Human Reproduction, Vol.24, No.6 pp. 1330 – 1338, 2009
Advanced Access publication on February 26, 2009
Pregnancies conceived using assistedreproductive technologies (ART) havelow levels of pregnancy-associatedplasma protein-A (PAPP-A) leading toa high rate of false-positive resultsin ﬁrst trimester screeningfor Down syndrome
D.J. Amor1,2,3,4,8, J.X. Xu1,2, J.L. Halliday1,2, I. Francis3, D.L. Healy5,6,
S. Breheny5,6, H.W.G. Baker4,7, and A.M. Jaques1
1Murdoch Childrens Research Institute, Royal Children’s Hospital, Parkville, Victoria, Australia 2Department of Paediatrics, University ofMelbourne, Melbourne, Victoria, Australia 3Victorian Clinical Genetics Services, Royal Children’s Hospital, Parkville, Victoria, Australia4
Melbourne IVF, East Melbourne, Victoria, Australia 5Department of Obstetrics and Gynaecology, Monash University, Clayton, Victoria,
Australia 6Monash IVF, Richmond, Victoria, Australia 7Department of Obstetrics and Gynaecology, University of Melbourne, Royal Women’sHospital, Parkville, Victoria, Australia
8Correspondence Address. Murdoch Childrens Research Institute, Royal Children’s Hospital, Flemington Rd, Parkville 3052, Australia. Tel: þ61-3-83-41-62-93; Fax: þ61-3-83-41-63-90; Email: [email protected]
background: First trimester screening (FTS) for Down syndrome combines measurement of nuchal translucency, free beta-humanchorionic gonadotrophin and pregnancy-associated plasma protein-A (PAPP-A). The aim of this study was to undertake a detailed analysisof FTS results in singleton pregnancies conceived using assisted reproductive technologies (ART) and non-ART pregnancies.
methods: A record linkage study compared outcomes in 1739 ART-conceived and 50 253 naturally conceived pregnancies.
results: Overall, signiﬁcantly lower PAPP-A levels were detected in ART pregnancies (0.83 multiples of median, MoM) than in controls(1.00 MoM) (t-test P , 0.001). This difference remained after excluding complicated pregnancies. Analysis of factors affecting PAPP-A levelssuggested fresh compared with frozen embryo transfers and use of artiﬁcial cycles compared with natural cycles for frozen transfers wereassociated with lower values. The adjusted odds ratio (AdjOR) for receiving a false-positive result was 1.71 (95% CI 1.44 – 2.04; P , 0.001)for ART pregnancies compared with non-ART pregnancies, and this leads to a higher AdjOR (1.24, 95% CI 1.03 – 1.49; P ¼ 0.02) for having achorionic villous sampling (CVS) or amniocentesis.
conclusions: ART pregnancies have reduced FTS PAPP-A levels leading to an increased likelihood of receiving a false-positive resultand having a CVS/amniocentesis. Lower PAPP-A may reﬂect impairment of early implantation with some forms of ART.
Key words: ART / PAPP-A / pregnancy screening / pregnancy complications / hormonal stimulation
based on maternal age alone was ﬁrst described in 1988 (Wald et al.,1988). Over the last 10 years, second trimester serum screening has
Pregnancy screening for Down syndrome (DS) and other chromo-
been progressively replaced by ﬁrst trimester combined screening.
some abnormalities has become part of routine antenatal care over
The ﬁrst trimester combined screen measures maternal serum levels
the last 20 years. The measurement of second trimester biochemical
markers in the blood of pregnant women to improve screening for DS
pregnancy-associated plasma protein-A (PAPP-A) at 9 – 12 weeks
& The Author 2009. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: [email protected]
gestation and measures nuchal translucency (NT) by ultrasound at
We hypothesized that in pregnancies conceived using ART, factors
11 – 13 weeks gestation. These measurements are combined with
exist that are absent in natural conceptions, potentially inﬂuencing the
maternal age, weight and gestational age to produce a risk estimate
marker levels, and consequently, risk results of the ﬁrst trimester com-
of the fetus having DS or trisomy 18 (T18) (Wald et al., 2003). For
bined screen. The aims of this study were to investigate in a large
pregnancies at increased risk, prenatal diagnostic testing, CVS or
population-based sample, the effect of ART on the individual
amniocentesis, can be offered. In the Victorian population, the sensi-
markers of the ﬁrst trimester combined screen (fb-hCG, PAPP-A
tivity of the ﬁrst trimester combined screen for DS is 91% (using a risk
and NT) and on the FPR. We also aimed to investigate the effect of
threshold of 1 in 300 at the time of ultrasound), with the proportion of
different ART modalities on these markers, and the impact of ART
unaffected pregnancies receiving an increased risk result (false positive
and screening results on the uptake of prenatal diagnostic testing
rate, FPR) being 3.9% (Jaques et al., 2007).
(CVS and amniocentesis) following ﬁrst trimester combined screening.
Pregnancies conceived using assisted reproductive technologies
(ART) now account for 3% of all live births in Australia (Wang
et al., 2007). Previous studies of second trimester serum screeninghave shown that serum markers in ART pregnancies differ from
The effect of ART (IVF, ICSI, GIFT, embryo cryopreservation and
natural conceptions, leading to an increased FPR (Barkai et al., 1996;
hormone treatment) on the FPR of the ﬁrst trimester combined screen
Ribbert et al., 1996; Frishman et al., 1997; Wald et al., 1999; Raty
and its three components (fb-hCG, PAPP-A, NT) was examined in
et al., 2002; Lambert-Messerlian et al., 2006). The effect of ART on
women with singleton pregnancies screened between February 2000
ﬁrst trimester combined screening has been examined in 13 studies
and June 2004 in Victoria, Australia. The pregnancies included did nothave a fetus or produce a child with a birth defect, and singleton pregnan-
that have yielded contradictory and inconclusive results, largely due
cies were deﬁned as the presence of one fetus after 20 weeks gestation.
to small sample sizes. Some studies found that fb-hCG (Liao et al.,
Data from three separate databases were linked to obtain the study popu-
2001; Niemimaa et al., 2001; Ghisoni et al., 2003; Bersinger et al.,
lation (Fig. 1). A fourth prenatal diagnosis database was linked to the study
2004) and NT thickness (Maymon and Shulman, 2004; Hui
population to investigate the uptake of CVS and amniocentesis. The four
et al., 2005b) were increased in ART pregnancies, yet others found
no differences in fb-hCG (Wojdemann et al., 2001; Maymon and
The Victorian Perinatal Data Collection Unit (VPDCU) is responsible
Shulman, 2002; Orlandi et al., 2002; Maymon and Shulman, 2004;
for the collection of information on all Victorian births 20 weeks’ ges-
Bellver et al., 2005; Lambert-Messerlian et al., 2006; Anckaert et al.,
tation or 400 g birthweight. Reporting to the VPDCU is mandatory
2008) or NT (Liao et al., 2001; Niemimaa et al., 2001; Wojdemann
and data collected includes obstetric factors, neonatal outcomes, birth
et al., 2001; Maymon and Shulman, 2002; Orlandi et al., 2002;
defects and previous pregnancy history. A unique registration number isassigned to each newborn. During the study period there were
Ghisoni et al., 2003; Lambert-Messerlian et al., 2006). PAPP-A levels
245 000 singleton births in Victoria.
have been decreased in some studies (Liao et al., 2001; Maymon
The Victorian Birth Defects Register collects data on all pregnancies
and Shulman, 2002; Orlandi et al., 2002; Bersinger et al., 2004;
diagnosed with a birth defect, comprising live births, stillbirths, neonatal
Maymon and Shulman, 2004; Hui et al., 2005a; Tul and Novak-Antolic,
deaths and terminations. For this study, all pregnancies that resulted in a
2006; Anckaert et al., 2008), but unaltered in others (Niemimaa et al.,
birth defect were identiﬁed and excluded from the study population.
2001; Wojdemann et al., 2001; Ghisoni et al., 2003; Bellver et al.,
The Victorian Clinical Genetics Service (VCGS) conducts all ﬁrst trime-
2005; Lambert-Messerlian et al., 2006).
ster combined screening tests in Victoria. The VCGS ﬁrst trimester
Figure 1 Flow diagram showing data linkage between three separate databases to obtain study population.
combined screening programme has been described elsewhere (Jaques
before oocyte collection. Hormone treatment accompanying frozen –
et al., 2006, 2007). Brieﬂy, fb-hCG and PAPP-A measurements were per-
thawed embryo transfers was deﬁned as any combination of estradiol
formed on a KRYPTOR analyser, and NT and crown– rump length (CRL)
and progesterone for artiﬁcial cycles for oligomenorrhoea/amenorrhoea,
measurements were made off-site by multiple ultrasonologists in accord-
and clomiphene with or without HCG for ovulation induction to time to
ance with the technique described by the Fetal Medicine Foundation
(Nicolaides, 2004). NT and biochemistry values were calculated fromCRL in ART and non-ART pregnancies because this was the only way
to directly compare ART and non-ART pregnancies, and because it has
Ethics approval was obtained from the Human Research Ethics Commit-
been shown that CRL and date of oocyte collection are practically equiv-
tees of the Victorian Department of Human Services, Mercy Health and
alent when calculating gestational age for ﬁrst trimester screening (FTS)
Aged Care, Royal Women’s Hospital, Freemasons Hospital, Epworth
(Gjerris et al., 2008). The biochemistry and risk estimates were calculated
Hospital, Monash University and Monash Surgical Private Hospital.
using software developed in house and results are monitored byUKNEQAS and audited by ascertaining pregnancy outcomes for calcu-lation of speciﬁcity and sensitivity as described elsewhere (Jaques et al.,
2006, 2007).The VCGS prenatal screening database contains comprehen-sive data for all women who had ﬁrst trimester combined screening in the
Women in the non-ART and ART groups were similar in terms of
maternal weight and gestational age at ultrasound (calculated from
The data on ART in Victoria were obtained from the three major pro-
CRL of the fetus) at the time of screening (Table I). Maternal
viders: Melbourne IVF, Monash IVF and Melbourne Assisted Conception
country of birth was Australia or Europe/UK for 91.7% of ART
Centre, accounting for 98% of ART pregnancies in Victoria. The dataset
mothers and 91.8% of non-ART mothers (P ¼ 0.90). The ART
included detailed information on the ART cycles, including the type of
group were older and more likely to be primigravid than the
ART treatment, indication for ART, whether embryos were fresh or
non-ART group. The measured CRL was less for fresh embryos
frozen-thawed, the use of donor gametes or embryos and the number
of fetal hearts present at ultrasound at 6 weeks gestation.
Data for all CVS and amniocentesis tests performed in Victoria and their
results are provided to the VPDCU by all four cytogenetic laboratories in
the State. Information regarding the uptake of CVS and amniocentesis was
PAPP-A levels were signiﬁcantly lower (P , 0.001) in ART pregnancies
available for the 41 756 pregnancies screened in 2002 – 2004. Excluded
(0.83 MoM) compared with non-ART pregnancies (1.0 MoM)
from this subset of analyses were 10 236 pregnancies screened in
(Table II). A small increase in the NT was detected in the ART
2000 – 2002 because these pregnancies were not linked to the prenatal
group (0.91 MoM) compared with the non-ART group (0.90 MoM)
(P ¼ 0.004). Analyses of fb-hCG levels showed no signiﬁcant differ-ence between ART (0.99 MoM) and non-ART (0.98 MoM)
Data coding and statistical analyses were conducted using SPSS (version
Adverse pregnancy complications, comprising pregnancies with
15; SPSS Inc., Chicago, IL, USA). Mann – Whitney U-tests were done to
adverse perinatal outcomes (stillbirth, neonatal death, prematurity,
test the difference between the characteristics (age, weight, CRL and
birthweight ,2500 g) and/or obstetric complications (pre-eclampsia,
gestational age of testing) of non-ART and ART groups, and chi-square
pregnancy-induced hypertension, gestational diabetes) were more
tests were performed to assess the difference in the proportion of
common in the ART pregnancies (21.0%) compared with the
women from the different groups who were primigravid. Independent
non-ART pregnancies (13.9%). The PAPP-A levels were lower in
samples t-tests were performed on the means of the log-transformed
the complicated pregnancies overall but there was still a signiﬁcant
multiples of median (MoM) values for fb-hCG, PAPP-A and NT. Results
difference between the ART and non-ART groups (Table II). Among
were then back-transformed and are presented as geometric means.
uncomplicated pregnancies, PAPP-A levels remained signiﬁcantly
The statistical signiﬁcance of differences in proportions between groups
reduced in ART pregnancies (0.85 MoM) compared with non-ART
and results were expressed as odds ratios (ORs) with 95% conﬁdence
pregnancies (1.02 MoM) (P , 0.001) (Table II).
intervals (CIs). Univariable logistic regression analysis was done toexplore associations between results and a number of covariables, e.g.
Analyses according to the aetiology of the infertility showed that
maternal age, maternal country of birth, gravidity, parity, birthweight and
PAPP-A levels were similarly reduced when the infertility was reported
gestational age. Multivariable analysis was adjusted for maternal age and
to be of female-only aetiology (0.82 MoM), male-only aetiology (0.85
gravidity as these were the only signiﬁcant confounders.
MoM) and when a combination of male and female aetiologies was
The ART study population was divided into subgroups to allow more
present in the couple (0.82 MoM). The differences between these cat-
in-depth analyses of the different exposures on outcome: (i) aetiology of
egories were not statistically signiﬁcant.
infertility (female-only aetiology, male-only aetiology, a combination of
Further analysis of the effect of ART on marker levels was under-
male and female aetiologies or unknown aetiology); (ii) ART procedure
taken using the subset of pregnancies that did not have adverse peri-
(IVF, ICSI or GIFT); (iii) embryo transfer (fresh or frozen – thawed); (iv)
natal and obstetric complications (Table III). Comparison of PAPP-A
presence or absence of exogenous hormone treatment and (v) presence
levels between non-ART pregnancies (1.02 MoM) and each subtype
of single or multiple fetal hearts at 6 weeks gestation. Although treat-
of ART showed that the reduction in PAPP-A applied to all three
ment protocols varied, hormone treatment accompanying fresh embryotransfers always was deﬁned as follicle stimulating hormone (FSH) or clo-
ART subtypes (IVF, 0.87 MoM; ICSI, 0.84 MoM; GIFT, 0.71 MoM).
miphene, with or without other drugs. All GIFT cycles were done with
Compared with non-ART pregnancies (1.02 MoM), PAPP-A levels
FSH. A small proportion (n ¼ 13) of fresh embryo transfers were done
were reduced for both fresh embryos transfers (0.79 MoM, t-test
without any hormone treatment to stimulate multifollicular development
P , 0.001) and frozen – thawed embryo transfers (0.95 MoM, t-test
Table I Characteristics of the study population
Values are given as median (range) and Mann – Whitney tests were performed for P-value for maternal age, maternal weight, CRL at NT scan, gestation at NT scan and gestation at bloodsampling. Primigravid is given as a percent and chi-square tests were performed for P-values.*Denotes P-value of ,0.05 for the different ART groups (All, IVF, ICSI, GIFT, fresh and frozen)versus non-ART comparisons, # denotes P-value of ,0.05 for IVF versus ICSI comparison, ^ denotes P-value of ,0.05 for fresh versus frozen – thawed comparison.
aP-value based on the independent samples t-test, values are compared with the non-ART group.bIncludes pregnancies with adverse pregnancy outcomes [neonatal death, preterm (,37weeks) and low birthweight (,2.5 kg)] and pregnancies with obstetric complications (pre-eclampsia, pregnancy-induced hypertension and gestational diabetes).
P ¼ 0.001), however, fresh embryo transfers were associated with a
Association between ART and false-positive
signiﬁcantly lower PAPP-A level when compared directly with
frozen – thawed embryos (t-test P , 0.001).
Table IV shows the comparison between the non-ART and ART
When examining the effect of hormone versus no hormone treat-
groups for the proportion of women receiving a false-positive result
ment irrespective of fresh or frozen – thawed embryo transfer, we
from ﬁrst trimester combined screening. Women conceiving using
found that transfer cycles that included any hormone treatment
ART had a signiﬁcantly increased likelihood of receiving a false-positive
resulted in lower PAPP-A levels (0.78 MoM) compared with those
result (OR 2.71, 95% CI 2.19 – 3.35; P , 0.001) compared with
without hormone treatment (0.99 MoM, t-test P , 0.001).
non-ART women. After adjusting for maternal age and gravidity,
ART women were still more likely to receive a false-positive result[adjusted OR (AdjOR) 1.71, 95% CI 1.44 – 2.04; P , 0.001]. The like-
lihood of receiving a false-positive result was higher for fresh embryo
Of the 1739 singleton ART pregnancies, 1604 had one fetal heart, 123
transfers than for frozen – thawed embryo transfers.
had two or more fetal hearts and results for 12 were not available. PAPP-A levels were 0.83 MoM for pregnancies with one fetal heart
and 0.97 MoM for pregnancies with multiple fetal hearts. Results forfb-hCG were 0.99 MoM for one fetal heart and 1.03 MoM for multiple
fetal hearts, and results for NT were 0.91 MoM for one fetal heart and
A higher proportion of women who conceived using ART (10.6%) had a
CVS or amniocentesis compared with their non-ART counterparts
Table III Effect of ART on marker levels—excluding complicated pregnancies
Fresh and frozen – thawed embryo transfer
Fresh and frozen – thawed embryo transfer by hormone treatmentb
Fresh without hormone treatment (n ¼ 11)
Frozen – thawed with hormone treatment (n ¼ 118)
Frozen – thawed without hormone treatment (n ¼ 455)
All embryo transfers by hormone treatmentb
aP-value based on the independent samples t-test, values are compared with the non-ART group. bHormone treatment accompanying fresh embryo transfers always included FSH or clomiphene, often in combination with other drugs. All GIFT cycles were done with FSH. Hormonetreatment accompanying frozen – thawed embryo transfers comprised various combinations of estradiol, progesterone, clomiphene and HCG.
Table IV Comparison of false-positive results
(5.3%) (OR 2.10, 95% CI 1.76 – 2.50; P , 0.001) (Table V). After adjust-
accounted for, the OR of having a CVS or amniocentesis was reduced
ing for maternal age, the ART group were still more likely to have a CVS
to 0.78 (95% CI 0.60 – 1.00; P ¼ 0.054) for the ART group compared
or amniocentesis compared with the non-ART group (OR 1.24, 95% CI
with the non-ART group, suggesting that the higher false-positive rate
1.03 – 1.49; P ¼ 0.023). When the increased risk result was also
was responsible for the higher uptake of invasive prenatal diagnosis in
the ART group. When examining the effect of hormone versus nohormone treatment, we found that after controlling for maternal age,
only women whose transfer cycles included hormone treatment were
more likely than non-ART women to have a CVS or amniocentesis
(AdjOR 1.38, 95% CI 1.11 – 1.71; P ¼ 0.003).
This is the largest and most comprehensive study to date on the inﬂu-
ence of ART conception on ﬁrst trimester combined screening, and is
important because of the increasing use of screening in all pregnancies.
This study comprises more than 1700 ART singleton pregnancies and
more than 50 000 non-ART singleton pregnancies, representing
almost as many ART pregnancies as all previous studies combined.
The primary effect of ART treatment was a signiﬁcant reduction in
the serum PAPP-A level compared with the non-ART conceptions,
whereas fb-hCG was not signiﬁcantly altered. A marginal increase in
NT was observed in ART pregnancies, however, this very small differ-
ence is likely due to operator effects, with ART pregnancies more
likely to be scanned by a small subset of operators linked to ART
clinics compared with non-ART pregnancies.
The pattern of markers observed in this study is similar to that
observed in several previous studies (Liao et al., 2001; Maymon andShulman, 2002, 2004; Orlandi et al., 2002; Hui et al., 2005b; Anckaertet al., 2008), but differs from others that detected no effect of ART on
PAPP-A levels (Niemimaa et al., 2001; Wojdemann et al., 2001;
Ghisoni et al., 2003; Bellver et al., 2005; Lambert-Messerlian et al.,
2006). Some studies have also suggested that ART results in an
increase in fb-hCG (Niemimaa et al., 2001; Ghisoni et al., 2003; Ber-singer et al., 2004). A likely explanation for the contradictory ﬁndings
of previous studies is their small sample sizes (sample size range 47 –
We have also shown that as a result of the decreased PAPP-A in
ART pregnancies, women conceiving using ART are more likely to
receive a false-positive result from the ﬁrst trimester combined
screen and are therefore more likely to have CVS or amniocentesis.
This ﬁnding is important because an increase in the uptake of CVS
and amniocentesis in healthy pregnancies will lead to an increase inparental anxiety and in procedure-related morbidity, including miscar-riage. The increase in false-positive results and uptake of prenatal diag-
nosis is seen for fresh and frozen – thawed embryo transfers, but only
in the subset of embryo transfers where the mother was given
hormone treatment around the time of embryo transfer. This study
only examined the FPR and not other indicators of screening test per-
formance such as sensitivity and speciﬁcity; therefore, no conclusion
can be drawn in relation to whether adjustments for calculation of
risk parameters need to be made. Following further investigation ofthe effectiveness of the ﬁrst trimester combined screen in ART preg-
nancies, it may be possible to modify screening protocols for ART
pregnancies in order to reduce the FPR without reducing the sensi-
The reduction in PAPP-A levels in ART pregnancies provides further
evidence that ART pregnancies are different from non-ART pregnan-
cies, an observation that may have implications beyond the combined
screen. PAPP-A is a growth factor that promotes growth by cleaving
insulin-like growth factor binding proteins (Lawrence et al., 1999)thereby increasing the bioavailability of insulin-like growth factors
(IGFs) (Conover et al., 2004). PAPP-A is present at low concen-
Novak-Antolic, 2006). We found signiﬁcantly lower PAPP-A levels for
trations in the blood of men and non-pregnant women, but is
embryos transferred fresh compared with frozen – thawed embryos.
detected at high concentrations in the blood of pregnant women
This ﬁnding is consistent with one previous study that separately exam-
(Lin et al., 1974), with blood levels rising soon after implantation
ined fresh and frozen – thawed embryo transfers and found a reduction
and increasing with gestation, peaking in the third trimester (Guibour-
in PAPP-A for fresh embryo transfers (Hui et al., 2005a). A second study
denche et al., 2003). In pregnant women, circulating PAPP-A originates
analysed PAPP-A levels in relation to the number of oocytes retrieved
at the interface between the placenta and the endometrium, where it
and found a greater reduction in PAPP-A following fresh embryo trans-
is produced by placental trophoblasts and decidualized endometrial
fers in stimulation cycles in which a greater number of oocytes were
stromal cells, and is hypothesized to regulate IGF-II bioavailability in
retrieved (Tul and Novak-Antolic, 2006), further suggesting that
the placenta and to facilitate implantation (Giudice et al., 2002).
hormone stimulation and the woman’s response to hormone treatment
Our data provide new insights into the possible mechanisms under-
lying reduced PAPP-A in ART pregnancies.
In contrast to previous studies, we also detected signiﬁcantly lower
One consideration is whether the presence of a ‘vanished twin’ in
PAPP-A levels for frozen – thawed embryos compared with non-ART
some ART pregnancies might explain the difference in results
pregnancies, indicating that the reduction in PAPP-A is not restricted
between ART and non-ART pregnancies. However, in our ART popu-
to fresh embryo transfers and prompting further analysis. The classiﬁ-
lation vanished twins appear to increase PAPP-A levels rather than
cation of fresh versus frozen – thawed embryos has potential to mask
decrease them; therefore, vanished twins are not responsible for
an effect of hormone treatment because in our study population a sig-
the observed reduced in PAPP-A levels in ART pregnancies. In fact,
niﬁcant proportion of frozen – thawed embryo transfers (20.6%) were
vanished twins may have the opposite effect, suggesting that the
accompanied by hormone treatment. When our data were stratiﬁed
number of fetal hearts at early ultrasound should be considered in
according to the presence or absence of any hormone treatment,
any modiﬁcation of screening protocols in ART pregnancies.
we found that reduced PAPP-A levels were seen only in the group
It has previously been suggested that a reduction in PAPP-A in ART
of pregnancies in which embryo transfer has been accompanied by
pregnancies might be an artefact of testing being undertaken at an
hormone treatment. In uncomplicated pregnancies where no
earlier gestation in this group (Maymon and Shulman, 2002). In our
hormone treatment was used, PAPP-A levels were normal.
population, there was no difference between the ART and non-ART
The endocrine changes that occur during early pregnancy, including
pregnancies in the timing of blood sampling or ultrasound. We did
the production of PAPP-A by the endometrium and placenta, are the
detect a slightly greater CRL for frozen – thawed embryos compared
result of a complex and poorly understood set of interactions between
with fresh embryos, equivalent to 1 day of gestational age. This
the corpus luteum, endometrium, placenta and embryo. Our data
difference may reﬂect a longer in vitro culture time for frozen –
suggest that the administration of exogenous hormones, occurring in
thawed embryos compared with fresh embryos, a difference that
fresh and artiﬁcial frozen – thawed ART cycles, interferes with the
would not affect the PAPP-A MoMs because these are adjusted for
normal endocrine changes of early pregnancy, resulting in reduced
PAPP-A levels are also known to be associated with adverse preg-
The contribution of different types of hormone treatment to the
nancy complications (hypertension, pre-eclampsia and gestational dia-
reduced PAPP-A in ART pregnancies is more difﬁcult to assess because
betes) and adverse perinatal outcomes (prematurity, low birthweight
treatment protocols are highly variable and more comprehensive treat-
and neonatal death) (Ong et al., 2000; Dugoff et al., 2004; Smith et al.,
ment data were not available. It is, however, notable that similarly
2006). ART is associated with an overlapping spectrum of pregnancy
reduced PAPP-A levels were seen for fresh embryo transfers that were
and perinatal complications (Maman et al., 1998; Schieve et al., 2002;
accompanied by FSH and/or clomiphene treatment, and for frozen–
Helmerhorst et al., 2004; Shevell et al., 2005); therefore, lower
thawed embryo transfers that were accompanied by treatment with
PAPP-A in ART pregnancies might simply be a predictor of these com-
various combinations of estradiol, progesterone, HCG and clomiphene.
plications that are known to be more common in the ART population
We propose a model whereby hormone treatment accompanying
(Maymon and Shulman, 2002; Bersinger et al., 2004). We were able to
embryo transfers results in abnormal levels of ovarian steroid hor-
analyse our data according to the presence or absence of these com-
mones and other factors yet to be identiﬁed, which in turn cause a
plications, and found that PAPP-A levels were reduced in ART preg-
reduction in PAPP-A production. Although PAPP-A is produced by
nancies with or without these complications.
the placenta, the reduction in PAPP-A is likely to be mediated via
It has also been suggested that lower PAPP-A levels in ART preg-
an effect of hormones on the endometrium because the effect is
nancies might be the result of metabolic impairments related to infer-
seen for hormone treatment administered prior to implantation and
tility in the mother (Maymon and Shulman, 2002). This hypothesis is
establishment of a placenta, possibly reﬂecting impairment of early
partly refuted by our data that show that PAPP-A is similarly
implantation with some forms of ART. Lower PAPP-A secretion
reduced in male-factor infertility compared with female-factor inferti-
should lower the availability of IGFs (Giudice et al., 2002) and
lity. However, the difference in PAPP-A results between frozen –
through this mechanism may directly contribute to low birthweight
thawed embryos transferred with and without hormone treatment
(Smith et al., 2002). It is well known that singleton babies born after
may be related to underlying ovulatory disorders and defective endo-
ART conception are at increased risk of being low birthweight or
metrial development in the mother. This requires further study.
small for gestational age (Schieve et al., 2002; Halliday, 2007), and
Our results support and extend the hypothesis that exogenous
those with low PAPP-A from FTS are also at increased risk of low
hormone treatment is the principal cause of reduced PAPP-A in ART
birthweight (Dugoff et al., 2004; Barrett et al., 2008). Moreover, it
pregnancies (Bersinger et al., 2004; Hui et al., 2005a; Tul and
has been observed that babies born from fresh ART cycles are at
increased risk of low birthweight and have a lower mean birthweight
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compared with babies born from frozen – thawed cycles (Wada
trimester biochemical screening for Down’s syndrome in singleton
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pregnancies conceived by assisted reproduction. Hum Reprod 2005;
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In conclusion, this study has provided conclusive evidence that ﬁrst
Liebaers I, Haentjens P, Bonduelle M. Neonatal outcome of 937 children
trimester maternal serum levels of PAPP-A are decreased in ART
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and comparison with outcome data of fresh ICSI and IVF cycles. Hum
bined screening. Our results highlight the importance of pre- and
post-test counselling for women carrying ART pregnancies. Further
Bersinger NA, Wunder D, Vanderlick F, Chanson A, Pescia G, Janecek P,
work should be undertaken to determine the viability of altering the
Boillat E, Birkhauser MH. Maternal serum levels of placental proteins
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Submitted on October 7, 2008; resubmitted on January 15, 2009; accepted on
in maternal mid-trimester serum screening for Down’s syndrome.
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