Haemaglobinopathies Flashcards by Aífe Hayden

What is a haemoglobinopathy

The normal production of structurally abnormal globin chains

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What may cause haemoglobinopathies
(4)

Point mutations
Deletion of triplets
Mutation of a stop codon
A fusion mutation or due to an insertion

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Give some examples of haemoglobin varients

Hb C
Hb S/C
Hb E
Hb D

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What is Hb C

B6 Glu>lys

Results in mild haemolytic anaemia

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What is Hb S/C

This accounts for 20-50% of patients with sickle cell disease

Causes less severe vaso-occlucive complications

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What is Hb E
(3)

Most common mutation

B26 Glu.lys

Homozygotes suffer from mild anaemia

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What is Hb D
(2)

B121 Glu>Gln

Mostly asymptomatic

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What percentage of west Africans have sickle cell disease

1 in 4

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Comment on the severity of sickle cell disease
(6)

HbS

Heterozygotes are asymptomatic

Homozygotes have SCD

Haemolytic anaemia of varying severity punctuated by painful sickling crises

Reduced life expectancy with most people suffering in Nigeria dying before 21 years of age

Approximately 20% of children die under 5 in Africa

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How does sickle cell disease confer protection against malaria
(3)

Hb S shape is difficult for the malaria parasite to use

Macrophages will also continuously break down these sickle cells

Parasites are unable to get to the end of their life cycle before being phagocytosed along with the sickle cell

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What causes structural/qualitative defects of Hb

Occurs when the amino acid sequence of one globin chain is altered

Mutation may arise due to addition, substitution, deletion or fusion to the base sequence in the globin chain -> this leads to a change in the amino acid sequence

Leads to sickle cell anaemia

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How does sickle cell anaemia come about
(5)

Due to a change in the base sequence of the B chain

An A is changed to a T

GAG&raquo_space;> GTG

This results in the replacement of glutamic acid by valine at position 6 on the B chain

This causes a change in the overall charge and polarity of the molecule

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How is HbS diagnosed in the lab
(7)

Positive solubility test

Blood film

Full Blood Count

DNA analysis to confirm genetic mutation

Electrophoresis

HPLC

General bloods

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How is a blood film used to diagnosed HbS
(7)

Sickle cells
Target cells
Polychromasia
Hypochromic microcytes
Reticulocytes
NRBC
Howel Jolly bodies if hyposplenic

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How is a full blood count used to diagnose HbS

Haemoglobin is low

MCV is low

MCH is low

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How is High Performance Liquid Chromatography used to diagnose Hb
(4)

Hb A is absent
Hb S 80-100%
Hb F 5-15%
Increased HbA2

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How is the Bio-Rad Varient II used in the primary screening of B-thalassaemia
(6)

Analyser utilises ion-exchange HPLC to investigate B-thalassaemia

A B-thalassaemia short program test kit is used with the analyser

Samples are diluted and mixed at the sampling station and then injected into the analytical cartridge

Hb is fractioned by type, based on its ionic interactions with the reagent

Results are shown as a chromatogram

The analyser is usually interfaced to the Laboratory Information System (LIS)

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How is the Sebia Capillarys 2 Flex Piercing - Confirmatory Testing used
(3)

Analyser uses capillary electrophoresis to confirm thalassaemia and other haemoglobinopathies

Works by separating charged molecules by their electrophoretic mobility in an alkaline buffer when voltage is applied

The Hb fractions are detected at 415nm at the cathode of the capillary and an electrophoretogram is produced, together with quantified fractions of Hb

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How is sickle cell anaemia treated
(4)

Drug treatments

Allogenic transplant

Ex vivo gene therapy

In vivo gene therapy

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How are drugs used to treat sickle cell anaemia

These aim to allow more cells to transit the microcirculation before sickling

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How are allogeneic transplants used to treat sickle cell anaemia

Bone marrow stem cells from a donor without SCD are transplanted

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How will Ex vivo gene therapy be used in the future to treat SCD

The patients bone marrow cells are modified by adding a B-globin gene using a retroviral vector or with gene editing

This will reactivate foetal haemoglobin (HbF) or correct the disease mutation

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How will Ex vivo gene therapy be used in the future to treat SCD

The patients bone marrow cells will be modified by adding a B-globin gene

This will be done using either a retroviral vector or with gene editing

This will reactivate foetal haemoglobin or correct the disease mutation

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How will In vivo gene therapy be used in the future to treat SCD

Direct gene editing in patients could circumvent the need for transplantation of modified patient cells if sufficient efficiency and safety can be achieved

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What are quantitative abnormalities of Hb

Heterogenous group of genetic disorders which result from a reduced rate of synthesis of the alpha or Beta globin chains

What causes quantitative abnormalities

May arise because of: Inadequate amount of mRNA Unstable mRNA mRNA that contains untranslatable nonsense message

What happens when there is quantitative abnormalities of Hb (Thalassaemias)?

There is a decrease in one of the globin chains So the other chain is in excess and thus the molecule is unbalanced and the chain in excess tries to form tetramers and precipitates within the cell

What are quantitative abnormalities of Hb called

Thalassaemias

Write a note on the types of thalassaemia (2)

Some mutations cause a complete absence of globin chain synthesis -> a0 or B0 thalassaemias In other mutations the globin chain is produced at a reduced rate and these are known as a+ or B+ thalassaemias

What are the three groups of thalassaemias

Thalassaemia major Thalassaemia intermedia Thalassaemia minor

What is thalassaemia major

Severe anaemia Transfusion dependant

What is thalassaemia intermedia

Phenotype ranging from severe anaemia to mild hypochromic microcytic anaemia

What is thalassaemia minor

Clinically asymptomatic carrier state

What is the most sever thalassaemia genetically

B0 B0

What haemoglobin is affected by a Thalassaemia?

HbA (aB) A2 (aB) F (ay)

What haemoglobin is affected by B Thalassaemia?

HbA (aB)

What haemoglobin is affected by y Thalassaemia?

HbF (ay)

What haemoglobin is affected by d Thalassaemia?

HbA2 (ad)

What happens in beta thalassaemia (4)

Excess alpha chain produced which are highly unstable and precipitate in red cells Degradation of excess a chains produce deleterious effects on RC membrane and electrolyte homeostasis - shortened RC survival alpha chain inclusions in red cell results in haemolysis in spleen Anaemia becomes apparent after 3-6 months of age when they switch from gamma to beta chain production occurs

How is B Thalassaemia diagnosed in the lab (6)

Full Blood Count Blood film HPLC Reticulocytes DNA analysis to confirm genetic mutation Family studies

How is B thalassaemia diagnosed on a blood film (7)

Hypochromic microcytes Target cells Anisopoikloctosis Polychromasia NRBC Pappenheimer bodies basophilic stippling

How is a FBC used to diagnose B Thalassaemia

Haemoglobin down MCV down MCH down

How is HPLC used to diagnose B thalassaemia

Reduced or absent HbA peak Increased Hb F Increased Hb A2

How is B thalassaemia treated (4)

Conventional therapies Pharmaceutical induction of y-globin Allogenic HSC transplantation Gene and cell therapy

What conventional therapies are used to treat B thalassamia

Transfusions and iron chelation therapy are used Frequent transfusions are needed which result in high iron which needs to be chelated to prevent overload

How is pharmaceutical induction of y-globin used to treat Thalassaemia

Drugs given to induce the production of y-globin

How is gene and cell therapy used to treat Thalassaemia

B or y-globin vectors to bring about the production of the genes

How is allogeic HSC transplantation used to treat thalassaemia

Haematopoeitic stem cell transplant

What percentage of people does thalassaemia affect

What percentage of thalassaemias are alpha thalassaemia

What are the different types of a thalassaemia (4)

Carrier: asymptomatic a-thal minor: only 2 alpha genes -> asymptomatic or mild microcytic anaemia Hb H disease -> only 1 working a gene -> haemolytic and microcytic anaemia -> splenomegaly no working alpha gene -> incompatible with life -> hydrops fetalis -> most common occurring type -> excess fluid builds up in the body before birth

How is alpha thalassaemia diagnosed in the lab (8)

FBC Blood film Reticulocytes H Prep HPLC Family studies DNA analysis General blood

How is FBC used to diagnose a- thalassaemia

Hb down MCV down MCH down

How is blood film used to diagnose a- thalassaemia

Hypochromic microcytes Target cells Anispoikilocytosis Polychromasia NRBC Pappenheimer bodies Basophilic stippling

How is general bloods used to diagnose a- thalassaemia

LDH and bilirubin up Haptoglobin down

How is H prep used to diagnose a- thalassaemia

Incubation with brilliant Cresyl Blue typical golf ball appearance (30-90%)

How is HPLC used to diagnose alpha thalassaemia

Hb H peak Hb Barts peak

What type of inclusions are seen in thalassaemias

Inclusion body Heinz body Howell-Jolly body

What is an inclusion body and how does it come about (3)

Seen in HbH disease (a type of a thal) Excess B chains precipitating as haemoglobin H inclusion bodies in the cell In B thalassaemia major, excess a chains can also precipitate as inclusion bodies

What is a Heinz body and how does it come about (4)

Type of inclusion body containing denatured haemoglobin Classically associated with G6DP deficiency but can be found in thalassaemias as well Typically larger than inclusion bodies If functional spleen is present these will lead to bite cells

What are Howell-Jolly bodies and how does it come about (3)

Type of inclusion body containing DNA Usually removed by splenic macrophages like Heinz bodies Can be seen when red cells fail to mature or when a functional spleen is absent

What is hereditary persistence of foetal Hb (3)

Rare, benign condition characterised by continued synthesis of HbF in adults No clinical symptoms of thalassaemia Little significance except when combined with other forms of thalassaemia or Haemoglobinopathies

What happens if hereditary persistence of foetal Hb is combined with sickle cell anaemi

It produces a milder form of disease due to the presence of Hb F

What is the one benefit of HbF

Its more resistant to denaturation than Hb A

How do we identify hereditary persistence of foetal Hb in the lab

Can be demonstrated on blood smears using Kleihauer Betke stain Only cells containing HbF will stain

Classify hereditary persistence of foetal Hb

Classified according to distribution of Hb F among red cells Pancellular HPFH Heterocellular HPFH

What is pancellular HPFH

Hb F uniformly distributed throughout red cells

What is heterocellular HPFH

Hb F found in only small number of cells

Haemaglobinopathies Flashcards

(68 cards)