Genetics I Cytogenetics

Introduction to Cytogenetic and Genetic Disorders

• Genetic Disorders Overview: Genetic abnormalities are categorized based on their scale and inheritance patterns. Normal human cells contain a chromosomal set of $46XX$ for females or $46XY$ for males ($2n = 46$).

• Chromosomal (Cytogenetic) Disorders: These involve abnormalities in the total number or structural arrangement of chromosomes. Any deviation from the normal $46$ chromosomes (e.g., $47XX$ or $45X,X$) is classified here. These abnormalities are present in every cell of the body (hair, brain, muscle, etc.) unless the individual is a mosaic.

• Mosaicism: An individual possessing differing chromosomal numbers within the same body. This results from mitotic errors during embryogenesis.

• Mendelian Disorders: Abnormalities affecting a single gene at the nucleic acid base pair level. These lead to protein abnormalities affecting bodily functions but do not alter the overall chromosome number or structure. Inheritance follows classic patterns: autosomal dominant/recessive and sex-linked dominant/recessive.

  • Example: Sickle cell disease involve the substitution of valine for glutamine in hemoglobin, resulting in abnormal protein function without structural chromosomal changes.

• Polygenic (Multigenic) Disorders: Diseases resulting from the inheritance of multiple genes combined with environmental modifications (e.g., Diabetes, Hypertension).

  • Dosage Effect: The more abnormal genes inherited, the more severe the clinical expression and the higher the chance of transmission.

  • Range of Severity: These conditions manifest across a spectrum (e.g., mild blood sugar elevation versus uncontrolled diabetes requiring combination therapy).

• Single Gene Disorders with Non-Classical Inheritance: Isolated gene abnormalities that do not follow Mendelian patterns.

  • Categories: Trinucleotide repeats (Fragile X), Mitochondrial inheritance (Leber Hereditary Optic Neuropathy), Genome Imprinting (Prader-Willi), and Gonadal Mosaicism.

Diagnostic Modalities for Genetic Disorders

• Karyotyping: The standard tool for identifying numerical or structural abnormalities. It involves the spread of chromosomes in pairs, arrested at metaphase, arranged by decreasing length.

  • Sample: White blood cells (WBC) are typically used because they are easily obtained and induced into mitosis using a mitotic spindle inhibitor like Colcemid.

  • Autosomes: The $22$ pairs of non-sex chromosomes. They are grouped by size and centromere location:

    • Group A: Chromosomes $1-3$

    • Group B: Chromosomes $4$ and $5$

    • Group C: Chromosomes $6-12$

    • Group D: Chromosomes $13-15$

    • Group E: Chromosomes $16-18$

    • Group F: Chromosomes $19$ and $20$

    • Group G: Chromosomes $21$ and $22$

  • Banded Karyotype: Uses Giemsa staining to create distinct banding patterns, allowing for the identification of balanced translocations and specific loci.

  • Coordinate System: Arms are divided into "p" (short) and "q" (long). Loci are identified by region, band, and sub-band (e.g., $17q11.2$).

• Fluorescent In-Situ Hybridization (FISH): Uses complementary DNA strands with fluorescent dye to visualize specific segments or numbers of chromosomes.

  • Advantages: Allows for "Interphase genetics" (cells do not need to be in mitosis) and aids in visualizing gene copies or translocations.

  • Interpretations: Three signals for a specific probe indicates trisomy; signals of mismatched length on paired alleles indicates translocation ($t$).

• Barr Body Analysis: Identifies inactive X chromosomes based on the Lyons Hypothesis.

  • Lyons Hypothesis: Only one X chromosome is active in any individual; all additional X chromosomes undergo pyknosis and become Barr bodies. This occurs randomly around the $16\text{th}$ day of embryonic life.

  • Calculation: $\text{Number of Barr bodies} = \text{Total X chromosomes} - 1$.

  • Clinical Findings: Normal males ($XY$) have $0$; normal females ($XX$) have $1$; individuals with $XXXX$ have $3$.

  • Sample: Buccal smear (squamous cells) is preferred over blood smears.

Mechanisms of Mutation

• Genome Mutations (Numerical Changes): Loss or gain of whole chromosomes through nondisjunction or anaphase lag.

  • Nondisjunction: Failure of chromosomes to separate at anaphase.

    • In Meiosis (Gametogenesis): Results in trisomy ($n+1$) or monosomy ($n-1$) upon fertilization.

    • In Mitosis (Embryogenesis): Results in mosaicism.

  • Anaphase Lag: One chromosome fails to reach the pole and is lost during nuclear membrane formation, resulting only in monosomy.

• Chromosome Mutations (Structural Changes):

  • Inversion: Two breaks in a single chromosome with rearrangement.

  • Isochromosomes: Centromere break leading to the loss of one arm and duplication of the other.

  • Deletion: Segment loss between two breaks.

  • Ring Chromosomes: Ends of both arms break and fuse into a circle.

  • Translocation: Transfer of segments between chromosomes.

    • Balanced: No loss of genetic material; phenotypic normality but risk to offspring.

    • Robertsonian (Unbalanced): Involves acrocentric chromosomes; results in one large and one very small chromosome (loss of segments).

Cytogenetic Disorders of the Autosomes

• Trisomy 21 (Down’s Syndrome): Most common chromosomal disorder; major cause of mental retardation.

  • Epidemiology: Incidence of $1:1500$ in mothers $< 20$ years old vs. $1:25$ in mothers $> 45$ years old.

  • Causes: $90\text{\%}$ meiotic nondisjunction, $4\text{\%}$ translocation ($t(14;21)$ or $t(21;22)$), $1\text{\%}$ mosaicism.

  • Clinical Features: Oblique palpebral fissures, epicanthic folds, flat facial profile, dysplastic/low-set ears, protruding tongue, Simean crease (single palpebral crease), clindodactyly (inward-curving $5\text{th}$ finger), hypotonia, and loss of Moro reflex.

  • Associated Conditions: Congenital heart disease ($40\text{\%}$: VSD, ASD, PDA, TOF), GI atresia/stenosis, acute leukemia ($150 \times$ Risk for AML, $33 \times$ Risk for ALL), and premature Alzheimer’s (onset around age $40$ due to amyloid precursor protein gene on chromosome $21$).

• Trisomy 18 (Edward’s Syndrome): Incidence $1:5,000$ to $1:10,000$. Most die before age $1$. Features: Prominent occiput, micrognatha, overlapping fingers ($2\text{nd}$ over $3\text{rd}$), and Rocker bottom feet.

• Trisomy 13 (Patau’s Syndrome): Very severe/rarely live to one year. Features: Microcephaly, Microphthalmia, severe bilateral cleft lip/palate, polydactyly, and Rocker bottom feet.

• DiGeorge Syndrome: Deletion at $22q11.2$ resulting in failure of the $3\text{rd}$ and $4\text{th}$ pharyngeal pouches.

  • CATCH 22/PATCH 22 Mnemonic: Cardiac abnormalities (TOF, VSD), Abnormal facies, Thymic hypoplasia (viral infection risk), Cleft palate, Hypocalcemia (seizures/tetany from parathyroid deficiency).

  • Psychiatric Risk: $25\text{\%}$ developing schizophrenia or bipolar disorder as adults; $30-35\text{\%}$ ADHD risk in children.

Cytogenetic Disorders of Sex Chromosomes

• Turner’s Syndrome (Gonadal Dysgenesis): Typically $45XO$. Phenotypically female with decreased female characteristics.

  • Clinical Features: Short stature (rarely over $5$ feet), primary amenorrhea, streaked ovaries (infertility), webbing of the neck, broad chest with wide-spaced nipples, and coarctation of the aorta. No Barr bodies are seen.

• Klinefelter’s Syndrome (Testicular Dysgenesis): Typically $47XXY$. Phenotypically male ($1$ Y chromosome) with combined female characteristics.

  • Clinical Features: Testicular atrophy, azoospermia/oligospermia ($< 1\text{ million}/\text{dm}^3$), gynecomastia, female distribution of hair, eunuchoid bodily habitus (tall/lanky), and mild mental retardation. Presence of at least one Barr body in males.

• Other Abnormalities:

  • Double Y Males ($47XYY$): Phenotypically normal, excessively tall, severe acne.

  • Multi X Females: Menstrual irregularities, increased mental retardation risk with additional X chromosomes.

Disorders of Sexual Development and Cancer Genes

• Hermaphroditism:

  • True: Both ovary and testis present (often mosaic $XX/XY$).

  • Pseudohermaphrodite: Divergence between phenotypic sex and gonads.

    • Female PH: Ovaries present but ambiguous/male phenotype due to androgen exposure (e.g., adrenal tumors).

    • Male PH (Testicular Feminization): Presence of testis ($XY$) but defective androgen receptors ($Xq12$). The body does not "read" the testosterone, leading to a female phenotype (vagina instead of penis).

• Oncogenes and Tumor Suppressors:

  • Philadelphia Chromosome: $t(9;22)$ in CML. Fuses ABL (chr $9$) and BCR (chr $22$), creating a fusion protein that stimulates unceasing mitosis.

  • Burkitt’s Lymphoma: $t(8;14)(q24)$.

  • Rb Gene: $13q14$ (Retinoblastoma).

  • P53: $17p13.1$.

  • WT-1: $11p13$ (Wilms tumor).

Questions & Discussion

• Q1: Patient with short stature, primary amenorrhea, wide spaced nipples, and negative Barr body.

  • Diagnosis: $45XO$ (Turner Syndrome). A normal female requires at least one Barr body.

• Q2: Newborn with poor suck, oblique palpebral fissures, Simian crease, and absent startle reflex.

  • Diagnosis: Down's Syndrome. Patau's and Edward's have different distinct facial/skeletal features.

• Q3: Karyotype of Down Syndrome ($47XX21+$). Which is incorrect?

  • Incorrect statement: Three Barr bodies. A female with Down's has two X chromosomes, thus only one Barr body.

• Q4: Mechanism for Turner Syndrome?

  • Options: Nondisjunction or Anaphase lag involving sex chromosomes. Both can result in the $45XO$ karyotype.

• Q5: Mode of inheritance where all children of affected mothers are affected, but no children of affected fathers are affected.

  • Mechanism: Mitochondrial inheritance (passed only via the mother).