2. Course title: Hematology (1) Course code: MLHE-201 Supervisor: Prof. Dr Magda Sultan Outcome : The student will know : -The types of hemolytic anemias -The diagnosis of hemolytic anemias -The types of hereditary hemolytic anemias. -The diagnosis of hereditary hemolytic anemias -The laboratory tests needed for diagnosis

3. 1-Hemolytic anemia(HA) HA = decreased levels of red blood cells in circulating blood (anemia) because of their accelerated destruction (hemolysis)

4. A red blood cell survives 120 days The spleen (part of the reticulo-endothelial system) is the main organ which removes old and damaged RBCs from the circulation. Breakdown of RBCs can exceed the rate that the body can make RBCs and so anemia can develop.

5. HEMOLYTIC ANEMIA Anemia of increased destruction Normochromic, normocytic anemia Shortened RBC survival Reticulocytosis - Response to increased RBC destruction Increased indirect bilirubin Increased LDH

6. Classification of hemolytic anemias Causes of hemolytic anemias can be either: 1 - hereditary (genetic ) 2 - acquired.

7. Hemolytic anemia Hereditary  Defects of hemoglobin  Thalassaemia, Sickle cell anemia  Defects of the red cell membrane  Hereditary spherocytosis, Hereditary elliptocytosis  Defective red cell metabolism (enzymes)  G6PD deficiency. P K deficiency Physiologic classification of anemia

8. Acquired  Immune mediated : Autoimmune, isoimmune, drugs  Microangiopathic: DIC, HUS  Hypersplenism  Miscellaneous: drugs, toxin, infection, burn, chemical

9. Mechanisms of hemolysis: - intravascular - extravascular

10. Intravascular hemolysis : Red cell destruction occurs in vascular space e.g eclinical states associated with Intravascular hemol acute hemolytic transfusion reactions severe burns physical trauma bacterial infections

11. Intravascular hemolysis : - laboratory signs of intravascular hemolysis: tests for hemolysis and aditionally: hemoglobinemia methemalbuminemia hemoglobinuria hemosiderinuria

12. Extravascular hemolysis :red cells destruction occurs in reticuloendothelial system e.g scyrestem - clinical states associated with extravascular hemolysis : autoimmune hemolysis delayed hemolytic transfusion reactions hemoglobinopathies hereditary spherocytosis hypersplenism hemolysis with liver disease - laboratory signs of extravascular hemolysis: tests for hemolysis

13. Signs of hemolytic anemia: Physical Symptoms of anemia Jaundice Pallor Splenomegaly / hepatosplenomegaly

14. Laboratory features (1) * Morphology: anemia *Peripheral blood smear microscopy: **fragments of the red blood cells ("schistocytes") can be present **some red blood cells may appear smaller and rounder than usual (spherocytes) **reticulocytes are present in elevated numbers. Normoblasts can be present.

15. Bone marrow smear microscopy: * Erytrhroid hyperplasia * The level of unconjugated bilirubin in the blood is elevated. * The level of lactate dehydrogenase (LDH) in the blood is elevated

16. Laboratory features (2) * The direct Coombs test is positive, if hemolysis is caused by an immune process. *Increased excretion of urobilinogen in the urine *Increased stercobilinogen in the stool. *Sometimes abnormal results of the osmotic fragility test

17. Hereditary Hemolytic anemia  Defects of hemoglobin  Thalassaemia, Sickle cell anemia  Defects of the red cell membrane  Hereditary spherocytosis, Hereditary elliptocytosis  Defective red cell metabolism  G6PD deficiency. P K deficiency

18. .  The abnormalities of the gene may result from substitution of single amino acid like sickle cell anaemia or decrease synthesis of the whole globin chain (thalassaemia) Haemoglobinopathy

19. Sickle cell anaemia  Substitution of glutamic acid by valine The Hb is stable when oxygenated state and become unstable and polymerized on deoxygenated state

20.  Polymerization will lead to precipitation of Hb. The cell become deformed (sickle shape) and very sticky leading to vascular occlusion and small infarction to the affected areas.  Short life span of cells leading to chronic anaemia, Sickle cell anaemia

21. The origin of the disease is a small change in the protein hemoglobin The change in cell structure arises from a change in the structure of hemoglobin. A single change in an amino acid causes hemoglobin to aggregate.

23. Diagnosis Laboratory: Hb 6-9 g/dl, high retics (5-15%), Normocytic anemia, target or anisocytosis Sickling test Hb electrophoresis Hb S 60-100 in SS Parents sickle trait Hb AS Sickle cell anaemia

24. Hgb Electrophoresis Amino acids in globin chains have different charges Separates hemoglobin according to charge 90% Hgb S, 10% Hgb F, small fraction of Hgb A2

27. Means defective synthesis of one or more of the globin chains which form normal hemoglobin. In globin chains which form normal hemoglobin. In very severe form this globin is totally absent. The defect may be in alpha chain (  thalassaemia), defect may be in alpha chain (  thalassaemia), Beta chain (  thalassaemia) or Delta chain (  Beta chain (  thalassaemia) or Delta chain ( thalassaemia) Thalassaemia

28. HEMOGLOBIN NORMAL ADULT RBC CONSISTS OF 3 FORMS OF Hb: - HbA - 2 α and 2 β globin chains - HbA2 – 2 α and 2 δ globin chains - HbF - 2 α and 2 γ globin chains THALASSEMIAS α and β

29. Different forms of thalassemia Alfa thalassemia Beta thalasemia: major, minor (trait), intermedia Delta/Beta thalassemia Hereditary persistentce of fetal hemoglobin (HPFH)

30. Beta Thalassaemia Defective  chain synthesis Excess  chain Precipitation cell membrane damage Circulating Red cell Anaemia Bone marrow expansion skeletal changes & hyper metabolism Erythropoietin increased Ineffective erythropoiesis Bone marrow blood transfusion Iron overload Complication and death Hemolytic  Iron absorption

31. Beta-Thalassemia major laboratory features Severe anemia Blood film: anisopoikilocytosis, hypochromia, target cells, basophylic stippling, reticulocytes moderately increased and normoblasts. Marrow: marked erythroid hyperplasia, increased sideroblasts Shortened red cell survival Haemoglobin electrophoresis : Fetal hemoglobin > 90%, HbA absent, HbA2 low/normal/high

32. HEREDITARY SPHEROCYTOSIS Defective or absent spectrin molecule Leads to loss of RBC membrane, leading to spherocytosis Decreased deformability of cell Increased osmotic fragility Extravascular hemolysis in spleen

33. Hereditary spherocytosis (HS) Laboratory features - hemolytic anemia - blood smear-microspherocytes - abnormal osmotic fragility test, acidified glycerol lysis time - negative direct Coombs test

34. G6PD DEFICIENCY Function of G6PD

35. Glucose 6-Phosphate Dehydrogenase Functions Regenerates NADPH, allowing regeneration of glutathione Protects against oxidative stress Lack of G6PD leads to hemolysis during oxidative stress Infection Medications Fava beans Oxidative stress leads to Heinz body formation,  extravascular hemolysis

36. G6PD DEFICIENCY DIAGNOSIS : QUANTITATIVE ASSAY DETECTING LOW ENZYME TREATMENT – SUPPORTIVE AND PREVENTATIVE

37. Assignement : Student name : الشيماء مصطفي عبد العاطي Title:Sickle cell anaemia Student name : اميره اسعد يوسف Title : Haemoglobin electrophoresis. Student name : اميره صلاح مرشدي Title :G6PD deficiency. Student name : انجي عبد الموجود Title : Spherocytic anaemia. Student name : بسمه مهدي رياض Title :Laboratory tests of hemolytic anaemias.

38. Training questions : What are the tests of hemolysis ? How to diagnose Sickle cell anaemia ? How to classify hemolytic anaemias ? Reference book : Essential Hematology. Dacie.