147 A PROSTAGLANDIN F_2α RECEPTOR ANTAGONIST MAY INHIBIT BAX EXPRESSION IN BOVINE PRE-IMPLANTATION EMBRYOS DURING EMBRYO FLUSHING

Abstract

Several in vivo studies have shown negative effects of prostaglandin F_2α (PGF_2α) on embryonic survival in cows. The PGF_2α receptor (FPr) was detected in bovine embryos (Scenna FN et al. 2006 Reprod. Fertil. Dev. 18, 180). Recently, it was reported that addition of a FPr antagonist (AL-8810) to the embryo flushing medium prevented the detrimental action of PGF_2α on pregnancy (Scenna FN et al. 2008 Reprod. Fertil. Dev. 20, 154). In the present study, we examined the morphological development in utero and gene expression of bovine pre-implantation embryos collected using flushing medium containing AL-8810. Embryos were collected from superovulated donors on Day 7 after artificial insemination with medium containing 1000 nm AL-8810 (n = 4, Sigma-Aldrich, St, Louis, MO, AL embryos) or with vehicle (n = 4, DMSO, Sigma-Aldrich, CON embryos). Following collection, the embryos were classified by quality and stage, and then 2 to 5 grade-1, stage 4 and 5 embryos (IETS classification) were transferred to recipient heifers (AL-embryos: n = 4, CON embryos: n = 4) for transient development in utero up to Day 15 of gestation. Embryos at Day 15 occasionally collapsed after uterine flushing due to breaches in trophectoderm (TE) tissue caused by the collection procedures. Only whole embryos were used and evaluated in this study. Using real-time PCR, the TE cells were examined to quantify the mRNA levels for 3 genes, interferon-τ (maternal recognition of pregnancy, INF-τ), connexin 43 (cell communication, CX 43) and Bcl-2-associated X protein (apoptosis, Bax). Data were analyzed using Student’s t-test and chi-square test. The average number of ova/embryos recovered at Day 7 was 12.0 ± 4.1 (AL embryos) and 8.8 ± 5.8 (CON embryos). There was no significance in the percentages of transferable embryos (AL-embryos; 67.0 ± 14.4%, 35/48 and CON-embryos; 79.6 ± 7.1%, 27/35; P > 0.05) between the 2 groups. The whole-embryo recovery rate on Day 15 for AL embryos (73.3%, 11/15) was >30 percentage points greater than that for CON embryos (41.2%, 7/17). The length of major axis of embryos and the diameter of embryonic disks were the same between AL embryos (59.6 ± 12.3 mm, 0.44 ± 0.03 mm, respectively) and CON embryos (49.3 ± 24.6 mm, 0.43 ± 0.05 mm, respectively). Transcripts of INF-τ and CX 43 in TE cells did not differ significantly between AL embryos and CON embryos (P > 0.05). However, the abundance of Bax transcripts in TE cells tended to be lower in AL embryos than in CON embryos (P = 0.07). These results suggest that addition of an FPr antagonist to the embryo flushing medium may promote the developmental capacity of pre-implantation embryos in utero.