108 Developmental potential and morphokinetics of in vitro-produced bovine embryos generated from oocytes of varying quality

Time-lapse incubation has been used in human and bovine embryos to select embryos for transfer. This system allows for continuous monitoring of embryos with fewer interruptions as well as analysis of morphokinetic parameters during development. Previous studies found that time intervals between divisions and reverse cleavage are related to the growth potential of bovine embryos. Additionally, while higher grade oocytes produce superior quality embryos and pregnancy rates, the differences in the morphokinetic parameters of embryos originating from oocytes of different grades has not been examined. In aim 1 of our study, we compared morphokinetic parameters, including timing of events, time between cell divisions, and incidence of reverse cleavage between embryos generated from different quality oocytes that developed to the morula stage or arrested earlier. In aim 2, we examined morphokinetic parameters of embryos that reach blastulation relative to embryos that did not. All embryos were generated from oocytes collected from abattoir-sourced ovaries, using a standard in vitro embryo production protocol. Oocytes were separated by grade (OG 1–4; IETS standards) before maturation. After fertilization, zygotes with two pronuclei were loaded into CultureCoins® and cultured in a MIRI® T6L incubator on Day 7. Timing of events from the second to eighth cell divisions (t2–t8) and morula and blastocyst formation were annotated manually for each embryo using the Miri-TL software. Reverse cleavage was also recorded. Aim 1 data were analyzed using a two-way ANOVA, and aim 2 data were analyzed using a Welch two-sample t-test. Reverse cleavage values were analyzed using Fisher’s exact test. In aim 1, the timing to the second, third, and fourth cell divisions were associated with oocyte grade and developmental outcome (P < 0.05). Timing between cell divisions (t3–t4, t2–t5, t2–t6) was also related to developmental outcome (P < 0.05). The timing between the second and fourth cell division varied by oocyte grade (P < 0.05). In all groups except embryos from grade 4 oocytes, a significantly higher number of embryos that arrested than embryos that developed to morula had at least one reverse cleavage event: OG1, 5/23 developed (dev) and 28/45 arrested (ar) (P = 0.002); OG2, 2/25 dev and 24/45 ar (P < 0.0001); OG3, 3/15 dev and 22/31 ar (P = 0.0016); and OG4, 5/15 dev and 12/23 ar (P = 0.32). In aim 2, arrested embryos took significantly longer to reach t2, t3, and t4 (28.9 ± 0.5 h, 33.7 ± 1 h, and 35 ± 0.9 h, repsectively) than embryos that developed to blastocyst (t2, 25 ± 0.6 h; t3, 30 ± 0.8 h, and t4, 31.5 ± 0.9 h). Reverse cleavage occurred more often in arrested embryos than in embryos that developed to blastocyst (dev, 6/24; ar, 87/139; P = 0.0007). We concluded that oocyte grade influences morphokinetic parameters of embryo development, including timing to and between cell divisions. We also found that embryos that reach the morula and blastocyst stage have different morphokinetic attributes than embryos that fail to develop. This finding may lead to improved selection of embryos for transfer.