2: Red Blood Cells

Section 2


Red Blood Cells




Normal Morphology (Discocytes and Normocytes)


Anemia


Polycythemia


Morphologic Changes Associated with Disease



Rouleaux


Agglutination


Polychromasia


Reticulocytes


Hypochromia


Anisocytosis


Macrocytosis


Microcytosis


Poikilocytosis



Crystalized Hemoglobin


Erythrocytic Ghosts (Lysed Red Blood Cells, Fading Erythrocytes)


Nucleated Red Blood Cells (Metarubricytes and Rubricytes)


Howell-Jolly Bodies


Basophilic Stippling


Siderotic Inclusions (Pappenheimer Bodies)


Heinz Bodies


Distemper Inclusions


Dysplastic Changes


Erythrocytic Parasites



Red Blood Cell Artifacts




Normal Morphology (Discocytes and Normocytes)






Diagnostic Significance:

Discocytes are the expected finding in healthy nonanemic dogs and cats but can be the predominant RBC morphology observed in many disease states as well.



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Anemia






Diagnostic Significance:

Two general mechanisms of anemia, each with several etiologies, exist. The first mechanism is decreased RBC production from bone marrow failure or disease. Differentials could include immune-mediated mechanisms, toxins, drug effects, infectious agents (feline leukemia virus [FeLV], Ehrlichia spp.), endocrine disease, chronic liver and renal disease, and neoplasia (primary bone marrow neoplasia or metastatic neoplasia). The second major mechanism of anemia is increased RBC loss, which could be secondary to immune-mediated RBC destruction, hemolysis (viral, bacterial, or ehrlichial infections; RBC fragmentation; oxidative RBC damage), and acute and chronic blood loss (coagulopathies, trauma, blood sucking endoparasites and ectoparasites).



Next Steps:

When anemia is diagnosed, careful blood smear assessment of RBC morphology and investigation for hemoparasites are important for classifying the anemia and identifying the cause and to assess for evidence of RBC regeneration. Additionally, evaluation of platelet numbers and morphology and the white blood cell (WBC) line may help identify important etiologies of anemia.



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Polycythemia






Diagnostic Significance:

The most common cause of polycythemia in dogs and cats is hemoconcentration secondary to decreased plasma volume (dehydration and endotoxic shock). A mild or moderate transient polycythemia may be secondary to epinephrine release (from excitement or fright), resulting in splenic contraction, especially in the dog. The last major category is from increased RBC production, which may be physiologically appropriate and inappropriate. Appropriate erythroid production is in response to hypoxia primarily secondary to cardiac disease, pulmonary disease, or both. Animals living at high altitudes and some canine breeds (Greyhound and other sight breeds) also have greater hematocrit values. Inappropriate polycythemia may result from inappropriate production of erythropoietin (responsible for stimulating RBC production in the marrow), as from renal cysts or tumors and some other types of neoplasia. Lastly, polycythemia vera (primary bone marrow neoplasia or leukemia) results in increased RBC production independent of erythropoietin.



Next Steps:

When polycythemia is identified, dehydration must first be excluded by physical examination and by checking blood chemistry parameters (total protein concentration, blood urea nitrogen [BUN]) and a urinalysis (urine specific gravity). If polycythemia is not secondary to dehydration, is persistent, and the patient does not live at high altitudes and is not a sight hound breed, then assessment for appropriate or inappropriate causes of increased RBC production should be considered and may include blood smear assessment, measurement of erythropoietin levels, bone marrow cytology, and exclusion of cardiac, pulmonary, and renal diseases.



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Morphologic Changes Associated with Disease



Rouleaux





Diagnostic Significance:

Rouleaux may be seen occasionally in healthy dogs and especially cats. Increased rouleaux formation is occasionally observed in sick or debilitated dogs and cats with increased fibrinogen or globulin concentration. Rouleaux is generally associated with inflammatory conditions but also in some noninflammatory conditions such as lymphoproliferative disorders resulting in increased globulin production. Lipemic blood samples may cause increased rouleaux as well.



Next Steps:

Careful attention should be made to not confuse rouleaux with RBC agglutination. Rouleaux are orderly linear stacks of RBCs, whereas RBC agglutination is formed by grapelike RBC aggregates. To aid in differentiating between rouleaux and agglutination, a saline dilution test is useful. Rouleaux may be easily dissociated by dilution of RBCs in saline, whereas true agglutination persists despite saline dilution (for method, see Appendix). If rouleaux is confirmed, determine plasma or serum total protein, albumin, globulin, fibrinogen values and lipemic index, and investigate potential causes of any abnormalities found.




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Agglutination





Diagnostic Significance:

Agglutination occurs when antibodies on one RBC bind to antigen on other RBCs, forming globular to amorphous, grapelike aggregates of RBCs. When present, RBC agglutination is supportive of immune-mediated hemolytic anemia (IMHA). Agglutination is not observed in most cases of IMHA, but when present, it occurs most commonly with immunoglobulin M (IgM) because of its pentavalent nature. However, extremely heavy IgG antibody coating of RBC membranes may cause agglutination. Agglutination is generally considered diagnostic of IMHA.



Next Steps:

If necessary, agglutination may be differentiated from rouleaux formation by performing a saline dilution (dispersion) test. The presence of RBC agglutination warrants careful assessment of the blood film for other supportive evidence of IMHA such as spherocytes, and also for underlying causes of IMHA such as neoplastic cells and hemoparasites.




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Polychromasia





Diagnostic Significance:

Significant polychromasia indicates a regenerative bone marrow response to anemia. Hemorrhage and hemolysis are the two main causes of regenerative anemia and increased numbers of polychromatophilic RBCs. Remember that polychromasia does not occur instantly after hemorrhage or hemolysis but takes 2 to 4 days to increase the number of polychromatophilic erythrocytes in peripheral blood and may not exceed the reference interval or achieve maximum values for 5 to 7 days. Therefore, early or acute hemorrhagic or hemolytic anemias may have minimal polychromasia. Also, polychromatophilic RBCs are often difficult to differentiate from normochromic RBCs when smears are stained with rapid stains (i.e., Diff-Quik).


In the cat, mild anemias that are regenerative may have minimal or no polychromasia. The lack of obvious regeneration (reticulocytosis) does not rule out either hemorrhage or hemolysis. Potential causes include early hemorrhage or hemolysis, immune-mediated disease directed at RBC precursors, and recent transfusion, as well as anemia caused by multifactorial causes. If polychromasia is not evident in a feline blood smear, a reticulocyte count is recommended, which is a more accurate way to determine regeneration in cats.



Next Steps:

Significant polychromasia may be used to call an anemia regenerative, but the absence of polychromasia should be interpreted cautiously. The anemia may appear nonregenerative in early acute hemorrhagic or hemolytic anemias, and polychromasia may be difficult to recognize with some stains. Thus, if the onset of anemia is unknown, serial complete blood count (CBC) examinations may be needed to evaluate for regeneration. Reticulocyte counts should also be performed in cases where regeneration is questioned.



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Reticulocytes




Distinctive Features:

When immature, anucleate erythrocytes are stained with a supravital stain such as new methylene blue (NMB), the stain penetrates the RBC membrane and binds to the ribosomes, staining them dark blue and causing them to clump, which identifies these cells as reticulocytes.


The dog has only one identifiable form of reticulocytes in peripheral blood, called aggregate reticulocytes. Cats, in contrast, have two: (1) punctate reticulocytes and (2) aggregate reticulocytes. Punctate reticulocytes contain few, small, dotlike (punctate), blue-black structures (small polyribosome aggregates) but no large aggregates of ribosomes. Aggregate reticulocytes contain one or more medium to large, blue-black structures that may appear as a cluster or network of aggregated structures.


The clinical usefulness of punctate reticulocyte counts in cats remains unclear. Many laboratories and all hematology instruments that report reticulocyte counts in cats, count and report only aggregate reticulocytes. Manual reticulocyte counts are done by counting the number of reticulocytes present in 1000 RBCs. The percentage is determined by dividing the number of reticulocytes counted by 10. The absolute reticulocyte count is determined by multiplying the reticulocyte % by the RBC count. The degree of reticulocytosis is best indicated by the absolute aggregate reticulocyte count (see the Appendix for more information).


Canine reticulocytes released from the bone marrow mature rapidly from aggregate reticulocytes to mature erythrocytes in approximately 24 hours. Punctate reticulocytes are present in low numbers in dogs. Feline aggregate reticulocytes released from bone marrow mature within 12 to 24 hours to the punctate reticulocyte stage. Feline punctate reticulocytes may take up to 2 weeks to develop into mature erythrocytes.


Usually, good correlation exists between the degree of polychromasia observed on a blood film and the aggregate reticulocyte numbers (with the exception of cats, see above). Punctate reticulocytes appear the same as mature erythrocytes on a Wright-stained blood film.



Diagnostic Significance:

The effectiveness of bone marrow response to anemia is determined by the number of aggregate reticulocytes in dogs and cats. Regenerative anemias occur secondary to blood loss and IMHA if the bone marrow is healthy. Some heavy metal toxicities, in particular, lead poisoning, result in excessive numbers (relative to the degree of anemia) of reticulocytes and nucleated RBCs (nRBCs). A mild to moderate anemia may be seen typically only in chronic lead poisoning. Also, with lead poisoning, basophilic stippling is noted within mature RBCs stained with Wright stain.



Next Steps:

If the anemia is regenerative (above reference interval), top considerations include blood loss (internal or external hemorrhage) or hemolysis. Note that chronic blood loss anemia may be nonregenerative because of iron deficiency. If the anemia is not adequately regenerative, it should be investigated if adequate time has elapsed for a regenerative response to have occurred. If deemed truly nonregenerative, further testing such as biochemical profile, urinalysis, and bone marrow cytology are indicated.



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Hypochromia





Diagnostic Significance:

Hypochromic RBCs are hemoglobin deficient mature erythrocytes. Increased hypochromia indicates iron deficiency anemia, which occurs secondary to low-grade, chronic blood loss. Iron deficiency anemia may be secondary to blood loss from the gastrointestinal tract (ulcers, neoplasia, parasites); blood loss in urine (urinary tract infection, calculi, or neoplasia); or external blood loss from blood sucking ectoparasites (fleas, ticks). Milk is low in iron; therefore, young, nursing animals are very susceptible to developing iron deficiency.



Next Steps:

Hypochromic RBCs must be differentiated from artifacts. Hypochromic RBCs show a gradual change from red to pale. Artifacts appearing as punched-out centers with a sharp demarcation from dark (red) to light (clear area) are called torocytes. Torocytes are artifacts and do not indicate iron deficiency anemia. Dogs often develop hypochromic RBCs with iron deficiency anemia but feline erythrocytes are less likely to become hypochromic. Significant hypochromia warrants ruling out sources of chronic blood loss. An iron profile may also be considered.



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Anisocytosis





Diagnostic Significance:

Increased anisocytosis occurs when significant numbers of small-diameter RBCs, large-diameter RBCs, or a combination of both are present together with normal-diameter RBCs. Anisocytosis is most often recognized with significant numbers of spherocytes in IMHA and large numbers of macrocytic polychromatophils in regenerative anemias.




Macrocytosis





Diagnostic Significance:

Macrocytosis secondary to reticulocytosis is expected in regenerative anemias. Macrocytosis is abnormal when secondary to feline leukemia virus (FeLV) infection in cats (myelodysplastic syndrome, erythroleukemia) or inherited disorders such as macrocytosis of poodles (poodle marrow dyscrasia).



Next Steps:

When significant numbers of macrocytic RBCs are evident, investigate for a regenerative anemia (expect concurrent polychromasia). In anemic cats with asynchronous regeneration (i.e., large number of nRBCs and absent or minimal polychromasia or reticulocytosis), consider FeLV testing to aid in identification FeLV associated myelodysplastic syndrome. Check for breed-associated disorders as well. Note that agglutination may cause a falsely increased mean corpuscular volume (MCV), suggesting macrocytosis; as aggregated RBCs are sometimes measured as a single RBC, careful examination of the blood film is important.



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Microcytosis





Diagnostic Significance:

Small RBCs may occur with acquired disorders (i.e., iron deficiency anemia, schistocytosis, bone marrow disorders); congenital disorders (i.e., familial dyserythropoiesis of English Springer Spaniels), and hereditary disorders (i.e., microcytosis of Akita and Shiba Inu dogs). Microcytes may be seen in both congenital and acquired liver disease, in particular liver shunts. MCV may be artifactually low if a short blood sample in an ethylenediaminetetraacetic acid (EDTA) tube is evaluated (excess EDTA). Spherocytes are small-diameter cells, which are included in this section, although these cells are not truly microcytic. Spherocytes may appear as small-diameter cells on a blood smear; however, this is caused by their spheric shape, which conserves volume after removal of membrane. Thus, spherocytes are not considered by many to be true microcytes.



Next Steps:

When significant numbers of microcytes are evident on blood smear review, rule out iron deficiency anemia, IMHA, microangiopathy, liver disease, and check for breed-associated disorders.



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Poikilocytes





Diagnostic Significance:

Many different forms of poikilocytes exist, as well as many different causes of poikilocytosis; a few important causes include the following:



The diagnostic significance of poikilocytes depends on which of the many different types are present and, in some cases, is dependent on the numbers of affected RBCs (i.e., low numbers of Heinz bodies may be normally seen in feline blood, but Heinz bodies are not normal in canine blood). Anemia is often associated with poikilocytes and may range from mild to moderate or severe. Some of the more important poikilocytes are discussed individually below.



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Leptocytes: Target Cells (Codocytes) and Folded Red Blood Cells




Diagnostic Significance:

Occasional target cells are normal in the peripheral blood of dogs. Increased numbers of target cells may be observed in blood from dogs with regenerative anemia, and sometimes, polychromatophilic cells appear as target cells, folded cells, or both. Leptocytes may be seen in pathologic states such as iron deficiency anemia, congenital dyserythropoiesis, liver disease, or nephrotic syndrome.

Related posts:

  1. 6: Extracellular Organisms
  2. 4: Platelets
  3. 1: General Assessment
  4. 5: Hematopoietic Neoplasia

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Jul 24, 2016 | Posted by in SMALL ANIMAL | Comments Off on 2: Red Blood Cells

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