7- Haemoglobinopathies

Qualitative and quantitative haemoglobin disorders

• Haemoglobinopathies- abnormalities in the structure or synthesis caused by globin gene mutations (haem normal)

• Qualitative- amino acid (missense single) deletion or substitution, sickle cell

• Quantitative- alpha and beta thalassaemia

Structural haemoglobin variants

• Most due to single point mutation, single AA substitution

• HbS glutamic acid (6) to valine

• HbC- glutamic acid (6) to lysine

• HbE- glutamic acid (26) to lysine

• All above on beta chain gene

• Homozygous- SS sickle cell disease, heterozygous trait (carrier)

• Two abnormal can be present (heterozygous HbSC)

Ways to observe abnormal haemoglobin

• Substitution may affect overall charge of molecule

• Seen by electrophoresis as speed of migration changes, thick bands if a lot

• High performance liquid chromatography- sample placed into mobile phase which enters the stationary phase (column), components are separated and converted into electrical signal by detector

• Genetic variants detected by PCR

Haemoglobin S

• Affects B chain, glutamic acid replaced by valine (uncharged)

• Less soluble than HbA

• They polymerise (bind) into long fibres which deforms the red blood cell to form sickle shape

• Reversible but permanent after a few cycles

Sickle cell disease vs trait

• Homozygous experiency sickling at much higher PO2

• Heterozygous at lower PO2, one normal B gene one abnormal, non HbS delays polymerisation, usually asymptomatic

• Prevelent in africa, mediterranean, parts of middle east and southern india

Pathophysiology of sickle cell anemia

• Cells sickle when PO2 e.g. higher altitude, deoxygenated HbS is 50x less soluble that HbA

• Repeated cycles of sickling and reversal, eventually irreversible

• Only survive 10-20 days, removed by extravascular haemolysis by macrophages in spleen, liver BM which results in anaemia

• Can obstruct small blood vessels are they are less flexible

• Continious cycle- structural change causes higher blood viscosity which leads to obstructions, tissue hypoxia which leads to more sickling, can eventually lead to tissue infration

Symptoms of sickle cell anaemia

• Asymptomatic in first year of life as fetal haemoglobin is present

• Classified as chronic haemolytic anaemia, may cause gallstones (bilirubin)

• Sickling can be spontaneous or caused by hypoxia, acidosis, hypotension, infection, dehydration, hypothermia

• Tissues become infracted- hands and feet, spleen (splenomegaly), liver (reduced extramedullary haematopoiesis)

• Patients more susceptible to infection

• Untreated crisis associated with higher morbidity and mortality

• Acute chest syndrome most common cause of death after age 2, lung infraction

•Strokes, microvascular occlusions

Laboratory findings

• Low haemoglobin, red cell count, haematocrit

• Normocytic, normochromic, reticulocytes present

• Sickle cells on blood film

• Elevated bilirubin and LDH (haemolysis)

• Electrophoresis- exclusively HbS or HbS and HbA

• Decreased solubility- cells are lysed open but HbS does not dissolve easily in plasma, will turn cloudy after adding sodium dithionite (HbA clear)

Types of thalassaemia

• Reduced production of globin due to gene deletion or point mutation, structure is normal

• Leads to ineffective erythropoiesis and shortened red cell life span

• Alpha and beta types (minor and major forms)

• Healthy at birth, become anaemic between 3-18 months, major more severe

• Most common in greece, mediterranean, asia, middle east, caribbean

Pathophysiology of alpha thalassaemia

• Normal ration is a 1:1 production of a and B chains

• Excess of one chain in thalassaemia leads to decreased haemoglobin production, ineffective erythropoiesis, chronic haemolysis

• In alpha thalassaemia excess B chains can combine to form a haemoglobin with four B chains (HbH)

• HbH has a high affinity for oxygen and does not unload easily, cells prematurely destroyed in the spleen

• In infants excess gamma chains combine to form haemoglobin Bart’s

Alpha thalassaemia inheritence

• Four alpha genes in normal individual

• Occurs when there is a deletion in any of the genes

• Carrier- loss of one gene, no symptoms

• Minor- loss of two genes, milk anaemia (losso of one from each chromatid or both from one)

• Major- loss of three genes, severe anaemia

• Barts hydrops fetalis- loss of four genes, incompatible with life

Laboratory and clinical findings in alpha thalassaemia major

• Chronic haemolytic anaemia- hyperbilirubinaemia, LDH, reduced haptoglobin, jaundice

• Hepatomegaly, splenomegaly

• Higher rates of haemolysis seen when patiets exposed to infections, fever, drugs

• Low haemoglobin, high reticulocyte count

• Low MCV and MHC

• Microcytic hypochromic, mishapen cells, target cells

• HbH inclusions seen with brilliant cresyl blue stain

• Electrophoresis- Hb Bart’s in neonates, HbH in adults (HbA/F also seen)

Laboratory findings in alpha thalassaemia minor

• Mild microcytic hypochromic, haemoglobin normal or slightly decreased

• Low MCV, MCH

• High RCC, target cells

Pathophysiology of beta thalassaemia

• Excess alpha chains are unstable and precipitate in the cell

• Bind to membrane cause damage which reduces flexibility

• Ineffective erythropoiesis due to destruction of these erythrocytes by BM macrophages

• Chronic extravascular haemolysis due to removal of damaged cells by spleen macrophages

Thalassaemia major pathophysiology and symptoms

• Presents around 6 months when HbF levels fall

• Reduced or absent B cell production, increased a chain, precipitation of Hb, cells removed by macrophages (ineffective EP)

• Haemolytic anaemia and tissue hypoxia, increased haematopoiesis

• Haemolytic anaemia, hepatosplenomegaly, skeletal abnormalities, iron overload and deposition

Beta thalassaemia minor laboratory findings

• One gene on each chromatid, carrier or major

• Low MCV, MHC

• Microcytic hypochromic

• Normal haemoglobin, normal RCC or slightly increased

• Target cells

•Electrophoresis (diagnostic)- HbA2 above 3.5%

Laboratory findings in B thalassaemia major (Cooley’s)

• Severe anaemia- decreased MCH, MCV, MCHC

• Microcytic, hypochromic, target cells, teardrop cells

• Electrophoresis- HbA decreased, HbF increased HbA2 variable

Haemaglobinopathy therapy

• Blood transfusions- control severe anaemia, reduce risk of sickle cell complications, supress abnormal erythropoiesis

• Chronic transfusions can cause iron overload, desferrioxamine chelates iron

• Splenectomy to supress haemolysis

• Bone marrow transplant

• Gene therapy with crispr to correct faulty gene or activate production of fetal haemoglobin