Effect of the microenvironment and embryo density on developmental characteristics and gene expression profile of bovine preimplantative embryos cultured in vitro

Abstract

The Well of the Well (WOW) system has been developed to culture embryos in small groups or to track the development of single embryos. In the present study, we aimed to examine the effects of the microenvironment provided by the WOW system and embryo density on developmental rates, embryo quality and preimplantative gene expression profile of the resulting embryos. Embryos cultured in a group of 16 reached the blastocyst stage at a significantly lower level than zygotes cultured in a group of 50 (22.2 vs 30.3%), whereas zygotes cultured in WOW were able to compensate against low embryo densities, reaching a blastocyst rate as high as embryos cultured in a group of 50 (31.3 vs 30.3%). Moreover, embryos derived from WOW culture did not differ in terms of differential cell counts and apoptotic cell index compared with controls. The gene expression analysis revealed 62 transcripts to be upregulated and 33 transcripts to be downregulated by WOW culture. Comparing the in vivo derived blastocysts with the blastocysts derived from WOW culture, and group culture, expression of ATP5A1, PLAC8 and KRT8 was more similar to the embryos derived from WOW culture, whereas expression of S100A10 and ZP3 genes was more similar to blastocysts cultured in a group. In conclusion, microenvironment as well as embryo density significantly affected developmental rates. While subsequent blastocysts did not differ in terms of differential cell counts and apoptotic cell index, significant differences were observed in terms of the relative abundance of transcripts in the resulting embryos.

Effect of supplementation of different growth factors in embryo culture medium with a small number of bovine embryos on in vitro embryo development and quality

“…In bovine, in disagreement with our results, some authors observed that blastocysts obtained after culture individually contained a lower number of cells than embryos cultured in large groups (O'Doherty et al, 1997;Larson and Kubisch, 1999). However, other authors did not find a significant decrease on blastocyst cell number of the embryos cultured singly (Goovaerts et al, 2009 and or in reduced groups (Hoelker et al, 2009). However, in the experiments of Goovaerts et al (2009 and, embryos were cultured with a cumulus cell coculture.…”

“…Other studies on bovine embryos cultured singly or in reduced groups did not show significant differences on apoptotic rate per embryo (Hoelker et al, 2009;Goovaerts et al, 2011). In the first of them, it is possible that the number of embryos maintained in reduced culture was too high (16 embryos) for observing a significant effect in apoptosis, although a reduction in the number of in vitro cultured bovine embryos, from 50 to 16 embryos, significantly decreased the blastocyst rate (Hoelker et al, 2009). In the second one, singly cultured embryos were cultured in the presence of a cumulus cell monolayer (Goovaerts et al, 2011) and it was observed that oviductal cell coculture diminished the superoxide anion level in embryo CM (Joo et al, 2001).…”

“…Brison and Schultz (1997) observed that culturing in groups or adding 0.1 pM TGF-a to medium reduced the apoptosis rate, using terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labelling (TUNEL) technique, when murine embryos were cultured singly. Other studies on bovine embryos cultured singly or in reduced groups did not show significant differences on apoptotic rate per embryo (Hoelker et al, 2009;Goovaerts et al, 2011). In the first of them, it is possible that the number of embryos maintained in reduced culture was too high (16 embryos) for observing a significant effect in apoptosis, although a reduction in the number of in vitro cultured bovine embryos, from 50 to 16 embryos, significantly decreased the blastocyst rate (Hoelker et al, 2009).…”

Effect of number of oocytes and embryos on in vitro oocyte maturation, fertilization and embryo development in bovine

“…In the present work, significant differences were observed when 10 embryos in 100 µL (1/10 oocyte µL -1 ) were in vitro cultured. In this sense, Hoelker et al (2009) detected significant differences in embryo development with 16 embryos (1/32 oocyte µL -1 ) in comparison with the control group (n = 50, 1/10 oocyte µL -1 ). However, other authors (Fujita et al, 2006) detected differences only when a lower number of embryos (n = 3, 1/5 oocyte µL -1 ) were cultured.…”

Safety in Assisted Reproductive Technologies: Insights from Gene Expression Studies During Preimplantation Development

“…Several variables during in vitro culture were seen to affect the success rate and the quality of the developing embryos. Embryo density during in vitro, for instance, significantly affected the expression of stress-related genes (de Oliveira et al, 2005), the developmental competence to blastocyst stage, as well as the gene expression patterns on a large-scale (Hoelker et al, 2009).…”