Laboratory Studies
A complete blood count (CBC) with platelet count is indicated. The hemoglobin concentration and hematocrit may help guide diagnosis and determine the presence and severity of anemia. White blood cell (WBC) and platelet counts may be decreased in primary marrow disturbances. Mean cell volume (MCV) is a calculated average red blood cell (RBC) volume. An MCV greater than 100 fL is macrocytosis by definition. Because evaluation of RBC size is key to the diagnosis of an anemia, the MCV is considered to be the most important of the RBC indices.
Peripheral blood smear morphology may be helpful. Round macrocytes suggest liver or marrow infiltrative disease, whereas oval macrocytes tend to suggest a megaloblastic disorder. This study provides clues to the etiology of macrocytosis. Hypersegmented neutrophils and macro-ovalocytes strongly suggest megaloblastic anemia. Nucleated RBCs, teardrop cells, decreased or large platelets, and immature WBCs are often present in myelophthisic disease and leukemias.
The reticulocyte count helps determine whether hemolysis is present; it can also indicate malfunctioning bone marrow. Marked reticulocytosis (>4%) is to be expected in hemolytic anemias. A reticulocyte count lower than 1% indicates inadequate marrow production. The reticulocyte count must be corrected for the degree of anemia present.
If the reticulocyte count is elevated, a Coombs test should be performed to aid in identifying the cause of hemolysis. A positive direct Coombs test finding is to be expected in autoimmune hemolytic anemias, hemolytic transfusion reactions, and some drug-induced anemias (eg, those caused by penicillin, methyldopa, some cephalosporins, or sulfonamides).
Lactate dehydrogenase (LDH) levels are elevated in both intravascular and extravascular hemolysis, including the ineffective erythropoiesis that occurs in megaloblastic anemias.
Because the haptoglobin binds free hemoglobin, a low or absent haptoglobin level indicates intravascular hemolysis.
If macro-ovalocytes and hypersegmented neutrophils are noted on peripheral smear, the vitamin B12 level may be low. If folate deficiency is the cause of the macrocytosis, the RBC folate level likely will be decreased. As in vitamin B12 deficiency, peripheral smear may reveal hypersegmented neutrophils and macro-ovalocytes.
Serum total homocysteine levels are almost always elevated in patients with folate deficiency because folate is required in the remethylation step that converts homocysteine to methionine. [24] Serum methylmalonic acid and homocysteine levels are increased early in vitamin B12 deficiency, even before hematologic manifestations or decreases in B12 levels are noted. [25]
Serum unconjugated bilirubin is expected to be elevated in hemolysis.
If vitamin B12 deficiency is the cause of the macrocytosis, the serum vitamin B12 level likely will be decreased. A Schilling test was previously considered the criterion standard for further investigation of a low vitamin B12 level. However, many institutions no longer offer the Schilling test. Alternative tests to the Schilling test are antibodies to parietal cells and intrinsic factor antibodies.
A serum folate level may be obtained, [26] although an RBC folate level is more reliable because it reflects the level over the lifespan of the RBC. However, although macrocytic anemia is a common reason for folate testing, in developed countries with folic acid fortification, most patients with macrocytosis do not have folate deficiency and folate testing may have limited diagnostic benefit. [27, 28]
A study designed to identify the underlying causes of macrocytosis by analyzing hematological features concluded that complete medical histories, analysis of red cell parameters, and peripheral blood smears were simple and inexpensive tools that can be helpful in settings with limited resources. [29]
Bone Marrow Biopsy and Aspiration
Bone marrow biopsy and aspiration are performed to determine whether the marrow is functioning adequately and also may reveal replacement of marrow with tumor, granuloma, or fibrosis. The bone marrow should be obtained before any vitamin B12 or folate therapy or blood transfusion because megaloblastic changes may reverse rapidly.
The bone marrow in megaloblastic anemias is usually hypercellular, with all cell lines proliferating. Marked erythroid hyperplasia may occur to the point at which the myeloid-erythroid ratio is reversed. Nuclear-chromatin dissociation with a young-appearing nucleus and abundant mature-appearing cytoplasm may occur. Granulocytic hyperplasia with giant metamyelocytes and bands is often noted.
Histologic Findings
On peripheral blood smear, large RBCs are evident. Depending on the etiology of the macrocytosis, peripheral smear may reveal nucleated RBCs, target cells, RBC fragments, hypersegmentation of neutrophils, immature WBCs, large platelets, or pancytopenia.
Depending on the etiology of the macrocytosis, the marrow may reveal hypercellularity, megaloblastic changes, fibrosis, infiltration by tumor or granulomatous disease, leukemic changes, or erythroid hyperplasia.
