Morphokinetic changes and apoptotic cell death in vitrified and non-vitrified in vitro-produced ovine embryos.

This article addresses morphokinetic changes and the extent of apoptosis in vitrified and non-vitrified in vitro-derived ovine blastocysts. Cumulus-oocyte complexes were collected after ovarian scarification obtain after slaughter and in vitro maturation was performed in TCM 199 medium supplemented with Earle's Salt, 10 % of FBS, and 5 µg/mL of LH/FSH at 38 °C for 24 h. After maturation, the oocytes were co-incubated with thawed ram semen (IVF) for 19 h.Embryo development was monitored with the aid of the Primo Vision Time-Lapse (TL) system. Twenty-five out of thirty-one ovine blastocysts that were vitrified using the Cryotop system at the early blastulation stage of development subsequently re-expanded. Both the vitrified (n = 25) and non-vitrified (control group: n = 28) blastocysts were examined for detection of apoptosis (TUNEL assay) and total blastomere counts at the time they attained the expanded blastocyst stage. Blastocyst formation occurred earlier in non-vitrified than in vitrified ovine embryos (147:49 ± 20:23 compared with 156:46 ± 19:24; hours:minutes post-insemination; mean ± SD; P < 0.05). The average number of blastocyst collapses was greater (2.45 ± 1.64 compared with 1.45 ± 1.64), but the number of weak contractions was less for vitrified than non-vitrified ovine blastocysts (P < 0.05). The mean number of blastomeres was greater (131.8 ± 38.6 compared with 91.5 ± 18.3; P < 0.05) while the number of TUNEL-positive cells (4.4 ± 1.6 compared with 6.3 ± 2.3) and apoptotic index (3.4 ± 1.2 % compared with 6.9 ± 2.6 %) were less (P < 0.05) in non-vitrified compared with vitrified blastocysts. Vitrification of ovine embryos was associated with a delayed blastocyst formation, greater numbers of apoptotic cells, significant reduction in the number of blastomeres, and higher/lower incidence of blastocyst collapse/weak contractions.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2023 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)