Anemias and thrombocytopenias (Proceedings)

Clinical approach to anemia

The typical clinical signs of anemia are weakness, exercise intolerance, inappetance or anorexia, pale mucous membranes, tachypnea, and icterus. The degree to which these clinical signs are manifested are dependent on the speed at which the anemia develops and the severity of the anemia. A patient with myelofibrosis that develops anemia over the course of many weeks may be surprisingly asymptomatic at a hematocrit of 18%, whereas a dog with acute immune-mediated hemolytic anemia and a hematocrit of 20% may be critical.

Physical examination findings in patients with anemia can be variable depending on the cause, but the clinician should look carefully for any of the following signs: icterus, which is an indication of hemolysis; splenomegaly, which can be seen in immune-mediated disease, hemotropich mycoplasma infection, Babesia, neoplasia (hemangiosarcoma and lymphoma), and extra-medullary hematopoiesis; purpura, which suggests that thrombocytopenia may be present and responsible for the anemia; and heart murmur, which is secondary to decreased blood viscosity with severe anemia.

Signalment can be an important predictor of disease. Certain breeds of dog may have hemolytic anemia associated with a genetic disorder. This includes pyruvate kinase deficiency in basenjis and beagles and phosphofructokinase deficiency in English springer spaniels. Also an older dog is more likely to have anemia associated with neoplasia and a middle-aged dog more often has immune-mediated disease.

There are a variety of oddities and miscellaneous points of interest when evaluating the complete blood count and hemogram. For instance: cat platelets are almost the same size as cat red cells and are often times counted as RBCs by most machines—this does not appreciably affect the RBC count, but the platelet counts are usually very low; cat RBCs cannot be detected in machines dedicated for counting human RBCs; Akitas normally have lower MCVs; some poodles may have very high MCVs (poodle macrocytosis); in the presence of anemia, macrocytosis usually reflects current or recent accelerated erythropoiesis; markedly regenerative anemias in dogs are usually accompanied by modest increases in MCV, whereas cats often have MCVs two times normal; prominent macrocytosis may be seen in FeLV-associated anemias whether regenerative or not and has also been seen in nonregenerative anemia in FIP; macrocytosis from Vitamin B12 deficiency (Cobalamin deficiency) is seen in a few breeds (esp. Giant schnauzer and Border collie); microcytosis is a feature of iron deficiency in adult dogs and cats; puppies are born with high MCV and hematocrit—the MCV is normal by 2-3 months, but the hematocrit declines to approximately 30% by 4-6 weeks and gradually increases to normal values by 6-12 months; kittens are born with hematocrit values of ~35% which fall to 25% by 3-4 weeks, and increase to normal by 16 weeks.

Red blood cell morphology can sometimes be indicative of specific disease processes. Some items to keep in mind include: polychromasia indicates increased release of immature cells; nucleated RBCs may accompany reticulocytosis, but are also seen in myeloproliferative disease, dysmelopoiesis, and extramedullary hematopoiesis; basophilic stippling, the spontaneous aggregation of ribosomes and organelles into basophilic granules, should lead to a suspicion of lead toxicity; Heinz bodies, which are hemoglobin precipitated by oxidative injury and are seen frequently in cats and are probably normal in cats in small numbers; spherocytes, red blood cells that are smaller and have lost their central pallor, which are formed when part of the RBC membrane is removed in an immune-mediated process and are the hallmark of immune-mediated hemolytic anemia; and acanthocytes, irregularly shaped RBCs, which are seen in liver disease, hemangiosarcoma, hypercholesterolemia, and feline hepatic lipidosis.

After detemining that a patient is anemic (low hematocrit/PCV/RBC mass), the first question to ask is: Is this regenerative or non-regenerative anemia? Determination of RBC regeneration is made by evaluating the reticulocyte count. The presence of nucleated RBCs and polychromasia are not reliable indicators of regeneration. Remember that with recent blood loss or hemolysis less than 2-4 days old, the regenerative response may be inapparent or inadequate. The easiest and best way to determine if the anemia is regenerative is to calculate the absolute reticulocyte count. This is done by multiplying the percent of reticulocytes by the RBC count. The absolute reticulocyte count should be >60,000 in cats and >100,000 in dogs (some use >60,000 in dogs).

Regenerative anemia is more common and is broadly categorized as hemolytic or hemorrhagic disease. Hemorrhage is characterized by evidence of external blood loss and low total protein, but this may not be true in all cases. With hemolysis, one often sees icterus, hemoglobinuria, bilirubinuria, and normal or increased total protein. When thrombocytopenia is present, remember that it typically requires a platelet count <50,000 in dogs and <10,000 in cats to have spontaneous hemorrhage associated with thrombocytopenia. The most common acquired cause of hemorrhage is a coaguloapthy associated with anticoagulant rodenticides. The identification of iron deficiency anemia should prompt the investigation of GI blood loss, such as from hookwork infestation or bleeding GI neoplasms.

Causes of hemolysis include immune-mediated hemolytic anemia; ehrlichiosis, secondary to vasculitis associated with the infection (a form of microangiopathic hemolytic anemia); Mycoplasma haemofelis, which can produce recurrent, cyclic hemolysis; babesiosis; cytauxzoonosis; oxidative injury which can lead to methemoglobinemia, Heinz body formation, and damage to membrane proteins and is caused by onions and acetaminophen, among other things; pyruvate kinase deficiency, , which is characterized by chronic, severe hemolyis and terminal myelofibrosis by 1-3 years; phosphofructokinase deficiency, which is characterized by mild to moderate chronic hemolysis with superimposed episodes of severe hemolysis precipitated by vigorous exercise or panting associated with overheating; microangiopathic disease, which is seen with hemangiosarcoma, heartworm disease, splenic torsion, disseminated intravascular coagulation, and other neoplasia; zinc toxicity, which is associated with the ingestion of pennies minted after 1983 and other zinc objects; and hypophosphatemia, which can be seen in re-feeding syndromes and during therapy for diabetic ketoacidosis.

Non-regenerative anemia can be caused by extra-marrow disease that inhibits erythropoiesis, either by the inhibition of EPO production, sequestration of iron, or secretion of cytokines that inhibit erythropoiesis. The presence of bicytopenias (non-regenerative anemia with thrombocytopenia or leukopenia) or pancytopenias (all cell lines decreased) supports the presence of intra-marrow disease. Causes of non-regenerative anemia include: anemia of chronic disease, which is the most common form of non-regenerative anemia and is usually mild; chronic renal failure; hypothyroidism and hypoadrenocorticism; FeLV infection; aplastic anemia, which is characterized by severe marrow depopulation and pancytopenias and can be caused by FeLV, estrogen toxicity, phenylbutazone toxicity, trimethoprim-sulfa, chemotherapy, canine ehrlichiosis, or may be idiopathic; myelodysplasia, which is most common in the cat; myeloproliferative disease, which is caused by purposeless, neoplastic proliferation of cells in the marrow; myelofibrosis and osteosclerosis, which are commonly a response to injury to the marrow; lymphoprolierative disorders; and chloramphenicol toxicity, which is less severe in animals than in people and is reversible in dogs and cats.

Specific diseases

      Autoimmune hemolytic anemia (AIHA)

The body has produced antibodies directed against normal RBC surface antigens; very uncommon.

      Immune-mediated hemolytic anemia (IMHA)

The body has produced antibodies directed against altered surface RBC antigens, either from drugs, toxins, viruses, bacterial proteins, etc...; the most common form. This is fairly common in dogs, rare in cats. It may be mediated by IgG or IgM in an extravascular process (the macrophages of the reticuloendothelial system remove the coated RBCs and destroy them) or by complement (produces intravascular hemolysis: this is still an IgG mediated process, but large amounts of IgG bind to the surface and activate the complement system). 74% of the cases are positive for IgG. Rarely is an underlying cause identified, even though this is not true AIHA. Subjectively more common in spring and early summer and in one study, 40% of the cases occurred in May and June. Females may be more commonly affected and the mean age of affected dogs in one study was 6.7 years.

Most are acute or peracute, and chronic cases are rare. Anorexia, lethargy, and exercise intolerance are common clinical signs and one may see pallor, icterus, fever, tachycardia, and hepatosplenomegaly. Most have PCV<20%, with a mild increase in MCV in dogs and marked increase in MCV in cats. Reticulocytes are present within 2-4 days and 33% are nonregenerative at presentation. Patients that have the immune-destruction localized to the marrow never have reticulocytosis. These are identified by seeing erythroid hyperplasia, and sometimes erythrophagocytosis, on a bone marrow aspirate/core biopsy. Spherocytosis is seen in 67% of the cases and 10% are hemoglobinemic or hemoglobinuric. Autoagglutination may be present and this can be quite dramatic and is documented by observing the RBCs clump when suspended in saline (1 drop of blood in 2-5 drops of saline). The autoagglutination may be visible macroscopically. A severe neutrophilic leukocytosis is common with a left shift and monocytosis and in one study, 100% were hyperbilirubinemic.

The Coombs' test quantitates Ig or Complement bound to RBC, but only 74% will have a positive test. Unnecessary to perform if they are autoagglutinating, although long-term monitoring of RBC cell-surface antibodies by flow cytometry may be an effective means to determine the efficacy of therapy and the appropriateness of long-term therapy. Bone marrow evaluation (biopsy, cytology) is necessary for patients with severe, acute, non-regenerative anemia that is suspected to be immune-mediated at the level of the marrow.

Prednisone is the mainstay of therapy and is initiated at 2-4 mg/kg PO q24h. Continue on the initial dose for 2 weeks beyond a normal PCV and then reduce the dose by 25 every 2-4 weeks for 3-5 reductions. Some dogs will relapse if tapered too fast. Prednisone inhibits the RES macrophages and does little to affect Ig production (it may decrease Ig production later on). Most patients show a response in 2-3 days, although it may take 5-7 days. This therapy is usually insufficient in patients that are autoagglutinating. Other options include azathioprine (Imuran), cyclosporine, intravenous gamma globulin, gold sodium thiomalate, and mycophenolate mofetil. For rapid control of patients with severe auto-agglutination, cyclosporine, mycophenolate mofetil, and IVGG are recommended.

Clinical approach to thrombocytopenia

The clinical signs of thrombocytopenia include spontaneous bleeding, which usually does not occur until the platelet count is <50,000, and usually <30,000, in dogs and <10,000 in cats; petechial and ecchymotic hemorrhages seen in the skin, mucous membranes, sclera; and, possibly, melena, epistaxis, intraocular bleeding, and hematuria.

The causes of thrombocytopenia are typically divided into categories of accelerated platelet removal (immune-mediated destruction), bone marrow disorders resulting in defective platelet production, increased platelet consumption (disseminated intravascular coagulation), and increased loss. Causes of accelerated platelet removal include primary immune-mediated thrombocytopenia, systemic lupus erythematosus, drug reaction (especially trimethoprim-sulfa), neoplasia, infections, and microangiopathy.

Causes of bone marrow disease with defective platelet production include myeloproliferative or lymphoproliferative disease, idiopathic bone marrow aplasia, drug-induced myelosuppression/marrow hypoplasia, chronic infection, hereditary disorders, and cyclic thrombocytopenia. Causes of increased platelet consumption include disseminated intravascular coagulation, hemolytic-uremic syndrome, and vasculitis. Cause of platelet loss include redistribution (splenomegaly, vascular pooling from hypothermia, splenic torsion, portal hypertension, hepatomegaly) and hemorrhage.

During the diagnostic work-up, we want to get a good history to exclude drug exposure or evaluate other exposure possibilities for disease. When evaluating the hemogram, it's important to remember that platelet counts <20,000 are usually from immune-mediated thrombocytopenia, although some cases of DIC may have platelet counts that low. Also, a low mean platelet volume (MPV) is seen in early immune-mediated disease and a large MPV suggests active bone marrow and is seen later in immune-mediated disease. Other tests that may be performed include an anti-nuclear antibody test for SLE, anti-platelet antibody test, coagulation profile, bone marrow aspirate/core biopsy, radiographs of thorax and abdomen, an occult heartworm test, s serology for FeLV, E. canis, RMSF, and a serum biochemistry profile (which actually has the lowest yield of all the tests listed).

In dogs, the most common cause of clinically significant thrombocytopenia is immune-mediated thrombocytopenia, which may be a primary or secondary immune-mediated destruction. It is characterized by the production of antiplatelet antibodies that bind to and destroy/damage the platelets. The antibody-coated platelets are destroyed by the reticuloendothelial system (fixed macrophages). IMT can be severe and acute or chronic. The clinical presentation varies depending on the severity of the thrombocytopenia, although platelet number alone if often a poor predictor of bleeding tendencies. Some dogs with platelet counts under 10,000 don't bleed, while others with platelet counts of 20,000 have significant bleeding. This is probably related to the formation of microplatelets, but can be associated with the degree of disruption of normal platelet function.

The diagnosis is often established by the presence of a platelet count <20,000 and no evidence of concurrent disease. A bone marrow aspirate/core biopsy may show increased megakaryocytes with IMT, but this test is often not performed. Antiplatelet antibody detection by flow-cytometry is currently used at KSU. In this test anti-canine IgG conjugated with fluoroscein isothiocyanate is incubated with the platelets and then is analyzed by flow cytometry. This requires small blood volumes (3-ml EDTA blood) and can be sent overnight (stable for 72 hours). This test can still have false positives and false negatives and is not frequently performed due to the minimal amount of information obtained. In most cases, the diagnosis of IMT is by elimination of all other causes of thrombocytopenia (normal PT and PTT, negative serology, no evidence of drugs, no obvious neoplasia, active bone marrow).

The therapy for IMT is much the same as for IMHA and other immune-mediated diseases. Prednisone is still the initial drug of choice, used at 2-4 mg/kg q24h or divided q12h. Other choices include azathioprine, cyclosporine, vincristine, intravenous gamma globulin, gold sodium thiomalate, and mycophenolate mofetil. Splenectomy is controversial and is not appropriate for the acute thrombocytopenic patient, but may be beneficial in the chronic, recurrent patient. Neither whole blood transfusion or platelet-rich plasma significantly increase the platelet count, but may be beneficial in reducing hemorrhage from mucosal surfaces. Primarily, transfusions are used to restore red cell numbers secondary to hemorrhage.

Idiopathic immune-mediate thrombocytopenia in the cat is rare, although reported in the literature. In most instances, thrombocytopenia is secondary to some other disease and cats should be extensively evaluated for some other primary disease.