THALASSEMIA
F.A. Rice, ART, CLS
March 1, 1996
Please send comments to: F.A. Rice
Outline:
Thalassemia was first described by Cooley and Lee in 1925. Most
of their patients, and those reported by others later, were of
Mediteranian ancestry. Hence this group of anemias are also known
as:
- Cooley's Anemia
- Mediteranian Anemia
Thalassemia is a genetically determined defect in hemoglobin synthesis. There is an inability to manufacture sufficient quantities of globin chains.
In the the adult there are three hemoglobin types normally present:
- Hgb A 2 alpha & 2 beta chains - 95% of total
- Hgb A2 2 alpha & 2 delta chains - 3% of total
- Hgb F 2 alpha & 2 gamma chains - 2% of total
During fetal life the majority of the hemoglobin is of the
Hgb F type.
During embryonic life at least two different hemoglobins are produced:
- Gowers 2 2 alpha & 2 epsilon chains
- Gowers 1 4 epsilon chains
The manufacture of each of these chains is controlled by
specific genes. During development from the embryo to the adult
these genes are switched on and off accounting for the different
hemoglobin types. In thalassemia there is a genetic failure in the
production of globin chains. Although a failure of production can
and does occur in the production of any of the five globin chain
types a failure of production of beta and alpha chains is the most
common. Hence the thalassemias are classified according to the type
of chain which is not produced. The most common being:
beta thalassemia - a failure of beta chain production
alpha thalassemia - a failure of alpha chain production
A further listing of hemoglobins, the characteristic
morphologies present and their identification can be seen here.
BETA THALASSEMIA
The genes controlling beta chain production are located on
chromosome 11. If both genes fail then the patient is said to have
beta thalassemia major. If only one gene fails then the patient has
beta thalassemia minor.
This is the most common of the thalassemias. Beta chain
production is less than normal due to the failure of one of the
genes coding for beta chains. Alpha chain production continues at a
near normal rate. The alpha chains combine with the available beta
chains resulting in decreased levels of hemoglobin A. There still
remains excess alpha chains and this stimulates the increased
production of delta chains. The alpha and delta chains combine to
form increased amounts of hemoglobin A2. If there is still an
excess of alpha chains the normal mechanism which switches off
gamma chain production does not function correctly and the rate of
gamma chain production is greater than in a normal adult. This
results in the formation of increased amounts of hemoglobin F.
Laboratory diagnosis of beta thalassemia minor
These patients are not severely anemic. The importance of
identifying heterzygous beta thalassemia is to prevent the
investigation and expense caused by confusion with iron deficiency.
As well these patients can then be provided appropriate genetic
counselling.
Hemoglobin, Hematocrit are decreased. The RBC count is not as
low as the hemoglobin and hematocrit, in fact it is usually normal.
This is due to the fact that the marrow can still produce the
cells but cannot fill them with hemoglobin. Hence the Hgb is low
and the empty cells occupy less space thus lowering the Hct
relative to the erythrocyte count.
Hemoglobin is very seldom, if ever, below 95 g/L. If the Hgb is
less than 93 g/L it is unlikely the patient has beta thalassemia
minor.
Morphology is microcytic and hypochromic. Anisocytosis is only
slight and not near as marked as in iron deficiency. This is
reflected in a near normal RDW and red cell histogram which is
shifted to the left but of near normal dimensions. There is slight
poik with an occasional target cell. A moderate basophilic
stippling is common.
Indecies show a characteristic discordance. The MCV is slightly
decreased and the MCH is decreased. The MCHC is normal. The RBC
number is usually normal.
The discriminant factor (DF) is a mathematic manipulation of
the indecies and generates a number which can be used to help
differentiate between thalassemia minor and iron deficiency. At
least three formulas are in use:
Mentzer: MCV
--- = if <13 then thal minor
RBC >13 then iron deficiency
Shine-Lal: (MCV)squared X MCH = if <1530 then thal minor
>1530 then iron deficiency
England-Frazer : MCV-(Hgb X 5)-RBC- 3.4 = if negative thal minor
positive iron def
The England-Frazer formula is the most often used.
White cell count and differential - normal
Retics - relative increase
PI decreased
Bone marrow - normal to slight erythroid hyperplasia.
Serum iron - normal
TIBC - normal
Ferritin - normal
Bilirubin - slight increase due to intramedullary hemolysis
FEP - normal
Hemoglobin studies:
Hgb A - decreased
Hgb A2 - increased
Hgb F - Sl increase to normal
These patients are well at birth but develop a life threatening
anemia by one or two months. They must be supported with blood
transfusions which result in iron overload. Unless the iron is
removed with appropriate chelation therapy these patients die of
hemosiderosis.
In beta thalassemia major there is a complete failure of beta
chain production. Hence there is very little, if any, Hgb A present. Delta and gamma chain production is increased. As a result there is
raised levels of Hgb A2 and Hgb F. Hemoglobin F has a very high
affinity for oxygen and is a poor oxygen deliverer. As a result the
only functional hemoglobin present is Hgb A2, hence the patient is
hypoxic which causes increased erythropoietin secretion. The excess
erythropoietin stimulates the marrow to the maximum, and ultimately
to the point that extramedullary hemopoiesis occurs with
splenomegaly. Even with in increased production of hemoglobins A2
and F there are still excess alpha chains and they precipitate in
the developing normoblasts. This results in intramedullary
hemolysis and their premature removal from the marrow by RE
cells.
Laboratory diagnosis of beta thalassemia major
Anemia is severe - Hgb 20 to 30 g/L. Hematocrit and RBC count
are also decreased hence the indecies are uniformly depressed.
The MCV, MCH and MCHC are all decreased.
The RDW is increased.
The RBC histogram is shifted to the right and abnormal in shape due
to the presence of NRBC and abnormal RBC shapes.
The morphology is severe hypochromic microcytic with marked
aniso and poik. Many target cells, schistocytes, leptocytes (large
thin flat cells). The presence of these cells often makes it
difficult to recognize the microcytosis. Moderate basophilic
stippling and many NRBC are common.
The WBC is invariably increased when first performed but is
normal after correction for NRBC. The differential is usually
normal.
Retic count - relative increase 5 to 10% but the PI is
decreased.
Bone marrow - erythroid hyperplasia with dyserythropoiesis.
Stainable iron is increased and a rare ringed sideroblast may be
seen.
FEP normal to increased
Serum iron - increased/normal
TIBC - decreased/normal
Ferritin - increased/normal
%saturation - increased
Hemoglobin electrophoresis -
Hgb A - decreased
Hgb F - increased
Hgb A2 - variable
ALPHA THALASSEMIA
There are four genes coding for alpha chain production. These
genes are located on chromosome 6. As a result there are at least
five forms of alpha thalassemia depending on the number and
location of the abnormal genes.
All genes are abnormal. There is no alpha chain production
hence no Hgb F production and death in utero.
Laboratory diagnosis of Hydrops Fetalis
At autopsy the cord blood shows severe anemia, less than 60g/L.
There is no Hgb A or Hgb F on electrophoresis, most of the
hemoglobin is hemoglobin Bart's which consists of 4 gamma
chains.
Three genes are abnormal and one gene is coding for alpha
chains. As a result there is limited production of Hgb F in utero
and Hgb A after birth. The excess gamma chains form Hgb Bart's and
the excess beta chains form Hgb H both of which are unstable and
precipitate in the cell resulting in the premature destruction in
the marrow and spleen with splenomegaly.
Laboratory diagnosis of Hemoglobin H Disease
The infant is anemic at birth with hemoglobin levels less than
90 g/L. The hematocrit and erythrocyte count are also decreased.
The Indecies are uniformly decreased with the MCV, MCH and MCHC
decreased. The RDW is increased. The RBC histogram is shifted to
the left as a result of the microcytes but the shape remains normal.
The morphology shows marked microcytosis and hypochromia.
Anisocytosis and poikilocytosis is moderate to marked. Basophilic
stippling is moderate due to the presence of precipitated Hgb H.
The may be an occasional NRBC but they are not as common as in beta
thalassemia major.
The retic count is only slightly raised with a decreased PI
The white cell count and differential are normal.
Hemoglobin H inclusion bodies can be demonstrated in a retic stain
Bone marrow shows erythroid hyperplasia.
Serum iron - increased
Ferritin - increased
TIBC - decreased
Hemoglobin electrophoresis -
Hgb Bart's increased at birth
Hgb Bart's 2 to 10% later
Hgb H 5 to 40%
Hgb A and A2 decreased
The clinical picture varies somewhat depending upon whether or
not the two deleted genes are on the same chromosome or not. In
alpha thalassemia 0 both genes are absent from the same chromosome.
In alpha thalassemia + one gene is missing from each chromosome.
The + form tends to be slightly more severe with slightly lower
hemoglobin levels.
Laboratory diagnosis of alpha thalassemia minor
In both forms however, there are only minor
changes with perhaps a mild anemia. The MCV and MCH are usually
boarder-line low with the MCH being depressed more often than the
MCV. The RDW tends to increase if an anemia develops as a result of
pregnancy etc. Hemoglobin electrophoresis is normal with increased
levels of Hgb Bart's if the cord blood is electrophoresed.
Only one of the four genes is abnormal. As a result there is a
near normal production of alpha chains with very few if any
clinical or laboratory changes.
Delta/Beta Thalassemia
The gene controlling delta chain production is located very
close to the beta gene on chromosome 11. If one gene is deleted
then the other may be affected.
Homozygous Delta/Beta Thalassemia is similar to beta
thalassemia, but symptoms are milder. Most patients survive to
adult life with minimal transfusion requirements.
Laboratory diagnosis of homozygous delta/beta thalassemia
Anemia - variable 40 to 100g/L
Morphology - hypochromic microcytic with marked anisocytosis and
poikilocytosis. Erythrocyte inclusions are common.
Hemoglobin electrophoresis shows 90 to 100% Hgb F.
Heterozygous Delta/Beta Thalassemia is similar to beta
thalassemia minor but symptoms tend to be less severe.
Laboratory diagnosis of heterozygous delta/beta thalassemia
Anemia - mild Morphology - hypochromic microcytic with slight
anisocytosis and poikilocytosis.
The indecies are discordant.
Hemoglobin electrophoresis -
Hgb A - decreased
Hgb F - 5 to 15%
Hgb A2 - normal
Hemoglobin Lepore
This is a mixup in the production of the non-alpha chain. The
carbon end is the amino acid sequence of the delta chain while the
N-terminal end is the amino acid sequence of the beta chain.
There are at least three variants depending on the length of
the mixed up segments:
1. Washington
2. Hollandia
3. Baltimore
Laboratory diagnosis of homozygous Hgb Lepore
Homozygous Hgb Lepore There is no normal beta or delta chain
production. The clinical and laboratory findings are identical to
Beta thalassemia major except on electrophoresis.
Hemoglobin electrophoresis:
Hgb F 80 to 90%
Hgb A absent
Hgb A2 absent
Hgb Lepore 10%
Hgb Lepore moves in the same position as Hgb S
Laboratory diagnosis of heterozygous Hgb Lepore
Heterozygous Hgb Lepore The clinical and laboratory findings
are identical to beta thalassemia minor except on electrophoresis.
Hemoglobin electrophoresis:
Hgb A decreased
Hgb A2 decreased
Hgb Lepore 10%
Hemoglobin E is not a thalassemia but is, rather a
hemoglobinopathy. Thalassemia is characterized by the inability to
produce sufficient numbers of globin chains. Hemoglobinopathies
are characterized by the production of abnormal globin chains.
Hemoglobin E is a beta chain variant in which lysine is
substituted for glutamic acid in position 26.
It is believed to be the most common hemoglobinopathy in the
world. Hemoglobin E occurs in Southeast Asia, especially
in Cambodia, Laos and Thailand. The importance of Hemoglobin E lies
in the fact that:
- Patients with hemoglobin E have microcytic blood pictures which may be confused with thalassemia or initial stages of iron deficiency.
- Patients who inherit both genes for Hemoglobin E and beta-thalassemia have severe anemia.
Laboratory diagnosis of Hemoglobin E
Patients who are homozygotes are mildly anemic - Hgb 90 to
100 g/L. Hematocrit and RBC count are also decreased hence the
indecies are uniformly depressed. The RDW is increased. The RBC
histogram is slightly shifted to the right. The morphology is
microcytic with anisocytosis and poikilocytosis with many target
cells. The presence of these cells may make it difficult to
recognize the microcytosis.
The WBC and differential is usually normal.
Retic count - slight increase 5 to 10%
Serum iron - increased/normal
TIBC - decreased/normal
Ferritin - increased/normal
%saturation - increased
Hemoglobin electrophoresis
- Hgb A - decreased
- Hgb E - increased
- Hgb A2 - variable
Patients who are heterozygotes are not anemic but have an
elevated RBC count and target cells are present.