Single-cell transcriptional landscapes of bovine peri-implantation development.

Supporting healthy pregnancy outcomes requires a comprehensive understanding of the molecular and cellular programs of peri-implantation development, when most pregnancy failure occurs. Here, we present single-cell transcriptomes of bovine peri-implantation embryo development at day 12, 14, 16, and 18 post-fertilization. We defined the cellular composition and gene expression of embryonic disc, hypoblast, and trophoblast lineages in bovine peri-implantation embryos, and identified markers and pathway signaling that represent distinct stages of bovine peri-implantation lineages; the expression of selected markers was validated in peri-implantation embryos. Using detailed time-course transcriptomic analyses, we revealed a previously unrecognized primitive trophoblast cell lineage. We also characterized conserved and divergence peri-implantation lineage programs between bovine and other mammalian species. Finally, we established cell-cell communication signaling underlies embryonic and extraembryonic cell interaction to ensure proper early development. These data provide foundational information to discover essential biological signaling underpinning bovine peri-implantation development.

In Vitro Culture Alters Cell Lineage Composition and Cellular Metabolism of Bovine Blastocyst.

Profiling bovine blastocyst transcriptome at the single-cell level has enabled us to reveal the first cell lineage segregation, during which the inner cell mass (ICM), trophectoderm (TE), and an undefined population of transitional cells were identified. By comparing the transcriptome of blastocysts derived in vivo (IVV), in vitro from a conventional culture medium (IVC), and in vitro from an optimized reduced nutrient culture medium (IVR), we found a delay of the cell fate commitment to ICM in the IVC and IVR embryos. Developmental potential differences between IVV, IVC, and IVR embryos were mainly contributed by ICM and transitional cells. Pathway analysis of these non-TE cells between groups revealed highly active metabolic and biosynthetic processes, reduced cellular signaling, and reduced transmembrane transport activities in IVC embryos that may lead to reduced developmental potential. IVR embryos had lower activities in metabolic and biosynthetic processes but increased cellular signaling and transmembrane transport, suggesting these cellular mechanisms may contribute to improved blastocyst development compared to IVC embryos. However, the IVR embryos had compromised development compared to IVV embryos with notably over-active transmembrane transport activities that impaired ion homeostasis.

Maternal restricted- and over- feeding during gestation perturb offspring sperm epigenome in sheep.

In brief: Inadequate maternal nutrition during gestation can have immediate and lifelong effects on offspring. This study shows that maternal restricted - and over- nutrition during gestation do not affect semen characteristics in F1 male offspring but alters offspring sperm sncRNA profiles and DNA methylome in sheep. Abstract: There is a growing body of evidence that inadequate maternal nutrition during gestation can have immediate and lifelong effects on offspring. However, little is known about the effects of maternal nutrition during gestation on male offspring reproduction. Here, using a sheep model of maternal restricted - and over - nutrition (60 or 140% of the National Research Council requirements) during gestation, we found that maternal restricted - and over - nutrition do not affect semen characteristics (i.e. volume, sperm concentration, pH, sperm motility, sperm morphology) or scrotal circumference in male F1 offspring. However, using small RNA sequencing analysis, we demonstrated that both restricted - and over - nutrition during gestation induced marked changes in composition and expression of sperm small noncoding RNAs (sncRNAs) subpopulations including in male F1 offspring. Whole-genome bisulfite sequencing analysis further identified specific genomic loci where poor maternal nutrition resulted in alterations in DNA methylation. These findings indicate that maternal restricted - and over - nutrition during gestation induce epigenetic modifications in sperm of F1 offspring sperm in sheep, which may contribute to environmentally influenced phenotypes in ruminants.

Single-cell transcriptional landscapes of bovine peri-implantation development.

Supporting healthy pregnancy outcomes requires a comprehensive understanding of the cellular hierarchy and underlying molecular mechanisms during peri-implantation development. Here, we present a single-cell transcriptome-wide view of the bovine peri-implantation embryo development at day 12, 14, 16 and 18, when most of the pregnancy failure occurs in cattle. We defined the development and dynamic progression of cellular composition and gene expression of embryonic disc, hypoblast, and trophoblast lineages during bovine peri-implantation development. Notably, the comprehensive transcriptomic mapping of trophoblast development revealed a previously unrecognized primitive trophoblast cell lineage that is responsible for pregnancy maintenance in bovine prior to the time when binucleate cells emerge. We analyzed novel markers for the cell lineage development during bovine early development. We also identified cell-cell communication signaling underling embryonic and extraembryonic cell interaction to ensure proper early development. Collectively, our work provides foundational information to discover essential biological pathways underpinning bovine peri-implantation development and the molecular causes of the early pregnancy failure during this critical period. Significance Statement: Peri-implantation development is essential for successful reproduction in mammalian species, and cattle have a unique process of elongation that proceeds for two weeks prior to implantation and represents a period when many pregnancies fail. Although the bovine embryo elongation has been studied histologically, the essential cellular and molecular factors governing lineage differentiation remain unexplored. This study profiled the transcriptome of single cells in the bovine peri-implantation development throughout day 12, 14, 16, and 18, and identified peri-implantation stage-related features of cell lineages. The candidate regulatory genes, factors, pathways and embryonic and extraembryonic cell interactions were also prioritized to ensure proper embryo elongation in cattle.

In Vitro Culture Alters Cell Lineage Composition and Cellular Metabolism of Bovine Blastocyst.

Profiling transcriptome at single cell level of bovine blastocysts derived in vivo (IVV), in vitro from conventional culture medium (IVC), and reduced nutrient culture medium (IVR) has enabled us to reveal cell lineage segregation, during which forming inner cell mass (ICM), trophectoderm (TE), and an undefined population of transitional cells. Only IVV embryos had well-defined ICM, indicating in vitro culture may delay the first cell fate commitment to ICM. Differences between IVV, IVC and IVR embryos were mainly contributed by ICM and transitional cells. Pathway analysis by using the differentially expressed genes of these non-TE cells between groups pointed to highly active metabolic and biosynthetic processes, with reduced cellular signaling and membrane transport in IVC embryos, which may lead to reduced developmental potential. IVR embryos had lower activities in metabolic and biosynthetic processes, but increased cellular signaling and membrane transport, suggesting these cellular mechanisms may contribute to the improved blastocyst development compared to IVC embryos. However, the IVR embryos had compromised development when compared to IVV embryos with notably over-active membrane transport activities that led to impaired ion homeostasis.

Establishment of bovine trophoblast stem cells.

Here, we report that a chemical cocktail (LCDM: leukemia inhibitory factor [LIF], CHIR99021, dimethinedene maleate [DiM], minocycline hydrochloride), previously developed for extended pluripotent stem cells (EPSCs) in mice and humans, enables de novo derivation and long-term culture of bovine trophoblast stem cells (TSCs). Bovine TSCs retain developmental potency to differentiate into mature trophoblast cells and exhibit transcriptomic and epigenetic (chromatin accessibility and DNA methylome) features characteristic of trophectoderm cells from early bovine embryos. The bovine TSCs established in this study will provide a model to study bovine placentation and early pregnancy failure.

High-resolution ribosome profiling reveals translational selectivity for transcripts in bovine preimplantation embryo development.

High-resolution ribosome fractionation and low-input ribosome profiling of bovine oocytes and preimplantation embryos has enabled us to define the translational landscapes of early embryo development at an unprecedented level. We analyzed the transcriptome and the polysome- and non-polysome-bound RNA profiles of bovine oocytes (germinal vesicle and metaphase II stages) and early embryos at the two-cell, eight-cell, morula and blastocyst stages, and revealed four modes of translational selectivity: (1) selective translation of non-abundant mRNAs; (2) active, but modest translation of a selection of highly expressed mRNAs; (3) translationally suppressed abundant to moderately abundant mRNAs; and (4) mRNAs associated specifically with monosomes. A strong translational selection of low-abundance transcripts involved in metabolic pathways and lysosomes was found throughout bovine embryonic development. Notably, genes involved in mitochondrial function were prioritized for translation. We found that translation largely reflected transcription in oocytes and two-cell embryos, but observed a marked shift in the translational control in eight-cell embryos that was associated with the main phase of embryonic genome activation. Subsequently, transcription and translation become more synchronized in morulae and blastocysts. Taken together, these data reveal a unique spatiotemporal translational regulation that accompanies bovine preimplantation development.

Blockchain Data Secure Transmission Method Based on Homomorphic Encryption.

To ensure the security of data transmission and recording in Internet environment monitoring systems, this paper proposes a study of a secure method of blockchain data transfer based on homomorphic encryption. Blockchain data transmission is realized through homomorphic encryption. Homomorphic encryption can not only encrypt the original data, but also ensure that the data result after decrypting the data is the same as the original data. The asymmetric encrypted public key is collected by Internet of things (IoT) equipment to realize the design of blockchain data secure transmission method based on homomorphic encryption. The experimental results show that the accuracy of the first transmission is as high as 88% when using the transmission method in this paper. After several experiments, the transmission accuracy is high by using the design method in this paper. In the last test, the transmission accuracy is still 88%, and the data transmission effect is relatively stable. At the same time, compared to the management method used in this article, the transfer method used in this paper is more reliable than the original transfer method and is not prone to data distortion. It can be seen that this method has high transmission accuracy and short transmission time, which effectively avoids the data tampering caused by too long time in the transmission process.

Oocyte specific lncRNA variant Rose influences oocyte and embryo development.

Fully grown mammalian oocytes store a large amount of RNA synthesized during the transcriptionally active growth stage. A large part of the stored RNA belongs to the long non-coding class which contain either transcriptional noise or important contributors to cellular physiology. Despite the expanding number of studies related to lncRNAs, their influence on oocyte physiology remains enigmatic. We found an oocyte specific antisense, long non-coding RNA, "Rose" (lncRNA in Oocyte Specifically Expressed) expressed in two variants containing two and three non-coding exons, respectively. Rose is localized in the nucleus of transcriptionally active oocyte and in embryo with polysomal occupancy in the cytoplasm. Experimental overexpression of Rose in fully grown oocyte did not show any differences in meiotic maturation. However, knocking down Rose resulted in abnormalities in oocyte cytokinesis and impaired preimplantation embryo development. In conclusion, we have identified an oocyte-specific maternal lncRNA that is essential for successful mammalian oocyte and embryo development.

The Epigenetics of Gametes and Early Embryos and Potential Long-Range Consequences in Livestock Species-Filling in the Picture With Epigenomic Analyses.

The epigenome is dynamic and forged by epigenetic mechanisms, such as DNA methylation, histone modifications, chromatin remodeling, and non-coding RNA species. Increasing lines of evidence support the concept that certain acquired traits are derived from environmental exposure during early embryonic and fetal development, i.e., fetal programming, and can even be "memorized" in the germline as epigenetic information and transmitted to future generations. Advances in technology are now driving the global profiling and precise editing of germline and embryonic epigenomes, thereby improving our understanding of epigenetic regulation and inheritance. These achievements open new avenues for the development of technologies or potential management interventions to counteract adverse conditions or improve performance in livestock species. In this article, we review the epigenetic analyses (DNA methylation, histone modification, chromatin remodeling, and non-coding RNAs) of germ cells and embryos in mammalian livestock species (cattle, sheep, goats, and pigs) and the epigenetic determinants of gamete and embryo viability. We also discuss the effects of parental environmental exposures on the epigenetics of gametes and the early embryo, and evidence for transgenerational inheritance in livestock.