362 DIFFERENT IN VITRO DEVELOPMENT AFTER AGGREGATION OF BOVINE AND FELINE PARTHENOGENETIC EMBRYOS

Abstract

Embryo aggregation has been shown to improve embryo development in several species. However, the effects seem to be different among species. Thus, the aim of this study was to compare the effect of embryo aggregation over in vitro development and blastocyst quality of bovine and feline parthenogenetic (PA) embryos. To this aim, bovine cumulus-oocyte complexes (COC) were collected from slaughterhouse ovaries, whereas cat ovaries were obtained from ovariectomized animals. The COC were in vitro matured in TCM199 supplemented following standard protocols for each species. After 24 h, cumulus cells and zona pellucidae were removed. Matured oocytes were selected and activated by 5 µM ionomycin treatment for 4 min followed by incubation in 1.9 mM 6-DMAP. Bovine and feline PA embryos were cultured in SOF medium in the well of well system in two different groups: only one PA embryo per microwell (1X); and three PA embryos per microwell (3X, aggregated embryos). Cleavage and blastocyst rates from all groups were assessed at Days 2 and 7, respectively. Size of blastocysts was measured at Day 7 using a millimetre eyepiece, and total cell number was determined by Hoechst 33342 staining. Blastocyst rates and embryo size were analysed by Fisher's test (P < 0.05) and total cell numbers by Kruskal–Wallis test with Dunn's correction (P < 0.05). Statistical differences were found in PA blastocyst rates between experimental groups (1X: 15/104, 24.6% v. 3X: 27/37, 62.2% for feline; and 1X: 21/113, 19.4% v. 3X: 20/32, 62.5% for bovine), but no differences were found between species. In addition, there was no statistical difference in the number of blastocysts obtained per oocyte used in any of the experimental groups. Bovine aggregated PA blastocysts were significantly larger than non-aggregated embryos (>200 microns, 1X: 2/20, 10% v. 3X: 9/19, 47.4%), but no differences were found in cell number. On the other hand, cat aggregated PA blastocysts had significantly higher cell numbers (1X: 122.4 ± 79.66 cells v. 3X: 259.8 ± 137.1 cells), but no differences were found in blastocyst size. This observation can contribute in the understanding of embryo physiology, suggesting that benefits of embryo aggregation in parthenogenic embryos vary among these species.