Genetics: C2 Flashcards
The human genome has approx. how much genes, which are the individual units of heredity of all traits?
25,000 genes
some sources est. 30,000-40,000 genes
What can contain one copy (N) of this genetic complement and are haploid?
Reproductive or germline cells
What can contain two complete copies (2N) and are diploid?
Somatic (non-germline) cells
What are organized into long segments of DNA, which, during cell division, are compacted into intricate structures with proteins to form chromosomes?
Gene
What has 46 chromosomes (22 pairs of autosomes, or non-sex chromosomes, and 1 pair of
sex chromosomes?
Each somatic cell
What can contain 22 autosomes and 1 sex chromosome, for a total of 23?
Germ cells (eggs, sperm)
At this, the full diploid chromosome complement of 46 is again realized in the embryo.
At fertilization
What is a functional unit that is regulated by transcription and encodes a RNA product, which is most commonly, but not always, translated into a protein that exerts activity within or outside the cell?
Gene
Genetic information in DNA is transmitted to daughter cells under two different circumstances:
1) Somatic cells divide by mitosis, allowing the diploid (2n) genome to replicate itself completely in
conjunction with cell division; and
2) Germ cells (sperm and ova) undergo meiosis, a process that enables the reduction of the diploid (2n) set of chromosomes to the haploid state (1n).
What has the function is to distribute and maintain the continuity of the genetic material in every cell of the body?
This results in equal distribution of the chromosomes to the two daughter cells.
Mitosis
Overview
● Prior to mitosis, cells exit the resting, or G0 state, and enter the cell cycle. After traversing a critical checkpoint in G1 cells undergo DNA synthesis (S phase), during
which the DNA in each chromosome is replicated, yielding two pairs of sister chromatids (2n –> 4).
● The process of DNA synthesis requires stringent fidelity in order to avoid transmitting errors to subsequent generations of cells.
● After completion of DNA synthesis, cells enter G2 and progress through a second checkpoint before entering mitosis.
● At this stage, the chromosomes condense and are aligned along the equatorial plate at metaphase.
● The two identical sister chromatids, held together at the centromere, divide and migrate to opposite poles of the cell.
● After formation of a nuclear membrane around the two separated sets of chromatids, the cell divides and two daughter cells are formed, thus restoring the diploid (2n) state.
Prior to mitosis, what happens to cells?
Cells exit the resting, or G0 state, and enter the cell cycle.
After traversing a critical checkpoint in G1, cells undergo:
DNA synthesis (S phase), during
which the DNA in each chromosome is replicated, yielding two pairs of sister chromatids (2n –> 4n)
The process of DNA synthesis requires
stringent fidelity in order to avoid transmitting errors to subsequent generations of cells.
After completion of DNA synthesis, cells enter what?
Enter G2 and progress through a second checkpoint before entering
mitosis.
At this stage, the chromosomes condense and are aligned along the equatorial plate at metaphase.
What are held together at the centromere, divide and migrate to opposite poles of the cell?
The two identical sister chromatids
After formation of a nuclear membrane around the two separated sets of chromatids, the cell divides and how many daughter cells are formed?
two daughter cells are formed, thus restoring the diploid (2n) state
duplicated chromosomes condense into microscopically visible bodies
Prophase
chromosomes continue to contract and line up on the metaphase plate (equatorial plate)
Metaphase
chromatids of the chromosomes separate at the centromeres and migrate to the opposite poles of the cell.
Anaphase
separated chromatids gather, a nuclear membrane forms around the separated sets and the cell divides, forming two daughter cells.
Telophase
Overview
Assortment and segregation of genes during meiosis.
● Meiosis occurs only in germ cells of the gonads.
○ It shares certain features with mitosis but involves two distinct steps of cell division that reduce the chromosome number to the haploid state.
○ In addition, there is active recombination that generates genetic diversity.
○ During the cell division, two sister chromatids (2n-›4n) are formed for each chromosome pair and there is an exchange of DNA between homologous paternal and maternal chromosomes.
○ This process involves the formation of chiasmata, structures that correspond to the DNA segments that cross over between the maternal and paternal homologues.
○ Usually there is at least one crossover on each chromosomal arm; recombination occurs more frequently in female meiosis than in male meiosis.
○ Subsequently, the chromosomes segregate randomly.
○ Because there are 23 chromosomes, there exist 2^23 (>8 million) possible combinations of chromosomes.
○ Together with the genetic exchanges that occur during recombination, chromosomal segregation generates tremendous diversity. and each gamete is genetically unique.
○ The process of recombination, and the independent segregation of chromosomes, provide the foundation for performing linkage analyses, whereby one attempts to correlate the inheritance of certain chromosomal regions (or linked genes) with the presence of a disease or genetic trait.
● After the first meiotic division, which results in two daughter cells (2n), the two chromatids of each chromosome separate during a second meiotic division to yield four gametes with a haploid state (1n).
○ When the egg is fertilized by sperm, the two haploid sets are combined, thereby restoring the diploid state (2n) in the zygote.
● In males, each primary spermatocyte forms four functional spermatids that develop into sperm, whereas in females, each oocyte forms only one ovum, the remaining products of meiosis being non - functional polar bodies.
● Processes fundamental to meiosis include chromosome
pairing, chromosome crossing over, and chromosome segregation.
○ These processes result in halving the chromosome number, regular distribution of chromosomes to daughter cells, and independent assortment of the genetic material from both the cross - over events and maternal - paternal homologue distribution in meiosis I, the ultimate result ensures genetic variability.
Meiosis occurs only in
germ cells of the gonads
○ It shares certain features with mitosis but involves two distinct steps of cell division that reduce the chromosome number to the haploid state.
○ In addition, there is active recombination that generates genetic diversity.
Meiosis
During cell division, what are formed for each chromosome pair and there is an exchange of DNA between homologous paternal and maternal chromosomes?
two sister chromatids
(2n -› 4n)
What are structures that correspond to the DNA segments that cross over between the maternal and paternal homologues?
Chiasmata
Usually how many crossover on each
chromosomal arm occur?
at least one crossover
What occurs more
frequently in female meiosis than in male meiosis?
Recombination
Subsequently, the chromosomes segregate
randomly.
Because there are 23 chromosomes, how many possible combinations of chromosomes are there?
2^23
(>8 million)
What can provide the foundation for performing linkage analyses, whereby one attempts to correlate the inheritance of certain chromosomal regions (or linked genes) with the presence of a disease or genetic trait?
The process of recombination, and the independent segregation of chromosomes
After the first meiotic division, which results in two daughter cells (2n), what happens to the 2 chromatids of each chromosome?
the two chromatids of each
chromosome separate during a second meiotic division to yield four gametes with a haploid state (1n).
When the egg is fertilized by sperm, the two haploid sets are combined, thereby restoring the diploid state (2n) in the zygote
What forms four
functional spermatids that develop into sperm?
Primary spermatocyte
What forms only one ovum, the
remaining products of meiosis being non - functional polar bodies?
Each oocyte
Processes fundamental to meiosis include:
(1) chromosome pairing,
(2) chromosome crossing over, and
(3) chromosome segregation.
These processes result in halving the chromosome number, regular distribution of chromosomes to daughter cells, and independent assortment of the genetic material from both the cross - over events and maternal - paternal homologue distribution in meiosis I, the ultimate result ensures genetic variability.
Each chromosome has a narrow waist which divides chromosomes into the short (p) and long (q) arms. This is the:
Centromere
Chromosomes are classified based on the:
(1) position of
centromere and
(2) size of the chromosomes
Positions of centromere:
1) Metacentric
2) Acrocentric
3) Submetacentric
● Centromere in the middle
● Chromosomes 1, 3, 16, 19 and 20
Metacentric
● Centromere close to one end
● Chromosomes 13, 14, 15, 21, 22 and Y
Acrocentric
● Centromere in an intermediate position
● Chromosomes 2, 4-12, 17, 18 and X
Submetacentric
Size of Chromosome
group A : chromosomes 1-3
group B : chromosomes 4-5
group C: chromosomes 6-12 and X
group D : chromosomes 13-15
group E : chromosomes 16-18
group F : chromosomes 19-20
group G : chromosomes 21-22 and Y
The nomenclature used to describe the chromosomes
and their bands, variants, and rearrangements is described in detail by the
International System of
Human Cytogenetic Nomenclature (ISCN)
What is used to describe the chromosome complement of an individual?
A shorthand notation
In the notation, what is specified first?
the number of chromosomes is specified first,
followed by the listing of the sex chromosomes
Thus, a normal female karyotype is designated as 46, XX and a normal male karyotype is designated 46, XY.
Any deviations of the autosomes are written where?
after the sex
chromosomes
What is referred to by its number, its short arm by the letter p, and its long arm by the letter q?
An individual autosome
A + or - sign written before a designated chromosome indicates that the chromosome is
extra (+) or missing (-)
for example. 47, XX + 21 describes a female with 47 chromosomes, including an extra chromosome 21 in addition to the 46 chromosomes of the normal karyotype
COMMONLY USED ABBREVIATION IN
CYTOGENETICS
del deletion
mos mosaic
der Derivative chromosome
p Short arm
die Dicentric chromosome
pat Paternal origin
dup duplication
(+) Gain of
fra Fragile site
q Long arm
h Secondary constriction (heterochromatin region)
r ring
i isochromosome
rep Reciprocal translocation
ins insertion
rea Recombinant translocation
Dir ins Direct insertion
Rob Robertsonian translocation
Inv ins Inverted insertion
(/) Separates cell lines in describing mosaics
mar marker
t Translocation
mat Maternal origin
ter Terminal end of a chromosome arm
(-) Loss of
EXAMPLES OF CYTOGENETIC NOMENCLATURE
46, XX Normal female
46, XY Normal male
45, X Monosomy X (Turner syndrome)
47, XXY Klinefelter syndrome
47, XXX Triple X female
47, XYY XYY syndrome
48, XXX, 49, XXXXY Two of the more common types of
polysomy X
46, XY / 47, XXY Mosaic Klinefelter syndrome
45, X / 46, XX Mosaic Turner syndrome
47, XY,+21 Trisomy 21 (Down syndrome)
46, XX/47, XX,+21 Mosaic down syndrome
47,XX,+18 Trisomy 18 (Edwards syndrome)
47,XY,+13 Trisomy 13 (Patau syndrome)
45, XX,-21 Monosomy 21
69, XXY Triploidy
92, XXXX Tetraploidy
46, XX,4p-
Deletion of short arm of chromosome No. 4 (Wolf-Hirschorn syndrome)
46, XX,5p-
Deletion of short arm of chromosome no. 5 (Cri-du-chat syndrome)
46, XX, t(4;12)(p14;p13)
Reciprocal translocation between
chromosomes 4 and 12, with
brek-points at p14 in chromosome 4 and p13 in chromosome 12
45, XY,-14,-21,+rob(14;21)(p11;p11)
Balanced carrier of dicentric
Robertsonian translocation between [chromosome 14 and 21]
46, XY,-14,+rob(14;21)(p11;p11)mat
Unbalanced complement, having
received (14;21) Robertsonian
chromosome from mother in place
of normal chromosome 14 (the
karyotype of Translocation Down
syndrome)
46, XX,r(15) Ring chromosome of No. 15
46, X,i(Xq) An isochromosome of the X long arm
What is the designation for the visual display of
chromosome studies?
Karyotype
It depicts all chromosomes in an individual cell
What do laboratory workers use to rearrange the images so that the chromosomes are lined up in pairs, typically beginning with the autosomes-chromosomes
1-22- and ending with the sex chromosomes?
Computer
This display is obtained by karyotyping
What refers to the systematic arrangement from a photograph or by computer of previously stained and banded chromosomes of a single cell by pairs?
Karyotyping
The usual procedure of producing a chromosome spread is to arrest mitosis in dividing cells in metaphase by the use of
colchicines
and then to stain the chromosomes
In this, the individual chromosome
takes the form of two chromatids connected at the centromere.
Metaphase spread
Because non-dividing chromosomes cannot be analyzed, what are required for chromosome analysis?
Live dividing cells
The cell type most commonly used is the
mitogenically
stimulated peripheral blood lymphocyte
What cell types are also used for special tests?
Skin fibroblasts
Bone marrow cells
Amniotic fluid cells
Chorionic villus cells
Tumor cells
Dividing cells are accumulated at what phase?
Metaphase
What is a drug that destroys the
mitotic spindle and is added to the culture medium toward the end of the culture period?
Colcemide
The cells are subjected to what treatment followed by fixation and spreading on microscope slides? The slides are then stained.
hypotonic
What may result in either a non-banded or a banded appearance of the chromosomes?
Staining techniques
● The main staining method used in routine clinical cytogenetics
● 300-400 alternating light and dark bands are produced allows for precise identification of every
chromosome and for detection and delineation of structural abnormalities
Giemsa or G banding
Stains constitutive heterochromatin, some of the
material on the short arms of the acrocentric chromosomes and distal part of the long arm of Y chromosomes
Centromere or C banding
Reverse staining of those produced by G and Q
banding
R banding
Stains black the nucleolar organizing regions which contain the multiple copies of genes coding for rRNA
NOR (silver)
● Produces a similar pattern to G banding
● Useful in the examination of the Y chromosome
Quinacrine or Q banding
Stains the heterochromatin of Chromosomes 1, 9,
15, 16 and Y
Distamycin A/ DAPI
What e helps doctors determine if a person has extra chromosomes, missing chromosomes, or chromosomes that have attached to one another in unusual ways?
A complete karyotype
What are first evaluated
microscopically, then their images are captured by a video camera and stored on a computer?
Metaphase chromosome spreads
○ Homologous chromosomes from a metaphase spread can be paired and arranged systematically into a karyotype.
○ The chromosomes are arranged by size in pairs, the largest autosome being designated chromosome 1 and the smallest chromosome 22.
○ The sex chromosomes (X and Y) make up the 23’ pair.
○ A description of the karyotype includes two or three parts: (1) the number of chromosomes, (2) the sex chromosome constitution, and (3) any abnormalities
noted.
○ A normal karyotype is 46, XX for females and 46, XY for males. If present, abnormalities are noted after the sex chromosome complement.
INDICATIONS FOR CHROMOSOME
ANALYSIS/KARYOTYPING
- The child with multiple congenital anomalies and/or dysmorphic features
- Pregnant women into advanced maternal age (>35 year)
- Patients with two major malformations and/or three minor
malformations - Patients with problems in early growth and development, including ambiguous genitalia or mental retardation
- Fertility problems or recurrent miscarriage (>/=3), stillbirth, and neonatal death
- First - degree relative with a known or suspected structural chromosome abnormality
What is a term used to describe an individual who has 2 different cell lines derived from a single zygote?
Mosaicism
Mitotic errors in early development give rise to two or more populations of cells with different chromosomal complement, in the same individual, a condition referred to as
mosaicism
Mosaicism can result from mitotic errors during what?
during the
cleavage of the fertilized ovum or in somatic cells
Mosaicism affecting the sex chromosomes is relatively common.
In the division of the fertilized ovum, an error may lead to one of the daughter cells receiving three sex chromosomes, whereas the other receives only one,
yielding, for example, a 45,X/47,XXX mosaic.
○ All descendent cells derived from each of these precursors thus have either a 47,XXX complement or a 45X complement.
○ Such a patient is a mosaic variant of Turner syndrome, with the extent of phenotypic expression dependent on the number and distribution of the 45X cells.
Autosomal mosaicism seems to be much less common than that involving the sex chromosomes. An error in an early mitotic division affecting the autosomes usually leads to a
nonviable mosaic due to autosomal monosomy
○ Rarely, the nonviable cell population is lost during embryogenesis.
○ Yielding a viable mosaic (e.g.46, XY/47, XY, +21).
Such a patient is a trisomy 21 mosaic with variable expression of Down syndrome, depending on the proportion of cells containing the trisomy
FACTORS THAT CAN CAUSE CHROMOSOMAL
ABNORMALITIES
Late Maternal Age
● The tendency to nondisjunction in elderly females has been attributed to a peculiarity of oogenesis
Radiation
● Induce many chromosome mutations
● 90% of the induced aberration are eliminated during embryonic development- more than half before implantation in the uterus
● 5% give rise to offspring with translocations or being aneuploid themselves
Autoimmune Disease
Viral Infection
● Site specific and may induce a mutational event in the
chromosome
Drugs
● Mutagenic drugs such as alkylating agents, nucleic acid
analogues, mitomycin and chlorpromazine
Cancer
● Most neoplastic cells are aneuploid
● Chromosomal rearrangements in association with a particular type of chromosome
The phenotypic anomalies that result from chromosomal
aberrations are mainly due to
Imbalance of genetic
information
What can be observed in 9.2 per 1000 newborns and in 50% of spontaneous abortions and stillbirths, 1 in 100 has a recognizable chromosomal abnormality and ¼ has a sex chromosomal abnormality?
Chromosomal aberrations
TWO TYPES OF CHROMOSOMAL ANOMALIES
Abnormalities of Chromosome Number
1) Aneuploidy
2) Polyploidy
3) Trisomies
4) Monosomies
Abnormalities in Chromosome Structure
1) Deletions
2) Translocation
3) Ring chromosome
4) Isochromosomes
5) Inversions
6) Duplications
7) Insertions
These arise from nondisjunction, that is, from the failure of two homologous chromosomes in the first division of meiosis or of two sister chromatids in either mitosis or the second division of meiosis to pass to opposite poles of the cell.
Abnormalities of Chromosome Number
What results in cells with abnormal chromosome numbers?
Nondisjunction
If nondisjunction occurs during an early cleavage division of a zygote, then what may occur?
chromosomal mosaicism (two or more cell lines differing in chromosome complement)
Somatic cells contain an abnormal number of normal chromosomes.
This is when a human cell has 23 chromosomes, the number of chromosomes in an ova or sperm.
Haploid cell
Chromosome number is not an exact multiple of the haploid set
Aneuploidy
Caused by either a non-disjunction, that is the failure of
paired chromosomes or sister chromatids to disjoin at anaphase, or an anaphase lag.
Aneuploidy
There is a complete extra set of chromosomes, increasing the chromosomes number to 69 (triploidy) or 92 (tetraploidy).
Polyploidy
What is caused by fertilization of 2 sperms, failure of one of the maturation divisions of either egg or sperm, or failure to complete the 15 zygotic division?
Polyploidy
What is the most common abnormalities of chromosome number?
Trisomies
Occurs when there are 3 representatives of a particular chromosome instead of the usual two:
Trisomies
Usually the result of meiotic non-disjunction
Trisomies
The most frequent and best known trisomy in human is:
Trisomy 21 or Down syndrome
Other relatively common
trisomies are
Trisomy 18 or Edwards syndrome
Trisomy 13 or Patau syndrome
Occurs when only one representative of a
chromosome is present
Monosomies
What is due to the non-disjunction or anaphase lag and is lethal early in development and only survive in mosaic form?
Complete monosomy
What are usually the offspring of a
translocation carrier?
Partial monosomy
What can result from chromosome breakage and reunion?
Abnormalities of Chromosome Structure
When a chromosome breaks, it can rejoin in its old form; what is the term?
Restitution
When a chromosome breaks, it can rejoin with another broken chromosome; what is the term?
Reunion
This leads to a structural rearrangement that can be
balanced or unbalanced.
Reunion
The amount of genetic material is presumed to be identical to that found in a normal cell, and there is a simple rearrangement of the distribution of this material.
Balanced or unbalanced?
Balanced
■ Types of balanced rearrangements include balanced reciprocal translocations,
Robertsonian translocations, and inversions.
■ Balanced chromosome rearrangements do not usually lead to any clinical change
This rearragement indicates loss or gain of chromosome material.
Unbalanced
Such unbalanced rearrangements of meiotic cells usually result in changes in the clinical phenotype due to chromosomal breakage and rearrangements.
Occurs when a piece of a chromosome is missing
Deletion
May occur as a simple deletions or as a deletion with duplication of another chromosomal segment.
May be located at the chromosome ends or in interstitial segments of the chromosome and are usually associated with mental retardation and malformations.
Deletions
What are the most commonly observed deletions?
4p-, 5p-, 9p-, 11p-, 13p-, 18p- and 18q-
What are defined as a small chromosome deletions that are detectable only in high-quality (pro)metaphase preparations, these deletions often involve several genes so that the affected individual may be identified by an unusual phenotype associated with a single gene mutation?
Microdeletions
Duchenne muscular dystrophy, Prader-Willi syndome
Involves the transfer of chromosomal material from one chromosome to another
Translocation
Occur with a frequency of 1/500 liveborn human infants
How is translocation acquired?
May be inherited from a parent or appear de novo
Result of breaks in non-homologous chromosomes with reciprocal exchange of the broken segments
Reciprocal translocation
This results from exchange of chromosome segments between non-homologous chromosomes
Balanced reciprocal translocation
An individual carrying such a rearrangement has a higher frequency of abnormal gametes as the result of a disturbance in chromosome pairing at meiosis. Such individuals themselves have a balanced chromosome complement and are clinically normal, but they may have a high risk of having congenitally malformed children and/or spontaneous abortions. Normal children may also be born. Such persons require careful genetic counseling.
Involve two acrocentric (centromere located at the end) chromosomes that fuse near the
centromeric region with subsequent loss of the nonfunctional, very truncated short arms
Robertsonian translocation
This is a specific type of unequal reciprocal translocation that occurs between acrocentric chromosomes, resulting in a new metacentric chromosome formed from two acrocentric chromosomes.
Robertsonian Translocation
Such rearrangements may be important in the transmission of Down syndrome when one of the chromosomes involved is chromsome 21. If the structural change occurs in the early embryo, mosaicism with some cells normal and some with the structural abnormality may result.
Both tips of a chromosome are broken and the ends of the centric fragment rejoins forming a ring
Ring chromosome
● Very rare but have been found in all human chromosomes
● The phenotype ranges from mental retardation and multiple congenital anomalies to normal
If a ring replaces a normal chromosome, the result is:
Partial monosomy
A horizontal division of the centromeres rather than from a vertical one as the formal process of mitosis
Isochromosomes
Seen in connection with x-chromosome
The segment between 2 breaks in a single chromosome is inverted and the order of the gene is reversed:
Inversions
The breaks are in the two opposite arms of the chromosome so that the intervening portion that contains the centromere is reversed
Pericentric inversions
Involve only chromosomal material from one
arm of a chromosorne
Paracentric inversions
These result from two chromosome breaks with inversions of the intervening segment and can be detected only by
By altered position of the centromere or by chromosome banding studies that show a changed banding sequence
Inversions result in disturbances in chromosome pairing and in the formation of unbalanced as well as balanced gametes.
● Presence of extra genetic material from the same chromosome
● May result from the abnormal segregation in carriers of translocations or inversions
Duplication
What is the addition of a chromosome segment and may be result of breakage reunion or
of replication error?
Duplication
The isochromosome shown in Figure 34-2D has both a deleted short arm and a duplicated long arm, or deletion and duplication.
This occurs when a piece of chromosome breaks at two
points and is incorporated into a break in another part of a chromosome
Insertions
READING AND WRITING KARYOTYPES
● If an abnormality is found, it is noted after the sex chromosome constitution.
○ For example, in a male with Down syndrome in which there is an extra chromosome 21, the
karyolype is written as 47, XY, +21
● To describe a translocation between two autosomes the small letter “t” is used outside parentheses and the chromosome numbers are inside: for example, 46, XY, t (9; 21)(q 34; p11).
○ This indicates a male with a normal number of chromosomes, and a reciprocal translocation between the long arm of chromosome 9 and short arm of chromosome 21 with designated breakpoints.
● In case of translocation, the chromosomes involved are
written in brackets preceded by a t, as in 45, XX, t(13q14q), indicating a female carrier of a translocation between the long arms of chromosomes 13 and 14.
● If the chromosome breaks are along an arm of a chromosome, the band position at which the break occurred is also indicated in the brackets, as in 45, XY, t(13q2.1-14q1.3), indicating a male carrier of a translocation within the long arms of chromosomes 13 and 14.
CHROMOSOMAL ABNORMALITIES
45, X Liveborn (Turner Syndrome)
47, XXY (Klinefelter Syndrome)
47, XXX
47, XYY
48, XXXX; 48, XXXY; 49, XXXXX; 49, XXXXY
The most consistent features found in girls with a
missing sex chromosome are short stature, usually
beginning before birth, and gonadal dysgenesis.
45, X Liveborn (Turner
Syndrome)
● Although eggs may be present in the newborn gonad, early attrition takes place and eggs have disappeared by puberty.
● The puffiness of the hands and feet usually disappears in childhood.
● Low posterior hairline and short and/or webbed neck, excessive pigmented nevi, deep-set nails, short fourth metacarpal, narrow maxilla, prominent ears, horseshoe kidney and heart defect (usually coarctation of the
aorta).
● Pubertal development usually does not occur in the absence of hormonal treatment.
The 45, X Liveborn (Turner Syndrome)
This condition occurs in about 1 in 500 males and is the most common cause of male infertility
47, XXY (Klinefelter Syndrome)
These males are tall with long limbs, have testes that
remain small after puberty, have gynecomastia, are dull
mentally (about 20% with IQ < 80), and are immature in
behavior.
47, XXY (Klinefelter Syndrome)
There are no characteristic major or minor malformations found in ____ females; almost all appear normal at birth.
47, XXX
● Height, weight, and head circumference are generally normal, although the head may be at a lower centile than height and weight.
● Language is usually delayed, and IQ is lower than normal, particularly lower than the IQ of siblings.
● Behavioral and social problems and emotional immaturity are the rule.
● Normal sexual development generally occurs although
apparently not frequent, both 47, XXX and 47, XXY individuals have been reported in offspring of females with this condition.
47, XXX
The karyotype has elicited much public interest because of the reports of association with criminality
47, XYY
● Occurrence of the karyotype is common, occurring in about 1 in 1000 male births.
● These males are tall and may have mild fine motor problems, impulsive behavior, and temper outbursts.
● Although most affected males blend into the general population, it is now recognized, after correction for earlier ascertainment bias, that there is an excess of berrant or criminal behavior as compared with 46, XY
males.
● Such behaviors may occur even with IQ scores as high
as 146.
● However, the average IQ of 47, XYY males is somewhat
lower than that of normal males
47, XYY
● In general, as additional X chromosomes are added, the
phenotypic consequences become more severe.
● Mental retardation is constant, dysmorphia is evident, stature tends to be small; and skeletal anomalies may occur.
● Facial features may suggest Down syndrome
48, XXXX; 48, XXXY; 49, XXXXX; 49, XXXXY
STRUCTURAL ABNORMALITIES
Fragile X
X/Y Translocations
Y-chromosome Syndromes
Overview: Structural Abnormalities
● The outcome of structural X chromosome abnormalities differs greatly for males and females.
● Large duplications and deletions in the female with an accompanying normal X almost always result in Turner Syndrome, whereas they are lethal in the male.
● A specific structural abnormality, isochromosome Xq (two copies of the X long arm joined with the missing short arm) is common among girls with Turner syndrome, comprising 20% of the total abnormal karyotypes.
● Microdeletions and duplications may show little effect in the carrier female but have syndromic consequences for the male.
● For example, microdeletion Xp22.32 in the male results in ichthyosis, chondrodysplasia punctata, short stature, and mental retardation.
● When the deletion is somewhat larger and includes the Kallman gene, inability to smell and hypogonadotropic hypogonadism occur in addition to the other features.
● In all instances of X microdeletion syndromes in the male, mothers should have chromosome analysis performed because the carrier rate is high.
● Clinical features in this syndrome vary even in the male, and carrier females often have minimal expression.
● This chromosome aberration also is not expressed in all who carry the defect, particularly females, making diagnosis of the condition difficult.
● The fragile X chromosome appears as a recurrent break
at the same site in Xg27 in 4 to 50% of cells.
● The syndrome is frequent, occurring in 2 to 6% of retarded males.
● Birth weight is usually high and the head relatively large.
● The face is long and narrow with prominent chin and ears.
● There is usually macroorchidism.
● Mental retardation is moderately severe.
○ About 15% exhibit autistic features.
● With the finding of CG trinucleotide repeat sequences in excess in the fragile X mental retardation 1 gene (FMR-1) in affected males, a molecular test has become available that can reliably diagnose carriers.
Fragile X
The XX with short stature, infantile testes, and mild intellectual difficulties is almost always the result of a cryptic translocation between the distal X short arm and distal Y short arm with transfer of the SRY gene (sex reversal Y) from the Y to the X.
X/Y Translocations
● Mosaicism for loss of the Y chromosome (45, X/46, XY) has varying results, probably depending on the percentage of cells missing a Y chromosome in the developing gonad.
● Peripheral blood cell mosaicism is compatible with normal male phenotype, with genital ambiguity, and with mixed gonadal dysgenesis or full Turner syndrome.
● In the latter case, surgical extirpation and pathologic
examination of the dysgenetic gonads are recommended because of the danger of gonadoblastoma.
● Deletion of the distal short arm of the Y when it includes the SRY gene results in a female with gonadal dysgenesis and usually also lymphedema, as in Turner
syndrome.
● However, these individuals are usually not short and may have excellent muscle strength.
Y-chromosome Syndromes
Overview: AUTOSOMAL STRUCTURAL ABNORMALITIES
● There are an almost unlimited number of different ways in which the 22 pairs of autosomes can be broken with pieces lost (deletions) or reattached (translocations,
duplications).
○ Therefore numerous described syndromes occur owing to imbalance of such chromosome
segments.
● Four such syndromes are summarized to illustrate certain points, all are microdeletion syndromes recognized by their recurring patterns of anomalies
before recognition of their chromosomal origin.
● Other well-known syndromes that have been shown to
be due to microdeletions are Williams syndrome (del 7q), Rubinstein-Taybi syndrome (del 16p), and Miller-Dieker lissencephaly syndrome (del 17 p).
What are now recognized to be the result of deletion of a small portion of the long arm of
chromosome 22 near the centromere?
DEL (22)(911q11)
Velocardiofacial (VCF or Sprintzen) syndrome and
DiGeorge syndrome
● The syndrome appears to be extremes along a clinical continuum not specifically related to the size of the deletion.
● The condition in some individuals is so mild that the individual blends into the normal population.
● The true incidence of this syndrome is unknown, but it may be as common as Down syndrome.
● Although the deletion of chromosome 22(q11) may be so small that it cannot be detected at the microscope without FISH, major malformations may be present (Table 34-1 and Fig. 34-3A).
● Mild to moderate mental retardation may be present,
and speech is hypernasal even without clefting.
● Absent thymus, tonsils, and adenoids and hypocalcemia
in infancy have been noted.
● Affect is often very bland, including facial expression;
phobias and psychosis may appear in adolescents and
adults.
DEL (22)(911q11) VCF (SPRINTZEN) AND DIGEORGE
MICRODELETION SYNDROMES
Common: cleft palate; conotruncal heart defects (interrupted aortic arch, tetralogy of Fallot, right side aortic arch, ventriculoseptal defect)
Also present: pharyngeal hypotonia, retrognathia, malar flatness, small anomalous ears, tortous retinal vessels, slender tapered digits, scoliosis, short stature
Face: long with narrow palpebral fissures, thickening of helical rims; nose is prominent with squared nasal root, philtrum is long, mouth is often held open, velopharyngeal incompetence with or without cleft palate.
Velocardiofacial/
(Sprintzen) DiGeorge
Face: narrow bifrontal diameter, almondshaped eyes, thin upper lip, down-turned mouth
Genitalia particular male,
are hypoplastic; hypogonadism; delayed puberty
Other: truncal obesity after
age 1 to 4 years
Prader-Willi
Small head: The occiput is flat, often with a pronounced occipital
groove. Eyes may be mildly wide spaced and deep set, nose is mildly prominent, mouth is large, teeth widely spaced. Ataxic gait. Happy demeanor.
Angelman
To note on velocardiofacial
syndrome:
Father and daughter with velocardiofacial syndrome.
Note mask-like facial features and lack of ability to purse the lips for whistling. The father had had repair of a congenital heart defect (tetralogy of Fallot); the child had a normal heart. A small deletion of chromosome 22 long arm is found in most cases, sometimes detected only with molecular-cytogenetic techniques.
Prader-Willi syndrome sample case
Twenty-year old man with Prader-Willi syndrome. He has a de novo deletion of the chromosome 15 homolog inherited from his father.
Angelman syndrome sample case
Twelve-year-old boy with angelman syndrome. He is ataxic and unable to walk without help, is happy and
alert, but is severely and mentally retarded and has no speech. He has the same de novo deletion as the patient in figure 34-38, but the
deletion occurred on the chromosome 15 inherited from his mother.
Case Impression
● Diagnosis: Down Syndrome (Trisomy 21)
● Pertinent information:
○ Michael’s mother was 46 years old when he was born
○ Physical characteristics included flat facial
features, chinky-eyed with an upward slant and low set ears
○ Delayed language development i.e. inability to talk and sing properly
● When a condition presents with multiple signs and symptoms then it is considered to be a syndrome. As in this case, where Michael exhibited dysmorphic features along with a language delay.
Autosomal Aneuploidy:
Of the three autosomal trisomies that may survive to a term birth, only one allows long-term survival
Trisomy 21 (Down syndrome)
Trisomies show remarkable variation in parental origin, example:
Paternal nondisjunction is responsible for nearly 50% of 47, XXY but only 5 to 10% of cases of trisomies 13, 14, 15, 21 and 22.
There is considerable variability in the meiotic stage of origin, e.g.
○ For example, among maternally derived
trisomies, all cases of trisomy 16 may be due to meiosis I errors.
○ Whereas for trisomy 21, one-third of cases are associated with meiosis I errors, and for trisomy 18, the majority of cases are apparently due to meiosis I nondisjunction.
○ In spite of this variation in parental and meiotic origin, nondisjunction at maternal meiosis I appears to be the most common source of trisomy
What is the most common source of trisomy?
Nondisjunction at maternal meiosis I
Molecular mechanism underlying nondisjunction, the source of trisomy and monosomy is mostly, if not all associated with
Alterations in genetic recombination
This is the process by which chromosomes
exchange genetic material during the first of the two meiotic divisions
Alterations in genetic recombination
What is the most important etiologic
factor in congenital chromosomal disorders?
Association between increasing maternal age and trisomy
exerted without respect to race, geography, or socioeconomic factors and likely affects segregation of all chromosomes.
Maternal age and Trisomy
Among women under the age of 25,
approximately 20% of all clinically recognized
pregnancies are trisomic; by age of 36, this
increased to 10% and by 42, to >33%.
This could possibly be explained by the age-dependent degradation of cell cycle proteins or meiotic proteins responsible for maintaining sister chromatid cohesion
This happens from the failure of two homologous chromosomes in the first division of meiosis or of two sister chromatids in either mitosis or the second division of meiosis to pass to the opposite poles of the cell.
Nondisjunction
Abnormalities in chromosome number
If these cells are gametes, fertilization results in a zygote with an abnormal chromosome number
If nondisjunction occurs during an early cleavage division of a zygote, what may result?
Chromosomal mosaicism (2 or more cell lines differing in chromosome complement)
What are the 3 types of Down Syndrome?
1) Full trisomy 21
2) Mosaic trisomy 21
3) Translocation type
94% of cases; all cells have an extra
chromosome
Full trisomy 21
2.4% of cases; some cells have an extra chromosome 21.
Mosaic trisomy 21
3.3% to 5% of cases; the extra chromosome 21 is attached to another chromosome, usually the D or G group of chromosomes; predominantly Robertsonian translocation between 14 and 21
Translocation type
Physical Manifestations
noted at birth:
○ Hypotonia
○ Poor moro reflex
○ Hyperflexibility of joints
○ Excess nuchal folds
○ Flat facial profile
○ Upslanting palpebral fissures
○ Small, low set ears
○ Dysplasia of pelvis
○ Clinodactyly (incurving of the 5th finger)
○ Transverse palmar crease (simian crease)
In addition to these features
the following may be observed:
○ Brachycephaly
○ Epicanthal folds
○ Brushfield spots
○ Depressed nasal bridge
○ Open mouth
○ Protruding tongue
○ Brachydactyly
○ Widened gap between the great and 2nd toes (saddle gap)
○ Dermatoglyphic abnormalities
○ Mental retardation
Other clinical manifestations related to DS:
abnormalities in the:
- thyroid
- cardiac
- intestinal
> Increase risk for leukemia
Increased susceptibility to infection and an
immunoglobulin imbalance
Widely recognized characteristic appearance in DS:
○ Depressed nasal bridge
○ Upward slanting eyes with epicanthal folds
○ Small mouth with protruding tongue
○ Smaller ears
○ Flattening of the back of the head
○ Microcephaly
○ Single crease in the palm
○ A gap between the first and second toes
● Children with Down syndrome are usually smaller, and
their physical and mental developments are slower.
● The majority function in the mild to moderate range of
mental retardation.
● Language development is also markedly delayed.
● The physical characteristics and limited intellectual capacity occurs because of the presence of an extra chromosome 21.
What is the most common cause of mental
retardation and congenital anomalies in a newborn?
DS
What is routinely being offered to
mothers 35 years old and above in centers overseas?
Prenatal dx
● For the translocation type of Down Syndrome, the parents need to be screened for a possible carrier state for a translocation.
● If one of the parents is a carrier; a future pregnancy can produce either a normal child, a translocation carrier or a child with Down Syndrome.
Thsi is used d/t increased incidence of the condition with increased maternal age that is used for prenatal diagnosis of Down Syndrome in older mothers.
Amniocentesis
Risk Factors and Incidence
● The risk of a 15 year old woman giving birth to a baby with Down Syndrome is low (1:1578) and gradually increases with maternal age.
● The risk sharply increased beyond 35 years old (1:384).
● After the age of 45, the risk increases to about 1 in 250.
● For the full trisomy type of Down Syndrome, the risk of another affected child is about 1% for the mother whose
first infant was born when the mother is less than 35 years old.
Prognosis
● Studies have shown that persons with Down Syndrome can lead productive lives.
● With an average lifespan of 55 years old, intensive early intervention is now offered to maximize the potentials of these individuals.
● In countries overseas, some of the adults are employed and lead independent lives.
● The Down Syndrome association of the Philippines, Inc. offers emotional support to families
Refers to the search for a genetic disorder in the entire population or in a high-risk population
Genetic Screening
Screening Technique Criteria
● Cost-effective
● High positive predictive value
● Should yield information that leads to disease prevention or a useful therapeutic intervention.
Examples
● Maternal serum markers to detect increased risk of Down syndrome
● Postnatal tests for phenylketonuria, and cholesterol levels in children to identify individuals at risk for familial hyperlipidemias.
Factors that determine which Genetic Disorder should be Screened Routinely
● Diseases frequency
● Severity of the disorder
● Cost of screening
● Whether treatment interventions can alter the course of the disease.
Used in individuals suspected to have a disease based on physical features, family history, or biochemical finding.
Genetic Testing
What do we look for in Genetic Tests?
gene abnormalities & presence/absence of key proteins whose production is directed by specific genes
Abnormalities in either could indicate an
inherited disposition to a disorder
Genetic testing includes:
- gene tests (DNA
testing) - biochemical tests (protein
testing).
In gene tests, DNA in cells taken from a person’s:
- blood
- body fluids
- tissues
These are examined for an abnormality that flags a disease or disorder
The abnormality can be:
can be relatively large - a piece of a chromosome, or even an entire
chromosome, missing or added.
sometimes the change is very small - as little as one extra, missing or altered chemical base within the DNA strand
Genes can be:
- amplified (too many copies)
- over-expressed (too active)
- inactivated
- lost altogether
Sometimes pieces of chromosomes become:
switched, transposed or discovered in an
incorrect location
1.
Focuses on chromosomal abnormalities along with specific genetic disorders for which there is increased risk of parental transmission:
Gene testing
A critical first step before initiating genetic testing is
to:
assure that the correct clinical diagnosis has been made, whether based on family history, characteristic physical findings, or biochemical testing.
This will prevent unnecessary testing and will direct testing towards the most probable candidate genes
Careful clinical assessment
Limitations to accuracy and interpretation of genetic tests:
○ Technical errors
○ Detect only the most common mutations
e.g. A negative result must be qualified by the possibility that the individual may have a mutation that is not included in the test
What is done for susceptibility to chronic disease?
Presymptomatic testing
This must be approached with caution and should be offered only to patients who have been adequately counseled and have provided informed consent.
Predictive molecular testing
Should be offered only to individuals with a suggestive personal or family medical history or in the context of a clinical trial.
Presymptomatic testing
What uses a variety of techniques to examine a
person’s DNA?
Gene tests
○ Some tests involve using probes - short strings of DNA - with base sequences complementary to those of the mutated gene.
- These probes will seek their complements within
an individual’s genome.
- If the mutated sequence is present in the
patient’s genome, the probe will find it and bind
to it, flagging the mutation.
○ Another type of gene tests involves comparing
the sequence of DNA bases in a patient’s gene
to a normal version of the gene
○ Biochemical tests look for the presence or absence of key proteins which signal abnormal or malfunctioning genes.
What can be predictive, discovering whether an individual has an inherited disposition to a certain disease, before symptoms appear?
Gene testing
Genetic tests can also:
- confirm a diagnosis if symptoms are present
- can determine whether a person is a
carrier for the disease (Carriers won’t get the disease, but can pass on the faulty gene to their children) - Prenatal testing (can help expectant parents know whether their unborn child will have a genetic disease or disorder)
- Newborn screening tests infants for abnormal or missing gene products
- Individuals in families at high risk for a disease live with troubling uncertainties about their own future as well as their children.
○ A negative test - especially one that is strongly predictive - can provide an enormous sense of relief.
○ A positive test can also produce benefits. In the best circumstances, a positive test enables the person to take steps to reduce risk. These steps could include regular screening for the disease or lifestyle changes, such as a change in diet or regular exercise. A positive test can relieve uncertainty, and can enable people to make informed decisions about their future.
What are the reasons for genetic testing?
- Predictive testing
- Carrier testing
- Prenatal testing
- Newborn screening
● Identifies people who are at risk of getting a disease before any symptoms appear.
● Include those that screen for some inherited predispositions of certain forms of cancer, such as colon and breast cancer.
● Being predisposed does not mean that the individual will get the disease, it means the person has a certain risk of developing the disease.
PREDICTIVE TESTING
● This can tell individuals if they are carriers of an inherited disorder that they may pass on to their children.
● A person who has only one abnormal copy of a gene for a recessive condition is known as a carrier.
● Carriers won’t get the disease, but can pass on the defective gene to their children.
● Cystic fibrosis and Tay-Sachs disease are examples of disorders for which parents can be carriers.
CARRIER TESTING
● Available to people at risk for having children with a chromosomal abnormality or an inherited genetic condition.
● Two procedures are commonly used in prenatal testing.
○ Amniocentesis involves analyzing a sample of amniotic fluid from the womb.
○ CVS (chorionic villus sampling) involves taking a tiny tissue sample from outside the sac where the fetus develops.
● Prenatal testing is often used to look for disorders such as Down syndrome, spina bifida, cystic fibrosis, and Tay-Sachs disease.
PRENATAL TESTING
The most widespread type of genetic testing, tests infant blooding, samples for abnormal or missing gene products.
○ For example, infants are commonly screened for Phenylketonuria (PKU), an enzyme deficiency that can lead to severe mental retardation if untreated.
NEWBORN SCREENING
How to decide whether to be tested?
● The decision to undergo testing is a very personal one.
● For many people, a pivotal consideration is whether there are preventive measures that can be taken if the test result is positive.
○ For example, those who test positive for
inherited forms of breast or colon cancer can benefit from preventive measures, screening for early detection, and early treatment.
● In contrast, there are no preventive measures or cures for Huntington’s disease, but a positive test for Huntington’s disease might help an individual make lifestyle decisions, such as career choice, family planning or insurance coverage.
● Because the decision about whether to be tested for genetic disease is complex, most people seek guidance from a genetic counselor trained to help individuals and
families weigh the scientific, emotional and ethical considerations that impact on this decision.
Follow-up care after testing
● Post-test interventions may include:
○ Cautious surveillance and appropriate health care screening
○ Specific medical interventions
○ Chemoprevention
○ Risk avoidance
○ Referral to support services
Prenatal Genetic Diagnosis: Techniques for Prenatal Diagnosis
● The advent of prenatal diagnosis through different techniques and procedures represents the most
important advance so far obtained in preventing the birth of infants with serious genetic defects and/or chromosome aberrations.
● Genetic counseling in an increasing number of cases can now be based on actual diagnosis in the uterus instead of calculated probability risks.
● Thus the risks in recessive diseases are no longer 1 in 4 and in dominant disease 1 in 2, but rather 100% or zero depending on antenatal tests.
The techniques currently in use or under investigation for prenatal diagnosis include the following:
○ Fetal tissue sampling
■ Amniocentesis, chorionic villi biopsy, other organ biopsies including liver and muscle biopsy;
○ Fetal visualization
■ Ultrasound, fetal echocardiography, embryoscopy, fetoscopy, magnetic resonance imaging and radiography;
○ Screening for neural tube defect (NTDs)
■ By measuring maternal serum a fetoprotein (MSAFP);
○ Screening for Fetal Down syndrome
■ By measuring MSAFP, unconjugated estriol (UE), and human chorionic-gonadotrophin;
○ Separation
■ Of the fetal cells from mother’s blood; and
○ Preimplantation biopsy of blastocysts obtained by in-vitro fertilization.
Fetal chromosome analysis should be offered in the
following situations:
○ Mother is 35 years old or older at the time of the delivery.
○ A previous child had a trisomic syndrome or other chromosome aberration.
○ A chromosome abnormality is present in either parent.
■ E.g. aneuploidy, balanced translocation, or clinically significant inversion
○ There is a fetus at risk for a serious X-linked disease.
○ Specific prenatal diagnosis is not available.
■ A parent is a fragile X carrier
○ There is an increased risk for Down
syndrome based on the result of maternal
serum screening in a woman who is younger than those ordinarily offered prenatal diagnosis on the basis of maternal age alone.
○ Ultrasound has identified an anatomic
abnormality in the fetus
■ E.g. omphalocele, hydrocephalus, renal
defects, etc.
Biochemical Studies
● Indicated when there is an increased risk of transmission of an error of metabolism.
● Often the increased risk is ascertained when:
○ Diagnosis of an inborn error of metabolism has been made in a previous child,
○ Both members of a couple are found to be carriers of an autosomal recessive disorder by screening based on ethnic origin, or
○ Carrier testing is done based on a history of a close relative with a specific disorder
● May be performed on amniocytes or chorionic villi.
● Study of other specific tissues, e.g. fetal white blood cells, liver biopsy, etc. may be indicated when expression on the gene product, protein or enzyme is confined to that tissue.
What are used to diagnose rare genetic diseases?
○ E.g. epidermolysis bullosa fetalis, for which specific genetic tests are not available
Routine and electron microscopy of biopsies and cells
MOLECULAR GENETIC STUDIES
● The use of molecular techniques in prenatal diagnosis is increasing rapidly.
● The clinical application of these methods is based upon the fact that the DNA complement is generally identical in every cell of the body, and therefore a hereditary defect detectable at the DNA level should be found in any nucleated cell in that individual.
● Enzymes (restriction endonucleases) that cleave DNA within a specific sequence recognition site are used to identify mutant genes, deletions within the gene, or to characterize DNA polymorphisms that are linked to the gene in question.
● Studies of DNA are currently applicable for the prenatal diagnosis of hemoglobinopathies, hemophilia A, Duchenne and Becker muscular dystrophies, cystic fibrosis, fragile X, myotonic dystrophy, neurofibromatosis, and a number of other disorders
When is Transabdominal Amniocentesis well established, safe, reliable and accurate procedure and is most commmonly used technique for obtaining fetal cells and amniotic fluid?
15 to 17 weeks of
pregnancy
Performed under ultrasound guidance, which at this
gestational age also affords the opportunity for
evaluation of fetal anatomy.
How many weeks does initiation and processing of cultures take?
2-3 weeks
but results of cytogenetic examinations are available in two weeks or less in many laboratories
What can be used for measurements of substances like hormones, enzymes, etc.?
supernatant amniotic fluid
Amniocentesis
● The results of laboratory studies on amniotic cell cultures are more than 99% accurate for most biochemical and cytogenetic studies.
● Significant maternal injury from amniocentesis is rare.
● Although injury to the fetus is also rare, there remains a small, procedurally related risk of fetal loss, estimated at less than 0.5% multicenter studies done on early amniocentesis have shown that the risk of miscarriage is higher, about 15%.
CHORIONIC VILLUS SAMPLING (CVS)
● Once removed, the villi are dissected from the decidual tissue.
● Biochemical assay and chromosome studies can be performed directly on cells from these tissue fragments, and DNA can be extracted from them.
○ In addition, tissue culture can be initiated for further studies.
● The major advantage of CVS when compared to standard amniocentesis is that the sample and laboratory results are available much earlier in the pregnancy.
○ The larger quantity of viable cells also makes these samples more suitable than amniotic fluid samples for rapid direct biochemical analysis or extraction of DNA.
○ Some studies claim a higher rate of fetal loss.
○ The risk of miscarriage is 1%.
■ However, with improved techniques, this
risk is reduced dramatically.
● As with amniocentesis, the risk of causing Rhesus factor sensitization may be avoided by administration of RH immune globulin with the procedure.
● Transabdominal chorionic villi sampling may also be performed in the second and third trimesters.
○ This is done when amniocentesis is precluded by severe oligohydramnios or a more rapid diagnosis is needed.
● The major advantage of this procedure is that earlier diagnosis is possible.
● At 9 to 11 menstrual weeks of gestation, a plastic catheter with a solid flexible stylet is inserted transcervically, or a needle is placed transabdominally, using ultrasound guidance, into the developing placenta.
● Chorionic villi fragments are removed by aspiration.
● The transcervical and transabdominal approaches appear to have comparable safety and accuracy.
● Maternal cell contamination is higher with CVS than with amniocentesis, particularly with transcervical CVS.
SAMPLING OF FETAL BLOOD AND TISSUES
● Fetoscopy for fetal blood sampling has been largely replaced by percutaneous umbilical blood sampling (PUBS), also known as cordocentesis.
● A technique for percutaneous sampling of fetal blood from the hepatic vein is also available.
● When fetal blood sampling is done under ultrasonic guidance, there is a lower rate of pregnancy loss following the procedure than after fetoscopy.
● An increased risk of fetal loss has been observed with common indications for fetal blood sampling such as multiple anomalies or intrauterine growth retardation in the fetus.
● Fetal blood sampling is a technique that follows rapid chromosome analysis.
● It is also useful for studying other fetal constituents such as platelets, hemoglobin, blood gasses, etc.
● A number of serious skin disorders, e.g. epidermolysis bullosa and serious genetic forms of ichthyosis may be diagnosed histologically by skin biopsy specimens
obtained percutaneously under ultrasonic guidance with or without fetoscopy.
● Liver and fetal muscle biopsies have also been performed.
● However, for many genetic conditions, fetal biopsy has been replaced by molecular genetic studies of amniocytes or chorionic villi.
ULTRASOUND
● This is the primary method for imaging fetal anatomy. The technique may be used throughout pregnancy, when there is clinical indication to monitor fetal growth, movement, position and morphology; to
assess amniotic fluid volume and to assess gestational age and placental location.
● During the early second semester, ultrasound is used to date the pregnancy, identify twins, diagnose some fetal structural anomalies, and examine the placenta and the amount of amniotic fluid.
● Many fetal organ systems and anatomic lesions can be visualized by ultrasound including some genito-urinary,
gastron-intestinal, skeletal and central nervous system anomalies.
● Fetal echocardiography may also identify many cardiac lesions.
● Ultrasound is also used to guide invasive sampling such as amniocentesis, chorionic villus sampling, amniocentesis, and various fetal biopsies.
● Transvaginal ultrasound has allowed improved resolution of the anatomy of the embryo during the first semester.
● The utility of this approach for earlier detection of certain fetal anomalies, like anencephaly, limb and other cardiac anomalies, is being evaluated.
FETAL ECHOCARDIOGRAPHY
● May be performed from 18 weeks & beyond to term.
● When used together with duplex and/or color flow
Doppler, it can identify a number of major structural cardiac defects and rhythm disturbances.
● This should be considered when any of the following lead to an increased risk for congenital heart disease:
○ An extracardiac malformation identified by ultrasound;
○ Exposure to potentially teratogenic agent;
○ Family history of congenital heart defects, particularly in a parent or sibling;
○ Suspected fetal chromosome abnormality or genetic disease associated with heart defects;
○ Maternal diseases associated with fetal structural heart defects, such as diabetes or PKU, or maternal disease associated with fetal cardiac arrhythmia, particularly heart block, such as lupus or other immune disorders; or
○ Suspected cardiac defects based on the findings of routine ultrasound.
EMBRYOSCOPY
● An experimental technique used in the first trimester of pregnancy.
● A rigid endoscope inserted via the cervix into the space between the chorion and amnion is used to visualize the embryo and offers the potential for diagnosis of structural malformations
FETOSCOPY
● A procedure in which a fine-caliber endoscope is inserted into the uterus was developed for fetal visualization and fetal blood sampling.
● Because of the risk of spontaneous abortion associated with fetoscopy, it is rarely used today and has been replaced by improved ultrasound resolution of fetal
anatomy.
MRI
As a fetal imaging technique, magnetic resonance imaging is being actively investigated, but its clinical application has been limited by fetal movements
RADIOGRAPHY
This has been largely replaced by ultrasonography for the detection of anatomic lesions but it is still indicated for certain disorders particularly osteochondrodysplasias, in the late second and third trimesters.
MATERNAL SERUM SCREENING FOR TRISOMY 21
● A number of studies have demonstrated that low maternal serum a-feto protein (MSAFP) concentrations are associated with an increased risk for trisomy 21 and perhaps other aneuploidies in the fetus.
● Results of MSAFP screening may be used along with maternal age to revise an individual patient’s risk of having a child with Down syndrome.
● A combination of laboratory tests (MSAFP, estriol and HCG), in conjunction with maternal age, has been used to evaluate the risk of Down syndrome and other chromosome abnormalities such as trisomy 18 and 45, X.
● More recently, decreased maternal serum levels of unconjugated estriol (UEs) and increased human chorionic gonadotropin (HCG) have also been associated with Down syndrome in the fetus.
● Of these, HCG is the most sensitive marker for detection of Down syndrome in the fetus.
FETAL CELL SEPARATION FROM MATERNAL BLOOD SAMPLES
● Separation of fetal cells from maternal blood samples obtained during early pregnancy is being actively investigated.
● In the future, these samples may provide a source of material for screening or diagnosis of chromosome abnormalities, particularly using fluorescent in situ hybridization or other genetic disorders by amplifying DNA.
PREIMPLANTATION GENETIC DIAGNOSIS
● Techniques to perform testing of cells obtained from biopsy of early cleavage stages or blastocysts of pregnancies conceived through in vitro fertilization are
currently being developed.
● These techniques may allow selective transfer into the uterus and implantation of those pregnancies that are not affected by a specific disorder.
● Both laboratory techniques for testing and biopsy are still experimental.
Diagnosis of Genetic Diseases
● Requires examination of genetic material (i.e., chromosome and genes).
● Two general methods are employed:
○ Cytogenetic analysis
○ Molecular analysis
● Cytogenetic analysis requires karyotyping
What should be offered to all patients who are at risk of cytogenetically abnormal progeny?
Prenatal chromosome analysis
Prenatal chromosome analysis can be performed on cells obtained by:
- amniocentesis,
- on chorionic villus biopsy,
- or on umbilical cord blood
Prenatal chr. analysis
Some important indications are the following:
○ Advanced maternal age (>34 years) because of greater risk of trisomies
○ A parent who is a carrier of a balanced
reciprocal translocation, robertsonian
translocation, or inversion (in these cases the gametes may be unbalanced, and hence the progeny would be at risk for chromosomal disorders)
○ A previous child with a chromosomal
abnormality
○ A parent who is a carrier of an X-linked genetic disorder (to determine fetal sex)
Postnatal chromosome analysis is usually performed on:
peripheral blood lymphocytes
Postnatal chromosome analysis
Indications are as follows:
○ Multiple congenital anomalies
○ Unexplained mental retardation or
developmental delay
○ Suspected aneuploidy (e.g., features of Down syndrome)
○ Suspected unbalanced autosome (e.g,
Prader-Willi syndrome)
○ Suspected sex chromosomal abnormality (e.g. Turner syndrome)
○ Suspected fragile X syndrome
○ Infertility (to rule out sex chromosomal
abnormality)
○ Multiple spontaneous abortions (to rule out the parents as carriers of balanced translocation; both partners should be evaluated)
MOLECULAR DIAGNOSIS/ANALYSIS
● Medical applications of recombinant DNA technology have come of age.
● With the rapid transfer of technology form “the bench to the bedside,” it is now clear that DNA probes can be powerful tools for the diagnosis of human disease, both genetic and acquired
Molecular diagnostic techniques have found application in virtually all areas of medicine.
These include the following:
○ Detection of inherited mutations that underlie the development of genetic diseases either prenatally or after birth.
○ Detection of acquired mutations that underlie the development of neoplasms
○ Accurate diagnosis and classification of
neoplasms, especially those that originate in the hematopoietic system.
○ Diagnosis of infectious diseases, including human immunodeficiency virus (HIV) disease.
○ Determination or relatedness and identity in transplantation, paternity testing, and forensic medicine
MOLECULAR DIAGNOSIS/ANALYSIS
● Many genetic diseases are caused by subtle changes in individual genes that cannot be detected by karyotyping
● Traditionally the diagnosis of single-gene disorders has depended on the identification of abnormal gene products (e.g, mutant hemoglobin or enzymes) or their clinical effects.
○ Such as anemia or mental retardation
(e.g.phenylketonuria).
● Now, it is possible to identify mutations at the level of DNA and offer gene diagnosis for several mendelian disorders.
● The use of recombinant DNA technology for the diagnosis of inherited diseases has several distinct advantages over other techniques.
● It is remarkably sensitive.
○ The amount of DNA required for diagnosis by molecular hybridization techniques can be readily obtained from 100,000 cells.
○ Furthermore, the use of PCR allows several million-fold amplification of DNA or RNA, making it possible to use as few as 100 cells or 1 cell for analysis.
● Tiny amounts of whole blood or even dried blood can supply sufficient DNA for PCR amplification.
● DNA-based tests are not dependent on a gene product that may be produced only in certain specialized cells (e.g. brain) or expression of a gene that may occur late in life. Because virtually all cells of the body of an affected individual contain the same DNA, each postzygotic cell carries the mutant gene.
● These two features have profound implications for the prenatal diagnosis of genetic diseases because a sufficient number of cells can be obtained from a few milliliters of amniotic fluid or from a biopsy of chorionic villus that can be performed as early as the first
trimester.
There are two distinct approaches to the diagnosis of single-gene diseases by recombinant DNA technology:
○ Direct detection of mutations
○ Indirect detection based on linkage of the diseased gene with a harmless “marker gene” - (linkage analysis)
MANAGEMENT OF GENETIC DISEASES
● The objectives of treatment of genetic disorders are to obtain and maintain optimal health in both the individual and the race.
● With recent advances in technology, the curative aspect of genetic diseases based on a genuine understanding
of the disease mechanisms have been made possible for many of the inherited biochemical diseases transmitted through single gene inheritance, affecting only a single gene enzyme.
● On the other hand, a great number of diseases determined by multifactorial inheritance or by chromosomal aberrations are less amenable to such maneuvers.
● These diseases are usually expressed in full prenatally, leaving little for the physician to accomplish postnatally.
● In such cases, reducing the number of births of affected individuals by genetic counseling until curative means are achieved, remains the procedure of choice.
● There are two areas which generally embrace technology designed to treat genetic disorders or to treat deleterious effects.
○ The first consists of measures to modify the environment in which the mutant organism lives. The potentially deleterious effect of the mutation is thus offset and the individual may lead a reasonably normal life.
○ The second area of concern introduces the less developed but intriguing genetic engineering whereby the genetic apparatus is attacked directly.
● To summarize, measures to manage genetic disorders include:
○ Controlling the external environment;
○ Regulating ingested food,
○ Modifying internal environment;
○ Removing diseased tissues or introducing normal donor issue;
○ Genetic engineering; and
○ Preventing the conception or birth of affected individuals by genetic counseling
CONTROL OF EXTERNAL ENVIRONMENT
● Protection from mechanical trauma is required for patients with fragile bones, weak muscles, lax joints or bleeding tendencies.
● Protection from thermal injury is necessary in rare genetic conditions like anhidrotic ectodermal dysplasia where hyperthermia may kill or cause irreversible brain damage.
● Ultraviolet radiation is harmful to albinos where pigmentary protection against ultraviolet rays is lacking, thus the specific protection against it.
● Protection against infectious disease is the first line of defense in patients with deficiencies of the cellular or humoral system
REGULATION OF INGESTED FOODS
● In metabolic disease that can be modified by food, the aim of treatment is to sustain nutrition while suppressing and preventing deficiencies.
● Diet therapy includes:
○ Augmentation intake of protective foods,
○ Total exclusion of nonessential foods that are toxic;
○ Restricted intake of essential foods that are toxic;
○ Supplementation of foods that are needed in greater than normal amounts;
○ Substitution of synthetic foods to fulfill the nutritional needs of an individual for whom a natural food is toxic; and
○ Oral replacement of missing enzymes when this is feasible.
Augmentation
● This is increased intake of certain foods, indirectly protecting against the complications of some metabolic diseases.
● Increased water intake, for example, protects patients (with sickle cell disease from dehydration which increases the tendency for rid calls to sickle.)
● In cystinuria, water reduces the danger of cystine crystallization in the urine.
Exclusion
● Dietary glucose malabsorption, fructose in fructose intolerance and galactose in galactosemia can cause severe metabolic disturbances that may lead to brain damage.
● This is prevented by strict dietary exclusion of the sugar concerned from the diet since early infancy.
● The same would apply to cholesterol and saturated fats which are non- essential foods that exacerbate the
hypercholesterolemia of type lI lipidemia.
Restriction
● When the toxic food in a metabolic disease is also an essential nutrient, depriving a palient of the nutrient can be complicated by malnutrition.
● The intake of the potentially toxic food should therefore not exceed its toxic threshold.
● Restriction of phenylalanine intake in phenylketonuria and galactose intake in galactosemia are examples of therapy in these inborn errors of metabolism.
Co-enzyme Supplementation
● Supplementation of specific foods will counteract the deficiency states that complicate certain genetic diseases.
● Supplementation with vitamins is useful in abnormalities of transport of vitamins leading to deficiency states.
● These are treatable by giving larger doses of vitamins and by parenteral administration.
● Administration of vitamins stimulate compensatory reactions to counteract some biochemical derangements.
Substitution
● Synthetic amino acid mixtures as a substitute for protein in phenylketonuria, maple syrup urine disease or propionic acidemia has preserved life, health and intellect of a number of patients
Replacement
● Restoring enzymes by oral therapy is possible only for enteral digestive enzymes such as in cystic fibrosis,
and in pepsinogen or lactase deficiency because few molecules are absorbed from the intestine before they are degraded by digestive enzymes.
MODIFICATION OF THE INTERNAL ENVIRONMENT
Chemical Modification
● Lowering the concentration of metabolites by chemical modification of the internal environment is the treatment of choice for some conditions with or without simultaneous dietary restriction.
● Hyperammonemia is aggravated by ammonia released by urease-forming microorganism in the colon, so
prevention of constipation is a prevention in any condition leading to hyperammonemia
● Chemical removal of copper retained in Wilson’s disease has been successful in preventing liver and brain damage complicating this disorder.
● Chelating agents like D-penicillamine flush sufficient copper out into the urine to deplete the excess copper stores in the patient’s tissues.
Pharmacological Modification
● Some drugs may be uniquely toxic to persons with certain genetic disorders.
● Glucose-6-phosphate dehydrogenase deficiency renders affected individuals prone to hemolytic crises whenever they are exposed to drugs that are oxidizing
agents.
● For the many conditions in which these drugs are harmful, the responsible drugs should be avoided.
● Pharmacological treatment of genetic disorders can be achieved by stimulation or suppression of physiological processes.
● Stimulation of the cellular immune system by transfer factor in some of the genetic immunodeficiency diseases has met with some success.
● Suppression of antihistamines has reduced attacks of angioneurotic edema due to C1 esterase deficiency.
Endocrinological Modification
● Replacement therapy of primary hormone deficiency is well recognized for genetic abnormalities of endocrine function.
● Timely administration of thyroid hormone in congenital hypothyroid states is essential for preservation of
normal brain development.
● Inborn errors of adrenal corticosteroid biosynthesis can lead to interruption of the feedback control of corticosteroid metabolism, with overproduction of androgenic by-products;
○ Therefore, in this disorder, replacement therapy also serves to suppress virilization.
SURGERY
● Neurosurgical shunt procedures can prevent or alleviate congenital hydrocephalus.
● Surgical shunting procedures on the gastrointestinal tract and portal venous system include colonic by-pass for the hyperammonemia syndromes and ileal by-pass to decrease fat absorption in Type Il lipidemia.
● Organ transplantation has been used extensively in the treatment of kidney disorders.
● Liver transplantation offers a more direct approach to enzyme deficiency disease, or to inborn errors of
metabolism.
● One disorder that has been cured metabolically by liver replacement in spite of its unknown pathogenesis
is Wilson’s disease, characterized by widespread accumulation of copper in the tissues.
● Serum ceruloplasmin, which is almost always very low in patients with Wilson’s disease, is restored to normal within a few days after transplantation.
● At least five other inborn errors with known and specific enzyme deficiencies have been treated by liver transplantation:
○ Alpha-antitrypsin deficiency
○ Tyrosinemia
○ Niemann-Pick disease
○ Type IV glycogen storage disease
○ Crigler-Naijar syndrome
● Transplantation of bone marrow, thymus and fetal liver have been successfully done for the cellular, metabolic and immunologic correction of various congenital and inherited disorders.
● Bone marrow transplantation has been used to correct inherited disorders resulting from the defective function of marrow-derived cells.
● It has also been used to reverse the cellular and immune defects in patients with various disorders including Wiscott-Aldrich syndrome, chronic
granulomatous disease (CGD), and Franconi’s anemia, etc.
● All centers doing bone marrow transplantation are able to cure aplastic anemia and fully correct the cellular
deficits by the same procedure.
● Even the worst form of Leukemia, acute myeloid leukemia occurring in children and young adults, is now often curable by marrow transplantation.
● The problem that often occurs, however, is host vs. graft reaction which limits the availability of donors and
contributes heavily to mortality when HLA-matched sibling donors are used.
● This has been benobiated in recent year by preparing the bone marrow so it will not initiate a graft vs. host reaction.
GENETIC ENGINEERING
● This refers to the possibility, based on work with bacteria and viruses, that the genetic apparatus can be altered in a direct fashion.
There are 3 approaches in genetic engineering:
a. Protein (enzyme) replacement
b. Enzyme induction or repression
c. Transformation or transduction of the gene
Protein Replacement
● This is the most accessible and best known area of application. It is exemplified by the use of injectable
insulin in diabetes mellitus, the diabetic today enjoys reasonable health and a relatively long life.
● Other examples of protein replacement are gamma globulin in congenital agammaglobulinemia and
antihemophilic globulin in hemophilia.
● All have in common a natural source of specific protein; in all instances, the catalyst can also reach its normal site of action. Any future advances in the techniques of protein replacement will depend on the twin criteria of exogenous availability and endogenous accessibility.
● At present, one can expect two sources of protein supply to be exploited. Synthetic sources will be one type, for which a primary structure of the protein to direct synthesis.
● Natural sources of protein are the alternate choice, of which species specificity will be of great importance if
sustained benefit from an exogenous supply is expected in the human host. At present an animal source for digestive proteins and insulin seems satisfactory and efforts to obtain homologous plasma are easily met.
Enzyme Induction and Repression
● Stimulation of increased formation of a deficient enzyme is possible for some inductible microsomal enzymes. In
Gilbert’s unconjugated hyperbilirubinemia, barbiturates
induce measurably increased glucuronyl
transferase activity, Phenobarbital also caused symptomatic and chemical improvement in some of the familial
cholestases.
● Inhibition of enzyme expression by preventing its activity with substrate analogues or by feedback suppression
with end products is the basis for the treatment of hyperuricemia with allopurinol of hypercholesterolemia
with clofibrate, and of congenital hyperplasia of the adrenals with mineralocorticoids.
Transformation and Transduction of the Gene
● Transduction is a form of recombination of genetic material in bacteria in which a bacteriophage transfers genetic information from one bacterium to another.
● Transformation is a form of recombination of genetic material in bacteria in which a bacterium incorporates DNA extracted from other bacteria into its own genetic material.
● The success of experiments in genetic information and transduction in microorganisms has encouraged some
to hope that techniques of directed mutation can become available to benefit man.
● Gene therapy by deliberate introduction of viral genes to man has been tried only once, in two sisters with
argininemia in an attempt to transduce a viral gene for arginase. The experiment generated considerable controversy about its advisability and results were not specific.
● One example exists where transcription to advantage in man. This is the control of herpes keratitis through the use of a uridine analogue placed on the cornea. The analogue, when incorporated by the replicating virus,
blocks further viral replication and thus halts the infection.
● The recombinant technique was made possible by the 1970 discovery of a new class of enzymes known as the
“site-specific restriction endonucleases”. These enzymes can, like molecular scissors, cleave DNA into specific pieces.
● In this technique, DNA from an organism is broken up with restriction enzymes and sliced into compatible sites
in viruses or bacterial cell elements known as plasmid.
● Transplanted into “host” cells, the hybrid DNA, now containing genetic information from another organism,
can replicate along with its host cell.
● One of the potential uses of this is a possible “microbial drug machine” that can make possible the large-scale
production of vital but hard to obtain proteins including insulin, growth hormones, blood clotting factors and
antibodies.
GENE THERAPY
● Will use normal genes to replace or supplement a defective gene, or to bolster immunity to disease.
Currently, gene therapy research is primarily concerned with establishing the safety of this approach, rather than
the effectiveness of the treatment.
● While there are hundreds of clinical trials and studies in progress, so far no cures have resulted.
GENETIC COUNSELING AND EDUCATION
1) Should begin before prenatal testing and should include:
● Description of the test
● Types of disorders that will be screened
● Limitations of the screening
● Exploration of what parents will do with the information Indication of when results will be available
2) Indications for Genetic counseling
● Advanced maternal (>35) or paternal (>50) age
● Consanguinity
● Previous history of a child with birth defects or a genetic disorder
● Personal or family history suggestive of a genetic disorder
● High-risk ethnic groups; known carriers of genetic mutations
● Ultrasound or prenatal testing suggesting a genetic disorder.
3) Refers to a communication process that deals with human problems associated with the occurrence or risk of a genetic disorder in a family.
4) Genetic risk assessment can be complex and often involves elements of uncertainty. Counseling therefore includes genetic education as well as
psychosocial counseling.
5) The genetic counselor will do the following:
● Gather and document a detailed family history
● Educate the patient about genetic principles related to disease risk, both for themselves and others in the family
● Assess and enhance the patient’s ability to cope with the genetic information gathered.
● Discuss how non genetic factors may relate to the ultimate expression of disease.
● Address medical management issues
● Assist in determining the role of genetic testing for the individual and family
● Ensure that the patient is aware of the risks, benefits, and limitations of the various genetic testing options.
● Refer the patient and other at-risk family members for additional medical and support services, if necessary.
6) Multidisciplinary team to provide broad-based support and medical care for those at risk and their family members;
● Composed of: medical geneticists, special physicians, genetic counselors, nurses, psychologists, social workers, and biomedical ethicists.
What are genetic counselors and what do they do?
● Genetic counselors are health care professional with specialized graduate degrees and experience in medical genetics and counseling.
● Genetic counselors work as members of health care teams providing information and support to individuals
or families who have genetic disorders or may be at risk for inherited conditions.
● Genetic counselors will help:
a. Assess the risk of a genetic disorder by
researching a family’s history and evaluating medical records
● Weigh the medical, social and ethical decisions surrounding genetic testing
● Provide support and information to help a person make a decision about testing
● Interpret the results of genetic tests and medical data
● Provide counseling or refer individuals and families to support services
● Serve as patient advocates
● Explain possible treatments or preventive measures
● Discuss reproductive options
How do I decide whether I need to see a geneticist or
other specialist?
● A genetics counselor may refer you to a geneticist– a medical doctor or medical researcher – who specializes in your disease or disorder. A medical specialist has completed a fellowship or has other advanced training in medical genetics.
● While a genetic counselor may help you with testing decisions and support issues, a medical specialist will make the actual diagnosis of a disease or condition.
Many genetic diseases are so rare that only a geneticist can provide the most complete and current information
about your condition.
● Along with a medical geneticist, you may also be referred to a physician who specializes in the type of disorder you have. For example, if a genetic test is
positive for colon cancer, you might be referred to annoncologist. For a diagnosis of Huntington’s disease, you
may be referred to a neurologist.
ETHICS OF COUNSELING
● Genetic Counseling is the process of conveying risk information about genes. Definition of genetic counseling agrees that it is the communication of information about diagnosed genetic conditions, in a way that allows the person to make as autonomous
decision as possible from that information while safeguarding the emotional and ethical character of the
persons who ask for the conclusion.
● Chadwick defines genetic counseling as advising adults preconception of the probability of their conceiving a child who will suffer from a genetic disorder
and advising adults, post-conception, by the use of some method for fetal screening as to whether or not a
fetus is suffering from a genetic disorder and altering adults to their options. This is called non-directive.
Genetic Counseling
● Is a communication process concerning the occurrence and risk of recurrence of genetic disorders within a
family. The aim of such counseling is to provide the consultants with the fullest comprehension of all the implications of the disease in question and all the possible available options.
● In particular, the counseling should provide a clear estimate of the risk about which the consultant inquires and such risk explained in a manner appropriate to the patient’s educational level. Some centers provide information on treatment and refer patients to institutions responsible for the education, medical supervision, or hospitalization of afflicted individuals,
while genetics centers, in general, will help in establishing a precise diagnosis in the patient. Thus, a further goal is helping families through their problems,
their decision making and their adjustments.
● Genetic counseling is increasingly becoming an important tool in pediatrics. A mother who has had a defective baby would want to know her chances of having another child with the same defect. This is especially so in cases of congenital malformation.
● A family with a known genetic disease plaguing its members would be interested to know the chances of the disease appearing if a member marries and has children. In some cases, advisability of marriage itself becomes a problem.
● In general, a geneticist or physician may be able to answer the above questions adequately although a
knowledge of the principles of genetics discussed at the beginning of the chapter will be of much value during
the interview.
● The role of the counselor is to assist consultants in recognizing their personal major priorities before any decision making. It is preferable to give to parents
whatever is known of the disorder in question including cause, clinical course and theoretical possibilities of
transmission, then let them make their decision on the matter. The final decision on whether to have another
child or not depends on many factors including the desire to have children, present size of the family, economic status, religious factors and ages of the
mother and father.
Accurate Diagnosis
● Genetic counseling should begin with the establishment of a genetic mechanism involved and for the estimation of the risk, the counselor must see to it that it is of the highest scientific quality. It is only prudent
in the absence of an accurate diagnosis, to refrain from guessing the recurrence risk.
● It is important to separate truly genetic conditions from those due to environment because the prognosis of the latter as far as subsequent children are concerned is generally much better. For example, cretinism occurring as a result of iodine deprivation of the mother can totally be prevented in future sibling with specific treatment of the mother. However, it is a known fact the same type of malformation may be the result of either a genetic factor, environmental factor, or both. Examples
of these are certain forms of congenital heart disease, congenital cataract, cleft lip and palate, microcephaly, and others.
● In order to determine more precisely the probable cause of the defect or anomaly, a meticulous history taking
should be included in the analysis of every propositus.
This, together with a good physical examination and laboratory examinations where necessary, may help the doctor in the counseling.
● If there is a history of similar disease or defect in another member of the family, it is better to prepare a pedigree chart of the family, such a chart indicating the patient as the propositus and including all members of the family tree. Since the purpose of the pedigree is to present history visually, some uniformity in recording is necessary so that pedigree from various sources can
be read quickly and informatively. The index case or propositus should be indicated first on the pedigree with
an arrow. The patient’s siblings are then recorded chronologically on the same line.
● All pregnancies, including miscarriages, abortions, or stillbirths, are listed. Then the parents, their brothers, sisters and children and the grandparents are added to the pedigree. The family history usually looks into 3 generations.
● Where a disease existing in a family is known to be usually transmitted by dominant inheritance and where the pedigree of the family suggests the same type of transmission, it can be told that the chances of a mating to be affected, where an affected member marries a
normal partner, will be 1:1.
● However, if the family member does not show any symptoms of the disease, it can be predicted that all children will be normal provided the disease in question has complete penetrance and expressivity. In a disease with low expressivity, laboratory tests may help the clinician in analyzing the situation. Genetic counseling in a family with a recessive trait is more difficult. A recessive trait is more difficult.
● A recessive trait can be handed down from generation to generation without being manifested. When a person heterozygote, the trait may appear but the family may think of it as a fresh mutation. If this trait is not a fresh mutation, one may expect every sibling of the affected member to have a one-fourth chance of having the disease; that is, the expected ratio of the affected to
clinically normal children, two-thirds would be expected to be gene-carriers, like their parents.
● If two affected partners (such as twodeaf-mutes) mate, all children will be affected if both of them carry the identical mutant recessive gene. If a recessive gene is rare, the chance that two unrelated carriers of the gene
will meet is remote. The rarer the gene, the higher will be the frequency of consanguineous marriages among
the parents of affected patients. Heterozygous carriers of the abnormal by appropriate biochemical tests.
● A man who manifests a sex-linked recessive trait, if married to a normal woman, will have sons who are normal but daughters who are all carriers. The sons of these female carriers will have a 1:1 chance to have the disease; those who will not manifest the disease will not transmit it. The daughters will all appear normal but have 1:1 chance of being carriers like their mother.
● Carriers for sex-linked recessive genes can often be recognized by specific biochemical or clinical tests. Test values for heterozygous may be intermediate between normal and abnormal.
● Mention has to be made about chromosomal aberrations. Since down syndrome is a common and well studied disease, more data are available upon
which to base genetic counseling as compared to other aberrations. The chance for the birth of a child with
Down syndrome does not rise until the mother is 30 years of age, and even at over 45 years, the mother’s risk does not exceed 3 percent.
● Similarly, it is known that irrespective of maternal age, the chance for another affected child after the birth of an infant with Down syndrome is low (about 1 to 2 percent).
Therefore, if chromosome studies are not available, the low risk for recurrence may be used. Table 9.8 gives the risk of Down syndrome according to maternal age. An appropriate risk for non-trisomy 21 is 1:200 for mothers under 35 years. Above this age, the risk is no different
from the risk for the general population.
● Among affected infants born to mothers under 30 years of age, the cause may not be trisomy but translocation involving chromosome No. 21. A parent carrying a 13-15/21 translocation has thirty percent chance of having an affected child.
● Some developmental anomalies are known to be caused by detectable chromosomal abnormalities or
by mutant genes. However, in most malformations, interaction between unidentified genetic, and environmental factors seem to be operative. Since exact
genetic factors involved are unknown, genetic prognosis must rely on studies of empiric recurrence risks which have been collected by numerous investigators. Table 9.9 gives a range of recurrence risk in medical genetics as given by Motulsky and Hecht. It summarized the range of different transmission and classify risk of recurrence in siblings as either total,
high, moderate, and low. Most congenital malformations of unknown etiology are classified under low risk
conditions.
Timing in Genetic Counseling
● Genetic counseling in the past has invariably been offered after the birth of a child with genetic disease or congenital malformation. Recent advances in genetic technology, mainly antenatal diagnosis and carrier detection, now enable the physician to counsel prospectively many patients at risk.
● Today in a variety of situations, prospective advice is both critical and important. Ideally, counseling should be
offered prior to conception or marriage. Those of eugenic inclination hold that genetic counseling should be offered early enough to provide options for those choosing their mates.
● Again the time to initiate counseling is not in mid-pregnancy when some carrier tests may be unreliable. It is also crucial to bear in mind the inability of parents anguished by recent loss to assimilate even the essence of counseling offered. When parental counseling immediately after such loss is important in the prevention of further pregnancies, there is a need to repeat the counseling offered some months later.
Available options to couples
● The role of the counselor in genetic counseling is to assist consultants in recognizing their personal priorities
crucial to their decision making. The counselor should not tell consultants what they ought to do or how they
should act, but help them recognize and anticipate issues and problems before they occur. This non-directive approach is endorsed by the WHO expert committee on genetic counseling.
● The intrinsic danger of directive approach is the insinuation by the counselor of his own religious, social,
racial, or eugenic dictates into the counseling offered. By helping to place the concerns of each family in relative perspective, their decision-making process is clarified, enhanced, and enriched.
The decision making process is simplified for the couple by preparing a list of the available options and discussing the merits of each option.
These options include:
- Have no further children- the advantage is that the couple will avoid the possibility of having another affected child. The disadvantage is that they may not be able to complete the family they
desire. - Take a chance- this refers to identification of a genetic disorder using a series of tests anytime during the pregnancy.
- Adoption- this will allow couples to complete their family without having a child with the disorders for which they are at risk. A disadvantage is that some couples find it difficult to accept a child that is not their own especially when infertility is not the basis for considering abortion.
- Divorce- for countries that allow divorce
- Artificial insemination- the husband may feel a closer bond to such a child than he might feel for an adopted child. The disadvantage and time-consuming undertaking.
- Termination- in most countries this is one of the available options presented to a couple during the process of genetic counseling. Termination is allowed before 24 weeks of gestation in the
majority of cases. In the Philippines, however, this is not an option.
HUMAN GENOME PROJECT
● Goal: to identify and sequence all of the DNA in human chromosomes.
● Project is initiated in 1990 in the U.S. with government funding and rapidly grew into an international consortium of academic centers and drug companies in
China, France, Germany, Japan, United Kingdom, and the U.S.
● Provide scientists with a detailed bluprint of our complex genetic code- large computer databases of genetic
information enable scientists to look for patterns and relationships among the actions of different genes.
● Has found out that the number of genes on the human genome is only about 31,000.
The DNA of a suspect
● An individual’s DNA is as unique as a set of fingerprints, and a DNA match can be used in a courtroom as evidence connecting a person to a crime.
● An international collaboration of scientists, governments, and drug companies from around the world. Scientists working on these projects have developed detailed maps that identify the chromosomal locations of the estimated 31,000 human genes. The
vast databases emerging from the project help scientists study previously unknown genes as well as many genes all at once to examine how gene activity can cause disease.
● Scientists expect that the project will lead to the development of new drugs targeted to specific genetic disorders.
Identification of genetic information
● Belonging to a specific individual has received a great deal of press coverage lately. Profiles (aka “DNA Fingerprints) are compiled from the results of DNA
testing for one or more genetic markers to identify unique characteristics of an individual.
● Information from genetic testing can affect the lives of individuals and their families. In addition to personal and
family issues, genetic disease or susceptibility may have implications for employment and insurance.
Therefore, careful consideration in the handling of this information is very important.
Critical issues include:
a. Privacy - the rights of individuals to maintain privacy. Some genetic tests are required or strongly encouraged for developing fetuses and newborn babies. If an infant is found to be a carrier or likely to develop or be affected by an
inherited disease, these findings may affect the future employability or insurability of the individual.
b. Informed consent - obtaining permission to do genetic testing. One must have knowledge of the risks, benefits, effectiveness, and alternatives to testing in order to understand the
implications of genetic testing.
c. Confidentiality - acknowledgement that genetic information is sensitive and access should be limited to those authorized to receive it. Future access to a person’s genetic information also
should be limited.
Gene Cloning
● Introduction of a DNA molecule into a replicating cell permits the amplification (that is, production of many copies) of the DNA. In some cases a single DNA
fragment to be cloned can be isolated and purified.
More commonly, to clone a nucleotide sequence of interest, the total cellular DNA is first cleaved with a specific restriction enzyme, creating hundreds of
thousands of fragments. The individual fragments therefore cannot be isolated, so all the resulting ‘insert’
DNA fragments are linked to form hybrid molecules.
● Each hybrid recombinant DNA molecule conveys its insert DNA into a single host cell, for example, a bacterium, where it is replicated (or amplified).
● As the host cell multiplies, it forms a clone in which every bacterium carriers copies of the same insert DNA is eventually released from its vector by cleavage (using the appropriate restrictions endonuclease) and is
isolated. By this mechanism, many identical codes of the DNA of interest can be produced.
PCR
● The polymerase chain reaction (PCR) is a test tube method for amplifying a selected DNA sequence and does not rely on the biologic cloning method.
● PCR permits the synthesis of millions of copies of a specific nucleotide sequence in a few hours. It can amplify the sequence, even when the targeted
sequence makes up less than one part in a million of the total initial sample.
● The method can be used to amplify DNA sequences from any source- bacterial, viral, plant, or animal.
Advantages of PCR
● The major advantage of PCR over cloning as a mechanism of amplifying a specific DNA sequence are sensitivity and speed. DNA sequences present in only trace amounts can be amplified to become the predominant sequence.
● PCR is so sensitive that DNA sequences present in an individual cell can be amplified and studied.
● Isolating and amplifying a specific DNA sequence by PCR is faster and less technically difficult than traditional cloning methods using recombinant DNA techniques.
Applications of PCR
● PCR has become a very common tool for a large number of applications. These include:
a. Comparison of a normal cloned gene with an unclone mutant form of the gene. PCR allows synthesis of the mutant DNA in sufficient quantities for sequencing protocol without laboriously cloning the altered DNA
b. Detection of low-abundance nucleic acid sequences. For example viruses that have a long latency period, such as HIV, are difficult to detect at the early stage of infection using conventional methods. PCR offers a rapid and sensitive method for detecting viral DNA sequences even when only a small proportion of cells is harboring the virus.
c. Forensic analysis of DNA samples. DNA isolated from a single human hair is sufficient to determine whether the sample comes from a specific individual.
● Within the DNA string, some areas are the same for all humans. These areas are said to be “conserved”. Other areas tend to vary across people. These areas are said to be polymorphic or variable. The variable regions (V) are usually interspersed among the conserved or
constant regions (C) as shown in the figure below.
● In forensic DNA tests, PCR is used to identify and copy one or more variable regions of DNA. The tests use “primers” (indicated by small arrows below) that
identify constant regions adjacent to the variable region of interest.
● The primers are short pieces of DNA (similar to the probes used for RFLP analysis) that are complementary to their target sequences. The primers serve as the starting points for copying of variable regions of the DNA sequence. The actual copying occurs in a test tube
which is placed in a device called a thermal cycler.
● The thermal cycler goes through a series of heating and cooling cycles. In each cycle, each DNA fragment from the target region is duplicated. As the cycles
continue, the quantity of target DNA increases exponentially.
APPLICATION OF DNA TESTING
● The purpose of DNA typing in forensic medicine is to match a sample from the crime site with a suspect.
Technically, application of the techniques described above do not actually determine whether the sample came from the suspect. Rather, statistical analysis of the test results yield a probability that the sample did not come from the suspect, and with DNA typing, that probability can be so miniscule as to be certain.
● Importantly, DNA testing has proven to be as powerful for exonerating suspects as it has for convincing them.
Indeed, in about one in three cases reported by FBI laboratories, DNA testing proved that the current suspect could not have committed the crime, which in many cases was followed by apprehension and conviction of the true perpetrator.
● A great diversity of criminal detection has benefited from DNA testing, and it has been especially valuable in solving rape and murder cases.
● Additional examples include robbery, assault, kidnapping, car accidents, extortion, and blackmail. It also has been successfully applied to parentage
determination and useful in settling certain immigration disputes that hinges on proving family relationships.
TECHNIQUES FOR DNA TESTING
● DNA typing is performed by demonstrating differences in length for specific DNA sequences. This can be done by digestion of DNA with restriction enzyme(s), followed by Southern blot hybridization using a probe for the polymorphic site.
● Polymerase chain reaction (PCR) techniques are becoming widely applied to the same task, and have several advantages over Southern blotting- for example, much less DNA is required and in many cases, typing can be done using partially degraded DNA.
● For PCR analysis, the primers are designed to flank the VNTR locus and the size of the PCR product is dependent on the number of repeats.
● The general term “DNA fingerprinting” is used to describe all these procedures for characterizing VNTRs, RFLPs, and other sequence polymorphisms.
● DNA has a number of real advantages over serological methods such as blood typing and HLA analysis for use in forensic investigations:
a. Unsurpassed discriminatory potential:
complete blood group in testing allows
discrimination of one person in several thousand and HLA typing one in several million. DNA typing can routinely provide exclusion probabilities on the order of one in billions.
b. Exquisite sensitivity: Standard DNA typing can be conducted with DNA extracted from the roots of a few hairs. In contrast to proteins, DNA can be amplified, and by using polymerase chain reaction methods, even smaller sample sizes are adequate. One important consequence of this great sensitivity is that it allows rather small
samples to be split and submitted for testing to more than one laboratory, which can identify the laboratory errors more commonly and serves to nullify objections that laboratory errors were
committed.
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old. Overall, DNA is remarkably robust as a sample for forensic testing, which, for example, has allowed it to be used on skeletonized remains for identification of soldiers missing in action. ● DNA typing has often been portrayed in the media and the courtroom as a controversial technology, largely because it has been so characterized by many defense attorneys. ● When DNA evidence demonstrates that the odds that someone other than your client committed the crime are one in a billion, there is really nowhere else to go but to attack the basic technology of DNA testing. With increased experience and standardization of testing methods, these assaults are heard less frequently. ● There have been some scientifically legitimate criticisms of DNA testing, based on concerns about allele frequencies in certain populations. These frequencies are used in calculation of probability of identification. ● The worry was that the chances of a random match may be higher than stated because the database used was inappropriate for the subpopulation of people containing the suspect. For example, the frequency of a specific allele under test may be 4% in Asians instead of 1% as it is in Northern Europeans. ● However, most experts concluded that such differences in allele frequency have rather little impact on the final diagnosis- it makes little difference whether the probability of innocence or guilt is one in 10 million or one in 100 million. ● Nevertheless, the basic premise of the argument is valid and has been incorporated into recommendations about how forensic DNA testing be conducted and interpreted. ● Ethical issues on genetic advancements: 1. Genetically engineered organism could harm people or the environment 2. Genes that affect characteristics other than those responsible for disease may be modified by some scientists 3. Discriminatory practices on those with undesirable genetic characteristics 4. Misperceptions that a person’s genes determine all aspects of a person’s life, including health and behavior.
