Tony Wynshaw-Boris, M.D., Ph.D.

SOM 204 Genetics Lecture 7

Clinical Cytogenetics
CBB Human Genetics Dr. Anthony Wynshaw-Boris

Clinical Cytogenetics
Began ~35 years ago with discovery that somatic cells in humans contain 46 chromosomes (1956) Simple procedurehypotonic treatment for spreading the chromosomes of individual cells Ability to microscopically examine and study chromosomes in single cells rather than tissue sections Between 1956 and 1959: visible changes in the number or structure of chromosomes could result in a number of birth defects Down, Turner, Klinefelter, and Cri du Chat syndromes Chromosome disorders: a large proportion of fetal loss, congenital defects and mental retardation Number of clinical indications for chromosome analysis

Clinical Indications for Chromosome Analysis

Problems of early growth and development Stillbirth and neonatal death Fertility problems Family history of chromosome abnormality Neoplasia Pregnancy in woman of advanced age (>35 years)

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Chromosome Identification
G bands (from Giemsa staining): most widely used banding technique. Q bands (quinacrine mustard and other fluorochromes): identical to G bands. Particularly useful for identifying the Y chromosomes. R bands ("reverse" banding): opposite to that in G and Q banding. Light bands produced on G and Q banding are dark on R banding, and dark bands on G and Q banding are light on R banding. T bands: specific for telomeric regions of the chromosome. C bands: constitutive heterochromatin located primarily on the pericentric regions of the chromosome.

Ideogram of G-Banded Metaphase Chromosomes (400 Bands)

G-Banded Metaphase Chromosomes (550 Bands)

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

High Resolution Banding X Chromosome

Molecular Cytogenetics: FISH (Fluorescence In Situ Hybridization

T FISH: Telomeres

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Spectral Karyotyping (SKY)

SKY Analysis- Medulloblastoma

Prader-Willi Syndrome 15q11-13 Deletion

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Chromosome Abnormalities
Abnormalities of chromosome number (diploid is normal, or euploid)
Triploidy and tetraploidy: multiple of haploid (3n or 4n) Aneuploidy: extra or missing chromosome

Abnormalities of chromosome structure

Unbalanced: more or less than normal complement
Deletions, duplications, marker chromosomes, ring chromosomes, isochromosomes

Balanced: no net loss of gain or genetic material, but packaged differently

Inversions, translocations (reciprocal, Robertsonian, insertions)

Mosaicism: two or more different chromosome complements present in an individual (commonly caused by mitotic nondisjunction in zygote)
Less severely affected If germline mosaicism, can have abnormal offspring

Abnormalities of Chromosome Number: Aneuploidy

Most common and clinically significant type of chromosome disorder 3-4% of all clinically recognized pregnancies Autosomes: Trisomy 21, 18 and 13 seen in live borns with trisomy 21 most common (Down syndrome) Sex chromosomes: both X and Y can be missing or in multiple copies (Turner and Klinefelter syndromes) Most common chromosomal mechanism: meiotic nondisjunction Failure of a pair of chromosomes to disjoin properly during one of the two meiotic divisions (usually meiosis I)

Male Patient with Down Syndrome (47 XY +21)

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Three Color FISH: Human Sperm Yellow: Ch. 18; Red: X; Green: Y
23,X and 23,Y 24,XX 24,XY

Three Color FISH: Amniocytes

Aqua: Ch. 18; Green: X; Red: Y Green: Ch. 18 Red: Ch. 21

Meiotic Nondisjunction and Aneuploidy

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Abnormalities of Chromosome Structure

Result from chromosome breakage and reconstitution in abnormal combination Less common than aneuploidy (1 in 375 newborns) Some rearrangements stable, others unstable Balanced if normal complement of chromosomal material Usually normal phenotype, but pose a risk to future generations Inversions, translocations (reciprocal, Robertsonian, insertions) Unbalanced if there is additional or missing material Can be associated with abnormal phenotype, but may be normal phenotype Deletions, duplications, marker chromosomes, ring chromosomes, isochromosomes

Types of Structural Abnormalities

Paracentric and Pericentric Inversions

Paracentric inversions low risk of liveborn child with abnormal karyotype

Pericentric inversion: high risk of liveborn child with abnormal karyotype (5-10%)

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Offspring of Carrier of Pericentric Inversion with Abnormal Karyotype

Quadrivalent Formation in Meiosis: Balanced 3, 11 Translocation

Cryptic Translocation Red: 3p telomere; Green: 11q

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Chromosome Paint Probes Balanced Translocation

Incidence of Chromosome Abnormalities: Newborn Survey

Chromosome Abnormalities: Spontaneous Abortions

Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Incidence of Chromosome Abnormalities: 10,000 Conceptions

Down Syndrome
Dysmorphic features Hypotonia Short stature Hands: short and broad, single palmar crease, incurved 5th digits Feet: wide space between 1st and 2nd toes, proximal plantar furrow Mental retardation: IQ 30-60, but verbal Congenital heart disease: present in 1/3 Duodenal atresia and tracheoesophageal fistula: more common Leukemia risk (15-fold elevation) Significant risk of Alzheimer disease at younger age Chromosomes: 95% trisomy 21; 4% Robertsonian translocation (between 21 and either 14 or 22); 1% 21q21q translocation, Mosaic Down syndrome, partial trisomy 21

Down Syndrome

Brachycephaly Flat occiput Short neck (loose skin) Flat nasal bridge Low-set ears Brushfield spots (iris margins) Open moth Protruding tongue Tongue furrows Epicanthal folds

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Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Dermatoglyphics- Down Syndrome

Down Syndrome- Maternal Age

Other Liveborn Trisomies

Trisomy 18: 1/7500 births Trisomy 13: 1/20-25,000 births

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Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Microdeletion/Duplication Syndromes

Mechanism of Duplication and Deletion via Repeat Sequences

Y Chromosome in Sex Determination and Differentiation

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Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Sex Determination and Differentiation

XX Male and XY Females Due to Unequal X-Y Exchange

X Inactivation- Barr Body

XX Female XX Female XY Male

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Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

X Inactivation Distribution

Xist RNA is Expressed from and Binds to the Inactive X

X Inactivation with Abnormal X Chromosomes

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Tony Wynshaw-Boris, M.D., Ph.D. SOM 204 Genetics Lecture 7

Disorders of Sex Chromosomes

Klinefelter syndrome (47,XXY; 1:1000 male births) Tall, thin, with long legs, hypogonadism with infertility, mild learning disabilities, psychosocial adjustment Klinefelter variants (48,XXXY; 49,XXXXY) Generally more severe phenotypes 47,XYY (1:1000 male births Normal phenotype and fertility, except for lower IQ Trisomy X (47,XXX; 1:1000 female births) Tall stature, usually fertile, lower IQ Turner syndrome (45,X and variants; 1:4000 female live births, 99% fetal demise, so in 1-2% of all conceptuses): Distinct phenotype High frequency of mutations or microdeletions on X that cause X-linked mental retardation

Klinefelter Syndrome- 47,XXY

Tall Thin Long legs Hypogonadism with infertility Mild learning disabilities Mild problems with psychosocial adjustment

Turner Syndrome- 45,X

Short stature Gonadal dysgenesis (streak gonads) Characteristic facies Webbed neck Low posterior hairline Broad chest with widely spaced nipples Frequent renal and cardiovascular problems Average to above average intelligence Nonverbal IQ<<Verbal IQ Growth hormone therapy adds 6-10 cm to adult height

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