HEMOLYTIC ANEMIADR SANDIP GUPTA

PGT,PEDIATRICS

B.S.M.C.H.

CLASSIFICATION

1.RED CELL ABNORMALITY(INTRACORPUSCULAR DEFECT)A.HEREDITARY1.Membrane defect(spherocytosis,ellipsocytosis)2.Enzyme defect3.Hemoglobinopathies(Thalassemia,SCD, other)

B.ACQUIRED1.Paroxysmal nocturnal hemoglobinuria

2.EXTRA CORPUSCULAR DEFECT

A.IMMUNE HEMOLYTIC ANEMIA1.AUTOIMMUNE HEMOLYTIC ANEMIAWarm antibodiesCold anti bodies2.ALLOIMMMUNE HEMOLYTIC ANEMIAHemolytic disease of newbornIncompatible blood transfusion B.NONIMMUNE HEMOLYTIC ANEMIAMicroangiopathic hemolytic anemias(DIC ,TTP,HUS)Trauma:prosthetic cardiac valveBurns,ECMO,snake biteInfection: malaria, babesiaChemical injury :lead,wilson disease,Hypersplenism

EVALUATION OF ANEMIALow Hgb/Hct

Low Hgb/Hct

Corr. Retic Ct >2%

Corr. Retic Ct >2%

Corr. Retic Ct <2%

Corr. Retic Ct <2%

Acute Blood Loss Acute Blood Loss MCV>100

MCV>100MCV 80-100

MCV 80-100

MCV<80

MCV<80

EVALUATE & TREAT APPRO-

PRIATELY

EVALUATE & TREAT APPRO-

PRIATELY

Evaluate for

Hemolytic Anemias

Evaluate for

Hemolytic Anemias

Evaluate for

microcytic anemias

Evaluate for

microcytic anemias

Evaluate for

macrocytic anemias

Evaluate for

macrocytic anemias

Evaluate for

normocytic anemias

Evaluate for

normocytic anemias

NOYES

STEP BY STEP APPROACH

1. Calculate for Corrected Reticulocyte CountRetic count: 10%Pt’s Hct 29Control Hct 40

Corrected Retic Count = % Retic x Pt’s HctControl Hct

= 10% x 29/ 40 = 7.73 % > 2% if no blood loss Indicates hemolysis

HEMOLYTIC ANEMIA

Premature destruction of RBCs

2. Confirm (+) hemolysis:

a) Corrected retic count > 2% b) Inc indirect bilirubins c) Inc LDH d) Low/absent haptoglobin

3. Look for cause of hemolysis- occult blood in urine, urine hemosiderin- peripheral blood smear- direct antiglobulin test, Hgb electrophoresis,

RBC enzyme analysis

ROLE OF PBS

1 Sickled cells

Bite cells

Schisto-cytes

Acantho-cytes

Sphero-cytes

Target cells

parasiteinclusions

DAT(+)

DAT(-)

Hgb electro-phoresis

G6PDlevel

PT/PTTCrea

platelets

Auto-ImmuneHemo-lytic

Anemia

Heredi-tary

Sphero-cytosis

Sickle CellDs

G6PDDeficient

VsUnstable

Hgbs

Thalas-semiasHemo-

globino-pathy

Liver Ds

LiverDs

MalariaBabe-siosisBarto-nella

TTP-HUSDIC

Prosthe-tic Valve

MalignantHTN

Hemolytic Anemia (CRC>2% + no blood loss)

HEMOLYTIC ANEMIAIntravascular Causes in Red

WITHIN THE RED CELL

1. Membrane defects

- HS

- HE

- Hereditary pyropoikilocytosis

- Hereditary stomatocytosis

2. Enzyme defects

-G6PD

-Pyruvate kinase

3. Hemoglobin defects

- SCA

- Thalassemias

- Unstable hemoglobin

NON-IMMUNE

1. Hypersplenism

2. Fragmentation syndromes - grafts / valves / AS - HTN / Pre-eclampsia - March hemoglobinuria - MAHA - TTP/HUS - DIC - hemangioma

2. Infections/Toxins (Malaria, Babeosis, Bartonella, Clostridium welchii, snakes, spiders)

3. Drugs

4. Liver dz (Spur cell)

5. PNH

AUTO-IMMUNE

1. Warm

2. Cold

3. Transfusion reactions

4. Drug associated

OUTSIDE THE RED CELL

Intravascular Hemolysis

RBC LYSIS

HBG

HAPTOGLOBIN

REMOVED BY LIVER

HEMOGLOBINEMIA

HEMOGLOBINURIA

HBG TAKEN UP BY RENAL TUBULAR CELLS

HEMOSIDERIN

CELLS SLOUGHED IN

URINE 1 WEEK LATER

Features specific to intravascular haemolysis:

• Haemoglobinaemia(haptoglobin and

haemopexin exhausted).• Methaemoglobinaemia.• Haemoglobinuria.• Haemosiderinuria.

HEREDITARY SPHEROCYTOSIS

Incidence:1/5000 in North European population

Autosomal dominant Defect in RBC

cytoskeleton(spectrin,ankyrin) Pathophysiology:A deficiency in

spectrin, ankyrin,protein 3, leads to weakening of the “vertical” interaction of the lipid bilayer & loss of membrane microvescicle . Loss of surface area,↑cation permeability, ATP use,& glycolysis leading to premature destruction in spleen.

CLINICAL FEATURES Neonatal period: anemia+ jaundice, more severe. Infancy&childhood:variable severity. Mild: asymptomatic Moderate: intermittent

jaundice,spleenomegaly,anemia. Severe:tranfusion dependeant,bone expansion,gall

stone LAB. DIAGNOSIS:Anemia(Hb:6-10g

%),PBS :Spherocytes lacking central pallor ,reticulocytes,MCV-N

MCH↑,MCHC >35,RDW>14.5,DCT: NegativeOsmatic fragility& Incubated osmotic fragility test.Differential diagnosis: autoimmune hemolytic anemia, G6PD def,Clostridial sepsis, wilsons disease.

TREATMENT Splenectomy was routine in past. Anemia,reticulocytosis, hyperbilirubinemia resolve. Transfusion requirement↓,risk of gall stone falls. Current approach is to spenectomize pts with severe hemolytic

anemia &those with s/s of anemia, growth failure, skeletal changes, leg ulcer, etramedullary hematopoiesis,

aplastic crises,cardiomegaly . Pt’s with Hb>10%& retic count<10% may not need splenectomy. Partial splenectomy in infants & young children with severe

hemolysis & transfusion dependent anemia has been recently advocated.

Suppoertive therapy : 1mg folic acid daily. Laparoscopic splenectomy. Lifelong antibiotic prophylaxis.

2.HEREDITARY ELLIPTOCYTOSIS

Equatorial Africa, SE Asia AD / AR Functional abnormality in one or more

anchor proteins in RBC membrane- Alpha & beta spectrin& defective spectrin heterodimer self association , Protein 4.1& glycophorinC.

Usually asymptomatic Mx: Similar to H. spherocytosis Variant:

3.SE-Asian ovalocytosis: Common in Malaysia , Indonesia… Asymptomatic-usually Cells oval , rigid ,resist invasion by malarial parasites SAO is associated with protein3 abnormality.

RED CELL ENZYMOPATHIES

Physiology: EM pathway: ATP production HMP shunt pathway: NADPH & Glutathione production

1. Glucose-6-Phosphate Dehydrogenase

( G6PD ) Deficiency Pivotal enzyme in HMP Shunt & produces NADPH to protect RBC

against oxidative stress Most common enzymopathy -10% world’s population 1% of indian males have G6PD deficiency Protection against Malaria X-linked recessive

Clinical Features: Acute drug induced hemolysis:

Aspirin, primaquine, quinine, chloroquine, dapsone…. Chronic compensated hemolysis Infection/acute illness Neonatal jaundice Favism

(Oxidised form)(Reduced form)

Inv:e/o non-spherocytic intravascular

hemolyisP. Smear: Bite cells, blister cells,

irregular small cells, Heinz bodies, polychromasia

G-6-PD level

Treatment: Stop the precipitating drug or treat

the infectionAcute transfusions if required

2. Pyruvate Kinase DeficiencyARDeficient ATP production, Chronic

hemolytic anemiaInv;

P. Smear: Prickle cellsDecreased enzyme activity

Treatment: Transfusion may be required

PAROXYSMAL NOCTURNAL HEMOGLOBINURIA

Clonal cell disorder Ongoing Intra- & Extravascular

hemolysis; classically at night Testing

Acid hemolysis (Ham test)Sucrose hemolysisCD-59 negative (Product of PIG-A gene)

Acquired deficit of GPI-Associated proteins (including Decay Activating Factor)

1.Warm AI Hemolysis: Can occurs at all age groups F > M Causes:

50% IdiopathicRest - secondary causes:

1.Lymphoid neoplasm: CLL, Lymphoma, Myeloma

2.Solid Tumors: Lung, Colon, Kidney, Ovary, Thymoma

3.CTD: SLE,RA4.Drugs: Alpha methyl DOPA, Penicillin , Quinine, Chloroquine

5.Misc: UC, HIV

Inv: e/o hemolysis, MCV P Smear: Microspherocytosis, n-RBC Confirmation: Coomb’s Test / Antiglobulin test

Treatment Correct the underlying cause Prednisolone 1mg/kg po until Hb reaches 10mg/dl then

taper slowly and stop Transfusion: for life threatening problems If no response to steroids Spleenectomy or, Immunosuppressive: Azathioprine, Cyclophosphamide

2. Cold AI Hemolysis

Usually Ig MAcute or Chronic formChronic:

C/F:Elderly patients Cold , painful & often blue fingers, toes, ears, or nose ( Acrocyanosis)

Inv: e/o hemolysis P Smear: Microspherocytosis Ig M

Other causes of Cold Agglutination: Infection: Mycoplasma pneumonia, Infec

Mononucleosis PCH : Rare cause seen in children in association with

viral infection. Demonstrable DONATH LANDSTEINER ANTIBODY

Treatment: Treatment of the underlying cause Keep extremities warm Steroids treatment Blood transfusion

NON-IMMUNE ACQUIRED HEMOLYTIC ANEMIA1. Mechanical Trauma

A). Mechanical heart valves, Arterial grafts: cause shear stress damage

B).March hemoglobinuria: Red cell damage in capillaries of feet

C). Thermal injury: burnsD). Microangiopathic hemolytic anemia (MAHA): by

passage of RBC through fibrin strands deposited in small vessels disruption of RBC eg: DIC,PIH, Malignant HTN,TTP,HUS

ACQUIRED HEMOLYSIS

2.InfectionF. malaria: intravascular hemolysis: severe called ‘Blackwater fever’Cl. perfringens septicemia

3.Chemical/Drugs: oxidant denaturation of hemoglobinEg: Dapsone, sulphasalazine, Arsenic gas, Cu, Nitrates & Nitrobenzene

THALASSEMIAS/ THALASSEMIA SYNDROME Epidimiology :

– Most Common genetic disorder in Pediatric ward– 7% of the world population is carriers of hemoglobin disorder– 1.5% of world population is carriers of ß Thalassemia gene (20 millions in India alone)– 8 to 10 thousand children born

in India with homozygous state for the Thalassemia in every year.

– There are around 65 to 67 thousand Thalassemia patients in our country.

– In India, Prevalence of defective ß gene varies from 1 to 17 %.

Hallmark of Thalassemia is decreased or absent synthesis of Globin chains of Hemoglobin i.e. it is quantitative disorder of Hb Synthesis.

Based on the chain affected Thalassemias are classified as α and ß Thalassemia.

If ß gene is absent, it is term as ß0 Thalassemia. If partially affected, it is called ß+ Thalassemia.

The genetic classification does not necessarily define the phenotype and the degree of Anemia does not always predict the genetic classification.

Thus for the management, the Thalassemias are classified into four groups, each for α & ß depending on clinical severity.

SALIENT FEATURES

α THALASSEMIA SYNDROMES :

Syndrome Clinical Features Hemoglobin Pattern

α-globin genes affected and genotype

Silent carrier No Anemia, normal red cells

1-2% Hb Bart’s(γ4) at birth 1- α/ αα

Thalassemia Trait Mild anemia, hypochromic

5-10 % Hb Bart’s(γ4) at birth, microcytic red cells

2- α/ -α, --/ αα

HbH Disease moderate anemia, Hepatosplenomegaly, malar prominence etc.

5-30 % HbH (ß4) red cells 20-30% Hb Bart’s(γ4) at birth

3--/ -α

Hydrops Fetalis/Hb Bart’s Syndrome

Severe anemia, Hepatosplenomegaly, Cardiac defect, Genito-Urinary Systems abnormality, PET in motherDeath in Utero

Mainly Hb Bart’s 90 %, small amount of HbH, gower 1, gower 2 and portland

4--/--

CLASSIFICATION, CLINICAL & HEMATOLOGICAL FEATURES OF ẞ THALASSEMIA :

Syndrome Clinical Features Hemoglobin Pattern

ß-globin genes affected and genotype

Heterozygous State–Silent Carrier

–Thalassemia trait

No Anemia, normal

Mild anemia, hypochromic, microcytic red cellsHb > 10 gm% RBC > 5.5 x 1012 per liter

Normal, HbF < 5%

Elevated HbA2 (3.6-8 %)

1ß+ / A

1 ß0 / A, ß+ / A

Homozygous State–Thalassemia Intermedia

–Thalassemia Major or Cooley’s Anemia

Moderate anemia, requires some transfusion Hb > 7-10 gm% RBC < 5.5 x 1012 per liter

Severe anemia, transfusion dependentHb < 7 gm% RBC < 4 x 1012 per liter

HbF elevated(20 -100 %)HbA2 < 3.5 %

HbF elevated (90%)HbA2 = 2% HbE = 30-40%

2ß+ / ß+

2ß0 / ß0, ß0 / ß+,

E / ß0

PRINCIPLES OF MANAGEMENTConfirmation of the Diagnosis

By HPLC Diagnose of Complication

Correction of Anemia – Packed Red Blood Cell (PRBC) transfusion

Management of Complications– Iron Overload and Chelation Therapy– Anemia/ Hypoxia

– Arrest of Growth– Infections– Hypersplenism

Pharmacological Methods – Increase gamma chain Synthesis (HbF)

Curative Treatment– Stem cell transplantation

Future Treatment– Gene Replacement therapy

Prevention of Disease

DIAGNOSIS OF COMPLICATION: It is not sufficient to diagnose the case as Thalassemia ONLY.

For complete management of the case, it is necessary to think about its genetic classification, clinical and patho-physiological stage in which it now belongs to.

THALASSEMIA

SpenomegalySkeletal Deformity

&Arrest of Growth

Iron Overload&

Chelation Therapy

Anaemia

Recurrent Blood BormeInfection

DEATH

Why need a Transfusion? Correct Anemia &prevention of hypoxia Reduce Hepatosplenomegaly &Hypersplenism

Reducing ineffective erythropoesis& GI absorption Reduce hemolytic facies& skeletal deformities. Improve growth

BT is Mandatory For All children with Thalassemia Major Thalassemia Intermedia , Hb < 7 gm % Evidence of growth retardation

Types of Transfusion Palliative(8.5g%) Hyper Transfusion(10g%) Super Transfusion(>12g%) Moderate Transfusion(9-10.5g%)

TRANSFUSION THERAPY IN THALASSEMIA

TRANSFUSION THERAPY (CONTD…)

Frequency of Transfusion– Every 3-4 weeks– Shorter interval of 2-3 weeks is more physiological – avg. time taken 3-4 hrs(@5mi/kg/hr).

Amount of Transfusion– 180 ml / kg. / yr in non spenctomised, non-sensitised pt. – 130 ml / kg /yr in spenctomised, sensitised pt. ( 30 % less)

Efficacy of Transfusion– Rate of fall of Hb should not exceed 1 gm / dl /week with spleen– Rate of fall of Hb should not exceed 1.5 gm / dl /week without spleen

Allo immunisation of RBC Hyperspenism Drag induced hemolysis Infection

TRANSFUSION THERAPY (CONTD…)

Adequacy of Transfusion– First decade : normal growth– No. of Normoblast < 5 / 100 WBC

Complication of Transfusion– Non hemolytic febrile Transfusion reaction NHFTR– Allo Immunisation– Plasma Borne Infection– Steps to prevent those infections– Allergic reactionINDICATION OF SPLENECTOMY Annual PRBCc>200-250ml/kg >1.5times basal requirement Massive spleenomegaly hypersplenism

IRON OVERLOAD Causes of Iron Overload

– Treatment with multiple transfusion One bottle blood increases iron store by 200 – 250 mg iron.

– Ineffective erythropoesis– Excessive dietary absorption of Iron

Consequence of Iron overload– Iron overload in Liver

Hepatomegaly, Fibrosis & cirrhosis– Iron overload in Spleen

Splenomegaly, Hypersplenisim– Cardiac complication

Failure & Arrhythmia– Endocrinal Dysfunction

Thyroid, Para-Thyroid, Pituitary, Pancreas, Gonads– Iron overload in Bones

Osteoporosis, Osteopenia

IRON CHELATION THERAPY Iron Chelation Therapy

– Goal Reduce the Iron store & sub sequently maintain it at low level ( < 1000 µg/ml )

– When to start Start after 15-20 transfusion or S.Ferritin > 1000 µg/ml (approx. 3 yrs of age) Needle Biopsy of Liver : 3.2Mg iron per gm of Liver tissue (

– Drugs presently used Inj. Deferrioxamine (SC/IV) : DFO/Desferal Oral Deferiprone Oral Deferasirox

– Newer Iron Chelator Desferrithiocin ( DFT) Hydroxy Benzyl Ethilene Diamine Diacetic acid (HBED) Pyridoxal iso nicotinyl Hydrazone (PIH) GT 56-252 40 SD02 (CHF 1540)

IRON CHELATION THERAPY (CONTD…) Inj. Deferrioxamine (SC/IV) : DFO/Desferal

Dose <2000ug/l→25mg/kg/d,2000-3000ug/kg/d →35mg/kg/d Mode of Delivery s/c:over 8-10 hrs as 10% sol Dipot DFO is more effective & latest. Recently I.V. is used in severe cardiac involvement. Toxicity/ adverse effect

Local reaction Visual abnormality ( 4-10 % of pt.) Sensori-neural hearing loss ( 4-38 % of pt.) Delayed linear growth Pulmonary Infarction

Auditory & Visual Toxicity is reversible Yersinia sp. Infection Vit-c in a dose of 50-200mg/d

CURATIVE TREATMENT

Stem Cell Transplantation This is the only curative therapy available today. Though expensive, it is cost effective as compared

to yearly cost of regular BT & chelation therapy Sources

Bone Marrow Cord Blood Fetal Liver Peripheral Blood

FUTURE TREATMENT Gene Therapy

Aim : Insertion of a normal copy of gene along with key regulatory sequences(LOCUS CONTROL

REGION) in the stem cells of recipients.

Two main approaches Somatic gene therapy in which non-

germ line cells are involved. Transgenic approach

in which transfuse gene can be expressed in subsequent generationsNeed high titre vectors for

sustained expression Lentiviral vector from HIV is a

hope.

SCREENING & PREVENTION

Premarital screening programmes Alternative is to screen pregnant woman in

early pregnancy. PRENATAL DIAGNOSIS: BY CVS AT 9-11WK Recently there has been attempt to isolate

fetal cells from maternal blood. PARENTERAL COUNSELLING