DNA methylation changes at infertility genes in newborn twins conceived by in vitro fertilisation

Background: The association of in vitro fertilisation (IVF) and DNA methylation has been studied predominantly at regulatory regions of imprinted genes and at just thousands of the similar to 28 million CpG sites in the human genome. Methods: We investigated the links between IVF and DNA methylation patterns in whole cord blood cells (n = 98) and cord blood mononuclear cells (n = 82) from newborn twins using genome-wide methylated DNA immunoprecipitation coupled with deep sequencing. Results: At a false discovery rate (FDR) of 5%, we identified one significant whole blood DNA methylation change linked to conception via IVF, which was located similar to 3 kb upstream of TNP1, a gene previously linked to male infertility. The 46 most strongly associated signals ( FDR of 25%) included a second region in a gene also previously linked to infertility, C9orf3, suggesting that our findings may in part capture the effect of parental subfertility. Using twin modelling, we observed that individual-specific environmental factors appear to be the main overall contributors of methylation variability at the FDR 25% IVF-associated differentially methylated regions, although evidence for methylation heritability was also obtained at several of these regions. We replicated previous findings of differential methylation associated with IVF at the H19/IGF2 region in cord blood mononuclear cells, and we validated the signal at C9orf3 in monozygotic twins. We also explored the impact of intracytoplasmic sperm injection on the FDR 25% signals for potential effects specific to male or female infertility factors. Conclusions: To our knowledge, this is the most comprehensive study of DNA methylation profiles at birth and IVF conception to date, and our results show evidence for epigenetic modifications that may in part reflect parental subfertility.