151 Multipolar zygotic divisions result in multinuclear and anuclear blastomeres in cattle

2021 ◽  
Vol 33 (2) ◽  
pp. 184
Author(s):  
T. De Coster ◽  
K. Smits ◽  
O. B. Pascottini ◽  
J. Vermeesch ◽  
A. Van Soom
Keyword(s):  
Time Lapse ◽  
Human Embryos ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Mixed Effect ◽  
Genetic Abnormalities ◽  
The Individual ◽  
Time Lapse Imaging ◽  
Nuclear Content

The mammalian zygotic cleavage is expected to result in two mononuclear blastomeres. However, zygotes undergoing multipolar divisions resulting in direct cleavage into three or four cells are frequently observed in bovine and human embryonic development and have been associated with decreased euploidy rates of resulting blastocysts and a lower pregnancy rate (Somfai et al. 2010 J. Reprod. Dev. 56, 200-207; https://doi.org/10.1262/jrd.09-097a; Zhan et al. 2016 PLoS ONE 11, 1-19; https://doi.org/10.1371/journal.pone.0166398; Sugimura et al. 2017 J. Reprod. Dev. 63, 353-357; https://doi.org/10.1262/jrd.2017-041). Therefore, multipolar zygotic divisions may underly genetic abnormalities by aberrant segregation of the chromosomal material resulting in multinucleated or anuclear blastomeres. These abnormal blastomeres have been observed in human cleavage-stage embryos (Nogueira et al. 2000 Fertil. Steril. 74, 295-298; https://doi.org/10.1016/s0015-0282(00)00642-7; Chatzimeletiou et al. 2006 Hum. Reprod. 20, 672–682; https://doi.org/10.1093/humrep/deh652), but the prevalence in bovine embryos and the direct association with the multipolar division in both bovine and human embryos remains unknown. We hypothesised that anuclear and multinuclear blastomeres also occur in bovine embryos, and we aimed to unravel the link between multipolar zygotic divisions and genome segregation errors by determining the nuclear blastomere content in a bovine model. Therefore, oocytes from 5 cows were matured and fertilized in vitro by the same bull according to our standard in vitro production procedure (Wydooghe et al. 2014 Reproduction 148, 519-529). The first cleavage was monitored the by time-lapse imaging. Forty-three blastomeres from 22 bipolar zygotic divisions, and 65 blastomeres from 20 multipolar zygotic divisions were collected immediately after the first cleavage, using pronase to isolate the individual blastomeres. The area of each blastomere was measured and the number of nuclei was determined after fixation and staining with Hoechst 33342. Generalized mixed effect models were built to identify the effect of the type of cleavage (bipolar vs. multipolar) on the number of nuclei (mononuclear vs. anuclear or multinuclear) in the blastomeres. Linear mixed models were built to determine the effect of the type of cleavage and the nuclear content on the size of the blastomeres. Embryos presented a greater number of blastomeres with a normal nuclear content (92.6 ± 0.4%) after a bipolar cleavage compared with multipolar division (73.2 ± 0.7%; P=0.03). Moreover, blastomeres presented a 28% larger blastomere area (P<0.001) after bipolar division compared with multipolar division. Notably, anuclear blastomeres tended to be smaller than multi- and mononuclear blastomeres (P=0.09 for both), while no difference was found between mono and multinucleated blastomeres (P=0.84). In conclusion, this is one of the first reports on the association between nuclear blastomere content in bovine embryos and the dynamics of the first zygotic division. Even though sample size was limited, these results confirm the hypothesised link between multipolar division and abnormal genome segregation as determined by multinuclear and anuclear blastomeres in the resulting blastomeres. Therefore, multipolar cell divisions at the zygotic division may underly at least some of the genetic abnormalities observed in embryos at early development.

288 KINETICS OF FERTILIZATION, DEVELOPMENT, AND SEX RATIO OF IN VITRO-PRODUCED BOVINE EMBRYOS OBTAINED WITH FOUR DIFFERENT BULLS

2007 ◽  
Vol 19 (1) ◽  
pp. 259 ◽  
Author(s):  
M. Alomar ◽  
H. Tasiaux ◽  
S. Remacle ◽  
F. George ◽  
D. Paul ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Time Lapse ◽  
Blastocyst Stage ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Chi Square ◽  
Cell Stage ◽  
Chi Square Test ◽  
Kinetics Of

The between-bulls variation in in vitro fertility and the shift of sex ratio toward male embryos are two problems affecting the in vitro production (IVP) of bovine embryos. Our objective was to evaluate the possible correlation between the kinetics of fertilization, embryo development, and the sex ratio of the resulting embryos. In a first experiment, and using frozen-thawed semen of 4 different AI bulls, the kinetics of pronucleus (PN) formation was evaluated at 8, 12, and 18 h post-in vitro insemination (hpi) after fixation and staining with Hoechst 33342. Fertilized oocytes were classified in 3 PN stages: PN1: showing the first signs of sperm head decondensation; PN2: with two pronuclei of different sizes, the two being far from each other; and PN3: showing two symmetric pronuclei of equal size, close to each other. Differences between bulls were observed at each time point, but were greater at 12 hpi than at 8 or 18 hpi. At 8 hpi and 12 hpi, bull C showed a significantly faster PN formation by comparison with the 3 other bulls (chi-square test: P < 0.05), whereas at 18 hpi, the proportion at each of the PN stages was similar to that of bulls A and D, with bull B showing delayed PN development. In a second experiment, a standard IVP procedure was conducted with the 4 bulls to determine cleavage and blastocyst rates. The timing of first cleavage was measured using time-lapse cinematography. Compared with those of bull B, the embryos generated with bull C led to significantly higher Day 7 blastocyst yields (31.3 � 9.5% vs. 21.9 � 6.7%; ANOVA: P < 0.05). Moreover, the embryos from bull C reaching the blastocyst stage cleaved faster (first cleavage at 23.1 � 2.1 hpi vs. 25.4 � 2.7 hpi for bull B; ANOVA: P < 0.05). In a third experiment, 65 to 76 Day 8 blastocysts were sexed per bull. Embryo sexing was performed by PCR using the co-amplification of a Y-specific bovine SRY sequence and an autosomal btRep-137 sequence. Only blastocysts obtained with bull C showed a shift in sex ratio toward male embryos (76.0% male embryos vs. 53.8% for bull B; chi-square test: P < 0.05), whatever the size of the blastocyst. The shift in sex ratio was already present at the 2-cell stage (64.2% male embryos; n = 53; chi-square test: P < 0.05). In conclusion, for 2 out of 4 bulls, a correlation was observed between the kinetics of PN formation, the timing of first cleavage, and the sex ratio of the resulting embryos.

Cleavage of Human Embryos: Options and Diversity

Acta Naturae ◽  
2016 ◽  
Vol 8 (3) ◽  
pp. 88-96
Author(s):  
Yu. K. Doronin ◽  
I. V. Senechkin ◽  
L. V. Hilkevich ◽  
M. A. Kurcer
Keyword(s):  
Time Lapse ◽  
Reproductive Technologies ◽  
Human Embryos ◽  
Imaging Data ◽  
Noninvasive Methods ◽  
Topographic Features ◽  
Time Parameters ◽  
Embryo Cleavage ◽  
Time Lapse Imaging ◽  
Developmental Dynamics

In order to estimate the diversity of embryo cleavage relatives to embryo progress (blastocyst formation), time-lapse imaging data of preimplantation human embryo development were used. This retrospective study is focused on the topographic features and time parameters of the cleavages, with particular emphasis on the lengths of cleavage cycles and the genealogy of blastomeres in 2- to 8-cell human embryos. We have found that all 4-cell human embryos have four developmental variants that are based on the sequence of appearance and orientation of cleavage planes during embryo cleavage from 2 to 4 blastomeres. Each variant of cleavage shows a strong correlation with further developmental dynamics of the embryos (different cleavage cycle characteristics as well as lengths of blastomere cycles). An analysis of the sequence of human blastomere divisions allowed us to postulate that the effects of zygotic determinants are eliminated as a result of cleavage, and that, thereafter, blastomeres acquire the ability of own syntheses, regulation, polarization, formation of functional contacts, and, finally, of specific differentiation. This data on the early development of human embryos obtained using noninvasive methods complements and extend our understanding of the embryogenesis of eutherian mammals and may be applied in the practice of reproductive technologies.

Noninvasive characterization of metabolites secreted in culture media by bovine embryos during in vitro production

Metabolomics ◽  
2016 ◽  
Vol 12 (5) ◽  
Author(s):  
Érika Cristina dos Santos ◽  
Camila Bruna de Lima ◽  
Kelly Annes ◽  
Marcella Pecora Milazzotto
Keyword(s):  
Culture Media ◽  
Bovine Embryos ◽  
In Vitro Production ◽  
Noninvasive Characterization ◽  
Vitro Production

Animal factors that influence the in-vitro production of bovine embryos

1999 ◽  
Vol 51 (1) ◽  
pp. 334 ◽  
Author(s):  
H Twagiramungu ◽  
N Morin ◽  
C Brisson ◽  
G Carbonneau ◽  
J Durocher ◽  
...  
Keyword(s):  
Bovine Embryos ◽  
In Vitro Production ◽  
Vitro Production

scholarly journals Live-cell time-lapse imaging and single-cell tracking of in vitro cultured neural stem cells – Tools for analyzing dynamics of cell cycle, migration, and lineage selection

Methods ◽  
2018 ◽  
Vol 133 ◽  
pp. 81-90 ◽  
Author(s):  
Katja M. Piltti ◽  
Brian J. Cummings ◽  
Krystal Carta ◽  
Ayla Manughian-Peter ◽  
Colleen L. Worne ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Neural Stem Cells ◽  
Single Cell ◽  
Cell Tracking ◽  
Time Lapse ◽  
Live Cell ◽  
Time Lapse Imaging

scholarly journals Resveratrol-supplemented holding or re-culture media improves viability of fresh or vitrified-warmed in vitro-derived bovine embryos

2021 ◽  
Vol 10 (14) ◽  
pp. e367101422097
Author(s):  
Arianny Rafaela Neto Silva ◽  
Thaisa Campos Marques ◽  
Elisa Caroline Silva Santos ◽  
Tiago Omar Diesel ◽  
Isabelle Matos Macedo ◽  
...  
Keyword(s):  
Culture Media ◽  
Cell Number ◽  
Time Dependent ◽  
Expansion Rate ◽  
Ros Generation ◽  
Hatching Rate ◽  
Protective Effects ◽  
Bovine Embryos ◽  
In Vitro Production

The effect of resveratrol supplementation on fresh (E1) or vitrified/warmed (E2) in vitro produced bovine embryos was investigated by evaluating the time-dependent response. After in vitro production, resveratrol (0.5 µM) was added to the incubation media and after two incubation periods with or without resveratrol, blastocysts were re-cultured for 24h. The rates of re-expansion, hatching, total cell number (TCN), apoptotic cells (ACN), reactive oxygen species (ROS) and intracellular glutathione (GSH) content were evaluated. For E1, the re-expansion rate differed at 6 and 10h within and between treatments (P<0.05), as did the re-expansion rate after 24h (P<0.01). The hatching rate increased after 10h with resveratrol (P<0.01) with differences within (P<0.05), but not between treatments after 24h of re-cultivation. At E2, hatching rate differed between treatments at 24h (P<0.01), with higher TCN in resveratrol-treated blastocysts after 10h (P<0.01). Resveratrol supplementation reduced ROS generation in E1 and E2 after 10h of incubation and increased GSH content (P<0.01). These results indicate that supplementation of holding re-cultivation medium with resveratrol for treatment of fresh or vitrified/warmed in vitro produced bovine embryos has a positive and time-dependent effect. The reduction of ROS content, the increase of GSH and the anti-apoptotic ability of resveratrol are responsible for its protective effects, allowing an extension of embryo storage time before transfer to recipients.

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