Objectives

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Overview of Anemia - II
Morey A. Blinder, M.D. Associate Professor of
Medicine and Pathology Immunology
Department of Internal Medicine Division of
Hematology
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Hematology Inpatient Consults (BJH)
Evaluation of anemia (n79)
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13
18
60
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Macrocytic Anemia with Low Reticulocyte Count

• Megaloblastic anemia
• Vitamin B12 deficiency
• Folate deficiency
• Non-megaloblastic macrocytic anemia
• Liver disease
• Hypothyroidism
• Drug-induced (DNA synthesis block)
• Myelodysplastic syndrome

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Folate and Cobalamin Daily Requirements
Diet Vitamin B12 (Cobalamin)
Folate Source Animal products Widespread Bo
dy stores 5 mg 5 mg Daily requirement 2-5
µg 50-200 µg Daily intake 10-20 µg 400-800
µg Dietary deficiency Rare Common
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Metabolic Testing for the Diagnosis ofVitamin
B12 and Folate Deficiency

• High Values
• Normal Vitamin B12 Folate Deficiency
  Deficiency
• Methylmalonic Acid lt 3 gt 95 lt 3
• Homocysteine lt 3 gt 95 gt 95

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Enteric Processing and absorption of Cobalamin
Food-Cbl
Stomach
Peptic digestion
H
Cbl R-binder
R-Cbl
Duodenum
Pancreatic enzymes
Cbl-TC complex
R-Cbl
IF Cb
OH -
Cbl-IF
Distal ileum
IF receptor
Cbl TC
Cbl-IF
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Vitamin B12 Deficiency Common Mechanisms

• Intragastric events
• Inadequate dissociation of cobalamin from food
  protein
• Total or partial gastrectomy
• Absent intrinsic factor secretion
• Proximal small intestine
• Impaired transfer of cobalamin from R protein to
  intrinsic factor
• Usurpation of luminal cobalamin
• Bacterial overgrowth
• Diphylobothrium latum (fish tapeworm)
• Distal small intestine
• Disease of the terminal ileum

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Pernicious Anemia

• Most common cause of vitamin B12 deficiency
• Occurs in all ages and ethnic backgrounds
• Associated with other autoimmune diseases
• Screen for thyroid disease every 1-2 years
• Pernicious anemia is a systemic disease
• Gastrointestinal tract involvement
• Neurologic involvement

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Pernicious Anemia Laboratory Diagnosis

• Anti-intrinsic factor antibodies
• Specific but not sensitive
• Anti-parietal cell antibodies
• Sensitive but not specific
• Schilling test
• Procedure
• Absorption of radiolabeled cobalamin Intrinsic
  factor
• Measure urinary excretion of radioactivity
• Specific but not sensitive

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Megaloblastic anemia
Macro-Ovalocytes
Hypersegmented Neutrophils
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Treatment of Vitamin B12 Deficiency

• Parenteral cobalamin
• 1 mg/day x 7 days
• 1 mg/week x 4 weeks
• 1 mg/month for life
• Oral cobalamin
• 1 mg/day for life

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Normo(macro)cytic Anemia with High Reticulocyte
Count

• Bleeding may have similar laboratory findings as
  hemolysis
• High reticulocyte count may lead to macrocytosis
• Diagnosis is usually ascertained
• Clinical manifestations of long term hemolysis
• Cholelithiasis
• Risk of aplastic crisis (Parvovirus B19)
• Classification
• Hereditary vs. acquired
• Extravascular vs. intravascular
• Immune vs.non-immune

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Hemolytic Anemia with Extravascular Hemolysis

• Extravascular (reticuloendothelial system)
• Hereditary
• Hemoglobinopathies (sickle cell anemia)
• Enzymopathies (G6PD deficiency)
• Membrane defects (hereditary spherocytosis)
• Acquired
• Immune mediated
• Autoimmune hemolytic anemia
• Non-immune mediated
• Spur cell hemolytic anemia
• Paroxysmal nocturnal hemoglobinuria (PNH)

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Glucose-6-Phosphate Dehydrogenase (G6PD)
Deficiency

• G-6-PD reduces NADP/oxidizes glucose-6-phosphate
• Detoxifies free radicals and peroxides

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Glucose-6-Phosphate Dehydrogenase Deficiency

• Sex-linked disorder
• 8 of African-American males
• Hemolytic anemia usually occurs in the presence
  of stress (infection or drugs)
• African form - mild hemolysis usually with drugs,
  infection, fever
• Mediterranean form - more severe
• Unique sensitivity to fava beans
• Chronic non-spherocytic hemolytic anemia

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Acquired Hemolytic Diseases

• Immune mediated hemolytic anemia
• Non-immune mediated

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Autoimmune Hemolytic Anemia

• Warm antibodies (IgG-mediated)
• Primary 45
• Secondary 40
• Lymphoproliferative disease
• Connective tissue disease
• Infectious disease
• Drug-induced 15
• Laboratory testing
• Normocytic/macrocytic anemia
• Peripheral smear spherocytosis

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Anti-Globulin (Coombs) Testing
Direct antiglobulin testing

Anti-C3d Anti-IgG
Patients RBCs
Indirect antiglobulin testing


Patients serum
RBCs
Anti-IgG
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Spherocytes Autoimmune Hemolytic Anemia
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Acquired Hemolytic Anemia with Intravascular
Hemolysis

• Mechanical damage (microangiopathy)
• Chemical damage
• Infection
• ABO incompatibility

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Non-immune hemolytic anemiaMicroangiopathic
Hemolytic Anemia

• Thrombotic thrombocytopenic purpura (TTP)
• Hemolytic uremic syndrome (HUS)
• Disseminated intravascular coagulation (DIC)
• Vasculitis
• Malignant hypertension
• Metastatic neoplasm with vascular invasion
• Preeclampsia/HELLP syndrome of pregnancy

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Schistocytes Microangiopathic Hemolytic Anemia
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Automated Cell Counting Deficiencies

• Abnormalities and inclusions in WBC
• RBC shape abnormalities
• RBC inclusions
• Platelet abnormalities and clumping

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Peripheral blood morphology
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Normal Peripheral Smear
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Normal Peripheral Smear
More information can be gained from examining the
blood smear than from any single hematologic
procedure
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Reticulocyte Polychromasia
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Reticulocyte Manual Count by Supravital Stain
Elevated Count
Normal Count
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Erythrocyte Inclusions with Wrights Stain
Inclusion Composition Appearance
Condition Basophilic Precipitated Evenly
dispersed Lead poisoning stippling ribosomes fine
or coarse granules thalassemia other
anemias Howell-Jolly Nuclear Dense, round
Post-splenectomy bodies fragment blue
granule Pappenheimer Iron-containing Small blue
granules Anemias bodies granules in
clusters Organism Small blue
inclusion Malaria Babesiosis
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Basophilic Stippling
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Howell-Jolly Body
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Malaria
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RBC Inclusions Composite
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Erythrocyte Distribution Abnormalities

• Rouleaux formation Stacking of RBCs due
  to increased plasma proteins coating
  RBCs
• Agglutination Antibody-mediated
  clumping Temperature dependent

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Rouleaux Formation
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Agglutination Reaction
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Variations in RBC Size and Shape

• Anisocytosis Variations in size (e.g.
  microcytes)
• Poikilocytosis Variations in shape (e.g. target
  cells)
• Hypochromia Increased central pallor due to
  decrease in hemoglobin

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Hypochromic Microcytic RBC
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Normal Hypochromic Microcytic
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Hypochromia without Anisocytosis Thalassemia
Trait
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Severe Hypochromia Iron Deficiency Anemia
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Mixed Population Treated Iron Deficiency Anemia
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Microcytic Hypochromia Alpha Thalassemia (?-/--)
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Microcytic Hypochromia Beta Thalassemia Major
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Microcytic Hypochromia Beta Thalassemia Major
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Macrocytic Anemia Macro-Ovalocytes
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Shape Abnormalities of Erythrocytes
Terminology Description Condition Target
cells Central hemoglobin target-shaped Liver
disease thalassemia Abnormal Hgb iron
deficiency Echinocyte Short spicules,
equally-spaced Uremia, hypokalemia,
artifact Acanthocyte Spiculated,
irregular Liver disease (alcohol), Post-s
plenectomy Spherocyte Spherical, no central
pallor HS, Immune hemolytic anemia Schistocyte
Fragmented RBC, helmet cells MAHA,
burns Ovalocyte Oval/elliptical
shaped Hereditary elliptocytosis, Megalob
lastic anemia Sickle cell bipolar spiculated
shape Hgb S-containing banana
shaped hemoglobinopathy Teardrop cell single
elongated extremity Myelophthistic
changes Bite cells Irregular gap in membrane
G6PD deficiency
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Target Cells

• Diagnostic possibilities
• Liver disease
• Hemoglobinopathy
• Thalassemia
• Iron deficiency
• Post-splenectomy
• Lipid disorders

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Echinocytes (Burr Cells)
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Acanthocytes (Spur Cells)
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Morphologic Changes in Liver Disease
Target Cells
Spur Cells
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Hepatorenal Syndrome Burr Spur Cells
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Spherocytes
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Spherocytes Autoimmune Hemolytic Anemia
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Spherocytes Hereditary Spherocytosis
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Schistocytes Microangiopathic Hemolytic Anemia
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Elliptocytes Hereditary Elliptocytosis
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Sickle Cell Anemia Hgb SS
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Hemoglobin SC Disease
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Hemoglobin S-Beta Thalassemia
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Teardrop Cells
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Bite Cells
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Heinz Bodies
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Morphology of Leukocytes

• Normal WBC populations
• Neutrophils (granulocytes)
• Lymphocytes
• Monocytes
• Eosinophils
• Basophils

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Neutrophil
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Eosinophil
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Neutrophil Eosinophil
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Monocytes
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Monocytes
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Small Lymphocyte
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Lymphocytes
Large
Small
Intermediate
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Basophils
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Granulocyte Inclusions or Variants
Terminology Description
Condition Dohle bodies Pale blue areas in
Infections, pregnancy, cancer
neutrophil cytoplasm Toxic Large purple
granules Infection Granulation in
neutrophil cytoplasm Vacuoles Transparent
areas Infection, Toxin in neutrophil
cytoplasm Hyper- 6 nuclear lobes
Megaloblastic anemia segmented Auer rods
Reddish long needle-like Acute myeloid
leukemia inclusions Ehrlichia Blue
inclusions in Ehrlichia sp.
monocytes/neutrophils
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Dohle Bodies
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Toxic Granulation
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Toxic Granulation
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Toxic Granulation and Vacuole Formation
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Hypersegmented Neutrophils
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Auer Rod Acute Myeloid Leukemia
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Ehrlichia
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Myeloid Leukemias and Leukemoid Reaction

• Bone marrow exam is almost always indicated
• Cytogenetic analysis
• Flow cytometry analysis

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Neutrophilia CML
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Pelger-Huet Abnormality
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Acute Myeloid Leukemia M1Myeloblasts without
Differentiation
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Acute Myeloid Leukemia M2Myeloblasts with Some
Differentiation
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Acute Myeloid Leukemia M3 Promyelocytic Leukemia
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Acute Myeloid Leukemia M4 Myelomonocytic Leukemia
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Acute Myeloid Leukemia M5 Monocytic Leukemia
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Acute Myeloid Leukemia M6Erythroleukemia
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Acute Myeloid Leukemia M7Megakaryocytic Leukemia
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Abnormalities of Lymphocytes
Variant Morphologic categories Atypical
lymphs Abundant cytoplasm, RBC
skirting Abnormal lymphs Nuclear
abnormalities i.e. clefts, folds,
notches Plasmacytoid lymphs Abundant
cytoplasm Hairy cells Cytoplasmic
projections Sezary cells Deeply folded
nucleus Prolymphocyte Large lymph with
prominent nucleolus
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Atypical (Reactive) Lymphocytes
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Atypical (Reactive) Lymphocytes
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Plasmacytoid Lymphocytes
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Plasma Cell Plasma Cell Leukemia
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Hairy Cell Hairy Cell Leukemia
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Sezary Cell
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Chronic Lymphocytic Leukemia (CLL)
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CLL Smudge Cells
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CLL Balloon Cells
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Acute Lymphocytic Leukemia L1
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Acute Lymphocytic Leukemia L2
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Acute Lymphocytic Leukemia L3 (Burkitts)