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Energy metabolism disorders during in vitro maturation of bovine cumulus-oocyte complexes interfere with blastocyst quality and metabolism
Follicular-fluid extracellular vesicles support energy metabolism of bovine oocytes, improving blastocyst development and quality
Abstract Extracellular vesicles (EVs) from follicular fluid (FF) seem to play a significant role in communication within ovarian follicles in several species. The present study aimed to examine the supporting effect of FF-derived small EVs (FF-sEVs) during in vitro maturation (IVM) of bovine cumulus–oocyte complexes (COCs) under conditions of disturbed energy metabolism. Bovine COCs were matured in vitro with inhibitors targeting lipid metabolism (etomoxir) or glucose metabolism (iodoacetate combined with dehydroepiandrosterone), in the presence or absence of FF-sEVs. Following maturation, oocytes and cumulus cells were analyzed by real-time quantitative polymerase chain reaction (qPCR) and stained to visualize lipid droplets. The uptake of FF-sEVs was visualized by fluorescent labeling. In vitro fertilization and embryo culture were followed by mass spectrometry analysis of hatched blastocysts. We demonstrate for the first time that FF-sEVs are transported from the medium into the oocytes, via the cumulus cells and through transzonal projections into the perivitelline space and ooplasm. Cumulus cells under metabolic stress conditions exhibit an increased FF-sEV uptake from the maturation medium. FF-sEV supplementation during metabolic stress conditions enhances the MII rate in oocytes and positively affects subsequent embryo development and quality revealed by altered metabolic activity. Lipid droplet parameters and gene expression in cumulus cells and oocytes are affected by FF-sEV supplementation, which is more pronounced in cumulus cells. Our findings show that FF-sEV supplementation during IVM under metabolic stress conditions significantly affects COCs, with a positive effect on further blastocyst quality. We provide novel insights into the role of FF-sEVs in oocyte maturation and blastocyst development.
Species and embryo genome origin affect lipid droplets in preimplantation embryos
Mammalian embryo development is affected by multiple metabolism processes, among which energy metabolism seems to be crucial. Therefore the ability and the scale of lipids storage in different preimplantation stages might affect embryos quality. The aim of the present studies was to show a complex characterization of lipid droplets (LD) during subsequent embryo developmental stages. It was performed on two species (bovine and porcine) as well as on embryos with different embryo origin [after in vitro fertilization (IVF) and after parthenogenetic activation (PA)]. Embryos after IVF/PA were collected at precise time points of development at the following stages: zygote, 2-cell, 4-cell, 8/16-cell, morula, early blastocyst, expanded blastocyst. LD were stained with BODIPY 493/503 dye, embryos were visualized under a confocal microscope and images were analyzed with the ImageJ Fiji software. The following parameters were analyzed: lipid content, LD number, LD size and LD area within the total embryo. The most important results show that lipid parameters in the IVF vs. PA bovine embryos differ at the most crucial moments of embryonic development (zygote, 8–16-cell, blastocyst), indicating possible dysregulations of lipid metabolism in PA embryos. When bovine vs. porcine species are compared, we observe higher lipid content around EGA stage and lower lipid content at the blastocyst stage for bovine embryos, which indicates different demand for energy depending on the species. We conclude that lipid droplets parameters significantly differ among developmental stages and between species but also can be affected by the genome origin.