114 ASYNCHRONOUS EMBRYO TRANSFER AFFECTS THE EXPRESSION OF IMPRINTED GENES IN EQUINE TROPHECTODERM

Abstract

Gene imprinting is a form of epigenetic modification that results in parent-of-origin specific monoallelic expression. Imprinted genes play important roles during fetal-placental growth with paternally imprinted genes generally promoting and maternally imprinted genes suppressing fetal growth. Imprinted genes are therefore believed to have important effects on trophoblast differentiation and placental development, and in adjusting fetal nutrition to maternal supply. The horse is an interesting model of early placental development because of its unusually long pre-implantation period (40 days), during which the conceptus is dependent on uterine secretions for nutrient provision. Moreover, horse embryos tolerate a wide range of uterine asynchrony following embryo transfer (ET), offering a unique tool to study maternal influences on conceptus development. This study examined the effect of asynchronous ET on the expression of imprinted genes in equine trophectoderm. Twenty Day 8 embryos were transferred to recipient mares that either ovulated on the same day (synchronous; n = 10) or 5 days after (asynchronous; n = 10) the donor mare. The conceptuses were recovered 6 or 11 days after ET (Day 14 or 19 of conceptus development; n = 5 per group). Bilaminar trophectoderm was isolated and mRNA expression for a range of genes known to be imprinted in equine trophectoderm (H19, PHLDA2, IGF2R, IGF2, PEG3, PEG10, SNRPN, INSR, and INS) was investigated by RT-qPCR. The effects of asynchronous ET and stage of pregnancy on gene expression were analysed by two-way ANOVA followed by independent-samples t-tests. IGF2, PEG3, PEG10, INSR, H19, and PHLDA2 all showed a significant up-regulation in gene expression between Days 14 and 19 of pregnancy; however, expression was higher in synchronous than asynchronous pregnancies at Day 19 (P < 0.05). IGF2R expression increased significantly from Day 14 to 19 in the synchronous pregnancies (P < 0.05), but did not differ between treatments at Day 19. SNRPN expression increased from Day 14 to 19, and was unaffected by asynchrony. INS mRNA was not detectable in trophectoderm. In conclusion, asynchronous ET had a significant effect on gene expression at Day 19 of gestation that was not evident at Day 14. This may be either a contributor to the delayed development that is observed in asynchronous pregnancies or a result/response; in either case, it may affect subsequent development.

This study was founded by EpiHealthNet (Project number 317146).