Approach to hemolytic anemias

for pediatrics UG teaching

1. Approach to the Diagnosis of Hemolytic ANEMIAS Dr M.Sanjeevappa Designated Associate Professor Dept of Paediatrics GMC ,Anantapur

2. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Hemolytic anemias are characterised by increase red cell destruction. Characteristic features are : A shortened red cell life span below the normal 120 days Elevated erythropoietin level and compensatory increase in erythropoiesis. Accumulation of hemoglobin degradation products that are produced as a part of process of red cell hemolysis.

3. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS PATHOGENESIS OF RED CELL DESTRUCTION : Two mechanisms : Extravascular Hemolysis – The site of destruction is mainly spleen and this is the major mechanism of red cell hemolysis. Red cells are taken up by the cells of RE system where they are destroyed and digested. Intravascular Hemolysis –This is the minor pathway of red cell destruction and red cells are destroyed in circulation releasing hemoglobin.

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7. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS CLINICAL FEATURES : Clinical signs and symptoms of hemolytic anemia depends upon the severity as well as duration of hemolysis. Pallor Jaundice Splenomegaly Gall stones Skeletal abnormalities in severe hemolysis Leg ulcers Dyspnoea Tachycardia and systolic murmur

8. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Hereditary hemolytic anemias : Type Etiology Associations Diagnosis Treatment Enzymopathies G6PD deficiency Infections, drugs, ingestion of fava beans Low G6PD activity Withdrawal of offending drug, treatment of infection Membranopathies Hereditary Spherocytes Hereditary Elliptocytosis family history, spherocytosis negative DAT. Splenectomy Hemoglobino - pathies . Thalassemia sickle cell disease Hemoglobin electrophoresis, genetic studies Folate, transfusions

9. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Acquired hemolytic anemias : Type Etiology Associations Diagnosis Treatment Immune-mediated Antibodies to RBC surface antigens. Idiopathic Drugs Autoimmune Infections, Transfusions Spherocytes Positive DAT Treatment of underlying disorder Steroids Splenectomy IV gamma globulins Microangiopathic Mechanical disruption of RBC in circulation TTP ,HUS , Pre eclampsia Eclampsia Malignant HTN Prosthetic valves Schistocytes Treatment of underlying disorder. Infection Malaria Babesiosis Clostridium infections Cultures Thick and thin smears Serologies Antibiotics.

10. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS INVESTIGATIONS : Investigations that establish ACCELERATED HEMOLYSIS : Shortened lifespan of red blood cells by 51Cr labeling Accelerated erythropoiesis: a. Erythroid hyperplasia in the bone marrow. b. Reticulocytosis.

11. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Continued…. Increase in catabolic products of hemoglobin in blood : a. Unconjugated hyperbilirubinemia b. Hemoglobinemia (intravascular hemolysis) c. Hemoglobinuria (intravascular hemolysis) d. Hemosiderinuria (intravascular hemolysis) e. Increased urinary urobilinogen f. Increased CO excretion

12. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Continued…. 4 . Increased concentration of products released from the breakdown of RBCs particularly LDH. 5 . Effects due to release of hemoglobin a. Depletion of plasma haptoglobin b. Depletion of plasma hemopexin.

13. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS CBC : Acute hemolytic episode : neutrophilia, thrombocytosis normoblastemia Chronic hemolysis : pancytopenia.

14. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS SPECIFIC TESTS TO ESTABLISH THE CAUSE OF HEMOLYSIS : PERIPHERAL BLOOD SMEAR FINDINGS: The most important investigation in hemolytic anemias. Morphological findings suggestive of hemolysis : Polychromatophilia, nucleated red cells, thrombocytosis and neutrophilia with mild shift to left. Spherocytes, Sickle cell, Target cells, Schistocytes (fragmented red cells, helmet cells, traingular cells) and acanthocytes

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16. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Bone marrow findings- Compensatory mechanism to hemolysis : Erythroid hyperplasia of bone marrow : Erythroid hyperplasia with normoblastic reaction. Reversal of M:E ratio Reticulocytosis : - Mild (2-10%)- Hemoglobinopathies - Moderate to marked (10-60%) Immune hemolytic anemias, Hereditary spherocytosis , G6PD deficient states

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18. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Tests for RBC membrane abnormality: i . Osmotic fragility, incubation osmotic fragility. ii. Identification of specific membrane abnormality

19. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS Identification of abnormal hemoglobin : Electrophoresis, isoelectrofocussing (IEF). Hb A2 by microcolumn chromatography Hb F by alkali denaturation Intracellular Hb F distribution Quantitation of alpha and beta-chains by HPLC or electrophoresis Hb S by sickling test, solubility test . Tests for unstable hemoglobins DNA analysis for detection of mutations. Screening tests or quantitative analysis for enzymopathies

20. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS HEREDITARY SPHEROCYTOSIS Hereditary spheroytosis is an inherited hemolytic anemia resultimg from red cell mebrane defect leading to microspherocytosis , splenomegaly and jaundice ETIOAPTHOGENESIS : Spectrin deficiency is the most common abnormality Mutation of b spectrin gene and point mutations affect the binding of spectrin to protein 4.1

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22. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS HEREDITARY SPHEROCYTOSIS - CLINICAL FEATURES Seen all over the world Autosomal dominant with variable penetrance M=F present in neonate, childhood or adulthood Intermittent jaundice is usual presentation O/E- splenomegaly is a constant feature Gall stones (pigment type) Chronic leg ulcers (rare)

23. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS HEREDITARY SPHEROCYTOSIS - LAB FINDINGS : PBF Findings- Microspherocytes which are small dense rbc without pallor MCV- Normal Reticulocytes- Increased Bone marrow- Erythroid hyperplasia with normoblastic reaction S. bilrubin - Increased (unconjugated ) U. bilrubin – Increased Fecal stercobilinogen - increased S. haptoglobins- Reduced

24. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS HEREDITARY SPHEROCYTOSIS - OTHER DIAGNOSTIC TESTS Osmotic fragility test- shift of curve to right Incubated osmotic fragiloty test Glycerol lysis test – Increased (rate of lysis)

25. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS GLUCOSE 6-PHOSPHATE DEHYDROGENASE DEFICIENCY Glucose6-phosphate dehydrogenase is the first enzyme in the hexose monophosphate shunt pathway (HMP) which protects red cells from oxidant injury Deficiency of G6PD may result in episodes of hemolysis following certain drug intake or chemical exposure or infection G6PD deficiency is a sex linked disease. Its prevalance is higher in tropical eastern countries. Prevalance is higher in kurdish jews (60-70%) and lower in japan (.1%)

26. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS CLINICAL AND HEMATOLOGICAL PRESENTATION OF G6PD DEFICIENCY : Acute hemolytic anemia - Occurs following exposure to drugs like primaquine, infections like pneumonia, typhoid and oxidative chemicals. Chronic non-spherocytic anemia - There is moderately severe enyme deficiency, hemolysis continues throughout life. Neonatal hyperbilrubinimia - Jaundice in G6PD deficient neonates, CF- Jaundice, kernicterus Favism- Common in children, caused by consumption of fava beans. Glucoside divisine and aglycone isouramil is responsible. Resulting in acute severe hemolysis within few hours . CF-headache, fever, chills and back pain.

27. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS G6PD DEFICIENCY -DIAGNOSTIC TESTS Peripheral blood film evaluation :Moderate anisopoikliocytosis with polychromatophilia, Microspherocytes and bite cell. Reticulocytosis (20-50%) Hemogobinuria and increase urobilinogen in urine Quantitative G6-PD assay and DNA analysis by PCR

28. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS G6PD DEFICIENCY -DIAGNOSTIC TESTS BLISTER CELLS BITE CELLS HEINZ BODIES

29. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS THE THALASSEMIAS Thalassemia syndrome are autosomal recessive disorders Thalassemia results from defects in the rate of synthesis of a or b chains, lead to reduced hemoglobin production and accumulation of a or b chains

30. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS SICKLE CELL DISORDERS: Sickling syndromes are characterized by the presence of HbS which imparts sickle shape to red cells in a state of reduced oxygen tension HbS is prevalent in Africa, Mediterranean countries and India. In India, seen common in tribals and in ethnic groups of MP, Orissa, AP, Maharashtra ( vidharba region), TN ( chetti tribes) and Kerala There is high prevalence of HbS in areas endemic to falciparum malaria.

31. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS SICKLE CELL DISORDERS - GENETICS Sickle mutation is caused by substitution of valine in place of glutamic acid in the 6th position of b-chain. Mutation results in clinical presentation 1. Sickle cell anemia - HbS-HbS , Homozygous state 2. Sickle cell trait - HbA-HbS , heterozygous state 3. Sickle cell disease- Refer to all diseases with HbS in combination with – normal ( HbA ), abnormal gene of b-thalassemia, a-thalassemia, HbD , HbE , HbC,HbQ

32. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS SICKLE CELL DISORDERS -Pathophysiology of vascular occlusion and hemolysis Polymerisation of deoxygenated HbS is the primary event in the pathogenesis of the disease Red cell containing HbS pass through microcirculation of spleen – various cycles of sickling and desickling – Irreversible sickeled RBCs – Extravascular hemolysis in spleen – Vascular stasis – vascular occlusion – splenic infarcts – hyposplenism (lead to infection) and autosplenectomy

33. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS SICKLE CELL DISORDERS - CLINICAL FEATURES Delay in puberty, growth and development Recurrent leg ulcers Avascular necrosis of femur head Dactylitis ( Hand –Foot syndrome ) Pneumonia, meningitis, Osteomylitis Jaundice and liver enlargement Pigment gall stones Acute abdominal pain ( infarcts of abdominal viscera) Priapism Acute chest syndrome (fever, chest pain, leucocytosis, appearance of pulmonary infilterate with sickle anemia ) Sickle retinopathy- Salmon patches- intra retinal hemmorhages

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35. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS CRISIS IN SICKLING SYNDROME Sickling crisis ( vaso -occlusive crisis) Hemolytic crisis Aplastic crisis Sequestration crisis

36. APPROACH TO THE DIAGNOSIS OF HEMOLYTIC ANEMIAS SICKLE CELL DISORDERS - HEMATOLOGICAL FINDINGS Anemia - moderately severe anemia with Hb 5- 10 gm PBF : Red cells- Sickle cells, target cells, ovalocytes, polychromtophila with nucleted RBCs. Howell-jolly bodies also seen, TLC- Mildly elevated , Platlets - Increased. Reticulocytosis- 3%-10% Bone marrow- Erythroid hyperplasia with normoblastic reaction. Sickling tests- Presence of HbS demostrated by using reducing agent like 2% sodium metabisulphite . Hb electrophoresis

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