Different pre-implantation phenotypes of bovine blastocysts produced in vitro.

Embryo transfer in cattle is performed with blastocysts produced in vivo or in vitro using defined media. However, outdated systems such as those that use serum and co-culture remain of interest for research purposes. Here, we investigated the effect of additional culture time on in vitro-produced embryos.

STAT3 signaling stimulates miR-21 expression in bovine cumulus cells during in vitro oocyte maturation.

MicroRNAs are potent regulators of gene expression that have been widely implicated in reproduction and embryo development. Recent studies have demonstrated that miR-21, a microRNA extensively studied in the context of disease, is important in multiple facets of reproductive biology including folliculogenesis, ovulation, oocyte maturation and early mammalian development. Surprisingly, little is known about the mechanisms that regulate miR-21 and no studies have characterized these regulatory pathways in cumulus-oocyte complexes (COCs).

Short-term effect of FSH on gene expression in bovine granulosa cells in vitro.

In reproduction, FSH is one of the most important hormones, especially in females, because it controls the number of follicles and the rate of follicular growth. Although several studies have examined the follicular response at the transcriptome level, it is difficult to obtain a clear and complete picture of the genes responding to an increase in FSH in an in vivo context because follicles undergo rapid morphological and physical changes during their growth. To help define the transcriptome downstream response to FSH, an in vitro model was used in the present study to observe the short-term (4h) cellular response.

Metabolic stress induces modifications in the epigenetic program of preimplantation bovine embryos.

The mammalian embryo is sensitive to and adapts to its metabolic environment. The mother's metabolic health and nutrient availability, for example, can modulate the oviductal fluid composition and thus embryo development. In this project, we induced energetic stress in bovine embryos during early culture to observe the epigenetic responses associated with metabolic stress, using a treatment paradigm known to decrease blastocyst rates.

Transcriptomic difference in bovine blastocysts following vitrification and slow freezing at morula stage.

Cryopreservation is known for its marked deleterious effects on embryonic health. Bovine compact morulae were vitrified or slow-frozen, and post-warm morulae were cultured to the expanded blastocyst stage. Blastocysts developed from vitrified and slow-frozen morulae were subjected to microarray analysis and compared with blastocysts developed from unfrozen control morulae for differential gene expression.

Spermatozoa DNA methylation patterns differ due to peripubertal age in bulls.

In the dairy industry, using semen as soon as the bull is mature enough to produce it is advantageous for breeding purposes. Mammalian spermatogenesis is a hormone-dependent developmental program in which a complex cascade of events must take place to ensure that germ cells reach the proper stage of development at the proper time. Conventional indicators of semen quality such as sperm cell motility and viability usually improve as bulls mature, meeting quality criteria satisfactorily at around 16 months.

Regulation of ATF1 and ATF2 transcripts by sequences in their 3' untranslated region in cleavage-stage cattle embryos.

The sequence of a 3' untranslated region (3'UTR) of mRNA governs the timing of its polyadenylation and translation in mammalian oocytes and early embryos. The objective of this study was to assess the influence of cis-elements in the 3'UTR of the developmentally important ATF1 and ATF2 transcripts on their timely translation during first cleavages in bovine embryos. Eight different reporter mRNAs (coding sequence of green fluorescent protein [GFP] fused to the 3'UTR of short or long isoforms of cattle ATF1 or -2, with or without polyadenylation) or a control GFP mRNA were microinjected separately into presumptive bovine zygotes at 18 hr post-insemination (hpi), followed by epifluorescence assessment for GFP translation between 24 and 80 hpi (expressed as percentage of GFP-positive embryos calculated from the total number of individuals).

Differences in cumulus cell gene expression indicate the benefit of a pre-maturation step to improve in-vitro bovine embryo production.

Study Question: Does the gene expression profile of cumulus cells (CC) accompanying oocytes with different degrees of chromatin compaction within the germinal vesicle (GV) reflect the oocyte's quality and response in culture during in-vitro embryo production (IVP).

Summary Answer: The transcriptomic profile of the CC is related to oocyte competence, setting the stage for the development of customized pre-maturation strategies to improve IVP.

What Is Known Already: Oocytes complete the acquisition of their competence during antral follicle development.

Meta-analysis of gene expression profiles in granulosa cells during folliculogenesis.

Folliculogenesis involves coordinated profound changes in different follicular compartments and significant modifications of their gene expression patterns, particularly in granulosa cells. Huge datasets have accumulated from the analyses of granulosa cell transcriptomic signatures in predefined physiological contexts using different technological platforms. However, no comprehensive overview of folliculogenesis is available.

Epigenetic modification with trichostatin A does not correct specific errors of somatic cell nuclear transfer at the transcriptomic level; highlighting the non-random nature of oocyte-mediated reprogramming errors.

Background: The limited duration and compromised efficiency of oocyte-mediated reprogramming, which occurs during the early hours following somatic cell nuclear transfer (SCNT), may significantly interfere with epigenetic reprogramming, contributing to the high incidence of ill/fatal transcriptional phenotypes and physiological anomalies occurring later during pre- and post-implantation events. A potent histone deacetylase inhibitor, trichostatin A (TSA), was used to understand the effects of assisted epigenetic modifications on transcriptional profiles of SCNT blastocysts and to identify specific or categories of genes affected.

Results: TSA improved the yield and quality of in vitro embryo development compared to control (CTR-NT).

Effect of vitrification using the Cryotop method on the gene expression profile of in vitro-produced bovine embryos.

The present study analyzed the changes in gene expression induced by the Cryotop vitrification technique in bovine blastocyst-stage embryos, using Agilent EmbryoGENE microarray slides. Bovine in vitro-produced embryos were vitrified and compared with nonvitrified (control) embryos. After vitrification, embryos were warmed and cultured for an additional 4 hours.

The effect of energy balance on the transcriptome of bovine granulosa cells at 60 days postpartum.

Dairy cows expend great amounts of energy during the lactation peak to cope with milk production. A state of negative energy balance (NEB) was suggested as a cause for the suboptimal fertility observed during this period, via an interaction with ovarian function. The objective of this study was to identify the impact of NEB on gene expression in granulosa cells of dairy cows at 60 days postpartum and to suggest a potential treatment to improve ovarian function.

Thyroid hormones alter the transcriptome of in vitro-produced bovine blastocysts.

Thyroid hormones (THs) have been shown to improve in vitro embryo production in cattle by increasing blastocyst formation rate, and the average cell number of blastocysts and by significantly decreasing apoptosis rate. To better understand those genetic aspects that may underlie enhanced early embryo development in the presence of THs, we characterized the bovine embryonic transcriptome at the blastocyst stage, and examined differential gene expression profiles using a bovine-specific microarray. We found that 1212 genes were differentially expressed in TH-treated embryos when compared with non-treated controls (>1.

Global gene expression in granulosa cells of growing, plateau and atretic dominant follicles in cattle.

Background: The physiological state of the dominant follicle is important as it may be linked to the competence of the oocyte within. The objective of this study was to analyze, by transcriptomic analysis, the changes occurring in granulosa cells from dominant follicles at different phases of follicular growth.

Methods: Granulosa cells were collected from slaughterhouse dairy cattle follicles with a diameter greater than 9 mm, and were classified at different phases of follicle growth based on flow cytometry profiles of DNA content after staining with propidium iodide.

Stable reference genes in granulosa cells of bovine dominant follicles during follicular growth, FSH stimulation and maternal aging.

The aim of the present study was to determine a set of reference genes in granulosa cells of dominant follicles that are suitable for relative gene expression analyses during maternal and follicular aging. Granulosa cells of growing and preovulatory dominant follicles were collected from aged and young cows (maternal aging study) and from FSH-stimulated follicles developing under different durations of FSH treatment (follicular aging study). The mRNA levels of the two commonly used reference genes (GAPDH, ACTB) and four novel genes (UBE2D2, EIF2B2, SF3A1, RNF20) were analysed using cycle threshold values.

Studying bovine early embryo transcriptome by microarray.

Microarrays represent a significant advantage when studying gene expression in early embryo because they allow for a speedy study of a large number of genes even if the sample of interest contains small quantities of genetic material. Here we describe the protocols developed by the EmbryoGENE Network to study the bovine transcriptome in early embryo using a microarray experimental design.

Sexual dimorphism in developmental programming of the bovine preimplantation embryo caused by colony-stimulating factor 2.

Physiology of the adult can be modified by alterations in prenatal development driven by the maternal environment. Developmental programming, which can be established before the embryo implants in the uterus, can affect females differently than males. The mechanism by which sex-specific developmental programming is established is not known.

Rapidly cleaving bovine two-cell embryos have better developmental potential and a distinctive mRNA pattern.

Mammalian embryos that rapidly reach the two-cell stage in culture have a higher probability of becoming viable blastocysts. Our goal was to separate two-cell bovine embryos based on their zygotic cleavage timing, and to assess their global mRNA levels. Following in vitro fertilization, all embryos that cleaved by 29.

Discovery, identification and sequence analysis of RNAs selected for very short or long poly A tail in immature bovine oocytes.

A major challenge in applying genomics to oocyte physiology is that many RNAs are present but will not be translated into proteins, making it difficult to draw conclusions from RNAseq and array data. Oocyte maturation and early embryo development rely on maternal storage of specific RNAs with a short poly(A) tail, which must be elongated for translation. To resolve the role of key genes during that period, we aimed to characterize both extremes of mRNA: deadenylated RNA and long polyA tails mRNA population in immature bovine oocytes.

Evolutionary conservation of the oocyte transcriptome among vertebrates and its implications for understanding human reproductive function.

Cross-phylum and cross-species comparative transcriptomic analyses provide an evolutionary perspective on how specific tissues use genomic information. A significant mRNA subset present in the oocytes of most vertebrates is stabilized or stored for post-LH surge use. Since transcription is arrested in the oocyte before ovulation, this RNA is important for completing maturation and sustaining embryo development until zygotic genome activation.

Molecular mechanisms and pathways involved in bovine embryonic genome activation and their regulation by alternative in vivo and in vitro culture conditions.

Understanding gene expression patterns in response to altered environmental conditions at different time points of the preimplantation period would improve our knowledge on regulation of embryonic development. Here we aimed to examine the effect of alternative in vivo and in vitro culture conditions at the time of major embryonic genome activation (EGA) on the development and transcriptome profile of bovine blastocysts. Four different blastocyst groups were produced under alternative in vivo and in vitro culture conditions before or after major EGA.

Differential gene expression profile in bovine blastocysts resulting from hyperglycemia exposure during early cleavage stages.

To understand the compromised survival of embryos derived from assisted reproductive techniques, transcriptome survey of early embryonic development has shown the impact of in vitro culture environment on gene expression in bovine or other living species. However, how the differentially expressed genes translate into developmentally compromised embryos is unresolved. We therefore aimed to characterize transcriptomic markers expressed by bovine blastocysts cultured in conditions that are known to impair embryo development.

Combining resources to obtain a comprehensive survey of the bovine embryo transcriptome through deep sequencing and microarrays.

While most assisted reproductive technologies (ART) are considered routine for the reproduction of species of economical importance, such as the bovine, the impact of these manipulations on the developing embryo remains largely unknown. In an effort to obtain a comprehensive survey of the bovine embryo transcriptome and how it is modified by ART, resources were combined to design an embryo-specific microarray. Close to one million high-quality reads were produced from subtracted bovine embryo libraries using Roche 454 Titanium deep sequencing technology, which enabled the creation of an augmented bovine genome catalog.

Analysis of the gene expression pattern of bovine blastocysts at three stages of development.

Blastocyst formation is a primordial event of pre-implantation development since it is required for pregnancy establishment and progression. The blastocyst plays a pivotal role because it is the stage at which the embryo starts coordinated cross-talk with the mother. It is also the terminal step before transfer in bovine; it reflects all stresses the embryo may have faced during the process of in vitro treatment.

Identification of follicular marker genes as pregnancy predictors for human IVF: new evidence for the involvement of luteinization process.

Multiple pregnancy represents an important health risk to both mother and child in fertility treatment. To reduce a high twin rate, restriction to one embryo transfer is needed. Morphological evaluation methods for predicting embryo viability has significant limitations.