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Epidemiological studies have reported a higher incidence of rare disorders involving imprinted genes among children conceived using assisted reproductive technology (ART), suggesting that ART procedures may be disruptive to imprinted gene methylation patterns. We examined intra- and inter-individual variation in DNA methylation at the differentially methylated regions (DMRs) of the  IGF2/H19  and  IGF2R  loci in a population of children conceived  in vitro  or  in vivo . We found substantial variation in allele-specific methylation at both loci in both groups. Aberrant methylation of the maternal  IGF2 / H19  DMR was more common in the  in vitro  group, and the overall variance was also significantly greater in the  in vitro  group. We estimated the number of trophoblast stem cells in each group based on approximation of the variance of the binomial distribution of  IGF2 / H19  methylation ratios, as well as the distribution of X chromosome inactivation scores in placenta. Both of these independent measures indicated that placentas of the  in vitro  group were derived from fewer stem cells than the  in vivo  conceived group. Both IGF2 and H19 mRNAs were significantly lower in placenta from the  in vitro  group. Although average birth weight was lower in the  in vitro  group, we found no correlation between birth weight and IGF2 or IGF2R transcript levels or the ratio of IGF2/IGF2R transcript levels. Our results show that  in vitro  conception is associated with aberrant methylation patterns at the  IGF2 / H19  locus. However, very little of the inter- or intra-individual variation in H19 or IGF2 mRNA levels can be explained by differences in maternal DMR DNA methylation, in contrast to the expectations of current transcriptional imprinting models. Extraembryonic tissues of embryos cultured  in vitro  appear to be derived from fewer trophoblast stem cells. It is possible that this developmental difference has an effect on placental and fetal growth.

plos genetics

Inter- and Intra-Individual Variation in Allele-Specific DNA Methylation and Gene Expression in Children Conceived using Assisted Reproductive Technology