Anaemia

Bill Gilmore

Normal red cell

Bill Gilmore

Red Cell Development

Stem Cells, Haemangioblast, mesenchymal
stem cells, haemopoietic stem cell, CFUGEMM, BFU-E, CFU-E
Early Erythroblast
Intermediate erythroblast
Late erythroblast
Reticulocyte
Mature red blood cell

Red Cell Development

Blasts
Nuclear:cytoplasmic ratio
large
Chromatin is fine and
meshy
Nucleoli present
Perinuclear halo present
(mitochondria)
Membrane highly
refractile
Cytoplasm blue

Changes
Nuclear:cytoplasmic ratio
lowers
Chromatin condenses
Nucleoli disappear
Perinuclear halo
disappears
Membrane become less
refractile
Cytoplasm become
haemoglobinised

The red blood cell

Biconcave disc
7 u in diameter
Lifespan 120 days
No nucleus, mitochrondria or other organelles
Function is to carry O2 from the lungs to the
tissues and CO2 from the tissues to the lungs

Haemoglobin
Protein moiety - in normal adult
haemoglobin A; 2 chains and 2
chains
Haem - iron containing protoporphyrin
ring

Haem Synthesis
Porphobilinogen

Glycine + succinyl CoA

Ala-synthase
- aminolaevulinic acid

Hydroxymethylbilane

Uroporphyrinogen III
Coproporphyri
nogen III

Protoporphyrinogen IX

Protoporpgyrin IX
Fe 2+

Ferrochelatase
Haem

Globin Synthesis
DNA

Transcription

Hn RNA

Splicing

5 cap

AAAA

Processing

5 cap
5 cap

AAAA

AAAA
Translation on cytoplasmic rib
- globin

Chromosome 16

Chromosome 11

Haemoglobin Function
Carries gases around the body.
Can bind O2, CO2, CO, Sulphur,
Cyanide
Methaemoglobin (Fe3+)
Carboxy Hb

Normal Red cell Metabolism

Energy comes from Embden-Meyerhof
Glycolyic Pathway
Produces 2,3 diphosphoglycerate from
Luebering-Rapoport shunt
Produces reducing power from
glycolysis and from pentose phosphate
pathway

Red Cell Membrane

Phospholipid Bilayer
Contains important proteins eg spectrin

Anaemia
Normal range for Haemoglobin
Men 150 (+ 20) g/l
Women 135 (+ 15) g/l

Clinically relevant anaemia =

Haemoglobin value below 105g/l

Aetiology of Anaemia
Decreased proliferation of new RBCs
Low EPO
Damage to Bone Marrow

Impairment of maturation of RBCs

Nutritional deficiency (iron, B12, folate)
Abnormal haem synthesis

Loss or destruction of RBCs

Haemorrhage
Haemolysis

Hypochromic anaemias

Nutritional deficiency of iron

Anaemia of chronic disorders
Sideroblastic anaemia
Iron overload

Figure 1. Note the profound central pallor of the hypochromic and microcytic RBC

Schrier, S. ASH Image Bank 2002;2002:100325

Copyright 2002 American Society of Hematology. Copyright restrictions may apply.

The Haematology Laboratory

General Haematology or Blood Science

Coagulation
Blood Bank
Stem Cell Transplantation
Specialist Haematology
Flow cytometry
Molecular haematology

General Haematology or Blood Science

Automated Blood Cell Counts

Reticulocyte Counts
Platelet counts
Eosinophil counts
Blood Cell Morphology
Blood Science includes automated
analysers for other blood parameters

Blood Specimens
Taken by venepuncture (branch of
Healthcare Science - Phlebotomy )
Without anticoagulant = serum / buffy
coat
With anticoagulant = plasma / cells in
suspension
Defibrinated blood blood taken and
fibrin removed by agitation with glass
beads

Anticoagulants
Ethylene Diamine Tetra Acetic acid
(EDTA) chelates Ca2+ ions, powerful,
very small amount required, can be
heat treated and dried to a negible
volume.
Sodium Citrate Chelates Ca2+ ions,
remains in liquid form used for
coagulation studies
Sodium Oxalate as with citrate

Initial Haematological tests

Full Blood Picture
Blood Cell Morphology differential
white cell count plus microscopial
observations

Normal Full Blood Picture

Microcytic Anaemia

Lowered Hb
Microcytosis MCV lowered
Hypochromasia MCH lowered
Morphology
Confirms microcytosis
Confirms hypochromasia

Full Blood Picture

Micro,Hypo

Macrocytic (Megaloblastic) Anaemia

Lowered Hb
Macrocytosis MCV and MCH raised
Lowered WCC and Platelet count
Morphology
Confirms macrocytosis
Confirms low WCC and platelets
Hypersegmented neutrophils present

FBC- Megaloblastic

Haemolytic Anaemia
Lowered Hb
May be Macrocytosis MCV and MCH
raised
Morphology

Confirms macrocytosis
Macrocytes are polychromasic (reticulocytes)
May contain nucelated Red blood cells
May be evidence of red cell damage

FBC- Haemolytic Anaemia

Microcytic Anaemia

Lowered Hb
Microcytosis MCV lowered
Hypochromasia MCH lowered
Morphology
Confirms microcytosis
Confirms hypochromasia

Further Investigation of Microcytic,

Hyopchromic blood picture

Serum iron
Total iron binding capacity
Serum ferritin
Transferrin receptor assays soluble
useful in the anaemia of chronic disease
Hb electropheresis useful for
thalassaemia
Perls Prussian Blue stain for iron
stores in BM useful for identifying ring

Macrocytic (Megaloblastic) Anaemia

Lowered Hb
Macrocytosis MCV and MCH raised
Lowered WCC and Platelet count
Morphology
Confirms macrocytosis
Confirms low WCC and platelets
Hypersegmented neutrophils present

Further investigation of a
megaloblastic anaemia
Serum B12 microbiological assay,
Euglena gracilis
Serum and red cell folate
microbiological assay, Lactobacillus
casei
Alternatively - A host of competitive
protein binding assays for serum B12
and serum/red cell folate

Investigation of absorption of
B12
Schilling test excretion in urine after
feeding 27Co or 58Co labelled B12
following by a wash out of unlabeled
vitamin. One isotope can be given alone
the other with Intrinsic Factor.
Whole body counting no flushing dose

Investigation of a probable B12

deficiency
Estimation of intrinsic factor in gastric
juice
Intrinsic factor antibidies
Parietal cell antibodies

Response to treatment (Therapeutic

Trial)
Observe the optimal haematological
response after administration of the
haematinic at day 0
Reticulocyte count rises on days 2 and
3 and reaches a peak between days 6
and 7
Red Cell count rises

Haemolytic Anaemia
Lowered Hb
May be Macrocytosis MCV and MCH
raised
Morphology

Confirms macrocytosis
Macrocytes are polychromasic (reticulocytes)
May contain nucelated Red blood cells
May be evidence of red cell damage

Diagnosis of a haemolytic
anaemia
Evidence of increased red cell break
down
Serum bilirubin
Urobilogen
Serum haptoglobin
Haemosiderin
51Cr red cell life span

Type of Haemolysis

Direct antiglobulin test (Direct Coombs)

Osmotic fragility
Haemosiderin and Hb in urine
Plasma Hb

If an hereditary disease is
suspected

Osmotic Fragility
Autohaemolysis + glucose
Red cell instability
Screening test for G6PDH deficiency
Glycolytic enzyme assays PK first
Glutathione
Electrophersis
5HbF
Sickling
Unstale Hb
Gel electropheresis for red cell membrane proteins