Placentas from pregnancies conceived by IVF/ICSI have a reduced DNA methylation level at the H19 and MEST differentially methylated regions.
OBJECTIVE
Does IVF/ICSI have an effect on the epigenetic regulation of the human placenta?


CONCLUSIONS
We found a reduced DNA methylation level at the H19 and MEST differentially methylated regions (DMRs), and an increased RNA expression of H19 in placentas from pregnancies conceived by IVF/ICSI when compared with placentas from spontaneous conception.


BACKGROUND
Changes in fetal environment are associated with adverse health outcomes. The placenta is pivotal for intrauterine environment. Animal studies show that epigenetic regulation plays an important role in these environment-induced phenotypic effects. Also, the preimplantation embryo environment affects birthweight as well as the risk of chronic adult diseases. Epigenetic processes are sensitive to the environment, especially during the period around conception.


METHODS
Placental tissue was collected from 35 spontaneously conceived pregnancies and 35 IVF/ICSI (5 IVF, 30 ICSI) derived pregnancies. We quantitatively analysed the DNA methylation patterns of a number of consecutive CpGs in the core regions of DMRs and other regulatory regions of imprinted genes, since these are involved in placental and fetal growth and development.


METHODS
By using pyrosequencing, the DNA methylation at seven germline-derived primary DMRs was analysed quantitatively. Five of these are maternally methylated (MEST isoform α and β, PEG3, KCNQ1OT1 and SNRPN) and two are paternally methylated [H19 DMR and the intergenic region between DLK1 and MEG3 (IG-DMR)]. The post-fertilization-derived secondary DMRs, IGF2 (DMR0 and 2) and IG-DMR (CG7, also called MEG3 DMR), and the MEG3 promoter region were examined as well. In case of differential methylation between the two groups, the effect on gene expression was assessed by quantitative real-time PCR.


RESULTS
Both the promoter region of MEST isoform α and β and the 6th CTCF binding site within the H19 DMR were significantly hypomethylated in the IVF/ICSI group. The phenomenon was consistently observed over all CpG sites analysed and not restricted to single CpG sites. The other primary and secondary DMRs were not affected. Expression of H19 was increased in the IVF/ICSI group, while that of IGF2 and MEST remained similar.


CONCLUSIONS
In the IVF/ICSI group, mostly ICSI pregnancies were investigated. The ICSI technique or male subfertility could be a confounding factor. Therefore, our results are less generalizable to IVF pregnancies.


CONCLUSIONS
The clinical effects of the observed placental hypomethylations on the developmental programming of the IVF/ICSI progeny, if any, are as yet unknown. Whether the hypomethylation is an adaptation of the placenta to maintain fetal supply and ameliorate the effects of environmental cues, or whether it is a deregulation leading to deranged developmental programming with or without increased vulnerability for disease, consistent with the developmental origins of health and disease hypothesis, needs further investigation.


BACKGROUND
Partly funded by an unrestricted research grant by Organon BV (now MSD BV) without any role in study design, data collection and analysis, or preparation of the manuscript. No conflict of interests to declare.


BACKGROUND
Dutch Trial Registry (NTR) number 1298.
Department of Obstetrics & Gynaecology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Centre, Maastricht, The Netherlands.
Ewka C M
EC
Nelissen
John C M
JC
Dumoulin
Antoine
A
Daunay
Johannes L H
JL
Evers
Jörg
J
Tost
Aafke P A
AP
van Montfoort
D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D005307:Fertilization in Vitro; D006801:Humans; D010920:Placenta / Q000378:metabolism*; D011247:Pregnancy; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011506:Proteins / Q000235:genetics* / Q000378:metabolism; D062085:RNA, Long Noncoding / Q000235:genetics* / Q000378:metabolism; D020554:Sperm Injections, Intracytoplasmic
C094335:H19 long non-coding RNA; D020033:Protein Isoforms; D011506:Proteins; D062085:RNA, Long Noncoding; C103637:mesoderm specific transcript protein