Thalassaemia: Pathogenesis and Lab Diagnosis

1. Thalassaemia:Pathogenesis and Lab Diagnosis Dr. M Sadequel Islam TalukderMBBS, M Phil (Pathology), MACPAssistant ProfessorDepartment of PathologyDinajpur Medical College Presentation at Seminar on observation of World Thalassaemia Day 2009 Presentation: 10 May 2009, Dinajpur Medical College

2. Pathogenesis Genetically determined Heterogenous group of disorder Reduced synthesis of one or more types of normal haemoglobin polypeptide chain Reduced haemoglobin involving affected chain

3. Normal Haemoglobin • HbA - α2β2 • HbA2 - α2δ2 • HbF – α2γ2

4. Each goblin chain have separate genetic control α –thalassaemia affect α-chain synthesis β –thalassaemia affect β-chain synthesis

5. β-Thalassaemia β Chain synthesis An absence or deficiency of β-chain synthesis of adult HbA Hb-A γ and δ chain Hb-A = α2β2

6. On the basis of synthetic ability β-genes are designated as • β gene – can synthesize normal amount of β-chain • β+ gene – can synthesize reduced amount of β-chain • β0 gene – cannot synthesize β-chain

7. β-thalassaemia major • Mutation of normal β-gene β0-gene  absence HbA  increased HA2 and HbF • genotype – β0β0 • β-thalassaemia intermedia • ↑HbA2 • ↑HbF • ↓HbA • Genotype β+β+ or β0β • β-thalassaemia minor • ↑HbA2 • HbA normal • HbF normal

8. Pathophysiology of β-Thalassaemia Various mutation in β-gene Complete or partial absence of β-chain Decreased adult HbA α-chain synthesis remain normal Free complementary α-chain – unstable and precipitate within normoblasts as insoluble inclusions Cell membrane damage & impaired DNA synthesis apoptosis i.e. ineffective erythropoeisis

9. 70-80% marrow normoblasts undergo apoptosis Inclusion bearing red cells undergo sequestration & destruction in spleen

10. Partial or lack of HbA synthesis ↓MCHC & MCH Hypochromia & microcytosis Normal Thalassaemia

11. Reticulocytes undergo intramedullary death Inadequate production + ineffective erythropoiesis + haemolysis Anaemia

12. ↑Haemolysis ↑demands of phagocytic function  hyperplasia of phagocytes Hepatosplenomegaly To compensate anaemia extramedullary haemopoiesis in liver, spleen & brain Organomegaly

13. ↑Erythropoiesismarrow expansion & thinning of cortex of skull bone Thalassaemia facies

14. α-Thassaemia An absence or deficiency of α-chain synthesis due to delation of α-genes.

15. Pathogenesis of α-Thalassaemia • In normal individual HbA, HbA2 and HbF need α-chain for their formation. • 4 genes of α-chain, each pair on short arm of chromosome 16 present with genotype α,α/α,α. • In α-thalassaemia, delation of α-genes reduction or absence of synthesis of α-chain depending on number of α-gene delation.

16. ↓α-chain synthesis free γ-chain in the fetus & β-chain in infant of 6 months, and continue in the rest of life. • Complementary 4γ and 4β are aggregated  Hb Bart (4γ ) and HbH (4β ), respectively.

17. Variants of α-Thalassaemia • Silent carrier • Delation of single α-gene • Genotype α/αα • Asymptomatic • Absence of RBC abnormality • Thalasaemia trait • Delation of 2 α-genes • Genotype --/αα • Asymptometic, minimal or no anaemia • Minimal RBC abnormalities

18. Hb H disease • Delation of 3 α-genes • Genotype --/- α • 75% reduction of α-chain • 25% α-chain synthesis small amount of HbF, HbA, & HbA2 • Fetus can survive • Severe anaemia • Severe RBC abnormalites • Hydrops fetalis • Delation of all α-genes • Genotype --/-- • Absence of α-chain synthsis • Only Hb Bart (γ4) is produced (High affinity for O2 and can not dissociate O2 to tissue)

19. Inheritance

22. Laboratory Diagnosis of Thalassaemia

24. Laboratory Findings • Hb concentration – Decreased • ESR – Mild increased • WBC – Neutrophilic leucocytosis or normal • RBC count – Markedly decreased • PCV – Markedly decreased • MCV, MCH, MCHC – reduced • Reticulocyte count – Increased • Platelet count – May be increased

25. Peripheral Blood Film Normal

26. RBC Marked anisocytosis, polikilocytosis, microcytosis, frequent target cells, basophilic stippling, fragmented cells or schistocytosis, polychromatic macrocytes and nucleated cells WBC Whin normal or neutrophilic leucocytosis Platelets Normal or increased Morphology of PBF

27. Thalassaemia slides

28. Haemoglobin Electrophoresis • Different types of haemoglobins contain different surface charge which determine the elecrtrophoretic mobility and gives a specific bands on the electrophoretic papers when haemolysate is undergone eletrophoresis. • HbF, HbA, HbA2, HbC, HbE, HbD, HbS, HbH and Hb Bart can be measured accurately by electrophoresis

31. Thanks