ANAEMIA – III
Dr GangadharDr Vaishali Jain
Classification of
Anemia
According to
Underlying
Mechanism
Desired Learning Outcomes
1.
2.
3.
4.
5.
6.
7.
Discuss aplastic anaemia in terms of etiopathogenesis, clinical features and
investigations
Briefly describe causes of pancytopenia
Outline the diagnostic approach for normocytic anaemia
Discuss pathogenesis of hereditory spheocytosis and
G6PD deficiency anaemia
Explain thalassemia in terms of pathogenesis, types
clinical features and investigations
Describe sickle cell anaemia with emphasis on pathogenesis,
clinical features and investigations
Explain etiopathogenesis and lab investigations for Paroxysmal
nocturnal haemoglobinuria
Lesson contents
1.
2.
3.
4.
5.
6.
7.
8.
Aplastic anaemia: Etiopathogenesis, clinical features,
investigations
Pancytopenia: Causes
Diagnostic approach for normocytic anaemia
Hereditory spherocytosis
G6PD deficiency anaemia
Thalassemia: Pathogenesis, types, clinical features, investigations
Sickle cell anaemia: Pathogenesis, clinical features, investigations
Paroxysmal nocturnal haemoglobinuria(PNH): Etiopathogenesis,
lab investigations
APLASTIC ANEMIA
• The bone marrow is “empty” (scattered
lymphocytes and a rare hematopoietic precursor)
and the blood shows pancytopenia (cells are of
normal size and shape)
• In bone marrow: No abnormal infiltrate (leukaemia)
or increase in fibrous tissue
• In blood : Not only an anemia, but a
thrombocytopenia and neutropenia
too…………..aplastic anaemia is misnomer
Aplastic anemia : Etiology
•
•
•
•
•
•
Idiopathic (50%)
Drugs (Chloramphenicol, alkylating agents, antithyroid)
Chemicals (Benzene, insecticides)
Radiation; Pregnancy
Viruses (CMV, EBV, hepatitis)
Fanconi anemia (autosomal recessive, charecterized by defect in DNA
repair leading to aplasia)
Pathogenesis
•
•
•
•
Affects quality or quantity of stem cells in bone marrow
Defective haematopoietic microenvironment
Deficiency of factors stimulating haematopoiesis
Inhibition of hematopoiesis by immunological mechanisms
Aplastic anemia : Clinical features
•
•
•
•
Insidious onset
Fatigue and tachycardia (from the anemia)
Petechiae and bruising (from the thrombocytopenia)
Infections (from the neutropenia)
** Lymph nodes, liver or spleen are not
enlarged. If enlarged, diagnosis other than
aplastic anaemia should be considered
Aplastic anemia : Investigations
• Peripheral blood examination: Any 2 of the following are diagnostic

Hb < 10 gm/dl

Neutrophil count <1,500/cmm

Platelet count < 50,000/cmm
•
Normocytic normochromic red blood cells
• Bone marrow examination:
1. Bone marrow aspiration: Dry tap

Hypocellular marrow

Erythroid and myeloid precursors and megakaryocytes – reduced

Abundant lymphocytes and red blood cells
2. Bone marrow biopsy: Necessary for diagnostic confirmation
 Hypocellularity
 Fat cells and sparse haematopoietic element
Bone marrow biopsy
Fat cells
Bony trabeculae
Aplastic anaemia
Aplastic anaemia: Bone marrow biopsy
PANCYTOPENIA : CAUSES
• Aplastic anaemia
• Megaloblastic anaemia
• Infiltration of bone marrow – leukaemia,
lymphoma, storage disorders
• Miliary tuberculosis
• Hypersplenism
Aplastic anaemia Vs Pancytopenia
• Comparing a distinct disease (aplastic anemia) with a
generalized blood finding (pancytopenia)
• Pancytopenia is decrease in all cell lines (white cells,
red cells, and platelets) in the blood
• Pancytopenia indicates that something serious is
going on in the marrow: Either the marrow isn’t
making enough cells, or it’s so full of other stuff (like
the fibrosis)
Bone marrow biopsy : Fibrosis resulting in pancytopenia
Fibrosis
Bony trabeculae
Diagnostic approach
HAEMOLYTIC ANAEMIA
HEREDITARY SPHEROCYTOSIS
• Pathogenesis:
Defective spectrin (a protein which attaches the
red cell membrane to the cytoskeleton)
The red cell membrane is unstable, leading to
increased fragility and the formation of
spherocytes, which get eaten by macrophages
Hereditary Spherocytosis: Peripheral blood smear
Normal RBCs
Spherocytes
• Morphology
• Mild normochromic, normocytic anemia
• Numerous spherocytes
G6PD DEFICIENCY ANAEMIA
• Pathogenesis:
 Glucose-6-phosphate dehydrogenase (G6PD) helps to
reduce free radicals formed during cell metabolism
 Without G6PD, free radicals attack the molecular bonds
between heme and globin, and globin becomes denatured,
forming a little blob called a Heinz body
 The spleen bites out Heinz bodies, leaving bite marks in
the cell
 Oxidizing substance (like a drug, or fava beans) triggers
free radical formation
G6PD deficiency anaemia :Peripheral blood smear
Bite cell
•
•
Schistocytes
Bite cells(caused by recent pitting of Heinz bodies)
Schistocytes (cell fragments)
G6PD deficiency anaemia : Peripheral blood smear
Bite cell
THALASSEMIA
Introduction
Thalassemia is a quantitative abnormality of hemoglobin
Haemoglobin structure
Relative proportion of normal haemoglobin
Adults
Newborn
HbA (α2β2) - 97%,
HbA (α2β2) - 20%
HbA2 (α2δ2) -
2.5%
HbF (α2γ2) - 80%
HbF (α2γ2) -
0.5%
In healthy adult,
• 2β genes encodes 2β globin chains
• 4α genes encodes 2α globin chains
Thalassemia
• Pathogenesis:
 It is inherited single gene disorder
 characterized by quantitative decrease in one of the globin
chains
 In α-thalassemia (deletion of α gene) - decreased amount of α chain—manifest
in fetal and adult life
 In β-thalassemia (point mutation in β gene) - decreased amount of β chain –
manifest in childhood and adult life
• Problem:
1. Decreased hemoglobin production (as decrease in globin chains)
2. Excess unpaired α chains (in β thalassemia) or β, γ, and δ chains
(in α thalassemia), which form tetramers and lead to premature
red cell destruction
β Thalassemia
Type
Genotype
β -Thalassemia β/β+, β/β0
minor
Anaemia RBC
morphology
Hb
Clinical
electrophoresis features
Mild
Hb A2>3.5%;
Hb F < 10%
Hb A- Normal
Microcytic
Hypochromic;
Occ.Target
cells;
Basophilic
stippling
Asymptomatic
β –Thalassemia Intermedia
β -Thalassemia β0/β+, β0/β0 Severe
Major
(Cooley’s
anaemia)
Microcytic
Hb A2 - Normal
Hypochromic;
Hb F = 10-95%
Many target cells; Hb A- reduced
Basophilic
stippling;
Many
normoblasts;
Severe
anisopoikilocytosis
Onset in
infancy;
Splenomegaly
+++; Marked
skeletal &
facial
changes;
Transfusion
dependent
β Thalassemia major: Peripheral blood smear
Normoblast
Target cell
β Thalassemia major: Peripheral blood smear
β Thalassemia major : Clinical features
Malar prominence
Splenomegaly
.
• Malar prominence is due to erythroid hyperplasia in bone marrow
• Splenomegaly due to extramedullary haematopoiesis
β Thalassemia major : Radiological findings
Erythroid hyperplasia in bone marrow result in,
• Skull bones
 Separation of the table and thickening of
the skull vault
 Bony trabeculae develop at right angles
to the tables of the skull bone giving rise
to ‘hair on end’/ ‘brush appearance’
 Medulla - less dense
• Tubular Bones of Extremities
Corterx – thinned
Medulla – less dense
Widening of the marrow cavity
α Thalassemia
Type
Silent
carrier
Genotype
αα/α-
α thalassemia α_/α_,
trait
αα/- -
Hb H disease α-/- -
Hydrops
foetalis
- -/- -
Anaemia RBC
Hb
Clinical
morphology electrophoresis features
Absent
None
Absent/
mild
Microcytic
Hypochromic
Moderate Microcytic
Hypochromic;
Target cells
Severe
Numerous
erythroblasts
Normal
Asymptomatic
Normal
Asymptomatic
Hb H
(β4)
Splenomegaly
Hb Bart’s
(γ4)
Splenomegaly
α Thalassemia: Clinical features
Hydrops foetalis
Thalassemia : Investigations
•
•
•
•
Hb - reduced
MCV; MCH – microcytic hypochromic
RDW – normal (In IDA- RDW increased)
Osmotic fragility test – red cells in thalassemia are resistant to
osmotic lysis
• Hb electrophoresis
• X-ray – skull; long bones
Prenatal diagnosis:
• Chromatography – globin chain synthesis studies
• Foetal DNA analysis – amniocentesis, chorionic villus biopsy
SICKLE CELL ANAEMIA
• Pathogenesis:
 Sickle cell anemia is a qualitative abnormality of
hemoglobin
 It is a type of hemoglobinopathy (diseases with
point mutations in one of the globin chain genes)
 The abnormal hemoglobin (called HbS) in sickle cell
anemia changes shape when it releases oxygen
 The HbS molecules then aggregate and polymerize,
forming the cell into a fragile, non-deformable, sickle
shape .
 glutamic acid is replaced by valine at 6th position in
the globin chains
 Sickle cells bust open more easily; they also stick
together in small vessels, leading to ischemia (often
occurring in hands, feet, lungs, and spleen)
 The spleen may undergo massive enlargement (due
to red cell sequestration) in early childhood
 By early adulthood, however, the spleen is reduced
to a small, fibrotic remnant (due to recurrent
hemorrhage and fibrosis); this is called
“autosplenectomy
Sickle cell anaemia :
clinical features
Leg ulcer
Dactylitis
Sickle cell anaemia : Investigations




Hb : reduced
Peripheral blood examination
ESR: low (inability of red cells to form rouleaux)
Identification of HbS:
1. Sickling test
Addition of reducing agent (2% sodium metabisulphite)
triggers polymerisation of HBS
2. Hb electrophoresis
3. High performance liquid chromatography (HPLC)
Sickle cell anemia : Periphral blood smear
 During crises (acute events)
Sickle cells are seen in the
peripheral blood
 After autosplenectomy
(Post-splenectomy blood picture)
- Nucleated red blood cells,
- Target cells,
- Howell- Jolly bodies, and
- Increased platelet count
Sickle cell anaemia
Sickle cell
Sickling test
Peripheral blood smear
Sickle cell anaemia : Peripheral blood smear
Sickle cells
Paroxysmal nocturnal haemoglobinuria (PNH)
• Acquired haematopoietic stem cell disorder
• Defective stem cells arise in case of aplastic anaemia
• Stem cell in PNH give rise to RBCs, WBCs and platelets
which are abnormally sensitive to complement
• In PNH, cell membrane proteins (CD55, CD59) are
deficient which normally inhibit lytic action of
complement
• Red cell destruction is more at night (as more
complement is activated at night) so haemoglobinuria
is observed in first voided urine in morning
Paroxysma nocturnal haemoglobinuria (PNH)
Laboratory investigations:
 Pancytopenia
 Reticulocyte count – increased
 Haemoglobinuria
 Haemosiderinuria
 Acidified serum test (Ham test) - acidification
of serum activates complement and causes
haemolysis of red cells
 Flow cytometric analysis of membrane proteins
PNH: Investigations
Mechanism of intravascular haemolysis
Mechanism of extravascular haemolysis in macrophages