The Effect of Vitrification on Expression and Histone Marks of Igf2 and Oct4 in Blastocysts Cultured from Two-Cell Mouse Embryos

OBJECTIVES Vitrification is increasingly used in assisted reproductive technology (ART) laboratories worldwide. In this study the effect of vitrification on the expression and modifications of H3 histones of Igf2 and Oct4 was investigated in blastocysts cultured from vitrified and non-vitrified two-cell embryos. MATERIALS AND METHODS In this experimental study, two-cell embryos were cultured in KSOM medium to reach the blastocyst stage. Expression of Igf2 and Oct4 and modifications of H3 histones in regulatory regions of both genes were compared with in vivo blastocysts, which comprise the control group. To gene expression evaluation, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and the ChIP assay method were carried out to assess expression and histone modifications of the two genes. RESULTS The expression level of Igf2 was significantly higher in both experimental groups than the control group. In the regulatory region of Igf2, H3K9 methylation decreased whereas H3K9 acetylation increased in the experimental group compared with the control group. In contrast, the expression level of Oct4 was significantly lower in experimental groups. The Oct4 gene promoter showed a significant increase in H3K9 methylation and decrease in H3K9 acetylation (P<0.05). CONCLUSIONS According to our results, both vitrification and cultivation conditions may lead to changes in expression level and modification of histones in Igf2 and Oct4. However, these effects were the same in vitrified and non-vitrified groups. Indeed, the embryo is most affected by culture environment and in vitro culture. Therefore, vitrification may be used as a low-risk technique for embryo cryopreservation in ART.