"Bone Marrow Evaluation" TUMOR TIDBITS, A BIWEEKLY VETERINARY ONCOLOGY E-LETTER Volume 2; Number 14; August 28, 2001. Editor: Kevin A. Hahn, DVM, PhD, Dipl - ACVIM (Oncology) & Overall Nice Guy! THIS WEEK'S TUMOR TIDBIT: BONE MARROW EVALUATION INTRODUCTION There is valuable information that can be obtained from marrow evaluation without advanced clinical pathology training. Interpretation of the bone marrow is critically dependent on the findings of a concurrent CBC. For example, if an animal has a high M:E ratio (more myeloid cells than erythroid), the CBC results may determine if it is due to an erythroid hypoplasia or a myeloid hyperplasia. The interpretation would depend on whether the animal had a peripheral anemia or a leukocytosis. CELLULARITY The cellularity of the marrow is determined by examining the relative amounts of fat and cellular material present in the marrow particles. A normal cellularity will have approximately 50% cells and 50% fat; slightly more in young growing animals, and slightly less in geriatric patients. Increased cellularity or hyperplastic marrow (>75% cells) is seen in marrows that are responding to peripheral cytopenias (e.g. regenerative anemia, leukopenia or thrombocytopenia). It is also noted in animals with an increased demand for leukocyte production (e.g. leukocytosis). Increased cellularity is also seen when the marrow is occupied by neoplastic cells (myelophthisic disease). Decreased cellularity (Hypoplastic or Aplastic Marrow) (>75% fat) is observed when there has been insult or injury to one or more of the hematopoietic cell lines. This may result in an aplastic anemia where all cell lines are decreased, or a cytopenia of only one or 2 of the different cell lines. There are many infectious agents, drugs, hazardous materials, and immune- mediated diseases that can cause hypoplastic or aplastic marrows. Unless previous exposure to drugs or infectious agents can be identified, most etiologies are undetermined and the disease is classified as idiopathic. MARROW IRON OR HEMOSIDERIN The bone marrow is a major site for storage iron to be used in hemoglobin synthesis. Iron or hemosiderin will appear as blue or black inclusions of amorphous material either in the background of the smear or within the cytoplasm of marrow macrophages. The presence or absence of stainable iron must be evaluated within a unit particle. The evaluation of iron content in a marrow is significant only in anemic animals. The evaluation of iron stores in the marrow of dogs is a judgment call. If abundant stainable iron is seen within a unit particle, it is assumed to be increased. In the cat stainable iron is normally not detected, so any amount of visible iron would be an indication of increased iron stores. Marrow iron stores are increased in the anemia of chronic inflammatory disease, hemolytic anemias, and in cases of decreased red cell production such as red cell aplasia or aplastic anemias. Marrow iron stores are also increased in animals that have received recent blood transfusions. Decreased storage iron is seen in cases of iron deficiency. In small animals this is almost exclusively associated with chronic blood loss. It should be noted that the absence of stainable iron in the marrow of cats couldn't be used as an indication of iron deficiency since stainable iron is not usually present in normal cats. MEGAKARYOCYTES Megakaryocytes are the largest cells in the bone marrow and are easily recognized with the 10x or 20x objective. Mature megakaryocytes have abundant amounts of pale blue cytoplasm that often contains fine eosinophilic granules. The cells appear to be multinucleated but actually contain a single, large, lobulated nucleus. Immature megakaryocytes are smaller cells, but are still at least 2 times larger than other hematopoietic precursors. They contain scant, deep blue cytoplasm and a round to slightly lobulated nucleus (4 lobes or less). In a normal marrow, the majority of the megakaryocytes should be mature. Increased numbers of immature cells are seen in a regenerative response to peripheral thrombocytopenias. One of the most common, and most useful reasons for a veterinarian to evaluate a bone marrow is to assess a patient with a peripheral thrombocytopenia. Megakaryocyte numbers in the marrow will allow classification of the disease as a production or a destruction problem. In a normal animal 1 to 3 megakaryocytes should be seen while examining the marrow particles on a low power objective (10x). If the patient is responding to a peripheral destruction or consumption of platelets, increased numbers of megakaryocytes will be seen (megakaryocytic hyperplasia), some of which will be immature. This patient has a much better prognosis, and is more likely to respond to appropriate therapy than one with a production problem where few to no megakaryocytes are found (megakaryocytic hypoplasia). ERYTHROID SERIES Erythroid precursors are characterized by a round nucleus with dark, dense chromatin. The immature erythroid precursors (rubriblasts) are large cells with deep blue cytoplasm. They have a round nucleus with clumped chromatin and one or more nucleoli. As these cells mature into rubricytes and metarubricytes they become smaller, the nucleus becomes dark and pyknotic, and the cytoplasm changes from deep blue to grey. In the normal marrow, the majority (>90%) of the erythroid precursors are mature cells, rubricytes and metarubricytes. In a normal marrow, the red cell series must mature all the way to anucleated, polychromatophilic erythrocytes (reticulocytes). If a bone marrow aspirate is performed to evaluate a peripheral anemia, we will want to determine if we have an erythroid hyperplasia or erythroid hypoplasia. Erythroid hyperplasia has increased numbers of erythroid precursors (myeloid / erythroid ratio of <1), in the presence of a peripheral anemia. This indicates the bone marrow is responding to a peripheral loss or destruction of erythrocytes. Increased numbers of rubriblasts may be seen, but these cells should not exceed more than 10% of the total nucleated cell population. Phagocytosis of erythroid cells by marrow macrophages may indicate an immune-mediated destruction or may be seen in patients who have had a recent blood transfusion. Erythroid hypoplasia, or decreased numbers of erythroid precursors (M / E ratio of > 2) in the presence of a peripheral anemia, would indicate the bone marrow is not responding. This suggests the anemia is due to an erythrocyte production problem or an immune-mediated destruction of erythroid precursors. MYELOID SERIES Immature myeloid cells (myeloblasts) have round to irregularly shaped nuclei with one or more nucleoli and paler cytoplasm than erythroid cells. The cytoplasm of these cells often contains fine eosinophilic granules (primary granules). The nuclei contain chromatin that is less clumped and more diffuse than the nuclei of the erythroid cells. In the normal marrow, the majority (>90%) of the myeloid cells will be myelocytes, metamyelocytes, bands, and segmented granulocytes. The neutrophil line is the most abundant cell type in the myeloid series. As the neutrophils mature to myelocytes, metamyelocytes, bands, and segmented neutrophils, the cytoplasm changes from light blue to clear, and the nucleus develops from bean-shaped to banded to segmented. If the bone marrow is evaluated because of abnormal leukocyte numbers in the peripheral blood, we will want to determine if there is a myeloid hyperplasia or a myeloid hypoplasia. Myeloid hyperplasia, or increased numbers of myeloid precursors (M / E ratio of > 2), in the presence of a peripheral leukocytosis or leukopenia, is most consistent with a marrow that is responding to a peripheral demand for neutrophils. This may be seen in a number of immune- mediated and inflammatory diseases. Increased numbers of immature myeloid cells may also be present, however, myeloblasts should not exceed more than 10% of the nucleated cell population, even in a strongly regenerative response. Myeloid hypoplasia, or decreased numbers of myeloid precursors (M / E ratio of <1), in the presence of a peripheral leukocytosis or leukopenia, would indicate a decreased production of neutrophils by the marrow. This is particularly significant if the patient has a peripheral neutropenia. There are a number of chemotherapeutic agents, drugs and infectious agents that may damage the bone marrow, resulting in myeloid hypoplasia. LEUKEMIAS A leukemia is a bone marrow neoplasm of the hematopoietic cells. Leukemias may be classified as a myeloproliferative disease (erythroid, myelogenous, or monocytic leukemia) or a lymphoproliferative disease (lymphoid leukemia) depending on the cell line from which the neoplastic population arises. Leukemias are also classified as acute or chronic depending on the immaturity of the neoplastic cell. In all types of leukemia, the bone marrow will be hypercellular. In acute leukemias the neoplastic cell is the blast cell or immature precursor. Acute leukemia is diagnosed whenever the blast cell population of the marrow exceeds 30% of the nucleated hematopoietic cells. An acute leukemia may involve any one of the hematopoietic cell lines (erythroid, myelogenous, monocytoid or lymphoid). Myeloproliferative diseases (erythroid, myelogenous, or monocytoid) may occasionally have 2 neoplastic cell lines occurring simultaneously (e.g. myelomonocytic). In cases of acute leukemia, it may be difficult to determine which type of leukemia is present without the use of special cytochemical stains. Therefore, whenever acute leukemia is diagnosed, extra marrow slides should be air-dried and mailed to a reference laboratory for classification. It is important to recognize that not all leukemia patients have blast cells in peripheral circulation. Some patients may have"aleukemic" leukemia. In chronic leukemias the neoplastic cell is the mature blood cell (erythrocyte, granulocyte, lymphocyte, monocyte, or platelet). Patients with chronic leukemia have an unexplained, marked elevation of the affected cell line in the peripheral blood. With the exception of the chronic lymphocytic leukemia, the diagnosis is sometimes difficult because the neoplastic cells are mature, and blast cells do not exceed 30% of the bone marrow population. Finding dysplastic changes in the hematopoietic cells in the peripheral blood or bone marrow may be an aid in diagnosis (see dysplastic changes above). Patients with chronic leukemia have a better short-term prognosis than those with acute leukemia. In general, patients with lymphoid leukemias have a better prognosis with regard to survival time and response to therapy than do patients with the comparable myeloproliferative disorder (e.g., acute lymphocytic leukemia vs. acute myelogenous leukemia). In lymphoid leukemia, hematopoietic cell lines other than lymphoid are not involved in the neoplastic process. Therefore, severe peripheral cytopenias are not usually encountered except in advanced cases where myelophthisic disease has occurred. MORE QUESTIONS ABOUT BONE MARROW EVALUATION? Don't hesitate to call or email us at Gulf Coast Veterinary Oncology! I can forward additional info by email if needed. IN 2 WEEKS: HOW TO GET A BIOPSY! ALL THE BEST, Kevin Hahn Kevin A. Hahn, DVM, PhD Diplomate ACVIM (Oncology) & Overall Nice Guy Gulf Coast Veterinary Specialists 1111 West Loop South, Suite 150 Houston, TX 77027 P: 713.693.1166 F: 713.693.1167 www.gcvs.com drhahn@g...