Articles on Medical Diseases and Conditions

Tag: Classification of Anemia

  • Anemia

    Although anemia may be defined as a decrease in Hb concentration, it may result from a pathologic decrease in the RBC count. Since mature RBCs are fully saturated with Hb, such a decrease means that total blood Hb value will also be affected.

    Before commencing transfusion therapy or an extensive workup for the etiology of anemia, one should consider the possibility of pseudoanemia Pseudoanemia may be associated with either a Hb value below reference range limits or a drop in Hb level of 2 gm/100 ml (20 g/L) or more from a previous value. Accuracy of the values, especially those below the reference range, should be verified by redrawing the specimen. It is also possible that a patient could have true anemia simultaneously with one of the conditions in the box on this page.

    One frequently overlooked cause of Hb decrease is iatrogenic blood loss due to laboratory test specimens. Several studies document that a surprising amount of blood can be withdrawn, especially in critically ill patients or those with diagnostic problems. Whereas the majority of patients contributed an average of 10-20 ml of blood per day, those in critical care units may average 40-50 ml/day and sometimes even as much as 150-500 ml per patient per day for several days’ time. In some cases this may total more than 1,000 ml during the hospital stay. (The data sometimes include blood withdrawn to clear arterial lines and sometimes not; this blood also may represent a considerable total quantity.)

    Some Conditions That Produce or Contribute to False Anemia
    Overhydration or rehydration of a dehydrated patient
    Specimen obtained with intravenous (IV) fluid running
    Fluid retention
    Pregnancy
    Hypoalbuminemia
    Posture changes (from upright to recumbent)
    Laboratory variation in hemoglobin assay (approximately ± 0.5 gm/dl) or laboratory error

    Classification of Anemia

    Classification of anemia is helpful because it provides a handy reference for differential diagnosis. There are several possible classifications; each is helpful in some respects.

    Anemia may be classified by pathogenesis. According to pathogenesis, three mechanisms may be responsible.

    1.
    Deficiency of vital hematopoietic raw material (factor deficiency anemia). The most common causes of factor deficiency anemia are iron deficiency and deficiency of vitamin B12, folic acid, or both.
    2.
    Failure of the blood-forming organs to produce or to deliver mature RBCs to the peripheral blood (production-defect anemia). This may be due to (1) replacement of marrow by fibrosis or by neoplasm (primary or metastatic); (2) hypoplasia of the bone marrow, most commonly produced by certain chemicals; or (3) toxic suppression of marrow production or delivery without actual marrow hypoplasia, found to a variable extent in some patients with certain systemic diseases. The most common of these diseases are severe infection, chronic renal disease, widespread malignancy (without extensive marrow replacement), rheumatoid-collagen diseases, and hypothyroidism. (These conditions may sometimes be associated with an element of hemolytic anemia.)
    3.
    RBC loss from the peripheral blood (depletion anemia). This is commonly due to (1) hemorrhage, acute or chronic (causing escape of RBCs from the vascular system), (2) hemolytic anemia (RBCs destroyed or RBC survival time shortened within the vascular system), or (3) hypersplenism (splenic sequestration).

    A second classification is based on RBC morphology. Depending on the appearance of the RBC on a peripheral blood smear, Wintrobe indices, or both, anemias may be characterized as microcytic, normocytic, or macrocytic. They may be further subdivided according to the average amount of RBC hemoglobin, resulting in hypochromia or normochromia. (Macrocytic RBCs may appear hyperchromic on peripheral smear, but this is an artifact due to enlarged and thicker cells that, being thicker, do not transmit light through the central portion as they would normally.) The box on this page lists the more common etiologies.

    Investigation of a Patient With Anemia

    Anemia is a symptom of some underlying disease and is not a diagnosis. There always is a cause, and most of the causes can be discovered by a relatively few simple procedures. Knowing the common causes of anemia, getting a good history, doing a thorough physical examination, and ordering a logical sequence of laboratory tests based on what the clinical picture and other findings suggest provide the greatest assistance in identifying which underlying disease is responsible. When anemia is discovered (usually by the appearance of a low Hb or Hct value), the first step is to determine whether anemia really exists. The abnormal result should be confirmed by drawing a second specimen. Then, if the patient is not receiving excess intravenous (IV) fluid that might produce hemodilution, the next step is to obtain a WBC count, differential, RBC indices, reticulocyte count, and a description of RBC morphology from the peripheral smear. It is wise for the physician to personally examine the peripheral smear, because many technicians do not routinely pay much attention to the RBCs. A careful history and physical examination must be performed. To some extent, the findings on peripheral smear and RBC indices (including the RDW, if available) help suggest areas to emphasize:

    1.
    If the RBCs are microcytic, the possibility of chronic blood loss must always be carefully excluded.
    2.
    If the RBCs are macrocytic, the possibility of megaloblastic anemia or reticulocytosis due to acute bleeding must always be investigated.
    3.
    If the RBCs are not microcytic, if megaloblastic anemia is ruled out in patients with macrocytosis, and if the reticulocyte count is significantly elevated, two main possibilities should be considered: acute blood loss and hemolytic anemia. The reticulocyte count is usually 5% or higher in these cases. However, the possibility of a deficiency anemia responding to therapy should not be forgotten.
    4.
    In a basically normocytic-normochromic anemia without significant reticulocytosis and in which either leukopenia or thrombocytopenia (or both) is present, hypersplenism, bone marrow depression, or a few systemic diseases (e.g., systemic lupus erythematosis) are the main possibilities. In patients over age 40 with normocytic-normochromic anemia and without WBC or platelet decrease, myeloma should be considered, especially if rouleaux or other abnormalities are present that are commonly associated with myeloma. The possibility of chronic iron deficiency should not be forgotten, even though the typical RBC morphology of iron deficiency is microcytic-hypochromic. Occasionally, patients with B12 or folate deficiency have an MCV in the upper normal range.
    5.
    Appearance of certain RBC abnormalities in the peripheral blood suggests certain diseases. A considerable number of target cells suggests one of the hemoglobinopathies or chronic liver disease. Marked basophilic stippling points toward lead poisoning or reticulocytosis. Sickle cells mean sickle cell anemia. Nucleated RBCs indicate either bone marrow replacement or unusually marked bone marrow erythropoiesis, most commonly seen in hemolytic anemias. Significant rouleaux formation suggests monoclonal gammopathy or hyperglobulinemia. Spherocytes usually indicate an antigen-antibody type of hemolytic anemia but may mean congenital spherocytosis or a few other types of hemolytic anemia. Schistocytes (burr cells) in substantial numbers are usually associated with microangiopathic hemolytic anemias or with uremia, alcoholism, and hypothyroidism. Macrocytes are frequently produced by reticulocytosis but are also associated with megaloblastic anemias, cirrhosis, chronic alcoholism, hypothyroidism, and aplastic anemia.

    Some Common Causes of Anemia According to RBC Morphology*
    MICROCYTIC
    Hypochromic

    Chronic iron deficiency (most frequent cause)
    Thalassemia
    Occasionally in chronic systemic diseases
    Normochromic
    Some cases of chronic systemic diseases
    (May be simulated by spherocytosis or polycythemia in some patients)

    NORMOCYTIC
    Hypochromic

    Some cases of anemia due to systemic diseases
    Many cases of lead poisoning

    Normochromic

    Many cases of anemia due to systemic disease (most common cause)
    Many cases of anemia associated with pituitary, thyroid, or adrenal disease
    Acute blood loss
    Hemolytic anemia
    Bone marrow replacement or hypoplasia
    Hypersplenism
    Distance-runner anemia (most persons)

    MACROCYTIC
    Hypochromic

    Some cases of macrocytic anemia with superimposed iron deficiency

    Normochromic

    Vitamin B12 or folic acid deficiency
    Malabsorption (vitamin B12 or folic acid)
    Chronic alcoholism
    Reticulocytosis
    Some cases of chronic liver disease and some cases of hypothyroidism
    Myelodysplasia syndromes/aplastic anemia
    Drug-induced

    *All patients with any disease do not fit into any one category.

    Once the basic underlying process is identified, the cause can usually be identified by using selected laboratory tests with the help of history, physical findings, and other diagnostic procedures. In general it is best to perform diagnostic laboratory studies before giving blood transfusions, although in many cases the diagnosis can be made despite transfusion. Blood specimens for the appropriate tests can usually be obtained before transfusion is actually begun, since blood for type and crossmatching must be drawn first. Serum can be saved or frozen for additional studies, if needed.

  • Bone Marrow Aspiration

    Bone marrow aspiration is of help in several situations: (1) to confirm the diagnosis of megaloblastic anemia; (2) to establish the diagnosis of leukemia or multiple myeloma; (3) to determine whether deficiency of one or more of the peripheral blood cellular elements is due to a deficiency in the bone marrow precursors (bone marrow hypoplasia); (4) to document a deficiency in body iron stores in certain cases of suspected iron deficiency anemia; and (5) in certain cases, to demonstrate metastatic neoplasm or some types of infectious disease (culture or histologic sections may be preferred to routine Wright-stained smears).

  • Platelet Count

    The manual procedure that employs a hemocytometer counting chamber and a standard microscope has approximately a 10%-20% error. A some-what similar method that makes use of a phase contrast microscope has a reported error of about 8%. Platelet counting machines can reduce the error even further. Reference values are 150,000-400,000/mm3 (150-400 Ч 109/L) for direct counts.

  • White Blood Cell Count

    White blood cell counts may be done either manually using a hemocytometer or with automated cell counters. The error produced by hemocytometer counts is about 4%-8% but may be higher with inexperienced personnel. Automated counters have approximately 2%-4% error. The machine has the disadvantage that WBC counts more than 100,000/mm3 become increasingly inaccurate unless a dilution is used. In addition, some of the abnormal lymphocytes of lymphocytic leukemia are exceptionally fragile and may be destroyed when the specimen is prepared for a machine count, thus giving a reading falsely lower than the true value. With either hemocytometer or machine counting, nucleated RBCs are counted as WBCs, and a correction must be made on the basis of the percentage of nucleated RBCs (to 100 WBCs) found on the peripheral smear.
    Reference values are most often quoted as 5,000-10,000/mm3(5-10 Ч 109/L). Several studies suggest that 4,500-11,000/mm3 would be more correct. However, there is a significant overlap of normal and abnormal between 4,500-5,000/mm3 and 10,000-11,000/mm3, especially in the latter area. There is some evidence that the normal range for African Americans may be at least 500/mm3 (.5 Ч 109/L) lower than the normal range for Europeans.

  • Reticulocyte Count

    Reticulocytes occupy an intermediate position between nucleated RBCs in the bone marrow and mature (nonnucleated, fully hemoglobinated) RBCs. After the normoblast (metarubricyte) nucleus is extruded from the cell, some cytoplasmic microsomes and ribosomes remain for 1-2 days that are not ordinarily visible on peripheral blood smears using Wright’s or Giemsa stain but that can be seen by using vital staining techniques and dyes such as methylene blue or cresyl blue. The material then is seen microscopically in the form of dark blue dots or thin short irregular linear structures arranged in loose aggregates or reticulum. The reticulocyte count is an index of the production of mature RBCs by the bone marrow. Increased reticulocyte counts mean an increased number of RBCs being put into the peripheral blood in response to some stimulus. In exceptionally great reticulocyte responses, there may even be nucleated RBCs pushed out into the peripheral blood due to massive RBC production activity of the bone marrow. Except in a very few diseases, such as erythroblastosis, peripheral blood nucleated RBCs are usually few in number and of a later maturity stage when they do appear. Reticulocytes are not completely mature RBCs; therefore, when reticulocytes appear in the peripheral blood, they may be slightly larger than normal RBCs and may be sufficiently large to be recognizable as macrocytes. When present in sufficient numbers, these macrocytes may increase the MCV index. Early reticulocytes sometimes appear blue-gray or gray with Wright’s stain in contrast to the red-orange appearance of the normal RBC; this phenomenon is called polychromatophilia and is produced by immature bluish cytoplasmic material to which reddish staining hemoglobin is added. In some conditions a reticulocyte may display small, evenly distributed dotlike aggregates of cytoplasmic ribosomes visible with Wright’s stain, a phenomenon known as basophilic stippling.

    Reference limits for the reticulocyte count are usually considered to be 0.5%-1.5%. Some investigators have reported somewhat higher values, especially in women. There is a substantial problem concerning reproducibility of reticulocyte counts, with statistical variation on the order of ±1 reticulocyte percent unit at normal reticulocyte levels of 0.5%-1.5% standard manual (i.e., statistical variation of more than 50%) and somewhat greater variation at levels of 5% or more. More recently, it has become possible to count reticulocytes using a fluorescent nucleic acid stain in a flow cytometer. This method generally has statistical error less than 15%. Howell-Jolly bodies and medical parasites may interfere if large numbers are present. Some automated cell counters can be adapted to count reticulocytes in the red cell counting channel with preliminary reports suggesting accuracy comparable to flow cytometry. There may be differences in the reference range produced by different equipment and reports.

    Reticulocytes are traditionally reported as a percentage of total RBCs (total RBC includes both mature RBCs and reticulocytes). Automated meter methods are frequently reported as the absolute (quantitative) number of retics. In some clinical situations the number of mature RBCs may decrease while the absolute (total) number of reticulocytes remains the same; this increases the reticulocyte percentage and gives a false impression of increased reticulocyte production. Therefore, some authorities recommend that reticulocyte counts be corrected for effects of anemia. This may be done by multiplying the reticulocyte count (percent) by the patient hematocrit and dividing the result by an average normal hematocrit (47 for men and 42 for women). An alternate method is to obtain the absolute number of circulating reticulocytes by multiplying the patient RBC count by the reticulocyte count (after converting reticulocyte % to a decimal fraction). If polychromatophilic RBCs are present, some experts recommend that the (already) corrected reticulocyte count be divided by 2 to correct for the longer stay of younger reticulocytes in the peripheral blood.

    As noted previously, reticulocyte counts are used as an index of bone marrow activity. Any substantial change in bone marrow RBC production theoretically should be reflected in reticulocyte count change. A normal reticulocyte count has traditionally been considered evidence against a substantial degree of hemolytic anemia and can be used as an index of success in therapy for factor-deficiency anemia. One difficulty is that it usually takes 48-96 hours, sometimes even longer, to establish a reticulocyte count elevation following acute episodes of blood loss, onset of hemolysis, or beginning of factor therapy. Also, reticulocyte counts are not above reference range in some patients with hemolytic anemia (as many as 20%-25% in some studies, depending on the etiology of the disorder), with the degree of reticulocyte response having some correlation with the severity of the hemolytic process. In some cases, failure to obtain expected degrees of reticulocyte response by be due to superimposed factor deficiency (e.g., iron or folate). Another problem may be failure to suspect a hemolytic process or blood loss because the hemoglobin level may remain within population reference range if increased RBC production balances RBC loss or destruction.

    In certain hemolytic anemias such as sickle cell anemia and congenital spherocytosis, temporary aplastic “crisis” may develop in which the anemia worsens because of a halt in RBC production rather than an increase in rate of hemolysis. These crises are sometimes due to parvovirus B-19 infection. The reticulocyte count will become normal or decrease after time is allowed for reticulocytes already in the peripheral blood to disappear.

    In certain anemias caused by ineffective erythropoiesis, the reticulocyte count is normal or decreased unless therapy is given. Some examples include deficiencies of iron, folic acid, vitamin B12, or pyridoxine, or in many patients with anemia associated with chronic disease. In the case of factor deficiency, blood transfusion or hospital diet may contain a sufficient quantity of the deficient factor to increase RBC production.

  • Examination Of Wright-Stained Peripheral Blood Smear

    This procedure gives a vast amount of information. It allows visual estimation of the amount of hemoglobin in the RBCs and the overall size of the RBCs. In addition, alterations in size, shape, and structure of individual RBCs or WBCs are visible, which may have diagnostic significance in certain diseases. Pathologic early forms of the blood cells are also visible. Finally, a good estimate of the platelet count can be made in most cases from the peripheral smear alone (normal is 7-25 platelets per oil immersion field, using 10 x oculars).

    Peripheral Smear Abnormalities

    Hypochromia (an increase in the RBC central clear area) raises the question of chronic iron deficiency.

    Macrocytes in considerable numbers suggest a differential diagnosis of megaloblastic anemia versus the other etiologies of increased MCV (see Table 2-1). Macrocytes in small numbers suggest reticulocytosis as the most likely cause. Some hematologists differentiate between types of macrocytes: oval macrocytes (folic acid or vitamin B12 deficiency; myelodysplasia; myeloid metaplasia) and round macrocytes (alcoholism; cirrhosis; hypothyroidism; reticulocytosis; aplastic anemia).

    Microcytes are small RBCs caused by any of the conditions listed under decreased MCV (see Table 2-2). Hypochromic microcytes suggest a differential diagnosis of chronic iron deficiency versus thalassemia minor or anemia of chronic disease.

    Causes of Spherocytosis
    ABO hemolytic disease of newborn
    Acute transfusion reactions (especially ABO type)
    Hereditary spherocytosis
    Transfused stored bank blood
    Autoimmune hemolytic anemia
    Thermal injury, especially in first 24 hr
    Physical RBC injury (as a component of microangiopathic hemolytic anemia)
    Toxins (Clostridium welchii sepsis and certain snake venoms)
    Hereditary elliptocytosis (10%-20% of cases)
    Occasionally in severe Heinz body hemolytic anemias

    Spherocytes are a type of microcyte in which the cell is round and lacks the central clear area. Spherocytes are a feature of congenital spherocytosis and are also found to varying degrees in certain other conditions.

    Polychromatophilic RBCs (discussed later) are reticulocytes, a sign of markedly increased RBC production, and therefore may be present in patients with acute bleeding, hemolytic processes, hematopoietic and nonhematopoietic malignancies, and factor deficiency anemia responding to therapy (see Reticulocytosis etiology box below). Sometimes a few polychromatophilic RBC may be present without a definite etiology.

    Schistocyte (“broken cell”; also called schizocyte) is a term given to a deformed or broken RBC. In general, the cells are smaller than normal, are misshapen, and have one or more sharp points protruding from the periphery. Names have been applied to various subgroups of misshapen RBCs, such as “burr cell,” “acanthocyte,” and “helmet cell.” Unfortunately, some of these names, especially burr cell, have been applied to different cells by different investigators, so that in most instances it might be preferable to use the noncommittal term, schistocyte. Some conditions associated with schistocytes are listed in Table 2-5.

    Some conditions associated with schistocytes

    Table 2-5 Some conditions associated with schistocytes

    Most Common Causes of Reticulocytosis
    Hemolytic anemia, chronic or acute (antibody induced, drug-induced, associated with abnormalities in Hb or RBC structure, etc.)
    Acute bleeding
    After treatment of vitamin B12/folate/iron deficiency

    Acanthocytes are a subgroup of schistocytes consisting of small spherical cells with several finger-like projections from the RBC surface distributed in an irregular manner. The ends of the projections tend to be slightly thickened. Acanthocytes are typically found in large numbers in hereditary abetalipoproteinemia, in moderate numbers in severe liver disease or in anorexia nervosa, and in small numbers in association with schistocytes of other types in other conditions.

    Red blood cell crenation (echinocytes) are RBCs that appear normal except for uniform small triangular projections arranged in a uniform manner around the circumference of the cell, like the outer edge of a gearwheel. When most of the RBCs have this appearance, they are most commonly artifactual; but in lesser numbers they may be found in liver disease, renal disease, hyperlipidemia, and in some RBC enzymopathies.

    Bite cells (degmacytes) are RBCs with a semicircular defect in one area of the outer edge. When present in significant number, bite cells are suggestive of hemolytic anemia due to an oxidizing agent (Heinz body anemia).

    Sickle cells are crescent-shaped RBCs pointed at one or both ends found in some patients with homozygous sickle cell anemia. Hemoglobin SC disease may sometimes display stubby sickled cells with a short thick bar protruding from the center that represents an Hb C crystal.

    Elliptocytes (ovalocytes) are oval RBCs found in varying numbers in persons with congenital elliptocytosis and occasionally in small numbers in normal persons. When seen on edge, the cells may look somewhat like short rods and, rarely, may superficially resemble an atypical sickle cell.

    Target cells consist of a peripheral ring and central disk of Hb. Target cells are found in large numbers in Hb C disease and in lesser numbers with certain other abnormal hemoglobins, in thalassemia, and in chronic liver disease.

    Teardrop cells look like RBCs in which one side has been gently pulled out to a sharp point while the opposite side is still rounded. These cells are most characteristically associated with myeloid metaplasia (myelofibrosis, Chapter 7) but can also be present in lesser numbers in other myeloproliferative syndromes, such as chronic myelocytic leukemia.

    Stomatocytes are RBCs that have a rectangular or slit-like central pallor configuration. This may be due to hereditary stomatocytosis or may be drug induced. A few stomatocytes may be found in normal persons and in a variety of diseases.

    Rouleaux are RBCs partially adhering to each other with the overall appearance of a partially spread out stack of coins. The RBC central clear area is usually absent. This appearance is similar to that normally seen in the very thick areas of a peripheral blood smear. However, with rouleaux there are a moderate number of free single RBCs intermingled with the RBC stacks, whereas there are no free RBCs in thick areas of the smear. Considerable rouleaux formation suggests the possibility of abnormal serum proteins (such as the monoclonal proteins of multiple myeloma).

    Red Blood Cell Inclusions (Fig. 2-1)

    Basophilic stippling describes a moderate number of small dark blue dotlike structures scattered fairly uniformly throughout the hemoglobinated area of the RBC. Stippling is derived from nuclear remnants, so that the cell represents a reticulocyte and thus may be seen in many of the same conditions as polychromatophilic RBCs. However, stippling is especially associated with lead poisoning.

    Abnormal RBC

    Fig. 2-1 Abnormal RBC. A, normal RBC; B, spherocyte; C, target cell; D, elliptocyte; E, echinocyte; F, sickle cell; G, stomatocyte; H, acanthocyte; I, J, K, L, schistocytes; M, teardrop RBC; N, distorted RBC with Hb C crystal protruding; O, degmacyte; P, basophilic stippling; Q, pappenheimer bodies; R, howell-Jolly body.

    Howell-Jolly bodies are small, round, blue-black inclusions that are considerably larger than basophilic stippling and ordinarily occur only one to an RBC. Howell-Jolly bodies may be present in any severe anemia but are more likely to be seen in severe hemolytic anemias and after splenectomy.

    Pappenheimer bodies are small dark-staining granular inclusions that tend to occur in small numbers, are irregularly distributed, and often occur in small groups. They actually are hemosiderin granules that can be confirmed with ferricyanide iron stains. They are found after splenectomy, in some patients with sideroblastic anemias, and occasionally in patients with severe hemolytic anemia.

    Three types of RBC inclusions cannot be seen with Wright’s or Giemsa stain. All three require supravital staining techniques or other special procedures. Reticulocytes (discussed in detail later) are the stage in RBC maturation just before full maturity. Their number serves as an index of bone marrow RBC production. Hemoglobin H inclusions can sometimes be seen on a reticulocyte preparation as uniformly distributed small round dots somewhat resembling basophilic stippling but of slightly differing sizes. If a reticulocyte is affected, the Hb H inclusions coexist with the more irregular and more linear reticulum structures. Heinz bodies also require a special staining procedure and may need RBC pretreatment with a strong oxidizing agent such as phenylhydrazine. Heinz body formation is most often found in anemias due to RBC enzyme defects, “unstable” hemoglobins (Chapter 5), and certain uncommon hemoglobins such as hemoglobin Koln and Zurich. The Heinz bodies are small, scattered, dotlike structures of varying size in the RBC derived from denatured hemoglobin.

    Limitations of the Peripheral Blood Smear Examination

    The peripheral smear is one of the most useful laboratory procedures in hematology. There obviously are many limitations; for example, a peripheral smear cannot demonstrate the presence of anemia per se, which must be detected by means of either the Hb level, Hct value, or RBC count. Also, many etiologies of anemia are associated with nonspecific peripheral blood changes. In some cases in which the peripheral smear is highly suggestive, it may not be so in early stages of the disease. Even if characteristic cell changes are present, there may be different underlying causes for the same morphologic type of anemia, different causes that call for different treatment. Finally, some conditions produce anemia without any demonstrable morphologic changes in the RBC on the peripheral smear. The same comments about RBCs are also generally applicable to the WBCs of the peripheral smear. However, it is often possible to predict leukocytosis by comparing the overall visual ratio of WBCs to RBCs. A differential count of the various WBC forms is done from the peripheral smear.

  • Cell Counting Instrument Artifacts

    Many laboratories perform Hb, RBC, and WBC determinations on electronic particle counting instruments. In certain cases, artifacts may falsely alter results.
    1.
    When WBC counts are substantially greater than 50,000/cu mm, the Hb, Hct, RBC, MCV, and MCH values may be falsely increased unless corrective measures are taken (the word “increase” as used here means increase from true values, which may or may not place the values outside of reference limits).
    2.
    Peripheral blood nucleated RBCs in substantial numbers will produce false increases of the WBC count unless manually corrected.
    3.
    Marked hyperlipidemia (>2,000 mg/dl of triglyceride) may increase Hb, MCH, and MCHC values.
    4.
    High titers of cold agglutinins may decrease the Hct and RBC count and increase MCV, MCH, and MCHC values. However, not all counting instruments react the same to cold agglutinins.
    5.
    Cryoglobulins may falsely increase the WBC count.
    6.
    Marked erythrocytosis may falsely decrease the RBC count and the Hct value from true levels and falsely elevate MCH and MCHC values.

  • Indices (Wintrobe Indices)

    Mean Corpuscular Volume (MCV)

    Measurement of the MCV uses the effect of the average RBC size on the Hct. If the average RBC size is increased, the same number of RBCs will have a slightly larger cell mass and thus a slightly increased Hct reading; the opposite happens if the average RBC size is smaller than normal. The MCV is calculated by dividing the Hct value by the RBC count.

    There is some disagreement in the literature on MCV reference ranges. Older sources and Coulter Company printed values are approximately :80-94 femtoliters (fL) for men and 81-99 fL for women. More recent reports are in substantial agreement on 80-100 fL for both sexes. Heavy smoking may increase the MCV as much as 3 fL.

    Conditions that increase the MCV are listed in Table 2-1. In my experience, the most common cause of macrocytosis is alcoholism with or without cirrhosis. The major causes for folic acid deficiency are dietary deficiency or malabsorption; for vitamin B12 deficiency, pernicious anemia; and for substantial degrees of reticulocytosis, acute bleeding or hemolytic anemia. Occasionally there are mixed disorders; for example, some patients with alcoholism, malignancy, myxedema, and drug-induced macrocytosis have folic acid deficiency, and some patients with sideroblastic or sideroachrestic anemia have pyridoxine deficiency.

    Some causes of increased mean corpuscular volume (macrocytosis)

    Table 2-1 Some causes of increased mean corpuscular volume (macrocytosis)

    It must be emphasized that a substantial number of patients with any disorder associated with macrocytosis will not display an elevated MCV when first seen by a physician. For example, 10%-20% of patients with megaloblastic anemia (folate or B12 deficiency) have normal range MCV (Table 2-1).

    Conditions that decrease MCV are listed in Table 2-2; the most frequent (in the U.S. population) is chronic iron deficiency. The incidence of decreased MCV in chronic iron deficiency ranges from 27%-76% (averaging about 65%), depending considerably on the degree of deficiency. Thalassemia minor (alpha or beta) comprises about 15% of patients with microcytosis but may be less frequent in some populations. The anemia associated with various chronic diseases (uremia, rheumatoid-collagen diseases, severe chronic infection, etc.) is usually normocytic; but according to the literature, it can be microcytic in about 15% of patients (range, 0%-36%). In my experience, incidence has been 7% (100 patients). Differential diagnosis of these conditions is discussed in the section on chronic iron deficiency.

    Some causes of decreased mean corpuscular volume (microcytosis)

    Table 2-2 Some causes of decreased mean corpuscular volume (microcytosis)

    Some reports in the literature indicate discrepancies when MCV data from microhematocrits are compared with results from automated cell counters such as the Coulter Counter. For example, one report noted that more than 30% of specimens in which the MCV fell below the lower reference range limit of 80 fL by Coulter Counter measurement were still within reference range when microhematocrits were used for the calculation. Another investigator found that macrocytes were reported on peripheral blood smear in only 65% of patients with elevated MCV by Coulter Counter measurement. These studies suggest that MCV values obtained using an automated cell counter are more sensitive to abnormality than other common hematologic parameters. On the other hand, in approximately 10%-20% of patients with an elevated MCV there was no adequate explanation for the abnormality (these patients usually had relatively small elevations, but small elevations do not imply nonsignificance). Also, a patient may have macrocytes in the peripheral blood smear with a normal MCV, since the MCV represents only the average RBC size.

    Mean Corpuscular Hemoglobin (MCH)

    The mean corpuscular hemoglobin (MCH) is based on estimates of the quantity (weight) of Hb in the average RBC. Calculation is done by dividing the blood Hb level by the RBC count. Reference values are 27-31 pg by manual methods and 26-34 pg by Coulter Counter.

    The MCH is influenced by the size of the RBC; a large RBC with normal Hb content will contain a greater weight of Hb than a smaller cell with a normal hemoglobin content. The MCH also depends on the amount of Hb in relation to the size of the cell; a hypochromic cell has a smaller weight of Hb than a normochromic cell of equal size. In general, the MCH level is increased in macrocytosis and decreased in microcytosis and in hypochromia, but there is some variation because of the interplay between the two factors of cell size and concentration of Hb.

    Recent articles have pointed out that MCH values from automated counting instruments closely parallel MCV, significantly more so than by calculation from manual measurements. Therefore, MCH levels from automated cell counters are said to add little if any useful information to that available from the MCV.

    Mean Corpuscular Hemoglobin Concentration (MCHC)

    The MCH concentration (MCHC) estimates the average concentration of Hb in the average RBC. The MCHC depends on the relationship of the amount of Hb to the volume of the RBC. Thus, the MCHC does not depend on cell size alone; a macrocyte with a normal amount of Hb has a normal MCHC. The MCHC is calculated by dividing the Hb value by the Hct value. Reference values are 32%-36% (320-360 g/L) (manual methods) or 31%% (Coulter Counter). Conditions that affect the MCHC are listed in Table 2-3.

    Some conditions that affect the mean corpuscular hemoglobin concentration (MCHC)

    Table 2-3 Some conditions that affect the mean corpuscular hemoglobin concentration (MCHC)*

    Red Blood Cell Distribution Width (RDW)

    Some of the newer electronic cell counting machines are able to sort out RBCs of different sizes and group them according to size (size histogram) as well as calculate the MCV. Normally, most RBCs are approximately equal in size, so that only one gaussian-type histogram peak is generated. Disease may change the size of some RBCs; for example, by fragmentation of RBCs (eg., in hemolysis) or by a gradual process of size change in newly produced RBCs (e.g., in folic acid or iron deficiency). In most cases the abnormal cell population coexists with normal (or at least, less affected) RBCs. The difference in size between the abnormal and less abnormal RBCs produces either more than one histogram peak or a broadening of the normal peak. The cell counting machines can calculate an index of the RBC size differences (anisocytosis) using data from the histogram and the MCV, called the RBC distribution width (RDW). Although the degree of abnormality determines whether or not the index value exceeds index population reference range, in general the RDW is elevated in factor deficiency (iron, folate, or B12), RBC fragmentation, and homozygous hemoglobinopathies (Hb SS, CC, and H) and is normal in thalassemia minor, anemia of chronic disease, and heterozygous trait combinations of abnormal hemoglobins with normal Hb A. The RDW index is never decreased. The RDW (like the MCV) is sometimes abnormal before anemia appears and may be abnormal even before the MCV. Different automated cell counters differ in the way they measure cell size and compute the index, and there may be differences in sensitivity of the index between instruments of different manufacturers and even between different instrument models of the same manufacturer (providing one source of confusion when data are evaluated in the literature and in patient reports). This means that each laboratory should obtain its own RDW reference range and also establish cutoff points for various diseases, which may be very difficult to do since some of the diseases are not common in every part of the country. Also, reports differ in percentage of patients with different diseases who have abnormal RDW (e.g., reports of elevated RDW in untreated pernicious anemia range from 69%-100%). Differentiation between various disorders affecting RBC using MCV and RDW are outlined in Table 2-4 (the diseases listed in each category do not include all patients with that disease).

    Red blood cell distribution width and mean cell volume

    Table 2-4 Red blood cell distribution width and mean cell volume

    FACTORS THAT AFFECT INTERPRETATION OF RED BLOOD CELL INDICES.

    1.
    As an index of RBC hemoglobin, the MCHC was often more reliable than the MCV when manual counting methods were used, because manual RBC counts are relatively inaccurate. Since this is not a problem with automated cell counters, MCHC is not frequently helpful except to raise the question of spherocytosis if the MCHC is elevated. Increase in MCHC is usually limited to relatively severe RBC abnormalities. Elevated MCHC may be a clue to a false increase in MCV and decrease in Hct value due to cold agglutinins or to a false increase in Hb level due to hyperlipemia. However, different counting instruments react differently to cold agglutinins.
    2.
    The MCV, MCH, and MCHC are affected only by average cell measurements either of size or of quantity of Hb. This is especially noticeable in the indices dependent on average RBC size (MCV and, to some extent, MCHC). There may be considerable variation in size between individual RBCs (anisocytosis), but average measurement indices do not reflect this, since they take into account only the average size.
    3.
    Although careful examination of a well-made peripheral blood smear yields a considerable amount of the same information as RBC indices, abnormality may be indicated by one and not by the other, so that the two techniques are complementary.
    4.
    Reference values for Hb, Hct, and indices for infants and children differ from adult values (see Table 37-1). There is some discrepancy in the literature regarding pediatric reference range values, more so than for adult reference ranges. Some of the reasons may be a more limited number of patients and the discrepancy between data derived from manual methods and data derived from automated cell counters.
    5.
    It usually is not necessary to repeat RBC indices for screening or diagnostic purposes after one set of values has been obtained.

  • Hematocrit (Hct)

    After centrifugation, the height of the RBC column is measured and compared with the height of the column of original whole blood. The percentage of RBC mass to original blood volume is the Hct. Anticoagulated whole blood is centrifuged in a special tube. Since whole blood is made up essentially of RBC and plasma, the percentage of packed RBCs after centrifugation gives an indirect estimate of the number of RBCs/100 ml of whole blood (and thus, in turn, is an indirect estimate of the amount of Hb). Hct thus depends mostly on the number of RBCs, but there is some effect (to a much lesser extent) from the average size of the RBC. In most automated cell counting systems the Hct is not measured directly but is calculated from the RBC count value and the mean corpuscular volume (MCV) value obtained from electric pulse height sizing of the RBCs. Reference values are 40%-54% for men and 37%-47% for women. The average error in Hct procedures is about 1%-2%. Microhematocrits are generally as accurate as the older standard Wintrobe (macrohematocrit) technique. The Hct may be decreased when going from upright to recumbent position and increased (1.5%-5.8% units) in the same manner as the Hb by heavy smoking.

    Useful relationships between Hb, Hct, and RBC count include:

    Useful relationships between Hb, Hct, and RBC count

    *at mean corpuscular hemoglobin concentration (MCHC) of 33; this factor varies from 2.7-3.2 depending on the MCHC value.

  • Red Blood Cell (RBC) Count

    The number of RBCs per cubic millimeter gives an indirect estimate of the Hb content of the blood. Manual blood cell counting chamber (hemocytometer) methods give errors of 7%-14% or even more, depending on the experience of the technician. Automatic counting machines reduce this error to about 4%. However, many smaller laboratories do not have these machines. Reference values are 4.5-6.0 million/mm3 (4.5-6.0 Ч 106/L) for men and 4.0-5.5 million/cu mm (4.0-5.5 Ч 106/L) for women.