HEMOLYTIC ANEMIA:

OVERVIEW

Sri Mulatsih
Hematology-oncology, Pediatric
Department, Sardjito Hospital,
Faculty of Medicine, Gadjah Mada
University
2009
 OVERVIEW

Normal RBC life span is 110-120 days

 OVERVIEW

With age:
– Decreased membrane lipid component
– Spherocytic shape
– Less pliable and deformable
– Trapped and removed by the RE system
 CLINICAL PRESENTATION
Patients may present with a variety of
symptoms due to anemia

– Related to tissue hypoxia

• Fatigue, weakness, lightheadedness, CP, SOB

– Related to cardiovascular compensation

• Tachycardia, palpitations

– Related to underlying etiology

CLINICAL PRESENTATION

JAUNDICE

SCLERAL ICTERUS

SPLENOMEGALY
 LABORATORY
EVALUATION
Initial
• CBCP with MCV
• Review peripheral blood smear
• Reticuloctye count (retic. count)
• LDH
• Billirubin-direct and indirect
• Consider haptoglobin, urinalysis
 LABORATORY
EVALUATION
Additional / Directed studies
• Coombs’ test
• Underlying disorders
• Osmotic fragility test
• Hgb electrophoresis
• G-6PD assays
• Enzyme assays
• Bone marrow aspirate / biopsy
– Reversed M:E ratio
 DEFINITION
Hemolysis
Any situation resulting in accelerated
destruction of RBC’s (lifespan < 120 days)

– Anemia may or may not be present

Site of Hemolysis

Extravascular
vs.
Intravascular
 EXTRAVASCULAR
HEMOLYSIS
• 90% OF CASES

• Inappropriate removal of RBC’s by the RE

system

• Exaggeration of the normal removal of

RBC’s
EXTRAVASCULAR
HEMOLYSIS
 INTRAVASCULAR
HEMOLYSIS
• 10% of all cases

• Red cells are disrupted in the general

circulation

• Free hemoglobin is released into the plasma

INTRAVASCULAR
HEMOLYSIS
 INTRAVASCULAR
HEMOLYSIS
By products of intravascular hemolysis
include:

Hemoglobinemia
Hemoglobinuria
Hemosiderinuria
Low haptoglobin
 HEMOLYSIS

Most hemolytic anemia’s have an element of

both extravascular and intravascular
destruction
Ex. sickle cell anemia
MECHANISM OF
HEMOLYSIS

INTRINSIC
VS.
EXTRINSIC
 INTRINSIC
• Reason for hemolysis is intrinsic to the
RBC itself:

– membrane disorders
– metabolic disorders
– hemoglobin variants
 Membrane Disorders:
Hereditary Spherocytosis
Most common cause of hereditary hemolytic
anemia in Caucasians

• 1/5000 incidence
• Heterogeneous molecular basis
• Most commonly AD inheritance
• Deficiency of membrane proteins
– Spectrin, ankyrin, band 3
 Membrane Disorders:
Hereditary Spherocytosis
 Membrane Disorders:
Hereditary Spherocytosis

• Often the hemolysis is well compensated

• Very diverse clinical presentation

• History of gallstones is common

• Splenectomy often very effective

 Membrane Disorders:
Hereditary Spherocytosis
 Hereditary Spherocytosis
• Common

• Diverse clinical presentation

– Sometimes diagnosed in adulthood

• Diverse molecular defects

• Spherocytes, osmotic fragility test

 Paroxysmal Nocturnal
Hemoglobinuria (PNH)
• Acquired stem cell disorder:

– Anemia, leukopenia, thrombocytopenia

– Hypocellular bone marrow
– Intravascular hemolysis
– Risk for thrombosis
 PNH
• Defect leads to the lack of a phosphatidylinositol-
glycan (PIG) anchor protein

– Lack of multiple related surface proteins which depend

on the missing anchoring protein

– Make rbc more prone to lysis by complement

• Chronic intravascular hemolysis
 PNH
• Previously diagnosed by:
– Ham’s test
– Sucrose hemolysis test

• Now use flow cytometry to detect cells

missing various cell surface antigens
/proteins
– CD-59
 INTRNSIC
• Reason for hemolysis is intrinsic to the
RBC itself:

– membrane disorders
– metabolic disorders
– hemoglobin variants
 INTRNSIC
• Mature RBC requires glucose metabolism for energy
requirements (ATP):
– Maintenance of protein integrity, cell deformability, shape
– Preservation of heme iron in ferrous form
– Modulation of oxygen affinity of hemoglobin

• Main players
– Glycolytic pathway-90% of RBC ATP
– HMP shunt- generates reduced glutathione
– Nucleotide salvage pathways
 Metabolic Disorders
• Due to defects in:

– Glycolytic pathway
• Pyruvate kinase deficiency most common
– Purine metabolism
– Pyrimidine metabolism
– Hexose-monophosphate shunt
 Metabolic Disorders:
G-6PD Deficiency
• HMP shunt functions to maintain
glutathione in a reduced state
• Generation of NADPH

– Essential co-factor for detoxifying H2O2

– Essential for ability of RBC to withstand
oxidant stress
– Failure leads to oxidation and damage to
hemoglobin, enzymes, and membrane
 Metabolic Disorders:
G-6PD Deficiency
• G-6PD catalyzes the first step in the HMP
shunt

• X-linked recessive inheritance

• Most common metabolic disorder of RBC’s

– Esp. in African American males (10-15%)
 Metabolic Disorders:
G-6PD Deficiency
• Hallmark is acute hemolysis following
exposure to a variety of oxidant drugs or
stressors
– Antimalarial -Acidoisis
– Sulfonamides -Infection
– Nitrofurantoin -fava bean
– Vitamin K
 Metabolic Disorders:
G-6PD Deficiency
• Suspect when bite cells seen on peripheral smear
in presence of acute hemolytic anemia

• Heinz body staining is often positive

• More definitive tests available

• Don’t send during acute crisis

 INTRINSIC
• Reason for hemolysis is intrinsic to the
RBC itself:

– membrane disorders
– metabolic disorders
– hemoglobin variants
 Hemoglobin variants
Numerous inherited abnormalities in the
hemoglobin molecule or synthesis make the
RBC prone to premature hemolysis

• Sickle cell syndromes

• Thallassemias
• Unstable hemoglobins
 INTRINSIC
• Reason for hemolysis is intrinsic to the
RBC itself:

– membrane disorders
– metabolic disorders
– hemoglobin variants
 EXTRINSIC
– Reason for hemolysis has nothing to do
with intrinsic properties of RBC itself
• Immune mediated

• Red cell fragmentation

• Infection
 EXTRINSIC:
IMMUNE MEDIATED
• Shortened RBC survival due to antibody or complement
coating the red cell surface

– Autoimmune
• Warm
• Cold agglutinin disease

– Drugs
• Multiple mechanisms
AUTOIMMUNE HEMOLYTIC
ANEMIA (AIHA)
• Typically either IgG or IgM / complement coat
the RBC surface:

– Recognized as abnormal by RE system

– Leads to hemolysis (IgG)

– Agglutination (IgM / C3)

AUTOIMMUNE HEMOLYTIC
ANEMIA (AIHA)

Spherocytes (IgG)or agglutination (IgM)

typically noted on the peripheral blood
smear
 AIHA
Warm Autoimmune hemolytic anemia
• RBC’s coated with IgG
• Reactive at room/warm temperatures
• Acute and severe
• Younger age group
• Fix complement poorly
• Spleen is primary site of destruction
• Spherocytes seen on blood smear
• Idiopathic, collagen vascular, lymphoproliferative
disorders
 AIHA
Warm Autoantibody hemolytic anemia
• Treatment is primarily steroids
• Splenectomy is often second line

• Other immunosuppressants are second/third

line
 AIHA
Cold agglutinin disease
• RBC’s coated with IgM which fixes complement
• Reactive at cold temperatures
• Older age group
• Intravascular site of destruction
• Agglutination noted on blood smear unless
warmed
• Post infectious, lymphoma, idiopathic
 AIHA
Cold agglutinin disease
• Treatment includes avoidance of cold

• Usually need to try catatonic therapy if

persistent

• Plasmapheresis in refractory cases

 AIHA
Coombs’ test
• Critical in the evaluation of AIHA

– Anti-globulin reagent prepared by injecting human globulin

into animals-antibody isolated

• IgG
• C3

– Mixed with patient rbc or serum and test for clotting in the
test tube
 Coombs’ Results
Examples
• DAT: warm antibody
– IgG 3+ (strong)
– C3 negative or weak

• DAT cold agglutinin

– IgG negative /weak
– C3 3+
 Extrinsic
• Reason for hemolysis has nothing to do
with the RBC itself
– Immune mediated

– Red cell fragmentation

– Infection
 RBC Fragmentation
Microangiopathic Anemias

• Schistocytes and microspherocytes noted on the blood

smear

• High shear stress leads to intravascular hemolysis

– Microvascular disease
• DIC, TTP etc.
– Heart valve
– Trauma / implanted devices
 RBC Fragmentation
Microvascular disease
DIC-microvascular thrombosis due to activation and consumption of
platelets, clotting factors

– RBC’s hemolyzed as they flow through the micro-environment of

the damaged vasculature

– Almost always secondary to underlying catastrophe

– Present with bleeding or clotting

– Treat underlying disease with blood product or anticoagulant

support
 RBC Fragmentation
Microvascular disease

• Thrombotic Thrombocytopenic Purpura (TTP)

– Microangiopathic hemolytic anemia
– Thrombocytopenia
– Mental status changes
– Renal failure
– Fevers

• Treatment is plasma exchange procedures

 Extrinsic
– Reason for hemolysis has nothing to do
with the RBC itself
• Immune mediated

• Red cell fragmentation

• Infection
 Extrinsic
Infection
• Clostridia sepsis
• Parasites—malaria
• Viruses
• Bacterial
 SUMMARY
Location of defect:

Intrinsic vs. Extrinsic

Membrane Hb Metabolic Immune Infectious Mechanical

 SUMMARY
Location of hemolysis:

Extravascular
vs.
Intravascular
 SUMMARY
• Initial evaluation and labs similar:
– CBC, MCV, Retic count, review blood smear
– LDH, billirubin, haptoglobin, UA

• Further workup and treatment are specific to the suspected

cause of hemolysis
– Ex. DAT, electrophoresis, osmotic fragility test

• BM usually not necessary, unless you need to rule out and

underlying lymphoproliferative or primary BM disorder
– Reversed or decreased M:E ratio
 Cases
A woman in her thirties has a long history of SLE. She has had many
complications of her disease. She now presents with worsening dyspnea,
shortness of breath, and chest pain. Cbc shows a hgb of 7.3 gm/dl,
hematocrit of 24%, normal wbc and platelets. You suspect AIHA. Which
of the following lab data would not support this diagnosis?

a. Macrocytosis / elevated MCV

b. Elevated indirect billirubin
c. Positive Direct Coombs test
d. Elevated LDH
e. Low reticulocyte count
 Cases
A previously healthy 34 year-old African American female was treated
with bactrim (trimethoprin/sulfamethoxazole) for a UTI. She presents 4
days later with extreme fatigue, lethargy and weakness. Lab evaluation
reveals a marked hemolytic anemia. You suspect G-6PD deficiency and
request to review the peripheral blood smear. Which of the following
findings would most support your diagnosis?

a. bite cells
b. schistocytes
c. spherocytes
d. sickle cells
e. target cells
 Cases
A previously well 34 year old male presents with fever, mental status
changes, thrombocytopenia, hemolytic anemia, and renal failure. You
suspect TTP, with a microangiopathic intravascular hemolytic anemia as
the underlying diagnosis. All of the following could be expected on
laboratory evaluation except:

a. low serum haptoglobin

b. schistocytes on peripheral blood smear
c. positive direct coombs’ test
d. hemoglobinuria
e. elevated LDH
 Cases
A 45 year-old male is referred for evaluation of a hemolytic anemia. He
has no obvious underlying infection, malignancy or vasculitis. You
suspect he has a AIHA. You order a direct Coombs’ test and are told it is
strongly positive with IgG, but negative with complement at room
temperature. This data:

a.)supports the diagnosis of cold agglutinin disease

b.)supports the diagnosis of warm autoimmune hemolytic anemia
c.)supports both diagnosis
d.)supports neither diagnosis as no underlying disease is identified