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Practical of Clinical Hematology

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1 Practical of Clinical Hematology
Lab 2 RBC’s Morphology

2 RBCs Abnormal morphology
Peripheral Blood Morphology

3 Abnormal erythrocyte morphology
Is found in pathological states that may be abnormalities in Red cell distribution. Size (anisocytosis). Hemoglobin content – Color Variation . Shape (poikilocytosis). The presence of inclusion bodies in erythrocyte.

4 Erythrocyte Distribution Abnormalities
Rouleaux formation Stacking of RBCs due to increased plasma proteins coating RBCs (resembling a stack of coins) Found in: - Hyperfibrinogenaemia - Hyperglobulinaemia Agglutination Antibody-mediated Irregular clumping , temperature dependent Found in: - Cold agglutinins - Warm autoimmune hemolysis 2



7 Variation in erythrocyte size (anisocytosis)
Anisocytosis: Variations in size (Microcyte and Macrocyte) Normocytic RBC’s Normal size of RBC (8 μm) with a range of 7 to 9 μm. The nucleus of a small lymphocyte (± 8 µm) is a useful guide to the size of a red blood cell).

8 Microcytic RBC cell smaller than the normal RBC ( <7 μm), and is associated with a decrease in hemoglobin synthesis Found in: Iron deficiency anemia. Thalassaemia. Sideroblastic anemia. Lead poisoning. Anemia of chronic disease.

9 Macrocyte RBC larger than the normal (<9 μm) and is the result of a defect in nuclear maturation or stimulated erythropoiesis. May be round or oval in shape, the diagnostic significance being different. Found in: Folate and B12 deficiencies (oval) Ethanol (round) Liver disease (round) Reticulocytosis (round)

10 Example : Film Study Most erythrocytes presented in the picture are microcytes (compare with the small lymphocyte). The degree of hemoglobinization is sufficient. Normal platelets and single ovalocytes are present. 1. microcyte 2. normocyte

11 Variation in erythrocyte color
A normal erythrocyte has a pinkish-red color with a slightly lighter-colored center (central pallor) when stained with a blood stain, such as Wright. The color of the erythrocyte is representative of hemoglobin concentration in the cell. Under normal conditions, when the color, central pallor, and hemoglobin are proportional, the erythrocyte is referred to as Normochromic.

12 Hypochromia Increased central pallor and decreased hemoglobin concentration, the central pallor occupies more than the normal third of the red cell diameter. Found in: Iron deficiency Thalassaemia any of the conditions leading to Microcytosis

13 Polychromasia Red cells stain shades of blue-gray as a consequence of uptake of both eosin (by hemoglobin) and basic dyes (by residual ribosomal RNA). Often slightly larger than normal red cells and round in shape - round macrocytosis. Found in: Any situation with reticulocytosis – for example bleeding, hemolysis or response to heamatinic factor replacement

14 What Abnormal Results Mean
This test is used to diagnose the cause of anemia. The following are the types of anemia and their causes: Normocytic/ normochromic (NC/NC) anemia is caused by sudden blood loss, prosthetic heart valves, sepsis, tumor, long-term disease or aplastic anemia. Microcytic/ hypochromic anemia is caused by iron deficiency, lead poisoning, or thalassemia. Microcytic/ normochromic anemia results from a deficiency of the hormone erythropoietin from kidney failure. Macrocytic /normochromic anemia results from chemotherapy, folate deficiency, or vitamin B-12 deficiency.

15 Shape Abnormalities of Erythrocytes
Poikilocytosis is the general term for mature erythrocytes that have a shape other than the round, biconcave disk. Poikilocytes can be seen in many shapes.(e.g. Acanthocyte, Spherocytosis,……)

16 Shape Abnormalities of Erythrocytes
Terminology Description Condition Target Cells Central Hemoglobin; target shaped Liver Disease; Thalassaemia, Abnormal Hb; Iron Deficiency Echinocyte Short specula's, equally-spaced Uremia, Hypokalemia, Artifact Acanthocyte Speculated, Irregular Liver disease (Alcohol), Post-spleenoctomy. Spherocyte Spherical, no central pallor HS, immune Hemolytic anemia Shistocyte Fragmented RBC, Helmet cells MAHA, burns Ovalocyte Oval / Elliptical shaped Hereditary elliptocytosis, Megaloblastic anemia. Sickle Cell Bipolar speculated shape “ banana” shaped Hb S-containing hemoglobinopathy Teardrop cell Single elongated extremity Myelophthistic changes Bite cells Irregular gap in membrane G6PD deficiency 2

17 Target cell Red cell with a “target” or bull’s-eye appearance. The cell appears with a central bull’s eye that is surrounded by a clear ring and then an outer red ring. Found in: Obstructive liver disease Severe iron deficiency Thalassaemia Haemoglobinopathies (S and C) Post splenectomy Lipid disorders

18 Spherocytosis: Red cells are more spherical. Lack the central area of pallor on a stained blood film. Found in: Hereditary spherocytosis Immune haemolytic anemia Zieve's syndrome Microangiopathic haemolytic Zieve's syndrome is an acute metabolic condition that can occur during withdrawal from prolonged alcohol abuse. It is defined by hemolytic anemia, hyperlipoproteinaemia (excessive blood lipoprotein), jaundice, and abdominal pain. The underlying cause is liver delipidization. This is distinct from alcoholic hepatitis, which however may be subsequent or copresent. microangiopathic hemolytic anemia (MAHA) :In all causes, the mechanism of MAHA is the formation of a fibrin mesh due to increased activation of the system of coagulation. The red blood cells are physically cut by these protein networks, and the fragments are identical to the schistocytes seen on light microscopy.

19 Stomatocytosis: Red cells with a central linear slit or stoma. Seen as mouth- shaped form in peripheral smear. Found in: - Alcohol excess - Alcoholic liver disease - Hereditary stomatocytosis Hereditary stomatocytosis describes a number of inherited autosomal dominant human conditions which affect the red blood cell, in which the membrane or outer coating of the cell 'leaks' sodium and potassium ions.

20 Ovalocyte : An elongated oval cell. They are a result of a membrane defect. Found in: Thalassaemia major. Hereditary ovalocytosis. Sickle cell anemia

21 Elliptocytosis: The red cells are oval or elliptical in shape. Long axis is twice the short axis. Found in: - Hereditary elliptocytosis - Megaloblastic anemia - Iron deficiency - Thalassaemia - Myelofibrosis Myelofibrosis, is a disorder of the bone marrow. It is currently classified as a myeloproliferative disease in which the proliferation of an abnormal type of bone marrow stem cell results in fibrosis, or the replacement of the marrow with collagenous connective tissue fibers.

22 Schistocyte: Found in:
red cell fragments that are irregular in shape and size. They are usually half the size of the normal RBC; therefore, they have a deeper red color. Found in: DIC Micro angiopathic haemolytic anemia Mechanical haemolytic anemia

23 Blister cell: pre keratocyte
Have accentric hallow area. Resemble a women's handbag and may be called pocket-book cell. Found in: Microangiopathic hemolytic anemia

24 Keratocytes (horn cell)
Part of the cell fuses back leaving two or three horn-like projections. The keratocyte is a fragile cell and remains in circulation for only a few hours. Found in: - Uraemia - Severe burns - EDTA artifact - Liver disease Also called helmet cells Uremia or uraemia (see spelling differences) is a term used to loosely describe the illness accompanying kidney failure (also called renal failure), in particular the nitrogenous waste products associated with the failure of this organ. This is not to be confused with uricemia, or hyperuricemia, a build up of uric acid in the blood.

25 Degmacyte "bite cell" An abnormally shaped red blood cell with one or more semicircular portions removed from the cell margin. These "bites" result from the removal of denatured hemoglobin by macrophages in the spleen. Found In: G-6-PD deficiency, in which uncontrolled oxidative stress causes hemoglobin to denature and form Heinz bodies, is a common disorder that leads to the formation of bite cells

26 Sickle Cells: Sickle shaped red cells Found in: Hb-S disease and trait
Sickle-cell disease (SCD), or sickle-cell anaemia (or anemia, SCA) or drepanocytosis, is an autosomal recessive genetic blood disorder with overdominance, characterized by red blood cells that assume an abnormal, rigid, sickle shape. Sickling decreases the cells' flexibility and results in a risk of various complications. The sickling occurs because of a mutation in the hemoglobin gene.

27 Echinocyte “Burr” (crenation ) cell:
Red cell with 30 or more, short blunt projections which are regularly distributed on their surface Found in: Usually artifactual— the result of slow drying under humid conditions. Sometimes are non - artifactual, indicating uremia or pyruvate kinase deficiency. Hemolytic anemia Uremia. Megaloblastic anemia Cells retain the central pallor. * Echinocytes (also called burr cells) have serrated edges over the entire surface of the cell and often appear crenated in a blood smear. Although often confused with acanthocytes, the projections of the red cell membrane are smaller and much more uniform in shape and distribution in echinocytes.

28 Echinocytes (Burr Cells)

29 Acanthocytosis (Spur Cells):
Red blood cells with irregularly spaced projections, these projections very in width but usually contain a rounded end Found in: - Liver disease - Post splenectomy - Anorexia nervosa and starvation Acanthocytes have only a few spicules of varying size that project from the cell surface at irregular intervals. They appear contracted and dense on stained peripheral blood smears. Spur cells appear to be the extreme form of acanthocytes and are seen in patients with severe liver disease . Spur cells are felt to be acanthocytes remodeled by the spleen, whereby the spicules become more blunt and the associated membrane loss makes the cell more spherocytic (spheroacanthocyte).

30 Acanthocytes (Spur Cells)

31 Dacryocytes (Teardrop):
Resembles a tear and usually smaller than the normal RBC. Found in: Bone marrow fibrosis Megaloblastic anemia Iron deficiency Thalassaemia

32 Envelope form cell Found in Thalassaemia Sickle cell anemia

33 Erythrocyte Inclusions with Wright’s Stain
Composition Appearance Condition Basophilic stippling Precipitated ribosomes Evenly dispersed fine or coarse granules - Lead poisoning Thalassaemia , other anemia. Howell-Jolly bodies DNA in origin Nuclear Fragment Dense, round blue granule Post – Splenectomy Pappenheimer bodies Iron-containing granules Small blue granules in clusters Anemia's Heinz bodies Denatured Hemoglobin Round blue precipitates G6PD Cabot Rings Remnants of Nuclear membrane Reddish-blue thread like rings Severe anemia, Lead poisoning. Organism Small blue inclusion Malaria Babesiosis

34 Howell-Jolly Bodies: Small round cytoplasmic red cell inclusion with same staining characteristics as nuclei Found in: Post splenectomy Megaloblastic anemia

35 Siderotic Granules (Pappenheimer Bodies)
These are iron containing granules in red blood cells that are seen because the iron is aggregated with mitochondria and ribosomes. They appear as faint violet or magenta specks, often in small clusters, due to staining of the associated protein. They are associated with severe anemias and thalassemias. Pappenheimer bodies can be increased in hemolytic anemia, infections and post-splenectomy.

36 Basophilic stippling:
Considerable numbers of small basophilic inclusions in red cells. Found in: - Thalassaemia - Megaloblastic anemia - Hemolytic anemia - Liver disease - Heavy metal poisoning.

37 Heinz Bodies: Represent denatured hemoglobin (methemoglobin - Fe+++) within a cell. With a supravital stain like crystal violet, Heinz bodies appear as round blue precipitates. Presence of Heinz bodies indicates red cell injury and is usually associated with G6PD-deficiency.

38 Heinz Body Preparation
Heinz Body Preparation. RBC are incubated supravitally in new Methylene blue to identify precipitates of oxidatively denatured hemoglobin.

39 Cabot Rings: Reddish-blue threadlike rings in RBCs of severe anemia's. These are remnants of the nuclear membrane or remnants of microtubules and appear as a ring or figure 8 pattern. Very rare finding in patients with Megaloblastic anemia. severe anemia's. lead poisoning. Dyserythropoiesis. A - Cabot ring B - Howell-Jolly body

40 Parasites of Red Cell: Two organisms are have a tendency to invade the RBCs. All 4 species of the malaria parasite will invade RBCs. We will see the Plasmodium of different species in RBCs. Theileria microti (Bebesia microti)

41 Malaria

42 RBCs Abnormal morphology
Depiction of red blood cell morphologies that may appear on a peripheral smear, showing: basophilic stippling, Howell-Jolly bodies, Cabot's ring bodies and Heinz's bodies.

43 Blood Film in Some Cases

44 Normal Peripheral Smear

45 Autoimmune Hemolytic Anemia

46 Hereditary Spherocytosis

47 Hereditary pyropoikilocytosis
Hereditary pyropoikilocytosis (HPP) is an autosomal recessive form of hemolytic anemia characterized by an abnormal sensitivity of red blood cells to heat and erythrocyte morphology similar to that seen in thermal burns. Patients with HPP tend to experience severe haemolysis and anaemia in infancy that gradually improves, evolving toward typical elliptocytosis later in life. However, the hemolysis can lead to rapid sequestration and destruction of red cells. Splenectomy is curative when this occurs. HPP has been associated with a defect of the erythrocyte membrane protein spectrin and with spectrin deficiency

48 Microangiopathic Hemolytic Anemia
microangiopathic hemolytic anemia (MAHA) is a microangiopathic subgroup of hemolytic anemia (loss of red blood cells through destruction) caused by factors in the small blood vessels. It is identified by the finding of anemia and schistocytes on microscopy of the blood film. Schistocytes

49 Sickle Cell Anemia Hb SS

50 Idiopathic myelofibrosis
Myelofibrosis, also known as myeloid metaplasia, chronic idiopathic myelofibrosis, osteomyelofibrosis and primary myelofibrosis is a disorder of the bone marrow. It is currently classified as a myeloproliferative disease in which the proliferation of an abnormal type of bone marrow stem cell results in fibrosis, or the replacement of the marrow with collagenous connective tissue fibers. Dacryocytes

51 Iron Deficiency Anemia
Severe Hypochromia

52 Treated Iron Deficiency Anemia
Mixed Population:

53 Alpha Thalassaemia (a-/--)
Microcytic Hypochromia

54 Morphologic Changes in Liver Disease
Target Cells Spur Cells

55 Hepatorenal Syndrome Burr + Spur Cells
Spur cell hemolytic anemia ,, is a form of hemolytic anemia that results secondary to severe impaired liver function or cirrhosis. Chronic liver disease impairs the liver's ability to esterificate cholesterol, causing free cholesterol to bind to the red cell membrane, increasing its surface area. This condition also creates rough or thorny projections on the erythrocyte named acanthocytes. [1] Burr + Spur Cells

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