What you can learn from RBC analysis

While patients occasionally present to a primary-care provider with signs and symptoms of anemia, the diagnosis is usually made from screening a complete blood count (CBC) in patients who are asymptomatic.¹ In either case, the next step is to develop an appropriate treatment plan by identifying the cause of the anemia. This can be accomplished most efficiently by categorizing the anemia's morphology.² Casanova et al were able to eliminate the need for exhaustive workups by developing a clinical prediction rule for their obstetric patients based solely on CBC results and serum ferritin levels.³ This approach may offer a cost-effective, time-efficient, and readily employable method for evaluating the majority of anemia cases in the primary-care settings.^4,5

RBC indices and morphology

Anemia can be classified by morphology based on (1) the size of the RBCs, (2) the amount of hemoglobin contained within the RBCs, and (3) the pathologic process causing the anemia.^1,5,6 Laboratory evaluation of the anemic patient begins with consideration of hemoglobin concentrations and hematocrit, while RBC morphology and indices are frequently overlooked. In clinical practice, however, these parameters can be used to guide further testing and establish a diagnosis in anemic patients.⁵

Beginning the evaluation

Once the clinician has the CBC results, the first index to consider is the mean corpuscular volume (MCV). From this information alone, you will be able to categorize the anemia as microcytic (MCV <75 fL), normocytic (MCV within normal range of 75-100 fL), or macrocytic (MCV >100 fL).

The MCV is an easily employable guide to the selection of additional tests. In one study, surveyed physicians regarded MCV as the single most useful RBC index in the evaluation of anemia.2 Respondents reported a low use of other RBC indices, including the RBC count itself, mean corpuscular hemoglobin (MCH), and mean corpuscular hemoglobin concentration (MCHC) data. Despite this finding, the MCHC is a useful index. An elevated MCHC can be a sign of spherocytosis (Figure 1), which may be associated with immune-mediated hemolytic anemia or may be an inherited defect of the erythrocyte cell membrane. Conversely, a low MCHC is often observed in iron deficiency anemia.¹

A reticulocyte count can also be telling. It measures circulating immature erythrocytes.⁷ Any elevation suggests release of immature RBCs into the circulation and is considered an indicator of effective erythropoiesis.^6,8 Conversely, a low reticulocyte count reflects insufficient bone marrow release of RBCs. The normal range of reticulocytes is 0.5%-1.5% of RBC populations.^1,9 Iron deficiency anemia usually presents with a decreased reticulocyte count, indicating inadequate RBC production.^1,6,10

Microcytic anemia: low MCV

Patients with a low MCV are likely to have iron deficiency anemia, although thalassemia and anemia of chronic disease are part of the differential.¹ When the MCV is low, the serum ferritin level can help to distinguish between anemias. A serum ferritin level <20 µg/L is diagnostic of iron deficiency anemia. Patients whose serum ferritin is >20 µg/L should have their serum transferrin receptor level measured to aid in selecting further determinations.^11,12 A low serum transferrin receptor level occurs in acute inflammation, protein deficiency and losses, and anemia of chronic disease. Patients with iron deficiency anemia have increased serum transferrin receptor levels.

The RBC distribution width (RDW) has low specificity as an independent index, but it is useful in conjunction with the MCV when considering the etiology of microcytic anemia.^2,7,8 The RDW is essentially the coefficient of variation in the measured RBC population.¹ A normal RDW (<15%) indicates a homogeneous RBC population. A low RDW would indicate a more homogeneous or clonal RBC population, while an elevated RDW implies a heterogeneous RBC population. For example, iron deficiency anemia presents with a low MCV due to production of microcytic RBCs (Figure 2). However, the RDW is often increased owing to the wide coefficient of variation in RBC size.^4,13 Note that an increased RDW is observed in iron deficiency anemia patients after treatment with iron therapy as newer RBCs are released from the bone marrow. Conversely, a patient with a low MCV that does not respond to iron therapy by production of increased RBCs of normal size would not exhibit an increased RDW. Thus, the value of this index lies in the post-treatment evaluation of anemias. A normal RDW may also be observed in mixed anemias, such as in patients with both iron deficiency (low MCV) and folate deficiency (high MCV).

Iron: a crucial marker

Although serum iron can be measured directly, such measurements have limited usefulness as an independent marker. This is because inflammation and diurnal variation contribute to unstable iron levels. Nevertheless, in the evaluation of microcytic anemias, a decreased serum iron is an important indicator of depleted total iron stores as would be seen in iron deficiency anemia.⁷

Serum ferritin, a storage protein, is a sensitive marker of iron storage.¹ As iron stores are depleted, the ferritin level also decreases.⁷ Ferritin is considered a nonspecific marker as well because it is also an acute phase reactant that may be increased in inflammatory processes.^6,7

The serum transferrin receptor level indicates the amount of iron functionally available in the tissue iron stores.^11,12 Unlike serum ferritin, the serum transferrin receptor level remains in the normal range during inflammatory processes, making it a more specific iron marker in patients with anemia.¹¹

Free erythrocyte protoporphyrin is a heme precursor and an indirect reflection of iron in the heme molecule.¹ Concentrations become elevated when serum iron levels are insufficient for RBC production, as occurs in iron deficiency anemia.⁷ Elevation may also be noted when there is interference in heme production, as happens in lead poisoning.^1,6

Normocytic anemia: normal MCV

In the presence of a normal MCV, the most likely diagnoses are anemia of chronic disease, hemorrhage, renal disease, liver disease, endocrine dysfunction, and hypoplastic marrow.¹ It is important to note that anemia of chronic disease may also present as a microcytic anemia (low MCV) depending on the duration of the primary illness. Similarly, early iron deficiency anemia may initially present as normocytic anemia.

For patients exhibiting a normal MCV, the next step in evaluation would be to assess the CBC findings for polychromasia or other abnormal morphology. Polychromasia dictates the need for a reticulocyte count. An elevated reticulocyte count is a sign of increased erythropoiesis1 and should lead to suspicions of hemolytic anemias.¹⁴

Since the early stages of both microcytic anemias and macrocytic anemias can present with a normal MCV, serum transferrin receptor, serum ferritin or serum B₁₂, and folate levels should be employed when history and physical examination correlate with those etiologies.^11,12

Macrocytic anemia: high MCV

A high MCV may result from hemolysis, primary vitamin B₁₂ or folate deficiency, intrinsic factor deficiency, myelodysplasia, or hepatic dysfunction, as occurs in alcoholism.^1,6 Oval macrocytes are commonly observed in myelodysplasia and vitamin B₁₂ or folate deficiencies (Figure 3).¹ Alcoholic liver disease is typically associated with large target-shaped RBCs.¹ A reticulocyte count and reticulocyte production index (RPI) may
also be helpful in distinguishing the causes.^9,14 Increases in the reticulocyte count and RPI indicate a good response by the bone marrow to anemia. A low reticulocyte count with low RPI indicates that RBC production is ineffective to compensate for the degree of anemia present. These measures will help to identify causes of anemia related to the bone marrow itself, such as iron deficiency, B₁₂ deficiency, and marrow failure.

Beyond iron and RBC indices

The presence of spherocytes on the peripheral smear indicates splenic culling or the extraction of immune complexes from the RBCs in the spleen. The direct antiglobulin test (DAT), formerly known as the Coombs test, detects the presence of antibodies bound to the surface of RBCs.¹ If the DAT is positive, an immune-mediated hemolysis is likely the cause of the anemia.¹⁴ 

Other RBC morphologies, such as target cells and sickled cells in sickle cell diseases, are pathognomonic for specific diseases. The RBC indices, such as MCV and RDW, can be variable in these instances.  

By evaluating the information obtained on the CBC, the etiology of most common anemias can be determined in the primary-care setting.² Categorizing anemia based on the MCV is the first step in a systematic approach to follow-up testing and will lead to a diagnosis in a majority of cases.³ Anemias not diagnosed by this approach can be referred to a hematologist for further evaluation.

Dr. Gunder is an assistant professor in the School of Allied Health Sciences at the Medical College of Georgia in Augusta. 

References
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