Bovine in vitro oocyte maturation and embryo culture in liquid marbles 3D culture system

Abstract

Despite the advances in in vitro embryo production (IVP) over the years, the technique still has limitations that need to be overcome. In cell cultures, it is already well established that three-dimensional culture techniques are more physiological and similar to the in vivo development. Liquid marble (LM) is a three-dimensional system based on the use of a hydrophobic substance to create in vitro microbioreactors. Thus, we hypothesized that the LM system improves bovine in vitro oocyte maturation and embryo culture. In experiment I, bovine cumulus-oocyte complexes (COCs) were placed for in vitro maturation for 22h in two different groups: control (conventional 2D culture) and LM (three-dimensional culture). We found that oocyte nuclear maturation was not altered by the LM system, however it was observed a decrease in expression of genes important in the oocyte maturation process in cumulus cells of LM group (BCL2, EIF4E, and GAPDH). In experiment II, the COCs were conventionally matured and fertilized, and for culture, they were divided into LM or control groups. There was a decrease in blastocyst rate and cell counting, a down-regulation of miR-615 expression, and an increase in the DNA global methylation and hydroxymethylation in embryos of LM group. Therefore, for the bovine in vitro embryo production, this specific three-dimensional system did not present the advantages that we expected, but demonstrated that the embryos changed their development and epigenetics according to the culture system.

Heat shock interferes with the amino acid metabolism of bovine cumulus-oocyte complexes in vitro: a multistep analysis

“…To examine the morphology of cumulus expansion both before and after maturation, oocytes were put under an inverted microscope (Nikon Eclipse Ti). Then, using Image J software (Version 1.46; National Institutes of Health, Bethesda, MD, USA), the cumulus area from 60 COCs in each treatment group was calculated and normalized to the control group (Ferronato et al 2023 ). To assess the cumulus expansion, 180 COCs (60 COCs per treatment) were used over the course of 3 separate days, with 2 replicates of 10 COCs each.…”

3D culture applied to reproduction in females: possibilities and perspectives

“…Thus, this type of system may improve de diffusion of the medium for COC in vitro maturation increasing the blastocysts rates. However, recently the LM system was used during oocyte maturation and embryo culture ( Ferronato et al, 2023 ). The LM system induced a decrease in transcripts related to oocyte maturation process in cumulus cells ( Ferronato et al, 2023 ).…”

“…However, recently the LM system was used during oocyte maturation and embryo culture ( Ferronato et al, 2023 ). The LM system induced a decrease in transcripts related to oocyte maturation process in cumulus cells ( Ferronato et al, 2023 ). Importantly, the LM system applied during IVC induced a decrease in blastocyst rate and total cell number while an increase in global DNA methylation and hydromethylation levels ( Ferronato et al, 2023 ).…”

“…The LM system induced a decrease in transcripts related to oocyte maturation process in cumulus cells ( Ferronato et al, 2023 ). Importantly, the LM system applied during IVC induced a decrease in blastocyst rate and total cell number while an increase in global DNA methylation and hydromethylation levels ( Ferronato et al, 2023 ). Although the results in bovine were not improved, the LM system demonstrated to induce epigenetic changes, which if combined with other changes in media and gas pressure could lead to better results.…”

Application of Enzyme-Linked Fluorescence Assay (ELFA) to Obtain In Vivo Matured Dog Oocytes through the Assessment of Progesterone Level

“…The collected oocytes require maturation from the moment of collection as in vitro maturation techniques for immature dog oocytes are not yet established [6]. Recent reports on dog oocytes indicate a relatively low rate of in vitro maturation at about 30% compared to approximately 80% in mice and 70% in both sows and cows [7][8][9][10]. One of the reasons for the low in vitro oocyte maturation rate in dogs is a specific physiologic trait in dog reproduction, in which dog oocyte maturation is completed in the oviduct after ovulation, whereas most animals ovulate matured or nearly matured oocytes [11].…”