The cumulative effect of assisted reproduction procedures on placental development and epigenetic perturbations in a mouse model

Abstract

Assisted reproductive technologies (ART) are associated with several complications including low birth weight, abnormal placentation and increased risk for rare imprinting disorders. Indeed, experimental studies demonstrate ART procedures independent of existing infertility induce epigenetic perturbations in the embryo and extraembryonic tissues. To test the hypothesis that these epigenetic perturbations persist and result in adverse outcomes at term, we assessed placental morphology and methylation profiles in E18.5 mouse concepti generated by in vitro fertilization (IVF) in two different genetic backgrounds. We also examined embryo transfer (ET) and superovulation procedures to ascertain if they contribute to developmental and epigenetic effects. Increased placental weight and reduced fetal-to-placental weight ratio were observed in all ART groups when compared with naturally conceived controls, demonstrating that non-surgical embryo transfer alone can impact placental development. Furthermore, superovulation further induced overgrowth of the placental junctional zone. Embryo transfer and superovulation defects were limited to these morphological changes, as we did not observe any differences in epigenetic profiles. IVF placentae, however, displayed hypomethylation of imprinting control regions of select imprinted genes and a global reduction in DNA methylation levels. Although we did not detect significant differences in DNA methylation in fetal brain or liver samples, rare IVF concepti displayed very low methylation and abnormal gene expression from the normally repressed allele. Our findings suggest that individual ART procedures cumulatively increase placental morphological abnormalities and epigenetic perturbations, potentially causing adverse neonatal and long-term health outcomes in offspring.

Morphologic and molecular changes in the placenta: what we can learn from environmental exposures

“…Intriguingly, the procedure of transferring blastocyst stage embryos alone can increase overall placental weight, strongly sμggesting that relatively brief procedures can contribute to placental phenotypes (14). In addition to overall increased placental weight, mouse IVF placentas also exhibit disproportionate overgrowth of the junctional zone, a tissue that secretes factors that may influence the growth of the placental vasculature (12–16).…”

“…Importantly, increased placental weight with ART was almost always associated with a significant reduction in fetal weight (11–15). In some instances, fetal weight was unchanged, which may be influenced by the genetic background of the embryo (14,15). Althoμgh reduced fetal weight may be indicative of reduced placental efficiency, it is not clear if reduced fetal weight is due to impaired placental function or if ART procedures also directly impact fetal growth processes.…”

“…Imprinted genes, which are genes that display monoallelic expression dependent on the parent-of-origin, are regulated by DNA methylation at discrete cis -acting elements known as imprinting control regions (ICRs). The placenta, relative to the embryo, is particularly sensitive to these epigenetic changes (14,25–28). In mid-gestation and term mouse placenta, ART procedures result in ICR hypomethylation and biallelic expression of many disease-relevant imprinted genes (14,25–28).…”

“…The placenta, relative to the embryo, is particularly sensitive to these epigenetic changes (14,25–28). In mid-gestation and term mouse placenta, ART procedures result in ICR hypomethylation and biallelic expression of many disease-relevant imprinted genes (14,25–28). In a different study, the relative expression of imprinted genes was significantly increased in mid-gestation placenta from ART and embryo culture groups by comparison with controls althoμgh no differences in ICR methylation were observed (29).…”

“…A global reduction in DNA methylation was detected in ART term mouse placentas using luminometric methylation assay (LUMA), an assay that measures the relative amount of global CpG methylation using methylation sensitive and methylation insensitive restriction enzymes (14). A global reduction in DNA methylation was also detected in ART extraembryonic tissues at E7.5 using methylated DNA immunoprecipitation (MeDIP) (12).…”