Prenatal genetics Flashcards
1
Q
What stages of oogenesis occur in the in the fetal ovary ?
A
- in the female embryo the primordial female germ cells migrate to the primitive ovary and rapidly proliferate to form the oogonia
- after month 7 most oogonia die- those that remain enter the 1st meiotic division to produce a primary oocyte
- the primary oocyte is arrested in prophase 1 of meiosis until puberty
2
Q
What stages of oogenesis occur in the in the adult ovary ?
A
at puberty groups of oocytes periodically resume meiosis
- oocytes are in metaphase II at ovulation and do not complete meiosis II until fertilisation
3
Q
What is the difference between cell division in oogenesis and spermatogenesis?
A
In oogenesis there is unequal cytokinesis in meiosis-most of the cytoplasm is retained by the oocyte and the polar body contains little more that the haploid nucleus
In spermatogenesis there is equal division
In meiosis I the primary spermatocyte divides to produce 2 secondary spermatocytes
in meiosis II the secondary seprmatocytes divide to produce 2 haploid spermatid (4 total)
4
Q
what is though to be responsible for the different mutation rates between the sexes?
A
there are 2 more mitotic divisions in the production of sperm than of an ovum
5
Q
what is the structure of the oocyte?
A
oocyte cytoplasm contains: mitochondria, ribosomes, DNA/RNA polymerases and protective chemicals
- plasma membrane is surrounded but a thick extracellular matrix called the zona pellucida
- the oocyte is surrounded but a layer of follicle cells called umulus cells which nurture the occyte just before and after fertilisation.
6
Q
what are the stages of spermatogenesis?
A
in male embryo development primordial germ cells migrate to the male testes and undergo proliferation to generate spermatogonia
some spermtogonia stop dividng and differentiate into spermatocytes- in meisosi 1 there is symmetrical division to produce 2 spermatocytes and in meiosis II 4 equally sized haploid spermatids are produced
7
Q
Where to sperm mature
A
haploid spermatids undergo morphological differentiation into sperm and escape in the lumen of the seminferous tubutles and then into the epididymus
8
Q
when does spermatogenesis occur?
A
starts in puberty and continues until death but the quality of sperm deteriorates
- get new sperm each month
9
Q
what is the structure of sperm?
A
sperm asr small with reduced cytoplasm
- head piece contains acrosomal vesical with digestive enzymes required to fertilise the oocyte
- mid piece with mitochondria and
- flagellum in tail provides propulsion
10
Q
Define fertilisation, where does it occur?
A
Process by which a sperm and egg fuse to create ans new individual and occur in the ampula of the fallopian tube
11
Q
What are the stages of fertilisation?
A
- sperm attaches to outer cumulus cells of the oocyte
- sperm reaches the zona pellucida (ZP)
- acrosome bursts releasing enzymes which digest the ZP so the sperm can come into contact with the oocyte membrane- acrosomal reaction
- cortical reaction occurs to prevent fertilisation by more than 1 sperm
- oocyte undergoes 2nd meiotic division to produce a secondary oocyte and a polar body
- sperm tail and mitochondria degrade- explaining maternal inheritance of mitochondria
- haploid sperm and oocyte nucelai form the pornuclei
- first mitotic division occurs
fertilised oocyte = totipotent zygote
12
Q
what occurs in the cortial reaction?
A
prevents fertilisation of an oocyte with more than 1 sperm
cortical granules within the oocyte fuse with the plasma membrane of the cell- enxymes in these granules are then expelled by exocytosis to the ZP. this causes crosslinking of gycoproteins in the ZP making the matrix impermeable to sperm
13
Q
Define embryogenesis?
A
process of cell division and differentiation of the human embryo
Embryo = fetus at 8 weeks gestation
14
Q
What happens to the oocyte post fertilisation?
A
fetilised egg is propelled along the oviduct and is prevented from adhering to the oviduct wall by the ZP (if this does not happen you can get an ectopic pregnancy)
in the uterus degrades and there is implantation of the blastocyst
15
Q
what are the stages of embryogenesis?
A
cleavage compaction blastocyst implantation gastrulation
16
Q
what happens in the cleavage stage of embryogenesis?
A
zygote divides rapidly to form a number of smaller cells- the blastomere
- controlled but zygotic genome
cells do not always divide at the same time so result in embryos with odd numbers of cells
17
Q
what happens in compaction?
A
at the 8 cell stage the cells latten against each other to from the tightly packed morula
compaction introduces some level of polarity
18
Q
what is the 16 cell embryo?
A
blastocyst with external polarised cells, internal non-polar cells
19
Q
what is the blastocyst
A
16 cells embryo with fluid filled cavity called the blastocoele
- outer layer of trophoblast cells with give rise to the fetal portion of the placenta (the chorion)
- innner cell mass (ICM) whihc congregate at one end- the embryonic pole
20
Q
What cells does the ICM give rise to?
A
all the cells of the organism plus the yolk sac, amnion and allantosis
21
Q
when can monozygotic twins arise?
A
at any time until the late blastocyst stage
22
Q
what happens in implantation?
A
at day 5 an enzyme is released whihc makes a hole in the ZP releasing the blastocyst- the blastocyst then attaches the the uterine membrane
- trophoblast cells rapidly divide to form the cytotrophoblast ans syncytiotrophoblast and invade the connective tissue of the uterus
- a fluid filled cavity forms within the ICM generating the amniotic cavity enclosed by the amnion
23
Q
what happens in gastrulation?
A
Gastrulation is the process in which the orientation of the body is established and the 3 germ layers are formed:
ectoderm
endoderm
mesoderm
The characteristic gastrulation is the formation of the primitive streak
24
Q
what is the primitive streak?
A
forms during gastrulation
starts as a primitive streak and then develops into a groove then a pit.
cells migrate through the primitive streak to the space between the epiblast and hypoblast to generate the 3 germ layers- 1st endoderm, 2nd mesoderm and 3rd ectoderm
25
what structures develop from the endoderm?
lungs, liver, thymus, endocrine glands
26
what structures develop from the mesoderm?
blood vessels, muscle tissue, connective tissue
27
what structures develop from the ectoderm?
skin and the CNS
28
What happens to the epigenetic imprint in the fetal germline?
All DNA methylation patterns are erased ans the pattern specific to the sex of the fetus is set- when fertilised the imprint is maintained stably in all somatic cells of the fetus and throughout life
29
what is IUI?
interuterine insemination- injection of sperm directly into the womb following stimulated ovulation of the woman
30
what is IVF?
adding sprm samples to eggs (havested from a women following stimulated ovulation) in a petri dish. They are incubated together and monitored for successful fertilisation. successfully developing embryo is transferred back to womans womb
31
what is ICSI?
microscopic injections of a sperm directly into an egg nuclei in aperi dish. Simillar to IVF but maximises the chance of fertilisation
32
what is MESA?
microsurgical epididymal sperm aspiration- surgical sperm extraction method and can be used with IUI, IVF or ICSI for men with azoospermia (no sperm in semen)
33
what is triploidy?
additional full set of chromosome
| 69,XX/XY
34
what is the incidence of triploidy?
1-3% of pregnancies
99.9% abort in 1st or second trimester- those that make it to term die in the neonatal period <1 month
advanced maternal age is not a risk factor
35
what is the proportion of diandry and digyny triploidy?
60-80% are diandry- most common
36
what is diandry triploidy?
2 maternal and 1 paternal set of chromosomes
37
what is the cause of diandry triploidy?
fertilisation of a normal egg by 2 sperm- failure of cortical reaction- 69,XXX or 69,XXY or 69,XYY- 69,YYY not viable so not seen
minority are due to fertilastion of a normal egg by diploid sperm by a failure of NDJ in spermatogenesis for a complete set of chromosomes
38
what is the characteristic feature of a diandric triploid?
partial hyadatiform mole which is a trophoblastic hyperplasia of the cystic villi
increased hCG levels
39
what are the clinical features of triploidy?
```
IUGR
head to chest fusion
limb growth abnormalities
macrocephaly- digyny
neural tube defect
hydrocephalus- digyny
syndactyly
heart/renal defects
cleft lip/palate
```
40
what is digyny triploidy?
2 maternal and 1 paternal set of chromosomes
non hydropic villi
NO MOLE and placenta is generally small
41
what causes digyny triploidy?
fertilisation of a diploid egg due to NDJ of a complete set of chromosome during meiosis
retention of a polar body in the fertilised egg- failure of meiosis II
fusion of 2 eggs and fertilisation by a haploid sperm
42
what causes the difference in presentation between digyny and diandry triploids?
due to the influence of differing imprinted states
43
what is the recurrence risk of triploidy?
usually sporadic
- digyny triploidy has been reported as recurrent in several families
- diandric recurrence risk is 1-1.5%
44
what is gestational trophoblastic disease?
family of diseases including
- non invasive: mole and partial mole
- malignant- invasive love, choriocarcinoma and placental site trophoblastic tumours
45
what is a complete mole?
46,XX/XY but only paternal chromosomes present
46
what are the characteristics of a complete mole?
high levels of hCG
bunch of grapes appearance
NO formation of a fetus
47
how do complete moles arise?
fertilisation of a enucleated egg with a haploid sperm and duplication of the haploid genome (46,XX)
remaining are fertilisation of an enucelated egg with 2 different sperm 46,XY or 46,XX
46,YY are not seen as unviable
48
How are molar pregnancies diagnosed?
high hCG
uterus may be enlarged
more vomiting
vaginal bleeding
49
what is the recurrence risk of a complete mole?
in in 100 after 1st
| 1 in 5 after 2nd
50
what is familial recurrent hyadatiform mole?
maternal affect
AR
results in recurrent molar pregnancies
the moles that are present are usually diploid and genetically bi parental which is different to sporadic miles which are diandric
51
what is a malignantgestational trophoblastic disease (GTD)?
- malignant transformation of a mole
- also known as a persistent mole
- usually arise form a complete mole
- patients present with elevated hCG after a previous molar pregnancy
also presents with abormal bleeding abdominal pain or swelling
52
What other malignancies may arise following molar pregnancy?
malignant choriocarcinoma- highly invasive and can cause symptoms locally e.g. uterine bleeding and from distant metastasis in the lungs, CNs and liver (most common)
diagnosed by raised hCG and reproductive history
53
what is the treatment for GTD?
suction evacuation recommended to remove complete and partial molar pregnancies.
- after molar pregnancy hCG levels should be monitored until they return to normal. usually ~6 weeks and not adviced to get pregnant during this period
if hCG does not return to normal, choriocarcinoma or metastasis is present chemotherapy with methotrexate is recommend
54
what is a dizygotic twin?
fraternal
2 different eggs fertilised by 2 different sperm
more common than identical twins
55
what are the risk factors for dizygotic twins?
increased maternal age
mutations in genes in the TGF9 pathway a.e.g BMP
can be familial
56
what is a monozygotic twin?
formed from complete cleavage of single embryo?
57
What are the different types of MZ twins and when are they formed?
DCDA -division at ~ day 4, morula stage
MCDA - division at day 4-7 st the blastocyst stage
MCMA- embryos that divid after the amnion is formed at day 9
58
are MZ twins identical?
have identical genes but rarely identical due to epigenetic and prenatal environmental post-zygoticmechanisms
59
what pregnancy complications are associated with twinning and what is the highest risk twin type?
6 fold increase in perinatal mortality
- preterm delivery, IUGR
MZ twins have poorer outcomes than DZ
MCMA have highest perinatal mortality rate
60
what is an anastomoses?
vascular connections between the feto-placental circulation in each twin
- risk on MCDA
61
what is TRAP?
Twin reversed arterial perfusion (TRAP sequence) is a rare condition of monochorionic twin pregnancies. It arises when the cardiac system of one twin does the work of supplying blood for both twins. The twin supplying the blood is known as the "pump twin" and develops normally in the womb. However, the increased pumping of the heart puts this twin at risk for cardiac failure. The other twin — known as the "acardiac twin" — lacks a heart or has one that is not fully formed. It usually has a poorly developed body and may also be missing a head, limbs and torso
62
what causes TRAP?
MC twins
| artery to artery and vein-verin anastomoses
63
What are the features of TRAP?
one twin is acardiac and perfused in a reverse direction by the pump twin
- acardiac twin has abn development of head, thorax and or limb
- 'normal' pump twin has poor prognosis and 50% chance of survival due to cardiac overload
some are chromosomally abn
64
What is TTTS?
twin twin transfusion syndrome
65
what causes TTTS?
Affects monochorionic twins
there is an imbalance in the blood exchange between the twins and the placenta. One twin — the donor twin — gives away more blood than it receives in return and runs the risk of malnourishment and organ failure. The recipient twin receives too much blood and is susceptible to overwork of the heart and other cardiac complications.
66
What are the features of TTTS?
dihydramnios and IUGR in donor twin
polyhydramnios and enlarged bladder and larger size in recipient twin
can result in preterm labour due to severe polyhydramnios
67
what is the testing and treatment for TTTS?
- amnio reduction by serial amniocentesis from recipient twin to reduce pressure
- laser ablation of connecting anastomoses (vessels)
amnio may be sent for testing but array is not indicated
68
What is a vanishing twin?
when 1 twin suffers early fetal death (1st trimester) and is not aborted
69
How may a vanishing twin be identified?
- areas of degenerate villi or amorphous debris- but often difficult to detect
- may be identified by the presence of an empty sac on USS
- can explain otherwise inexplicable anomalous maternal serum and alpha fetoprotein and CVS chromosome result
70
what is a fetus papyraceaus?
when a twin suffers death and isn't aborted but unlike in a vanishing twin the dead fetus persist (fetal death occurs beyond the first trimester)
71
What is zygosity testing?
determine the degree of identity in the genome of twins
72
when is zygosity testing performed?
generally requested when one twin has developed a clinical phenotype that is though to be genetic in origin
also used to in transplantation where a twin is a HLA match- if they are identical there is less likely to be GVHD in the recipient
73
How is zygosity calculated?
Use QF-PCr to detect microsatellie markers
the liklihood of zygosity is then calculated using bayes.
the prio risk of monozygosity is 1/3 and dizygosity is 2/3
- if sex is know this can be added (1/2)
- the probability of being dizygotic but with the same alleles by chance is cal;calculated from the possible combination of parental alleles.
74
What is the method for QF-PCR
- amplifies microsatellite markers on target chromosomes for semi-quantitative detemination of ploidy
- multiplexed and uses conditional to allow multiple PCR reactions to be run together
- products are differentiated by using different colour dyes and product sizes using capilliary electrophoresis
75
how is QF-PCR quantitative?
reaction is stopped in the exponential phase- PCR reagents are not depleted
In the exponential phase the level of product is proportional to the amount of starting material and can be used to determine the relative CN of the chromosomes tested
BPG recommend 24-26 cycles
76
how many markers are required for reporting QF-PCR?
| what are the requirements for chosing markers?
Need 2 informative markers- remaining can be inconclusive but not contradictory to the result being reported.
recommended to have at least 4 markers per chromosome to increase the chance of an informative result
when chosing markers need?
- high heterozygosity in the generalpopulation
- SNP checked
- majority are 4bp repeats as these have fewer stutter peaks than dinucleotide repeats
77
What sex chromosome markers are used for QF-PCR?
sexing may or may not be included depending on local policy- risk of sex selection of fetus
AMEL- present on X and Y and gives peaks 4bp apart. cannot detect sex chr no. but can detect their ration e.g. 1:1 XY or 2:1 XXY
SRY- Y chromosome specific non-polymorphic marker to confirm presence or Y
TAF9- present on chr3p and Xq used to determine X chromosome copy number based on the assumption that 3p will have a CN of 2
- can be used to distinguish XO from XX with all homozygous markers
78
how is peak ratio determined for QF-PCR?
peak height/area of smaller peak divided by the peak height/area of the larger peak
79
what is normal PCR ratio?
1:1 alleles with a range from 0.8-1.4
for alleles >24bp apart a ratio of 1.5 is acceptable because of preferential amplification of the smaller allele
80
what is a trisomy PCR result?
1:1:1 - triallelic and confirms meiosis 1 NDJ
2: 1 ratio 1.8-2.4
1: 2 ratio 0.46-0.65
81
what is the significance of a biallelic PCR result
a biallelic pattern could indicate meiosis II NDJ or mitotic NDJ and CPM should be considered for CVS - report cautiously
- meiotic NDJ not confirmed
- follow-up karyotype on cultured cells which contain the mesenchyme and are more representative of the fetus
- suggest detailed USS and AF sampling
82
what are the inconclusive ratio in PCR and their implication?
0.6-0.8 and 1.4-1.8
can be resolved by ruinning extra single chromosome markers
- cannot report as sample as trisomic if there is a single inconclusive marker or single normal marker for that chromosome
may be due to preferential amplification of the smaller allele and can result in inconclusive ratios the greater the distance between the 2 markers.
83
doe results need to be confirmed?
2018 BPG
trisomic results do not need confirmation by another method but sample ID must be confirmed before reporting- usually by repeating the original sample
normal result do not need to be repeated
84
can single allele result be reported
occasionally if normal nut this should be stressed on the report- ideally another method e.g. karyotype of FISH should then be used as follow-up to confirm the result for the chromosome
85
how may MCC present on PCR results?
- skewed/inconclusive results for all chromsomes
- normal female results
- need to request maternal blood
if there is a mixed genotype there will be 3 alleles - 1 fetal (pat) specific, 1 mat and fetal shared and 1 mat specific.
- in a normal sample: the sum of the peak of the maternal and fetal specif alleles should add up to the fetal+mat shared allele ans give a normal ratio
AMEL can be used to quantify the fetal genotype in a male- if a normal result with non skewed XY is obtained a normal result can be issued
86
what types of MCC result can and cant be reported?
BPG
- low level MCC - majority fetal genotype with no inconclusive markers- report
- single fetal genotype with no inconclusive alleles- report
inconclusive alleles with mixed genotype- do not report
QF-PCR is sensitive to MCC to ~10%
87
how can the risk of MCC be reduced for CVS and AF?
AF-MCC may be obvious due to bloodstaining wither before or after centrifuging
- if normal female result obtained need to request mat sample before reporting
- could have a tissue plug resulting in MCC with no obvious bloodstaining
- culturing will remove MCC from blood as the culture will favour growth of the amniocytes, but contamination from mat tissue may grow
CVS- carfeul dissection to remove maternal decidua required to prevent MCC risk
- analysis of cultured cells may help but need to be cautious as maternal tissue may still be present and could predominate
88
How is mosaicism detected on PCR?
presence of extra peaks, skewerd markers for just one chromsome
markers may be subtle with alleles still in the normal or inconclusive range
diploid/triploid mosaicism can be difficult to distinguish from MCC
89
what are the causes of mosaicism?
mitotic rescue of a trisomic cell line
mitotic NDJ and generation of a trisomic cell line in a normal conceptus
Need to be wary of CPM in CVS samples
90
what may cause a single abnormal marker?
```
SMM
PSP
SMD
partial chromosome imbalance
CNV
```
91
can markers 2bp apart be used for reporting?
No - stutter peaks may be incorporated into the peak for the shorter allele resulting in misinterpretation of allele ratios
92
what is an SMM?
somatic microsatellite mutation
- single marker with 1:1:1 ratio and unequl peak heights (2 will add up to the larger one)
- due to mutation of an allele and mosaicism for the de novo and orginal allele
- caused by defective proofreading during DNA rep in early embryogenesis
cant use as an informative marker for reporting, lower annealing temp to distinguish from a PSP
93
what is a PSP?
polymorphsim to primer binding site resulting in efficient amplifcation of an allele ans a single inconclusive result 0.6-0.8 or 1.4-1.8
repeating with a lowered anealing temp should normalise the ratio
94
what is an SMD?
submicroscopic micrsatellite dup
- trisomic result fro a single marker- all other consistent with normal
- some are inherited and benign- list on known inherited SMDs available
- if flanked by 2 normal markers assumed to represent an SMD and not reported
- parents may be requested to check if SMD is inherited and not associate with abn phenotype
distinguish from PSP be repeating at lower annealing temp
detail findings on report if not considered benign and extra testing with flanking markers may be required
95
How may a partial chromosome imbalance appear on PCR?
nromal and abnormal results for the same chromosome- 2 or more consecutive markers showing the same result
follow up with karyotype, array, parental bloods
96
what are the considerations for PCR in twin pregnancies?
twins can be identified as dizygotic or probably monozygotic which may be clinically useful
- liklihood of monozygosity is determined by the number of informative markers and the maternal/paternal relatedness
sampling the same twin twice cant be excluded
which twin the report is for should be clearly identified e.g. twin 1 upper left
97
what are the advantages of PCR over FISH?
cheaper
needs less sample
less labour intensive
can detect MCC in all pregnancies not just male
can detect UPD
may detect some unbalanced rearrangements
98
what should be included on PCR reports?
- assumption that fetal material was tested
- result is consistent with trisomy as only specifc markers tested
- include interpretative sentence e.g. consistent with a diagnosis of dowm syndrome
2 alleles results- risk that it may represent CPM- follow up karyotype, AF and USS
- should perform karyotype follow-up of abnormal results to determine the structure and recurrence risk
99
what are the limitations of FISh for aneuploidy?`
- does not give structure
- higher resolution that G-banding
- cant detect MCC in female pregnancies but may be suspected if there is mosaicism
- more expensive ans time consuming that PCR
- higher failure rate that PCR
- need to scan lots of cells are there can be a range of artefactural staining e.g. monomsomy from co-localisation of 2 signals or trisomy from 2 cells being on top of each other.
need to scan even more if referral indicates mosaicism
100
what is the problem with using centromere probes in FISH?
repetitive sequence probes are targeted to the alpha satellite sequences found at the centromere - composed on array on tandomly repeated monomers
the are sufficiently different to allow distinction of all chromosome except
chr13 and 21 and chr14 and chr22
so need to look for 5 signals for a trisomy and follow-up testing required to confirm the chromosome involved.
target specific probes targeted to specific sequences on the q arms of chromosomes are therefore preferred
101
what percentage of pregnancies have major congenital abnormalities and what percentage of these are chromosomally abnormal?
1-1.5% of pregnancies have major congenital abnormalities
| 20-25% of these are chromsomal
102
what are the benefits of prenatal microarray?
higher resolution than karyotype increases diagnostic yeild
- BPG recommend minimum resolution of 400Kb
- 2.4% increase in diagnostic rate for all and 7% for abscan referrals
- carriers of familial rearrangements not detected
- culturing generally not required (except small samples or MCC cases
103
what are the limitations of prenatal array?
increased resolution can increase the pick up of VUS which can be difficult to interpret- may need MDT. should only report variants that are class 4 or 5 in prenatals, may also need to request parents to aid interpretation
- may miss low level mosaisicism
- may not detect triploidy
- may detect IFs- need reporting policy (report IFs that are clinically actionable to the family)
- culturing may be required for follow-up as array does not give structure
104
what was the EACH study?
Evaluation of ArrayCGH in prenatal diagnosis of fetal anomalies
- multi centre study comparing aCGH to karyotype in fetus with 1+ USS abn of NT>3.5mm
- 500 recrutied
- QF-PCR followed by aCGH for those with a -ve result
- ISCA arrays
- variants categorised as benign, VUS or pathogenic (arrays are now using a 5 clas system as with SNV interpretation)
- only reported variants associated with a known clinical syndrome of significant chromosomal imbalance
105
what were the aims of the EACH study?
- to optimise prenatal DNA extraction techniques
- incoporate aCGH into standard lab service
- all CNVs recorded in a standard way and ina central database
106
what were the findings of the EACH study?
CMA is a robust, cost effective, acceptable diagnostic technique for prenatals with a NT >3.5mm and US abn and should replace karyotype
1-3% increased diagnosis compared to karyotype
107
what are the recommendations for use of microarrays in pregnancy?
2015 working group on behalf of the joint committee on genomics in medicine
- gudiance on reporting in prenatal samples e.g. whether come low penetrances susceptibility loci should be reported
```
recommends that:
NIPA1 dels and dups are not reported
15q13 dups not reported
STS dups not reported
16q13 dups not reported
het del of gene that cant be linked to phenotype i.e. carrier status should not be reported
```
108
what percentage of pregnancies are chromosomally abnormal?
1%
109
what is a very early conceptus and what percentage end in miscarriage?
at the morula to blastocyst stage many abnormal embryos arrest
~ 30% of pregnancies fail to implant and anothe 30% implant transiently - this results in little disruption to the menstrual cycles
cause is commonly polyploidy or monosomy and frequency increases with advancing maternal age
110
what gestation distinguishes miscarriage from still birth?
fetus is incapable of surviving independently < 24 weeks, therefore below this is classed as miscarriage and above this is a still birth
111
what are the common cause of 1st trimester pregnancy loss?
50% are chromosomally abnormal
of these..
I.e majority are trisomy, triploidy and monosomy X
```
60% single trisomy
12-15% monosomy X
12% triploidy
3% tetraploidy
~5% structural rearrangement
~5% multiple aneuploidy
```
112
what percentage of 1st trimester pregnancies end in miscarriage?
~10-15% usually towards the end where the placenta takes over the nourishment of the fetus so the body recognises abnormality
113
what are the most common trisomies in pregnancy loss?
```
T16
T22
T21
T15
T13/18
```
only T21,18 and 13 are seen in liveborns non-mosaically
the frequency of trisomy in pregnancy is greater than in live birth as many are lethal and will result in miscarriage
114
what is the prevalence of T16 in miscarriage
T16 is seen in 1% of pregnancies and accounts for 10% of pregnancy loss
those that survive are mosaic or CPM and only in placenta- this can result in placental insufficiency and IUGR
115
When are most trisomies derived from?
Majority are from an error (NDJ) in maternal meiosis I
- T18 associated with NDJ in meiosis II
significantly associated with maternal age
116
what is the miscarriage rate in the second trimester?
2-5% with 20% chromosome abn rate
117
what are the common chr abn in 2 trimester miscarriage and what percentage are chromosomally abnormal?
20% chromosomally abnormal
- more likely to be viable trisomies (21, 18, 13) as others lost in 1st trimester)
- increased proportion of structural rearrangments
118
what percentage of 3rd trimester pregnancies result in miscarriage?
0.5%
119
what are the genetic findings in 3rd trimester miscarriage?
reduced proportion of aneuploidy (T21, T18, T13 and 45,X)
increased proportion if sex chr abn and structural rearrangements and there is less genetic imbalance than in a trisomy
120
what percentage of the following chr abn are lost during pregnancy?
```
T16
T21
T18 + T13
45,X
tertraploid
triploid
balanced rearrangement
unbalanced rearrangement
normal karyotype
```
```
T16- 100% unless mosaic
T21 - 80%
T18 + T13 - 95%
45,X- 95-99%
tertraploid- almost 100%
triploid- almost 100%
balanced rearrangement- 15%
unbalanced rearrangement- 95% (depending on size of imbalance)
normal karyotype- 8%
```
121
what is the role of tetraploidy on miscarriage?
4 copies- usually due to failed early mitotic division resulting in duplicated genome and 92,XXX or 92,XXYY
2-3% of spontaneous miscarriage
122
what is the role of triploidy on miscarriage?
10% of miscarriages
2/3 are diandric due to dispermy
- partial hyadatiform mole and IUGR
- most miscarry in 1st trimester a few make it to second trimester
1/3 digyny due to diploid egg from NDJ of complete chromosome set in oogenesis or failure to expel polar body
- misacarry in 1st trimester
- severe IUGR, macrocephaly and small placenta (non molar)
123
what is the role of 45,X on miscarriage?
95-99% die in utero
| hypothesised that complete 45,X is lethal and that all TS patients have some level of mosaicism
124
what is the role of monosomy on miscarriage?
Only seen for chr 21 ~1 in 1000 karyotyped abortions
125
what is the role of uniparental diploidy on miscarriage?
diandric diploidy- complete mole due to 2 sperm fertilising and empty egg
- increased risk of choriocarcinoma
digyny diploidy- ovarian teratoma. fatal. may contain fully differentiated tissue e.g. hair and nails
126
what is the role of structural rearrangements on miscarriage?
likelihood of miscarriage of abnormal offspring is dependent on the size and genetic content of the imbalanced region if unbalanced
balanced rearrangements seen in the same frequency in liveborns
127
what is the role of chromosome mosaicism and CPM on miscarriage?
CPM with the abnormality only in the placenta can still result in pregnancy loss due to abnormal development of the placenta which can have a physiological effect on the fetus
128
What referrals are eligible for testing following a miscarriage?
3rd consecutive recurrent miscarriage (<24weeks)-n RCOG = test the POc and follow- up in parents if an abnormality is detected. Only test parents by karyotype in exceptional circumstances
pregnancy loss with significant fetal abnormalities or termination (irrespective of gestation)
still birth (pregnancy loss >24 weeks)
129
how many women experience recurrent miscarriage?
1%
130
What causes are there for miscarriage other than genetics?
epidemiological factors
parental/ embryonic factors
infections
acquired thrombophilic defects
anatomical abnormalities (uterus, cervix)
maternal endocrine and immune anomalies (thyroid dysfunction)
UNEXPLAINED
131
what is the genetic testing methodology for POCs
QF-PCR for common aneuploidies and sex
- some centres also include other trisomies common in miscarriage 16, 22, 14, 15
ArrayCGH
Karyotype no longer performed as tissue can be hard to culture and prone to infection or failure plus array provides a higher resolution and expecting to find imbalance.
Donaghue et al 2017- PCR and array = lower failure rate and higher yeild that karyotype or MLPA strategies
132
when is the 1st trimester USS offered and what are the aims
11-14 weeks
- pregnancy dating
- ID multiple pregnancies
- placental location
- ID fetal anomalies (50%) can be detected at 1st trimester scan
133
what is the nuchal translucency?
subcutaneous fluid under the skin at the nape of the neck
- most powerful measure for detecting aneuploidy
- included in the combined scrore (with mat age, PAPP-A and B-hCG)
- requires a highly trained sonographer to be measured and variation in measurement has been reported
134
what is the significance of the NT measurement?
increasing size is correlated with increased risk of chromosomal abn
>3.5mm is significant and may be due to chromosomal abnormality (T21, 18, 13, XO), cardiac defect/failure or delayed development of the lymphatic system
can either regress or evolve into cystic hygroma, oedema, fetal hydrops in second trimester
135
what are the USS soft markers 'normal variants'
USS markers that may be associated with anuploidy but are also seen in normal pregnancies
- absent nasal bone (seen in 2% of normal fetus and 9% in normal afro-Caribbean pregnancies)
- dilated cisterna magna
- echogenic foci in the heart
- 2 vessel cord
- echogenic bowel with density lower than bone
136
what is the significance of normal variants?
DS screening programme has increased the detection of aneuploidy at first trimester therefore if normal variants are detected at the second trimester and the mother had a normal combined screen score they are not referred for genetic testing
137
what second USS abn are offered genetic testing?
ventriculomegaly >10mm
echogenic bowel with density greater than bone
nuchal fold >6mm
renal pelvic dilation
138
what major congenital abnormalties can be detected in 1st trimester and what conditions do they indicate?
holoprocencephaly- failure of brain to divide into 2 lobes (T13, 18, SHH gene)
Exomphalos- herniation of fetal abdomen 9t18, 13 or BWS)
megacystis - enlarged fetal bladder (T13 and 18)
skeletal abn e.f. T21, 18 and X) associated with shortened long bones
139
What are the USS abn associated with iso(12p)?
diaphragmatic hernia
short limbs
abnormal hands and feet
140
What are the USS abn associated with wolf hirschorn?
IUGR
microcephaly
cleft palate
heart defect
141
What are the USS abn associated with cri-du-chat?
heart defect
| cleft lip/palate
142
What are the USS abn associated with Miller Dieker?
lissencephaly (LISS1)
143
What are the USS abn associated with 22q11.21?
cardiac abn (TOF)
renal abn
polyhydramnios
cleft lip and palate
144
What are the USS abn associated with Williams?
IUGR, cardiac defects
145
What are the USS abn associated with BWS?
overgrowth, exomphalos
146
What are the USS abn associated with AS?
asymmetrical IUGR
147
What are the USS abn associated with achondroplasia?
short long bones
| frontal bosing
148
What are the USS abn associated with T21?
```
cardiac defects-VSD
duodenal atresia
IUGR
hydrops
echogenic bowel
renal anomalies
syndactyly
```
149
What are the USS abn associated with T18?
```
rocker bottom feet
talipes
clenched hands
exomphalos
VSD
oesopageal atresia
IUGR
renal abn
neural tube defect
```
150
What are the USS abn associated with T13?
```
holoprosencephaly
diaphragmatic hernia
cleft lip/palate
IUGR
renal abn
```
151
What are the USS abn associated with 45,X?
```
cystic hygroma
hydrops
increased NT and nuchal fold
fetal oedema
IUGR
renal abn
2 vessel cord
```
152
What are the USS abn associated with triploidy?
IUGR
holoprosencephaly
2 vessel cord
partial mole in diandric
IUGR, macrocephaly and small placental in digyny
153
what is PIGD?
PIGD aims to select embryos that are unaffected for transfer to the uterus to prevent the birth of a baby with a disease causing genetic mutation.
154
at what stages can cells be sampled for PIGD
1. biopsy of polar body just prior to conception (1st polar body) or just after fertilisation (2nd polar body)
2. biopsy of day 3 cleavage stage embryo (5-cells)
3. biopsy of trophoectoderm from bastocyst (day 5-6 embryos)
155
what techniques are used fro PIGD?
used to be FISH but now being replaced by array (structural) and PCR (molecular diagnosis by linkage) can also do direct mutations analysis by sanger
156
what diseases has PGD been added to?
CF, HD, NF1, DMD/BMD, chromosomal rearrangments
157
what is PGS
Preimplantation genetic screening-screens embryos for aneuploidies with the aim of selecting euploid embryos for transfer
the use for PGS is controversial, at least 11 randomised control trials have found no clinical benefit and 1 study has shown that PGS significantly lowered the birth rate from IVF when applied to women of advanced maternal age
Not currently supported by or offered by the NHS
158
Which group is in charge of determining the clinical criteria for offering PIGD in the NHS?
NHS reference group for medical genetics published a clinical commissioning policy for PGD which outlines the conditions under which PGD can be commissioned
159
what are the aims of the PGD NHSE clinical commissioning group?
- provide guidance, consistency and clarity in the commissioning of PGD services
- imporve equality of access to PGD
- ensure PGD is offered for diseases where there is sufficient evidence of clinical benefit and cost effectiveness
- reduce variation in clinical practice and conditions referred for PGD
160
what are the requirements for a couple to receive PGD?
NHS will commission up to 3 courses of PGD for a couples
- at risk of having a child with a sever genetic disease >10% risk
- fertile- different to IVF treatment for infertility
- HEFA licence for the disease
- genetic counselling
- female partner <40yrs and BMI between 19 and 30
- both partners should be non-smokers
161
what are the exclusions for PGD?
not offered for sex selection for family balancing in the UK
- PGS is not commissioned in the NHS as there is no evidence that it provides clinical benefit
- not to be used to address fertility problems or recurrent miscarriage with unknown aeitiology
162
what is the HFEA and what do they do?
human fertility embryology authority
- licence and monitor UK fertility clinics and all research involving embryos on the UK
- licence that diseases for which PGD can be offered
163
what is the method for PGD?
IVF--> biopsy cells --> genetic test --> transfer unaffected embryos
164
what are the stages in IVF?
in IVF a women undergoes ovarian stimulation then oocyte removal. The oocyte can then be fertilised by a sperm in a petri dish. In IVF the sperm and the egg and incubated in the same petri dish and the sperm are left to fertilise the egg. Improved fertilisation can be achieved by ICSI where the sperm is injected into the egg.
sperm for ICSI are morphologically selected- sperm with a large vacuole are more likely to result in aneuploidy
165
what are the different stages that cells can be biopsied for PGD
- 1st and 2nd polar body
- cleavage stage embryo (day 3)
- blastocyst (day 5-7) trophoectoderm
- balstocyst- blastocoele fluid
166
what are the advantages and disadvantages of the sampling 1st and 2nd polar body?
longer time to complete genetic testing without requiring crypreservation,
can only detect maternal mutations
generally only used in countries where testing cleavage stage embryos is illegal - Austria, Switzerland and Germany
167
what are the advantages and disadvantages of the sampling cleavage stage embryo (day 3)?
well assessed methodology
may hamper embryo viability
60% show mosaicism for anueploidy but rectified by the blastocyst stage
168
what are the advantages and disadvantages of the sampling blastocyst (day 5-7) trophoectoderm?
more cells to test = more DNA for analysis
| may not be representative of the inner cell mass which goes on to develop into the embryo
169
what are the advantages and disadvantages of the sampling balstocyst- blastocoele fluid?
less invasive and safe and contains DNA from ICM and trophoectoderm
only ~ 50% of samples contain DNA suitable for analysis and there can be low concordance with cleavage stage embryos
170
what is the non-invasive preimplantation genetic screening approach?
embryoonic DNA is isolcated from the blastocyst culture medium combined with blastocoele fluid can be used for genetic testing.
- aspiration of blastocoele fluid is less technically challenging and invasive than TE cell biopsy
origin of DNA in the blastocoele fluid is still under investigation and is routinely discarded during hr vitrification process to prevent formation of crystals
171
what is PGD used for?
Monogenic disorders
- 1st PGD cases where directed at the Y chromosome for X-linked disease
- HD is the most commonly tested for AD disease - direct and indirect testing is available
- CF- due to the high number of mutations a muliplex linkage based approach has been used so that the same design can be used for all mutations.
172
how can PGD be applied to mitochondrial disease?
PGD can be used to select embryos with a low level of the mutatn mtDNA
less prone to allele drop our as there is a high copy number of mtDNA
when there is a high level of mut mtDNA or homoplasmy PGD is not suitable
main issue is even if an embryo is selected with a level of heterpolasmy below that associated with disease they may still be affected due to the mitochondrial bottleneck which could result in higher levels in the fetus
173
what is PGH?
PG haplpotyping
used for diagnosis of monogenic disorders by using linkage to determine if the high or low risk allele has been inherited
- same test can be used for all families rather than needing to design mutation specific tests for each mutation
- can be used for disease where the causative mutation has not been determined
- ideally need DNA from parents and an affected proband to ID the high risk allele but an unaffected family member can be used for exclusion testing
- useful if mutation is not amenable to PCR detection
- need markers either side of gene and should determine that there are enough informative markers before offering test
- need to determine the risk of a double recombination resulting in low risk haplotype carrying the causative mutations
- where there is an informative marker either side of the gene the risk of a double recombination is much lower than if the informative markers are only on one side.
174
what is MDA- mutiple displacement amplification?
used bacteriophage fro whole genome amp meaning lots of DNA can be generated from small samples for genotyping multiple polymorphic markers
175
what are the limitations of PGD that can lead to misdiagnosis?
```
confusion of embryo and cell line number
transfer of the wrong embryo
mat/pat contamination
allele drop-out
probe/primer failure
chromosomal mosaicism
```
176
how can PGD impact epigenetics?
children born from ART are at a greater risk of suffering from and imprinting disorder- 10% increased risk of BWS
this is because the epigenetic genome undergoes 2 waves of reporgramming
- in primordial germ cells
- during preimplantation genetic development
ART is through to interfere with this
however there is not a large cohort to study and this also makes analysis of trangenerational effects on imprinting difficult to study
- most parents will still chose to have PGD despite the associated risks
177
what are some of the ethical concerns of PGD?
- morally acceptable? many people put higher value on the fetus than embryo making PGD preferable to PND
- the risk of designer babies if exaggerated and improbable providing there is adequate legislation
- need to consider the severity of disease, age of onset (number of disease free years), treatments available
- variable penetrance diseases?
178
what is non-disclosure testing?
not permitted in most centers
client may not want to know if they have the disease but this is unethical to uphold if you find they have zero risk as they will be undergoing unnecessary and risky procedures for no reason
179
Is HLA typing?
used to conceive a child that is a HLA to a sibling so they may be a stem cell donor for a critically ill sibling if no other donor is available
preferential procedure may be to harvest embryonic stem cells from matched embryos
180
what is the reported rate of mosaicism in CVS and AF?
1-2% in CVS
0.1-0.3% in AF
lower level in AF likely represents pregnancy loss in the time between CVS and AF sampling and CPM
181
give a brief description of embryology?
fertilisation --> zygote--> early blastocyst
- trophpectoderm goes on to form the placenat and extra-embryonic material
- ICM form the amnion and embryo
the amniotic cavity and embryo are formed from a different cell lineage from the chorion. The chorion differentiates early and is rapidly dividing do more likely to contain mitotic errors.
182
what cells are in the AF?
AF contains cells shed from the embryo- skin, fetal urine, cells from the fetal respiratory tract
cell are derived directly from the phenotype and are more likely to be a true representative of the fetal genotype
183
How can mosaicism develop?
Mitotic NDJ
| Trisomy rescue
184
what is level I , II and III mosaicism?
level 1- 1 cell in 1 culture - pseudomosaic
level 2- > 1 cell in 1 culture - may be pseudomosaic or true mosaicism
level 3 - > 1 cell in > 1 culture- likely to represent true fetal mosaicism and reported
type 1 and 2 may be a cultural artefact
185
what are the BPG required work up for exclusion of mosaiscism in AF and CVS? Hsu and Benn 1999
basic- 20 cells from 2 cults, 1 of whihc may contain an abnormal metaphase
moderate- 20 cells from an additional separate culture
extensive - 20 cells from 2 additional separate cultures
186
how can mosaicism be excluded in FBS?
single cell abn can be clinically significant and 30 additional metaphases should be screened
If FISH is used 50 cells should be screened for exclusion of mosaicism
187
what are the BPG for reporting mosaicism?
pseudomosaicism is considered an artefact by application of Hsu and Benn
if level II mosaicism is sen on CVS it may be appropriate to mention on report and recommend additional follow-up with USS and AF
for level III mosaicism the number of cells in each cell line should be included in the report with the caveat that the levle may be different in other tissues
if a chr involved in UPD is involved UPD studies should be activated
188
describe pallister killian
iso(12p)
- normal karyotype in peripheral blood due to the high turnover rate of cells and instability of the iso(12p) meaning it is lost from these cells
- can be detected in skin, AF, CVS
- only ever seen as mosaic
189
what is a marker chromosome?
small piece of extra chromosomal material - smaller than chr 22 and unable to be clearly identified by G-banding alone
- usually present in additional to the normal chromosome complement and termed supernumary marker chromosome or extra structurally abnormal chromosome
190
what structure can a marker have?
ring
inv dup
centric minute
191
what determines the phenotypic consequence of a marker chromosome?
```
size
genetic content
level of mosaicism
euchromatic/heterochromatic
result in copy number gain of the region in the marker
```
192
what is the most frequent marker chromosome?
markers derived from chromosome 15 are the most common
193
what causes cat eye syndrome?
+ psu idic (22)(q11.2)
194
what causes Emanuel syndrome?
+ der(22)t(11;22)(q23;q11)
195
When may markers be identified?
prenatal or postnatal array- detect presence of duplication and investigated by follow-up karyotype to confirm as a marker
karyotype investigation of an infertile male/ POF female
196
what should be the management if a marker is detected?
- determine genetic content
- check to see if it is mosaic
- test parents to inform on pathogenicity and recurrence risk
- if marker involves a chromosome involved in UPD it is relevant to check for UPD for the sister chromosome for UPD
197
How can the genetic content of a marker be determined?
arrayCGH best first option
If the marker only contains heterochromatin may require FISH to determine origin
chr 15 most common
198
what is the consideration of inheritance of a marker?
if inherited more likely to be benign
- phenotype expected to be similar to parent
however may be different if mosaic as level and tissue distribution can affect penetrance
199
how can cyto banding help ID a marker?
provide structural info
main limitation is resolution
can be used to determine level of mosaicism and is more sensitive than array
200
what is the use of C-banding to investigate a marker chromosome
C-banding can be used to determine the amount of heterochromatic and determine if a marker is acrocentric or not.
can also determine the number of centromeres to ID is mono or dicentric
markers that are all het are expected to have low risk as they contain little active genetic material
201
what is the use of silver staining to investigate a marker?
stains ribosomal RNA in the nucelolar organiation region (NOR)
- found on short arms of acrocentric chromosomes
202
what is the use of FISH when investigating a marker?
can use sequential FISH
- if acrocentric start with 15
- limited benefit if inhertied from normal parent so may just want to start with chromosomes involved in UPD (6,7,11,14,15,)
- costly and inefficient
- if abn pheno may also want to FISH for chr 12, 18 and 22 and iso chro associated with severe pheno
203
Markers derived from sex chromosomes?
- if karyotype is correlated with TS the marker is likely to be derived from a sex chr e.g. 45,X + mar
- important to determine if Y chr derived due to the risk of gonadoblastoma
- if X chr derived can test for Xist- if present marker will be inactive and more likely benign
204
what the use of array to investigate a marker?
- can ID full extend of marker and hence gene content and clinical significance
- may miss low level mosaic and het markers
- can ID markers consisting or more than 1 chromosome
- cannot ID the structure
205
what should be considered if a chr15 marker is identified?
need to distinguish between benign small marker idic(15) and larger pathogenic marker idic(15) which contains the PWS/AS region and is associated with LD, epilepsy and ASD
206
what should be considered if a ring chr marker is detected?
may see dynamci mosaicism and loss, gain, doubling up in different cells resulting in generalised ring syndrome and short stature
207
what is cffDNA derived from?
cells apoptosed from the outer layer of the placenta - cytotrophoblasts and syncytiotrophoblast - therefore it is at risk of CPM
208
from what gestation can cffDNA be tested?
can be observed from ~ 5 weeks but generally need to wait till 8-10 weeks before there is enough for reliable testing (high enough fetal fraction)
benefit is testing in early pregnancy
209
what size cffDNA, how does this differ from maternal?
cffDNA is ~200-300bp long
| shorter than mat cffDNA and this difference can be used to enrich for cffDNA for genetic testing ang
210
what are the components of CVS?
inner mesenchyme, diofferentiates later and more closely related to fetus
outer cytotrophoblast
211
when can CVS be sampled?
early CVS is associated with fetal limb defects
taken in 1st trimester 10-13+6 weeks gestation
212
what is the miscarriage risk associated with CVS?
1-2% according to RCOG, risk influenced by experience of operator and due to infection, amniotic leakage or damage to the placenta
213
How is CVS sampled?
10-25mg of CVS is taken from the placenta via a needle and syringe under USS guideance
214
what is the advantage and disadvantage of CVS compared to AF?
can be taken in earlier gestation giving more time for pregnancy decision making and follow-up testing
Risk of CPM
slightly increased miscarriage rate
215
what proportion of CVS show CPM?
1.5-2%
216
what is the amniotic fluid and what cells does it contain?
fluid filled sac that surround the developing fetus and facilitates symmetric growth and lung development
cells shed from fetal skin, fetal urine and fetal respiratory tract = more representative of true fetal genotype
217
how is AF taken?
transabdominally using syringe
| 10-20mls
218
what is the miscarriage risk for AF?
0.5-1% RCOG
219
what gestation can AF be taken?
2nd and 3rd trimester
from ~15 weeks, early AF has an increased miscarriage risk and low test success as not enough fetal cells. late gestation AFs are also more dirty and prone to failure
220
when can fetal blood be sampled?
from ~ 18-20 weeks whennveins are sufficiently developed
221
how is fetal blood sampled?
blood is taken by needle from umbilical cord or fetal blood vessels
222
what is the miscarriage risk for fetal blood sampling?
2%
223
for what reasons may fetal blood be sampled?
most commonly for:
- suspected fetal blood disorders e.g. fetal anemia
- inconsistent CVS and AF result follow-up
- fetocide/MTOP
- immunologic tests
224
what other prenatal samples may be taken?
pleural effusion when excess fluid removed
| POC from miscarriage or TOP
225
what is the main MCC risk in CVS and AF?
CVS- need to carefully dissect to remove the maternal decidua
AF- bloodstaining may be maternal (or fetal) also risk of tissue plug
- discarding the first draw of AF can reduce the risk of MCC
226
what is the impact of culturing on MCC for CVS and AF?
culturing conditions for AF favors the growth of the amniocytes to reduce or eliminate mat blood cells
In CVS and POC the culturing process does not reduce the risk of MCC as the maternal and fetal tissue will both be grown
227
what confluency of cultures is required for karyotype?
75% confluent monolayer- can be less for a targeted test of a few cells
228
According tot CMGS MCC BPG when should MCC be suspected?
- normal female in bloodstained sample
- XX/XY mixed genotype
- presence of mixed genotype on PCR
- mosaicism in a female
- uncertainty about the ID of a tissue sample in culture
- slow growth, especially when originating from a single piece of tissue or a small number of colonies.
229
when should MCC testing be performed?
if MCC suspected
for all monogenic tests- need to test mate sample against prenatal. lab use QF-PCR whihc has sensitivity to ~10% therefore can only state that significant MCC has been excluded
BP for the testing lab to do the MCC and needs to be on the DNA aliquot that was tested.
sensitivity of MCC assay may be lower than of the test so confirmation of result that are the same genotype as the mother may require confirmation y another method e.g. linkage following positive PCR result
should not issue report until MCC has been completed
230
How common in mosaicism in AF?
0.1-0.3% (lower than CVS)
231
what is the clinical relevance of CPM?
- observation of aneuploidy in CVS but not in AF = risk of UPD by trisomy rescue (chr 6, 7, 11, 14, 15)
most pregnancies with CPM continue to term with no abn but in some cases the abn in the placenta can lead to placental insufficiency and IUGR or other abn
232
what factors play a role on the impact of CPM?
Origin of error – somatic errors are associated with less severe consequences.
Level of mosaicism – there is a correlation between a high number of aneuploid cells and poor pregnancy progress.
Specific chromosomes – the influence of CPM on fetal growth is chromosome specific. Certain chromosomes carry imprinted genes involved in growth or placental function, which may contribute to impaired pregnancy progress when CPM is detected. CPM involving sex chromosomes usually has no adverse effect on foetal development.
Type of chromosome abnormality – marker chromosomes are more often confirmed in the fetus than trisomies.
233
How can CPM risk be minimised in prenatal testing?
- test mutltiple villi and chop/digest to ensure mesenchyme and trophoblast are sampled
- follow-up mosaic CVS with long term cult as mesenchymal core culture results are more likely to reflect a true fetal mosaicism than direct preparation
- follow-up mosaic with AF and detailed USS
decision regarding pregnancy should not be made on the
234
what is the difference between meiotic and mitotic CPM and what are the associated aneuploidies?
Mitotic CPM – Mitotic non-disjunction can occur in a trophoblast cell or a non-fetal cell from the inner cell mass creating a trisomic cell line in the tissue which is destined to become the placental mesoderm. Trisomy 2, 3, 7 and 8 are derived mitotically.
Meiotic CPM –Due to trisomy rescue of a trisomic conceptions Trisomy 16 and 22 are derived meiotically.
235
what factors influence the pattern of normal and abnormal cells in the developing embryo?
- reduced or improved replicative rates of the normal and abnormal cell lines
- abnormal cells may fail to differentiate or function properly
- may be no selection against abnormal cells but there presence could compromise the pregancy e.g. placedntal insufficiency, IUGR
236
what are the 3 types of CPM and there prevalence?
Type I, the abnormal cell line is confined to the cytotrophoblast (40%)
Type II, it affects only the mesenchymal cells of the stromal villous core (40%)
Type III, it involves both tissues. (7%)
Type I and II CPMs are usually the consequence of nondisjunction events, and type III is the result of “trisomy rescue”
type III carries a risk of CPM in the rescued diploid fetus
237
which mosaics are more likely to involve the fetus and hence considered high risk?
trisomies 8, 9, 12, 13, 15, 18, 20, and 21
238
what is the impact of CPM on arrayCGH
Karampetsou et al. (2014) report a case of CPM involving a deletion of exons 7-10 of the STS gene detected by array-CGH using DNA extracted from uncultured CVS.
therefore suggested that reports on uncultured CVS should include the rider that CPM has not been ruled out and ideally confirmation should be on cultured CVS or AF
