombocytopenia (Table 1)
There are four major mechanisms that result in thrombocytopenia:

• Abnomal platelet production
• Accelerated platelet removal
• Abnormal distribution of platelets
• Some combination of the above

click table to enlarge

Abnormal Platelet Production
Abnormal platelet production is rare and is virtually always accompanied by another cytopenia, such as anemia and/or neutropenia. Platelet production defects include:

• Pure megakaryocytic hypoplasia—immune or infectious etiologies
• Marrow panhypoplasia—drug, infectious or toxic etiologies
• Dysthrombopoiesis (myelodysplasia or megakaryocytic leukemia). Diagnosis requires bone marrow aspirate at least; marrow core biopsy is recommended. Multiple aspirates on different days may be required.

Infectious causes include ehrlichiosis, feline leukemia virus (FeLV) infection, and feline immunodeficiency virus (FIV) infection. Vaccination-induced interference with platelet production has been reported in dogs with measles, distemper and parvoviral vaccinations, and in cats with feline panleukopenia vaccination. Drug-induced suppression of thrombopoiesis usually affects other cell lines:

• Neutropenia usually occurs at day 5 and thrombocytopenia at days 8–10 after exposure to drug.
• Anemia is not usually seen because of longer red cell lifespan.
• Commonly implicated drugs are estrogen, sulfadiazine and nonsteroidal anti-inflammatory drugs.

Both primary and secondary immune-mediated thrombocytopenias occur.

Primary immune-mediated thrombocytopenia

• Is most common in dogs with some breed predisposition, such as cocker spaniels, English sheepdogs, German shepherds and poodles.
• Is associated with the presence of antiplatelet antibodies, which cause accelerated destruction by the mononuclear phagocyte system.
• May also be associated with anti-megakaryocyte antibodies that impair thrombopoiesis.
• Is associated with variable clinical signs, including hemorrhage from the mucous membranes, skin, genitalia, nose or gastrointestinal tract. Many severely thrombocytopenic patients remain asymptomatic.

Laboratory findings in primary immune-mediated thrombocytopenia include:

• Severe thrombocytopenia, often less than 30,000/µL.
• The presence of increased proportions of small or large platelets.
• Megakaryocytic hyperplasia in the marrow if antibodies are directed against circulating platelets; reduced numbers of megakaryocytes if the disease is marrow directed.

Establishing the diagnosis is mainly by exclusion.

• Rule out pseudo-thrombocytopenia due to platelet clumping or EDTA anticoagulation.
• Splenomegaly suggests a secondary process.
• Anemia suggests a concurrent disease process.
• Drug exposure, infection, recent vaccination neoplasia or previous transfusion must be considered.

Secondary immune-mediated thrombocytopenia is the most common cause of canine thrombocytopenia and the most common canine hemostatic disorder. It is associated with underlying conditions, including:

• Systemic autoimmune disease
• systemic lupus erythematosus (SLE)
• immune-mediated hemolytic anemia
• rheumatoid arthritis
• pemphigus
• juvenile-onset polyarthritis of Akitas
• Neoplasia
• multicentric or metastatic
• hematologic
• Infectious diseases including
• FIV, FeLV in cats
• Ehrlichiosis
• Rocky Mountain spotted fever
• Vaccination with modified live virus distemper vaccines (transient)
• Protozoal infections such as leishmaniasis and babesiosis
• Dirofilariasis
• Histoplasmosis
• Immune-complex vasculitis

Alloimmune thrombocytopenia occurs when maternal antibodies to paternal antigens are transferred across the placenta or in colostrum, and cause platelet destruction in the neonate. It is not reported in dogs and cats. Post-transfusion purpura has been described in dogs receiving DEA1-incompatible erythrocytes or plasma. The resultant thrombocytopenia appears to resolve within hours.

Secondary nonimmune thrombocytopenia

• Causes include disseminated intravascular coagulation, thrombotic thrombocytopenic purpura and hemolytic uremic syndrome.
• Disseminated intravascular coagulation (DIC)—occurs secondary to systemic inflammation (acute, subacute, chronic). It is associated with vascular damage, sepsis, release of tissue thromboplastin from diseased or neoplastic tissue.
• Thrombotic thrombocytopenic purpura—is severe thrombocytopenia, intravascular hemolysis with schistocytosis and neurologic signs.
• Hemolytic uremic syndrome is severe thrombocytopenia, intravascular hemolysis with schistocytosis and renal dysfunction.

Abnormal Platelet Distribution
Abnormal platelet distribution is associated with hypersplenism and endotoxemia. Hypersplenism is a pathologic condition in which a large proportion of circulating platelets become sequestered in the spleen. One or more cytopenias is usually present. Splenomegaly is sometimes present, while endotoxemia may cause splenic pooling of platelets.

Drug-Induced Thrombocytopenia
Decreased platelet production, accelerated platelet removal and platelet sequestration can all be associated with the use of various pharmaceuticals. Implicated drugs include antibiotics, antimicrobial agents, anticonvulsants, anti-inflammatory agents, chemotherapeutic agents, antiviral drugs and diuretics.

click table to enlarge

Drug-induced thrombocytopenias may be marrow-mediated or peripherally mediated. Drug-induced marrow-mediated thrombocytopenia may cause bone marrow suppression, which is most often associated with direct cytotoxic effects on progenitor cells. Immune-mediated drug-induced destruction of megakaryocytes has been reported in dogs.

Drug-induced peripherally mediated thrombocytopenia can be either immune-mediated or nonimmune. Immune-mediated drug-related thrombocytopenia can occur when some drugs provoke an immune response against unadulterated platelets, such as methyldopa, levodopa and gold. Other drugs provoke antibody formation against drug-platelet antigen complexes, such as heparin and quinine. Still other drugs cause immune-mediated thrombocytopenia after prolonged drug therapy or the reintroduction of a previously used drug.

• Secondary exposure results in antibody formation
• Drug absorbed or adsorbed to the platelet surface
• Platelet destroyed as an innocent bystander by immune activity

Conclusion
Platelet numbers should be evaluated both by a hematology analyzer and by examination of a stained peripheral smear. Microscopic examination of the smear is essential to confirm thrombocytopenia (‹10 platelets per 1000x field without significant clumping of platelets, particularly in the "feathered edge" of the smear). If thrombocytopenia is an isolated finding without clinical signs, a separate sample should be evaluated to rule out spurious thrombocytopenia due to clumping or clot formation.

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Use your IDEXX Practice Developer Points toward the purchase of a new LaserCyte® Hematology Analyzer.

With LaserCyte's 5-part differential and reticulocyte count, you can expand your in-house services.

Protect sample integrity and provide immediate answers during the patient visit.
The CBC is a time-sensitive test. Hematology samples are composed of living cells and should be analyzed as soon as possible to prevent cell distortions due to shipment or storage, and to prevent artifacts created by long-term exposure to EDTA.^†

• Samples should be processed within three hours to avoid artificially increased Hct and MCV, and decreased MCHC
• Platelet counts should be performed within one hour after collection
• Red cells crenate, reticulocytes mature, and lymphocytes' nuclei become distorted

LaserCyte's gold-standard laser-flow cytometry delivers immediate value.

Medical

• Diagnose and treat rather than wait for the lab report for confirmation
• Obtain results more quickly—patients' conditions continually change, as does the sample when it leaves the clinic
• Provide the immediate patient care your clients appreciate—recommend treatment, advise clients, rule out diseases without having to call back.
• Control problematic sample draws—redraw bad samples while the animal is still present

Economics

• Receive actual platelet counts and reticulocytes without added cost—reference laboratories often charge more for a CBC and may not include a platelet count or a reticulocyte count
• Increase practice revenue—availability drives diagnostics

Use your Practice Developer Points to Practice What's Possible®

Practice Developer members can use their points to acquire practice-building tools, such as:

• Diagnostic instrumentation including the LaserCyte Hematology Analyzer
• Extended maintenance agreements
• Medical textbooks
• Staff training on important medical topics presented by industry experts
• Lunch or dinner for you and your staff

Contact us

• To redeem points, or find out how to pay for your LaserCyte with Practice Developer call 1-800-340-4579 or e-mail diagnosticedge@idexx.com.
• Not yet enrolled in Practice Developer or not sure? Call 1-800-548-6733, ext. 4987 or e-mail diagnosticedge@idexx.com for more information.

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At the American Animal Hospital Association Annual Meeting, Tampa, Florida

Lunch will be provided at the above session. To register, send us an e-mail with your name, practice, clinic address and how best to contact you. We will send you more specific location information. Seating is limited, so sign up early!

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I often see the phrase "enlarged platelets noted" on the comprehensive CBC from my reference laboratory. My technicians and I also notice this on the blood films we examine in our practice. Is this an important observation or should I just ignore the phrase?

Enlarged platelets are commonly encountered in our population of sick animals, but are uncommon in healthy animals. The interpretation is different for different species.

In the dog and most other species, the finding of enlarged platelets indicates an increased rate of thrombopoiesis (platelet production) in the marrow. In the cat, this finding is extremely equivocal; enlarged platelets are commonly encountered in the peripheral blood film of clinically normal, as well as clinically ill, animals, and it does not correlate well with an increased rate of thrombopoiesis in all cases.

The value of finding enlarged platelets in the dog and non-cat species is directly related to the interpretation of the peripheral blood platelet pool. If there is a decrease in platelet numbers (thrombocytopenia), the possible causes include decreased production at the bone marrow level, peripheral blood destruction as with immune-mediated thrombocytopenia, and increased peripheral blood utilization/consumption during coagulation.

When enlarged platelets are easily found in the case of thrombocytopenia, peripheral blood destruction or utilization/consumption is likely since the bone marrow has the ability to produce platelets. The thrombocytopenia simply indicates that the marrow is not capable of keeping up with the peripheral destruction or utilization/consumption process. If no enlarged platelets are seen, decreased bone marrow production is much more likely, and detailed evaluation of the marrow with fine needle aspiration and possible core biopsy is essential to further characterize the thrombocytopenia. These procedures are much less likely to be of help in characterizing the cause for the thrombocytopenia if enlarged platelets are observed; a different testing protocol would be employed.

In many CBCs in dogs and other non-cat species where there is a normal platelet count, "enlarged platelets" are still seen in the peripheral blood film. Like thrombocytopenia, this morphologic finding suggests that there is likely an increased rate of thrombopoiesis at the bone marrow level. The primary difference in this situation is that the bone marrow is keeping up with the peripheral demand for platelets. Many of the patients we evaluate on a regular basis are animals with inflammatory disease. A critical component of inflammation is the activation of the coagulation system and the utilization of platelets in the process. Many of these cases with enlarged platelets and normal platelet counts are from animals with active inflammatory disease, which is sometimes difficult to characterize solely with a CBC.

Would you like to submit a hematology question for our clinical pathologists to answer in an upcoming issue of Diagnostic Edge?

Responding to this month's question is Dennis B. DeNicola, DVM, PhD, DACVP, Chief Veterinary Educator, IDEXX Laboratories.

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Follow these steps for your whole-blood sample collection for use with the LaserCyte Hematology Analyzer

1. Use the IDEXX VetCollect tube.
2. Use a syringe or vacuum collection system with the appropriate gauge needle for the patient.
3. Be sure to fill the tube with the required amount of blood.
4. Draw the sample gently and transfer if necessary:
• If using a syringe, immediately transfer the sample to the IDEXX VetCollect tube.
• Push the syringe needle through the rubber stopper of the vacuum collection tube.
• The vacuum will draw the correct sample volume. If the blood does not initially flow into the tube, lightly depress the syringe plunger to start the sample flowing.
• Use a minimum of a 21-gauge needle will ensure that the red blood cells are not damaged during the transfer from syringe to tube.
5. Gently invert the tube for 30 seconds to mix.

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... and we want you to hear from us! Pass the following link along to your friends and colleagues. They can register for the Diagnostic Edge to learn more about hematology, stay informed of the latest IDEXX hematology products and services, and respond to various customer and market surveys that we will soon offer. Registration is easy at www.idexx.com/diagnosticedge.

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