Amniocentesis and CVS

Methods of chromosomal evaluation Non invasive:

Fetal cells from maternal blood preimplantation embryos (PGD)

Invasive: amniotic fluid (amniocentesis) placenta (chorionic villus tissue) Fetal blood

Invasive techniques Amniocentesis:

Late – second trimester after 15 weeks Early – earlier than 15 weeks

Chorionic villus sampling (CVS) Abdominal Trans cervical Trans vaginal

Fetal blood sampling

karyotypefish

PCR

What can be evaluated? Chromosomal aberrations:

Trisomy, Monosomy, Polyploidy, Marker chromosome, Deletion, duplication, inversion, translocation,

ring chromosome . Genetic aberrations (DNA) Infectious disease Biochemical markers (AFP)

Amniocentesis First introduced by Serr and Fuchs and Riis

in the 1950s for fetal sex determination Only at the late 70th a static ultrasound

was used to locate the placenta and amniotic fluid pocket

Only In 1983, Jeanty reported a technique of amniocentesis ’’under ultrasound vision’’

Mid Trimester Amniocentesis Per coetaneous 20-23g needle Ultrasound guided Usually 20cc amniotic fluid Results – 2 to 3 weeks

complications Pregnancy loss 0.3-1.0%. Increase risk:

Needle larger than 18g Multiple needle insertion Discoloration of the fluid High AFP, multiple late abortions, previous

vaginal bleeding Placental perforation – recent studies didn’t

find correlation

Complications Leakage of amniotic fluid (better prognosis

than spontaneous leakage) Amnionitis Vaginal bleeding Needle puncture of the fetus Long term complications:

Respiratory distress?? Isoimmunization??

Amniocentesis and HIV positive women Increased rate of vertical transmission Chemoprophylaxis previous to

amniocentesis appears to be beneficial in preventing vertical transmission

Multiple Gestation Three methods:

Indigo carmine injection to the first sac A single needle puncture sampling technique

(Jeanty 1990)

Simultaneous visualization of two needles on each side of the separating membrane (Bahado-Singh 1992)

Abortion risk – probably higher Detailed description of fetus

position and placental location

Early Amniocentesis: 9-14 weeks Introduced at late 80th 10-14 weeks gestation Only the amniotic (inner) sac should be

aspirated Approximately 1 cc for gestational age Higher rate of pregnancy loss, talipes

equinovarus, and post procedural amniotic fluid leakage

laboratory failure op to 20%

Chorionic villus sampling Was developed in the 80th

percutaneous transabdominal with 19-20g needle

Chorionic villus sampling Was developed in the 80th

percutaneous transabdominal transvaginal transcervical

15-30mg each aspiration 20mg ideal for cytogenetic testing 30-40mg for cytogenetic and other direct

molecular and biochemical tests

CVS results Direct analysis examines the trophoblast

cells of the placenta (very rapidly dividing cells) Results in few hours greater vulnerability to mitotic error

Cultured analysis examines the fibroblast like cells of the villus stroma or mesenchymal core. Approximately 7-10 days Accurately reflect the chromosomes of the

fetus. 

Risk of invasive procedure Early amniocentesis:

High pregnancy loss High fetal malformations High rate of multiple needle insertions (4.7%) High rate laboratory failures (1.8%)

Late amniocentesis: “Low” pregnancy loss (0.3-1%) Low rate of multiple needle insertions (1.7%) Low rate laboratory failures (0.2%)

Risk of invasive procedure - CVS Transabdominal CVS as safe as second

trimester amniocentesis Trans abdominal and transcervical CVS are

equally safe and efficacious, provided that centers have expertise with both approaches

In approximately 3–5% of cases, clinical circumstances will support one approach over the other

Limb reduction – not after 9 weeks

mosaicism True chromosomal mosaicism is when two

or more abnormal cells lines are detected in two or more culture flasks from the same individual.

Pseudomosaicism is a term used to describe two abnormal cell lines that are found in only one culture flask (not reported to the patient)

mosaicism Most often involving trisomic cell and

normal cells 1-2% of pregnancies undergoing CVS 0.1% of pregnancies undergoing

amniocentesis Clinical outcome of chromosomal

mosaicism is strongly dependent on the specific chromosome involved and the number of trisomic cells in both the placenta and the fetus

Mosaicism (trisomic cells) in CVS

Option of an additional prenatal diagnostic procedure (amniocentesis or fetal blood sampling)

Mosaicism (trisomic cells) in CVS

Four possible conditions: Mosaicism only in the placenta not affecting

the fetus or placental function. Mosaicism only in the placenta not affecting

the fetus but alter placental function (IUGR) Trisomy cells are both in the placenta and in

the fetus Trisomy cells in the placenta and uniparental

disomy in the fetus

Mosaicism (trisomic cells) in amniotic fluid Probably there are trisomic cells in the

fetus The true level and distribution of trisomic

cells cannot be accurately assessed with any prenatal procedure

Ultrasound is often the best judge of how a baby is developing

Uniparental Disomy Arises when an individual inherits two

copies of a chromosome pair from one parent and no copy from the other parent Maternal UPD – two copies from the mother Paternal UPD – two copies from the father

How does UPD happen? Loss of a chromosome from a trisomic

zygote, "trisomic rescue" Duplication of a chromosome from a

monosomic zygote, "monosomic rescue" Fertilization of a gamete with two copies

of a chromosome by a gamete with no copies of the same chromosome, called gamete complementation.

Trisomic rescue following an error in meiosis

heterodisomy

Trisomic rescue followed an error in meiosis II

isodisomy

UPD - health concerns in people for two possible reasons: Parental imprinting in the case of

heterodisomy and isodisomy Unmasking of recessive conditions in some

cases of isodisomy

Clinical consequences of UPD molecular UPD testing should be

considered for certain chromosomes (including 6, 7, 11, 14, 15) that are known to have adverse phenotypic imprinting effects. 

Factors considered when trying to predict the outcome of mosaicism the chromosome involved

A mosaic finding 18 or 21 is likely to have worse implications

mosaic finding for trisomy 15 or 16 is likely to have less implications (trisomy 15 or 16 cells cannot survive )

Factors considered when trying to predict the outcome of mosaicism The tissues affected and level of

trisomy in those tissues The tissue affected cannot be evaluated The level of trisomy can be only estimated   

Factors considered when trying to predict the outcome of mosaicism method of ascertainment  

CVS shows that the placenta is affected Amniotic fluid suggests that at least one fetal

tissue may be affected Fetal blood sampling confirms the diagnosis of

chromosomal mosaicism

Factors considered when trying to predict the outcome of mosaicism ultrasound findings   presence/absence of uniparental

disomy   number of previous case reports

known in the literature  

Thank you