Transcript thalassemia occurs when one or more of the 4 alpha chain genes

Sickle cell anemia
and thalassemias
Paul R. Earl
Facultad de Ciencias Biológicas
Universidad Autónoma de Nuevo León
San Nicolás, NL, Mexico
[email protected]
Genetic explanations.
Thalassemia is caused by
impaired production of either
the  or  hemoglobin chain.
Alpha () thalassemia occurs
when one or more of the 4 alpha
chain genes fails to function.
A) The loss of one gene
diminishes the production of the
alpha protein only slightly. This
condition is so close to normal
that it can be detected only by
specialized laboratory techniques.
A person with this condition is
called a silent carrier because of
the difficulty in detection.
B) The loss of 2 genes (2-gene
deletion alpha () thalassemia)
produces a condition with small
RBCs, and at most a mild anemia.
People with this condition look and
feel normal. The condition can be
detected by routine blood testing,
however.
C) The loss of 3 alpha genes produces a
serious hematological problem (3-gene
deletion  thalassemia). Patients with
this condition have a severe anemia,
and often require blood transfusions to
survive. The severe imbalance between
the alpha chain production (now
powered by one gene, instead of 4) and
beta chain production (which is normal)
causes an accumulation of beta chains
inside the RBCs. Normally, beta chains
pair only with alpha chains.
D) The loss of all 4 alpha genes
during fetal life causes death in
utero or shortly after birth.
Rarely, 4 gene deletion alpha
thalassemia has been detected in
utero, usually in a family where
the disorder occured in an earlier
child. Repeated transfusions can
keep victims alive.
E) There are 2 genes for the beta
chain of hemoglobin in thalassemia. Unlike -thalassemia,
-thalassemia rarely arises from the
complete loss of a beta globin gene
that is present, but produces little
beta globin protein. The types of
genes can be analyzed in each
case.
Even when the affected gene produces
no beta chain, the condition is mild
since one of the 2 beta genes functions
normally. The red cells are small and a
mild anemia may exist. People with the
condition generally have no symptoms.
The condition can be detected by a
routine laboratory blood evaluation.
(Note that in many ways, the one-gene
beta thalassemia and the two-gene
alpha thalassemia are very similar, from
a clinical point of view. Each results in
small red cells and a mild anemia).
The sickle cell disorders usually
result from S mutation
homozygocity which is an A for
T oligonucleotide substitution
at codon 6 of the beta globin
gene leading to the
nonsynchronous replacement
of valine for glutamic acid.
Clinical classification
of the thalassemias.
Alpha thalassemia has 4
manifesta-tions (a-d) according to
the number of defective genes.
a) Silent carrier state. This is the onegene deletion alpha thalassemia
condition. People with this condition
are hematologically normal. They are
detected only by sophisticated
laboratory methods.
-Thalessia has 3 states (a-c).
a) Thalassemia minor (known as
thalassemia trait) in people who have
small red cells and mild or even no
anemia. These patients are usually only
detected through routine blood testing.
b) Thalassemia intermedia in people
with anemia able to survive without
blood transfusions.
c) Thalassemia major patients require
chronic transfusions.
Carrier detection.
The carrier detection procedure of a
preventive program should be designed
to be precise enough to secure all
couples at risk.
Heterozygous -thalassemia, either the
0 or + type, is characterized by high
RBC counts, microcytosis, hypochromia,
increased hemoglobin A2 (HbA2) levels
and unbalanced -globin/non--globin
chain synthesis. However, this
phenotype can be modified, causing
problems in carrier identification.
Heterozygous -thalassemia:
Phenotypic modifications
Normal RBC indices  and  interactions
Normal HbA2 level
Iron deficiency
Coinheritance of  and 
thalassemias
Some mild -thalassemia
mutations -thalassemia
Normal RBC indices and HbA2
Silent -thalassemia mutations
-Globin gene triplication
The preliminary selection of individuals
at risk of being heterozygous for a form
of thalassemia is based on the
determination of mean corpuscular
volume (MCV) and mean corpuscular
hemoglobin (MCH) values. However,
double heterozygotes for both  and 
anemias could have normal MCV and
MCH values, and thus could be missed.
Quantitaion of the HbA2 level should
also be tested for.
Molecular diagnosis.
To date at least 150 molecular
defects have been defined in
-thalassemias. The common
polymerase chain reaction
(PCR) procedures used are
given in the table:
Reserve oligonucleotide hybridization uses
membrane-bound allele specific probes
that hybridize to the complementary PCR
sequence prepared by using the patient’s
DNA as the starting template. Up to 20-30
mutations have been screened in one step!
Primer-specific amplification of the target
DNA can detect mutants. Only the normal
primer amplifies normal DNA,while DNA
from homozygotes is amplified only by the
-thalassemia primer and DNA from
heterozygotes by both primers.
-Globin gene analysis.
This analysis is carried out to
define double heterozygotes
for  and  thalassemias with
normal HbA2 that can be
confused with -thalassemia.
Sickle cell anemia.
It commonly results from
homozygosity for the HbS.
Sometimes it is caused by
compound heterozygosity for the
HbS mutation, and HbC and other
variants like HbO Arab. Dot blot
analysis with allele specific probes
or primer specific amplification
may be particularly useful.
Prenatal diagnosis.
For some years, the diagnosis of
thalassemia was obtained either
indirectly by polymorphism analysis
or direcly by oligonucleotide
hybridization on electrophoretically
separated DNA fragments.
Nowadays, thalassemias are
detected directly by the analysis of
amplified DNA from fetal
trophoblasts and amniotic fluid cells.
Counselling for
hemoglobin disorders.
Couples at risk for hemoglobin disorders
may be identified retrospectively after the
birth of an affected child, or prospectively
by analyzing childless spouses. Prospective
identification allows parents to have a
disease-free family. Education of the public
by mass media, posters, lectures, etc. was
carried out in Sardinia, Italy. Special
meetings were held with physicians and
especially with pediatricians and
obstetricians, family planning associations,
nurses and social workers.
Future prospects.
Simplification and automation of some PCR
procedures such as primer-specific amplification
for carrier screening and prenatal diagnosis are
expected. Oligonucleotide microchip assay is a
new application for detecting mutations in
medicine. If mutation detection is used, all
hematologic steps would be skipped.
A strong advance would be fetal diagnosis by
analysis of fetal cells in the maternal circulation.
Point mutations responsible for -thalassemia
and sickle cell anemia can be successfully
identified in fetal cells involving magnetically
activated cell sorting using anti-transferrin
receptor antibody.