Session 5

Question 1

What’s the difference between thalassaemia and sickle cell disease?

Answer 1

Difference - thalassaemia has reduced rate of synthesis of
normal α- OR ß- globin chains (the α- and ß- thalassaemias) whereas sickle cell disease has normal rate of synthesis but produces abnormal haemoglobin.
Similarity - they are both genetic

Question 2

What populations is thalassaemia prevalent in?

Answer 2

Thalassaemia is a heterogeneous group of genetic disorders with varied expression worldwide – traditionally prevalence in S Asian, Mediterranean, Middle east (Beta) and Far East (alpha)

Question 3

How are the globin chains in haemoglobin encoded in DNA?

Answer 3

The α-globin chains are encoded by a pair of α-globin genes on chromosome 16 whilst
the γ-(gamma),δ-(delta) and β-globin genes are together in a cluster on chromosome 11.
• Normal expression of these genes under tight control to ensure a 1:1 ratio of α to non–α globin chains.
• Defects in the regulation of the expression of these genes can result in abnormalities in
the relative and absolute amounts of the globin chains resulting in thalassaemia

Question 4

What causes hypochromic microcytic red cells in thalassaemia?

Answer 4

Low level of intracellular haemoglobin accounts for hypochromic microcytic red cells

Question 5

Why can thalassaemia described as a haemolytic anaemia?

Answer 5

The relative excess of the other globin chain (eg in ß-thal get insoluble aggregates of alpha chains) contributes to the defective nature of the red cell as the aggregates get oxidised and damage the red cell membrane
• Most of the maturing erythroblasts are destroyed within the bone marrow and there is excessive destruction of mature red cells in the spleen
• So… as well as defective Hb production this is a form of haemolytic anaemia also as the red cells are destroyed

Question 6

What are the types of α-thalassaemia?

Answer 6

Silent carrier state
α-Thalassemia trait
Haemoglobin H (HbH) disease
Hydrops fetalis

Question 7

What is the silent carrier state of α-thalassaemia?

Answer 7

Deletion of a single α-globin gene.

It is asymptomatic, without anaemia

Question 8

What is α-Thalassemia trait?

Answer 8

Deletion of two α-globin genes.
It may affect both genes of one chromosome or one gene of each chromosome
There is minimal or no anaemia and no physical signs; findings are identical to those of β-thalassemia minor (microcytosis and hypochromia)

Question 9

What is Haemoglobin H (HbH) disease?

Answer 9

Deletion of three α-globin genes.
Tetramers of β-globin, called HbH, are formed
There is moderately severe anaemia, resembling β-thalassemia intermedia (microcytic, hypochromic anaemia with target cells and Heinz bodies in the blood film)

Question 10

What is hydrops fetalis?

Answer 10

Deletion of all four α-globin genes
In the foetus, excess of γ-globin chains form tetramers (Hb Bart) that are unable to deliver the oxygen to tissues. Usually intrauterine death

Question 11

What are the different types of β-Thalassaemia?

Answer 11

• β-Thalassaemia major
• β-Thalassaemia minor or β-thalassemia trait
• β-Thalassaemia intermedia

Question 12

Describe how genes can be altered in β-thalassaemia?

Answer 12

β-thalassaemia
chromosome 11 - often gene mutations, rather than deletions
β0=total absence of production
β+= reduction of globin production

Question 13

What is β-Thalassaemia major?

Answer 13

Severe transfusion-dependent anaemia that first becomes manifest 6 to 9
months after birth as synthesis switches from HbF (α2γ2) to HbA (α2β2)
Homozygous
Either type βo or β (βo/βo or β+/β+)

Question 14

What is β-Thalassaemia minor / β-thalassemia trait?

Answer 14

Usually asymptomatic with a mild anaemia (very microcytic and hypochromic)
Heterozygous
One normal gene (βo/β or β+/β)

Question 15

What are the consequences of thalassaemia?

Answer 15

• Extramedullary haemopoiesis is an attempt to compensate but results in
splenomegaly, hepatomegaly and expansion of haemopoiesis into the bone cortex
..this impairs growth and causes classical skeletal abnormalities
• Reduced oxygen delivery leads to stimulation of EPO which further contributes to
the drive to make more defective red cells
• Iron overload is major cause of premature death and occurs due to:
• Excessive absorption of dietary iron due to ineffective haematopoiesis
• Repeated blood transfusions required to treat the anaemia
• Reduced Life expectancy

Question 16

What is the treatment for thalassaemia?

Answer 16

• Transfusions of red cells from childhood
• Iron chelation
• Folic acid
• Immunisation
• Holistic care – cardiology, endocrine, psychological, ophthalmology input to
pick up and manage complications
• Stem cell transplantation in some – replace the defective red cell
production
• Pre-conception counselling for at risk couples and antenatal screening

Question 17

What is sickle cell disease?

Answer 17

• Inheritance of the sickle ß-globin chain
• A point mutation causes substitution of valine for glutamic acid in position 6 in the ß chain
• HbSS = homozygous sickle cell anaemia, is most common cause of severe sickling syndrome
• HbS can also be co-inherited with another abnormal Hb eg HbC (HbSC ) or ß-thal (HbS
ß-thal) to cause a sickling disorder
• HbS carrier state causes a mild asymptomatic anaemia and is found in up to 30% of W African people as it confers protection against malaria

Question 18

Why do symptoms arise in sickle cell disease?

Answer 18

• Symptoms of anaemia usually mild ie the anaemia is well tolerated as HbS readily gives up Oxygen in comparison to HbA
• Problems come in low oxygen state when the deoxygenated HbS forms polymers and the red cells form a sickle shape
• Irreversibly sickled red cells are less deformable and can cause occlusion in
small blood vessels – ‘sticky’

Question 19

What are the three types of crises experienced by patients with sickle cell disease?

Answer 19

1. Vaso-occlusive
Painful bone crises
Organ – chest, spleen
2. Aplastic (often triggered by parvovirus infection) (more common in children)
3. Haemolytic

Question 20

What is the end result and often cause of death for patients with sickle cell disease?

Answer 20

End organ damage
as a result of chronic or acute thromboses or oxygen deprivation. Can lead to stroke, retinopathy, blindness, lung problems (pneumonia, infarcts, acute chest syndrome), iron overload in heart and liver, atrophic spleen, skin ulceration, kidney problems, osteomyelitis

Question 21

How can you tell the difference between someone with iron deficiency and someone with β-Thalassaemia minor or β-thalassemia trait

Answer 21

both will show very microcytic and hypochromic RBCs but only the patient with iron deficiency will be very anaemic. Patient with β-Thalassaemia minor or β-thalassemia trait will be asymptomatic with very mild anaemia.

Question 22

What can trigger crises in sickle cell patients?

Answer 22

cold weather, stress, infections

Question 23

Why might someone with sickle cell disease be taking antibiotics?

Answer 23

they have atrophic spleen due to many small infarcts so are considered asplenic so take antibiotics to help with their immune system.

Question 24

What are the consequences of sickle cell disease?

Answer 24

• Reduced Life expectancy – though improving in UK, and life expectancy
with HbSS now 67 years. Commonest causes of death in UK: Stroke,
multi-organ failure, acute chest syndrome
• Acute and chronic pain problems
• Stroke, cognitive and neurological problems, kidney failure, priapism (sustained painful erection) – all a result of capillary occlusion and tissue hypoxia

Question 25

What is haemolysis?

Answer 25

Abnormal breakdown of red blood cells

Question 26

What are the two different types of haemolysis?

Answer 26

• Within blood vessels (intravascular haemolysis)

* Or in the spleen or wider RES (extravascular haemolysis)

Question 27

What capacity does bone marrow have to cope with haemolysis?

Answer 27

The bone marrow can compensate for a decrease in lifespan by increasing red cell production but has capacity to increase roughly 6x so if rate of destruction exceeds
this……. anaemia develops

Question 28

What does haemolytic anaemia cause?

Answer 28

Results in:
• Symptoms of anaemia – severity worse if Hb very low or if a sudden fall in Hb rather than in chronic disease
• Accumulation of bilirubin leading to jaundice and associated risk of complications
such as pigment gallstones.
• Overworking of the red pulp leading to splenomegaly
• Massive sudden haemolysis (as can happen in an incompatible blood transfusion) can cause cardiac arrest due to:
- Lack of oxygen delivery to tissues
- Hyperkalaemia as a result of release of intracellular contents.

Question 29

How can haemolytic anaemias be categorised?

give several examples for each

Answer 29

Inherited (defective gene) or acquired (damage to cells).
Examples of inherited causes of haemolytic anaemia:
Glycolysis defect - Pyruvate kinase deficiency limits ATP production
Pentose-P pathway- G6PDH deficiency leads top oxidative damage
Membrane protein - e.g hereditary sperocytosis
Haemoglobin defect - e.g sickle cell disease

Examples of acquired haemolytic anaemia:
Mechanical damage - e.g macroangiopathic anaemia or from heart valves
Antibody damage - e.g Autoimmune haemolytic anaemia
Oxidant damage - from Exposure to chemicals or oxidants
Heat damage - e.g severe burns
Enzymatic damage - e.g from snake venom

Question 30

What is autoimmune Haemolytic anaemia?

Answer 30

• In this condition autoantibodies (an Immunoglobulin protein produced by person’s own B lymphocytes) bind to the red cell membrane proteins
• Broadly classified as:
• Warm autoimmune haemolytic anaemia (IgG, maximally active at 370C)
• Cold autoimmune haemolytic anaemia (IgM, maximally active at 40C)
• Causes can be infections (eg chest infections in children causing the cold form) or cancers of the lymphoid system (eg B cell lymphoma)
• The spleen recognises the red cell as ‘abnormal’ and removes it .. So reducing the life span

Question 31

What causes an overproduction of cells?

Answer 31

Either caused by Myeloproliferative disorders or as a physiological reaction

Question 32

What are myeloproliferative disorder?

Answer 32

Classed as neoplasms (cancers)
3 conditions:
•Essential Thrombocythaemia (too many platelets)
•Polycythaemia Vera (too many red cells)
•Myelofibrosis (hardening of bone marrow)
•All of these disorders involve dysregulation at the multipotent haematopoietic stem cell

Question 33

What are the clinical features of myeloproliferative disorders?

Answer 33

• Overproduction of one or several blood elements with dominance of a transformed clone.
• Hypercellular marrow / marrow fibrosis.
• Cytogenetic abnormalities.
• Thrombotic and/or haemorrhagic diatheses.
• Extramedullary haemopoiesis (liver/spleen).
• Potential to transform to acute leukaemia.
• Overlapping clinical features.

Question 34

What mutation do many patients with myeloproliferative disorders have?

Answer 34

Many patients have a specific point mutation in one copy of the Janus kinase 2 gene (JAK2)
- a cytoplasmic tyrosine kinase on chromosome 9, which causes increased proliferation and survival of haematopoietic precursors.
We now have specific drugs targeting the aberrant protein

Question 35

Give general information on Polycythaemia.

Answer 35

• Diagnostic criteria = High haematocrit (>0.52 in men, >0.48 in women) OR raised red cell mass
• JAK2 V617F mutation is present in approximately 95% PRV patients
• No reactive cause found
• Some patients also have high platelets &neutrophils
• Median age 60 yrs
• Male=Female

Question 36

What are the clinical feature of polycythaemia vera?

Answer 36

Clinical features:
•Significant cause of arterial thrombosis
•Venous thrombosis
•Haemorrhage into skin or GI tract
•Pruritis (itching)
•Splenic discomfort , splenomegaly
•Gout
•In some transformation to myelofibrosis or acute leukaemia

Question 37

How do we manage polycythaemia vera?

Answer 37

Venesection to maintain the Hct to <0.45
• Aspirin 75 mg unless contraindicated
•Manage CVS risk factors
• Sometimes drugs to reduce the overproduction of cells should be considered

Question 38

Define polycythaemia?

Answer 38

•Polycythaemia = an increase in circulating red cell concentration typified by a persistently raised haematocrit (Hct).

Question 39

What can cause polycythaemia?

Answer 39

This increase can be:
•Relative = normal red cell mass with ↓ plasma volume or
•Absolute = ↑red cell mass:
- Primary = polycythaemia vera
- Secondary – driven by erythropoietin EPO production
*Physiologically appropriate – in response to tissue hypoxia
*Physiologically inappropriate

Question 40

What would cause secondary polycythaemia?

Answer 40

Physiologically appropriate EPO production
Pathological EPO production
Other causes of EPO in blood

Question 41

What would cause physiologically appropriate EPO production?

Answer 41

Central Hypoxia –
Chronic lung disease
R to L shunts
Training at altitude
CO poisoning

Question 42

What can cause ectopic EPO production?

Answer 42

Hepatocellular carcinoma
Renal cell cancer
Cerebellar haemangioblastoma
Uterine tumours
Phaeochromocytoma

Question 43

What would cause pathological EPO production?

Answer 43

Renal Hypoxia –
Renal artery stenosis
Polycystic disease

Question 44

What would be considered an “other” cause of EPO in the blood / secondary polycythaemia?

Answer 44

Erythropoietin injections (can be done to cheat in sport)

Question 45

When looking at polycythaemia what order of questions should you use to evaluate possible causes?

Answer 45

firstly is it absolute (increase in number of erythrocytes) or relative (decrease in plasma volume)?
Then if its absolute, is it primary (abnormality originates in bone marrow. polycythaemia is the only example) or secondary (caused by increased levels of erythropoietin)?
if secondary, is it a physiological response to hypoxia (e.g high altitude or from chronic lung disease) or is it from abnormal EPO production (e.g from renal carcinoma or renal artery stenosis)?

Question 46

What are the features essential thrombocythaemia?

Answer 46

Excess platelets in blood
Large and excess megakaryocytes in bone marrow
Thrombotic events
Screen for Jak2 and CALR mutations

Question 47

How is essential thrombocythaemia managed?

Answer 47

• Any cardiovascular risk factors should be aggressively managed
• Aspirin
• High risk patients:
• > 60 years, platelet count >1500 or disease-related thrombosis/haemorrhage
• return the platelet count into the normal range with drug such as hydroxycarbomide

Question 48

What could be the possible causes that should be excluded before coming to essential thrombocythaemia?

Answer 48

• Infection
• Inflammation (Inflammatory bowel disease, Rh arthritis)
• Other tissue injury (e.g. surgery, trauma, burns)
• Haemorrhage
• Cancer
• Redistribution of platelets -Post-splenectomy and hyposplenism

Most of these conditions cause the release of cytokines which cause the release of platelets and CRP so we can test CRP too.
•Ensure it is persistent rather than transient before investigating for ET

Question 49

What is myelofibrosis?

Answer 49

Heavily fibrotic marrow, little
space for haemopoiesis
Blood film shows red cells
looking like tear drops
A cause of massive splenomegaly
\+/- hepatomegaly
due to extramedullary
haematopoiesis
•clonal haemopoietic stem cell proliferation
•May be end result of PV or ET, or
•Primary disease (PMF)
•PMF starts with proliferative phase when all counts may be high
•Then in all cases progressive pancytopenia due to bone marrow fibrosis and hypersplenism

Question 50

What are the clinical features of myelofibrosis?

Answer 50

• Patients with advanced disease experience severe constitutional symptoms –fatigue, sweats, weight loss
• The consequences of massive splenomegaly
• pain, early satiety (feeling full due to pressure on stomach from large spleen), splenic infarction
• Progressive marrow failure requiring transfusions of blood products
• Transformation to leukaemia
• Early death

Question 51

What is chronic myeloid leukaemia?

Answer 51

Usually presents with very high WCC, this may be incidental finding
•Patients may present with symptomatic splenomegaly, hyperviscosity (sticky blood) or bone pain
•Disease of adults, v rare in children
•Blood film and marrow will show excess of all myeloid
series from blast through to fully mature neutrophils

Question 52

What is pancytopenia?

Answer 52

= Reduction in white cells, red cells and platelets

Question 53

Why might someone get pancytopenia?

Answer 53

Reduced production = most common

or Increased removal of cells

Question 54

What may cause increased removal of cells leading to pancytopenia?

Answer 54

• Immune destruction possible but rare to cause a pancytopenia
• Splenic pooling ~
  Hypersplenism in
  Massive splenomegaly
• Haemophagocytosis =
  Chewing up of the cells in the bone marrow .. v v rare

Question 55

What would cause reduced production of cells leading to pancytopenia?

Answer 55

B12/folate deficiency
Drugs –chemotherapy, antibiotics, anticonvulsants, psychotropic drugs, DMARDs
Viruses –EBV, viral hepatitis ,HIV, CMV
Bone marrow infiltration by malignancy (blood cancers or other cancers)
Marrow fibrosis
Radiation
Idiopathic aplastic anaemia
Congenital bone marrow failure eg Fanconi’s anaemia, dyskeratosis congenital –present in childhood

Question 56

What is aplastic anaemia

Answer 56

Pancytopenia with a hypocellular bone marrow in the absence of an abnormal infiltrate and with no increase in fibrosis

Mortality is high as cure is difficult – immune treatments and bone marrow transplantation

Deaths often due to neutropenic infection or bleeding

Question 57

What role do platelets have?

Answer 57

Key role in Haemostasis to facilitate clot formation, initially via a platelet ‘plug’:
Adhesion to damaged endothelial wall and to vWF
Activation–change in shape from disc and release of granules
Aggregation– clumping together of more platelets to form the plug

Question 58

How can platelet orders be divided?

Answer 58

Quantitative –low (thrombocytopenia)

Qualitative –normal number but defective function

Question 59

How should a clinician evaluate the cause of a patients thrombocytopaenia? Perhaps use a diagram to display your answer.

Answer 59

Is it acquired (common) or is it inherited (rare syndromes)?
If its acquired, is it from decreased platelet production (B12 or folate deficiency, acute leukaemia or aplastic anaemia, liver failure (decreased production of thrombopoietin), sepsis, cytotoxic chemotherapy), increased platelet consumption (Massive haemorrhage, disseminated intravascular coagulation, thrombotic thrombocytopenic purpura) or increased platelet destruction (autoimmune thrombocytopenic purpura, drug induced e.g from heparin, hypersplenism resulting in increase in destruction and splenic pooling of platelets)?

Question 60

What are the consequences of severe thrombocytopenia?

Answer 60

Patients generally not symptomatic until the platelet count <30
Easy bruising
Petechiae, purpura
Mucosal bleeding
Severe bleeding after trauma
Intracranial haemorrhage

Question 61

What is the most common cause of immune platelet destruction?

Answer 61

Immune thrombocytopenic purpura -autoantibodies against Glycoprotein GPIIb/IIIa and GPIb/IX

Question 62

What is another cause of immune platelet destruction? (not Immune thrombocytopenic purpura)

Answer 62

Can be secondary to autoimmune disease eg SLE and lymphoproliferative disorders eg lymphoma, Chronic lymphocytic leukaemia

Question 63

How is immune platelet destruction treated?

Answer 63

Treated with immunosuppression (corticosteroids or Intravenous immunoglobulin first line).
Platelet transfusions do not work (as the transfused platelets get destroyed too)

Question 64

How do we classify disorders of platelet function?

Answer 64

• Hereditary (very rare)
• eg Bernard Soulier syndrome, Glanzmann’s thrombasthenia
• Acquired (very common)
• Aspirin/NSAIDS/clopidogrel–all drugs designed to inhibit the normal function of platelets
• Uraemia
• Other causes

Question 65

Why might spherocytosis cause a low platelet count?

Answer 65

Spherocytosis causes enlargement of red pulp in spleen so platelets spend longer in the splenic circulation and are broken down as a result of hypersplenism.

Question 66

What are the 4 main types of leukaemia?

Answer 66

acute/chronic myeloid

acute/chronic lymphoid

Question 67

What do we know about the genetic change that drives chronic myeloid leukaemia?

Answer 67

Philadelphia chromosome present in people with the condition. . It involves a reciprocal translocation between chromosomes 9 and 22. This translocation causes an oncogenic gene fusion (BCR-ABL) with tyrosine kinase activity that results in proliferation, differentiation and inhibition of apoptosis. Understanding the molecular basis of this translocation has led to the development of targeted cancer therapy through drugs which inhibit the ATP-binding site of the tyrosine kinase. Use of these oral drugs has dramatically improved survival rates in patients with CML.. Imatinib was developed to combat this

Question 68

What is pyruvate kinase deficiency and how does it cause anaemia?

Answer 68

Pyruvate kinase deficiency is an inherited metabolic disorder (typically autosomal recessive but there is also a dominant form) due to mutations in the PKLR gene. There are four pyruvate kinase isoenzymes, two of which are encoded by PKLR (isoenzymes L and R expressed in liver and erythrocytes, respectively). Mutations in the PKLR gene therefore cause a deficiency in pyruvate kinase in erythrocytes. This enzyme catalyses the final step in glycolysis, transferring the phosphate from phosphoenolpyruvate to ADP to form ATP. This is the second ATP producing “pay-back” step of the process. Since red blood cells lack mitochondria, pyruvate kinase deficiency inhibits their only metabolic pathway which can supply ATP for cellular processes. The sodium potassium ATPase pump activity is inhibited from insufficient ATP to provide energy and the red cells lose potassium to plasma. Water moves down its concentration gradient out of cells causing them to shrink resulting in cellular death and haemolytic anaemia. Most affected individuals with pyruvate kinase deficiency have only a mild deficiency in enzyme activity and do not require treatment. Individuals with a more severe deficiency may require regular blood transfusion.

Question 69

What is microangiopathic haemolytic anaemia (MAHA)

Answer 69

Microangiopathic haemolytic anaemias (MAHA) are a group of acquired haemolytic anaemias where red cells are damaged by physical trauma. Often the trauma results from red cells getting snagged as they try to pass through small vessels laden with fibrin strands in situations where there is an increased activation of the coagulation cascade. Such conditions include Disseminated intravascular coagulation (DIC) and Thrombotic thrombocytopenic purpura. Another example is Haemolytic Uraemic Syndrome (HUS) seen more commonly in children after developing E coli diarrhoea. Red blood cells can also be damaged by the shear stress produced by a defective heart valve such as in aortic valve stenosis

Question 70

What is disseminated intravascular coagulation?

Answer 70

Disseminated intravascular coagulation (DIC) - a condition where bleeding and clotting occur at the same time in the patient e.g. in malignancy, obstetric complications, trauma, and sepsis

Question 71

What is thrombotic thrombocytopenic purpura?

Answer 71

This is an autoimmune disease characterised by isolated thrombocytopenia which can take an acute (short-lived) or chronic course. It presents due to symptoms if the platelet count is very low or as an incidental finding on a blood count. It is mainly due to anti-platelet auto-antibodies but T-cell activity against the platelets and megakaryocytes are contributory. Sometimes there is an obvious trigger, such as an acute infection. This is common in children and is associated with a better outcome with often spontaneous improvement in the platelet count and no relapse. Other causes in adults are associated autoimmune diseases such as rheumatoid arthritis or systemic lupus erythematosus (SLE), underlying lymphoid cancers such as lymphoma or leukaemia, or carriage of the human immunodeficiency virus (HIV). No obvious cause is found in 80% of adult cases, Fatal intracranial haemorrhage can occur though is rare (1:500). It is important that ITP is recognised as a cause of severe thrombocytopenia by doctors as the treatment is very different to the alternative causes such as bone marrow failure, Treatments include corticosteroids, other immunosuppressive drugs, intravenous pooled human Immunoglobulin, splenectomy and more recently thrombopoietin receptor agonists.

Question 72

What is G6PDH deficiency and how does it cause anaemia?

Answer 72

Glucose-6-phosphate dehydrogenase (G6PDH) deficiency is an X-linked recessive inborn error of metabolism. G6PDH is the rate limiting enzyme of the pentose phosphate pathway which supplies reducing energy by maintaining NADPH levels. NADPH is required to protect against oxidative stress by maintaining the level of reduced glutathione. Since the pentose phosphate pathway is the only source of reduced glutathione in red blood cells, these are particularly affected by defects in the glucose-6-phosphate dehydrogenase enzyme. Patients with G6PDH deficiency are therefore at risk of haemolytic anaemia in states of oxidative stress such as infection or exposure to certain chemicals or medications. Damaged red cells are phagocytosed in the spleen and metabolism of the excessive haemoglobin to bilirubin can lead to jaundice.

Question 73

What is hereditary spherocytosis and how does it cause anaemia?

Answer 73

Hereditary spherocytosis is an inherited autosomal dominant disease resulting in abnormalities in erythrocyte membrane proteins which impede the ability of the cell to change shape. Mutations in the genes coding for 4 different proteins have been shown to cause the disease:
• Spectrin (an actin crosslinking and molecular scaffold protein that links the plasma membrane to the actin cytoskeleton).
• Ankyrin (links integral membrane proteins to the underlying spectrin-actin cytoskeleton).
• Band 3 (facilitates chloride and bicarbonate exchange across the plasma membrane and is also involved in a physical linkage of the plasma membrane to the underlying cytoskeleton (via binding with ankyrin and protein 4.2)).
• Protein 4.2 (an ATP-binding protein which may regulate the association of protein 3 with Ankyrin).

Although these 4 proteins all have different functions, their common role is in facilitating “vertical interactions” between the cytoskeleton and the lipid bilayer of the plasma membrane. Disruption of this interaction mutation of the genes coding for these proteins results in a local disconnection of the cytoskeleton and membrane. This is followed by vesiculation of the unsupported membrane components leading to progressive reduction in membrane surface area and production of a “spherocyte” shape to the red blood cell. Red cell membranes are normally very flexible but become increasingly less deformable as surface area is lost. The poor deformability of spherocytes means that they become trapped and damaged as they pass through the spleen (extravascular haemolysis) resulting in a reduction in the lifespan of erythrocytes and haemolytic anaemia. The deficit in cell deformability of spherocytes only appears to be a problem in the spleen rather than small capillaries since the red cells return to a nearly normal lifespan following splenectomy.
In addition to anaemia, symptoms include jaundice and splenomegaly. Howell-Jolly bodies may be seen within red blood cells. Patients with mild symptoms do not usually require treatment but for those with severe symptoms partial or full splenectomy improves the anaemia. The spherocytes still remain but are no longer lysed by the spleen.

Question 74

What are Haemoglobinopathies?

Answer 74

Haemoglobinopathies are inherited disorders where expression of one or more of the globin chains of haemoglobin is abnormal. There are two main categories:
• Abnormal haemoglobin variants: result from mutations in the genes for α or β globin chains that alter the stability and/or function of haemoglobin (e.g. Sickle cell disease).
• Thalassaemias: result from reduced or absent expression of normal α or β-globin chains. This leads to a reduced level of haemoglobin rather than the presence of an abnormal haemoglobin.

Haemoglobinopathies are typically autosomal recessive diseases. Carriers (heterozygotes) of the disease usually show very mild or no symptoms whereas homozygotes, who have inherited an abnormal gene from both parents, show the symptoms of the disease. The UK has a genetic sickle cell and thalassaemia screening programme in place.

Question 75

Explain the normal structure of haemoglobin, how its coded and how expression changes?

Answer 75

The haemoglobin molecule consists of a tetramer of four globin polypeptide chains; two alpha chains and two non-alpha chains (β, δ or γ), held together by noncovalent interactions with each globin chain complexed with an oxygen binding haem group. The globin chain compositions of the 3 main normal types of haemoglobin present in adult are as follows:
Haemoglobin type - Globin chains - Proportion of total Hb in adult
A - 2α/2β - >95%
A2 - 2α/2δ - 2-3.5%
F - 2α/2γ - <1%
Fetal haemoglobin (haemoglobin F) is the main form expressed before birth but the synthesis of haemoglobin A also commences before birth and steadily increase to become the dominant form over the first few months of life and into adulthood.
The α-globin chains are encoded by a pair of α-globin genes on chromosome 16 whilst the γ-,δ- and β-globin genes are together in a cluster on chromosome 11. The expression of these genes are under tight control to ensure a 1:1 ratio of α to non–α globin chains. Defects in the regulation of the expression of these genes can result in quantitative abnormalities in the relative and absolute amounts of the globin chains resulting in thalassaemia whilst defects in the coding regions can result in abnormal variants with structural defects that alter the stability and/or function of haemoglobin.

Question 76

What are the symptoms of sickle cell disease?

Answer 76

The consequences of sickle cell formation include:
• Vaso-occlusive episodes due to occlusion of small capillaries from sickle cells getting trapped. This leads to recurrent acute pain and syndromes such as stroke or acute chest syndrome as
well as chronic kidney disease and joint damage from avascular necrosis.
• Anaemia due to sickle cells undergoing haemolysis resulting in a shortened erythrocyte lifespan from ~120 days to ~20-30 days.
• Jaundice and gallstones due to increased bilirubin resulting from chronic haemolysis.
• Splenic atrophy due to splenic infarction with an associated susceptibility to infection by encapsulated bacteria such as Streptococcus pneumoniae and Streptococcus meningitidis

Question 77

What is sickle cell disease and explain how it causes change of red blood cell shape?

Answer 77

The most common haemoglobin variant of clinical significance is haemoglobin S (HbS). The HbS variant is found mainly in people of Black African descent but is also common in Arab, Mediterranean, and South Asian populations. Heterozygous individuals for HbS have some resistance to malaria due to changes in the red blood cell making it difficult for the Falciparum parasite to grow. Homozygous individual however develop sickle cell disease and combinations with other haemoglobinopathies produce sickling syndromes of variable severity such as sickle-β-thalassaemia, HbS/C or HbS/E.
The HbS variant has an uncharged valine instead of a charged glutamic acid at position 6 of β-globin making haemoglobin S more prone to polymerise at low oxygen tension. This leads to the formation of long twisted haemoglobin polymers that can result in the deformation the red blood cell membrane leading to the cell taking on a sickle shape. After repeated episodes of sickling, damage occurs to the red cell membrane causing it to lose elasticity. Such damaged cells fail to return to a normal shape when normal oxygen tension is restored.

Question 78

What are myoproliferative neoplasms and what are the four types?

Answer 78

Myeloproliferative neoplasms (MPNs) are a group of diseases of the bone marrow in which excess cells are produced. They arise from genetic mutations in the precursors of the myeloid lineage in the bone marrow.
There are 4 major types of MPN, depending on the type of cells that are overproduced:
1. Polycythaemia vera (excess erythrocytes)
2. Essential thrombocythaemia (overproduction of megakaryocytes leading to excess platelets)
3. Primary myelofibrosis (initial proliferative phase followed by replacement of haematopoietic tissue by connective tissue leading to impairment of the generation of all blood cells (pancytopenia))
4. Chronic myeloid leukaemia (excess granulocytes)

Question 79

What is polycythaemia? Give an overview (Symptoms, treatment, causes etc)

Answer 79

Polycythaemia is a disease state in which the volume percent of erythrocytes in the blood (the haematocrit) exceeds 52% (males) or 48% (females). This situation can arise from either an increase in the number erythrocytes (termed absolute polycythaemia) or to a decrease in the plasma volume (termed relative polycythaemia).
Polycythaemia vera (PV) is a specific form of polycythaemia which arises from a myeloproliferative neoplasm in the bone marrow resulting in overproduction of erythrocytes (although in some case white blood cells and platelets can also be overproduced). Most (~95%) cases are caused by mutation of the gene coding for Janus Kinase 2 (JAK2), a cytoplasmic tyrosine kinase whose normal function is to stimulate signalling pathways leading to erythrocyte production in response to the hormone erythropoietin. Multipotent stem cells harbouring the JAK2 mutation survive longer and proliferate continuously.
The clinical features associated with polycythaemia vera essentially result from the blood being thicker and include:

• Thrombosis (venous & arterial)
• Haemorrhage (skin or GI tract)
• Headache and ‘dizziness’
• Plethora
• Burning pain in the hands or feet (Erythromelalgia)
• Pruritus
• Splenic discomfort, splenomegaly
• Gout
• Arthritis
• May transform to myelofibrosis or acute leukaemia
Treatment consists primarily of phlebotomy to maintain the haematocrit below 45%. Aspirin due to its anti-platelet effects may be prescribed unless contraindicated. Cytoreduction using agents such as hydroxycarbamide (oral antimetabolite that inhibits DNA synthesis) should be considered if the patient has poor tolerance of venesection, shows symptomatic or progressive splenomegaly or other evidence of disease progression, e.g. weight loss and night sweats.
Secondary causes of polycythaemia are far more common than PV and are driven by increased stimulation by erythropoietin. Causes of this are logical if you consider the oxygen dissociation curve: living at high altitude; chronic hypoxia (severe chronic obstructive pulmonary disease, heavy long-term cigarette use or cyanotic heart disease); renal disease. It can also be caused by tumours secreting erythropoietin.

Question 80

What is essential thrombocytopenia?

Answer 80

Thrombocytosis is an increase in the platelet count compared to the normal range of a person of the same gender and age. Thrombocytosis is a common reaction to infection and inflammation and is seen often in hospitalised patients. However, it can also result from a myeloproliferative neoplasm, in which case the thrombocytosis is generally referred to as essential thrombocythaemia. This rare chronic blood cancer is characterised by the overproduction of platelets by megakaryocytes in the bone marrow. Around half the cases of essential thrombocythaemia are caused by the same JAK2 mutations found in Polycythaemia vera. Mutations in the thrombopoietin receptor can also result in the disease. The most common symptoms of essential thrombocythaemia are as follows:
• Numbness in the extremities
• Thrombosis (most often arterial e.g. stroke or peripheral gangrene)
• Disturbances in hearing and vision (related to microvascular complications)
• Headaches
• Burning pain in the hands or feet (Erythromelalgia)
Patients are typically classified as low and high risk for bleeding/blood clotting (based on age, medical history, blood count and lifestyle) and treated accordingly. Low risk patients are usually treated with aspirin whilst high risk patients are also given hydroxycarbamide and/or other treatments that reduce platelet count.

Question 81

Give an overview of thrombocytopenia?

Answer 81

Thrombocytopenia
In contrast to thrombocytosis (increased platelets) the condition of an abnormally low level of platelets is called thrombocytopenia. Thrombocytopenia can arise for a number of reasons and can either be inherited or acquired.
Inherited thrombocytopenia
A number of rare inherited syndromes (the details and names of which are beyond the scope of this unit) are associated with a decrease in platelet number. These include Fanconi anaemia, Bernard-Soulier syndrome, (associated with large platelets) and Alport syndrome.
Acquired thrombocytopenia
Acquired thrombocytopenia is much more common and is typically detected from a full blood count. Some individuals with a very low platelet count may experience bleeding gums, nosebleeds, and women may have heavier or longer menstrual periods. Spontaneous bleeding under the skin may sometimes also be visible as bruising and petechiae (small red/purple spots on the skin) are usually seen on the feet, shins and forearms. Acquired thrombocytopenia can result from a decrease in platelet production, increased consumption of platelets (e.g. due to DIC) or an increased destruction of platelets.
Treatment depends on the specific cause of the disease and focuses on eliminating the underlying problem (e.g. discontinuing a drug or treating underlying sepsis).

Question 82

Give an overview of primary myelofibrosis

Answer 82

Primary myelofibrosis
Primary myelofibrosis is a myeloproliferative neoplasm where the proliferation of mutated hematopoietic stem cells results in reactive bone marrow fibrosis eventually leading to the replacement of marrow with scar tissue (collagen deposition). Mobilisation of mutated progenitor cells from bone marrow can also occur and these cells can colonise the liver and spleen leading to extramedullary haemopoiesis (i.e. the production of blood cells outside the medulla of the bone). For this reason patients with primary myelofibrosis often show an enlarged liver and spleen. Similar to polycythaemia vera and essential thrombocythaemia, mutations in the JAK2 gene are often associated with the disease. The term “myelofibrosis” alone usually refers to primary myelofibrosis. The term secondary myelofibrosis is used when the disease has developed as a consequence of polycythaemia vera or essential thrombocythaemia.
Symptoms include:
• Hepatosplenomegaly
• Bruising
• Fatigue (and other symptoms related to anaemia)
• Weight loss
• Fever
• Increased sweating
• Portal hypertension
Treatment options are largely supportive and generally unsatisfactory (median survival for the disease is ~5 years). Hydroxycarbamide, folic acid, and allopurinol may have some benefits and patients with advance disease may require blood transfusions and sometimes splenectomy. Recently the drug ruxolitinib, an inhibitor of JAK2 has been shown to significantly reduce spleen volume and improve symptoms of myelofibrosis.

Question 83

What is aplastic anaemia?

Answer 83

Aplastic anaemia is a rare disease resulting in damage to bone marrow and hematopoietic stem cells leading to pancytopenia (a deficiency of all three blood cell types: red blood cells (anaemia), white blood cells (leucopenia), and platelets (thrombocytopenia). The term “aplastic” simply refers to an inability of the stem cells to generate mature blood cells. Aplastic anaemia can be caused by genetic causes, auto immunity or exposure to chemicals, drugs, or radiation.

Question 84

Give information on leukaemia specifically chronic myeloid.

Answer 84

In simple terms, the best way to think about the types of leukaemia is whether they are ‘acute’ or ‘chronic’ and what lineage do the abnormal clonal malignant cells arise from. Acute leukaemias rapidly cause bone marrow failure due to large numbers of immature blast cells overwhelming the ability of the tissue to produce mature blood cells. Chronic leukaemias are more often slow to cause symptoms and may even be picked up as a chance finding on a blood count. In chronic leukaemias (in contrast to acute leukaemia) there is often differentiation i.e. the malignant clonal cell may be a mature cell, as in the commonest leukaemia B-CLL (chronic lymphocytic leukaemia).
Chronic myeloid leukaemia (CML) which accounts for 15–25% of leukaemia in adults is characterized by the unregulated growth of myeloid cells in the bone marrow leading to the accumulation of mature granulocytes (mainly neutrophils) as well as myelocytes in blood. It is associated with a specific chromosomal translocation called the Philadelphia chromosome involving a reciprocal translocation between chromosomes 9 and 22. This translocation causes an oncogenic gene fusion (BCR-ABL) with tyrosine kinase activity that results in proliferation, differentiation and inhibition of apoptosis. Understanding the molecular basis of this translocation has led to the development of targeted cancer therapy through drugs which inhibit the ATP-binding site of the tyrosine kinase. Use of these oral drugs has dramatically improved survival rates in patients with CML.