SCBM343: Clinical Pathology

“Pathology of RBC and WBC”

RED BLOOD CELL
Niwat Kangwanrangsan, Ph.D.
[niwat.kan@mahidol.ac.th]

Department of Pathobiology, Faculty of Science, Mahidol University 2

272 Rama VI Road, Rachathevi, Bangkok. 10400

Objectives Outline:
• Discuss the morphology and normal value of red blood cell • Morphology and normal value of red blood cell
• Discuss the pathophysiology changes in the quantity of red
• Pathophysiological changes in the quantity of red blood
blood cell
cell
• Discuss the abnormal morphology of red blood cell
• Abnormal morphology of red blood cell
• Categorize and discuss about laboratory test procedures
• Diagnosis of red blood cell disorders
used in the diagnosis of red blood cell disorders
Red Blood Cell (RBC) Erythropoiesis

Factors involved in release of RBC Destruction of RBC

• Hypoxia
• Erythropoietin
• Thyroid hormone
• Androgen
• Growth hormone
• Prostaglandins
 Red cell disorders

A. Anemia - decrease in production

- increase in destruction

B. Polycythemia (erythrocytosis)

A. Anemia
“a reduction in the oxygen-transporting capacity of
blood, which usually stems from a decrease in the red
cell mass to subnormal levels”

Pathology - blood loss (hemorrhage)

- increased red cell destruction (hemolysis)
- decreased red cell production
Anisocytosis Poikilocytosis
“red blood cells with
• “aniso” = unequal abnormal shape”
• red blood cells present in different size
• normally found in patients with anemia,
thalassemia, folic acid or Vit.B12 deficiency
A1: Anemia of blood loss A2: Hemolytic anemia
(hemorrhage)
■ Hereditary spherocytosis
• acute blood loss may results in “normocytic” and
■ Sickle cell anemia
normochromic” (recovery by compensatory rise in erythropoietin)
 reticulocytosis ■ Thalassemia
(acute blood loss >20% of blood volume  hypovolemic shock*)
■ Glucose-6-phosphate dehydrogenase (G6PD) deficiency
• chronic blood loss  iron deficiency anemia
■ Immunohemolytic anemias

■ Malaria

A3: Anemia of diminished erythropoiesis

B. Polycytemia
■ Iron deficiency anemia “increase in red cells per unit volume of peripheral
■ Anemia of chronic disease blood, usually associated with an increase in
hemoglobin concentration”
■ Megaloblastic anemia
■ Aplastic anemia
■ primary polycythemia (autonomous proliferation)
■ Myelophthisic anemia
■ secondary polycythemia (elevated level of
erythropoietin)
 Lab tests for red cell disorders
Complete blood count (CBC)
(blood indicies; Hb, Hct, MCV, MCH,… / Platelet / WBC count / Differential WBC)

Reticulocyte count

Special tests of blood cells

Clotting test

Measures of proteins and other substances

Abnormal morphology of red cells

■ anisocytosis ■ acanthocytes
■ poikilocytosis ■ echinocytes
■ macrocytosis ■ stomatocytes
■ microcytosis ■ tear drop cells
■ hypochromia ■ cigar cells
■ polychromasia ■ bite cells
■ spherocyte ■ red cell rouleaux WHITE BLOOD CELL
■ ovalocyte ■ Howell-Jolly bodies
■ target cells ■ malaria
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Outline: Lineages of Hematopoiesis

 Normal WBC
- blood-forming / hematopoiesis
- typical life span
- structure and function

 Alterations of leukocytes
- quantitative/qualitative leukocyte disorders
- leukocytosis/leukopenia
- morphological abnormality
- common diagnosis

 Summary
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Typical life span of blood cells

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 White blood cell (WBC) count  Total white cell count
manual automated “number of white cells per cubic millimeter of blood”

* reference range  based on age, sex, and ethnic background

Adults: 4.0–10.0 x 103 cells/mm3

Children: 5.0–15.0 x 103 cells/mm3
Newborns: 10.0–30.0 x 103 cells/mm3

[mm3 = L]

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 Differential white cell count  Absolute white cell count

“ratio (%) of each cell type in peripheral blood” “total number of each cell type”
ne - 58.0% = %cell type counted x total white cell count
ly - 33.5%
mo - 6.0%
eo - 2.5%
ba - 0.0%
------------
= 100.0%
------------

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 Neutrophil Eosinophil
• PMN • 12-17 M
• 10-15 M • Nucleus: 2-3 lobes
• Nucleus 2-5 lobes • Cytoplasm: Pink, irregular borders
• Cytoplasm : Pale blue to pink • Granules: Red to orange, round
• Abundant 2o granules • 0-5 % of total WBC count
• 50-70 % of total WBC • Fight parasitic worm,
• Phagocytosis, chemotaxis allergic reactions
• Life span: hrs to 3 days • Life span : 10-12 days

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Basophil Lymphocyte

• 10-14 M • 7-18 M
• Nucleus: 2 lobes • Nucleus : Round to oval, indented
• Cytoplasm : Lavender to colorless • Cytoplasm : Scant to moderate,
• Granules: Deep purple to black, sky blue, vacuoles
irregular • Granules : Few azurophilic
• 0-1% of total WBC count • 20-40% total WBC count
• Release histamine
• CMI (T cell), HMI (B cell)
• Life span: hrs to 3 days
• Life span : days to yrs

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Monocyte Alterations of WBC
• 12-20 M “range from defects in the quality and quantity (leukopenia) to
increase in number of leukocytes (leukocytosis)”
• Cytoplasm : Blue-gray
• Nucleus : Round, A. Quantitative leukocyte disorders
horse-shoe- or kidney-shaped - result from decreased production / increased destruction
• Granules : Fine, ground glass of WBC in circulation
appearance - response to infections

• 3-8% of total WBC count

B. Qualitative leukocyte disorders
• Phagocytosis, transform to mØ “disruption of WBC function”
• Life span : Month - result from congenital or acquired disorders

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A. Quantitative disorders Neutrophilia

(leukocytosis, granulocytosis)
 Leukocytosis
 >7,500 cells/cu.mm (>75%)
- Neutrophilia
- Lymphocytosis  Most common hematologic abnormalities
- Monocytosis  mechanisms
- Eosinophilia, Basophilia - increased production in bone marrow
 Leukopenia - accelerated maturation process
- Neutropenia - increases release from bone marrow
- Lymphocytopenia - redistribution in circulation (“demarginalization”)
- decreases splenic trapping
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Shift to the Left Leukemoid Reaction

• Persistent neutrophilia of 30,000-50,000 /L
• Shift to the left
• WBC immature forms
(increased leukocyte alkaline phosphatase)
• Acute infection

metamyelocytes, myelocytes, band cells

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Neutropenia
 < 2,000 cells/cu.mm.

< 500 cells/cu.mm  severe neutropenia

 reduced granulopoiesis / marrow injury

 Infections: hepatitis, typhoid, malaria

 Marrow aplasia

 Nutrition: Deficit folic acid, vit B12

 Others: SLE, antileukocyte Ab, immunodef.

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Severe granulocytopenia Acute Infection

(Agranulocytosis)
• Neutrophilia :
 secondary to arrested hematopoiesis in bone
marrow or massive cell destruction in circulation -15,000-25,000 cells/cu.mm.  good prognosis
- Increased production (colony stimulating factor)
 marrow suppression by chemotherapeutic drugs
 recurrent and persistent life-threatening infection
• Neutropenia  poor prognosis
(particularly of the respiratory system)
- Alcoholic + malnutrition, marrow aplasia
 septicemia, fever, tachycardia, ulcers in mouth and
colon, sepsis (death within 3-6 days)

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Chronic Infection Eosinophilia
• Neutrophilia • > 450 cells/cu. mm.
> 25,000 cells/cu.mm.  good prognosis • Allergic disorders (type I hypersensitivity)
Shift to the left  poor prognosis • asthma, hay fever, exfoliative dermatitis,
* increased production of colony stimulating factor drug reactions
• Parasitic diseases (particularly metazoan parasites)
• Neutropenia • Neoplasms : myeloproliferative disorders, Hodgkin
disease
Shift to the left  poor prognosis
* septicemia, fungemia

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Basophilia
• > 50 cells/cu.mm.
• Rare
• Response to inflammation and immediate
hypersensitivity reactions
• Myeloproliferative disorders

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Eosinopenia
• Migration of cells into inflammatory site
• May be seen in Cushing syndrome (steroid), and
as a result of stress, shock, trauma, burns
Basopenia
• Found in hyperthyroidism (Graves disease), acute
infection, long-term steroid treatment
•May be seen during ovulation and pregnancy
Monocytosis
• > 800 cells/cu.mm.
• often transient and not related to a dysfunction of
monocyte production
• commonly occur with neutropenia
• Infections (chronic): TB, rickettsia, bacterial endocarditis
(particularly in the late stage and recovery stage)
• Myeloproliferative disorders
• Hodgkin's disease
• can be seen in normal newborn (physiologic)

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Lymphocytosis
• > 5,000 cells/cu.mm.

• Physiologic: infants & young children (4 mo – 4 yr)

• Acute infections: hepatitis, typhoid, CMV

• Chronic infection: congenital syphilis

• Metabolic disorders : thyrotoxicosis

• Malignancy: lymphocytic leukemia

• Infectious mononucleosis ("atypical" lymphocytosis)

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 Lymphocytopenia
• Immunodeficiency: AIDS, agammaglobulinemia (B-cell),
DiGeorge syndrome (T-cell)

• Lymphocyte destruction: steroid, radiation,

chemotherapy, Hodgkin lymphoma, renal failure, heart
“Atypical” lymphocytes failure (elevated cortisol), TB, SLE, aplastic anemia
- Larger, more cytoplasm & nucleoli
- Cytoplasm tends to be indented by
surrounding RBC's
- Associated with infectious mononucleosis
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Pancytopenia Abnormal neutrophils

Nucleus :
• reduction in all cellular components of blood • 0-2 lobes
granulocyte/monocyte/lymphocyte/erythrocyte/megakaryocyte - Pelger Huet anomaly
- Pseudo Pelger Huet
• found in leukemias
anomaly
(progenitor cell undergoes malignant transformation) • > 5 lobes
- Hypersegmented
neutrophils
- Polycyte
Cytoplasm :
• Inclusion bodies
- May-Hegglin anomaly
- Dohle bodies
• Abnormal azurophilic
granules
- Alder Reilly anomaly
- Chediak-Steinberg
Higashi syndrome
- Toxic granulation
• Vacuole

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Hypersegmentation Pelger-Huet anomaly
• PMN >5 lobe, large PMN • Hereditary, autosomal dominant, neutrophils :
• Maturation abnormality - Mostly bilobed in heterozygote (normal function)
• Abnormal DNA synthesis - Unilobate in homozygote (fatal)
• Deficiencies of folic acid & vit B12 - Excessively coarse clumping of chromatin
• Megaloblastic anemias • WBC count is normal
• Myeloproliferative disorders • Mostly bands
• Chemotherapy • Infections, leukemia
• Polycyte : - PMN >5 lobes, normal size • Metastatic cancer
- Recovery phase of infection
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May-Hegglin anomaly Dohle Inclusion Body

• Inherited : rare autosomal dominant, large, • Blue, small, irregularly shaped in cytoplasm (free
prominent Dohle-like bodies ribosomes or RER) of N, E
• Leukopenia • Acquired :
• Thrombocytopenia - Infections
- Trauma, burns
• Giant platelets
- Cancer
• Cyst-like cytoplasmic
- Pregnancy
inclusions - Cytoloxic drugs (poisoning)
- Chemotherapy
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Activated Neutrophil - Dohle Body Chediak (Steinberg) Higashi Syndrome
• Giant azurophilic granules in neutrophil (rare)
• Abnormal lysosomes & leukocyte function
• Autosomal recessive: Chediak-Higashi syndrome
- infections
- albinism, photophobia,
hepatosplenomegaly
- pancytopenia
- lymphadenopathy
- neurologic defects

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Toxic Granulations B. Qualitative disorders

• Increased azurophilic (1o) B1. Defects in cell maturation
granules
• Bacterial infections
• Cytoplasmic vacuolization
• Severe infection, burn, drug

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 B2. Defects in cell activation Common Diagnosis
• Neutrophilia - Infection, inflammation, stress
• Eosinophilia - Allergies
• Basophilia - Polycythemias, allergic reactions
• Monocytosis - Lymphoproliferative disorders
Leukemia
• Lymphocytosis - Infection, immunodeficiencies
Lymphoproliferative disorders
Leukemia

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References:
1. Pathophysiology : The Biologic Basis for Disease in Adults and Children
(2010)
6th Edition, Kathryn McCance, Sue Huether, Valentina Brasher, Neal Rote. Mosby, Elsevier

2. Pathophysiology (2009 )
Ivan Damjanov. Saunders, Elsevier

3. Blood Cells: An Atlas of Morphology (2007)

Gene Gulati, Jaime Caro. American society for Clinical Pathology Press

4. Clinical Hematology Atlas (2013)

4th Edition, Rodak BF and Carr JH. Elsevier
LABORATORY

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Part 1: Blood smear & Differential count

1. blood collection
2. blood film preparation
3. fixation
4. staining (Wright’s or Giemsa’s)
5. observation

count the proportion of WBC

on the smear
 Part 2a: Defects on RBC Morphology
normocytic normochromic red cells
Sample slides

• Slide #2 Thalassemia

• Slide #7 Homozygous Hb CS

• Slide #8 Iron deficiency

• Slide #9 Hereditary ovalosis

• Slide #10 Hereditary spherocyte

• Slide #11 Hereditary acanthocyte

• Slide #xx Malaria

Part 2b: Defects on WBC Morphology
Sample slides

• Slide #16 Megaloblastic anemia (Macro-, hypersegmented)

• Slide #19 Neutrophilia (Drum stick, hypersegmented)

• Slide #20 Eosinophilia

• Slide #21 Lymphocytosis (Atypical lymphocyte)

• Slide #22 Monocytosis (monocyte vacuolization)

• Slide #24 Acute myeloblastic leukemia (Auer rods)

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