Insights Into The Molecular Mechanisms Underlying The Implantation Failure Following Ovarian Stimulation In Mice

Several studies have revealed the link between ovarian stimulation and implantation failure. Despite extensive research on this issue in the last few years, the underlying molecular mechanisms are still largely unknown. The objectives of the present study are to identify the molecular mechanisms by which ovarian stimulation affects uterine receptivity by using two doses of gonadotropins 10 iu and 5 iu during window of implantation in mice(3.5 dpc) at both m RNA expression and promoter DNA methylation levels as well as to examine the key histological features of the endometrium. Affymetrix Gene Chip? Mouse Genome Array was used for the global m RNA expression analysis and Affymetrix Mouse Promoter 1.0 R Tiling Arrays was used for mapping of promoter DNA methylation profiles. Results of gene chips were validated by q RT-PCR for m RNA expression levels and by pyrosequencing analysis for promoter DNA methylation levels.The results show that there are a large number of differentially regulated genes as well as promoter DNA methylation levels between both stimulated groups and unstimulated controls. This large degree of gene disturbance highlights the need for further rethinking about ovarian stimulation in order to optimize the procedures. Several pathways which are important for implantation were found to be altered in the uteri of stimulated groups, such as hedgehog signaling pathway, arachidonic acid metabolism, complement and coagulation cascades and focal adhesion. Ovarian stimulation provides some stress factor to the uterine environment, manifested by higher cellular and metabolic activities in both stimulated groups. High estrogen levels induce higher inflammatory response by genetic and epigenetic modifications of uterine genes which ultimately lead to apoptosis and predisposing endometrium to the cancer related inflammation. Moreover, ovarian stimulation alters prostaglandin production through genetic and epigenetic modifications of endometrial genes, suggesting that inappropriate prostaglandin production could be a possible cause for implantation failure following ovarian stimulation.Endometrial histology was also dramatically altered upon ovarian stimulation which renders the endometrium less or non-receptive for embryo implantation. Both mild and high doses of gonadotropins have deleterious effects on uterine receptivity with more negative impact being induced by high doses. In conclusion, high estrogen level resulting from ovarian stimulation is the main reason for unsuccessful implantation in stimulated cycles. In addition, the obtained data do not support fresh embryo transfer technology and more importantly raise the concerns about the safety of ovarian stimulation protocols due to an increased incidence of endometrial cancer.