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EMBRYO SCREENING BY PREIMPLANTATION GENETIC DIAGNOSIS – PGD
Pre-implantation
Genetic Diagnosis (PGD), also known as Embryo Screening,
is a state-of-the-art procedure used in conjunction with in vitro fertilization.
PGD is generally recommended to detect numerical or structural anomalies
in the chromosomes of embryos, as well as conditions caused by single
gene defects. When embryos are affected by certain chromosomal conditions,
these can prevent implantation to the uterine lining, lead to pregnancy
loss, or result in the birth of a child with physical problems and/or
mental retardation.
PGD can help prevent adverse outcomes by identifying affected embryos
as they are developing in the laboratory and before they are transferred
to the womb during the IVF cycle.
PRE-IMPLANTATION GENETIC
DIAGNOSIS (PGD) FOR ANEUPLOIDY:
Normal human cells (i.e. embryonic cells) contain 46 chromosomes in
23 pairs. We receive 23 chromosomes from each parent. The first 22 pairs
of chromosomes are the same for men and women. The 23rd pair determines
our sex. A female has two "X" chromosomes, whereas a male
has an "X" and a "Y." As such, the woman can only
pass an X to her child in her egg. The man passes either the X or the
Y in the sperm, therefore determining the sex of the child. If an error
occurs, and the egg or sperm has an extra or missing chromosome, the
embryo created by that egg or sperm will have an extra or a missing
chromosome resulting in a condition called aneuploidy. Having an extra
chromosome is known as trisomy (tri = three of a given chromosome) and
having a chromosome missing is known as monosomy (mono = one of the
chromosome). When aneuploidy involves the larger chromosomes, the embryo
may not attach to the wall of the uterus or may stop developing soon
after attaching and miscarry. However, if the aneuploidy involves chromosomes
such as the 13, 18, 21, X or Y, the pregnancy may still carry on until
birth, even though the pregnancy has a chromosomal disorder. The most
common of these is an extra number 21, known as Down syndrome or trisomy
21 (three 21 chromosomes). Other common aneuploidies are Klinefelter
syndrome (XXY), trisomy 13, and trisomy 18. The features of the chromosome
condition depend upon which chromosome is extra or missing, but can
include physical differences and mental retardation.
ANEUPLOIDY AND MATERNAL
AGE:
As a woman advances in age, the chance of aneuploidy in her pregnancies
increases because her eggs are as old as she is. Females have all of
their eggs in the fetal stage, and as a result, they are born with all
the eggs they will have in their lifetime. In males, sperm is made every
65-75 days; therefore, the sperm is not as old as the man. Based on
this, the theory regarding aneuploidy risk and advancing maternal age
is that, over time, the chromosomes in the egg are less likely to divide
properly, which results in the egg having an extra or missing chromosome.
Therefore, the risk of aneuploidy increases with maternal age.
The purpose of preimplantation genetic diagnosis for aneuploidy,
therefore, is to select for transfer only chromosomally normal embryos
so as to achieve more pregnancies, reduce the number of pregnancy losses,
and reduce the number of affected offspring.
HOW IS PGD PERFORMED ?
Two procedures are currently available in order to do PGD of embryos.
The PGD team of doctors, geneticists and embryologists will decide which
procedure to use.
 Polar
Body Biopsy: The maturing egg produces two small cells,
called the polar bodies, which degenerate after fertilization. The chromosomal
content of these cells allows us to infer the chromosomal content of
the egg. In order to test the polar body, an opening is made in the
covering of the egg and the polar body is carefully removed. The polar
body is then analyzed while the egg is kept in culture in an incubator.
Analysis of polar bodies provides information only from the mother.
Chromosome abnormalities that may occur after fertilization, when the
sperm meets the egg, will not be detected via Polar body biopsy.
 Embryo
Biopsy or Blastomere Biopsy: To test an embryo, a blastomere
or embryonic cell is removed via a microscopic opening ade in the covering
of the embryo during its third day of development (5 to 8-cell stage).
The embryo is then kept in culture in an incubator while the cell is
analyzed by PGD.
THE PGD ANALYSIS:
 The
biopsied cells are analyzed using a technique called FLUORESCENCE IN-SITU
HYBRIDIZATION OR FISH.
This technique uses probes, small pieces of DNA that are a match for
the chromosomes we want to analyze, to count the chromosomes present.
Each probe is labeled with a different fluorescent dye. These fluorescent
probes are applied to the biopsied cell and attach to the chromosomes.
Under a fluorescent microscope, the number of chromosomes of each type
(color) can then be counted. The geneticist, therefore, can distinguish
normal cells from cells with aneuploidy. Testing of the cells destroys
them because they must be glued to a glass slide and repeatedly heated
and cooled. As such, one cannot use them for another purpose or return
them to the embryo. This analysis causes
no extra inconvenience to the patient as it is accomplished in one day.
The picture displays various chromosomes during FISH analysis.
ADVANTAGES OF PGD
1. Reduction in the
Chance of Having a Child with Aneuploidy: According
to current figures, the chance for a woman delivering a baby with aneuploidy
is on average 1% if she is 35-39 years of age and ~3.5% if she is 40-45.
PGD does lower the chance of having an affected baby. However, we are
unable to test all of the chromosomes at present. We therefore recommend
that prenatal testing be performed in the resultant pregnancy via chorionic
villous sampling or amniocentesis in order to confirm our diagnosis
from PGD and to rule out other aneuploidies for which we do not test.
2. Increased Implantation Rate:
It is well known that the pregnancy rate after in vitro fertilization
decreases dramatically with maternal age. Aneuploid embryos have much
lower survival rates than normal embryos, and half of them (the ones
missing a chromosome) seldom implant. It appears likely that the decrease
in pregnancy rates with maternal age is mostly caused by a corresponding
increase in the number of aneuploid embryos. By performing PGD for aneuploidy
and transferring only chromosomally normal embryos, we might be able
to increase the pregnancy rates noticeably. In several recent studies,
an increase in implantation rates after PGD has been demonstrated. In
addition, the implantation rate increased from 18% in controls to 23%
in PGD patients when we the number of chromosomes tested was increased.
3. Reduction in Pregnancy Losses:
In women 35 and older, approximately 35% of pregnancies are miscarried.
Aneuploidy accounts for 50%, or more of these losses. By transferring
only chromosomally normal embryos, the number of pregnancies going to
term should increase. Recent studies have detected a significant reduction
in pregnancy losses after PGD, from 23% to 9%. The increase in implantation
rate and the significant decrease in pregnancy loss rate resulted in
a significant increase in ongoing pregnancies and delivered babies.
ISSUES ASSOCIATED WITH
PGD
1. The Risk of Embryo
Biopsy: While PGD is a relatively new procedure in IVF,
the micromanipulation techniques required to perform it have been in
use for many years. The risk of accidental damage to an embryo during
removal of the cell(s) in the hands of an experienced embryologist is
very low, and it is currently calculated at less than 1.0% Other Assisted
Reproduction procedures such as Intracytoplasmic Sperm Injection (ICSI),
Fragment Removal and Assisted Hatching are all performed by making microsurgical
openings in the covering of the egg or embryo and none have been found
to have other than mostly positive effects on implantation and viable
pregnancy rates.
2. Removal of Cells from the Embryo:
No part of the future fetus will be affected because one or two cells
are removed from an embryo approximately two days after fertilization.
At this develeopmental stage all cells in an embryo remain totipotent
(until about the fourth day). These cells have not differentiated yet,
meaning that each cell by itself can grow into a whole and perfect fetus.
The biopsy procedure merely delays continued cell division for a few
hours, after which the embryo reaches the same number of cells as before
and continues its normal development. It is possible that embryo biopsy
may lower embryo implantation rates slightly, while selection of chromosomally
normal embryos via PGD may increase them. Therefore, the balance between
potential biopsy damage and beneficial effects of PGD seems to be positive.
3. Misdiagnosis: The accuracy
of PGD for aneuploidy is approximately 90%. This means that the error
rate is 10%. Within this chance of misdiagnosis, there is a false negative
rate, a false positive rate, the chance for no result and the chance
for mosaicism. A mosaicism is defined as the embryo having cells with
different chromosome make-up. Typically, all cells of the embryo have
the same chromosomal make-up as they originate from the same fertilized
egg. However, it is possible for cells of the same embryo to have differing
numbers of chromosomes.
When the cell analyzed has a different chromosomal complement than all
the others in the embryo a misdiagnosis occurs. Due to the chance of
misdiagnosis as well as the presence of aneuploidies, for which testing
is not available, we recommend prenatal testing as stated earlier.
SUMMARY OF REASONS TO CONSIDER
PGD
1. Recurrent miscarriage
2. > 2 unsuccessful IVF cycles
3. Family history of structural chromosomal condition
4. Unexplained Infertility
5. Advanced Maternal Age
6. Family history of X-linked disease
7. Severe Male Factor Infertility
8. Single-gene defects
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