Case 7 - Embryology & Genetics

Case 7 - Embryology & Genetics

1. Embryology

   • Gamete Production
     • Oogenesis
       • In meiosis, one primary oocyte gives rise to 4 daughter cells
         • One of these develops into secondary gamete --> Mature ovum
           • Other 3 develop into polar bodies --> discarded
             • Oogenesis
           • Metaphase I - Anaphase I: Nondisjunction can occur here
           • Pause for secondary oocyte after meiosis I
             • Second meiosis occurs in fertilisation after sperm has entered oocyte membrane to form female pronucleus
         • Gametes derive from PCGs which form in epiblast in week 2 & arrive in gonad in week 5
     • Spermatogenesis
       • Components of Sperm
         • Green = Sperm, Blue = Microproteins from bulbourethral gland
   • Fertilisation
     • 1. Sperm binds to Zona pellucida
       • CAPACITATION: Occurs in uterine tube, glycoprotein coat & seminal plasma proteins removed, acrosomal region exposed
     • 2. Contents of acrosome released by exocytosis
       • ACROSOME REACTION: Occurs after binding to ZP, induced by ZP proteins, across & trips released to fertilise ovum, makes sperm hyperactive
     • 3. Hydrolytic enzymes released help sperm burrow through zona pellucia & enables binding & fusion with egg
     • 4. Sperm nucleus enters egg cytoplasm
       • ZONA PELLUCIDA HARDENING: When outer membranes fuse, chemicals released to harden ZP & defer other sperm
     • Order of penetration: Corona radiata --> Zona pellucia --> Oocyte membrane
     • Fertilisation restores diploid number of chromosomes upon microtubule union of maternal & paternal DNA, this is then replicated
     • Site: Upper third of ampulla
   • Implantation
     • Week 1 = Cleavage: Zygote --> Morula --> Blastocyst
       • Morula: Inner cells = inner cell mass, Outer cells = trophoblast (forms blastocoele cavity)
       • NB: Any blastomeres can form own embryo (IVF treatment)
     • Week 2 = Bilaminar Embryonic Disc: Embryo implants into endometrium & inner cell mass --> epiblast & hypoblast
       • Hypoblast borders blastocoele cavity --> Mesoderm & yolk sac formed
         • Upon implantation, trophoblast --> Cytotrophoblast & syncytiotrophoblast
           • Trophoblasts invade & cells come into contact with arteries & uterine glands
     • Week 3 = Primary germ layers form & folding begins
   • Gastrulation - (3 weeks)
     • 1. Epiblast changes from circular to oval shape, with long head end & midline groove called primitive streak
     • 2. Cells of epiblast migrate towards midline & when they get there, they turn & go inwards & descend to hypoblast beneath it
     • 3. Some cells migrate through all way down to hypoblast, invaginate & occupy it, eventually displace hypoblast cells & become endoderm
     • 4. Other cells migrate down & stop in space between 2 layers, forming the mesoderm
     • 5. Other cells migrate to middle layer & stay in midline to form notochord
     • 6. Remaining cells don't migrate through primitive streak but remain in upper layer & form ectoderm
     • 7. Ectoderm cells occupying upper layer near midline = neural ectoderm cells & cells alongside that = neural crest cells & remainder = skin or surface ectodermal cells
   • Neurulation (4 weeks)
     • 1. Notochord releases Shh protein - inducing formation of floor of neural tube
     • 2. Ectoderm cells migrate to form neural plate over notochord - lateral edges become more elevated & form neural fold with depression called neural groove
     • 3. Flat embryo folds to become 3D- left & right end meet & fuse at midline to make tube with outer layer = ectoderm, middle layer = mesoderm & inner layer = endoderm (tube open at front & back)
       • NB: Neural tube defects caused by arrest of normal development & closure of neural tube
       • 4. Endoderm - lining of gut tube (one end mouth, one anus)
       • 5. Ectoderm - Outer layer - skin
       • 6. Mesoderm - Cartilage, bone, muscle, connective tissue, blood etc.

2. Chromosomal Abnormalities

   • Autosomal Dominant
     • Genotype of affected children = AA, Aa - 50% offspring risk
     • Male to male transmission seen, along with incomplete penetrance & variable expression, males & females equally affected as disorder transmitted by both sexes
       • Incomplete penetrance: Mutation present but compensation leads to complete lack of characteristics of disease
       • Variable expression: Difference in severity of conditions found amongst individuals with same mutation
     • Examples: Huntington's, Long QT, FAP, Marfan syndrome, breast cancer & neurofibromatosis
   • Autosomal Recessive
     • Genotype of affected children = aa - 25% offspring risk
     • Common in consanguinity - Marriage between 2 individuals relates as second cousins or closer
     • Examples: Cystic fibrosis, Sickle cell anaemia, thalassaemia, tay sachs
   • X-Linked
     • Males more commonly affected as no second X to compensate for damage on inherited X chromosome
     • Father has trait --> All daughters will be carriers & sons will be unaffected
     • Mother has trait --> All sons affected
       • Daughter carrier, non-carrier & son affected/non-affected = 25%
     • Examples: DMD, haemophilia A, red-green colourblindness, Fabry's disease
   • Common Abnormalities
     • Down's Syndrome (Trisomy 21)
       • Developmental delay & intellectual disability
       • Distinctive features: Upslanting palprebal fissures, low set ears, single palmar crease, flattened nose
       • Short stature & obesity
       • Increased risk of CHD, thyroid disorders, dementia, hearing loss, seizures
       • 95% Trisomy 21, 3-4% extra 21 from RT, 1-2 Mosaicism
         • Recurrence Risks: T21 1% (10% after 2 pregnancies), RT 15-15% & Mosaicsm 1-2%
     • Edward's Syndrome (Trisomy 18)
       • 75% miscarries or stillborn, 2nd most common trisomy
       • Intrauterine growth retardation, severe developmental delay, seizures, structural abnormalities
       • Distinctive features: Clenched hands & rocker-bottom foot with prominent calcaneus
     • Patau Syndrome (Trisome 13)
       • High rates of miscarriage, 20% survival rate at 1 year
       • Severe developmental delay, seizures, microcephaly, seizures, neural tube defects
     • Klinefelter Syndrome (47, XXY)
       • Primary hypogonadism, pubertal gynacomastia, small testes, delayed development of secondary sexual characteristics, relatively tall stature & infertility
       • Decreased X-Linked disease, Increased risk of diseases associated with women
       • Treatment: Testosterone, surgery, fertility treatment
     • Turner's Syndrome (45, X)
       • Short stature, neck webbing, broad 'shield' chest, widely shaped nipples, lymphedema of hand & feet, low set ears
       • Lack of sexual development at puberty, infertility, amennorhoea, heart defects, normal intelligence
       • Treatment: Hormone therapy (GH --> Oestrogen), surgery for webbing, IVF
   • Unusual patterns of inheritance
     • Imprinting: One gene 'switched on/off" - inactive gene is imprinted
       • 15q11-13 = PW (Deletion paternal) & AS (Deletion maternal)
     • Uni-Parental Disomy (UPD): Two copies of chromosome from one parents (imprinted genes cause issues) e.g. Prader WIlli & Angleman's
     • Mosaicism: After normal fertilisation, cell lines later on have different genetic makeup
     • Mitochondrial Inheritance: Maternal mitochondria passed to offspring e.g. LHON (blindness)
   • Testing of Abnormalities
     • Detection
       • Chromosomes are encouraged to enter mitosis then prevented from separating (cosmid removes spindle) uncoiled, Giesma stained & organised into karyotype
       • Also: Array-CGH very accurate
     • Definitions
       • Malformation: Occur during formation of structures in 3rd-8th week due to environmental/genetic factors
       • Deformation: Occur due to mechanical factors over prolonged periods from intrinsic or extrinsic factors as structures grow
       • Disruption: Occur due to destructive forces, altering pre-existing structures e.g. rubella & limb amputation
   • Translocations
     • Reciprocal Translocation
       • When any 2 chromosomes swap piece of chromosomal material - Balanced as no material lost or gained
       • Genetic Counselling issues: Half of offspring carry translocation --> miscarriage
       • Not all are inherited so carrier testing done to predict recurrence risk
     • Unbalanced Translocation
       • 50% of offspring of carrier - individual family risk needed to assess personal miscarriage risk
     • Robertsonian Translocation
       • 45 Chromosomes present rather than 45 due to acrocentric chromosomes joining together
       • E.g. 14-21 (translocation trisomy 21 & 14) & 13-14 (normal phenotype, trisomy 13 - patau syndrome)
       • Commonly form between 13, 14, 15 & 22
   • 'Broken Chromosomes'
     • Terminal Deletion: 1p36 (straight eyebrows), 4p & 5p (Cri-Du-Chat)
     • Interstitial Deletion: 15q11-a13 (PW syndrome), 22q11.2 (DiGeorge)
       • Di George: Heart, palatal, statue, psychiatric abnormalities
     • Others: Translocation (above), duplications & inversions

3. Genetics Screening & Diagnostics

   • First Trimester Screening (9 - 13 weeks)
     • Blood Test (for free b-hCG & PAPP-A)
       • Free b-hCG = Low in Trisomy 21 & 18, normal or high in Trisomy 13
       • PAPP-A = High in Trisomy 21, low in Trisomy 18 & normal in trisomy 13
     • Ultrasound (for NT)
       • NT between 2.5mm and 3.5mm = Normal (High = DS)
     • Also check viability & gestational age
     • NB: Raised hCG may indicate miscarriage or ectopic pregnancy
   • Second Trimester Screening (14 - 18 weeks)
     • Maternal Serum Screen (Triple) = AFP (alpha ferroprotein), UE3 (oestriol) & free b-hCG
       • AFP = High in T21, Low in T18 & T13
         • Elevated by leakage into amniotic fluid by exposed foetal capillaries in NTD
       • Free b-hCG = Low in T21, T18 & T13
       • UE3 = Low in T21, T18 & T13
       • NB: b-hCG may give false reading in multiple births as high
     • Quadruple marker: Inhibin A raised in DS/T13
       • This is preferred in the obese as combined is harder in those with BMI >25
   • Chorionic Villus Sampling (11-14 weeks)
     • Less acute, more risk but quicker than amniocentesis
     • Two routes: Transcervical & transabsominal
     • Needle, guided by ultrasound, inserted through abdomen to take sample of CV cells from placenta - tested for missing, extra or abnormal chromosomes
     • Miscarriage risk: Less than 2%
     • CVS & amniocentesis given to all over 35yrs
   • Amniocentesis (15 - 18 weeks)
     • Needle, guided by ultrasound, inserted into abdomen to take small sample of amniotic fluid around baby & tested for missing, extra or abnormal chromosomes
     • More accurate, less risky but slower than CVS
     • Miscarriage risk 1% - risk of foetal respiratory damage
   • Rhesus Disease
     • When mother = RdD-ve & baby RhD+ve & mother previously sensitised to RhD +ve blood
     • Antibodies produced by mother & destroy blood cells --> anaemia & jaundice
     • 2 doses of Anti-D given to these mothers

4. Social & Environmental Affects on Pregnancy

   • Alcohol
     • Can lead to damage to foetal cell development, organisation & maturation in CNS (especially in first 3 months before liver development)
     • Foetal Alcohol Syndrome
       • 2-5% USA & Western Europe
     • Also increases risk of miscarriages & stillbirths
   • Smoking
     • Increased risk of SIDS (Sudden Infant Death Syndrome)
     • Low birth weight in at least 20% infants as nicotine & CO reduce availability for oxygen for foetus --> Low foetal tissue oxygenation
   • Age (Increasing maternal age)
     • Fertility - Decrease in ovarian reserve & quality with age, they become less sensitive to FSH stimulation
     • Gestational diabetes - Insulin resistance increases in pregnancy due to changes to pancreatic cells by hormones e.g. oestrogen
     • Pre-eclampsia - Hypertension, Proteinuria & Non-dependant oedema that progresses from mild to severe as symptoms of CNS dysfunction, liver distention, or particularly high bp/proteinuria measurements made
     • Miscarriage & Stillbirth: Probably due to high rates of chromosome abnormalities incompatible with life - 50% rate by age 45
     • Problems with placenta - This can lead to low birth weight due to impaired transplacental flux of nutrients
       • Placenta Previa --> Haemorrhage (alcohol also increases this risk)
       • Placenta percreta (invades uterus --> rupture)
     • Chromosomal abnormalities - At 20 yrs, risk of DS 1/1000, at 40 yrs - 1/80
   • Teratogenesis
     • Three Principles: Dosage, Time & length of exposure, Genotype of embryo
     • E.g. Rubella: 0-12 weeks = 100% risk of foetal infection, 13-16 weeks = 15% Deafness/Retinopathy, 16 weeks + Normal
     • E.g. X-Radiation: Babies develop microcephaly, spina bifida, blindness, cleft palate & limb defects
     • Drugs: All should be avoided if possible for 1st trimester, prescribed only if benefit to mother exceeds risk to child
       • Thalidomide in 1957-1961 for morning sickness --> Limb defects

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