Blood cell morphology

BLOOD CELL MORPHOLOGY

1. BLOOD CELL MORPHOLOGY PRESENTED BY MISS TASMIA ZEB

2. INTRODUCTION: Study of morphology of blood cells in a wellspread and well-stained blood film yields invaluable diagnostic information. Therefore the blood film should be examined carefully and systematically. It is preferable that the film should be mounted with a cover glass using a neutral mounting medium. It provides not only good refraction but also preserves the blood film. First it should be examined under low power (x10) objective. This will give an idea of the quality of film and distribution and staining of cells and platelet aggregates. It also gives the idea about rouleaux formation, presence of agglutinates, dimorphic population of cells and presence of some haemoparasites e.g., microfilariae. Then select a suitable area and switch to dry high power (x40) objective. Oil immersion (x100) objective should be reserved for the study of finer details of the cells. There are three types of cells in the blood, RBC, WBC and platelets. Each of these should be studied systematically.

3. MORPHOLOGY OF RED BLOOD CELLS Normal red blood cells appear as circular discs of about 6-8.5 µm in diameter, roughly equal to the size of nucleus of a small lymphocyte. They have bright reddish colour (due to haemoglobin) at the periphery, which becomes pale towards the centre because of the biconcave shape of RBC. The central pale area normally does not exceed one third of the total area of RBC. In a normal blood film RBC lie separately in the central area of the film. RBCs are examined for their distribution, size, shape, colour (Hb content) and inclusions.

4. MORPHOLOGY OF RED BLOOD CELLS Abnormalities in these characters may be artefactual or may arise in disease because of: 1. Changes in plasma proteins and development of antibodies to RBC surface antigens. 2. Abnormal erythropoiesis 3. Inadequate haemoglobin formation 4. Damage to red cells in circulation 5. Increased erythropoiesis

5. ABNORMALITIES OF DISTRIBUTION Rouleaux formation: Rouleaux formation (stacking of RBC on top of each other) is seen when fibrinogen concentration of blood is increased e.g., in infections, pregnancy and tissue necrosis. But it is characteristically seen in conditions with abnormal globulin production e.g., in multiple myeloma. The degree of rouleaux formation is directly proportional to ESR. Agglutination: Agglutination is defined as random aggregation of RBC. These form clusters of varying number of cells. This results from bridging of cells by antibody molecules, particularly IgM, against antigens on surface of RBC circulating in plasma. These may have been produced endogenously (autoantibodies) as in cold haemagglutinin disease or rarely have been introduced from out side e.g., infusion of large amounts of mis-matched plasma. In incompatible blood transfusion agglutinates seen comprise of cells of donor origin.

6. ABNORMALITIES OF SIZE Anisocytosis: If the size of RBC varies, in the same blood film, beyond normal limits, it is termed anisocytosis. It is expressed as + to +++. It is a non-specific feature of several haematological disorders. Microcytosis: When the average size of RBC in a blood film is less than normal it is termed microcytosis. The degree of microcytosis is directly proportional to decrease in MCV. It seldom occurs alone but is usually accompanied with hypochromia. Microcytosis is commonly seen in iron deficiency anaemia and thalassaemia. Sometimes small cells with no central pale area are seen. These usually have normal MCV. These are termed spherocytes.

7. ABNORMALITIES OF SIZE Macrocytosis: When the average size of RBC is more than normal, it is termed macrocytosis. The degree of macrocytosis is directly proportional to increase in MCV . Common causes of macrocytosis are liver disease, megaloblastic anaemia, aplastic anaemia, refractory anaemia, obstructive airway disease, excess of alcohol, treatment with hydroxyurea and hyperglycaemia. In patients whose marrow is responding by increased haematopoiesis and there are lot of polychromatic cells, these appear as macrocytes.

8. ABNORMALITIES OF COLOUR The only true variation in colour is the hypochromia. It results from decreased haemoglobinisation of RBCs, commonly seen in iron deficiency anaemia and thalassaemia. Degree of hypochromia is proportional to MCHC. Leptocytes may appear hypochromic because of flattening. Spherocytes appear hyperchromic because of loss of central pale area and increased thickness of the cell. Macrocytes may also appear hyperchromic because of increased thickness.

9. ABNORMALITIES OF COLOUR Target cells: have a central haemoglobinised area, surrounded by a pale ring and then a peripheral haemoglobinised area. These also result from increased membrane surface due to increase in its cholesterol and phospholipid content. These are characteristically seen in thalassaemias, HbC disease, HbD disease, HbE disease, obstructive liver disease, postsplenectomy and iron deficiency anaemia. If an artefact, then these are confined to only a portion of blood film. Dimorphism: It is the term used when two distinct populations of RBC are seen in the blood film. One population may be normal and the other abnormal, particularly hypochromic microcytic or macrocytic. It is seen in sideroblastic anaemia, when a patient has been transfused or when a patient is receiving haematinics for treatment of deficiency anaemia.

10. ABNORMALITIES OF SHAPE Poikilocytosis: When the shapes of RBCs vary more than expected in normal individuals, in the blood film, it is termed poikilocytosis. RBC of abnormal shape is termed a poikilocyte. Poikilocytosis is also a non-specific feature seen in several haematological disorders, abnormal erythropoiesis, megaloblastic anaemia, MDS, iron deficiency anaemia, thalassaemia, and myelofibrosis. However specific types of poikilocytes are diagnostic of specific disorders. Spherocytes: When RBCs are more spheroidal than normal, these are termed spherocyte. These may result from genetic defects of red cell membrane as in hereditary spherocytosis, because of interaction between Ig or complement coated red cells with macrophages as in immune haemolytic anaemias, ABO haemolytic disease of newborn and from action of certain bacterial toxins e.g., Cl.perfringens. Spherical forms may be seen when anticoagulated blood is allowed to stand for a long time e.g., banked blood.

11. ABNORMALITIES OF SHAPE Elliptocytes and Ovalocytes: About 10% RBC in a normal blood film, particularly at the tail end, appear oval and less commonly elliptical in shape. Their proportion is higher in iron deficiency anaemia, megaloblastic anaemia and myelofibrosis. In iron deficiency these are usually more elongated (pencil cells), whereas in megaloblastic anaemia these are macrocytic as well (oval macrocytes).In myelofibrosis ovalocytes are somewhat pointed on narrow side (tear drop cells). If this shape is seen in vast majority of cells and in central area of the film then the condition is termed Elliptocytosis or Ovalocytosis. This results from a hereditary membrane defect. Stomatocytes: When RBCs have a 'mouth' like slit, these are called stomatocytes. Few stomatocytes are usually seen in normal blood film. Their number is increased in alcoholism, liver disease and Rh null disease. These are numerous in a hereditary membrane defect.

12. ABNORMALITIES OF SHAPE Schistocytes: These are fragmented red blood cells of various shapes and sizes. Large cells from which portions are fragmented some times appear as helmets and are called helmet cells. Schistocytes are increased in conditions like iron deficiency anaemia, megaloblastic anaemia and thalassaemia but are characteristically increased when RBCs are exposed to mechanical trauma. This occurs when RBCs are passing through meshes of fibrin as in DIC, or through narrowed vessels as in microangiopathy or through prosthesis. Echinocytes and Burr cells: Echinocytes or crenated cells have evenly distributed blunt spicules of uniform size on their surface. These are formed if anticoagulated blood is allowed to stand for long periods e.g., over night at room temperature or if the film is prepared on a slide that has fatty material on it or if pH of the blood is raised.These are also seen in patients who have uraemia or have been on cardiopulmonary bypass. Burr cells are also echinocytes but their spicules are reversible

13. ABNORMALITIES OF SHAPE Acanthocytes: These are small densely staining RBC with thorn like projections. Generally the projections are fewer, of varying sizes, variable number and more blunted than echinocytes. These may be hereditary or acquired. Hereditary causes include McLeod phenotype and disorders of lipid metabolism. The acquired causes include spur cell anaemia and chronic liver disease. Pyropoikilocytes: These are seen in a rare hereditary disorder, pyropoikilocytosis, and comprise microspherocytes and fragments of RBC. Their number greatly increases when blood is heated to 45°C. Sickle cells: These are thin, elongated, deeply staining red cells with elongated ends. These may be straight, curved or of various other shapes. These are produced by polymerisation of HbS in sickle cell disease.

14. INCLUSIONS IN RBC Hb crystals: Some abnormal Hb, particularly C and S polymerise to form crystals inside RBC. Polymerisation of HbS gives a distinct shape to RBC, sickle cell. HbS and HbC occurring together polymerise to form straight crystals with parallel sides and one blunt projecting end or multiple crystals projecting from a common centre. HbC crystals are hexagonal with blunt ends. Howell-Jolly bodies: These are small rounded fragments of the nucleus staining reddish-blue to blue-black resulting from incomplete extrusion of the nucleus. These contain DNA and are <1 µm in diameter. These usually occur singly in RBC but may be multiple. Most common cause is splenectomy or splenic atrophy but these are also seen in alcoholism, sickle cell anaemia, and megaloblastic anaemia. Basophilic stippling or punctate basophilia: These are fine to coarse, deep blue to purple, small but multiple inclusions of varying sizes. These represent aggregated ribosomes. These are seen in thalassaemia, megaloblastic anaemia, liver disease, lead poisoning, unstable Hb, pyrimidine 5nucleotidase deficiency and infections.

15. INCLUSIONS IN RBC Pappenheimer bodies: These are small, dark staining, irregular granules composed of haemosiderin occurring near the periphery of the cells. Their presence is related to iron overload. These stain positively with perl’s stain. These are seen in sideroblastic anaemia, dyserythropoietic anaemia and thalassaemia. Cabot rings: This is thin reddish blue, ring like structure occupying varying portion of RBC. It may be twisted to form figure of 8. Its origin is not clear. These are commonly seen in severe anaemia of any type but most commonly in megaloblastic anaemia, lead poisoning and dyserythropoietic anaemias. These may occur alone but are usually associated with punctate basophilia and Howell-Jolly bodies. Parasites: These include malarial parasites and Babesia.