hemolytic anemia (cell membrane defect)
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Hemolytic Anemia
Dr. Rafi Ahmed GhoriFCPS
Professor MedicineLiaquat University of Medical & Health Sciences, Jamshoro
Classification of Hemolytic Anemias
1. Abnormalities of RBC interior1. Enzyme defects.2. Hemoglobinopathies
2. RBC membrane abnormalities1. Hereditary spherocytosis etc.2. Paroxysmal nocturnal hemoglobinuria.3. Spur cell anemia.
3. Extrinsic factor1. Hypersplenism.2. Antibody: immune hemolysis.3. Microangiopathic hemolysis.4. Infections, toxins, etc.
Intracorpuscular
Extracorpuscular
Hereditary
Acquired
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Red Cell Membrane Disorders
Usually detected by morphologic abnormalities of RBC on blood film.
Three types of inherited RBC membrane abnormalities– Hereditary sphercytosis.– Hereditary elliptocytosis (including hereditary
pyropoikilocytosis).– Hereditary stomatocytosis.
Red cell membrane has a lipid bilayer to which a ‘skeleton’ of filamentous proteins is attached via special linkage proteins.
RED CELL CYTOSKELETON
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
SPLENIC ARCHITECTURE
HEREDITARY SPHEROCYTOSISOsmotic Fragility
0
20
40
60
80
100
0.3 0.4 0.5 0.6
NaCl (% of normal saline)
% H
emo
lysi
s
Normal HS
Hereditary Spherocytosis
Usually has an autosomal dominant inheritance pattern and incidence of approximately 1:1000 to 1:4500.
In ~20% patients, the absence of hematologic abnormalities in family members suggests either autosomal recessive inheritance or spontaneous mutation.
Hereditary Spherocytosis
Sometimes clinically apparent in early infancy but often escapes detection until adult life.
Hereditary SpherocytosisClinical Manifestations
– Anemia.– Splenomegaly.– Jaundice.
Hereditary SpherocytosisClinical Manifestations
– Prominent jaundice accounts for disorder’s prior designation as “congenital hemolytic jaundice” and is due to an increased concentration of unconjugated (indirect-reacting) bilirubin in plasma.
Hereditary SpherocytosisClinical Manifestations
– Jaundice may be intermittent and tends to be less pronounced in early childhood.
– Due to increased bile pigment production, pigmented gallstones are common, even in childhood.
– Clinical course may be complicated by crisisHemolytic crisis (infection).Megaloblastic crisis (folate deficiency, especially in
pregnancy).Aplastic crisis (parvovirus infection).
Hereditary SpherocytosisDiagnosis
– Must be distinguished primarily from the spherocytic hemolytic anemias associated with RBC antibodies.
– If present, family history of anemia and/or splenectomy.
– Immune spherocytosis positive direct Coombs test.
Hereditary SpherocytosisDiagnosis
– Splenomegaly can also be seenCirrhosis.Clostridial infections.G6PD deficiency.
Hereditary Spherocytosis
Treatment– Splenectomy reliably corrects the anemia.– Splenectomy in children should be postponed
until the age of 4-years.– Polyvalent pneumococcal vaccine should be
administered at least 2-weeks before splenectomy.
– In patients with severe hemolysis folic acid (5-mg/d) should be administered prophylactically.
Hereditary SpherocytosisTreatment
– Acute, severe hemolytic crises requires transfusion.
Hereditary ElliptocytosisHeterogeneous disorders in elliptocytic red
cells.Functional abnormality of anchor proteins
in red cell membrane (alpha spectrin or protein 4.1).
Autosomal dominant or recessive.Less common than hereditary spherocytosis
(1/10,000).
Red cell membrane has a lipid bilayer to which a ‘skeleton’ of filamentous proteins is attached via special linkage proteins.
Hereditary ElliptocytosisClinical presentation depends upon degree
of membrane defect.Asymptomatic diagnosed on blood film.Chronic compensated hemolytic state.
Hereditary ElliptocytosisTreatment
– Same as hereditary spherocytosis only in chronic hemolytic anemia.
Differential diagnosis includes– Iron deficiency anemia– Thalassaemia.– Mylofibrosis.– Mylodysplasia.– Pyruvate kinase deficiency.
Paroxysmal Nocturnal Hemoglobinuria
Clonal cell disorder Ongoing Intra- & Extravascular hemolysis; classically
at night Testing
– Acid hemolysis (Ham test)– Sucrose hemolysis– CD-59 negative (Product of PIG-A gene)
Acquired deficit of GPI-Associated proteins (including Decay Activating Factor)
GPI BRIDGE
Paroxysmal Nocturnal Hemoglobinuria
GPI Proteins
GPI links a series of proteins to outer leaf of cell membrane via phosphatidyl inositol bridge, with membrane anchor via diacylglycerol bridge
PIG-A gene, on X-chromosome, codes for synthesis of this bridge; multiple defects known to cause lack of this bridge
Absence of decay accelerating factor leads to failure to inactivate complement & thereby to increased cell lysis
HEMOLYTIC ANEMIAMembrane abnormalities -
Enzymopathies
Deficiencies in Hexose Monophosphate Shunt– Glucose 6-Phosphate Dehydrogenase Deficiency
Deficiencies in the EM Pathway– Pyruvate Kinase Deficiency
G6PD DEFICIENCYFunction of G6PD
G6PD
GSSG 2 GSH
NADPH NADP
2 H2O H2O2
6-PG G6P
Hgb
Sulf-Hgb
Heinz bodies
Hemolysis
InfectionsDrugs
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
Glucose 6-Phosphate DehydrogenaseDifferent Isozymes
0
0.2
0.4
0.6
0.8
1
0 20 40 60 80 100 120
RBC Age (Days)
G6P
D A
cti
vit
y (
%)
Normal (GdB) Black Variant (GdA-)
Mediterranean (Gd Med)
Level needed for protection vs ordinary oxidative stress
HEMOLYTIC ANEMIACauses
INTRACORPUSCULAR HEMOLYSIS– Membrane Abnormalities– Metabolic Abnormalities– Hemoglobinopathies
EXTRACORPUSCULAR HEMOLYSIS– Nonimmune – Immune
EXTRACORPUSCULAR HEMOLYSIS
Nonimmune
Mechanical Infectious Chemical Thermal Osmotic
Microangiopathic Hemolytic AnemiaCauses
Vascular abnormalities– Thrombotic thrombocytopenic purpura– Renal lesions
Malignant hypertension Glomerulonephritis Preeclampsia Transplant rejection
– Vasculitis Polyarteritis nodosa Rocky mountain spotted fever Wegener’s granulomatosis
Microangiopathic Hemolytic Anemia
Causes - #2
– Vascular abnormalities AV Fistula Cavernous hemangioma
Intravascular coagulation predominant– Abruptio placentae– Disseminated intravascular coagulation
IMMUNE HEMOLYTIC ANEMIAGeneral Principles
All require antigen-antibody reactions Types of reactions dependent on:
– Class of Antibody– Number & Spacing of antigenic sites on cell– Availability of complement– Environmental Temperature– Functional status of reticuloendothelial system
Manifestations– Intravascular hemolysis– Extravascular hemolysis
IMMUNE HEMOLYTIC ANEMIAGeneral Principles - 2
Antibodies combine with RBC, & either1. Activate complement cascade, &/or2. Opsonize RBC for immune system
If 1, if all of complement cascade is fixed to red cell, intravascular cell lysis occurs
If 2, &/or if complement is only partially fixed, macrophages recognize Fc receptor of Ig &/or C3b of complement & phagocytize RBC, causing extravascular RBC destruction
IMMUNE HEMOLYTIC ANEMIA
Coombs Test - Direct Looks for immunoglobulin &/or complement of
surface of red blood cell (normally neither found on RBC surface)
Coombs reagent - combination of anti-human immunoglobulin & anti-human complement
Mixed with patient’s red cells; if immunoglobulin or complement are on surface, Coombs reagent will link cells together and cause agglutination of RBCs
IMMUNE HEMOLYTIC ANEMIA
Coombs Test - Indirect
Looks for anti-red blood cell antibodies in the patient’s serum, using a panel of red cells with known surface antigens
Combine patient’s serum with cells from a panel of RBC’s with known antigens
Add Coombs’ reagent to this mixture If anti-RBC antigens are in serum, agglutination
occurs
HEMOLYTIC ANEMIA - IMMUNE
Drug-Related Hemolysis Alloimmune Hemolysis
– Hemolytic Transfusion Reaction – Hemolytic Disease of the Newborn
Autoimmune Hemolysis– Warm autoimmune hemolysis– Cold autoimmune hemolysis
IMMUNE HEMOLYSISDrug-Related
Immune Complex Mechanism– Quinidine, Quinine, Isoniazid
“Haptenic” Immune Mechanism– Penicillins, Cephalosporins
True Autoimmune Mechanism– Methyldopa, L-DOPA, Procaineamide, Ibuprofen
DRUG-INDUCED HEMOLYSIS
Immune Complex Mechanism
Drug & antibody bind in the plasma Immune complexes either
– Activate complement in the plasma, or– Sit on red blood cell
Antigen-antibody complex recognized by RE system Red cells lysed as “innocent bystander” of destruction
of immune complex REQUIRES DRUG IN SYSTEM
DRUG-INDUCED HEMOLYSIS
Haptenic Mechanism
Drug binds to & reacts with red cell surface proteins
Antibodies recognize altered protein, ± drug, as foreign
Antibodies bind to altered protein & initiate process leading to hemolysis
DRUG-INDUCED HEMOLYSIS
True Autoantibody Formation
Certain drugs appear to cause antibodies that react with antigens normally found on RBC surface, and do so even in the absence of the drug
ALLOIMUNE HEMOLYSISHemolytic Transfusion Reaction
Caused by recognition of foreign antigens on transfused blood cells
Several types– Immediate Intravascular Hemolysis (Minutes) - Due to preformed
antibodies; life-threatening– Slow extravascular hemolysis (Days) - Usually due to repeat
exposure to a foreign antigen to which there was a previous exposure; usually only mild symptoms
– Delayed sensitization - (Weeks) - Usually due to 1st exposure to foreign antigen; asymptomatic
INCOMPATIBLE RBC TRANSFUSIONRate of Hemolysis
0
20
40
60
80
100
0 1 2 3 4 5 6 7Weeks Post-Transfusion
Su
rviv
ing
Ce
lls
(%
)
Normal Immediate Intravascular Hemolysis
Slow Extravascular Hemolysis Delayed Extravascular Hemolysis
ALLOIMMUNE HEMOLYSISTesting Pre-transfusion
ABO & Rh Type of both donor & recipient Antibody Screen of Donor & Recipient,
including indirect Coombs Major cross-match by same procedure
(recipient serum & donor red cells)
ALLOIMMUNE HEMOLYSISHemolytic Disease of the Newborn
Due to incompatibility between mother negative for an antigen & fetus/father positive for that antigen. Rh incompatibility, ABO incompatibility most common causes
Usually occurs with 2nd or later pregnancies Requires maternal IgG antibodies vs. RBC antigens
in fetus
ALLOIMMUNE HEMOLYSISHemolytic Disease of the Newborn -
#2
Can cause severe anemia in fetus, with erythroblastosis and heart failure
Hyperbilirubinemia can lead to severe brain damage (kernicterus) if not promptly treated
HDN due to Rh incompatibility can be almost totally prevented by administration of anti-Rh D to Rh negative mothers after each pregnancy
AUTOIMMUNE HEMOLYSIS
Due to formation of autoantibodies that attack patient’s own RBC’s
Type characterized by ability of autoantibodies to fix complement & site of RBC destruction
Often associated with either lymphoproliferative disease or collagen vascular disease
AUTOIMMUNE HEMOLYSISWarm Type
Usually IgG antibodies Fix complement only to level of C3,if at all Immunoglobulin binding occurs at all temps Fc receptors/C3b recognized by macrophages; Hemolysis primarily extravascular 70% associated with other illnesses Responsive to steroids/splenectomy
AUTOIMMUNE HEMOLYSISCold Type
Most commonly IgM mediated Antibodies bind best at 30º or lower Fix entire complement cascade Leads to formation of membrane attack complex,
which leads to RBC lysis in vasculature Typically only complement found on cells 90% associated with other illnesses Poorly responsive to steroids, splenectomy;
responsive to plasmapheresis
HEMOLYTIC ANEMIASummary
Myriad causes of increased RBC destruction Marrow function usually normal Often requires extra folic acid to maintain
hematopoiesis Anything that turns off the bone marrow can
result in acute, life-threatening anemia