Dataset Information

Suboptimal culture conditions induce more deviations in gene expression in male than in female bovine blastocysts

ABSTRACT: Background: Since the development of in vitro embryo production in cattle, different supplements have been added to embryo culture media to support development, with serum being the most popular. However, the addition of serum during embryo culture can induce high birth weights and low viability in calves (Large Offspring Syndrome). Global gene expression analysis of bovine embryos through RNA sequencing produced in different conditions (in vivo, in vitro with serum added and in vitro without addition of serum) can provide valuable information to optimize culture media for in vitro embryo production, which is necessary to obtain healthy offspring. Results: Here, we used RNA sequencing to examine the effect of in vitro embryo production, either in serum or in serum-free conditions, on the global gene expression pattern of individual bovine blastocysts. For this purpose, we compared in vitro produced with in vivo derived embryos. The embryos produced in serum had ten times more genes differentially expressed than the embryos produced in serum-free conditions (1,106 vs. 110). Importantly, in vitro production appeared to have a different impact on the embryos according to their sex, with male embryos showing more deviations in their gene expression pattern than their female counterparts. In particular, embryos produced in the presence of serum showed the largest difference, with male embryos having eight times more genes differentially expressed than the females (1,561 vs. 200). This difference was reduced when the embryos were produced in serum-free conditions, where the male embryos had only about double the number of genes differentially expressed than their female counterparts (64 vs. 35). The pathways affected by in vitro production differed depending on the supplementation. While embryos produced in serum conditions had altered genes involved in RNA processing and mitosis, embryos produced in serum-free conditions showed aberrations in genes involved in lipid metabolism. Conclusions: Serum supplementation had a major impact on the gene expression pattern of the embryos. On the contrary, the transcriptome of embryos produced in serum-free conditions resembled more in vivo derived embryos, although genes involved in lipid metabolism were altered. Male embryos appeared to be most affected by in vitro production, especially after serum supplementation.

ORGANISM(S): Bos taurus

PROVIDER: GSE74675 | GEO | 2016/01/27

SECONDARY ACCESSION(S): PRJNA301141

REPOSITORIES: GEO

ACCESS DATA

Similar Datasets

Project description:Despite many improvements with in vitro culture systems, the quality and developmental ability of mammalian embryos produced in vitro is still lower than their in vivo counterparts. To bring knowledge to answer this question, we used a mass spectrometry (MS) approach to compare the protein content of bovine embryos that were conceived in vivo or produced in vitro. A total of 38 pools of grade-1 quality bovine embryos at the 4-6 cell, 8-12 cell, morula, compact morula and blastocyst stages developed either in vivo or in vitro were analyzed by nano-liquid chromatography coupled with label-free quantitative mass spectrometry, allowing the identification of 3,028 proteins. Multivariate analysis of quantified proteins showed a clear separation of embryo pools according to their in vivo or in vitro origin at all stages. Three clusters of differentially abundant proteins (DAPs) according to embryo origin were evidenced, including 463 proteins more abundant in vivo than in vitro across development, and 314 and 222 proteins more abundant in vitro than in vivo before and after the morula stage, respectively. The functional analysis of proteins found more abundant in vivo showed an enrichment in carbohydrate metabolism and cytoplasmic cellular components. Proteins found more abundant in vitro before the morula stage were mostly localized in mitochondrial matrix and involved in ATP-dependent activity while those overabundant after morula stage were mostly localized in the ribonucleoprotein complex and involved in protein synthesis. Oviductin and other proteins previously shown to interact with early embryos in vitro were among the most overabundant proteins after in vivo development. In conclusion, the maternal environment led to faster degradation of mitochondrial proteins at early embryo developmental stages, lower abundance of proteins involved in protein synthesis at the time of embryonic genome activation and a global upregulation of carbohydrate and small molecule metabolic pathways compared to in vitro produced embryos. Furthermore, our data confirm that embryos developed in vivo uptake large amounts of oviduct fluid-derived proteins as soon as the 4-6 cell stage. These data provide new insight into the molecular contribution of the mother to the developmental ability of early embryos and will help designing better in vitro culture systems.

Project description:Differentiation of totipotent embryonic cells into the first cell lineages (trophectoderm (TE) and inner cell mass (ICM), and further into epiblast (EPI) and primitive endoderm (PE)), although well studied, is only beginning to be understood at the molecular level. An in depth understanding of the regulation of these early differentiation events would enable improvements in the quality of in vitro produced embryos, as well as the study and application of bovine embryonic stem cells. The objective of this work was to perform single cell RNASeq on bovine blastocysts derived in vivo (IVV), in vitro in a conventional in vitro production system (IVC), and in vitro in reduced nutrient media (IVR). All in vitro culture was serum free. In vivo embryonic cells could be assigned based on Leiden clustering analysis, although there was some overlap in expression of marker genes. The two TE groups identified clearly separated in a uniform manifold approximation and projection (UMAP) plot from EPI groups. Putative epiblast cells could be identified. In in vitro embryos produced in a conventional system, considerable overlap in expression of marker genes made Leiden clustering difficult, but a selected count could be used for UMAP to tentatively identify one TE, 2 EPI and 2 PE groups. In assigned cell types we observed higher expression of the identifying marker genes, but some marker genes were expressed in all cell types. In IVC embryos, TE markers clustered while PE and EPI marker genes overlapped. In IVR embryos, again there was considerable overlap between marker gene expression. Like IVC embryos, TE was easily identified by clustering in IVR, but EPI and PE were too similar to distinguish. In summary, the methods used here could partially distinguish putative cell types in bovine blastocysts, although marker gene expression was not always discreet. This may not be surprising given differentiation events are in progress in the blastocyst stage embryo. TE cells were more easily distinguished than EPI and PE cells, which would support this hypothesis.

Dataset Information

Suboptimal culture conditions induce more deviations in gene expression in male than in female bovine blastocysts

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure