Erythrocytes and Hemoglobin

Erythrocytes (Red Blood Cells)

• Erythrocytes are biconcave discs without a nucleus or organelles.
• Their diameters are larger than some capillaries.
• Erythrocytes are filled with hemoglobin (Hb) for gas transport
• They significantly contribute to blood viscosity
• The lifespan of an erythrocyte is 120 days.

Adult Haemoglobin (HbA)

• Haemoglobin contains four polypeptide subunits.
• Each HbA subunit features a globin chain with haem.
• Two alpha (α) and two beta (β) chains make up HbA (α2β2).
• Haemoglobin A has two alpha and two beta polypeptide chains and four haem groups
• Iron-containing haem pigment in the globin chains

Haemoglobin Assembly

• Alpha and beta-globin polypeptides are translated from their respective mRNAs.
• Upon haem binding, the protein folds into its native three-dimensional structure.
• The alpha globin chain is located on chromosome 16
• The beta globin chain is located on chromosome 11
• Normally, equal amounts of alpha and beta chains are produced (1:1).

Haemoglobinopathies

• These are common inherited diseases involving genetic defects of haemoglobin.
• Mutations or deletions in the α- or β-globin genes cause them.
• They fall into quantitative and qualitative defects.
• Gene mutations reduce the rate of synthesis of normal α- or β-globin chains, resulting in thalassemia as quantitative defects (reduced amounts); examples include α- and β-thalassaemia.
• Changes in Hb structure result from the synthesis of an abnormal haemoglobin with an altered amino acid sequence as qualitative defects; examples include HbS, HbE, and HbC.

Sickle Cell Disease

• First described in 1910 with abnormally elongated red blood cells in anaemic patients and links sickle cells to the clinical symptoms.
• Sickle cell anaemia is a structural variant of normal adult haemoglobin called HbS.
• HbS is the most common pathological haemoglobin variant worldwide, affecting about 5-7% of the global population.

Genetics of Sickle Cell

• A point mutation in sickle cell disease results in a base change from A to T.
• This mutation causes a substitution of glutamic acid to valine in the beta globin chain at the sixth position (6Glu→Val). causing it to become HbS.
• Inheritance of the affected gene from both parents results in sickle cell anaemia.
• Sickle cell is recessive
• Inheritance of HbS from only one parent results in a heterozygous child who can be an asymptomatic carrier - sickle cell trait

Sickle Cell Anaemia

• Under well-oxygenated conditions, HbS is fully oxygenated and soluble.
• When HbS is deoxygenated, it loses solubility and polymerises into rigid chains, distorting the red cell shape.
• Polymerisation of HbS is reversible, and sickle cells may revert to normal shape after re-oxygenation.
• After numerous oxygenation/deoxygenation cycles, red cells become irreversibly sickled.
• Sickle cells can block blood vessels and are eliminated in the liver and spleen.

Clinical Findings

• Sickle cells are found on blood film.
• Haemoglobin concentrations range from 60-90 g/L.
• Haemoglobin profile typically shows >80% HbS and 5-15% HbF.
• Other symptoms found are anaemia, dactylitis, necrosis and leg ulceration
• Chronic haemolysis causes excess haemoglobin molecule breakdown, leading to high bilirubin levels
• Splenic injury

Capillary Electrophoresis

• This technique separates haemoglobin types based on charge in an alkaline buffer.
• It is a screening test to identify normal and abnormal haemoglobins.
• Normal adult haemoglobin types include HbA (95-98%), HbA2 (2-3%), and HbF (0.8-2%).
• In sickle cell anaemia, the Hb S level is usually greater than 80%.

Treatment of Sickle Cell Anaemia

• Treatment aims to reduce the symptoms of sickle cell disease, associated pain, and complications.
• Hydroxyurea (hydroxycarbamide) increases HbF amounts but can decrease other blood cells and is not used during pregnancy.
• Folic acid increases red blood cells.
• Other treatments include blood transfusions, pain relief, and blood and bone marrow transplant.

Gene Therapy

• Casgevy (Exagamglogene autotemcel (exa-cel)) is a gene therapy approved for sickle cell disease and beta thalassaemia using the gene-editing tool CRISPR.
• Nonviral cell therapy designed to reactivate foetal haemoglobin synthesis.
• CRISPR-Cas9 gene editing of autologous CD34+ haematopoietic stem and progenitor cells (HSPCs) at the erythroid-specific enhancer region of BCL11A.
• The therapy "turns on" the production of HbF.
• Bone marrow stem cells are edited to express HbF before transplanting these edited stem cells back into the patient.