[en] The field of assisted reproduction has been developed to treat infertility in women, companion animals, and endangered species. In the horse, assisted reproduction also allows for the production of embryos from high performers without interrupting their sports career and contributes to an increase in the number of foals from mares of high genetic value. The present manuscript describes the procedures used for collecting immature and mature oocytes from horse ovaries using ovum pick-up (OPU). These oocytes were then used to investigate the incidence of aneuploidy by adapting a protocol previously developed in mice. Specifically, the chromosomes and the centromeres of metaphase II (MII) oocytes were fluorescently labeled and counted on sequential focal plans after confocal laser microscope scanning. This analysis revealed
a higher incidence in the aneuploidy rate when immature oocytes were collected from the follicles and matured in vitro compared to in vivo. Immunostaining for tubulin and the acetylated form of histone four at specific lysine residues also revealed differences in the morphology of the meiotic spindle and in the global pattern of histone acetylation. Finally, the expression of mRNAs coding for histone deacetylases (HDACs) and acetyl-transferases (HATs) was investigated by reverse transcription and quantitative-PCR (q-PCR). No differences in the relative expression of transcripts were observed between in vitro and in vivo matured oocytes. In agreement with a general silencing of the transcriptional activity during oocyte maturation, the analysis of the total transcript amount can only reveal mRNA stability or degradation. Therefore, these findings indicate that other translational and post-translational regulations might be affected.
Overall, the present study describes an experimental approach to morphologically and biochemically characterize the horse oocyte, a cell type that is extremely challenging to study due to low sample availability. However, it can expand our knowledge on the reproductive biology and infertility in monovulatory species.
Deleuze, Stefan ; Université de Liège > Dép. clinique des animaux de compagnie et des équidés (DCA) > Obstét. et path. de la reprod. des anim. de comp. et équidés
Dellenbach, P. et al. Transvaginal, sonographically controlled ovarian follicle puncture for egg retrieval. Lancet. 1(8392), 1467 (1984).
Humaidan, P. et al. Ovarian hyperstimulation syndrome: review and new classification criteria for reporting in clinical trials. Hum Reprod. (2016).
Pincus, G., & Enzmann, E. V. The Comparative Behavior of Mammalian Eggs in Vivo and in Vitro: I. The Activation of Ovarian Eggs. J Exp Med. 62(5), 665-675 (1935).
Emery, B. R., Wilcox, A. L., Aoki, V. W., Peterson, C. M., & Carrell, D. T. In vitro oocyte maturation and subsequent delayed fertilization is associated with increased embryo aneuploidy. Fertil Steril. 84(4), 1027-1029 (2005).
Nichols, S. M., Gierbolini, L., Gonzalez-Martinez, J. A., & Bavister, B. D. Effects of in vitro maturation and age on oocyte quality in the rhesus macaque Macaca mulatta. Fertil Steril. 93(5), 1591-1600 (2010).
Requena, A. et al. The impact of in-vitro maturation of oocytes on aneuploidy rate. Reprod Biomed Online. 18(6), 777-783 (2009).
Franciosi, F. et al. In vitro maturation affects chromosome segregation, spindle morphology and acetylation of lysine 16 on histone H4 in horse oocytes. Reprod Fertil Dev. (2015).
Franciosi, F. et al. Changes in histone H4 acetylation during in vivo versus in vitro maturation of equine oocytes. Mol Hum Reprod. 18(5), 243-252 (2012).
Choi, Y. H., Gibbons, J. R., Canesin, H. S., & Hinrichs, K. Effect of medium variations (zinc supplementation during oocyte maturation, perifertilization pH, and embryo culture protein source) on equine embryo development after intracytoplasmic sperm injection. Theriogenology. (2016).
Hendriks, W. K. et al. Maternal age and in vitro culture affect mitochondrial number and function in equine oocytes and embryos. Reprod Fertil Dev. 27(6), 957-968 (2015).
Carnevale, E. M. The mare model for follicular maturation and reproductive aging in the woman. Theriogenology. 69(1), 23-30 (2008).
Ginther, O. J. The mare: a 1000-pound guinea pig for study of the ovulatory follicular wave in women. Theriogenology. 77(5), 818-828 (2012).
Chiang, T., Duncan, F. E., Schindler, K., Schultz, R. M., & Lampson, M. A. Evidence that weakened centromere cohesion is a leading cause of age-related aneuploidy in oocytes. Curr Biol. 20(17), 1522-1528 (2010).
Duncan, F. E., Chiang, T., Schultz, R. M., & Lampson, M. A. Evidence that a defective spindle assembly checkpoint is not the primary cause of maternal age-associated aneuploidy in mouse eggs. Biol Reprod. 81(4), 768-776 (2009).
Larionov, A., Krause, A., & Miller, W. A standard curve based method for relative real time PCR data processing. BMC Bioinformatics. 6(62) (2005).
Choi, Y. H., Hochi, S., Braun, J., Sato, K., & Oguri, N. In vitro maturation of equine oocytes collected by follicle aspiration and by the slicing of ovaries. Theriogenology. 40(5), 959-966 (1993).
Hassold, T., & Hunt, P. To err (meiotically) is human: the genesis of human aneuploidy. Nat Rev Genet. 2(4), 280-291 (2001).
Akiyama, T., Nagata, M., & Aoki, F. Inadequate histone deacetylation during oocyte meiosis causes aneuploidy and embryo death in mice. Proc Natl Acad Sci U S A. 103(19), 7339-7344 (2006).
Homer, H. A. et al. Mad2 prevents aneuploidy and premature proteolysis of cyclin B and securin during meiosis I in mouse oocytes. Genes Dev. 19(2), 202-207 (2005).
Rambags, B. P. et al. Numerical chromosomal abnormalities in equine embryos produced in vivo and in vitro. Mol Reprod Dev. 72(1), 77-87 (2005).
Nabti, I., Marangos, P., Bormann, J., Kudo, N. R., & Carroll, J. Dual-mode regulation of the APC/C by CDK1 and MAPK controls meiosis I progression and fidelity. J Cell Biol. 204(6), 891-900 (2014).
Shomper, M., Lappa, C., & FitzHarris, G. Kinetochore microtubule establishment is defective in oocytes from aged mice. Cell Cycle. 13(7), 1171-1179 (2014).
Luzzo, K. M. et al. High fat diet induced developmental defects in the mouse: oocyte meiotic aneuploidy and fetal growth retardation/brain defects. PLoS One. 7(11), e49217 (2012).
Ma, P., & Schultz, R. M. Histone deacetylase 2 (HDAC2) regulates chromosome segregation and kinetochore function via H4K16 deacetylation during oocyte maturation in mouse. PLoS Genet. 9(3), e1003377 (2013).
Yang, F., Baumann, C., Viveiros, M. M., & De La Fuente, R. Histone hyperacetylation during meiosis interferes with large-scale chromatin remodeling, axial chromatid condensation and sister chromatid separation in the mammalian oocyte. Int J Dev Biol. 56(10-12), 889-899 (2012).
Luciano, A. M. et al. Oocytes isolated from dairy cows with reduced ovarian reserve have a high frequency of aneuploidy and alterations in the localization of progesterone receptor membrane component 1 and aurora kinase B. Biol Reprod. 88(3), 58 (2013).
Luciano, A. M., Lodde, V., Franciosi, F., Ceciliani, F., & Peluso, J. J. Progesterone receptor membrane component 1 expression and putative function in bovine oocyte maturation, fertilization, and early embryonic development. Reproduction. 140(5), 663-672 (2010).
Terzaghi, L. et al. PGRMC1 participates in late events of bovine granulosa cells mitosis and oocyte meiosis. Cell Cycle.1-14 (2016).
Susor, A., Jansova, D., Anger, M., & Kubelka, M. Translation in the mammalian oocyte in space and time. Cell Tissue Res. 363(1), 69-84 (2016).
Chen, J. et al. Genome-wide analysis of translation reveals a critical role for deleted in azoospermia-like (Dazl) at the oocyte-to-zygote transition. Genes Dev. 25(7), 755-766 (2011).
Ma, J., Flemr, M., Strnad, H., Svoboda, P., & Schultz, R. M. Maternally recruited DCP1A and DCP2 contribute to messenger RNA degradation during oocyte maturation and genome activation in mouse. Biol Reprod. 88(1), 11 (2013).
