HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

251 CLINICAL PATHOLOGY

MODULE 7
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

MODULE OUTLINE
A. HEAMATOLOGY
1. INTRODUCTION TO BLOOD DISORDERS
1. General Introduction
2. Blood formation Haemopoiesis
3. Full haemogram and ESR
2. ANAEMIA INTRODUCTION
a) Classification
b) Effects of anaemia
c) Adaptation to anaemia
d) Pathophyisology
e) Clinical features of anaemia
f) Causes of anaemia
g) Mechanisms of anaemia formation
3. APLASTIC AND DYSHEAMPOIESIS ANAEMIAS
a) Aplastic anaemia
b) Iron (Fe) Deficiency anaemia
c) Megaloblastic anaemia
d) Pernicious anaemia
e) Haemosiderosis and Iron overload

4. HEAMOLYTIC ANAEMIAS
a) Haemolytic mechanism
b) Inherited (Intrinsic) haemolytic

b) Inherited (Intrinsic) haemolytic

anaemia
1. Membranopathies
2. Enzymopathies
3. Haemogloginopathies
c) Sickle cell disease (SCD)
d) Acquired (Extrinsic) haemolytic
anaemia
1. Immune haemolytic anaemia
2. Non-immune haemolytic anaemia
5. HAEMORRHAGIC ANAEMIAS
a) Introduction
b) Causes and mechanism of
haemorrhagic anaemia
1. Platelet disorders
2. Disorders of blood vessels
3. Disorders of coagulation
a.

b.

6. LEUKAEMIA

a) Introduction
b) Acute leukaemia
c) Chronic leukaemia

B. LYMPHORETICULAR SYSTEM
1. LYMPHADENOPATHY
2. SPLEENOMEGALLY
3. LYMPHOMAS
a) Hodgkins Disease
b) Non-Hodgkins Disease
c) Aids Related Lymphomas
d) Burkitts Lymphoma
e) Multiple Myeloma

Page 5 of 154
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

HAEMATOLOGY

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HAEMATOLOGY
INTRODUCTION TO BLOOD DISORDERS
LEARNING OUTCOMES
At the end of the lesson the learner should be competent to: 1. Describe the process of formation of blood cells
2. Describe the components of full haemogram
3. Outline the causes of increased and decreased blood cells
4. Outline factors that affect production of blood cells
5. Outline factors that influence production of erythropoietin
1.0. INTRODUCTION
Haematology is the study of diseases of blood and blood forming organs. The
study of haematology takes into consideration of the blood and bone marrow
entailing the following abnormalities:
1. Abnormalities of the cells
a. Erythrocytes (red blood cells) - RBCs
b. Leucocytes (white blood cells) WBCs
c. Platelets
2. Lymphocytes
3. Disorders of cell proliferation and production.
4. Disorders of clotting and fibrinolysis
2.0. CONCEPTS & PRINCIPLES
Concept

Principles

Blood cells

Formation, Structure and functions

Full heamogram Red cells, White blood cells

Anaemia

Causes, Pathophysiology, Clinical features and investigations

Haemolytic anaemia and haemorrhagic anaemia
Microcytic and macrocytic anaemia

White blood cells

What are the associated disorders.

Platelets

What are the disorders?

3.0. BLOOD FORMATION HAEMOPOIESIS

3.1. Introduction
Haematopoiesis provides the vehicle for the transport of haemoglobin. Normal
erythropoiesis requires the production of an adequate number of erythrocytes at
a rate determined by the bodys needs. The cells should be mature with
adequate content of haemoglobin for oxygen uptake, transport and exchange.
Disorders of erythropoiesis will be diminished production, defective production
and production of erythrocytes with intrinsic defects.

Disorders of erythropoiesis will be diminished production, defective production

and production of erythrocytes with intrinsic defects.
3.2. Requirements
The erythropoietin process requires: 1. An intact and integrated system of enzymes
a. Erythropoietin
b. Thyroxin
c. Androgens
2. Factors for multiplication, maturation and release of erythrocytes
a. Nutrients proteins, carbohydrates
b. Iron
c. Vitamin B12
d. Folate
e. Vitamin B6 (pyrodoxine)
f. Riboflavin
g. Vitamin E
h. Copper
3.3. Sites of Production
A. Sites are derived from a mesenchymal stem cell
a) 1st few weeks of life (gestation) embryogenic sac
b) 6th week the liver
c) 12th 16th week the liver becomes the main site and remains active until a
few weeks before birth. The spleen is a minor organ of erythropoiesis (and
continues until gestation). The thymus and lymph nodes are though to be
a minor role. Production of red cells by the liver persists for 1-2 weeks
post-delivery.
d) From 20th week erythropoiesis commences in the bone marrow and rapidly
increases rapidly during the last trimester.
e) 28th week majority of the production is in the bone marrow
In childhood and infancy all the bone marrow is red marrow.

Diagram 1: Sites of erythropoietic tissue in foetus and throughout life

4.0. FORMATION OF BLOOD CELLS

a) Red cell formation erythropoiesis
b) White cell production granulopoeisis
c) Monocytes/macrophages
d) Platelets
e) Lymphocytes
4.1. Red cell formation (Erythropoiesis)

are the
4.1.1. Development ofWhat
Red Blood
cells

physical
characteristics
of 1) progressive reduction in cell size; 2)
A developing red blood
cell undergoes
progressive reduction in blood?
the size of the nuclei with condensation of chromatin
with cessation of differentiation; 3) loss of cytoplasmic RNA and 4) concurrent
production of haemoglobin.

Maturation of reticulocytes to form erythrocytes normally takes about 48-72

hours of which 24 hours is usually in the circulation. Severe anaemia shortens the
maturation period leading to appearance of immature cells in circulation. Each
day 2 x 1011 erythrocytes enter the circulation after passing the vascular barrier
at a rate dictated by the porosity of sinusoidal wall and/or the ability of the cells
to squeeze through a narrow gap.
The reticulocyte count (R.C) is an informative clinical measurement of
erythropoietic activity which is expressed as a percentage (%) of the total red
cell count. In adults the normal value is 0.5-2.0%. It can be an absolute value of
25-75 x 109/L or reticulocyte index, which is the reticulocyte count (%) divided

cell count. In adults the normal value is 0.5-2.0%. It can be an absolute value of
25-75 x 109/L or reticulocyte index, which is the reticulocyte count (%) divided
by the haematocrit (%).
The number of mitoses reduces in megaloblastic haemopoiesis seen in vitamin B12
and folic acid deficiency. Vitamin B12 and folic acid are responsible for DNA
synthesis hence deficiency leads to formation of megaloblasts, which have large
nuclei with abundant cytoplasm (macrocytes) ending up in reduced red cell
production. Iron deficiency leads to increased mitoses producing small red blood
cells (microcytes).
Diagram 2: Diagrammatic Illustration of erythropoiesis
Pro-erythroblast (Pronormoblast)
- Earliest recognizable cell
Grey

Takes 16 hours, 3 cell divisions

occur, Fe is added
Early normoblast

Basophilic erythroblast
(2 divisions) Takes 16 hours

Blue

Polychromatic erythroblast
(4 divisions)

Takes 16 hours

Late normoblast

Pyknotic nucleus extraction

Hb is synthesized
(8 divisions)

Pink

Reticulocyte
Takes 42-72 hours
Red cell in circulation

4.1.2. Regulation of Erythropoeisis

There has to be a balance between the rate of production of new cells and rate
of destruction. Erythropoiesis is controlled by the amount of oxygen carried by
red cells and end-product-feedback mechanisms. This can be encountered in
situations of anoxia, haemolysis, haemorrhage and high altitude.
Important factors needed are: 1. Erythropoietin finely controls erythropoiesis by determining the rate of
normoblast formation, cell maturation, Hb synthesis and release of red
cells into the circulation.
2. Nutrients - proteins and carbohydrates, iron, vitamin B12 and folate,
vitamin B6 (pyridoxine), riboflavin, vitamin E and copper
3. Hormones androgens and thyroxine

vitamin B6 (pyridoxine), riboflavin, vitamin E and copper

3. Hormones androgens and thyroxine
4.1.3. Erythropoietin production
Increased production of erythropoietin can be physiological or pathological.
1. Physiologically appropriate (compensatory)
a) Hypoxia

b) Anaemia

a. High altitude

b. Cardiovascular shunt
c. Chronic pulmonary disease
d. Massive obesity
a. Haemolytic
b. Haemorrhagic
c. Megaloblastic
d. Iron deficiency
e. Aplastic anaemia

2. Physiologically inappropriate (Pathological)

a) Renal disease

b) Tumours

c) Endocrine
d) Drugs

a. Cysts
b. Hydronephrosis
c. Carcinoma
d. Renal transplant
a. Hepatoma
b. Adrenocortical adenoma
a. Adrenal cortical tumours
a. Corticosteroids
b. Growth hormone
c. Androgens
d. Thyroxine

4.2. White cell production (Granulopoiesis)

The white blood cell precursor is the myeloblast from the stem cell.
Diagram 3: Granulopoiesis
Stem cell

Myeloblast

Myeloblast

Promyelocyte

Myelocyte

Discrete pink granules

Large pink tear drop

to

Purple
Black tear
drop

Neutrophils

myelocyte Eosinophil

myelocyte Basophil

myelocyte

4.3. Monocytes/Macrophages
Monocytes and macrophages are phagocytic cells that develop from monoblasts
and promonocyte
4.4. Platelets
Platelets develop from the stem megakaryocytes within a period of 10 days
4.5. Lymphocytes
Lymphocytes develop from lymphoblasts in the bone marrow with further
replication in the lymph vesicles of the lymph glands.

THE FULL HEAMOGRAM AND ESR

1.0. INTRODUCTION
The full haemogram consists of information on the various cells in plasma.
These are the red blood cells, white blood cells, platelets, inclusions and the
erythrocyte sedimentation rate (ESR).
1. The red blood cells
a. Number
b. Haemoglobin (Hb) content
c. Size and shape
2. White blood cells

3. Platelets

a. Total count
b. Differential count

3. Platelets

a. Number
b. Distribution

4. Erythrocyte sedimentation rate

5. Inclusions
2.0. THE RED BLOOD CELLS (ERYTHROCYTES)
Mature erythrocytes are non-nucleated cells that lack the usual cell organelles.
The normal erythrocyte is a biconcave disc 7.2 m in diameter with a thickness
of 2.4 m at the periphery and 1 m at the centre. The biconcave shape enables
the red cells to easily pass through capillaries whose minimum diameter is 3.5
m. More than 90% of the weight of the erythrocytes is taken up by haemoglobin.
The life span of erythrocytes is 120 days.
Diagram 4: Red Blood Cell

3.0. THE RED BLOOD CELLS INDICES (FEATURES).

The red blood cells have several features that can be described using the red cell
indices which include: a) Primary Measurements
b) Absolute values
c) Morphological changes
d) Hb content variation
3.1. Primary Measurements
The primary red cell measurements include: a) Packed cell volume, PCV (Haemotocrit)
b) Haemoglobin concentration [Hb]
c) Red cell count (RBCs)
3.1.1. Packed cell volume (PCV - Percentage)
Estimation of PCV can be done by: 1. Modern automated blood cells counters where cells are passed through an
aperture of impedance counter or a beam of light of a light scattering
instrument generating an electrical pulse the height which is proportional
to the cell volume.
2. Manual techniques that involve centrifuging of whole blood and estimate

to the cell volume.

2. Manual techniques that involve centrifuging of whole blood and estimate
the proportion of packed cells
The normal values of PCV in females averages 41 + 5(36 - 46) % while that of
males averages 45 + 5(40 - 50) %. Increased PCV is seen when the red cells are
distorted and hence more are trapped in plasma e.g. in spherocytosis, iron
deficiency anaemia, thalassaemia and sickle cell disease (SCD).
3.1.2. Hb concentration [Hb]
Hb estimation can be done by automated and spectrophotometric techniques.
The normal values are: - females average 14.0 + 2.5 (11.5-16.5) gm/dl and
males average 15.5 + 2.5 (13 -18.) gm/dl.

3.1.3. Red cell count (x1012/L)

The normal values of red cell count are females average 4.3 + 0.5 (3.8 4.8) x
1012/L and males average - 5.0 + 0.5 (4.5- 5.5) x 1012/L. Several methods are
used in the process of red cell counting.
a) Automated techniques
In automated techniques the cells are counted as they pass in a stream through
an aperture in impedance or light scattering technology. There are two
techniques namely impedance technique and light scattering.
Impedance technique Wallace Coulter (1956) Coulter Counter
Red cells are poor electrical conductors whereas some diluents are good
conductors and blood is highly diluted in a buffered electrolyte solution. The
blood solution is passed through an aperture tube with a constant source of
direct current electricity between two electrodes.
Blood passes through the aperture displacing some conducting fluid and increase
electrical resistance that changes potential between the electrodes. The height
of the pulse produced corresponds to the cell count.
Light scattering technique
Red cells and other cells can be counted by means of electro-optical detectors. A
diluted cell suspension flows through an aperture so that cells pass in a single
file through the light beam. The scattered light is detected by a photomultiplier
or photodiode that convert it into electrical impulses that are counted and
accumulated. The amount of light scattered is proportional to the surface area
and volume.

3.2. Absolute values

3.2. Absolute values

Absolute values are independent of age, sex and altitude but very minimal
variations may be seen. The absolute values of red blood cells are: 1. Mean cell volume (MCV)
2. Mean cell haemoglobin (MCH)
3. Mean cell haemoglobin concentration (MCHC)
3.2.1. Mean Cell volume (MCV)
The mean cell volume (MCV measures the average volume of the red blood cells.
The normal values are 85 + 8 (78 93) fl (this range indicates normal volume
cells). MCV of less than 83 fl reflects small cells microcytes (microcytic cells)
and that of over 101 fl depicts large cells macrocytes (macrocytic cells)
MCV

CV (%)
P
RBC x 1012/L

PCV x 10-2 x 10-1/L

RBC x 1012/L

PCV
RBC

x 10-15/L

3.2.2. Mean Cell (Corpuscular) Haemoglobin (MCH).

The mean cell (corpuscular) haemoglobin (MCH) measures the average amount of
haemoglobin per cell. The normal values 29.5 + 2.5 (27 32) pg (normal Hb
content normochromes normochromic cells). An MCH of less than 27 means
the red cells have less Hb (hypochromic cells) and that of more than 32 the cells
have excess Hb (hyperchromic cells)
MCH

=
=

Hb gm/dl
RBC x 1012/L

b x 10-12/L
H
RBC

3.2.3. Mean Cell Haemoglobin Concentration (MCHC)

MCHC measures the concentration of Hb in gm/dl of red cells. The normal value
is 33 + 3 (30 36) gm/dl
MCHC = Hb gm/dl x 100
PCV (%)
=

b
H
PCV

gm/dl

3.3. Morphological changes

Red blood cells vary in shape and size. Normally they are biconcave with an

3.3. Morphological changes

Red blood cells vary in shape and size. Normally they are biconcave with an
average diameter of 7.0 micrometers (6.0 8.5) and they stain heavily at the
periphery and lightly at the central (pale). The central pallor is approximately
1/3 of the total surface area of the cell Normochromia.
3.3.1. Changes in shape and size
Abnormal variation in size is called anisocytosis while irregularities in shape are
referred to as poikilocytosis.
3.3.2. Size
Abnormal variation is size the cells can be large (macrocytes) or small
(microcytes)
Macrocytes
Are large abnormal cells seen in: 1. Megaloblastic anaemia
2. Aplastic anaemia
3. Liver disease
Microcytes
Are small cells seen in all forms of anaemia but predominate in iron (Fe)
deficiency and thalassaemia.
3.3.3. Shape
The abnormal shapes are: 1. Poikilocytes are abnormal sized cells seen in severe anaemia
2. Schistocytes are helmet shaped cells with red cell fragments seen as a
result of mechanical injury to the red cells.
3. Spherocytes are red cells with globular shape and reduced diameter. Is a
form of membranopathy and exhibits un-uniform staining of the red cells.
4. Sickle cells sickle shaped cells due to errors of Hb structure

5. Target cells large floppy cells with central area of thickening due to
deficient haemoglobinization or liver disease. They are also seen in SCD
and Thalassaemia
6. Elliptocytes are hereditary oval cells
7. Inclusions abnormal structures that may be iron granules (siderocytes),
Heinz bodies, Basophilic stippling and Howell-Jolly bodies
3.4. Hb content variation

3.4. Hb content variation

The Hb content may vary by increasing or reducing. A reduction in Hb content
can result form.
1. Reduced circulating RBCs (number of red blood cells).
2. Reduced size of red cells
3. Reduced Hb concentration
The reduction in Hb in megaloblastic anaemia is due to reduced number of RBCs
and in Fe deficiency anaemia it results from reduction in cell size, MCH and
number of red blood cells.
Reduction in Hb concentration leads to hypochromia where there is peripheral
staining (ring staining) as evident in Fe deficiency, sideroblastic anaemia,
anaemia of chronic infection due to defective haem synthesis and thalassaemia
due to defective globin synthesis.
Anisochromia is a situation where some cells are normochromic and others are
hypochromic as seen in Fe deficiency anaemia responding to treatment,
following transfusion and sideroblastic anaemia.
Nutritional Requirements for Erythropoiesis
1. Metals - Iron (formation of heam molecule), cobalt and manganese
2. Vitamins - B12 and Folate (biosynthesis of nucleic acid), Vit. C
(facilitate iron turn over in the body) and Vit. B6, E and riboflavin
3. Amino acids for development of globin
4. Hormones - erythropoietin, androgens and thyroxine

4.0. THE LEUCOCYTES (WHITE BLOOD CELLS-WBCS)

4.1. Introduction
There are two main groups of leucocytes namely granulocytes and
agranulocytes. The granulocytes are further grouped as neutrophils, eosinophils
and basophils while the agranulocytes are monocytes and lymphocytes.
These groups of cell differ because of their precursor cells, morphology and
functions. The normal WBC count is 4.0 11.0 x 109/L while the differential
count is outlined in the table below.
Table 1: WBC Count
Cell type
Neutrophils
Lymphocytes
Monocytes
Eosinophils
Basophils

Number (x 109/L)
2.0 7.0
.0 3.0
1
0.2 1.0
0.02 0.5
0.02 0.1

ercentage (%)
P
40 80%
20 40%
2 10%

1 6%
< 1 - 2%

Basophils

0.02 0.1

< 1 - 2%

4.2. Granulocytes
4.2.1. Introduction
Granulocytes originate form the bone marrow and comprise of three groups of
cells namely the neutrophils, eosinophils and basophils. Neutrophils are chief
cells in inflammation where they phagocytose and digest foreign bodies,
microorganisms, damaged tissues and dead cell where eosinophils are increased
in atopic hypersensitivity and parasitic infestation where they play a defensive
role. Increased WBC count above normal is termed as leucocytosis and reduced
below normal is leucopenia.
Leucocytosis is accompanied by increased number of young cells in the
circulation (myelocytes and metamyolocytes) a situation described as a Shift to
the Left. This reactive neutrophils leucocytosis is physiological (compare with
chronic granulocytic leukaemia CGL). Toxic granulation is seen in severe
infections or toxic states where there is morphological damage of neutrophils. A
Shift-to-the-Right there are more mature cells in circulation than the young
ones.

A leukeamoid reaction is a situation where there is outpouring of immature

forms of leucocytes (myelocytes, promyelocytes and myeloblasts) seen in severe
chronic infections e.g. haematogenous tuberculosis, severe haemolysis and solid
cancers of the breast, kidney and lungs.
Causes of Leucocytosis

TASK
What are the causes of
leucocytosis?
Causes of Leucopenia

10-1 - deci,
10-2 - centi,
10-3
-6
- milli,
10 - micro,
10-9
-nano,
10-10
-pico,
-15
-18
10 -femto,
10 -alto
4.2.2. Neutrophils
granulocytes

1. Infections
2. Drugs
3.
Megaloblastic
anaemia
4. Hyperspleenism
5.
Leucoerythroblastic
anaemia
6. Acute leukaemia
7. Aplastic anaemia

Neutrophils have a size of 10-20 micrometers with a segmented nucleus of 2-5

lobes and their granules stain faintly pink with Romanwsky dyes. Production of

Neutrophils have a size of 10-20 micrometers with a segmented nucleus of 2-5

lobes and their granules stain faintly pink with Romanwsky dyes. Production of
neutrophils is controlled by injury and infection. Neutrophils ingest and kill
bacteria by opsonization and phagocytosis with the assistance of IgA. Alteration
of blood flow and mediators of inflammation influence the movement of the
neutrophils in coordination with other factors.
Diagram 5: Neutrophil

1. Agranulocytosis
Agranulocytosis is a situation with marked neutropenia and severe infection of
drug induced origin. The mechanism is immunological in nature. It may also be
seen in infectious mononucleosis, mycoplasmal pneumonia and systemic lupus
erythromatosus (S.L.E). The risk is the chances of infection even with the normal
commensals like the gut coliforms and fungi.
2. Neutrophil Leucocytosis (Neutrophilia)
Neutrophilia is increased neutrophils in circulation above 7.0 x 109/L
Causes
a) Bacterial infarction commonest cause
b) Tissue necrosis without infection
a. Myocardial infarction
b. Burns and crush injury
c. Cancer (rapid)
c) Excessive haemorrhage
d) Acute haemolysis
e) Drug reactions e.g. with steroids
3. Neutrophil Leucopenia (Neutropenia)
Neutropenia is reduced number of neutrophils in circulation and can be part of
pancytopenia.
Causes:
1. Bone marrow failure (BMF)
a.

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Cause
defective
production
(predictable
e.g.
chloramphenicol;
idiosyncratic
reactions
e.g.
phenylbutazone, frusemide, chlorpromazine, phenothiazines,
sulphonamides
and
antithyroid
drugs;
ineffective
myelopoiesis e.g. chloramphenicol and phenytoin).
Immune destruction

Combination of the two mechanisms (Analgesics

phenylbutazone; Antimicrobials tetracycline streptomycin,
novobiocin,
chloramphenicol;
Anticonvulsants;
Antihistamines

chlormpheniramine,
promethazine,
mepyramine; Antithyroids thiouracil, carbimazole;
Antidiabetic agents tolbutamide, chlorpropamide;
Tranquillisers; Miscellaneous barbiturates, metronidazole,
PAS, penicillamine, phenindione)

2. Overwhelming infection
3. Excessive peripheral destruction e.g. drugs, antibodies and hyperspleenism.
4. Excessive consumption e.g. in anaphylactic shock.
4. Defects of Neutrophil Function
The integrity of the function of neutrophils depends on response of chemostatic
stimuli, phagocytic activity and intracellular destruction.

The integrity of the function of neutrophils depends on response of chemostatic

stimuli, phagocytic activity and intracellular destruction.
Disorders of chemotaxis and opsonization
Disorders of neutrophil chemotaxis and opsonization presents with: - recurrent
infections (bacteraemia) with Strep, pneumonia, Neisseria, Haemophilus
influenzae and Pseudomonas and recurrent pulmonary infections and meningitis.

Disorders of locomotion and ingestion

Disorders of locomotion causes the Lazy leucocyte syndrome and
corticosteroids, exercise and adrenaline cause reduced margination of
neutrophils whereas dialysis causes increased margination.
4.2.3. Eosinophil Granulocytes
Eosinophils have two nuclei with granules that stain red with the eosin. They
respond to chemostatic to stimuli and they migrate to sites of allergic reactions
or parasitic infection. Eosinophils have phagocytic capabilities.
Diagram 6: Eosinophil

Eosinophil leucocytosis (Eosinophilia)

Eosinophilia is increased eosinophils numbers
Causes
1. Parasitic infestations
a. Intestinal parasites Amoeba, Hook worms, Tape worms, Ascaris
b. Tissue parasites Filariae and Trichnosis
2. Allergic disorders/hypersensitivity
a. Asthma
b. Hay fever
c. Food sensitivity
d. Urticaria
e. Angioneurotic oedema
3. Drugs
4. Loefflers Syndrome
5. Skin disorders
a. Urticaria
b. Eczema
c. Pemphigus
d. Psoriasis
e. Dermatitis herpetiforms

e. Dermatitis herpetiforms
6. Malignant disorders

a. Lymphoma Hodgkins
b. Carcinoma
c. Melanoma
d. Leukaemia (CGL)

7. Tropical eosinophilia (possibly microfilarial)

8. Miscellaneous conditions
a. Polyarteritis nodosa
b. Sarcoidosis
c. Scarlet fever
d. Chorea
e. Erythema multifornate
f. Ulcerative colitis
g. Hypoadrenalism

Eosinophil leucopenia (Eosinopenia)

Eosinophilia is reduced eosinophils. The circulating eosinophil level has a diurnal
variation with adrenocortical activity (increased adrenaline level leads to
decreased eosinophils)
Causes
1. Increased secretion of
glucosteroids
2.
Therapeutic

glucosteriods and ACTH

4.2.4. Basophils
Basophils resemble other mature granulocytes but have coarse intensely
basophilic granules which fill the cytoplasm and over overlie an obscure nucleus.
Granules of basophils contain histamine and 5HT.
Diagram 7: Basophil

Basophilia

Basophilia
Basophilia is increased number of basophils.
Causes of Basophilia
1. Chromic myeloid leukaemia
2. Polycytheamia vera
3. Myxodema
4. Ulcerative colitis
5. hodgokins disease
6. Following spleenectomy
7. Myelosclerosis

4.3. Agranulocytes
4.3.1. Monocytes
Monocytes are precursors of macrophages. Macrophages are phagocytic cells
involved in immune response. They are distributed in the lymph nodes, spleen,
liver and the bone marrow. Macrophages have a fixed phagocytic activity in the
reticulo-endothelial system. They have receptors for IgG, produce humoral
mediators, and also do activate neutrophils and their release.
Diagram 8: Monocyte

Monocytosis
Increased monocytes (monocytosis) can be caused by: 1. Bacterial endocarditis
2. Brucellosis
3. Tuberculosis
4. Typhoid fever
5. Rickettsial illness
6. Protozoal diseases Malaria,
Trypanosomiasis, Kala azar
7. Hodgkins disease
8. Carcinoma
9. Monocytic and myelomonocytic
leukaemia

9. Monocytic and myelomonocytic

leukaemia
A monocytosis is a striking feature of chronic malaria, recovery phase of
infections and chronic illness

4.3.2. Lymphocytes (Immunocytes)

Lymphocytes play a significant role in both the humoral immunity and cellmediated immunity.
Lymphocytosis
Lymphocytosis is increased number of lymphocytes in circulation. It results
mainly from viral diseases and chronic conditions.
Causes
a) Viral infections
i. Infectious mononucleosis
ii. CMV
iii. Mumps
iiii. Rubella
v. Whooping cough (Pertusis)
b) Chronic infections
i. Bacterial infections tuberculosis, syphilis, brucellosis
ii. Infectious hepatitis
c) Thyrotoxicosis (usually relative)
d) Malignancy
i. Gross lymphocytosis e.g. chronic lymphocytic leukaemia
(CLL)
Lymphopenia
Lymphocytosis is decreased number of lymphocytes in circulation.
Causes
1. Stress
2. Elderly states
3. Increased adrenocortical activity
4. Administration of corticosteroids
5. Cytotoxic drugs

4.4. BLOOD PLATELETS

Platelets perform a number of functions in the body. The normal platelet count is
150- 400 x 109/l
4.4.1. Function

150- 400 x 10 /l
4.4.1. Function
1. Help maintain integrity of the vascular endothelium
2. Forms primary haemostatic plug following injury
3. Activate blood coagulation system
4. Provide mediators for vessel repair, inflammation and regulation of
vascular toxicity.
Platelet disorders include: a) Thrombocytosis increased platelets count (reactive)
b) Thrombocythaemia increased platelet count (neoplastic)
c) Thrombocytopenia decreased platelet count
d) Defective function (qualitative platelet disorders)
4.4.2. Causes of Thrombocytosis
1. Myeloproliferative disorders
i. Primary thrombocythaemia
ii. Polycythaemia vera
iii. Chronic myelocytic leukaemia (CML)
2. Secondary thrombocytosis
i. Bleeding
ii. Inflammatory disorders
iii. Rheumatoid arthritis
iiii. Malignant diseases
v. Post spleenectomy
vi. Spleenic atrophy- SCD
vii. Drugs- Epinephrine
viii. Haemolytic anaemia
ix. Post thrombocythemia

5.0. ERYTHROCYTE SEDIMENTATION RATE (ESR)

5.1. Introduction
Erythrocyte sedimentation rate (ESR) is a measure of suspension stability of red
cells in blood which influences the distance red blood cells fall in I hour that is
measured in millimetres from the surface of meniscus to upper limit of red cell
layer in a column of blood.
The sedimentation rate is influenced both by the proteins of the acute phase
response in inflammation and by anaemia. The rate of fall of red cells is
influenced by a number of inter-reacting factors but basically it depends upon
the difference in specific gravity between red blood cells and plasma and greatly
to the extent to which red cells form rouleaux that sediment more rapidly than
single cells.
Rouleaux formation and red cell clumping are controlled by the concentration of
plasma proteins fibrinogen and acute phase proteins such as alpha1-acid
glycoprotein, alpha1- antitrypsin, haptoglobin, ceruloplasmin, C-reactive proteins

plasma proteins fibrinogen and acute phase proteins such as alpha1-acid

glycoprotein, alpha1- antitrypsin, haptoglobin, ceruloplasmin, C-reactive proteins
and glycoproteins. Rouleaux formation is enhanced by immunoglobulins and
retarded by albumin. .
Increased sedimentation rate of clusters of cells shows reduced fluid friction
resulting from decreased surface: volume ratio leading to reduced surface
tension of the red cells. Anaemia alters the ratio of red cells to plasma,
encourages rouleaux formation and accelerates sedimentation.
Clinical evaluation (prognostic aid in rheumatoid arthritis, acute renal failure,
tuberculosis and assist to distinguish between serious and functional symptoms
Sedimentation takes place in three stages:
1. Preliminary stage that lasts a
few minutes
a. Rouleaux formation
b. Aggregation
2. Period of sinking of the
aggregates at a constant speed
3. Slowing of sedimentation
rate as the aggregated cells pack at
the bottom of the tube
5.2. Technique
Blood is collected in anticoagulation. Add to Wintrobe haemotocrit tube to 10cm
mark (Wintrobe) and Westegren (200mm)
5.3. Causes of Increased ESR
1. Non-specific index of organic
disease
2. Acute or chronic infections
3. Neoplasms
4. Collagen disease
5. Renal insufficiency
6. Any condition altering
plasma proteins
7. Physiological pregnancy and
old age
6.0. LABORATORY TESTS
Investigations of blood disorders depend on examination of the peripheral blood
and the bone marrow.
6.1. Peripheral Blood
The main parameters examined on the peripheral blood film stained by a
Romanowski method include: 1.

Haemoglobin
concentration
whole blood)

(Hb)
(g/dl

concentration
whole blood)
2. Cell count

3.

(g/dl

a. Red cell
count
b.
White
cell
count
(total
and
differential)
c. Platelets

Absolute
cell
measurements
a. Mean red
cell
volume
(MCV)
b.
Mean
cell
haemoglobin
(MCH)
c. Mean cell
haemoglobin
concentration
(MCHC)

4. Packed cell volume (PCV)

5. Shapes of red cells
6. Colour of red cells
6.2. Bone Marrow Examination
Examination of the bone marrow is important in explaining abnormalities of the
peripheral blood. Bone marrow tissue is obtained by aspiration and a trephine
biopsy performed at the posterior iliac crest. The bone marrow may exhibit
reduced, normal or increased function.

Page 5 of 154
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

ANAEMIA - INTRODUCTION
LEARNING OUTCOMES
At the end of the lesson the learner should be competent to: 1. Classify anaemia
2. Describe the effects of anaemia
3. Explain how the body adapts to anaemia
4. Explain the pathophysiology of anaemia
5. Outline the clinical features of anaemia
6. Outline causes of anaemia
7. Describe the mechanisms of anaemia formation
1.0. INTRODUCTION
Anaemia is the reduction in haemoglobin (Hb) concentration in the blood

Anaemia is the reduction in haemoglobin (Hb) concentration in the blood

below the reference range, normal (with respect to age, sex and altitude).

2.0. CLASSIFICATION (AETIOLOGY)

1. Excessive loss or destruction of red cells
a. Blood Loss haemorrhage (post haemorrhagic anaemia)
b. Destruction Haemolytic anaemia.
2. Under production (Failure of production of cells).
a. Diminished production with marrow
hyperplasia (Dyserythropoeitic anaemia)
b. Diminished production with marrow
hypoplasia - hypoplastic anaemia and aplastic
anaemia

3.0. EFFECTS OF ANAEMIA

The effects of anaemia are greatly influenced by its severity, duration and rate
of development of anaemia. In moderate cases of anaemia that evolves slowly
symptoms such as dyspnoea only appear on exertion and when the Hb in such
case drops further, clinical signs remain mild. In very rapidly developing
anaemia, compensatory mechanisms fail to adjust adequately resulting in shock.
1. Anaemia reduces oxygen
carrying capacity of blood
resulting in tissue hypoxia.
compensatory
include:

2. The
adjustments

Reduced
anterior tone

Increased
stroke
Increased heart
rate

Increased
cardiac output

These
adjustments
contribute
to
the
clinical

Reduced
circulation time

pictures of the
patient
producing
features such as:

patient
producing
features such as:
Improved tissue
perfusion

1)
bounding pulse, 2)
Wide pulse pressure,
3) Cardiomegally and 4) Haemic murmur

3. Tissue hypoxia stimulates erythropoietin production which advances

erythroid hyperplasia
4. The response of the bone marrow to erythropoietin leads to conversation of
yellow fatty marrow to red marrow increasing cell production (effective
erythropoiesis). This is seen in haemolytic anaemia and haemorrhagic
anaemia.
5. If bone marrow hyperplasia occurs but maturation of cells is slow
precipitating destruction of the red cell mass, the red cells mass fails to
increase (ineffective erythropoiesis).
6. Compensatory hyperplasia fails to occur despite erythropoietin stimulation
of the bone marrow leading to aplastic anaemia.

Diagram 1: Effects of Anaemia

Reduction in Hb

Tissue hypoxia

Compensatory changes

Circulatory

Biochemical

Heart rate and

Increased content

yperplasia
H
Output increased;
to
Dilatation of arterioles
And increased rate of
Circulation.

Increased tissue perfusion

of 2,3 DPG in red cells

Marrow

(attempt
correct red
cell deficit)

ffinity of Hb for O2
A
Reduced in peripheral
Circulation

Easier transfer to tissues

ABSENCE OF SYMPTOMS AT REST

4.0. ADAPTATION TO ANAEMIA

4.1. Oxygen utilization
The function of red cells is to carry oxygen between lungs and tissues. Tissue
oxygenation depends on cardiac output, ventilatory function, the state of
capillaries and erythron.
a. Ventilation responds to pH, partial pressure of CO2
(PCO2) and hypoxia.
b. Cardiac output depends on venous return
depending on tissue metabolism which modifies
resistance to blood flow in microvasculature.
c. Erythron performance is influenced by change in
Hb, arterial oxygen saturation and oxygen affinity of
circulating Hb.
Anaemia leads to low arterialvenous oxygen difference therefore to compensate
for this there is: a) Modulation of oxygen affinity
b) Redistribution of flow between different organ
c) Increased cardiac output
d) Reduction of mixed venous oxygen tension to increase arterio-venous
oxygen difference.

Diagram 2: Steps in Oxygen utilization

Steps
Ambient oxygen tension

Ventilation

Factors involved
Altitude

Alveolar ventilation
as to blood diffusion
G
Ventilation/perfusion ratio
Anatomical shut

 as to blood diffusion
G
Ventilation/perfusion ratio
Anatomical shut
Circulation

Tissue perfusion

ardiac output
C
Blood (Hb)
Oxygen dissociation curve
Intercapillary distance

Diagram 3: Oxygen utilization

4.2. Intrinsic red cell adaptation

The reduced oxygen carrying capacity of the Hb leads to increased production of
2,3 PDG (diphosphoglycerate) in the red cells and shifting of the dissociation
curve to the right.
Increased production of 2,3 PDG is seen in: 1. Anaemia
2. Alkalosis
3. Renal failure
4. Hypoxia
5. Pregnancy
6. Cyanotic heart disease (CHD)
7. Thyrotoxicosis
8. Red cell enzyme deficiencies
Reduced 2, 3 PDG is seen in: 1. Acidosis
2. Cardiogenic/septicaemia shock
3. Hypothyroidism.

2. Cardiogenic/septicaemia shock
3. Hypothyroidism.
4.3. Local changes in tissue perfusion
The changes that occur in tissue perfusion as a compensatory strategy are:
1. Shunting of blood from less vital tissue to vital tissues attains increased
tissue perfusion.
2. Vasoconstriction of vessels in the skin and kidney
3. Increased flow to the myocardium, brain and muscle.
4.4. Cardiovascular system
A Hb of less than 7-8g/dl leads to increased cardiac output at rest and after
exercise because of vasodilatation and reduced blood viscosity.
Because of the low haemoglobin there is: 1. Increased stroke volume
2. The hyperkinetic circulation results in tachycardia, arterial and
capillary pulsation, wide pulse pressure and haemic murmur
3. Reduced circulation time
4. Increased left ventricular stroke work
5. Increased coronary flow and cardiac output
In Severe cases this leads to cardiomegally, pulmonary oedema, ascites,
peripheral oedema and congestive cardiac failure (CCF)
4.5. Pulmonary function
Severe anaemia is associated with dyspnoea due to cardiac failure and response
to hypoxia, which is centrally mediated.
5.0. PATHOPHYSIOLOGY OF ANAEMIA
Reduced level of haemoglobin leads to lowered oxygen-carrying capacity of blood
a situation that in turn triggers compensatory physiological adaptation
mechanisms such as: 1. Increased release of oxygen from haemoglobin
2. Increased blood flow to tissues
3. Maintenance of blood volume
4. Redistribution of blood flow to maintain cerebral blood supply.
Hypoxia leads to impaired tissue and organ function and the degree of functional
impairment depends on tissue oxygen requirements e.g. the heart, central
nervous system and skeletal muscles during exercise are most affected by
anaemia.
6.0. PATHOLOGICAL COMPLICATIONS OF ANAEMIA
In anaemia, the effects of degenerative arterial disease are usually aggravated
such that symptoms like angina pectoris and claudication of the lower limbs
become worse. In severe anaemia the effects of hypoxia are well exhibited in
many body organs.

become worse. In severe anaemia the effects of hypoxia are well exhibited in
many body organs.
Diagram 4: Pathological Complications
HYPOXIA

Fatty change in
to
Myocardium

Fatty change in liver

Circulatory compensatory
Changes, weakens myocardium
Leading to acute cardiac failure

Kidneys sensitive
hypoxia

Increased
erythropoietin by
renal peritubular cells

Marrow hyperplasia
7.0. CLINICAL FEATURES OF ANAEMIA
Clinical Features of Anaemia
Development of symptoms and signs of anaemia depends on four main factors
1. The speed of onset of anaemia
2. Severity of anaemia
3. The age of the patient
4. The haemoglobin dissociation curve
Symptoms
1. Tiredness
2. Easy fatigability
3. Generalised muscular weakness
4. Lethargy
5. Headache
6. In older patients
a. Cardiac failure
b. Angina pectoris
c. Intermittent claudication
d. Confusion
e. Visual disturbances
Signs
1. Pallor
2. Cardiovascular system
a. Hyperdynamic circulation
i. Tachycardia
ii. Collapsing pulse
b. Cardiomegally

i. Tachycardia
ii. Collapsing pulse

b. Cardiomegally
c. Murmur (midsystolic flow murmur)
d. Dyspnoea on exertion
e. Congestive cardiac failure

3. Central nervous system

a. Faintness
b. Giddiness
c. Headache
d. Tinnitus
e. Drowsiness
f. Numbness
g. Tingling sensation of the hands and feet
4. Ocular manifestations
a. Retinal haemorrhage
5. Reproductive system
a. Menstrual disturbances
b. Loss of libido
6. Renal system

a. Mild proteinuria
b. Impaired concentration of urine

7. Gastrointestinal system
a. Anorexia
b. Flatulence
c. Nausea
d. Constipation
e. Weight loss
8.0. CAUSES OF ANAEMIA
Classification of Anaemia
Anaemia can be classified in two ways based on pathophysiology and
morphology.
a) Pathophysiologic classification
i. Anaemia due to increased blood loss
1. Acute blood loss
2. Chronic blood loss
ii. Anaemia due to impaired red cell production
1. Cytoplasmic maturation defects
a. Deficient heam
synthesis iron deficiency
b. Deficient globin
synthesis - Thallassaemia

synthesis - Thallassaemia
2. Nuclear maturation defects
a. Vitamin B12
(megaloblastic anaemia)
b. Folic acid deficiency
What is your understanding of the concept of
(megaloblastic
anaemia)
anaemia?
What is the impact of anaemia on body
3. Defects in stem cell proliferation
functions?
a. Aplastic anaemia
b. Anaemia of chronic
disorders
4. Anaemia of chronic disorders
5. Bone marrow infiltration
6. Congenital anaemia
iii. Increased red cell destruction (Haemolytic
anaemia)
1. Extrinsic (extra corpuscular) red cell
abnormalities
2. Intrinsic (intra corpuscular) red cell
abnormalities.
a. Membranopathies
b. Enzymopathies
c.
Haemoglobiniopathi
es
b) Morphological classification based on the size of the red cell, Hb content
and red cell indices.
1. Microcytic, hypochromic
2. Normocytic, normochromic
3. Macrocytic normochromic.
Diagram 5: Causes of Anaemia

9.0. MECHANISMS OF ANAEMIA FORMATION

9.1. Introduction
The various aetiological agents of anaemia formation do cause anaemia via four
main mechanisms that depend on the life cycle of the red blood cells comprising
of the bone marrow cycle and the circulation cycle. Normal haemoglobin level
depends on the red cell balance sheet established between production (bone
marrow) and losses (spleen, liver, lymph nodes and bone marrow).
The mechanisms are: 1. In the Bone Marrow
a. Hypopalstic anaemia
b.
Dyserythropoietic
anaemia
2. In the circulation
a. Haemorrhagic anaemia
b. Haemolytic anaemia
9.2. Hypoplastic Anaemia
In hypoplastic anaemia there is actual diminution in the productive marrow
(hyoplasia or aplasia) or marrow replacement by tumour. This produces
hypoplastic and aplastic anaemia which result in pancytopenia in the peripheral
blood.
9.3. Dyserythropoietic Anaemia
The marrow is unable to produce sufficient normal red blood cells due to
deficiency of essential factors such as iron, folate and vitamin B12.
9.4. Haemorrhagic Anaemia
There is loss of red blood cells to haemorrhage or bleeding
9.5. Haemolytic Anaemia

9.5. Haemolytic Anaemia

There is excessive destruction of red blood cells by the macrophage system
(especially the spleen) which destroys normal red blood cells as in
hyperspleenism and autoimmunity and abnormal red blood cells such as
hereditary spherocytosis and sickle cell disease.
Page 5 of 154
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

APLASTIC AND DYSHAEMOPOIETIC ANAEMIAS

LEARNING OUTCOMES
At end of the lesson the learner should be competent to: 1. Describe the causes and pathology of aplastic anaemia
2. Describe causes of bone marrow failure
3. Describe functions and metabolism of iron
4. Outline the causes of iron deficiency
5. Outline the causes of folic acid and vitamin B12 deficiency
6. Describe pathophysology of dyshaemppoietic (deficiency) anaemias
7. Describe pathology of dyshaemppoietic (deficiency) anaemias
8. Describe clinical features of dyshaemppoietic (deficiency) anaemias
9. Outline the effects of iron overload and tissue damage.
10. Outline the investigations in dyshaemppoietic (deficiency) anaemias

APLASTIC ANAEMIA (BONE MARROW FAILURE)

1.0 INTRODUCTION

Aplastic anaemia presents as pancytopenia with reduced bone marrow cellular

activity hypocellularity (aplasia). It results from reduction in number of
pluripotent stem cells. There is failure or suppression of the multipotent myeloid
stem cells and resultant neutropenia, anaemia and thrombocytopenia
(pancytopenia). Bone marrow failure may absolute or relative.
2.0 CAUSES

1. Primary
a. Congenital

e.g. Fanconis anaemia Fanconis anaemia is

an inherited as an autosomal recessive and is
associated with skeletal, renal and central nervous
system abnormalities. Presents usually between the
ages of 5 and 10 years.
b. Idiopathic (65% cases)
2. Secondary
a. Bone

marrow infiltration or replacement

i.
Hodgkins
and
Non-Hodgkins
lymphoma
ii. Acute leukaemia
iii. Myeloma
iiii. Secondary carcinomas

iii. Myeloma
iiii. Secondary

carcinomas

v. Myelofibromatosis
b. Chemicals

e.g. benzene

c. Drugs

Chemotherapeutic e.g. alkylating

agents
and
antimetabolites/cytotoxic
drugs
(vincristine, busulfan, doxorubicin)
ii. Idiosyncratic e.g. chloromphenicol,
chlorpromazine, gold, carbimazole,
phenytoin, tolbutamide, NSAIDS and
streptomycin
i.

d. Insecticides
e. Ionising

radiations/irradiation

f. Infections

Viral infections (Hepatitis non-A,

non-B and non-G, measles, HIV,
parvo virus, Epstein Barr virus)
ii. Bacterial e.g. tuberculosis
i.

g. Pregnancy
h. Paroxysmal

nocturnal haemoglobinuria

3.0 PATHOGENESIS

1. A

primary defect in the number or function of stem cells

of antigenically altered stem cells by immune (T cell-mediated)
mechanisms. Stem cell alterations may be secondary to environmental
insults, drug exposure or infections.

2. Suppression

4.0 MORPHOLOGY

(PATHOLOGY)

Hypocellular bone marrow and the haemopeoitic cells are replaced by fat
cells
Secondary effects
o Granulocytopenia (leads to infections)
o Thrombocytopenia (leads to bleeding)
5.0 CLINICAL

FEATURES

Anaemia
Bleeding ecchymosis, bleeding gums, epistaxis
Infections
Laboratory
o Pancytopeia - reduced red Hb, platelets and
white blood cells
o Absence of reticulocytes
Severe aplastic anaemia has a poor prognosis and is associated with any
two of the following: 9
o Neutrophil count < 0.5 X 10 /L
9
o Platelet count of < 20 X 10 /L
9
o Reticulocyte of < 40 X 10 /L

Reticulocyte of < 40 X 10 /L

6.0 INVESTIGATIONS

The laboratory diagnosis is based on: 1. Pancytopeia

a. Reduced

red Hb
b. Reduced platelets
c. Reduced white blood cells
2. Absence of reticulocytes
3. Hypocellualr marrow or aplastic bone marrow with increased fat spaces
7.0 DIFFERENTIAL

1. Aplastic

DIAGNOSIS OF PANCYTOPENIA

anaemia

2. Drugs
3. Megaloblastic

anaemia
4. Bone marrow infiltration or replacement
5. Hyperspleenism
6. Systemic Lupus Erythromatosus (SLE)
7. Disseminated tuberculosis
8. Overwhelming sepsis

DYSHEMOPOIETIC (DEFICIENCY) ANAEMIAS

The cause of anaemia in dyshaemopoietic anaemia is deficiency of an
element that is essential for proper haemoglobin synthesis or maturation and
development of erythroblasts. They are associated with a hypercellular
marrow and can be divided into normoblastic and megaloblastic on the basis
of the type of erythroblast maturation in the bone marrow. The term
dyshaemopoiesis means disturbed erythropoiesis.
Diagram 1: Pathogenesis of Dyshaemopoietic anaemia

A.
IRON
DEFICIENCY
due to: 1. Blood loss
2.
Deficient
intake
3.Increased
requirement

DYSHAEMOPOIETIC
ANAEMIA

1
.
V

BONE MARROW
Impairment
synthesis
haemoglobin
NORMOBLASTIC
maturation
MEGALOBLASTIC
maturation

of
of

requirement

.
V
I
T
A
M
I
N
B

HYPERCELLULARITY
1
2

maturation

NORMAL KIDNEY

D
E
F
I
ERYTHROPOIETIN
C
IRON
E DEFICIENCY ANAEMIA
N
C
1.0 INTRODUCTION
Y
Iron is a transition metal (atomic number 26, anatomic weight 55.85) that is an
2
essential nutrient required by every human
cell.
.
F
O
2.0 FUNCTIONS OF IRON
L
I
1. Carrier for oxygen and electrons
C
2. Catalyst for oxygenation
and hydroxylation
A
3. Catalyst element in C oxidative metabolism, cellular growth, oxygen
transport and storage I
D
The capacity for iron to change
between ferrous and ferric oxidation states
D
highly influences the many E
functions that iron supports. Iron does not exists a
free cation but in a variety ofF iron compounds.
I
C
Iron deficiency is the commonest
nutritional disorder throughout the world with
a much higher prevalence inI the developing world. The factors responsible for
iron deficiency are legion. E
N
C
Y

3.0 FUNCTIONS OF HAEM CONTAINING PROTEINS

1. Synthesis of haemoglobin (combines with 4 haem groups linked to four

globin
chains)
The
main
effects of anaemia
2. Myoglobin in muscles (1 Haem)
from ashypoxia
and(iron
thecompounds)
3.result
Mitochondria
cytochromes
4.compensatory
Enzymes for DNAmechanisms.
synthesis, drug detoxification, sterol synthesis,
How
do these factors
contribute
catacholamine
metabolism,
collagen formation and hydroxylation of
todrugs.
the pathophysiology of

anaemia?
4.0 IRON METABOLISM
4.1. Biochemistry
Iron (Fe) is an important element in human metabolism as it plays a central role
in erythropoeisis and intracellular processes of all tissues in terms of its
importance in oxygen and electron transport system, which are concerned with
cellular energy production. Iron derives its functional properties from its ability
to exist in both ferrous and ferric states. Physiologically active iron compounds in

importance in oxygen and electron transport system, which are concerned with
cellular energy production. Iron derives its functional properties from its ability
to exist in both ferrous and ferric states. Physiologically active iron compounds in
the body are haem proteins while others are specialized proteins of iron
transport and storage. Specialized proteins enable iron remain in solution at
neutral pH. Ferric iron is completely insoluble.
Excess Fe stimulates the synthesis of ferritin (small stores are formed) and the
storage compounds of ferritin, which are haemosiderin (found in the reticulo
endothelial system, R.E.S comprising of the liver, spleen, bone marrow) and the
parenchymal cells. Fe is transported in plasma bond to transferrin (-globulin)
synthesized by the liver.
4.2. Distribution
The body concentration of iron in the human body is 40-50mg Fe/kg body weight
(which is usually less in females) of which 30 mg Fe/Kg is stored as Hb iron that
is contained in circulating red blood cells. 6-10mg Fe/Kg is present in tissues
throughout the body in form of myoglobin and in heme enzymes cytochromes,
catalases and peroxidases.
The distribution in percentages is as follows: Largest component (65%) is circulating haemoglobin (200 mg of iron in 450
ml of whole blood)
Storage proteins (30%) haemosiderin and ferritin found in the R.E.S cells
of the liver, spleen and bone marrow (which gain iron by breaking down
red cells) and in parenchymal liver cells (gain iron from the plasma irontransporting protein transferrin).
Myoglobin (3.5%)
Haem and non-haem enzymes and transferrin-bound (1.5%)
4.3. Absorption
Physiological control of iron balance is maintained by regulation of iron
absorption, which occurs mainly in the duodenum and proximal jejunum. The
intestinal mucosa is extremely sensitive to body requirements of iron forming an
important regulatory mechanism. Inorganic iron is usually best absorbed from the
gastro-intestinal tract in a fasting state. Iron in food is released into gastric juice
as ferrous, ferric or as a haem complex and the amount available for absorption
depends on its release (the digestive process) and interaction with the G.I.T
lumen. Ascorbic acid makes iron more soluble (into ferrous state).
Iron absorption is influenced by a number of factors, which are dietary, mucosal
and luminal factors.
Table 1: Factors Influencing Iron Utilization
Factors

Increased absorption

Dietary

Increased haem iron

Increased animal foods
Ferrous iron salts

Luminal Acid

pH

Decreased absorption
Decreased haem iron
Decreased animal foods
Ferric iron salts
Insoluble

iron

complexes

Ferrous iron salts

Luminal Acid
(tea,
Mucosal

Ferric iron salts

pH

Insoluble

Iron deficiency
Increased erythropoeisis
Pregnancy
Anorexia
Ineffective erythropoeisis
Anoxia

iron
complexes
hosphates, bran)
p

Iron overload
ecreased erythropoeisis
D
Acute or chronic inflammation

4.4. Iron Transport

Iron is transported bound to transferrin in plasma and extra cellular fluid. 5-6mg
Fe/Kg in females and 10-12 mg Fe/Kg of iron in males is in storage form of
ferritin and heamosiderin mainly in hepatocytes and macrophages in the liver,
bone marrow, spleen and muscle. Transferrin is the sole physiological carrier of
iron between body tissues. Iron is transported in plasma bound to -globulin,
transferring which the liver synthesizes.
4.5. Ferritin
Primary iron storage protein
Provides a reserve of iron
Water soluble
4.6. Haemosiderin
Haemosiderin is water insoluble golden yellow-to-brown granular pigment
composed of aggregates of ferritin, which is usually derived from haemoglobin. It
stores less iron than ferritin.
4.7. Iron balance
Between amount absorbed and the amount lost. The balance is maintained by
regulating the absorptive intake by intestinal mucosal cells (mucosal block).

5.0 CAUSES
A. Deficient intake
1. Inadequate dietary intake
a. Mal nutrition
b. Poor economic state
2. Decreased absorption

a. Partial or total gastrectomy

b. Achlorhydria
c. Malabsorption
d. Celiac disease
e. Duodenal by-pass

3. Impaired transport
B. Increased blood loss
1. Uterine menorrhagia, malignancy, abortions

1. Uterine menorrhagia, malignancy, abortions

2. Gastrointestinal tract
a) Oesophageal varices
b) Peptic ulcers
c) Hookworm infestations (A. duodenalae 0.2 mls/worm/day;
N. americanas 0.03 mls/worm/day)
d) Malignancy stomach, large bowel, rectum, caecum
e) Ulcerative colitis
f) Chronic aspirin ingestion
g) Small intestine diverticuli
h) Haemorrhoids
3. Renal/urinary tract haematuria, haemoglobinuria
4. Nose - repeated epistaxis
5. Lungs - haemoptysis
C. Increased requirements (physical demands)
1. Prematurity
2. Infants
3. Adolescent
4. Pregnancy
5. The elderly

6.0 PATHOGENESIS OF FE DEFICIENCY ANAEMIA

Iron deficiency anaemia develops when the supply of iron is inadequate for the
requirements of haemoglobin synthesis. The negative iron balance is initially
countered by mobilization of stored iron but when the stores are depleted or
exhausted the state of iron deficiency ensues.
Diagram 2: Pathogenesis of Fe Deficiency anaemia
(1) Iron Deficiency

(2) Mobilization of reserves (haemosiderin first and then ferritin)

(3) Exhaustion of reserves (there will be increased absorption if Fe is

available in the diet and may postpone exhaustion)

(4) Deficient synthesis of haemoglobin

(5) ANAEMIA

7.0 PATHOPHYSIOLOGY
The pathophysiology ranges from diminished iron stores to severe anaemia and
deficiency of tissue iron containing enzymes. There is depletion of the iron stores
and iron deficient erythropoeisis.

8.0 CLINICAL PRESENTATION

8.1. Introduction
Patients with iron deficiency may present with no signs or symptoms but only
with abnormal laboratory test results; features of the underlying disorder
responsible for the iron deficiency and manifestations of anaemia
Marked deficiency of iron which reduces synthesis of functional iron compounds
and cause anaemia may either be asymptomatic or produce a variety of clinical
manifestations. Because iron deficiency is often of insidious onset and prolonged
duration, adaptive circulatory and respiratory responses may minimize the
manifestations permitting tolerance of low haemoglobin levels in the body. As in
all other forms of anaemia, the prominence of signs and symptoms of iron
deficiency anaemia depends on both the degree and rate of development of
anaemia.
8.2. Features of Iron deficiency anaemia
Symptoms
Palpitations
Tinnitus
Headache
Irritability
Weakness
Dizziness
Easy fatiquability
8.3. Features Independent of anaemia
These are features that result from depletion of functional iron compounds in
nonerythroid tissues resulting in impaired proliferation, growth and function of
the structures and organs affected.
Epithelial tissues have high iron requirements due to their rapid rates of
growth and turnover. This leads to: -

 Epithelial tissues have high iron requirements due to their rapid rates of
growth and turnover. This leads to: 1. Glossitis
2. Angular stomatitis
3. Postcricoid web or stricture (may become
malignant)
4. Gastric atrophy
5. Plummer-Vinson
6. Patterson-Kelly syndrome
7. Koilonychia
Blue sclera (due to thinning of the sclera due to reduced collagen synthesis
making the choroids visible)
Impaired immunity
Diminished exercise tolerance
Neuropsychological abnormalities
9.0 HAEMATOLOGICAL EFFECTS (CHANGES SEEN)
1. Blood

a) MCV decreased
b) MCH decreased
c) MCHC decreased
d) Microcytic, hypochronic
e) Poikilocytosis
f) Decreased serum Fe levels
g) Anisocytosis
h) Target cells.

2. Bone marrow

a) Decreased iron
b) Marrow cellular
c) Marrow hyperplasia due
stimulation
d) Increased erythroblasts.

to

erythropoeitin

3. Effects on other tissues - affects haem-containing cytochromes in

epithetical cells, muscle, liver and the kidney.
a) Decreased mitochondrial protein synthesis
b) Epithelial abnormalities leading to atrophy of buccal/gastric mucosal
surfaces and oesophageal mucosal web formation (Peterson- Kelly
syndrome.)
c) Koilonychia
d) Atrophic gastritis
e) Glossitis (atrophic)
f) Dysphagia (anaemia, dysphagia and Glossitis make up the PlummerVinson Syndrome)
g) Achlorhydria
h) Lymphocyte function impaired.
i) Impaired immune response
j) Angular stomatitis

10.0 INVESTIGATIONS

10.0 INVESTIGATIONS
1. Full heamogram and ESR (what values will you get)
2. Stool
3. Urine
4. Barium studies
5. Serum Fe levels
6. Total iron binding capacity (TIBC)
7. Serum ferritin (decreased in rheumatoid arthritis, chronic renal
failure and inflammatory bowel disease)
8. Marrow iron
9. Endoscopy
10. Recto/sigmoscopy
11. Ultrasound
12. Liver function tests

IRON OVERLOAD
1.0 INTRODUCTION
Absence of a physiological pathway for excretion of excess iron leads to
accumulation of Fe in the tissues of the body resulting in lethal tissue damage.
Iron overload denotes excess total body iron resulting from an iron supply that
exceeds body iron requirements. The body iron requirements are limited and
since humans lack the physiological means of excreting excess iron, the iron
usually accumulates in body tissues. Local iron overload may be due to
sequestration of iron e.g. pulmonary haemosiderosis. When the amount of iron
that accumulates in the body surpasses the bodys capacity for safe storage
lethal damage to tissues results.
2.0 CAUSES
1. Congenital
defects
e.g.
atransferrinae
mia

mia
2.
Blood
transfusion
Congenital anaemia
Sideroblastic anaemia
Hypoplastic anaemia
3. Excess Fe
absorption
Idiopathic haemochromatosis
Ineffective erythropoeisis
4. Excess
intake

Fe

Prolonged oral iron therapy

High Fe alcoholic drinks
Excess parental iron therapy
5. Secondary
to
liver
disease
Alcoholic cirrhosis
Porto-caval anastomasis
3.0 IRON AND TISSUE DAMAGE:
Haemachromatosis is deposition of iron in the parenchymal cells of the liver,
heart, endocrine glands that results in extensive tissue damage and fibrosis.

4.0 CLINICAL PRESENTATION

a) Increased skin pigmentation
b) Hepatic disease
c) Diabetes mellitus
d) Gonadal insufficiency
e) Endocrine disorders e.g. pituitary gland, adrenal gland
f) Abdominal pain
g) Cardiac dysfunction
h) Arthropathy
i) Occasionally neurologic and psychological abnormalities
5.0 DISORDERS OF FE METABOLISM
Fe released from the breakdown of Hb by the macrophages is bound to
apotransferin to form transferrin and is returned to the bone marrow where
transferrin is given up to normoblasts in the bone marrow and used to form
haem.
Chronic diseases such as chronic infection, TB, Rheumatoid disease, S.L E., Renal

haem.
Chronic diseases such as chronic infection, TB, Rheumatoid disease, S.L E., Renal
failure with anaemia and Disseminated carcinoma block the release of Fe by
macrophages. It presents with mild microcytic anaemia; shows low plasma levels
of Fe. There is low Total Iron Binding Capacity (TIBC) and low plasma transferrin
leads to low Fe supply to the bone marrow hence delayed maturation of
erythroblasts.

SIDEROBLASTIC ANAEMIA
1.0 INTRODUCTION
A sideroblast is a normoblast stained by Perls Prussian blue method to show
cytoplasmic iron granules, which are usually arranged in rings. A pathological
sideroblast is where there is abnormally large accumulation of Fe. In
sideroblastic anaemia there is defective haem synthesis in this all hence their
failure to utilize Fe.
2.0 CLASSIFICATION
1. Hereditary sideroblastic anaemia
A rare X-linked disorder that manifests in childhood or early adult
life
2. Acquired sideroblastic anaemia
Primary (idiopathic)
Secondary
o Drugs, chemicals and toxins
chloromphenicol,
isoniazid,
cycloserine, alcohol and lead
o Haematological - polycythaemia vera,
acute
leukaemia,
myeloma,
lymphoma and haemolytic anaemia
o
Miscellaneous

carcinoma,
myxoedema, rheumatoid arthritis,
S.L.E

MEGALOBLASTIC ANAEMIA
1.0 INTRODUCTION

1.0 INTRODUCTION

Megaloblastic anaemias are a group of haematological disorders resulting from

impaired DNA synthesis characterized by distinctive morphological abnormalities
of developing red cells (haemopoietic precursors) in the bone marrow. The
maturation of the nucleus of the red cells is delayed relative to maturation of
the cytoplasm. The cell division is slow but cytoplasmic development progresses
normally resulting in abnormally large nucleated red cell precursors called
megaloblasts.
The megaloblasts are morphologically and functionally abnormal cells with the
resulting mature red cells that are released into the peripheral circulation being
abnormal in size and shape with the most prominent abnormality being
macrocytosis.

This affects production of the red cells, granulocytes and platelets with the
distinct abnormality of haemopoeisis being the megaloblastic change.

2.0 FEATURES OF MEGALOBLASTIC ANAEMIAS

The following features are common: 1. Abnormally large red cells (megaloblasts) whose nuclear maturation lags
behind the cytoplasmic maturation.
2. Ineffective erythropoeisis
3. Prominent anisocytosis
4. Increased MCV
5. Abnormal granulopoeisis.

3.0 CAUSES

Anaemia

1.
Vitamin B12 deficiency.
2.
Folic acid deficiency
3.
Cytotoxic drug therapy
4.
Inherited
enzyme
defects

These are the commonest cause of macrocytic anaemia. Other causes of

A. Loss
megaloblastic
anaemia without megaloblastic marrow change
include: - alcoholic
UNDERPRODU
liver disease, severe haemorrhage and anaemia with
CTIONextramedullary
haemopoeisis.

Haemorr
Haemol
Iron
hage
ysis
Deficiency
4.0 EFFECTS OF B12 AND FOLIC ACID DEFICIENCIES

Megalobla
stic

Vitamin B12 and folic acid play an important role in cell function (normal cell
division and maturation) as well as being co-enzymes in biochemical reactions.
Platelet
Coagulat
Intrinsic
Extrinsi
Acquir
Inherit
Acquir
Inherit
Deficiencies
of folic
tissues with high rate
of cell
Disord
ion acid and vitamin B12 affect
(Heredita
c

division and maturation) as well as being co-enzymes in biochemical reactions.

Platelet
Coagulat
Intrinsic
Extrinsi
Acquir
Inherit
Acquirand vitamin
Inherit
Deficiencies
of folic
acid
B12 affect
tissues with high rate
of cell
Disord
ion
(Heredita
c
ed
ed
ed
ed
turnover
e.g.
Haemopoietic
system.
They
produce
profound
effects
in
ers
Defects
ry)
(Acquirmany
organs as outlined below.
ed)
1. Blood
Membranop
athies

2. Bone Marrow

Warm

Enzymopat neutropenia
Haemoglobino
a. Pancytopenia,
and slight reduction in
hies
pathies
platelets
b. Reduced Hb and MCV
c. Anisocytosis (varying shapes)
d. Macrocytes
e. Poikilocytes (abnormal Shapes)
Immune
Nonf. Decrease Reticulocyte count.
causes
immune

a. Maximal cellularity
b. Autoimmu
Megaloblastic change due to Iso-immune
delay in nuclear
maturation,
accumulation of many cells in an early
ne
stage of development and many immature cells die in
the marrow.
Cold

Rh

ABO

incompatibility
3.
Neurological Changes antibodies
antibodies
incompatibil
a. Common in B12 deficiency
ity
b. S.A.C.C.D. sub-acute combined cord degeneration as
a result of: i. Discontinuous demyelination of long
pyramidal tracts
ii. Involvement of the posterior
column of mid thoracic spinal cord
iii. Involvement
of
the
large
peripheral nerves
iiii. Focal demyelination may occur in
the
cerebral
hemispheres

psychiatric
disturbances
megaloblastic madness

4. Other Tissues

a. Epithelial changes seen include: i. Atrophy of Epithelium of tongue

glossits and oral ulceration.
ii. Uterus and cervix (B12)
iii. Villous atrophy
iiii. Sterility resulting from disturbed
maturation of germ cell in the
gonads (B12)
v. Haemosideron deposition in renal
tubes
vi.
Spleenomegally
(slight
to
moderate)

Diagram 2: Effects of B12 and Folate Deficiency in Organs

FOLIC ACID/FOLATE DEFICIENCY ANAEMIA

1.0 INTRODUCTION
Folic acid (pteroyl glutamic acid) consists of pteridine, para aminobenzoic acid
(PABA) and glutamic acid.
2.0 BIOCHEMISTRY
Folate of folic acid is a water-soluble B-complex vitamin called pteroyl glutamic
acid (PGA), which exists in polyglutamate form (conjugated folates). For its
metabolic action as a co-enzyme in DNA and RNA synthesis, polyglutamates must
be reduced to dihydrofolate and tetrahydrofolate forms. The Average daily
requirement of folic acid is 100-200 g.

3.0 SOURCES
Folate exists in plants, bacteria and animal tissues
Main dietary sources - fresh leafy vegetables, fruits, liver, kidney and to a
lesser extend muscle meat, cereals and milk
Small amount synthesized by bacteria in human large bowel (not available
to the body since folate is absorbed in the small intestine).
4.0 ABSORPTION, TRANSPORT AND STORAGE
Folic acid is absorbed mainly from the duodenum and upper jejunum, less extent
from the lower jejunum and ileum in monoglutamate and diglutamate forms
after being broken down from polyglutamate forms by folate conjugase enzyme
in the mucosal cells. Synthetic folic acid preparations in polyglutamate form are
absorbed rapidly as mono and diglutamate forms. Carrier proteins transport folic
acid. Folate is labile and is largely destroyed by cooking and canning
5.0 FUNCTION

5.0 FUNCTION
Folate plays an essential role in cellular metabolism by acting as a co-enzyme in
two reactions involving DNA and RNA synthesis.

6.0 CAUSES OF DEFICIENCY

1. Poor diet/Dietary
a. Especially poverty
b. Psychiatric disturbance
c. Alcoholism
d. Dietary fads
e. Scurvy and kwashiorkor
f. Goats milk anaemia
g. Elderly persons
h. Delayed infancy weaning
i. Recurrent infections
2. Malabsorption
a. Major factors

b. Minor factors

i. Gluten sensitive enteropathy

ii. Tropical spruce
iii. Celiac disease
iiii. Congenital malabsorption
i. Partial gastroectomy and jejunal
resection
ii. Chronic diseases
iii. Lymphoma
iiii. Systemic infectious

c. Drugs alcohol, oral contraceptives, cholestyramine.

3. Excess Requirements
a. Physiological

b. Pathological

i. Pregnancy
ii. Prematurity and infancy
iii. Malignancies
1. Leukaemia
2. Carcinoma
3. Lymphoma
4. Myeloma
5. Sarcoma
iiii. Blood disorders (haematological)

5. Sarcoma
iiii. Blood disorders (haematological)
1. Haemolytic anaemia
(SCD, thalasseamia)
2.
Sideroblastic
anaemia
v. Inflammatory
1. Tuberculosis
2. Malaria
3. Chrons disease
4. Psoriasis
5.
Exfoliative
dermatitis
6. Rheumatoid arthritis
vi. Metabolic
Homocystinuria
(sulphar containing amino acid cystine)
vii. Excessive urinary excretion
1. Congestive cardiac
failure (CCF)
2. Acute liver disease
3.
Chronic
haemodialysis
and
peritoneal
dialysis
4. Antifolate drugs the mechanism is uncertain
a. Anticonvulsants (phenytoin, barbiturates, promidine)
b. Oral contraceptives
c. Nitroforantoin
d. Tetracycline
e. Anti-TBs (les well documented)
f. Alcohol.
5. Mixed causes

a. Liver disease
b. Alcoholism
c. Intensive care units

7.0 DIAGNOSIS
1. History and physical examination
2. Investigations
a. Full heamogram
b. Serum folate (normal 6-12 ng/ml [6-12 g/L)
c. Red cell folate (20-50 times more than serum folate)
3. Therapeutic trial

VITAMIN B12 DEFICIENCY ANAEMIA

1.0 INTRODUCTION
Vitamin B12 is a complex compound with different chemical forms called the
cobalamins.
It has a cobalt atom and a corrin ring similar in structure to the porphyrin of

cobalamins.
It has a cobalt atom and a corrin ring similar in structure to the porphyrin of
haem. There are four major forms of cobalamine: 1. Hydroxycobalamin (tissue, plasma and commercial/therapeutic
preparations
2. Methylcobalamin (co-enzyme)
3. Cyanocobalamin is the stable form (therapeutic preparation).
4. Deoxycobalamin
2.0 SOURCES
1. Animal protein kidney, liver, heart, muscle meats, fish, eggs, cheese and
milk.
2. Cooking has little effect
3. Synthesized in the human large bowel by microorganisms
4. Average daily requirements 2-4 g.
3.0 ABSORPTION AND TRANSPORT
After ingestion vitamin B12 in food is released and forms a stable complex with
gastric R-binder a glycoprotein found in various secretions (saliva, milk, gastric
juice, bile), phagocytes and plasma. In the duodenum B12-R-binder complex is
digested releasing vitamin B12 which then binds to the intrinsic factor (IF) a
glycoprotein produced by the parietal cells of the stomach
Secretion of IF parallels that of hydrochloric acid. Vitamin B12-IF complex reaches
the distal ileum where it binds to specific receptors on the mucosal wall
facilitating absorption. After several hours in the mucosal cells the IF is
destroyed and vitamin B12 released and transferred to another protein,
transcobalamin (TC) II. (Others are TCI and TC III). Vitamin B12-TCII complex is
secreted into the portal circulation and taken to the liver, bone marrow and
other cells.
4.0 STORAGE
The principal storage site is the liver (2 mg) and other tissues kidney, heart and
brain. The body stores of vitamin B12 are adequate for 2-4 years and the major
source of loss is via bile and shedding of intestinal epithelial cells.

5.0 FUNCTIONS
Vitamin B12 plays a significant role in general cell metabolism especially: 1. Normal haemopoiesis due to DNA synthesis
2. Maintenance of integrity myelin sheath of the nervous system
3. Aids in intracellular conversion of transport form of Folic acid (5 methyltetralyhydrofolate) to polyglutamate form.
4. Purine and pyrimidine synthesis
5. Mitosis and maturation of cells.
Deficiency of vitamin B12 results in megaloblastic haemopoeisis due to DNA
synthesis failure and neurological disturbances resulting from impaired RNA
synthesis in nerve cells.

Deficiency of vitamin B12 results in megaloblastic haemopoeisis due to DNA

synthesis failure and neurological disturbances resulting from impaired RNA
synthesis in nerve cells.
6.0 CAUSES OF B12 DEFICIENCY
1. Dietary
a. Vegetarians/Vegans
b. Long continued poor diet (extremely) rare.
2. Malabsorption
a. Gastric causes

partial

1. Deficient production of IF
2. Chronic gastritis
3. Gastrectomy total or

4. Congenital IF deficiency
or abnormality
5. Antibodies against IF
b. Intestinal causes

Intestinal
stasis
encourages growth of
bacteria which extract
B12 from food - Stagnant
loop syndrome seen in
jejunal
diverticulosis,
ileocolic
fistula,
anatomical blind loop,
strictures and chronic
obstruction.
2. Ileal resection
3. Crohns disease
4. Chronic tropical sprue
5. TC II deficiency
6. Zollinger

Ellison
syndrome
7.
Fish
tape
(Diphyllobothrium
latum) absorbs free and
bound vitamin B12
8. Drugs anticonvulsants,
neomycin, slow K and
metformin
1.

7.0 PATHOPHYSIOLOGY
Vitamin B12 and folic acid are essential coenzymes in the DNA synthetic pathway.
A deficiency of these nutrients results in deranged or inadequate synthesis of
DNA but synthesis of RNA and proteins in usually not affected. Disordered DNA
formation leads to enlargement and maturation of the cytoplasm without
matching nuclear maturation, which is delayed. This results in effects red blood
cells and rapidly dividing cells e.g. myeloid cells and the mucosal epithelium of
the gastrointestinal tract.

matching nuclear maturation, which is delayed. This results in effects red blood
cells and rapidly dividing cells e.g. myeloid cells and the mucosal epithelium of
the gastrointestinal tract.
Anaemia results from: Poor erythropoeisis
Production of abnormal cells that are susceptible to accelerated
haemolysis
Poor formation of red blood cells and platelets together with premature
destruction of cells affects granulocyte and platelet precursor cells resulting in
pancytopenia. The classical disease resulting from B12 and folic acid deficiency is
pernicious anaemia.
Atrophic gastritis results from reduced regenerative activity of the mucous
membranes of the alimentary and genital tracts, as they cannot meet the
demands of the high cell turnover. There is achlorhydria and absent intrinsic
factor.
8.0 BIOCHEMICAL BASIS OF MEGALOBLASTIC ANAEMIA
Inhibit DNA synthesis by reducing actually of enzymes in purine/pyrimidine
synthesis. In vitamin B12 deficiency disturbs folate metabolism and in folate
deficiency there is no polyglutamate hence decreased DNA synthesis.
9.0 CLINICAL FEATURES
Abnormal findings due to B12 and folic deficiency
Anaemia
Effects of B12 and folic acid on other tissues
10.0 DIAGNOSIS
1. Accurate and detailed history
2. Physical examination
3. Laboratory
11.0 LABORATORY DIAGNOSIS OF MEGALOBLASTIC ANAEMIA
a) General tests
1. Full heamogram the red cell indices
2. Bone marrow findings
3. Marrow cellularity
4. Erythropoeisis
5. Marrow iron
6. Biochemical findings
7. Serum bilirubin
8. LDH
9. Serum iron and ferritin
10. Urea and electrolytes
11. Uric acid

11. Uric acid

b) Specific tests
1. For B12 deficiency

2. Folate deficiency

a. Serum vitamin B12 assay (microbiological

assay or radiological assay)
b. Schilling Test detects B12 deficiency and
distinguish and detect lack of IF and
malabsorption
a. Serum folate
b. Red cell folate

3. Therapeutic trial
4. Test for causes of B12 folate deficiency
a. B12 deficiency
b. Serum antibodies against parietal cells, IF
and immunoglobulins
5. Gastric secretion IF and acid
6. Gastric biopsy, endoscopy
7. Barium meal and follow through
8. Fist tape work ova
9. Proteinuria
10. Folate deficiency - small intestines function - glucose, vitamin A,
fats and B12 absorption:

PERNICIOUS ANAEMIA (PA)

1.0 INTRODUCTION
Pernicious anaemia is a chronic disorder with a familial tendency common in
middle-age and elderly individuals of either sex where there is lack of secretion
of the intrinsic factor (IF) or production of antibodies against the IF and the
parietal cells (autoimmunity). The antibodies may be present in gastric juices. It
may be associated with other autoimmune disorders such as adrenal
insufficiency, malabsorption and hypopathyroidism. It causes megaloblastic
anaemia of insidious onset with a slow progress the clinical manifestations are
mainly due to vitamin B12 deficiency.
These include: Anaemia
Glossitis
Neurological disturbances such as neuropathy, sub acute combined cord
degeneration (SCCD) and retrobulbar neuritis
Gastrointestinal manifestations such as diarrhoea, anorexia, weight loss
and dyspepsia
Hepatospleenomegally
Congestive heart failure
Haemorrhagic manifestations
2.0 CAUSES

2.0 CAUSES
Parietal cell antibodies are found in: a. Atrophic gastritis
b. Chronic active Hepatitis
c. Thyroid disease
d. Addisons (renal) disease
e. Diabetes mellitus
f. Rheumatoid arthritis
g. Iron deficiency anaemia
3.0 PATHOLOGY

Morphological changes are found in the bone marrow, alimentary tract and the
central nervous system.
Bone marrow
Megaloblastic hyperplasia
Giant myelocytes
Alimentary canal
Atrophic gastritis - the tongue is shinny, glazed and red
Fundal atrophy with absent parietal cells
Central nervous system
Demyelination of the dorsal and lateral tracts of the spinal cord leading to
spastic pareisis and sensory ataxia.

Page 5 of 154
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

HAEMOLYTIC ANAEMIA
LEARNING OUTCOMES
At the end of the lesson the learner should be competent to: 1. Outline the causes of intrinsic and extrinsic (acquired) haemolytic anaemia
2. Describe the pathophysiology of haemolysis in acquired haemolytic anaemia
3. Describe the pathophyisology and pathology of sickle cell disease
4. Describe the clinical features and complications of sickle cell disease
5. Outline the pathology of thalassaemia
6. Explain the pathophysiology of the clinical features in these disorders
7. Describe the investigations and the expected results
1.0 INTRODUCTION
Haemolytic anaemia occurs in situations favouring increased red cell destruction
and shortening of life span of red cells. The red cell destruction could be
extravascular or intravascular haemolysis. The balance between increased
haemolysis and the consequent increased production of red cells by the bone
marrow determines the rate and magnitude of the ensuing anaemia once the
compensatory mechanisms fail.
a) Extra-vascular haemolysis occurring in macrophages in the reticuloendothelial system (spleen, bone marrow and the liver) with the
destroyed cells being phagocytosed by macrophages)
b) Intravascular haemolysis occurs in the circulation (and the contents
released into the plasma) due to mechanical trauma, cell membranes
damage by antibody and toxic chemicals.
2.0 HAEMOLYTIC MECHANISMS
Haemolytic processes can occur as a result of: a) Structural and functional abnormalities of the cell membrane.
b) Excessive physical trauma of the red cells
c) Rigidity of red blood cells secondary to precipitation
d) Abnormal molecular configuration of Hb.

The red blood cells possess some characteristics that favour their movement
without being destroyed by the phagocytic cells. These characteristics include:
a) Deformability

without being destroyed by the phagocytic cells. These characteristics include:

a) Deformability
b) Maintenance of the Surface: volume ratio
c) Maintenance of the integrity of the cell membrane
d) Cell membrane facilitation of movement of K+ and Na+
e) Maintained membrane proteins
f) Maintained membrane lipid.

Table 1: Mechanisms of Haemolysis

Mechanism

Example

a) Abnormalities of red cell membrane

i. Genetic membrane abnormalities
Spherocytosis
and elliptocytosis
ii. Alteration of lipid constitution

Liver disease
iii. Altered suphydryl reactivity
Oxidant drugs
iiii. Altered properties interaction
I
immune
Haemolytic anaemia
with complement or Ig.
v. Increased permeability, reduced plasticity Glycolytic enzyme
defects
b) High rigidity causing abnormal flow

drugs

i. Aggregation of Hb molecules
ii. Decreased solubility of Hb
iii. Inclusions (Heinz bodies)

c) Direct physical trauma

d) Turbulent flow clearage by

S CD
HbC disease
hallasaemia and
T

March/Karate haemoglobinuria

fibrin strands

Cardiac haemolytic anaemia

Microangiopathic haemolytic anaemia.

3.0 CONSEQUENCE OF HAEMOLYSIS

The consequences of haemolysis include haematological and biochemical changes
which affect the compensatory mechanisms. If the life span of the red cells is
reduced to 20 days (1/6) bone marrow production in increased by 6 times. This
strikes a balance between red cell destruction and formation and hence anaemia
is not evident (compensated erythropoeisis). If life span is shortened to 10 days
(1/12) marrow production is increased maximally by 8 10 times the normal
hence the balance is not attained and anaemia ensues.
In situations when the life span of red cells is shortened
the hyperplastic
marrow is susceptible to deficiency of essential factors thus underlying
conditions such as infections and pregnancy may limit the capacity of the
marrow to respond. Such conditions compensated haemolysis and hence
haemolytic anaemia.

conditions such as infections and pregnancy may limit the capacity of the
marrow to respond. Such conditions compensated haemolysis and hence
haemolytic anaemia.
a) Compensatory mechanisms: i. Increased Erythropoietin production (kidney) and
reticulocyte count
ii. Erthyroid hyperplasia of the bone marrow (Myeloid:
Erthyroid ratio)
iii. Demand for Fe/Folate
iiii. Extramedullary haemopoiesis (from early life) in
spleen, liver and lymph nodes
b) Hb catabolism
i. Hb is broken down into haem and globin components and the
haem iron is transported to the bone marrow for storage
ii. Residual porphyrin rings are broken down into bilirubin
leading to jaundice
iii. Polypeptide chains of globin are hydrolysed to amino acids
and taken into the circulation.
4.0 PATHOLOGICAL CHANGES IN HAEMOLYTIC ANAEMIA
The pathological changes seen in haemolytic anaemia are associated with
reduced RBC life span and bone marrow reaction
1. Reduced RBC life span
a. Measurement of red cell life
span using radioactive
sodium chromate. In normal
red cells there is gradual
loss of radioactivity
attaining 50% level at 30
days compared 50% level at
3 days in haemolytic
anaemia.
b. Effects of increased
degradation of haemoglobin
are: i. Increased
Bilirubin
levels bilirubina
emia,
jaundice,
urobinoge
n and
increased
stercobili
nogen.
ii.
Increased
iron
haemosid
erin may
be
deposited

be
deposited
in the
bone
marrow,
spleen
and liver
iii. Amino
acids
derived
from
protein
fractions
are reutilized
c. Effects of liberation of
haemoglobin

small
amounts of haemoglobin
are liberated into blood.
The
free
haemoglobin
rapidly forms a complex
with haptoglobin (a
globulin). This prevents the
harmful effects of free
haemoglobin
on
the
kidneys.
In
severe
intravascular
haemolysis
e.g. in malaria and ABO
incompatibility there is
excess
Hb
in
blood
(haemoglonimnaemia) and
haptoglobin reserves are
exhausted leading available
free Hb in plasma which
passes over into the kidney
(haemogloninuria).
2. Reactive bone marrow changes
There is marked morrow hyperplasia with extension of the red marrow into the
long bones.
5.0 CLASSIFICATION OF HAEMOLYTIC ANAEMIA
A. Hereditary (Intrinsic/Corpuscular) Causes
a. Membranopathies (abnormalities of red cell membrane)
i. Hereditary spherocytosis
ii. Hereditary elliptocytosis (ovalocytosis)
b. Enzymopathies (red cell enzyme defects)
i. Glucose-6-phosphate dehydrogenase (G6PD) deficiency
ii. Pyruvate Kinase (PK)
c. Haemoglobinopathies

ii. Pyruvate Kinase (PK)

c. Haemoglobinopathies
i. Sickle cell
ii. Thalassaemia
B. Acquired (Extrinsic/Extra-corpuscular) Causes
a. Immune
i. Auto-immune haemolytic anaemia - warm Abtype and cold
Ab type
ii. ISO-Immune haemolytic anaemia
1. Rhesus (Rh) incompatibility seen in Haemolytic
disease of the new born (HDN)
2. ABO incompatibility
b. Non-immune
i. Trauma

1. Microangiopathy
2. Valve prostheses
3. Body surface
4. Mechanical damage

ii. Hyperspleenism
iii. Physical agents drugs, chemical, toxins
iiii. Bacterial Infections (bacteraemia)
v. Parasitic disorders malaria
vi. Membrane defects e.g. liver disease
vii. Defective red cell maturation
Diagram 1: Classification of Haemolytic anaemia

INHERITED HAEMOLYTIC ANAEMIAS

This is anaemia caused by inherited or intrinsic/corpuscular factors which can be
grouped into three groups namely: A. Membranopathies (membrane disorders)
B. Enzymopathies (enzyme disorders)
C. Haemoglobinopathies (haemoglobin disorders)

A. MEMBRANOPATHIES
1.0 INTRODUCTION
The RBCs have a bilayer layer in which a number of integral membrane proteins
are embedded. The membrane is extensively and tightly connected with the
complex network of fibrous proteins forming the cytoskeleton in the cytoplasm
giving the red cells the two essential structural-functional characteristics of
biconcave disc shape and deformability.
The membrane consists of 50% proteins and 50% lipid (the lipid (the lipid consists
of cholesterol and phospholipid in equal proportions).
The red blood cells regulate their volume and water content through Na+ and K+
ions movement into and out of the cells.
The surface area of the red cell is about 140 m2. The cells become spherocytes
as Surface: Volume ratio decreases for example when there is loss of cell
membrane surface (microspherocytes) and gain in red cell volume
(macrospherocytes). An increase in the Surface: Volume ratio leads to the
appearance of target cells in the peripheral blood. Osmotic fragility test tests
the Surface: Volume ratio.
2.0 PATHOPHYSIOLOGY
The structural/functional of the abnormal membrane is recognized by
macrophages in the R.E.S, which destroy the abnormal red cells.
3.0 HEREDITARY SPHEROCYTOSIS (HS)
Hereditary spherocytosis is an autosomol dominant trait in which the red cell
membrane is abnormal. It is a cause of chronic anaemia

3.1. Pathogenesis
In hereditary spherocytosis there is a deficiency in the structural protein of the
red cell membrane called spectrin, which results in spheroidal contour and
smaller size of red cells that are not flexible. The membrane has reduced

In hereditary spherocytosis there is a deficiency in the structural protein of the

red cell membrane called spectrin, which results in spheroidal contour and
smaller size of red cells that are not flexible. The membrane has reduced
phospholipid and cholesterol content and the defect favours excessive
permeability to Na+ resulting in reduced surface: volume ratio and hence the
shape of the red cells change form a disc to a sphere. The rigid red cells have
reduced deformability and cannot pass through the spleen. They also have
increased osmotic fragility and permeability.
This process is countered by increased glycolytic process producing ATP (Energy)
needed by Na pump in order to pump Na ions from the cell resulting in an
increase cell turnover and loss of membrane lipid producing microspherocytes. In
the spleen there is less glucose available hence the destruction of cells due to
lack of deformability characteristic to enhance their passage in the red pulp of
the spleen. This may be explained two-fold red cells with decreased
deformability are retained in the spleenic pulp for unduly long time or the cells
find the environment of acidic pH and low glucose concentration in the spleenic
pulp unfavourable.
3.2. Clinical Features

Mild to moderate anaemia

Jaundice
Spleenomegally
Pigment stones
Chronic leg ulcers
Crises may occur as haemolytic crises during infections and pregnancy and
hypoplastic crisis due to reduced erythropoietic activity

3.3. Diagnosis

What is the
pathophysio
logy
of
these
symptoms?
. History and physical
examination

1
2

. Laboratory
a)
Full
hae
mog
ram

hae
mog
ram
b)
Feat
ures
of
Hae
moly
sis
incr
ease
retic
uloc
yte
coun
t,
incr
ease
urob
ilino
genu
ria.
3
.
Increase
osmotic
fragility
4
.
Negativ
e Direct
Coomb
s test

4.0 HEREDITARY ELLIPTOCYTOSIS (HE)

Hereditary elliptocytosis (ER) resembles hereditary spherocytosis except it is
milder and the cells appear elliptical (oval). Elliptocytosis may be acquired in
cases of iron deficiency and myeloproliferative diseases.

B. ENZYMOPATHIES
1.0 NORMAL CELL METABOLISM
The red blood cells loss their mitochondria during the development process when
the nucleus is extruded and therefore mature red blood cells have no capacity
for oxidative energy production but use pay as you eat energy producing
system by burning glucose as the main sources of fuel. There are two enzyme
systems that guide glucose metabolism process in two pathways hexose
monophosphate (HMP) shunt and Embden-Meyerhof glycolytic pathway, which are
controlled by two enzymes glucose-6-phosphate dehydrogenase and pyruvate
kinase respectively.
Defects in the enzymes results in defective cell metabolism hence the stability

Defects in the enzymes results in defective cell metabolism hence the stability
and integrity of the red cells is compromised.
Diagram 2: Enzymatic Pathways

Glucose
ATP
Hexokinase
ADP
Glucose-6-phosphate

ADP
Generation

Multiple
enzymatic steps

Phosphoenol pyruvate

6 phosphogluconate

NADP

NADPH

Glutathione reductase

ADP
ATP
Pyruvate

Oxidised
glutathione

Reduced glutathione

The energy produced maintains the integrity of the red cells and prevents
formation of spherocytes by maintaining the internal environment of the cells by
regulating the pump action while reduced glutathione maintains stability of
haemoglobin hence preventing formation of Heinz bodies (precipitation of globin
chains e.g. seen in infections). NADPH converts ferric state (Fe+++) of iron to the
ferrous (Fe++) state suitable for Hb synthesis and prevents oxidation of iron of
the haem.
2.0 HMP(AEROBIC GLYCOLYSIS)
HMP is stimulated by oxidants and involves reduction of NADP to NADPH. It
maintains an adequate level of decreased glutathione in red cell, which protects
the red cells against oxidant damage. Glutathione in reduced state maintains
stability of Hb by disposing H2O2 that would oxidize Hb to methaemoglobin
(which is a non-functional Hb)
3.0 PYRUVATE KINASE DEFICIENCY (PKD)
3.1. Introduction
Pyruvate kinase deficiency is an autosommal recessive trait (haemolysis occurs
only in homozygous state). It affects the ME pathway reducing utilization of
glucose for production of ATP in the red cell. It is the 2nd commonest cause of

only in homozygous state). It affects the ME pathway reducing utilization of

glucose for production of ATP in the red cell. It is the 2nd commonest cause of
inherited haemolytic anaemia.
3.2. Pathogenesis
There is accumulation of glycolytic intermediates 2, 3-DPG, phosphoenol
pyruvate with decreased availability of energy (poor glycolysis) and increased
reticulocytosis. The cells may use oxidative pathways using residual mitochondria
in young cells.
3.3. Clinical features
Presents in early childhood with:

Neonatal jaundice and haemolysis

Anaemia
Spleenomegally
Bone changes
Aplastic crises.

4.0 G6PD DEFICIENCY

4.1. Introduction
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is sex linked affecting
males (female carriers). It is carried on the X-chromosome and affects millions
of people worldwide. In Kenya it is encountered in western Kenya (Western and
Nyanza provinces). There are several variants of G6PD with the A-(negative being
more common and significant in black males. This variant confers protection
against malaria. A-G6PD sufferers have a shorted red cell life span but without
anaemia and these individuals develop haemolysis on exposure to oxidant stress
e.g. viral and bacterial infections, drugs (antimalarials, sulphonamides,
nitrofurantoin, aspirin and vitamin K ), metabolic acidosis and on ingestion of
fava beans (favism).

4.2. Pathogenesis
G6PD deficiency leads to decrease in glutathione destroying the integrity of the
red cells which are easily haemolysed under the stimulus of oxidative stress. The
oxidants damage cellular constituents; Hb is oxidized to methaemoglobin (nonfunctional Hb). Precipitation of globin produces Heinz Bodies, which consist of
denatured Hb and stromal proteins).
4.3. Clinical disorders/presentation
a) Drug induced Haemolysis.
b) Favism
c) Neonatal jaundice
d) Chronic Haemolytic anaemia.

c) Neonatal jaundice
d) Chronic Haemolytic anaemia.
e) Haemolysis associated illness (intercurrent).
Drugs
Haemolysis occurs 3 days after taking the drug and stops when reticulocytosis has
occurred (Reticulocytes have more G6PD) and Hb level may increase to normal.
1. Antimalarials such as aminoquinolones (primaquine and chloroquine) and
quinine
2. Sulphones Dapsone
3. Nitrofurans - Nitrofurations
4. Analgesics - ASA and Acetaphenetidin
5. Miscellaneous - Vit K, Chloromphenocal and quinidine
Favism
Favism is sensitivity of blood to the broad bean (Vicia fava) and consumption of
bean results in intravascular haemolysis.
Chronic Haemolysis
Rare/mild
Worsened by drugs.
Neonatal jaundice
Seen 2 5 days after birth
Kernicterus is common if exchange transfusion is not done.
The jaundice disappears but the deficiency remains.
Infections
Respiratory
Infection hepatitis
Diabetic acidosis

C. HAEMOGLOBINOPATHIES
1.0 INTRODUCTION
Haemoglobin molecule has four globin subunits linked at a specific site to a haem
group composed of an iron atom surrounded by a porhyrin ring. It consists of 2
pairs of coiled polypeptide chains and 4 prosthetic haem groups. Four
polypeptide chairs exist viz and , , d (determined by sequences of amino
acids).
Haemoglobin comprises of the haem component (made up of pyrrole rings and
contains Fe) that transports oxygen and globin, which is made up of chains of
amino acids and facilitates oxygen transportation by the haem by providing a
suitable environment. 100mls of aerated blood (oxygenated) carries 19 mls of
oxygen in Hb and 0.3 mls in solution of plasma.
Normal Hb consists of a pair of alpha (

) chains and another pair of either

Normal Hb consists of a pair of alpha (

) chains and another pair of either
beta chains (
) or gamma chains or delta ( ). The distribution of Hb types in
a normal red blood cell. A normal red cell has HbA (adult) which accounts for,
96-98%, HbF (foetal) that takes 0-10% and HbA2 that is 1.5 3%

The haemoglobin molecule undergoes structural changes during oxygen uptake

and release. In a deoxygenated state the beta chains rotate apart facilitating the
functional properties of haemoglobin.
Important features of the oxygen transport system are:
1. Haemoglobin has a high affinity for oxygen in the lungs and a low affinity
for oxygen in tissues.
2. Myoglobin has a higher affinity for oxygen than haemoglobin at low oxygen
concentrations.
3. Haemoglobin transports carbon dioxide back to the lungs where it is
expelled.
4. Haemoglobin releases its oxygen preferentially to exercising muscle rather
than to resting muscle

Diagram 3: Structures of Haem

2.0 HB VARIANTS
Haemoglobin variants have haemoglobin with abnormalities in amino acid
sequence of globin chains. These result from disorders of haemoglobin synthesis.
The beta chains contain abnormal amino acid sequence for example
haemoglobin S, C, D, D, and E.
Inheritance is via simple Mendelian pattern.
Substitution of external (polar) amino acids produce clinical disease in
homozygous state while changes at the internal (non-polar) results in
clinical disease in heterozygous state
Classification of Hb variants
1. HbS (Sickle cell syndromes)
2. Unstable haemoglobins
3. Thalassaemia
4. Low oxygen affinity haemoglobins

SICKLE CELL DISEASE, SCD (Hbss)

1.0 INTRODUCTION
Sickle cell disease (SCD) is an inherited disorder with cardinal features of chronic
haemolytic anaemia and recurrent painful episodes (crises). SCD results from
substitution of external amino acids as a result of mutant sickle cell Hb (HbS).
There are two varieties Hbss a homozygous state that is responsible for sickle
cell disease and HbSs a heterozygous state that results in sickle cell trait (may
develop a clinical disease under certain circumstances).
2.0 PATHOGENESIS
Sickle cell disease is caused by a single base mutation of adenine to thymine
resulting in substitution of valine for glutamic acid at the sixth position of the
amino acids of the beta-globin chain sequence. It has a similar geographical
distribution with falciparum malaria.
The red blood cells have reduced solubility with polymerisation of Hb when
deoxygenated, sickling, poor deformability and occlusion of the microvasculature
by sickle red cells.

by sickle red cells.

3.0 PATHOPHYSIOLOGY
Manifestations of SCD occur as a result of the effects of deoxygenation, HbS
polymerisation, cellular sickling and increased blood viscosity.
HbS Polymer
Oxygenated HbS is as soluble as HbA but the increase in surface hydrophobicity
of HbS tetramer (due to presence of valine instead of glutamic acid) causes
decreased solubility of deoxygenated molecule.
Polymerisation
Solubility of HbS is 17 mg/dl, which is lower than that of HbA (34 mg/dl).
Deoxygenation results in supersaturation of deoxygenated HbS solution, which
favours aggregation of HbS molecules resulting information of HbS polymer.
Cellular Sickling
Cellular sickling occurs in deoxygenated state of HbS

Cation Homeostasis and Cell Dehydration

Sickle cells become dehydrated and extreme dehydration of red blood cells
promotes HbS polymerisation. Dehydration occurs following of failure of energy
driven and gradient driven systems that affect movement of potassium and water
from sickle cells resulting in dehydration, polymerisation and sickling.
Oxidative Damage
Hb oxygenation and deoxygenation generates methaemoglobin S and oxidative
radicals producing oxidative stress, which causes damage and inhibition of
several enzyme systems. Oxidative damage destroys the lipids and proteins
making up the red cells.
Sickle cell adherence
There is increased adherence of sickled red blood cell to the vascular
endothelium. The adherence of sickled red blood cells to activated or injured
endothelial cells involves additional adhesive mechanisms with enormous
contribution of platelets and neutrophils.
Neutrophils
Polymorphonuclear neutrophils influence vaso-occlusion of vessels due to poor
deformability of PMNS compared with normal erythrocytes, increased adherence
to the endothelium and activation of PMN by sickle erythrocytes.
Immune Deficit
Sickle cell disease patients have immune deficiency due to: Defective spleenic function

Sickle cell disease patients have immune deficiency due to: Defective spleenic function
Opsonization
Abnormalities of the alternate complement pathway
Decreased antibody tires/levels
4.0 EFFECTS OF SCD
At the molecular level, HbS has low partial pressure of oxygen which leads
to low solubility resulting in formation and aggregation of crystals.
In the red cells HbS causes distortion of the red cells (sickling) which
precipitates destruction of red cells leading to chronic haemolytic
anaemia and eventually pigment gallstones.
In the tissues it causes blocking of the microcirculation leading to trapping
of RBCs in the liver and spleen (acute anaemia sequestration crises),
bone necrosis, bone infection (osteomyelitis by salmonella), stroke and
skin ulcers (e.g. legs).
5.0 PATHOLOGY
When the alpha chains move apart to give up oxygen the amino acid substitution
leads to locking of the adjacent ends of alpha chains with the abnormal beta
chains. The Hb molecules become stuck in rows distorting the red cells forming
sickle shapes in deoxygenated blood. This facilitates destruction of red blood
cells. Hbs gives up oxygen more readily than HbA hence provides good tolerance
to exercise at low Hb levels.
Mechanisms of Haemolysis
Premature destruction of the red blood cells can be extracellular or
intracellular. Extracellular haemolysis accounts for 2/3 of the destruction of
red cells. IgG antibodies mediate the process of red cell destruction.
Extracellular haemolysis is carried out by the macrophages through the process
of phagocytosis resulting from:

Impaired deformability of the cells

Changes in sickle cell membrane due to oxidative damage and sickling
Impaired inactivation of the complement system
Mechanical trapping of the cells

Intracellular haemolysis accounts for 1/3 of cases of red cell destruction, which
is usually fuelled by increased mechanical fragmentation and changes in proteins
in the cell membranes.
6.0 CLINICAL FEATURES
1. Life expectancy is reduced
2. Chronic anaemia due to chronic haemolysis
3. Exacerbation of anaemia due to: a. Aplastic crisis that presents with abrupt fall in Hb,
reticulocytosis and reduced red cell precursors.
b. Sequestration in the spleen and renal associated with
hyperhaemolysis
c. Spleenic sequestration (presents with acute
exacerbation of anaemia, persistent reticulocytosis,
tender enlarged spleen and hypovolaemia).

exacerbation of anaemia, persistent reticulocytosis,

tender enlarged spleen and hypovolaemia).
4. Acute phase episodes
5. Multi-organ failure involving the lungs, brain, kidney, liver, heart and
haemopoietic system which can be precipitated by vaso-occlusion and fat
embolus.
6. Psychological issues
7. Growth and development retardation
8. Infections
9. Crises

7.0 SCD CRISES

There are five types of crises that do occur in SCD: 1. Infarctive, Vaso-occlusive
(thrombosis) - painful crises
2. Sequestration crises
3. Aplastic crises
4. Haemolytic crises
5. Megaloblastic crises
6. Combination crises.
7.1. Infarctive Crisis
The infective crisis results from blockage of small blood vessels causing ischemia
and infarction.
Clinically it affects: Small bones of hard (ductilitys and & foot syndrome)
Heads of long bones
Lungs
Eyes
Abdominal pain
Pianism.
7.2. Sequestration Crisis
Sequestration crisis occurs in young children where there is sudden massive
pooling of blood in the spleen resulting is severe anaemia and circulatory

Sequestration crisis occurs in young children where there is sudden massive

pooling of blood in the spleen resulting is severe anaemia and circulatory
collapse.
7.3. Aplastic Crisis
Aplastic crisis occurs due to bone marrow failure causing severe anaemia with
associated infections.
7.4. Haemolytic Crisis
Haemolytic crisis occurs as a result of accelerated rate of haemolysis usually
peopled by infection and can also be part of the infective crises.
7.5. Megaloblastic Crisis
This results from the arrest of haemopoiesis due to severe folic acid deficiency
and a megaloblastic marrow.

8.0 SICKLE CELL TRAIT

Sickle cell trait is the heterozygous state of Hobs and accounts for 12 20% of
the cases.
9.0 COMPLICATIONS OF SCD
Complications of SCD are quite many and they arise from the chronic process of
disease punctuated with chronic haemolytic anaemia, which destroys organs due
persistent hypoxia of tissues, recurrent SCD crises (vaso-occlusive (thrombosis),
sequestrative, haemolytic, aplastic and megaloblastic) and acute infections.
A. Haematological - Haemolytic anaemia
B. Hepatobillary:
1. Jaudice/Icterus tendency to form gallstones
2. Obstructive jaundice- gall stones
3. Hepatic infarcts Liver failure
4. Infected Hepatic Infarcts Abscess.
5. Hepatic fibrosis due to repeated crises causes
reduced liver function leading to increased
jaundice.

C. Cardiopulmonary:
1. Congestive cardiac failure (CCF) secondary to
chronic anaemia and chronic hypoxia.

1. Congestive cardiac failure (CCF) secondary to

chronic anaemia and chronic hypoxia.
2. Cardiomegally Due to chronic anaemia and CCF
3. Myocardial infarction- vaso-occlusive/infective crisis
4. Pulmonary infarcts due to portal hypertension
leading to right ventricular failure (cor
pulmonalae)
5. Impaired pulmonary function pulmonary disease is
the leading cause of death in SCD. It can be acute
or chronic.
6. Aspiration pneumonia
7. Acute chest syndromes (dyspnoea, chest pain, fever,
tachypnoea and leucocytosis that occurs due to
vaso-occlusion, ischemia and infections)
8. Pulmonary fat embolus
9. Obstructive lung disease
10. Pulmonary hypertension
11. Airway hyperreactivity
D. Renal
Haematuria

from
microinfarcts
whose
development
is
favoured
by
the
acidic
environment, which favours sickling of red cells.
2. Renal failure due to renal infarcts/fibrosis.
3. Nephrotic syndrome
1.

E. Obstetric/gynaecologic
1. Delayed menarche
2. Dysmenorrhoea
3. Ovarian cysts
4. Pelvic infections
5. Fibrocystic disease of the breast
6. Foetal complications due to reduced
placental blood flow
a. Spontaneous abortions
b. IUGR
c. Pre-ecclampsia
d. Low birth weight
e. IUFD
7. Maternal
a. Crises are recurrent
b. Severe anaemia
c. Infections
d. Death

F. In males - there is priapism which if recurrent causes damage of the

penis and delayed development of secondary sexual characteristics.
G. Skeletal
MUCOUS
MEMBRANES

1. Ductilitys leads to shortening of fingers

and toes
BONE
hands
MARROW
and foot syndrome.
2. A vascular neurosis of head of femur and

MUCOUS
MEMBRANES
of the
alimentary
tract and
genitalia
REGENERATIO
N of the
destroyed and
aging mucous
membrane
cells cannot
balance
H. Occular
surface cell
losses
THINNING
(ATROPHY)
which results
in functional
I. Skin
deficiencies of
the affected
mucous
membranes

and toes
BONE
hands
MARROW
and foot syndrome.
2. A vascular neurosis of head of femur and
humerus.
Blood cell
3. Bossing of
precursors
skull. of all
4. Bone destruction
blood series
following
are
bone infarcts
formation.
affected
5. Osteonecrosis
6. Gouty arthritis
Many
Increased
7. Periarticular infarction
immature red
GROWTH
8. Effusion
cells are
FACTORS
9. Bone marrow infarction leads to anaemia
destroyed in
cause
and pancytopenia
the marrow
increased
due to their
cellularity
poor
1. Blindness ocular infracts, anterior
development
chamber
ischemia,
tortuosity
of
and the red
conjuctival
vessels,
renal
artery
cells released
occlusion, proliferative retinopathy and
into blood
retinal haemorrhage.
have a
2. Recurrent retinnitis
shortened life
span
HYPERCELLUL
MEGALOBLA
1.
Ulcers/Necrosis
ARITY OF THE
STIC
2. Leg ulcers lower
BONE
leg especially around
APPEARANC
the
lateral
and
MARROW
medial
malleolus and
E OF THE
skin.
Occurs
due
to
ischemia,
trauma and
RED CELLS
infections (organisms staph. aureas,
pseudomonas, strep. and bacteroids.

J. Reticulo-endothelial system
1. Autospleenectomy decreased immunity.
K. Central nervous system pathology occurs due cerebral thrombosis and
subarachnoid haemorrhage causing: 1. Transient ischaemic attacks (TIA)
2. Cerebro-vascular accident (CVA) presents
with hemiplegia, seizures, coma, speech
defects and visual impairment.
3. Cerebral infarction due to ischemia
4. Cerebral haemorrhage
5. Convulsions/seizures
6. Unexplained coma
7. Spinal cord infarction and compression
8. CNS infections
9. Vestibular dysfunction
10. Sensory hearing loss
11. Cerebral thrombosis
L. Reduced Immunity
1. Compliment activation is defective hence
decreased opsonization activity.
2. Hypospleenism leads to decreased
macrophage function, reduced antibody
production and defective opsonization.
3. Reduced Cell Mediated Immunity (CMI)
4. Intravascular haemolysis makes Fe more

production and defective opsonization.

3. Reduced Cell Mediated Immunity (CMI)
4. Intravascular haemolysis makes Fe more
readily
available
for
bacterial
metabolism.
5. Necrotic tissue following infarcts is a good
focus for infection

1.0 INTRODUCTION

THALASSAEMIA

Thalassaemia syndromes are heterogeneous groups of inherited anaemias

characterized by defective synthesis of the globin chains. Thalassaemia is a
Greek term that means the sea in blood. They are inherited as pathologic
alleles of one or more of the globin genes located on chromosomes 11 and 16.
This may occur as a result of total deletion or rearrangement of the loci points
that interferes with transcription, processing or translation of globin mRNA
The overall effects of the pathology in thalassaemia include reduced or absent
production of one or more globin chains and imbalanced synthesis of individual
and
chains. The main impact of thalassaemia results in defects in
and
chains, which are utilized in formation of HbA 2 2.. The impaired synthesis of
the globin chains results in hypochromic and microcytic red blood cells.
Unpaired , and chains are highly insoluble and form homotetramers, which
are incapable of releasing oxygen normally, are relatively unstable and
precipitate readily as the cell ages. Excess
globin chains occur in
thalassaemia and excess globin chains are seen in thalassaemia.
thalassaemia
In
thallassaemia there is reduced synthesis of b globin chains resulting in
diminished synthesis of HbA hence producing red blood cells that are
hypochromic and have abnormal shapes. The red cells contain less Hb resulting
in severe anaemia. Accumulation of a globin leads to deposition and
precipitation of the same. There are three varieties of thalassaemia namely
thalassaemia major, intermedia and minor.

2.0 PATHOPHYSIOLOGY
Reduced synthesis of b globin of HbA ( 2)
The red cells are microcytic and hypochromic
3.0 CLINICAL FEATURES

3.0 CLINICAL FEATURES

Occur after 6 months

Pallor
Irritability
Growth retardation
Abdominal swelling spleenomegally and hepatomegally
Jaundice
Facial and skeletal changes

4.0 LABORATORY

Red blood cells microcytes, hypochromia

Reduced Hb
Poikilocytes
Tear drop cells
Target cells
Reticulocyte count 2-8% (less than normal)
Increased white blood cells
Normal platelet count

5.0 EFFECTS
1. Skeletal
Osteoporosis
Bossing of the skull
Pathologic fractures
Overvgrowth of maxilla leading to malocclusion
2. Liver and gall bladder
Gall stones
Hepatomegally
3. Spleen
spleenomegally
hyperspleenism
4. Heart cardiomegally
5. Lungs abnormality of pulmonary function
6. Kidneys enlarged due to extramedullary haemopoeisis and urine contains
urates and uric acids.
7. Growth and endocrine
Growth retardation
Delayed menarche
Poor breast development
Decreased libido
8. Infections
Strep. pneumoniae
H. infleunzae
N. meningitis

 N. meningitis

ACQUIRED/EXTRINSIC/EXTRACORPUSCULAR CAUSES
Acquired haemolytic anaemias are caused by a variety of extrinsic or external
factors, which can be classified into various groups.
1.0 CLASSIFICATION OF ACQUIRED HAEMOLYTIC ANAEMIA

1.0 CLASSIFICATION OF ACQUIRED HAEMOLYTIC ANAEMIA

I. Immune haemolytic anaemia
i. Autoimmune haemolytic anaemia
1. Warm antibody type
2. Cold antibody type
ii. Isoimmune haemolytic anaemia
1. Haemolytic disease
newborn (HDN)
2. Transfusion reactions
a.
incompatibility
b.
incompatibility

of

the
ABO
Rh

II. Non-immune haemolytic anaemia

i. Infections
ii. Chemicals
iii. Drugs
iiii. Hyperspleenism
v. Mechanical damage (fragmentation syndromes)
vi. Burns
vii. Acquired cell membrane disorders

IMMUNE HAEMOLYTIC ANAEMIA

1.0 AUTOIMMUNE HAEMOLYTIC ANAEMIA
1.1.1. Introduction
Autoimmune haemolytic anaemia may occur as a primary (idiopathic) disorder
or may co-exist with another disease (secondary) or follow administration of
certain drugs. Autoimmune haemolytic anaemias are disorders in which
autoantibodies against antigens on the erythrocyte membrane cause a shortened
red blood cell life span. The autoantibodies are capable of damaging red cells
(membrane constituents). The precipitating factors are defective suppressor Tcells, drugs and infections.
There are three types of anti-erythrocyte autoantibodies: 1. Cold agglutinins (cold antibody type), which are mainly IgM capable of
causing clumping of red blood cells at low temperatures.
2. Cold haemolysins (IgG Donath-Landsteiner antibodies), which fix to the RBC
membranes in the cold activating the haemolytic complement cascade
when the cells are warmed to 37oC.
3. IgG warm antibody, which bind to erythrocytes at 37oC but fail to
agglutinate the RBCs.
1.1.2. Causes of Autoimmune Haemolytic Anaemia

1.1.2. Causes of Autoimmune Haemolytic Anaemia

1. Warm autoantibody haemolytic anaemia
a. Primary (idiopathic)
b. Secondary
i. Connective tissue disease
ii. Lympoproliferative disease
c. Drugs
e.g.
penicillin,
ampicillin,
methicin,
cephaloridine and carbenicillin.
2. Cold autoimmune haemolytic anaemia
a. Primary (idiopathic)
b. Secondary
i. Lympoproliferative disease
ii. Infections
e.g.
mycoplasma
pneumoniae, infectious mononucleosis and post
viral infection.

1.1.3. Immunologic Mechanisms of Red Cell destruction

Normally, red blood cells change from biconcave disk to a sphere at the end of
its life span where it triggers off the immune response initiating phagocytosis and
proteolysis by the macrophages, which then hand over the RBCs to the
lymphocytes. Lymphocytes of the immune system do ignore numerous selfantigenic determinants on the RBC membrane.
In autoimmune haemolytic anaemia: There is derangement of the immunological tolerance
Failure to develop tolerance of blood group antigens that occurs during
foetal life before the foetus develops the capacity to respond to
immunogens
Failure of the body to purge all self-reactive lymphocytes
Failure of immunoregulation by the T cell population. Humans posses T cell
population of CD4+ helper cells which augment the activity of B cells and
the CD8+ cytotoxic and suppressor cells.
1.1.4. Pathophysiology
Antibody production
Opsonized RBCs are recognized and cleared from the circulation by
macrophages in the spleen (main site) and liver (minor site).
Spleenic environment is conducive to immune clearance
1.2. Warm Antibody type
Is due to IgG class of autoantibodies that bind strongly to red blood cells at
37oC for example Rh iso-antibodies.
This results in microspherocytic red blood cells which undergo phagocytic
destruction in the spleen, liver and marrow (extravascular haemolysis)

 This results in microspherocytic red blood cells which undergo phagocytic

destruction in the spleen, liver and marrow (extravascular haemolysis)
Clinical example is the Coombs test
Coombs Test
Direct antiglobulin test where anti-human globulin serum will agglutinate
red blood cells, which have been sensitised or coated with an antibody
globulin.
Direct Coombs test patients (infant) red blood cells are treated with
antihuman globulin serum after being washed with saline to remove traces
of serum

Indirect Coombs test tests presence of antibodies in patients serum (the

mother). Normal compatible red cells are incubated with the patients
serum at 37oC for one hour. If the antibody is present it is coated on the
red cells. The red blood cells are then washed in saline and tested with
antihuman globulin serum.
A positive Coombs test is seen when there are Rh antibodies and some ABO
incompatibilities.
1.3. Cold Antibody type
IgM antibodies bind to red blood cells below 37oC facilitating autoagglutination of red blood cells and sensitising them to complement
activation and eventually destruction (Intravascular haemolysis) Coombs
tests.

1.4. Clinical Features

Jaundice
Anaemia
Spleenomegally (2 8 cm below the costal margin)
Signs of CCF
Lymphadenopathy
Fever
Hypertension
Renal failure
Rash
Petechiae and ecchymosis
Laboratory - reduced Hb, increased reticulocyte count (RC); Bone marrow
hyperplasia and megaloblastic features; Increased serum unconjugated
bilirubin
1.5. Therapy
Transfusion
Corticosteroids
Spleenectomy

 Corticosteroids
Spleenectomy
Plasma exchange

2.0 ISO-IMMUNE HAEMOLYTIC ANAEMIA

2.1. HDN (Haemolytic disease of the new born)
Foetal red blood cells enter maternal circulation provoking formation of
maternal antibodies (which are mainly IgG) to blood group antigens
foreign to the mother. The antibodies that are developed cross the
placental barrier to destroy the foetal red blood cells.
This follows shortly before or during labour, obstetric manipulation,
abortions e.g. Rh-positive baby Rh-negative mother.
When mild it results in transient jaundice and anaemia (congenital
Haemolytic anaemia); if severe it causes icterus gravis neonatorum with a
possible danger of development of Kernicterus.
In very severe cases there is severe anaemia leading to intra-uterine foetal
death (IUFD) or neonatal death due to anaemia and congestive cardiac
failure resulting in hydrops feotalis (Erythroblastosis feotalis)
2.2. Transfusion reactions
Are best illustrated by ABO incompatibilities

NON-IMMUNE HAEMOLYTIC ANAEMIA

Non-immune haemolytic anaemia can result from: 1. Fragmentation
2. Heat de-naturation
3. Mechanical damage
4. Osmotic attack
5. Hyperspleenism
6. Infection
7. Alteration of RBC surface
8. Liver disease
9. Renal disease
10. Venoms, bites, stings, toxins
11. Drugs and chemicals
1.0 MECHANICAL DAMAGE (FRAGMENTATION SYNDROMES)

Fragmentation haemolysis or microangiopathy occurs when mechanical forces

disrupt physical integrity of the red cell membrane. The result is
microangiopathic haemolytic anaemia (MAHA). Intravascular haemolysis due to
mechanical destruction of red blood cells due to pathological changes in small
blood vessels. There is widespread deposition of fibrin in small blood vessels due
to injury (microangiopathy) or activation of clotting mechanism. The fibrin mesh
fragments the red blood cells
Seen in AGN, APH, pre-eclampsia, malignant hypertension, renal cortical
necrosis, septic shock prosthetic heart valves, S.L.E and P.N. It is associated with

Seen in AGN, APH, pre-eclampsia, malignant hypertension, renal cortical

necrosis, septic shock prosthetic heart valves, S.L.E and P.N. It is associated with
thrombrocytopenia
Causes
1. Damaged microvasculature
a. Associated with pregnancy pre-eclampsia/eclampsia
b. Malignancy
c. Vasculitis
d. Abnormalities of renal vasculature
e. D.I.C
2. Atrio-ventricular malformations
3. Cardiac abnormalities replaced valves, prosthesis, drugs

2.0 MARCH HAEMOGLOBINURIA

March haemoglobinuria is an acute haemoglobinuria that follows marches, long
distance running, marathon and karate resulting in red cell destruction in the
feet and hands
3.0 HEAT DENATURATION
Normal red blood cells will undergo budding and fragmentation when
exposed to a temperature of 49oC in vitro.
Blood for transfusion should warmed to body temperate to prevent budding
and fragmentation
Patients with severe burns and heat stroke may have haemolytic anaemia
4.0 MECHANICAL TRAUMA
March haemoglobinuria in soldiers after a long march, joggers after running on a
hard road and in karate or conga drumming following long periods of practice.
5.0 OSMOTIC ATTACK
Abrupt changes in osmolality of cell can cause haemolysis. Fresh water drowning
may be associated with swelling of red cells.
6.0 HYPERSPLEENISM
In all organs of the monocyte-macrophage system (reticulo-endothelial system
R.E.S) blood cells leaving arterial bed are unloaded into channels, which the red
cells must pass through the wall of the sinusoids to re-enter the circulation. The
normal RBC is a discocyte having a surface area 40% larger than a sphere of that
volume. This large surface area (8 um) allows the red cell diameter to twist,
elongate and deform sufficiently so as to squeeze through 2-3 um slits. The
excess surface area is also called the SA: V ratio, which is 1.4.

elongate and deform sufficiently so as to squeeze through 2-3 um slits. The

excess surface area is also called the SA: V ratio, which is 1.4.
Any condition that reduces the SA: V reduces the ability of the red cells to
traverse sinusoidal slits because a plump sphere cannot deform adequately.
Interference with interaction of cytosol and the cell membrane impairs red cells
ability to deform e.g. Heinz bodies lie adjacent to the cell membrane interfering
with the smooth movement of the membrane over cytosol (tank treading)
hence the cells are selectively blocked from leaving the spleenic cords to enter
the sinuses.

Inflammation or infection enhances the ability of the spleenic macrophages to

attack and ingest red blood cells.
The spleen provides a double filter (cords and sinuses) and blood cells must be
remarkably deformable to pas through. Slow passage of blood cells through the
spleen permits selective action by macrophages that detect several sorts of
alterations in blood cells. Macrophages hold, retard, modify (pitting function)
or remove (culling function) of the red cells identified
All activities of the spleen are accentuated in a large spleen and if the increased
activity hyperspleenism ensues.
The spleen enlarges and there is delay in release of leucocytes and platelets
trapped in the spleen increasing red cell destruction. The hyperspleenism
syndrome comprises of spleenic enlargement, decrease in one or more of the cell
types in peripheral blood, normal or hyperplastic marrow and correction of cell
defects by spleenectomy.
Causes: a) Congestive Spleenomegally
b) Leukaemia/lymphoma
c) Tuberculosis
d) Malaria
e) Kala-azar
f) Brucellosis
g) S. L. E
h) Feltys syndrome (Rheumatoid arthritis, spleenomegally and leucopenia)
7.0 INFECTIONS
The mechanisms by which infections cause haemolysis are: 1. Direct parasitization of red cells e.g. malaria, toxoplasmosis, bacterial
infection
2. Immune mechanisms e.g. cold agglutins
3. Induction of hyperspleenism e.g. malaria, schistosomiasis
4. Alteration of red cell surface topology e.g. Haemphilus influenzae
5. Release of toxins and enzymes e.g. HIV.
8.0 DIRECT PARASITIZATION
This occurs in malarial infection by P. falciparum, P. vivax and P. malariae. The
sporozoites are injected by the mosquito in its saliva and make their way to liver
cells where after 1-2 weeks they become merozoites, which burst out of their

This occurs in malarial infection by P. falciparum, P. vivax and P. malariae. The

sporozoites are injected by the mosquito in its saliva and make their way to liver
cells where after 1-2 weeks they become merozoites, which burst out of their
cells and into the blood stream. The parasite attaches on a specific receptor on
the red cell surface. This allows engulfment of the red cells by the parasite
which immediately co-opts the red cells metabolic machinery, degrades and
ingests Hb, grows eventually bursting out of the red cells and the cycle begins
again.
The red cells are lysed both intravascularly by direct parasitic destruction and
extravascularly as a result of changes in the spleenic microvasculature and
activation of monocyte-macrophage system. Plasmodium falciparum alters the
spleenic function and non-parasitized cells inciting their premature destruction.
a) Malaria
Intravascular and extravascular (spleen) haemolysis of infected red
blood cells
There is dyshaemopoiesis associated with reduced haemoglobin,
hyperplastic
marrow,
monocytosis,
neutropenia
and
thrombocytopenia.
D.I.C e.g. black water fever
b) Toxoplasmosis
c) Bacterial infarction cause depressed erythropoeisis and acute haemolysis
Severe infections cause: o D.I.C due to effects of endotoxins produced
by Gram negative organisms
o Mechanical haemolytic anaemia
o
Intravascular
haemolysis
(Clostridium
welchii)
o Microspherocytosis
o Fragmentation of red cells
o Renal failure
Mechanism of haemolysis includes
o D.I.C
o Direct destruction of red blood cells e.g. by
lecithinase, proteoyltic toxins.
9.0 ALTERATION OF RED CELL SURFACE BY BACTERIAL PRODUCTS
Infections may alter the red cell surface facilitating haemolysis e.g. H.
influenzae. Bacterial products cause haemolysis by direct damage to the cells or
release of enzymes by organisms. The enzymes degrade the phospholipids of the
bilayer cell membrane and structural membrane proteins e.g. clostridium
infections.
There is haemolysis that is less understood in infections such as HIV, CMV and
visceral leishmaniasis.
10.0 CHEMICALS
Chemical can cause haemolysis due to their oxidant effects and effects of their
metabolites e.g. enzymopathies. The notable chemicals include Pb, Cu and

Chemical can cause haemolysis due to their oxidant effects and effects of their
metabolites e.g. enzymopathies. The notable chemicals include Pb, Cu and
potassium chlorate.
11.0 VENOMS, BITES, STINGS, TOXINS
Results in intravascular haemolysis
Involves bee/wasp stings, spiders and snake venoms
There is bleeding as a result of development of DIC
12.0 DRUGS
Drugs cause haemolysis via two main processes: )
A
b
V
s
d
r
u
g
s

Talking Point
1. Who are predisposed to iron deficiency
anaemia?
2. What is the heamogram picture in
sideroblastic anaemia?
3. Describe the aetiology of anaemia of chronic
disorders?
4. What are the causes of hypochromic
anaemias?
5. State the laboratory diagnosis of
hypochromic anaemias
6. What is the pathophysiology of anaemia in
Ab vs. drugs
chronic disorders?
7. What is the heamogram picture in anaemia of
Drugs + RBC
drug/red cell complex (antigen)
chronic disorders?
8. What are the functions of folate and vitamin
B12?
b production.
9. How do we getAfolate
and vitamin B12?
10. How would you make the diagnosis of iron
Is mediated mainly by IgG and the haemolysis not severe, it ceases with
deficiency
withdrawal
of drugs.
highindoses
11. What
is the Examples
relevant are
history
iron of penicillin and cephalosporins.
Ab combine
with
the
drug/plasma
protein
complex (antigen complex), which
deficiency
binds to red blood cells facilitating their destruction. The anaemia ensuing is

i
)
A
b
V
s
r
e
d
b
l
o
o
d
c
e
l
l
s

withdrawal
of drugs.
highindoses
11. What
is the Examples
relevant are
history
iron of penicillin and cephalosporins.
Ab combine
with
the
drug/plasma
protein
complex (antigen complex), which
deficiency
binds12.
to What
red blood
cells
facilitating
their
destruction.
The anaemia ensuing is
is the pathophysiology of the clinical
severe/moderate.
Examples
- digoxin,
quinidine, phamacetin, sulphonamides and
features of iron
deficiency
disorders
chlorpropramide.
13. Explain the basis of investigations in iron
deficiency anaemia.
14. How does iron deficiency anaemia cause
Ab vs. normal
RBCs
impaired
immunity?
15.- What is Plummer-Vinson syndrome and
Drugs induce
autoantibodies
against red blood cells inducing haemolysis e.g.
Patterson-Kelly
syndrome?
methyldopa.
16. What is the explanation of the blood picture
seen in iron deficiency anaemia?
17. What signs are you most likely to elicit in a
13.0 BURNS
patient with severe chronic iron deficiency
anaemia?
Severe burns cause intravascular haemolysis with microspherocytosis and
18. What are the clinical features of B12 and
fragmentation of red blood cells in the peripheral blood
folic acid deficiency anaemia?
19. What is the haemogram picture in
megaloblastic anemia?
14.0 CELL MEMBRANE DISORDERS
20. Compare and contrast pernicious anaemia
and B12 deficiency anaemia.
Cell membrane defects are seen in:
21. What are the physical examination findings
Lipid disorder cholesterol and phospholipids
in megaloblastic anaemia?
Liver disease
22. What are the pathology, clinical features
Vitamin
E deficiency
- Vitamin
and investigations
in aplastic
anaemia? E prevents auto-oxidation of the
fattyblood
acidspicture
in red cell
membrane.
23.unsaturated
What is the
in aplastic
anaemia?
24. What are the causes of bone marrow
15.0 LIVER
DISEASE
failure?
How does it present?
25. Compare and contrast pernicious anaemia
Haemolysis
inanaemia.
liver disease is not profound but contributes significantly to
and aplastic
the severity of anaemia when associated with defects in red cell
production and blood loss.
Due to spleenomegally secondary to portal hypertension
Abnormal membrane SA: V ration impair membrane fluidity
Acute alcoholism interferes with red cell intermediary metabolism
16.0 RENAL DISEASE
Anaemia in renal disease can occur as a result: Impaired red cell production
Impairs platelet function leading to occult loss
Disease of small renal arterioles causes fragmentation haemolysis in seen in
pre-eclampsia and malignant hypertension
Uraemia shortens the life span of red cells
Reduced activity of enzymes G6PD and glutathione peroxidase

Page 13 of 155
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

HEAMORRHAGIC ANAEMIA
Learning Outcomes:
At the end of the lesson the learner should be competent to: 1. Outline the relevant history in a patient with bleeding disorder
2. Describe causes and features of thrombocytopenia
3. Describe the defects of platelets
4. Describe disorders of platelet function
5. Describe disorders of blood vessels that result in haemorrhage
6. Describe the causes and pathology of disorders of coagulation.

1.0 INTRODUCTION
Haemorrhagic disorders can be inherited (congenital) or acquired which fall
under three main classes of disorders of (1) platelets, (2) blood vessels and (3)
coagulation and fibrinolysis.
Coagulation defects and trauma usually initiates bleeding whose persistency and
severity characterizes the disease(s). Disorders of platelet disorders and blood
vessels result in spontaneous bleeding (purpura) into the skin, mucous
membranes and from the skin surface
In clinical practice, patients with haemorrhagic disorders do present with various
forms of bleeding which the clinician needs to get in-depth history in order to be
in a position to get adequate and relevant information regarding the bleeding
disorders.

2.0 HISTORY IN A PATIENT WITH BLEEDING DISORDER

1. Epistaxis
2. Gingival haemorrhage
3. Skin haemorrhage petechiae and bruises
4. Tooth extractions ages at extractions
a. Deciduous
b. Permanent molar, other
c. Duration of bleeding packing, resuturing, transfusion
d. Estimated blood loss
5. Bleeding from minor cuts
6. Excessive bleeding from major cuts
7. Haemoptysis spontaneous or associated with respiratory illness
8. Haematemesis
9. Haematuria

7. Haemoptysis spontaneous or associated with respiratory illness

8. Haematemesis
9. Haematuria
10. Haematochezia
11. Melena
12. Central nervous system bleeding haemorrhagic stroke
13. Venipuncture bleeding sites
14. Ophthalmic bleeding subconjuctival, retinal, retrobulbar
15. Menses/menstrual flow
16. Childbirth
How do these
17. Haemathroses joints involved, frequency
characteristics affect the
18. Previous surgical procedures
normal functioning and
19. Bleeding at circumcision
destruction of the red
20. Telengiectasis mucous membranes, skin, gastro-intestinal tract
blood cell?
21. Connective tissue disorders
22. Wound healing
23. Medications iron, birth control pills, aspirin
24. Family history of bleeding

3.0 CAUSES OF HAEMORRHAGIC ANAEMIA

The causes of haemorrhagic anaemia includes: 1. Disorders of platelets
2. Disorders of the blood vessels
3. Disorders of coagulation and
fibrinolysis

4.0 PLATELET DISORDERS

4.1. Introduction
Platelets contain dense bodies that contain ADP, serotonin, calcium and alpha
granules (containing fibrinogen and platelets factor 4). Platelet aggregation
leads to synthesis of prostaglandins. An estimated 15 X 106 megakaryocytes/kg
body weight produce about 1500 platelets each. The normal platelet count is 150
400 X 109/L. Platelets disorders can result from thrombocytopenia and platelet
defects (structure and function)

4.2. Thrombocytopenia
4.2.1. Introduction
Thrombocytopenia results when the total platelet count is less than 140 x 10
9
/L. Thrombocytopenia occurs when platelets are lost from the circulation faster
than they can be replaced by the bone marrow. This could be because of failure
of production, increased rate of removal from the circulation or a combination of
the two.
Spontaneous bleeding will occur when the platelet count is less than 20 x 10 9/l
while a platelet count of 20 50 x 10 9/l will result in bleeding when infections
precipitate the process of bleeding. Bleeding is uncommon if the platelet count
is over 50 X 109/L.
4.2.2. Causes and Mechanisms
Causes of thrombocytopenia can be inherited or acquired. Impaired production
of platelets leads to reduced mean platelet volume (MPV) and may present with
anaemia and/or leucopenia.

Causes of thrombocytopenia can be inherited or acquired. Impaired production

of platelets leads to reduced mean platelet volume (MPV) and may present with
anaemia and/or leucopenia.
1. Impaired production of platelets leads to reduced mean platelet volume
and may present with anaemia and/or leucopenia.
a. General bone marrow failure due to: 1. Hereditary bone marrow aplasia
2. Leukaemia
3. Aplastic anaemia
4. Megaloblastic anaemia
5. Myeloma
6. Myelofibromatosis
7. Bone marrow infiltration by solid tumours
8. Radiation
9. Toxins
10. Uraemia
11. Alcoholism
b. Selective reduction in megakaryocytes (cells used to
produce platelets
1. Hereditary
2. Dyshaemopoiesis
3. Drugs - Drugs e.g.
chloromphenicol,
cotrimoxazole,
phyenylbutazone, Gold,
cytotoxics
4. Chemicals
5. Viral infections mumps,
rubella,
measles,
varicella,
cytomegalovirus,
infectious
mononucleosis, chicken
pox,
haemorrhagic
fevers,
hepatitis,
parvovirus and HIV
2. Excessive destruction of platelets (diminished platelet survival - there is
increased destruction of the platelets and the MPV is increased and the
platelets appear larger than normal.
a. Immune mediated
1. Platelet alloantibodies neonatal and post-transfusion
purpura
2. Platelet autoantibodies - Idiopathic thrombocytopenia purpura
(I.T.P) and following bone marrow transplant
3. Post transfusion purpura
4. Other mechanisms
a) S.L.E, autoimmune disorders
b) Malaria
c) AIDS
d) Drugs - Analgesics/anti-inflammatory drugs (aspirin,
paracetamol, phacetin, gold salts), Antinicrobials
(penicillins, sulphonamides, trimethoprim, P.A.S,),
Sedatives/anticonvulsants
(diazepam,
sodium
valproarte), Diuretics (frusemide, chlorothiazides,

Sedatives/anticonvulsants
(diazepam,
sodium
valproarte), Diuretics (frusemide, chlorothiazides,
acetazolamide),
Antidiabetics
(chlorpropamide,
tolbutamide)
and
Others
(digoxin,
heparin,
methlydopa, oxyprenolol, quinine, quinidine)
e) Other viral infections
b. Coagulation
1.
Disseminated
intravascular coagulation (D.I.C)
2.
Thrombosis
thrombocytopenia purpura
3. Haemolytic uraemia
syndrome
4. Platelet aggregation
e.g. drugs - heparin
c. Sequestration hyperspleenism
d. Dilutional - Loss of platelets form the systemic circulation and
massive or exchange transfusion
4.2.3. Pathogenesis
Thrombocytopenia can be caused by two general mechanisms: 1. Platelet under production
2. Increased platelet destruction
a. Immune
b. Coagulation
c. Sequestration
d. Dilutional loss
Thrombocytopenia due to underproduction is usually accompanied by
pancytopenia. Thrombocytopenia resulting from platelet destruction due to
immune or nonimmune mechanisms ensues when the rate of platelet destruction
surpasses bone marrow ability to produce platelets. In sequestration there is
redistribution of platelets, which results in pooling of platelets in the spleen.
Haemodilution is caused by administration of colloids, crystalloids or blood
products resulting in decrease in the number of red cells, white cells and
platelets.
4.2.4. Hypersensitivity to Drugs
Hypersensitivity to drugs develops after a single dose (in a previously sensitised
individual) or protracted treatment course. The drugs and their metabolites act
as haptens. Binding of the drugs and their metabolites to plasma proteins forms
the antigenic complexes with the antibodies produced being mainly IgG. The
antigen-antibody-platelet complex formed is destroyed by phagocytosis
.
Examples of drugs: - Chlorothiazides, Digitoxin, Methyldopa, PAS (Para
Aminsalicylic Acid), Paracetamol, Quinine, Quinidine, Sulphonamide
4.2.5. Autoimmune Thrombocytopenia

4.2.5. Autoimmune Thrombocytopenia

There is development of IgG autoantibodies to the platelets. Antibodies bind to
platelets encouraging destruction by phagocytosis. May occur as ITP or in
association with other diseases such as S.L.E and myasthenia gravis
4.2.6. Clinical Features
The commonest symptoms and signs of haemorrhage in platelet disorders are of
purpuric type: spontaneous skin purpura and ecchymosis, bleeding from mucous
membranes, particularly the nose, multiple small subcutaneous bruises and
menorrhagia. The important causes of death are intracranial haemorrhage and
massive gastro-intestinal bleeding.
5.0 PLATELET DEFECTS
Platelet defects which can be in terms of structure and function can be acquired
or hereditary. Acquired platelet dysfunction can be due to chronic renal failure
and drugs such as A.S.A, dextran and carbenicillin.
5.1. Platelet Function
Disorders of platelet function do reduce the ability and capacity of the platelets
to effectively participate in blood coagulation hence resulting in bleeding
disorders.
The disorders of platelet function include: a. Disorders of platelet adhesion due to lack of von Willebrand
factor and some necessary glucoproetins and deficiency of platelet
collagen receptors.
b. Disorders of platelet aggregation
c. Disorders of platelet secretion of four types of granules: - the
dense granules (containing ADP, ATP, calcium, serotonin), -granules
(contain proteins) lysosomes and microperoxisomes.
d. Disorders of platelet procoagulant activity, which provides a
surface for attachment of coagulation factor X and prothrombin.
5.2. Hereditary
Deficiency of platelet membrane glycoprotein complex and platelet receptor for
von Willebrand factor(vWF)
5.3. Acquired
1. Myeloproliferative disorders
a. Thrombocythaemia
b. Chronic myeloid leukaemia
c. Polycythaemia vera
d. Myelofibrosis
2. Acute leukaemia
3. Chronic renal disease
4. Chronic hypoglycaemia
5. Drugs NSADS (aspirin, indomethacin, ibobrufen, meclofenamic acid,
piroxicam),
-lactam
antibiotics
(penicillins,cephalosporins),

5. Drugs NSADS (aspirin, indomethacin, ibobrufen, meclofenamic acid,

piroxicam),
-lactam
antibiotics
(penicillins,cephalosporins),
nitrofurantoin, heparin, cardiovascular drugs (propranolol, nifedipine,
verapamil, quinidine), psychotropic drugs (amitriptyline, chlorpromazine,
promethazine) anaesthetics (lidocaine, halothane), narcotics (heroin),
dextrans and food/food additives (ethanol, onions, turmeric and clove).
6. Acquired storage pool deficiency
a. Autoimmune disease
b. D.I.C
c. Severe burns
d. Valvular heart disease/cardiopulmonary bypass surgery
e. Renal transplant rejection

6.0 DISORDER OF BLOOD VESSELS

6.1. Introduction
Disorders of
haemorrhage
petechial and
Easy bruising
vessels.

increased vascular fragility can play a role in causation of

even though the bleeding is massive but most often it induces
purpuric haemorrhages.
and bleeding into the skin characterize disorders of the blood

6.2. Causes
1. Congenital

a. Connective tissue disease e.g. Marfans syndrome and

osteogenesis imperfecta
b. Hereditary telegiectasia

2. Acquired

a. Severe infections which cause vasculitis and DIC due

to septicaemia, meningococcal infections, measles
and typhoid

3. Allergic reactions

a. Autoimmune disorders e.g. SLE and Rheumatoid

arthritis
b. Drug reactions deposition of immune complexes in the
vessel wall and production of hypersensitivity
vasculitis

4. Others

a. Poor vascular support impaired formation of collagen

e.g. in scurvy and loss of perivascular supporting
tissue e.g. in Cushings syndrome.
b. Senile purpura
c. Easy bruising syndrome

7.0 COAGULATION DISORDERS

Classification
1. Hereditary
a.

1. Hereditary
a.

Haemophilia
i.

Haemophi
lia
A
(Classical
)
ii.
Haemophi
lia
B
(Christma
s disease)
b. Von
Willebrands disease
2. Acquired

a. Auto-antibodies to clotting
factors
b. Vitamin K. deficiency
c. Vitamin K. antagonism
d. Vitamin C deficiency
e. Liver disease
f. Alcohol
g. D.I.C.
h.
Haemolytic
uraemia
syndrome.

HEREDITARY COAGULATION DISORDERS

A. HAEMOPHILIA A (CLASSIC)
1.0 INTRODUCTION
Haemophilia A is the most common with an incidence 1:10000. It is a sex-X-linked
recessive disease that obeys the Mendelian pattern primarily affecting the males,
as the females are carriers.
Carrier Female
X
1
X

XX

1Y
X
Affected male

Normal Female
X

Normal Male
X

1X
X
XY
Carrier female

Affected Male
1
X

XX1
Carrier female

X1

X1X

XY

normal male

carrier female

XY

normal male

2.0 PATHOLOGY
Haemophilia A (classic) results from Factor VIII deficiency. Factor VIII
(Antihaemophilic factor) is a large protein synthesized in the liver. It regulates
activation of Factor X during the process of coagulation of blood.

3.0 CLINICAL FEATURES

1. Haemarthrosis especially in the weight bearing joints
2. Bleeding into muscles of the calf, thigh, fore arm and the Iliopsoas sheath
3. Microscopic haematuria
4. Intracranial haemorrhage (low incidence)
5. Intramuscular haematoma that can lead to ischemia, infarction, gangrene
and paralysis.
6. Patients bleed easily following trauma, surgery, circumcision and tooth
extraction
7. Epistaxis
8. Petechial haemorrhage
9. Ecchymosis
What
is
the
pathogenesis
of
4.0 DISEASE GRADINGthe
clinical
features
of
Depends on level of inherited
deficiency
1. Severe <1% Factor VIII activity
spherocytosis?
2. What history will be 2. Moderate 1-5% Factor VIII activity
relevant in such 3. Mild > 5% Factor VIII activity
patients
Usually the clinical
features
correlate
3. What will
you with the grading of the disease i.e. if the
disease is severe; patients bleed even without trauma/injury while in mild
find on physical
disease they bleed after considerable trauma.
1.

examination

5.0 INVESTIGATIONS
Do the coagulation screen
1. Bleeding time (2 7 minutes)
2. Clothing time (3-7 minutes)
3. Platelet count (150 400 X 109/L)
4. Prothrombin time; PT (11-15 seconds)
5. Thrombin time (6-9 sec)
6. KCCT; Kaolin Cephalin Clotting Time (35-45 seconds)
7. Full heamogram and ESR
8. Clotting Factor Assay

7. Full heamogram and ESR

8. Clotting Factor Assay
6.0 DIAGNOSIS
1. Accurate and comprehensive history
2. Accurate physical examination
3. Laboratory investigations

B. HAEMOPHILIA B (CHRISTMAS DISEASE)

Haemophilia B (Christmas disease) is an inherited sex linked recessive disorder
that results from deficiency of clotting Factor IX which is a protein synthesized in
the liver and requires vitamin K for activity. Its clinical presentation resembles
that of Haemophilia A.
C. von WILLEBRANDS DISEASE
1.0 INTRODUCTION
von Willebrand disease (Dr. Erich von Willebrand 1926) is inherited as Mendelian
autosomal dominant gene and affects both sexes equally with an incidence of
1:800 1000.
2.0 PATHOLOGY
The lesion is a quantitative or qualitative abnormality of Von Willebrand (vWB)
VIIIR factor which is a plasma glucoprotein whose functions are:
a. A factor for platelet adhesion to
vascular subendothelium hence its deficiency
results in prolonged bleeding time, low platelet
adhesion (this differentiates it from Haemophilia).
b. Is a carrier protein for Factor VIII that
is a coagulation protein and protects it from
premature destruction.
Describe the full haemogram picture
expected
spherocytosis
vWB
factorinishereditary
synthesized
by vascular endothelium and megakaryocytes and its
abnormalities include reduced synthesis of all oligomers associated with amino
acid defects, which prevents formation of complexes.
3.0 CLINICAL PICTURE
Excessive bleeding from cuts, injuries, epistaxis, gastro-intestinal tract,
gums, menorrhagia and haemoarthrosis.
Blood picture
o Increased bleeding time
o Increased KCCT
o Defective platelet adhesion
o Defective platelet aggregation
o Normal platelet count
o Reduced levels of Factor VIII
4.0 DIAGNOSIS
1. Accurate and comprehensive history
2. Accurate physical examination

1. Accurate and comprehensive history

2. Accurate physical examination
3. Laboratory investigations Do a coagulation screen

ACQUIRED BLEEDING DISORDERS

Acquired bleeding disorders include: 1. Vitamin K deficiency/antagonism
2. Liver disease
3. D.I.C.
4. Auto antibodies against clotting factors
5. Vitamin C deficiency
6. Alcohol
7. Haemolytic uraemia syndrome
1.0 VITAMIN K DEFICIENCY
1.1. Introduction
Vitamin K is a fat-soluble vitamin that is essential for synthesis of functional
factors II, VII, IX, X. and protein C and S. These factors are thereby able to bind
calcium ions and become biologically active and protein C and S act as natural
anticoagulants by inhibiting the active forms of factors V and VIII.
Its source is green vegetables and synthesis by colonic flora (bacteria). Vitamin K
is absorbed in the upper part of the small intestine in the presence of bile and i
stored in the liver. It plays a role in carboxylation of precursors of factor II
(prothrombin). Vitamin K is usually converted to the active form Vitamin K
epoxide.
Vitamin K is important in modification of blood coagulation proteins, factors VII,
IX and X, and prothrombin and plasma regulatory proteins proteins C and S
making them relevant in the process of blood clotting.
Diagram 1: Vitamin K Metabolism

Vitamin K

Liver hepatocytes

Vitamin K- epoxide (active form)

Warfarin

1.2. Causes of deficiency

1.2. Causes of deficiency

1. Inadequate dietary intake
2. Mal absorption
3. Loss of storage sites hepatocellular disease
4. Broad-spectrum antibiotics
Embden-Meyrhof
Hexose
Monophosphate
(EM)
Glycolytic
(HMP) shunt
Pathway

Aaerobic 10%)
1.3. Risk(Anaerobic
groups
90%)
1. Neonates
2. Malnutrition
3. Antibiotic therapy especially cephalosporins
4. Parenteral nutrition without K supplement e.g. p surgery
G6PD
5. Post operative
6. Diseases - billiary obstruction, celiac disease and pancreatic insufficiency
7. Chronic liver disease.
1.4. Clinical features
Patients present with bleeding tendencies and the blood picture reveals
prolonged prothrombin time (PT) and reduced levels of Factor II, VII, IX and X.

Pyruva
2.0 VITAMIN K ANTAGONISM
te
Kinase
There is interference with vitamin K metabolism e.g. Warfarin overdose. The oral
anticoagulants (coumarin and phenindione derivatives) are competitive inhibitors
of the vitamin K-epoxide reductase enzyme complex decreasing the availability
of reduced vitamin K in the hepatocytes effectively inhibiting synthesis of the Kdependent factors.
3.0 LIVER DISEASE
3.1. Introduction
All the clotting factors synthesized in liver except FVIII hence liver disease leads
to disturbed (reduced) synthesis of clotting factors, production of structural
abnormal clotting factors, disturbed storage and cholestasis (a feature of liver
disease impairs absorption of vitamin K).
3.2. Causes
1. Anatomic defects lead to bleeding
a. Portal hypertension which presents with varices,
spleenomegally and secondary thrombocytopenia
b. Peptic ulceration
c. Gastritis
2. Hepatic function abnormalities
a. Decreased synthesis fibrinogen, prothrombin, factor V,
VII, IX, X, XI.
b. Decreased synthesis of coagulation inhibitors, protein C

VII, IX, X, XI.

b. Decreased synthesis of coagulation inhibitors, protein C
and S.
c. Impaired Vitamin K absorption and metabolism.
d. Failure to clear activated coagulation proteins- D.I.C
and fibrinolysis
3. Vitamin K deficiency - due to intrahepatic or extrahepatic cholestasis
4. Thrombocytopenia due to hyperspleenism associated with portal
hypertension
5. Functional abnormalities of platelets and fibrinogen
6. Iatrogenic

a. Massive transfusion- dilution platelets, coagulation proteins.

b. Infusion of activated coagulation protein.
c. Bleeding from Heparin effects.

3.3. Blood picture

Prolonged prothrombin time (PT) and mild thrombocytopenia
3.4. Clinical Picture
Bleeding tenderness
4.0 VITAMIN C DEFICIENCY
Vitamin C deficiency leads to defective collagen synthesis.
5.0 ALCOHOL
Direct toxin on the marrow
Inhibit platelet production and function e.g. in liver cirrhosis.
6.0 HAEMOLYTIC ANAEMIA SYNDROME
The damage is immunological and deposition of immune complexes and
activation of complement.
7.0 DISSEMINATED INTRAVASCULAR COAGULOPATHY (D.I.C).
7.1. Introduction
D.I.C is an acute, sub acute or chronic explosive life threatening
thrombohaemorrhagic disorder that occurs as a complication of a number of
diseases or disorders and results is severe bleeding episodes. It is characterized
by formation of microthrombi throughout the microcirculation resulting in
consumption of platelets, fibrin and coagulation factors. Secondarily, this leads
top activation of the fibrinolytic mechanisms.
D.I.C can present with signs and symptoms related to infarction (due to
microthrombi) and bleeding (due to depletion of elements necessary for

D.I.C can present with signs and symptoms related to infarction (due to
microthrombi) and bleeding (due to depletion of elements necessary for
haemostasis and activation of the fibrinolytic system).
7.2. Causes
D.I.C is commonly associated with: 1. Obstetric catastrophes/complications
a. Abruptio placentae
b. Retained dead foetus
c. Septic abortion
d. Amniotic fluid embolism
e. Toxaemia
2. Disseminated malignancy
a. Ca pancreas
b. Ca prostate
c. Ca lung
d. Ca stomach
e. Acute promyelocytic leukaemia
3. Massive tissue injury
a. Traumatic
b. Severe burns
c. Extensive surgery
4. Infections

5. Miscellaneous

a. Gram-negative sepsis
b. Meningococcemia
c. Malaria
d. aspergillosis
a. Shock
b. Liver disease.
c. Heat stroke
d. Acute intravascular haemolysis
e. Vasculitis

7.3. Pathogenesis & Pathophysiology

The pathogenesis of D.I.C revolves on two main mechanisms release of tissue
factor or thromboplastic substances into circulation and widespread injury to
endothelial cells.
There occurs activation of the clotting system resulting in deposition of fibroin in
the small vessels of many organs such as the kidneys, lungs, brain, adrenals and
placenta. Subsequently it causes tissue necrosis and organ dysfunction
(multiple). The bleeding tendencies in DIC result from consumption of clotting
factors and platelets and enhancement of the fibrinolytic system.
Diagram 2: Pathophysiology of DIC

Release of
Tissue factor

Activation of Microagiopathic
Vascular
Plasmin Haemolytic

occlusion
Anaemia

Platelet aggregation

Proteolysis of
Clotting factors
Fibrin split
products

Inhibition of thrombin,
platelet aggregation and
fibrin polymerization

7.4. Etiological factor and disorders that lead to D.I.C

A. Liberation of toxins.
i. Obstetric syndromes such as abortion (2nd trimester),
placenta abruption, amniotic fluid in embolism which lead to
retained dead foetus
ii. Haemolysis
iii. Fat embolism
iiii. Tissue damage due to burns, front bite, head injury and
gunshot wounds.
B. Endothelial damage as in acute glomerulonephritis ( AGN) and aortic
aneurysm
C. Vascular damage with decreased blood flow
D. Infections
i. Bacterial streptococci, staphylococci and gram negative
bacilli
ii. Viral Rubella, Varicella and Arbovirus
iii. Parasitic malaria and kala azar
iiii. Ricketsia - Rocky mountain fever
v. Mycotic

iiii. Ricketsia - Rocky mountain fever

v. Mycotic
7.5. Clinical features
Clinical features vary with the stage or severity of the syndrome.
Extensive skin and mucous membrane bleeding.
Haemorrhage from multiple sites surgical incision, vene-punctual,
catheter sites.
7.6. Laboratory

Thrombocytopenia
Presence of schisocytes and fragmental red blood cells
Prolonged prothrombin time (PT)
Reduced Fibrinogen levels.
Reduced coagulation proteins

Page 13 of 155
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

LEUKAEMIA
1.0 INTRODUCTION
Leukaemias are primary neoplastic (malignant) tumour disease of blood forming
(haemopoetic) cells affecting proliferation of precursors of the white blood cells.
Leuko means white, aemia means blood.
It is a clonal disorder of the bone marrow that results in excessive production
and proliferation of white blood cells. The proliferation of leukaemic cells occurs
in the bone marrow or certain lymphoid tissues resulting in their spill over into
the peripheral circulation raising the total white blood cell count.
The occupation of the bone marrow by leukaemic cells results in bone marrow
failure, which manifests with anaemia, thrombocytopenia and neutropenia and

The occupation of the bone marrow by leukaemic cells results in bone marrow
failure, which manifests with anaemia, thrombocytopenia and neutropenia and
involvement of other organs such as the liver, spleen, lymph nodes, meninges,
brain and the skin.
Diagram 1: Formation of Blood Cells

2.0 CLASSIFICATION
Leukaemias are classified on the basis of kinetic behaviour of the cells and the
cell types that are predominant in the disease picture. Both classifications are
used together.
1. Kinetic behaviour of the cells, which determines the rate progression or
natural history of the disease.
a. Acute leukaemia
Acute leukaemias have rapid progression and involve proliferation of
immature cells the primitive blast cells
b. Chronic leukaemia
Chronic leukaemias have a slow progression and involve proliferation of the
maturing cells.
2. Pattern of differentiation and the cell types predominantly involved

maturing cells.
2. Pattern of differentiation and the cell types predominantly involved
a. Myeloid leukaemia involves differentiation of the
myeloblats mainly the granulocytes and monocyte
series.
i. AML (Acute Myeloid Leukaemia)
ii. CML (Chronic Myeloblastic Leukaemia)
b. Lymphocytic leukaemia involves the lymphoblasts of
the B and T lymphocyte series
i. ALL (Acute Lymphocytic Leukaemia)
ii. CLL (Chronic Lymphoblastic Leukaemia)

3.0 AETIOLOGY
The aetiology is unknown but several factors have been implicated.
1. Genetic factors
Leukaemias tend to have increased frequency or incidence with a variety of
congenital disorders such as Downs, Blooms, Klinefelters, Fanconis and
Wiskott-Aldrich syndromes.
2. Environmental factors

a. Ionising radiation common in AML, ALL and CML

X-rays
Post Hiroshima bombing
Chernobyl bombing
b. Chemical carcinogens benzene and aromatic
hydrocarbons (AML)
c. Drugs chemotherapy
3. Infections viral infections e.g. The Human T cell Leukaemia virus 1 (HTLV1)
4.0 AGE INCIDENCE
Acute Leukaemia has two peaks of 5 -14 years and 55-75 years and the cell
patterns do vary with AML being predominant in adults and ALL during childhood.
Chronic leukaemia occurs in individuals over 40 years of age with CLL > CML
5.0 PATHOPHYSIOLOGY
There is malignant transformation of a single clone of cells belonging to the
myeloid or lymphoid series followed by proliferation of the transformed clone.
This results in the basic defect that affects the DNA conferring the heritable
malignant characteristics, which interfere with normal differentiation and
maturation of leucocyte precursors which proliferate uncontrollably at the
expenses of normal marrow proliferation. Cellular replication and expansion
follows malignant transformation of a haemopoetic or lymphoid progenitor. The
cells fail to mature beyond myeloblast stage (AML) and lymphoblast stage (ALL).
The leukaemic cells proliferate in the bone marrow and spill over into the
general circulation where they multiply and seed other organs such as the liver,
spleen, lymph nodes, skin, viscera and the central nervous system.

general circulation where they multiply and seed other organs such as the liver,
spleen, lymph nodes, skin, viscera and the central nervous system.
Leukaemic cells accumulate in the bone marrow and suppress normal
haemopoietic stem cells. In a leukaemic marrow there is an imbalance between
cell proliferation and cell destruction as there is excessive proliferation and
defective terminal differentiation.
6.0 PATHOLOGY
a. Bone marrow replacement
b. Bone marrow failure
c. Low erythropoeisis

ACUTE LEUKAEMIA
1.0 INTRODUCTION
Acute leukaemias are characterized by predominance of undifferentiated
leucocyte precursors or leukaemic blasts. They may be derived from the myeloid
stem cells acute myeloblastic leukaemia (AML) or from the lymphoid stem cells
acute lymphoblastic leukaemia (ALL).
The incidence in childhood is 80% ALL and adulthood 80% AML.
The cells are poorly differentiated and immature cells with the degree of
differentiation dependant on the type of leukaemia. Many of the cells are in the
resting (non-dividing) stage.
Therapeutic induction depends on destruction of leukaemia cells and not
correcting their behaviour

2.0 CLINICAL FEATURES

The clinical features are divided into two groups: 1. Those due to bone marrow failure (BMF)
2. Those due to organ infiltration
3.0 BONE MARROW FAILURE (BMF)
These are represented in the triad of anaemia, bleeding manifestations and
infections.
1. Anaemia

a. Pallor
b. Constitutional features/general features (sudden onset,
fever, weakness/fatigue)
c. Dyspnoea

2. Bleeding tendencies due to thrombocytopenia

a. Features purpura, spontaneous bruises, mucous
membrane
bleeding,
menorrhagia,
petechial
haemorrhage, ecchymoses, fundal haemorrhage and
prolonged haemorrhage after surgery.

haemorrhage, ecchymoses, fundal haemorrhage and

prolonged haemorrhage after surgery.
b. Sites skin, CNS. Lungs, viscera
3. Infections due to neutropenia
a. Common sites - Infections of the mouth, throat, skin,
respiratory system and perianal.
b. Common organisms Gram-negative microorganisms, E.
coli, Pseudomonas spp, Proteus, Klebsiella and
Candida.
4.0 ORGAN INFILTRATION
Organ infiltration occurs due to replacement of the bone marrow and other
cell disease
tissuesSickle
with leukaemic
cells manifests
hence disrupting their functions.
with a spectrum of signs and
symptoms
such as
1. Pain
and tenderness
overchronic
bones e.g. sternal tenderness due to bone infarcts
haemolytic
anaemia,
painful
or subperiosteal infiltrates in children with ALL
sickling crises,
leg ulceration, common in ALL
2. Generalized
lymphadenopathy
recurrent
respiratory
tract
3. Hepatomegally
infections,
myocardial
4. Heamaturia and blockage of ureters
insufficiency,
5. Gum
hypertrophyjaundice and
crises
affect
the abdomen,
6. Chloromas
localized
tumour-forming masses in the skin or orbit due to
bones
and
joints
resulting
infiltration by tumour cells
from blockage
of small (presents
blood
7. Meningeal
involvement
with increased intracranial pressure,
vessels
by
sickled
cells
and
headache, nausea/vomiting,
blurring of vision, diplopia)
infarcts from the
spleen(CVA)
and due to intracranial haemorrhage.
8. Cerebro-vascular
accident
bones.
9. Others
a. Testicular swelling in ALL
b. Mediastinal compression in T cell ALL

5.0 CLASSIFICATION
5.1. Acute Myeloid Leukaemia (AML)
AML is classified on the basis of the morphological appearance of the bone
marrow.
Table 1: FAB (French-American-British) classification of AML

Type
Differentiation
M0 Undifferentiated cells
M1

Undifferentiated myeloid

Description of Cells
Cells are poorly differentiated that
their myeloid nature is not apparent
Very primitive myeloid cells

M2
M3

Myeloid
with
differentiation
Promyelocytic

Further myeloid differentiation

M4

Myelomonocytic

M5

Monocytic

Both granulocytic and

differentiation
Large primitive monocytes

M6

Erythroleukaemia

Many red cell precursors

M7

Megakaryoblastic

Primitive megakaryocytes.

granulocytic Myeloblasts with granules

monocytic

5.2. Acute Lymphocytic Leukaemia (ALL)

ALL is predominantly a disease of children.
Table 2: FAB classification of ALL
Type
Occurrence
L1 Childhood-ALL (B-ALL and T-ALL)

L2

Adult ALL (mostly T-cells)

L3

Burkitt type-ALL (uncommon

affects both children and adults)

Description of Cells
Homogenous small lymphoblasts
Cells have a scanty cytoplasm,
small regular nuclei with a small
inconspicuous nucleoli
Heterogeneous lymphoblasts
Variable amounts of cytoplasm
Irregular nuclei
Large nucleoli
Large homogenous lymphoblasts
Oval/round nucleus with prominent
nucleoli
Cytoplasmic vacuolation

6.0 DIAGNOSIS
1. History
2. Physical examination
3. Laboratory findings
6.1. History
6.2. Physical Examination
6.3. Laboratory Features
1. Blood picture

a. Anaemia
b. High leucocyte count (WBC) more than 100 x 10 9/L
c. Cells are large, primitive, have nucleoli
d. High nuclear: cytoplasmic ratio
e. Cells show features of immaturity and poor
differentiation
f. Thrombocytopenia

2. Bone marrow do bone marrow aspirate or trephine biopsy

a. High cellularity
b. Leukaemic cells
c. Reduced erythropeiotic cells
d. Reduced megakaryocytes
e. Abnormall chromosomes of diving leukaemic cells
(chromosome 9 and 22)
3. Cytochemistry use of staining procedures to check for enzymes present
e.g. acid phosphotase in ALL.
4. Others

e.g. acid phosphotase in ALL.

4. Others
a. Cell surface markers used in classification in ALL
b. Serum uric acids increases due to rapidly growing number of
leukaemic cells.
Table 3: Differences and Similarities between AML and ALL
Feature
Common age

AML
ALL
Adults between 15 40

years

20% of childhood
leukaemias
Physical findings
Spleenomegally

Hepatomegally

Lymphadenopathy

Bony tenderness

Gum hypertrophy

Laboratory
Myeloblats and

promyeloblasts
dominate in blood and
bone marrow

Moderate to severe
thrombocytopenia
Response
to
Remission rate low

therapy
Short period of remission

Median survival

12 18 months

Children under 15 years

80% of childhood
leukaemias
Marked spleenomegally
Marked hepatomegally
Marked
lymphadenopathy
Bony tenderness
Meningeal involvement
Lymphoblats dominate
in the blood and bone
marrow
Moderate to severe
thrombocytopenia

Remission rate high

Prolonged period of
remission
Children without CNS
prophylaxis 33 months
With CNS prophylaxis
60 months
Adults 12 18 months

CHRONIC LEUKAEMIAS
1.0 INTRODUCTION
Chronic leukaemias are haematologic malignancies with well-defined leukaemic
cells (the cell types can easily be recognized).
It is divided into two main types: a. Chronic myeloid (granulocytic) leukaemia CLL/CGL
b. Chronic lymphocytic leukaemia CLL
2.0 CHRONIC LYMPHOCYTIC LEUKAEMIA (CLL)
2.1. Introduction
CLL, which accounts for 25% of all leukaemias, is predominantly an incurable
disease of the elderly (over 50 years) and affects more males than females in a
2:1 ration. It also affects middle-aged individuals. CLL mainly affects B cells but
a small percentage of T cells may be affected.

2:1 ration. It also affects middle-aged individuals. CLL mainly affects B cells but
a small percentage of T cells may be affected.
CLL is characterized by accumulation in the blood and bone marrow of small
lymphocytes that have failed to mature normally hence they are immunologically
incompetent.
2.2. Clinical Features
The disease has an insidious onset.
1. Features of anaemia
2. Generalized lymphadenopathy the lymph nodes are soft, rubbery,
homogenous, non-tender and have a symmetrical involvement. The nodal
architecture is lost.
3. Spleenomegally
4. Hepatomegally
5. Haemorrhagic manifestations due to thrombocytopenia
6. Recurrent infections especially the respiratory system
bronchopneumonia
7. Less common features
a. Mediastinal pressure
b. Pressure effects on the bladder, oedema
c. Disturbed vision
d. Bone and joint pains
e. Tonsilar enlargement

2.3. Diagnosis
Diagnosis is based on the minimum diagnostic criteria of a persistent circulating
lymphocyte count of > 5 X 109/L and bone marrow lymphocytosis of > 30%.
2.4. Investigations/Laboratory
1. Blood picture

2. Bone marrow

3. Others

a. Anaemia mild to moderate normocytic normochromic

anaemia with reticulocytosis.
b. White blood cells increased > 15 X 109/L and
granulocytopenia in advanced disease.
c. Platelets normal or moderately reduced
a. Increased lymphocyte count
b. Reduced myeloid precursors
c. Reduced erythroid precursors
d. Fatty marrow replaced by leukaemic cells
a. Low serum immunoglobulins IgG, IgM and IgA.
b. Positive Coombs test if haemolysis is occurring.

2.5. Classification/Staging
This is based on physical examination findings and the blood picture. It is used to
assess the extent of the disease and indicate the likely prognosis.

This is based on physical examination findings and the blood picture. It is used to
assess the extent of the disease and indicate the likely prognosis.
Rai Staging
Stage 0 Lymphocytosis only (in blood and bone marrow)
Stage I
Lymphocytosis with lymphadenopathy
Stage II
Lymphocytosis with hepatic and/or spleenic enlargement
Stage III
Lymphocytosis with anaemia (Hb < 11 gm%)
Which microorgansism
Stage IV
lymphocytosis
with thrombocytopenia (platelet count of < 100 X
9
will
cause
problems
in a
10 /L)
sickler?
Why?
Explain
Note: Lymphocytosis white cell count > 15 X 109/L of which > 40% are
with clinical illustrations.
lymphocytes

Binet Staging
Stage A < 3 lymphoid areas involved
Stage B >
3 lymphoid areas
Stage C A
ny number of involved
lymphoid areas

Hb > 10 gdL-1
platelets < 100 X 109/L
Hb < 10 gdL-1
Platelets < 100 X 109/

Note: Lymph node areas are (1) cervical (2) axillary (3) inguinal lymph nodes (4)
spleen and (5) liver. The number of sites range from 0 5.
Revised Classification integrated and Rai System
Clinical stage

Good prognosis Hb> 10 g/dl (No anaemia)

Platelets >100x109/L (No
thrombocytopenia)
< 3 sites /areas of lymphoid
enlarge

Intermediate prognosisNo anaemia

No thrombocytopenia <100 x 109/L
>3 sites of lymphoid tissue
involved

Clinical stage B

Clinical stage C

Poor prognosis

Anaemia and /or hrombocytopenia

Regardless of no of site
lymphoid enlargement
Platelet count of < 100x109/L

of

2.6. Indications of treatment

1. Increasing or troublesome lymphadenopathy
2. Systemic symptoms
3. Bone marrow failure
4. Autoimmune complications e.g. haemolytic anaemia and immune
thrombocytopenia.

3.0 CHRONIC MYELOID LEUKAEMIA (CML, CGL)

3.1. Introduction
Chronic myeloid (myelogenous, granulocytes) leukaemia accounts for 20% of all
leukaemias. It affects both sexes equally and has the peak incidence in the 3rd
and 4th decades of life. It is characterized by the uncontrolled proliferation of
myeloid cells as a result of mutations in a single pluripotential haemopoietic
stem cell usually on chromosome 9(Philadelphia chromosome).
It develops in three phases namely: a. Chronic phase a 2 6 year period of slow evolution
b. Accelerated phase which leads to an increase in immature cells
c. Blast phase when there is transformation to an acute leukaemia especially
the myeloblasts and rarely lymphoblasts.
3.2. Clinical Features
1. Features of anaemia
2. Symptoms due to hyper-metabolism
a. Weight loss
b. Lassitude
c. Night sweats
d. Anorexia
3. Massive spleenomegally (leads to abdominal discomfort, some infarcts may
form resulting in abdominal pain)
4. Marked hepatomegally
5. Bleeding tendencies
6. Less common

a. Gout due to high uric acid levels form breakdown of

nucleic acid in leukaemia cells.
b. Visual disturbance
c. Neurological manifestations
d. Priapism

3.3. Investigations/Laboratory
1. Blood

a. Anaemia normocytic normochromic

b. Increased white blood cells
c. Platelets low or normal or increased

2. Bone marrow bone marrow aspiration

a. Hypercellularity
b. Replacement of fat spaces by proliferating myeloid
cells
c. Myeloid cells predominate
d. Reduced erythropoiesis
e. Megakaryosites are small in size.
3. Others

a. Increased serum uric acid/hyperuriceamia

b. Low alkaline phosphotase in the neutrophil leukaemic
cells
c. Increased B12 due to increased B12 binding protein.

4.0 OTHER MYELOPROLIFERATIVE DISORDERS

Myeloproliferative disorders are a group of neoplastic proliferation of
multipotent haematopoietic stem cells.
4.1. Polycythaemia Vera
4.1.1. Introduction
Polycythaemia Vera (PV) is characterized by increased production of all myeloid
elements resulting in pancytosis, elevated haemoglobin concentration and
spleenomegally.
Primary Polycythaemia
Polycythaemia rubra vera or polycythaemia vera denotes primary or idiopathic
polycythaemia only.
Secondary Polycythaemia (Erythrocytosis)
Causes
1. High altitude
2. Cardiovascular disease
3. Pulmonary disease with alveolar hypoventilation
4. Heavy smoking
5. Inappropriate increase in erythropoeitin
a. Renal cell carcinoma
b. Hydronephrosis
c. Hepatocellular carcinoma
d. Haemangioblastoma
e. Massive uterine leiomyoma

d. Haemangioblastoma
e. Massive uterine leiomyoma
6. Relative or spurious polycythaemia may result from plasma loss e.g. in
burns and dehydration (vomiting and water deprivation)
Secondary polycythaemia is not associated with spleenic enlargement or
increased leucocytes unlike polycythaemia vera.
4.1.2. Clinical Features
Polycythaemia vera is a disease of late middle age and is slightly more common
in males. It runs a chronic but slow progressive course. Clinical features are due
to the effects of hyperviscosisty, hypervolaemia, hypermetabolism and decreased
cerebral perfusion.
Headache, vertigo, tinnitus, visual alterations, syncope or coma
Risk of thrombosis due to accelerated atherosclerosis
Increased risk of haemorrhage due to increased plasma volume and
intrinsic platelet dysfunction
Spleenomegally
Pruritus especially after a bath
Increased risk of urate stones and gout due to hyperuricaemia
4.1.3. Laboratory Findings
1. Raised haemoglobin concentration above 17.5g/dL in males and 15.5 g/dL
in females
2. Erythrocytosis above 6 million/ l in females and 5.5 million/ l in males
3. Heamatocrit (PCV) above 55% in males and 47% in females
4. Mild to moderate leucocytosis
5. Thrombocytosis with defective platelet function
6. bone marrow erythroid hyperplasia or panhyperplasia

Page 13 of 155
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

THE LYMPHO-RETICULAR SYSTEM

LEARNING OUTCOMES
At the end of the lesson the learner should be competent to: 1. Describe the causes, pathology, clinical features and investigations in
disorders of the lymph nodes.
2. Describe the causes, pathology, clinical features and investigations in
disorders of the spleen.
3. Describe the causes, pathology, clinical features and investigations in

2. Describe the causes, pathology, clinical

disorders of the spleen.
3. Describe the causes, pathology, clinical
lymphomas
4. Describe the causes, pathology, clinical
TalkingBurkitts
Pointlymphoma
5. Describe the causes, pathology, clinical
1. Describe
all the possible
multiple
myeloma

features and investigations in

features and investigations in
features and investigations in
features and investigations in

patterns
of
SCD
inheritance THE
overLYMPHOID
four SYSTEM
generations of a family.
2.
Explain
the
1.0 INTRODUCTION
pathophysiology of the
cardinal
features
of lymphoid organs (lymph nodes,
The lymphoid
tissue consists
of peripheral
haemolytic
anaemia.
spleen, MALT,
NALT, GALT
and SALT) and central lymphoid organs thymus
In a table format explain
and bone3.marrow.
the pathophysiology of the
The lymphoid
organs are principally
complications
of involved
SCD in: stating
their
clinical
a) Production and maturation of lymphocytes (thymus T
features.
lymphocytes; bone marrow B lymphocytes
4. Compare and contrast the
b) Antigen presentation and immune response
laboratory
of
c) Filtrationfeatures
and phagocytosis
of microorganisms and particulate
inherited
haemolytic
material.
anaemias.
5. Describe the process of
2.0 LYMPHOID
Hb TISSUES
formation.
6. Describe the process of
oxygen carriage by Hb
7.
How
would
you
LYMPH NODES
What areinvestigate
the lymphoidand
tissues?
Draw
manage
their structures
and state their
SCD crisis?
1.0 INTRODUCTION
functions.
8. Outline complications of
SCD
occur asoraoval
result
Lymph nodes
arethat
bean-shaped
structures 1-2 cm long. The basic structure
comprises has
of infarctive
the medullaprocesses.
and cortex (central cortex and paracortex) covered by
a connective
tissue capsule.
A lymphand
node is served by an afferent lymphatic
9. Describe
the causes
that drainsprocess
into the of
lymph
node and an efferent lymphatic one that drains out of
haemolysis.
the node.10. What investigations will
be relevant in a patient
with SCD? How would you
2.0 STRUCTURE
make the diagnosis of
The cortex
SCD?
has several lymphoid follicles and is responsible for T cell
(paracortex) and B cell (central cortex) activities while the medulla, which has
sinuses and phagocytic cells, deals with filtration and presentation of antigens by
the dendrtitic antigen presenting cells (APCs).

3.0 FUNCTION
1. To mount an immune response in the body
2. Active phagocytosis of particulate material
Lymph nodes are usually secondarily involved in a number of systemic diseases,
local injuries and infections and they also form the site for some primary
neoplasms. Lymph node enlargement may be localized or generalized with the
enlargement resulting from either reactive hyperplasia or neoplasm. Disorders of
lymph nodes are reactive lymphadenitis, malignant lymphomas, lymph node
metastatic tumours and plasma cell disorders.

4.0 REACTIVE LYMPHADENITIS

Lymph nodes undergo reactive changes in response to stimulation by microbial
infections, drugs, environmental pollutants, tissue injury, immune complexes and
neoplasms. Lymph node enlargement due to primary inflammatory reactions
would be preferably called reactive lymphadenitis whereas that due to immune
reactions is termed lymphadenopathy, however the term lymphadenopathy is
used generally to refer to all cases of enlarged lymph nodes. The lymph nodes
frequently affected are the cervical, axillary, inguinal and mesenteric groups.
Acute lymphadenitis is usually mild and transient with the lymph nodes being
acutely inflamed, enlarged, tender, fluctuant (if extensively involved) with the
overlying skin being red and hot.
5.0 LYMPHADENOPATHY

Outline
causes
lymphadenopathy

of

6.0 SPLEENOMEGALY
Structure
The spleen consists of the connective tissue framework, vascular channels,
lymphatic tissue, lymph drain channels and cellular components of haemopeitic
and reticuloendothelial systems.
Histologically the spleen has two main components

Histologically the spleen has two main components

The white pulp the unit of lymphoid tissues (lymphoid follicles) called the
Malphigian bodies.
The red pulp the vascular network of sinuses and pulp cords that forms
the major part of the organ.
Blood supply is through the trabecular arteries that arise form the spleenic
artery and drains via the corresponding veins. Blood flows through the spleen in
two systems the open and closed systems.

Figure 1: Blood flow in the Spleen

Functions
1. Haemopoeisis during foetal life (12th week to birth), severe
haematological stress (thalassaemia, chronic haemolytic anaemias) and
granulopoeisis.
2. Humoral control of haemopoeisis the spleen elaborates humoral factors
which control haemopoeisis (suppress or stimulate)
3. Cell sequestration, phagocytosis and pooling
a. Cleans and reconditions cells for recirculation

which control haemopoeisis (suppress or stimulate)

3. Cell sequestration, phagocytosis and pooling
a. Cleans and reconditions cells for recirculation
b. Removes from the circulation damaged or effete cells and
foreign matter
Sequestration cells are held in spleen temporarily before returning into the
circulation
Phagocytosis irreversible removal by macrophages
Pooling increased amount of blood that is in continuous exchange with the
circulation
4. Immunological function the spleen is the largest lymphoid tissue
containing 25% of T cells and 10 15% of B cells in the body. It is also an
antibody production site
5. Blood reservoir
6. Plasma volume control has neurohumoral mechanisms, which controls the
fluid equilibrium between interstitial and intracellular compartments.
Spleenomegally
The spleen should enlarge 1.5 2 times to be palpable but a normal spleen is
palpable if displaced downwards e.g. by an effusion.
Clinical detection of spleenomegally
Moves downwards and medially on inspiration and enlarges in similar
manner
Has a spleenic notch on the medial margin
Does not push through from the right loin (compare with right kidney)
Difficult to get the upper boarder
Dull to percussion
Imaging procedures radionuclide scans, ultrasound scans, CT scan and MRI
Differential diagnosis of a mass in the right upper quadrant
1. Spleen
2. Right kidney
3. Tumours spleenic flexure and colon
4. Masses arising from the stomach
5. Retroperitoneal masses
Causes of Spleenomegally
The degree of spleenomegally varies with the disease entity: Mild enlargement (up to 5 cms) occurs in chronic heart failure, acute
malaria, typhoid fever, bacterial endocarditis, SLE, rheumatoid arthritis
and thalassaemia minor.

Moderate enlargement (up to umbilicus) occurs in hepatitis, cirrhosis,

lymphomas, infectious mononucleosis, haemolytic anaemia, spleenic
abscesses and amyloidosis.

Massive enlargement (below umbilicus) occurs in CML, TSS. Leishmaniasis,

Gauchers disease and myefibrosis.

 Massive enlargement (below umbilicus) occurs in CML, TSS. Leishmaniasis,

Gauchers disease and myefibrosis.
1. Infections

a. Malaria
b. Acute bacterial sepsis
c. Bacterial endocarditis
d. Viral EBV, Coxsackie, HIV, Hepatitis
e. Chronic bacterial TB, syphilis, typhoid, brucellosis
f. Chronic parasitic
i. TSS (massive)
ii. Hydatid
iii. Kala azar (massive)
iiii. Schistosomiasis
v. Trypanosomiasis

2. Circulatory Disturbances
a. Portal hypertension
b. Biliary cirrhosis
c. Portal vein obstruction
d. Cardiac failure
e. Budd Chiari syndrome
3. Diseases of the blood
a. Thalassaemia
b. Hereditary spherocytosis
c. Sickle cell disease
d. Red cell defects
e. Acute leukaemia
f. CML (massive spleenomegaly) and CLL
g. Polycythaemia vera
h. Megaloblastic anaemia
4. Neoplasms

a. Hodgkins lymphoma and Non-Hodgkins lymphoma

b. Multiple myeloma
c. Myeloproliferative disorders (CML, myelofibrosis)
d. Metastatic tumours

5. Storage Diseases and Degenerations

a. Gauchers disease (massive)
b. Amyloidosis
c. Idiopathic
Explain
the
pathophysiology
of
6. Disorders of immunoregulation/connective tissue
disorders
these
clinical
a. S.LE. and Rheumatoid arthritis
features.
b. Feltys syndrome
Page 13 of 155
HAEMATOLOGY AND LYMPHORETICULAR SYSTEM

MALIGNANT LYMPHOMAS
1.0 INTRODUCTION
Lymphomas are malignant tumours of the lymphoreticular system originating the

Lymphomas are malignant tumours of the lymphoreticular system originating the

lymphocytes and histiocytes and arising from the lymph nodes or tissues rich in
lymphoid tissues e.g. the spleen, bone marrow, pharynx and the G.I.T. it is
usually a disease of the T-lymphocytes but the B-lymphocytes are affected in
some instances.
2.0 CLASSIFICATION
1. Hodgkins Lymphoma /Hodgkin Disease (after Thomas Hodgkin An English
Physician 1798 1866)
2. Non-Hodgkin Lymphoma (NHL)
3.0 HODGKINS LYMPHOMA/DISEASE
3.1. Introduction
Lymphoid cells are widely distributed in lymph nodes and a number of organs
excluding the central nervous system. There are three major cell types namely
the natural killer (NK) cells, B cells and T cells that are capable of undergoing
expansion, maturation and selection on exposure to an appropriate antigenic
stimulation. Hodgkins disease arises from lymph nodes and involves the extranodal sites secondarily. It occurs at any age but there are two peaks; one in early
adult life (15-35 years) and the other in old age after the 5th decade of life. The
average incidence is 30 cases/million and affects the more males than females in
a ratio of 2:1.

3.2. Aetiology
The aetiology is unknown but associated factors are: Infections EBV, HLTV-1, Herpes and HIV
Genetic HLA related

3.3. Clinical Presentation

1. Progressive and painless lymph node enlargement starting with the cervical
lymph nodes and spreads to involve the axillary, abdominal, mediastinal
and pharyngeal.
2. Spleenomegally
3. Hepatomegally
4. Recurrent infections due to reduced cell mediated immunity
5. Constitutional features fever, weight loss, pruritus/itchness, night sweats,
malaise and fatigue.
6. Anaemia
7. Involvement of other organs skin, G.I.T, brain, retina.

3.4. Spread

3.4. Spread
1. Spreads from the cervical lymph nodes to other lymphoid tissues along the
central axis and the spleen.
2. Spreads to other organs bone marrow, liver, alimentary tract.

3.5. Pathology
Macroscopy
1. Lymph nodes
Initially they are enlarged, discrete, soft and rubbery but later they become
firm producing pressure effects on the trachea and mediastinum.
2. Enlarged spleen
3. Others involvement of the kidney, liver, lungs, bone marrow, vertebrae
and G.I.T.
Microscopy
The diagnosis is made by histological examination of lymph node biopsy. The
lymph node tissue may be partly or completely replaced by tumour cells with the
typical cells characteristic of HD being the REED STEINBERG (RS) cells.

3.6. Diagnosis
1. Physical examination
2. Investigations
a. Laboratory

i. Full haemogram normocytic

normochromic anaemia, increased
lymphocytes and raised ESR
ii. Liver biochemistry
iii. Urea and electrolytes
iiii. Uric acid levels raised

b. Chest X-ray
c. CT scan of the pelvis and abdomen
d. Bone marrow aspirate or trephin biopsy
e. Lymph node biopsy for histology (RS cells)

3.7. Classification/Staging
This is done by histological examination and it is for prognosis purposes.
1. Rye Classification
Type

Features

1. Rye Classification
Type
1.

Features

Lymphocyte predominant
(10%)
lymphocytes

Lymphocytes are predominant; Few RS cells

seen,
nodular
and
diffuse
patterns,
and macrophages

2. Nodular sclerosing (30-60%)

Lymph node capsule is thickened, thick bands

of collagen divide the node into nodules, RS
cells are seen, and cellular infiltration is mixed
lymphocyte depleted or dominant.

3. Mixed Cellularity (20-30%)

Numerous RS cells seen, cellular infiltration

by neutrophils, oesinophils, macrophages,
plasma cells and lymphocytes.

4. Lymphocyte depleted (< 2%)

Dominated by RS cells, large pleomorphic

neoplastic cells, sparse lymphocytes and
disordered connective tissue.

2. Ann Arbor Classification

Stage Features
I

I nvolvement of a single lymph node

single
extralymphatic organ or site (IE)

region

(I)

or

II

involvement of two or more lymph node regions on the same side of the
diaphragm (II) or one or more lymph node regions plus an
extralymphatic site (IIE)

III

Involvement of lymph node regions on both sides of the diaphragm (III)(the

spleen is
included in stage III e.g. cervical lymphadenopathy +
spleenomegally = stage III)

IV

Involvement of one or more extralymphatic organs e.g. lung, bone, bone

marrow with or without lymph node involvement.

Note:
All stages are subclassified as A (asymptomatic) of B (fever, night sweats
and loss of > 10% of body weight)
x denotes bulky disease (lymph node mass > 10 cm in diameter or
involves the mediastinum- a mass > 1/3 of the intrathoracic diameter at
T10)
4.0 NON-HODGKINS LYMPHOMA (NHL)
4.1. Introduction

4.0 NON-HODGKINS LYMPHOMA (NHL)

4.1. Introduction
NHL encompasses all malignancies originating from the lymphoid system with the
exception of Hodgkins disease. NHL is a malignant neoplasms of the immune
system and are more common than HD. They are frequent in young adults (20-40
years) but can occur at any age. Majority arise in lymph nodes (65%) and 35% in
extra-lymph node sites. They have the potential to spread to other lymph nodes,
spleen, liver, bone marrow and spill over into the peripheral blood. It involves
mainly B-cells (90%) and T-cells (10%).

4.2. Aetiology
Unknown but associations exist.
1. Viral infections EBV, HLTV-1
2. Genetic association chromosomal translocations
3. Immunodeficiency - HIV
4. Immunosuppression chemotherapy, radiation
5. Autoimmune disorders SLE

4.3. Pathogenesis
Genetic alterations in the tumour clone and biological alterations in the
host
Accumulation of gene alterations in the tumour genome
Infection of the tumour clone by oncogenic virus by e.g. EBV and HIV
Stimulation and selection of tumour cells by an antigen
Immunodeficiency of the host

4.4. Clinical Features

1. Superficial lymphadenopathy that is widespread
2. Constitutional features fever, night sweats, weight loss (.10%)
3. Oropharyngeal involvement involvement of the Waldeyers ring
4. Abdominal disease enlargement of liver, spleen and mesenteric and
retroperitoneal lymph nodes.
5. Other organs skin, brain

4.5. Investigations
1. Blood cell count (haemogram)
2. Urea/electrolytes
3. Liver biochemistry
4. Chest X-ray
5. CT scan abdomen/pelvis
6. Bone marrow aspirate/trephin biopsy
7. Lymph node biopsy
8. Raised uric acid levels
9. Hypercalcaemia
10. Lymphagiography
4.6. Staging
R.E.A.L (Revised European American Lymphoma) Classification - 1994

4.6. Staging
R.E.A.L (Revised European American Lymphoma) Classification - 1994
1. Leukaemias and Lymphomas of B-cell origin
a. Indolent B-cell malignancies
b. Aggressive B-cell malignancies
2. Leukaemias and Lymphomas of T-cell origin
a. Indolent T-cell malignancies
b. Aggressive T-cell malignancies

Table 1: Differences between Hodgkins Disease & Non-Hodgkins Lymphoma

Feature
Hodgkins
Non-Hodgkins
Cell derivation
T or B cell
90% B and 10% T
Nodal involvement
Localized may speared Generalized
Extranodal spread
Uncommon
Common
Bone marrow involvement Uncommon
Common
Constitutional symptoms Common
Common
Spill over
Never
May spread to blood
Prognosis
Better
Bad

Diagram 1: Complications of Lymphomas

Figure 1: Complications of Lymphomas

Physical bulk

Bone Marrow
Failure

Spleenic Infiltration

Immunological Disturbances

Hyperspleenism
Autoimmune
Haemolysis

5.0 BURKITTS LYMPHOMA

5.1. Introduction
Burkitts lymphoma is a high-grade malignant neoplasm of the lymphoreticular
tissues. It is a tumour of B-lymphocytes commonly affecting children. It
frequently involves facial bones and periorbital region. There are three varieties
African (endemic) Burkitts lymphoma, Sporadic (non-endemic) and a subset of
aggressive lymphomas occurring in patients infected with HIV.
5.2. Incidence and Distribution
The geographical distribution corresponds to that of Plasmodium falciparum
malaria where reduced cellular immunity is reduced hence plays a role in tumour
development. It is associated with EBV. Burkitts lymphoma is rare below 2 years
of age and affects male and females in the ratio of male:female being 2:1.

5.3. Pathology
The common clinical presentation is: 1. Facial swelling jaws and periorbital tumour
2. Abdominal swelling ovaries and retroperitoneal masses
3. The most common sites of tumour deposit are maxilla and mandible of the
skeletal system. The other sites are: a. Lymphoreticular tissue cervical, mediastinal,
abdominal lymph nodes and the spleen.
b. Cardiovascular system deposits in the pericardium and
is associated with mediastinal involvement.
c. Respiratory system is rarely involved
d. Central nervous system is involved in 30% of the cases
affecting the brain and meninges more than the spinal
cord.
e. Alimentary system salivary glands, palate, tonsilar
deposits, stomach, small intestines and appendix.
f. Endocrine glands direct extension to the pituitary
gland and deposits to the thymus and adrenals.
g. Genito-urinary tract ovaries are frequently involved;
kidneys have hydronephrosis and renal impairment
h. Other organs subcutaneous tissues may be affected as
a late manifestation of the disease.

5.4. Clinical Presentation

1. Most common jaw swelling seen in 75% of the cases. More often the
maxilla is affected first then the mandible. The tumour of the maxilla
spreads to the orbit and leads to exophthalmia. Facial tumours of the
mandible and maxilla are almost always painless and initially they present
as swellings of the gum and loose teeth, which may cause the patient to
visit a dentist.

mandible and maxilla are almost always painless and initially they present
as swellings of the gum and loose teeth, which may cause the patient to
visit a dentist.
2. Second common presentation is abdominal swelling in 60% of the patients.
It affects the kidney, ovaries, mesenteric lymph nodes and retroperitoneal
lymph nodes. Frequently you may have ascites.
3. CNS presentation is seen in 30% of patients paraplegia, multiple cranial
nerve palsies and CSF pleocytosis.
4. Others involvement of the thyroid, testis, breasts, peripheral nerves,
bones and rarely anaemia.

5.5. Diagnosis
1. History and physical examination
Burkitts lymphoma is a rapidly growing tumour within 2 3 weeks
Rapidly growing abdominal swelling within six months in 2 20 year
olds
2. Histology
Take tumour biopsy and tumour aspirate histological examination
reveals undifferentiated lymphoid cells with many mitotic figures
with many macrophages giving the STARRY SKY appearance.

5.6. Differential Diagnosis

1. Jaw swelling

2. Orbital swelling

a. Dentiginous cyst
b. Fibroma
c. Ameblastoma
a. Retinoblastoma
b. Chloroma

3. Abdominal swelling
a. Nephroblastoma
b. Neuroblastoma

5.7. Staging
A
B
C
D
AR

Single extra-abdominal mass

Multiple extra-abdominal masses
Abdominal mass with or without facial tumour
Abdominal mass with site of tumour other facial or marrow involvement
Abdominal mass with > 90% of tumour ressected

5.8. Complications
1. Pressure symptoms
a. Intestinal obstruction due to large abdominal tumour
b. Kidney renal failure
c. Respiratory obstruction jaw/thyroid

a. Intestinal obstruction due to large abdominal tumour

b. Kidney renal failure
c. Respiratory obstruction jaw/thyroid
d. Extramedullary spinal cord compression paraplegia
2. Metabolic disturbances
a. Hyperuricaemia especially after treatment has started
b. Hypoglycaemia
c. Metabolic acidosis
6.0 AIDS RELATED LYMPHOMAS
NHL is the most lethal complication of HIV disease with most of the cells being of
B cell origin. This is usually due to: Inadequate immunologic control of persistent viruses with a capacity to
transform.
Alteration in regulatory environment for lymphoid growth control.
Accumulation of genetic abnormalities in target cells.
The definite AIDS associated malignancies Kaposis sarcoma, Non-Hodgkins
lymphoma and squamous cell carcinoma of the cervix, anus and conjunctiva
while the probable ones being leiomyosarcoma and Hodgkins disease.

MULTIPLE MYELOMA
1.0 INTRODUCTION
Multiple myeloma is a neoplastic disease of the plasma cells and its precursors
and widespread deposits of the tumour cells in the bone marrow characterize it
but proliferation of the tumour cells is multifocal.
Multiple myeloma primarily affects the elderly (peak incidence 5th-6th decades)
and increases in incidence with age. Multiple myeloma is rare under the age of
40 affecting more males than females.

2.0 AETIOLOGY
Myeloma is a monoclonal proliferation of B-cells. The aetiology is unknown but it
is thought that its induction requires prolonged exposure to foreign antigens in
predisposed individuals. It is associated with antigenic stimulation, genetic
predisposition and translocation of chromosomes.

3.0 PATHOLOGY
Talking
Point affects principally the bone marrow but in the course of the
Multiple
myeloma
disease other organs are involved through invasion by the tumour and its
1. What
the mechanisms of
products
(the are
M component).
haemolysis in acquired haemolytic
anaemiatissue is in the form of reddish nodules throughout the bone
The neoplastic
2.
What
the
clinicalare
features
in these
marrow. These
nodules
osteoclytic
affecting bone absorption and rarefaction
situations
resulting in spontaneous fractures. The affected bone shows no osteoblastic
3. What
are theosteoclats,
relevant which respond to osteoclast stimulating factor
activity.
Activated
investigations?
Explain
the the bone tissue. Widespread proliferation of
produced by myeloma cells
destroys

resulting in spontaneous fractures. The affected bone shows no osteoblastic

3. What
are theosteoclats,
relevant which respond to osteoclast stimulating factor
activity.
Activated
investigations?
Explain
the the bone tissue. Widespread proliferation of
produced by myeloma cells
destroys
parameters
expected.
plasma cells in the bone marrow leads to diffuse osteoporosis. Increased bone
lysis 4.
leads to increased mobilization of calcium from the bone resulting in
hypercalcaemia. Localized bone mass lesions may be palpable over the skull,
clavicles and sternum. The vertebral column may be affected collapsing causing
cord compression symptoms.
3.1. Osseous (Bone marrow) lesions
The disease involves multiple bones mainly those with red marrow skull, spine,
ribs and pelvis but later the long bones of the limbs become affected. On X-ray
(radiologically) the lesions appear as punched out, rounded 1-2cm sized defects
in the affected bone.
3.2. Extraosseous lesions
1. Blood atypical plasma cells, anaemia (nornocytic normochromic), marked
red cell rouleaux formation due to hypervisocity and an elevated ESR.
2. Myeloma kidney there is myeloma nephrosis due to infiltration of light
chains (Bence Jones proteins) which are precipitated in the distal
convoluted tubules.
3. Myeloma neuropathy infiltration of the nerve trunk roots by tumour cells
4. Systemic amyloidosis
5. Liver and spleen involvement.

4.0 PLASMA PROTEINS

Multiple myeloma cells synthesize and secrete immunoglobulins and therefore Ig
(mainly IgA, IgM and IgG) levels are raised. Mainly there is an increase in the
monoclonal Ig but a reduction in the normal Ig. The immunoglobulins, which are
mainly made of light chains, join to form monomers/dimmers seen urine as
Bence Jones proteins. The Paraprotein appears as a dense MM band on
electrophoresis

5.0 CLINICAL FEATURES

1. Bone pain mainly involves the back and ribs, pathological fractures.
2. Susceptibility to infections especially pneumonias and pyelonephritis. The
commonest organisms include Streptococcus pneumoniae, Staphylococcus
aureas, Klebsiella, E. coli, and gram-negative Bacilli, Herpes varicella and
Herpes simplex. This is due to reduced immunity reduced production and
increased destruction of antibodies (Ig) and granulocyte dysfunction and
neutropenia.
3. Renal failure due to hypercalcaemia, glomerular deposits, hyeruricaemia,
precipitation of Bence Jones proteins and infiltration with tumour cells.
4. Anaemia
5. Hypercalcaemia
6. Bleeding tendencies
7. Neurological symptoms
8. Hyperviscoisty syndrome producing headache, fatigue, visual disturbances

6. Bleeding tendencies
7. Neurological symptoms
8. Hyperviscoisty syndrome producing headache, fatigue, visual disturbances
and haemorrhages.
9. Amyloidosis
10. Extramedullary disease involving the lymph nodes, skin, liver, spleen,
lungs and meninges.

6.0 DIAGNOSIS
Classic triad of: 1. Marrow plasmatosis (>10%)
2. Radiological evidence of lytic bone lesions
3. Demonstration of serum and/or urine M component (MM band and Bence
Jones proteins respectively)
Diagnostic Criteria
1 major + 1 minor or at least 3 minors
Major Criteria
Plasmacytoma on tissue biopsy
Marrow plasmacytosis > 30%
Monoclonal protein
o IgG
o IgA
o BJP

3.5 gm/dl
>
> 2 gm/dl
> 1 gm/24 hours

Minor Criteria

Marrow plasmacytosis 10 29%

Monoclonal protein present but less than the levels defined above
Lytic bone lesions
Decrease in uninvolved immunoglobulins
o IgM
< 50 mg/dl
o IgA
< 100 mg/dl
o IgG
< 600 mg/dl

7.0 INVESTIGATIONS
1. Haematological

2. Immunological

3. Biochemical

What will
a. Blood count anaemia, increased ESR and rouleaux on
you need
blood film, reduced wbc count
to know
b. Bone marrow aspirate/trephine biopsy Bone marrow
about
infiltration with plasma cells
these
situations
?
a. Serum MM band
b. Urine Bence Jones Proteins (BJP)
c. Serum Ig increased IgG, IgA

a. Hypercalcaemia

3. Biochemical

a. Hypercalcaemia
b. Uraemia
c. Increased creatinine
d. Hypoalbuminaemia

4. Radiological

a. X-rays show lytic bone lesions punched out osteolytic

lesions; pathological fractures and soft tissue masses
b. MRI

8.0 STAGING
Stage
I

Criteria

Hb > 10 gm/dL, serum calcium < 12 mg/L, Normal bone on X-ray or

solitary lytic lesion, Low M-component IgG < 5 gm/dL, IgA < 3
gm/dL, urine light chains (BJP) < 4 gm/24 hours

II

Intermediate

III

Hb < 8.5 gm/dL, Calcium > 12 mg/L, advanced lytic bone lesions, high Mcomponent IgG > 7 gm/dL. IgA > 5 gm/dL and urine light chains >
12gm/24 hours.

Note: All stages are further classified as A or B where A serum creatinine <
2gm/dL and B serum creatinine is > 2gm/dL.

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