methylation status of h19/igf2 differentially methylated region in in vitro human blastocysts donated by healthy couples

backgrund: imprinted genes are a unique subset of few genes, which have been differentially methylated region (dmr) in a parental origin-dependent manner during gametogenesis, and these genes are highly protected during pre-implantation epigenetic reprogramming. several studies heve shown that the particular vulnerability of imprinting genes during suboptimal pre- and peri-conception microenvironments often occur by assisted reproduction techniques (art). this study investigated the methylation status of h19/igf2 dmr at high-quality expanding/expanded human blastocysts donated by healthy individuals to evaluate the risks linked to art. method: methylation levels of h19/igf2 dmr were analyzed by bisulfite conversion and sequencing at 18 cpg sites (cpgs) located in this region. result: results showed that the overall percentage of methylated cpgs and the proportion of hyper-methylated clones of h19/igf2 dmr in analyzed blastocysts were 37.85±4.87% and 43.75±5.1%, respectively. for validation of our technique, the corresponding methylation levels of peripheral human lymphocytes were defined (49.52±1.86% and 50%, respectively). conclusion: considering the absence of in vivo produced human embryos, it is not possible to conclude that the methylation found in h19/igf2 dmr is actually normal or abnormal. regarding the possible risks associated with art, the procedures should be optimized in order to at least reduce some of the epigenetic risks.