Transcription profiles of oocytes during maturation and embryos during preimplantation development in vivo in the goat

RNA sequencing performed on goat matured oocytes and preimplantation embryos generated invivo enabled us to define the transcriptome for goat preimplantation embryo development. The largest proportion of changes in gene expression in goat was found at the 16-cell stage, not as previously defined at the 8-cell stage, and is later than in other mammalian species. In all, 6482 genes were identified to be significantly differentially expressed across all consecutive developmental stage comparisons, and the important signalling pathways involved in each development transition were determined. In addition, we identified genes that appear to be transcribed only at a specific stage of development. Using weighted gene coexpression network analysis, we found nine stage-specific modules of coexpressed genes that represent the corresponding stage of development. Furthermore, we identified conserved key members (or hub genes) of the goat transcriptional networks. Their association with other embryo genes suggests that they may have important regulatory roles in embryo development. Our cross-mammalian species transcriptomic comparisons demonstrate both conserved and goat-specific features of preimplantation development.

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Amino acid depletion and appearance during porcine preimplantation embryo development in vitro

Reproduction ◽

10.1530/rep.1.00727 ◽

2005 ◽

Vol 130 (5) ◽

pp. 655-668 ◽

Cited By ~ 37

Author(s):

Paul J Booth ◽

Peter G Humpherson ◽

Terry J Watson ◽

Henry J Leese

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Embryo Development ◽

Preimplantation Embryos ◽

Type I ◽

Preimplantation Embryo Development ◽

Type Iv ◽

Stage Of Development ◽

Amino Acid Depletion

Preimplantation embryos can consume and produce amino acids in a manner dependent upon the stage of development that may be predictive of subsequent viability. In order to examine these relationships in the pig, patterns of net depletion and appearance of amino acids byin vitroproduced porcine preimplantation embryos were examined. Cumulus oocyte complexes derived from slaughterhouse pre-pubertal pig ovaries were matured for 40 h in defined TCM-199 medium (containing PVA) before being fertilised (Day 0) with frozen-thawed semen in Tris–based medium. After 6 h, presumptive zygotes were denuded and cultured in groups of 20, in NCSU-23 medium modified to contain 0.1 mM glutamine plus a mixture of 19 amino acids (aa) at low concentrations (0.02–0.11 mM) (NCSU-23aa). Groups of 2–20 embryos were removed (dependent on stage) on Day 0 (1 cell), Day 1 (two- and four-cells), Day 4 (compact morulae) and Day 6 (blastocysts) and placed in 4 μl NCSU-23aafor 24 h. After incubation, the embryos were removed and the spent media was analysed by HPLC. The net rate of amino acid depletion or appearance varied according to amino acid (P< 0.001) and, apart from serine and histidine, stage of development (P< 0.014). Glycine, isoleucine, valine, phenylalanine, tryptophan, methionine, asparagine, lysine, glutamate and aspartate consistently appeared, whereas threonine, glutamine and arginine were consistently depleted. Five types of stage-dependent trends could be observed: Type I: amino acids having high rates of net appearance on Day 0 that reached a nadir on Day 1 or 4 but subsequently increased by Day 6 (glycine, glutamate); Type II: those that exhibited lower rates of net appearance on Days 0 and 6 compared with the intermediate Days 1 and 4 (isoleucine, valine, phenylalanine, methionine, arginine); Type III: amino acids which showed a continuous fall in net appearance (asparagine, aspartate); Type IV: those that exhibited a steady fall in net depletion from Day 0 to Day 6 (glutamine, threonine); Type V: those following no discernable trend. Analysis of further embryo types indicated that presumptive polyspermic embryos on Day 0 had increased (P< 0.05) net rates of leucine, isoleucine, valine and glutamate appearance, and reduced (P< 0.05) net rates of threonine and glutamine depletion compared with normally inseminated oocytes. These data suggest that the net rates of depletion and uptake of amino acids by pig embryos vary between a) amino acids, b) the day of embryo development and, c) the type of embryos present at a given stage of development. The results also suggested that the net depletion and appearance rates of amino acids by early pig embryos might be more similar to those of the human than those of the mouse and cow.

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l-Carnitine affects preimplantation embryo development toward infertility in mice

Reproduction ◽

10.1530/rep-16-0290 ◽

2016 ◽

Vol 152 (4) ◽

pp. 283-291 ◽

Cited By ~ 5

Author(s):

Christiana Kyvelidou ◽

Dimitris Sotiriou ◽

Tania Antonopoulou ◽

Margarita Tsagkaraki ◽

George J Tserevelakis ◽

...

Keyword(s):

Embryo Development ◽

Preimplantation Embryo ◽

Day Treatment ◽

Intraperitoneal Administration ◽

Blastocyst Development ◽

Cell Stage ◽

Preimplantation Embryo Development ◽

Early Stages

l-Carnitine (l-Cn), despite the beneficial role as energy-generating substance delivering long-chain fatty acids to the β-oxidation pathway in mitochondria, has been accused to cause an endometriosis-like state to BALB/c mice manifested by increased inflammatory cytokines in serum and peritoneal fluid, accumulation of immune cells in the peritoneal cavity and uterine walls and most importantly, correlating to infertility. Exploring this type of infertility, the effect of l-Cn on preimplantation embryo development, ovarian integrity and systemic maternal immunity was studied. Using nonlinear microscopy analysis, which was shown to be a powerful tool for determining embryo quality by quantitatively estimating the lipid body (LB) content of the cells, it was shown that in vitro and in vivo administration of l-Cn significantly decreased LB mean area in zygotes. Daily intraperitoneal administration of 2.5mg l-Cn for 3, 4 and 7days to mice significantly decreased the percent of normal zygotes. However, only the 7-day treatment persisted by affecting 2- and 8-cell stage embryos, while almost abolishing blastocyst development. Such effects were accompanied by abnormal ovarian histology, showing increased numbers of corpora luteus and elevated progesterone concentration in the serum. In addition, it was shown that the 7-day l-Cn treatment pushed maternal systemic immunity toward inflammation and immunosuppression by increasing CD11b-, CD25- and CD11bGr1-positive cells in spleen, which opposed the necessity for immunostimulation at these early stages of pregnancy. In conclusion, the results presented here demonstrated that elevated doses of l-Cn affect early stages of embryo development, leading to infertility.

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Involvement of Nlrp9a/b/c in mouse preimplantation development

Reproduction ◽

10.1530/rep-19-0516 ◽

2020 ◽

Vol 160 (2) ◽

pp. 181-191 ◽

Cited By ~ 2

Author(s):

Satoko Kanzaki ◽

Shiori Tamura ◽

Toshiaki Ito ◽

Mizuki Wakabayashi ◽

Koji Saito ◽

...

Keyword(s):

Asymmetric Cell Division ◽

Culture Conditions ◽

Preimplantation Development ◽

Preimplantation Embryos ◽

Dependent Manner ◽

Blastocyst Development ◽

Triple Mutant ◽

Cell Stage

Nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing proteins (NRLPs) are central components of the inflammasome. Accumulating evidence has shown that a reproductive clade of NRLPs is predominantly expressed in oocyte to cleavage stage embryos and participates in mammalian preimplantation development as a component of a multiprotein complex known as the subcortical maternal complex (SCMC). Nlrp9s belong to the reproductive class of NLRPs; Nlrp9b is unique in acting as an inflammasome against rotavirus in intestines. Here we generated mice carrying mutations in all three members of the Nlrp9a/b/c gene (Nlrp9 triple mutant (TMut) mice). When crossed with WT males, the Nlrp9 TMut females were fertile, but deliveries with fewer pups were increased in the mutants. Consistent with this, blastocyst development was retarded and lethality to the preimplantation embryos increased in the Nlrp9 TMut females in vivo. Under in vitro culture conditions, the fertilized eggs from the Nlrp9 TMut females exhibited developmental arrest at the two-cell stage, accompanied by asymmetric cell division. By contrast, double-mutant (DMut) oocytes (any genetic combination) did not exhibit the two-cell block in vitro, showing the functional redundancy of Nlrp9a/b/c. Finally, Nlrp9 could bind to components of the SCMC. These results show that Nlrp9 functions as an immune or reproductive NLRP in a cell-type-dependent manner.

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Embryos generated from oocytes lacking complex N- and O-glycans have compromised development and implantation

Reproduction ◽

10.1530/rep-12-0084 ◽

2012 ◽

Vol 144 (4) ◽

pp. 455-465 ◽

Cited By ~ 11

Author(s):

Patricia Grasa ◽

Heidy Kaune ◽

Suzannah A Williams

Keyword(s):

Embryo Development ◽

Subsequent Development ◽

Early Embryo ◽

Preimplantation Development ◽

Ovulation Rate ◽

Preimplantation Embryos ◽

Small Litter ◽

Delayed Implantation

Female mice generating oocytes lacking complexN- andO-glycans (double mutants (DM)) produce only one small litter before undergoing premature ovarian failure (POF) by 3 months. Here we investigate the basis of the small litter by evaluating ovulation rate and embryo development in DM (Mgat1F/FC1galt1F/F:ZP3Cre) and Control (Mgat1F/FC1galt1F/F) females. Surprisingly, DM ovulation rate was normal at 6 weeks, but declined dramatically by 9 weeks.In vitrodevelopment of zygotes to blastocysts was equivalent to Controls although all embryos from DM females lacked a normal zona pellucida (ZP) and ∼30% lacked a ZP entirely. In contrast,in vivopreimplantation development resulted in less embryos recovered from DM females compared with Controls at 3.5 days post coitum (dpc) (3.2±1.3 vs 7.0±0.6). Furthermore, only 45% of mated DM females contained embryos at 3.5 dpc. Of the preimplantation embryos collected from DM females, approximately half were morulae unlike Controls where the majority were blastocysts, indicating delayed embryo development in DM females. Post-implantation development in DM females was analysed to determine whether delayed preimplantation development affected subsequent development. In DM females at 5.5 dpc, only ∼40% of embryos found at 3.5 dpc had implanted. However, at 6.5 dpc, implantation sites in DM females corresponded to embryo numbers at 3.5 dpc indicating delayed implantation. At 9.5 dpc, the number of decidua corresponded to embryo numbers 6 days earlier indicating that all implanted embryos progress to midgestation. Therefore, a lack of complexN- andO-glycans in oocytes during development impairs early embryo development and viabilityin vivoleading to delayed implantation and a small litter.

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Whole-Genome DNA Methylation Dynamics of Sheep Preimplantation Embryo Investigated by Single-Cell DNA Methylome Sequencing

Frontiers in Genetics ◽

10.3389/fgene.2021.753144 ◽

2021 ◽

Vol 12 ◽

Author(s):

Zijing Zhang ◽

Jiawei Xu ◽

Shijie Lyu ◽

Xiaoling Xin ◽

Qiaoting Shi ◽

...

Keyword(s):

Dna Methylation ◽

Single Cell ◽

Embryo Development ◽

Molecular Mechanisms ◽

Preimplantation Embryo ◽

Preimplantation Embryos ◽

Whole Genome ◽

Cell Stage ◽

Preimplantation Embryo Development ◽

Developmental Block

The early stages of mammalian embryonic development involve the participation and cooperation of numerous complex processes, including nutritional, genetic, and epigenetic mechanisms. However, in embryos cultured in vitro, a developmental block occurs that affects embryo development and the efficiency of culture. Although the block period is reported to involve the transcriptional repression of maternal genes and transcriptional activation of zygotic genes, how epigenetic factors regulate developmental block is still unclear. In this study, we systematically analyzed whole-genome methylation levels during five stages of sheep oocyte and preimplantation embryo development using single-cell level whole genome bisulphite sequencing (SC-WGBS) technology. Then, we examined several million CpG sites in individual cells at each evaluated developmental stage to identify the methylation changes that take place during the development of sheep preimplantation embryos. Our results showed that two strong waves of methylation changes occurred, namely, demethylation at the 8-cell to 16-cell stage and methylation at the 16-cell to 32-cell stage. Analysis of DNA methylation patterns in different functional regions revealed a stable hypermethylation status in 3′UTRs and gene bodies; however, significant differences were observed in intergenic and promoter regions at different developmental stages. Changes in methylation at different stages of preimplantation embryo development were also compared to investigate the molecular mechanisms involved in sheep embryo development at the methylation level. In conclusion, we report a detailed analysis of the DNA methylation dynamics during the development of sheep preimplantation embryos. Our results provide an explanation for the complex regulatory mechanisms underlying the embryo developmental block based on changes in DNA methylation levels.

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Effects of embryo culture on global pattern of gene expression in preimplantation mouse embryos

Reproduction ◽

10.1530/rep.1.00297 ◽

2004 ◽

Vol 128 (3) ◽

pp. 301-311 ◽

Cited By ~ 193

Author(s):

Paolo Rinaudo ◽

Richard M Schultz

Keyword(s):

Gene Expression ◽

Preimplantation Embryos ◽

Expression Analysis Systematic Explorer ◽

Cell Stage ◽

Global Pattern ◽

Preimplantation Mouse Embryos ◽

Preimplantation Mouse ◽

Global Patterns ◽

The One

Culture of preimplantation embryos affects gene expression. The magnitude of the effect on the global pattern of gene expression, however, is not known. We compared global patterns of gene expression in blastocysts cultured from the one-cell stage in either Whitten’s medium or KSOM + amino acids (KSOM/AA) with that of blastocysts that developed in vivo, using the Affymetrix MOE430A chip. The analysis revealed that expression of 114 genes was affected after culture in Whitten’s medium, whereas only 29 genes were mis-expressed after culture in KSOM/AA. Expression Analysis Systematic Explorer was used to identify biological and molecular processes that are perturbed after culture and indicated that genes involved in protein synthesis, cell proliferation and transporter function were down-regulated after culture in Whitten’s medium. A common set of genes involved in transporter function was also down-regulated after culture in KSOM/AA. These results provide insights as to why embryos develop better in KSOM/AA than in Whitten’s medium, and highlight the power of microarray analysis to assess global patterns of gene expression.

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Spatiotemporal dynamics of OCT4 protein localization during preimplantation development in mice

Reproduction ◽

10.1530/rep-16-0277 ◽

2016 ◽

Vol 152 (5) ◽

pp. 417-430 ◽

Cited By ~ 10

Author(s):

Atsushi Fukuda ◽

Atsushi Mitani ◽

Toshiyuki Miyashita ◽

Hisato Kobayashi ◽

Akihiro Umezawa ◽

...

Keyword(s):

Nuclear Localization ◽

Splice Variants ◽

Protein Localization ◽

Preimplantation Development ◽

Preimplantation Embryos ◽

Sequencing Analysis ◽

Dependent Manner ◽

Cell Stage ◽

Protein Levels ◽

Oct4 Protein

Spatiotemporal expression of transcription factors is crucial for genomic reprogramming. Pou5f1 (Oct4) is an essential transcription factor for reprogramming. A recent study reported that OCT4A, which is crucial for establishment and maintenance of pluripotent cells, is expressed in oocytes, but maternal OCT4A is dispensable for totipotency induction. Whereas another study reported that OCT4B, which is not related to pluripotency, is predominantly expressed instead of OCT4A during early preimplantation phases in mice. To determine the expression states of OCT4 in murine preimplantation embryos, we conducted in-depth expression and functional analyses. We found that pluripotency-related OCT4 mainly localizes to the cytoplasm in early preimplantation phases, with no major nuclear localization until the 8–16-cell stage despite high expression in both oocytes and early embryos. RNA-sequencing analysis using oocytes and early preimplantation embryos could not identify the splice variants creating alternative forms of OCT4 protein. Forced expression of OCT4 in zygotes by the injection of polyadenylated mRNA clearly showed nuclear localization of OCT4 protein around 3–5-fold greater than physiological levels and impaired developmental competency in a dose-dependent manner. Embryos with modest overexpression of OCT4 could develop to the 16-cell stage; however, more than 50% of the embryos were arrested at this stage, similar to the results for OCT4 depletion. In contrast, extensive overexpression of OCT4 resulted in complete arrest at the 2-cell stage accompanied by downregulation of zygotically activated genes and repetitive elements related to the totipotent state. These results demonstrated that OCT4 protein localization was spatiotemporally altered during preimplantation development, and strict control of Oct4 protein levels was essential for proper totipotential reprogramming.

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Inhibition of DRP1 Impedes Zygotic Genome Activation and Preimplantation Development in Mice

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.788512 ◽

2021 ◽

Vol 9 ◽

Author(s):

Yuanyuan Li ◽

Ning-Hua Mei ◽

Gui-Ping Cheng ◽

Jing Yang ◽

Li-Quan Zhou

Keyword(s):

Embryo Development ◽

Preimplantation Embryo ◽

Preimplantation Embryos ◽

Dependent Manner ◽

Zygotic Genome Activation ◽

Preimplantation Embryo Development ◽

Cell Embryo ◽

Embryo Stage ◽

Genome Activation ◽

Zygotic Genome

Mitochondrion plays an indispensable role during preimplantation embryo development. Dynamic-related protein 1 (DRP1) is critical for mitochondrial fission and controls oocyte maturation. However, its role in preimplantation embryo development is still lacking. In this study, we demonstrate that inhibition of DRP1 activity by mitochondrial division inhibitor-1, a small molecule reported to specifically inhibit DRP1 activity, can cause severe developmental arrest of preimplantation embryos in a dose-dependent manner in mice. Meanwhile, DRP1 inhibition resulted in mitochondrial dysfunction including decreased mitochondrial activity, loss of mitochondrial membrane potential, reduced mitochondrial copy number and inadequate ATP by disrupting both expression and activity of DRP1 and mitochondrial complex assembly, leading to excessive ROS production, severe DNA damage and cell cycle arrest at 2-cell embryo stage. Furthermore, reduced transcriptional and translational activity and altered histone modifications in DRP1-inhibited embryos contributed to impeded zygotic genome activation, which prevented early embryos from efficient development beyond 2-cell embryo stage. These results show that DRP1 inhibition has potential cytotoxic effects on mammalian reproduction, and DRP1 inhibitor should be used with caution when it is applied to treat diseases. Additionally, this study improves our understanding of the crosstalk between mitochondrial metabolism and zygotic genome activation.

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Mitochondrial membrane potential in 2-cell stage embryos correlates with the success of preimplantation development

Reproduction ◽

10.1530/rep-13-0288 ◽

2014 ◽

Vol 147 (5) ◽

pp. 627-638 ◽

Cited By ~ 22

Author(s):

Kouji Komatsu ◽

Akira Iwase ◽

Miki Mawatari ◽

Jingwen Wang ◽

Mamoru Yamashita ◽

...

Keyword(s):

Membrane Potential ◽

Mitochondrial Membrane Potential ◽

Mitochondrial Membrane ◽

Developmental Process ◽

Preimplantation Embryos ◽

Cell Stage ◽

Hormonal Stimulation ◽

Maternal Mrna

Hormonal stimulation in superovulation induces female mice to ovulate more oocytes than spontaneous ovulation. Because the superovulated oocytes contain a number of oocytes that normally regress before spontaneous ovulation or immature oocytes, the development of some embryos that derive from these oocytes by IVF is prevented. Therefore, the quality of superovulated oocytes should differ from that of spontaneously ovulated oocytes. In this study, we evaluated the quality of superovulated oocytes, by examining 1- and 2-cell stage embryos, in which the development mainly depends on the maternal mRNA, proteins, and mitochondria that are contained in the oocytes, and we then measured the mitochondrial membrane potential (ΔΨm) of the 1- and 2-cell stage,in vivo-fertilized, and IVF embryos. The ΔΨmof 1-cell stage IVF embryos was lower than that ofin vivo-fertilized embryos; however, there was no difference between IVF embryos. During the developmental process from 1- to 2-cell stage, the ΔΨmofin vivo-fertilized embryos was highly upregulated, whereas a number of IVF embryos remained unchanged. As a result, 2-cell stage embryos were divided into two groups: high- and low- ΔΨm2-cell stage IVF embryos. The development of low-ΔΨm2-cell stage IVF embryos tended to be arrested after the 2-cell stage. These results indicated that the upregulation of ΔΨmduring the 1- to 2-cell stage was important in the development of early preimplantation embryos; there were some defects in the mitochondria of superovulated oocytes, which prevented their development.

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The Usefulness of Retinoic Acid Supplementation during in Vitro Oocyte Maturation for the in Vitro Embryo Production of Livestock: A Review

Animals ◽

10.3390/ani9080561 ◽

2019 ◽

Vol 9 (8) ◽

pp. 561 ◽

Cited By ~ 4

Author(s):

Abdelnour ◽

El-Hack ◽

Swelum ◽

Saadeldin ◽

Noreldin ◽

...

Keyword(s):

Retinoic Acid ◽

Embryo Development ◽

Oocyte Maturation ◽

Mammalian Species ◽

Antioxidant Properties ◽

Reproductive Function ◽

Embryonic Growth ◽

Oxidative Damages

Retinoic acid (RA) is an indigenous metabolite and descriptive physiologically functioning constituent of vitamin A. Retinoids were documented as vital regulators for cell development and distinction, embryonic growth, and reproductive function in both male and female livestock. Previously, RA has been shown to have several positive impacts in vivo and in vitro and critically control many reproductive events, such as oocyte development, follicular growth, and early embryonic growth. In addition, RA manages apoptotic signaling and oxidative damages in cells. Recently, RA has been used widely in assisted reproductive technology fields, especially during in vitro embryo development in various mammalian species, including buffaloes, bovine, goats, sheep, pigs, and rabbits. However, the optimum concentration of RA greatly differs based on the condition of maturation media and species. Based on the obtained findings, it was generally accepted that RA enhances nuclear oocyte maturation, cleavage and maturation rates, blastocyst formation, and embryo development. As such, it possesses antioxidant properties against reactive oxygen species (ROS) and an anti-apoptotic effect through enhancing the transcription of some related genes such as superoxide dismutase, prostaglandin synthase, glutathione peroxidase, peroxiredoxins, and heme oxygenase. Therefore, the current review concludes that an addition of RA (up to 50 nM) has the potential to improve the oocyte maturation media of various species of livestock due to its antioxidant activity.

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