77 OVIDUCT-EMBRYO INTERACTIONS: TWO-WAY TRAFFIC OR A ONE-WAY STREET? TRANSCRIPTOMIC RESPONSE OF THE BOVINE OVIDUCT TO THE PRESENCE OF AN EMBRYO

Abstract

Despite clear evidence of a two-way interaction between the developing conceptus and the uterine endometrium in early pregnancy, the evidence for reciprocal cross-talk during the transit of the embryo through the oviduct is less clear. The aims were (1) to characterise the transcriptome of the bovine oviduct at the initiation of embryonic genome activation (EGA), (2) to examine the effect, if any, of the presence of an embryo on the oviduct transcriptome, and (3) to compare gene expression in the ampulla and isthmus of the oviduct ipsilateral to the corpus luteum. The oestrous cycles of cross-bred beef heifers were synchronized and those recorded in standing oestrus were randomly allocated to control group, nonbred (n = 7), or AI group (n = 11). All heifers were slaughtered on Day 3 after oestrus. The oviducts from each animal were isolated, straightened, and cut in half (ampulla and isthmus). Each portion was flushed with 500 μL of PBS to confirm the presence of an oocyte/embryo and was then opened and scraped longitudinally to obtain epithelial cells. Cells were snap-frozen in liquid nitrogen for microarray analysis. All recovered oocytes and embryos were located in the isthmus of the oviduct ipsilateral to the corpus luteum. The recovery rate was 72.7% (8/11) and 83.3% (5/6) for pregnant and cyclic animals, respectively. The stage of the recovered embryos was as follows: 4-cell stage (n = 1), 8-cell stage (n = 5), and 8–16 cell stage (n = 2), whereas in the cyclic group all recovered structures were unfertilized oocytes. The cells of the isthmus from ipsilateral and contralateral oviducts from 5 cyclic and 5 pregnant animals (8-cell embryos) and the ipsilateral ampulla cells from the pregnant animals were used for microarray analysis (Affymetrix Bovine ST array, Affymetrix, Santa Clara, CA, USA). Array data were analysed using BioConductor packages in R and custom CDF files downloaded from MBNI. Preprocessing of raw data was performed with RMA, and differential expression was assessed by linear modelling implemented in the limma package. Genes displaying P < 0.05 after adjustment for multiple testing were considered differentially expressed. A total of 18 809 probe sets were assessed for differential expression. Comparison of pregnant and cyclic oviduct epithelium revealed no significantly altered genes. However, comparison of the isthmus and ampulla of the ipsilateral oviduct in pregnant animals revealed 4011 (P < 0.05) and 2327 (P < 0.01) differentially expressed genes. Some of the gene ontologies involved in biological processes included fatty acid metabolism, cell adhesion, cell morphogenesis, cellular developmental process, and reproduction. In conclusion, we have characterised the transcriptome of the bovine oviduct epithelium at the initiation of embryonic genome activation on Day 3 post-oestrus in pregnant and cyclic heifers. Although large differences in gene expression were observed between the isthmus and ampulla, data suggest that the presence of an 8-cell embryo had no effect on the transcriptome of the cells of the isthmus, although a local effect at the precise position of the embryo cannot be ruled out.