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Southern Distinct Population Segment (sDPS) green sturgeon spawn solely in one stretch of the Sacramento River in California. Management of this spawning habitat is complicated by cold water temperature requirements for the conservation of winter-run Chinook salmon. This study assessed whether low incubation and rearing temperatures resulted in carryover effects across embryo to early juvenile life stages on scaling relationships in growth and metabolism in northern DPS green sturgeon used as a proxy for sDPS green sturgeon. Fish were incubated and reared at 11 °C and 15 °C, with a subset experiencing a reciprocal temperature transfer post-hatch, to assess recovery from cold incubation or to simulate a cold-water dam release which would chill rearing larvae. Growth and metabolic rate of embryos and larvae were measured to 118 days post hatch. Reciprocal temperature transfers revealed a greater effect of low temperature exposure during larval rearing rather than during egg incubation. While 11 °C eggs hatched at a smaller length, log-transformed length-weight relationships showed that these differences in developmental trajectory dissipated as individuals achieved juvenile morphology. However, considerable size-at-age differences persisted between rearing temperatures, with 15 °C fish requiring 60 days post-hatch to achieve 1 g in mass, whereas 11 °C fish required 120 days to achieve 1 g, resulting in fish of the same age at the completion of the experiment with a ca. 37-fold difference in weight. Consequently, our study suggests that cold rearing temperatures have far more consequential downstream effects than cold embryo incubation temperatures. Growth delays from 11 °C rearing temperatures would greatly increase the period of vulnerability to predation in larval green sturgeon. The scaling relationship between log-transformed whole-body metabolism and mass exhibited a steeper slope and thus an increased oxygen requirement with size in 11 °C reared fish, potentially indicating an energetically unsustainable situation. Understanding how cold temperatures affect green sturgeon ontogeny is necessary to refine our larval recruitment estimations for this threatened species.

Xenopus embryos provide a favorable material to dissect the sequential steps that lead to dorsal-ventral (D-V) and anterior-posterior (A-P) cell differentiation. Here, we analyze the signaling pathways involved in this process using loss-of-function and gain-of-function approaches. The initial step was provided by Hwa, a transmembrane protein that robustly activates early β-catenin signaling when microinjected into the ventral side of the embryo leading to complete twinned axes. The following step was the activation of Xenopus Nodal-related growth factors, which could rescue the depletion of β-catenin and were themselves blocked by the extracellular Nodal antagonists Cerberus-Short and Lefty. During gastrulation, the Spemann-Mangold organizer secretes a cocktail of growth factor antagonists, of which the BMP antagonists Chordin and Noggin could rescue simultaneously D-V and A-P tissues in β-catenin-depleted embryos. Surprisingly, this rescue occurred in the absence of any β-catenin transcriptional activity as measured by β-catenin activated Luciferase reporters. The Wnt antagonist Dickkopf (Dkk1) strongly synergized with the early Hwa signal by inhibiting late Wnt signals. Depletion of Sizzled (Szl), an antagonist of the Tolloid chordinase, was epistatic over the Hwa and Dkk1 synergy. BMP4 mRNA injection blocked Hwa-induced ectopic axes, and Dkk1 inhibited BMP signaling late, but not early, during gastrulation. Several unexpected findings were made, e.g., well-patterned complete embryonic axes are induced by Chordin or Nodal in β-catenin knockdown embryos, dorsalization by Lithium chloride (LiCl) is mediated by Nodals, Dkk1 exerts its anteriorizing and dorsalizing effects by regulating late BMP signaling, and the Dkk1 phenotype requires Szl.

Although the effects of genetic and environmental perturbations on multicellular organisms are rarely restricted to single phenotypic layers, our current understanding of how developmental programs react to these challenges remains limited. Here, we have examined the phenotypic consequences of disturbing the bicoid regulatory network in early Drosophila embryos. We generated flies with two extra copies of bicoid, which causes a posterior shift of the networks regulatory outputs and a decrease in fitness. We subjected these flies to EMS mutagenesis, followed by experimental evolution. After only 8-15 generations, experimental populations have normalized patterns of gene expression and increased survival. Using a phenomics approach, we find that populations were normalized through rapid increases in embryo size driven by maternal changes in metabolism and ovariole development. We extend our results to additional populations of flies, demonstrating predictability. Together, our results necessitate a broader view of regulatory network evolution at the systems level.

Neural crest cells exemplify cellular diversification from a multipotent progenitor population. However, the full sequence of early molecular choices orchestrating the emergence of neural crest heterogeneity from the embryonic ectoderm remains elusive. Gene-regulatory-networks (GRN) govern early development and cell specification toward definitive neural crest. Here, we combine ultradense single-cell transcriptomes with machine-learning and large-scale transcriptomic and epigenomic experimental validation of selected trajectories, to provide the general principles and highlight specific features of the GRN underlying neural crest fate diversification from induction to early migration stages using Xenopus frog embryos as a model. During gastrulation, a transient neural border zone state precedes the choice between neural crest and placodes which includes multiple converging gene programs. During neurulation, transcription factor connectome, and bifurcation analyses demonstrate the early emergence of neural crest fates at the neural plate stage, alongside an unbiased multipotent-like lineage persisting until epithelial-mesenchymal transition stage. We also decipher circuits driving cranial and vagal neural crest formation and provide a broadly applicable high-throughput validation strategy for investigating single-cell transcriptomes in vertebrate GRNs in development, evolution, and disease.

Loss of Cdx2 in vivo leads to stunted development of the allantois, an extraembryonic mesoderm-derived structure critical for nutrient delivery and waste removal in the early embryo. Here, we investigate how CDX2 dose-dependently influences the gene regulatory network underlying extraembryonic mesoderm development. By engineering human induced pluripotent stem cells (hiPSCs) consisting of wild-type (WT), heterozygous (CDX2-Het), and homozygous null CDX2 (CDX2-KO) genotypes, differentiating these cells in a 2D gastruloid model, and subjecting these cells to single-nucleus RNA and ATAC sequencing, we identify several pathways that are dose-dependently regulated by CDX2 including VEGF and non-canonical WNT. snATAC-seq reveals that CDX2-Het cells retain a WT-like chromatin accessibility profile, suggesting accessibility alone is not sufficient to drive this variability in gene expression. Because the loss of CDX2 or TBXT phenocopy one another in vivo, we compared differentially expressed genes in our CDX2-KO to those from TBXT-KO hiPSCs differentiated in an analogous experiment. This comparison identifies several communally misregulated genes that are critical for cytoskeletal integrity and tissue permeability. Together, these results clarify how CDX2 dose-dependently regulates gene expression in the extraembryonic mesoderm and reveal pathways that may underlie the defects in vascular development and allantoic elongation seen in vivo.

Without new transcription, gene expression across the oocyte-to-embryo transition (OET) relies instead on regulation of mRNA poly(A) tails to control translation. However, how tail dynamics shape translation across the OET in mammals remains unclear. We perform long-read RNA sequencing to uncover poly(A) tail lengths across the mouse OET and, incorporating published ribosome profiling data, provide an integrated, transcriptome-wide analysis of poly(A) tails and translation across the entire transition. We uncover an extended wave of global deadenylation during fertilization in which short-tailed, oocyte-deposited mRNAs are translationally activated without polyadenylation through resistance to deadenylation. Subsequently, in the embryo, mRNAs are readenylated and translated in a surge of global polyadenylation. We further identify regulation of poly(A) tail length at the isoform level and stage-specific enrichment of mRNA sequence motifs among regulated transcripts. These data provide insight into the stage-specific mechanisms of poly(A) tail regulation that orchestrate gene expression from oocyte to embryo in mammals.

Embryo-maternal crosstalk is essential to establish pregnancy, with the equine embryo moving throughout the uterus on days 9-15 (ovulation = day 0) as part of this interaction. We hypothesized that the presence of a mobile embryo induces local changes in the gene expression of the endometrium. On Day 12, the endometrial transcripts were compared among three groups: uterine horn with an embryo (P+, n = 7), without an embryo (P-, n = 7) in pregnant mares, and both uterine horns of nonbred mares (NB, n = 6). We identified 1,101 differentially expressed genes (DEGs) between P+ vs. NB and 1,229 DEGs between P- vs. NB. The genes upregulated in both P+ and P- relative to NB were involved in growth factor pathway and fatty acid activation, while downregulated genes were associated with oxytocin signaling pathway and estrogen receptor signaling. Comparing the transcriptome of P+ to that of P-, we found 59 DEGs, of which 30 genes had a higher expression in P+. These genes are associated with regulating vascular growth factors and the immune system, all known to be essential in early pregnancy. Overall, this study suggests that the mobile embryo influences the endometrial gene expression locally.

Non-invasively evaluating gene expression products in human pre-implantation embryos remains a significant challenge. Here, we develop a non-invasive method for comprehensive characterization of the extracellular RNAs (exRNAs) in a single droplet of spent media that was used to culture human in vitro fertilization embryos. We generate the temporal extracellular transcriptome atlas (TETA) of human pre-implantation development. TETA consists of 245 exRNA sequencing datasets for five developmental stages. These data reveal approximately 4,000 exRNAs at each stage. The exRNAs of the developmentally arrested embryos are enriched with the genes involved in negative regulation of the cell cycle, revealing an exRNA signature of developmental arrest. Furthermore, a machine-learning model can approximate the morphology-based rating of embryo quality based on the exRNA levels. These data reveal the widespread presence of coding gene-derived exRNAs at every stage of human pre-implantation development, and these exRNAs provide rich information on the physiology of the embryo.

Introduction:Urtica simensis has been used to treat various diseases such as malaria, hypertension, diabetes, gonorrhea, gastritis, body swelling, and wound infections. However, the safety of consuming U. simensis leaves during pregnancy has not been evaluated yet. Therefore, this experimental study was conducted to evaluate the toxic effects of U. simensis leaf extract on the prenatal development of embryos and fetuses in pregnant rats. Methods: Fifty pregnant Wistar albino rats were randomly assigned to five groups of 10 gravid rats for each experiment. Groups I–III were given 70% ethanol leaf extract of U. simensis at doses of 250, 500, and 1000 mg/kg daily from 6th to 12th days of gestation. Groups IV–V were kept as pair‐fed and ad libitum controls. The developing embryos and fetuses were retrieved on 12 days and 20 days of gestation, respectively. Embryos were evaluated for growth and developmental delays. Fetuses were also assessed for growth retardation and external and visceral anomalies. Results: In the embryonic experiment, somite numbers (p = 0.001) and morphological scores (p = 0.029) were significantly decreased in pregnant rats given 1000 mg/kg of U. simensis leaf extract. Embryonic developments of the caudal neural tube (CNT) (p = 0.001), otic system (p = 0.025), olfactory system (p = 0.013), and limb buds (p = 0.026) were significantly delayed in pregnant rats given 1000 mg/kg of extract. Oral administration of 500 mg/kg of U. simensis leaf extract also caused significant developmental delays in the CNT (p = 0.021) and olfactory system (p = 0.032). In the fetal experiment, fetal resorption (p = 0.015) was significantly increased whereas crown rump length (p = 0.012) and fetal weight (p = 0.019) were significantly decreased in pregnant rats given 1000 mg/kg of U. simensis leaf extract. Conclusions: The embryotoxic effects of U. simensis leaf extract were evidenced by significant developmental delays. The fetal toxic effects of U. simensis leaf extract were also shown by significant decreases in fetal growth indices. Therefore, pregnant women should be well informed of the possible toxic effects of consuming U. simensis leaf during pregnancy. [ABSTRACT FROM AUTHOR]

Summary Accumulation of triacylglycerols (TAGs) is crucial during various stages of plant development. In Arabidopsis, two enzymes share overlapping functions to produce TAGs, namely acyl‐CoA:diacylglycerol acyltransferase 1 (DGAT1) and phospholipid:diacylglycerol acyltransferase 1 (PDAT1). Loss of function of both genes in a dgat1‐1/pdat1‐2 double mutant is gametophyte lethal. However, the key regulatory elements controlling tissue‐specific expression of either gene has not yet been identified. We transformed a dgat1‐1/dgat1‐1//PDAT1/pdat1‐2 parent with transgenic constructs containing the Arabidopsis DGAT1 promoter fused to the AtDGAT1 open reading frame either with or without the first intron. Triple homozygous plants were obtained, however, in the absence of the DGAT1 first intron anthers fail to fill with pollen, seed yield is c. 10% of wild‐type, seed oil content remains reduced (similar to dgat1‐1/dgat1‐1), and non‐Mendelian segregation of the PDAT1/pdat1‐2 locus occurs. Whereas plants expressing the AtDGAT1pro:AtDGAT1 transgene containing the first intron mostly recover phenotypes to wild‐type. This study establishes that a combination of the promoter and first intron of AtDGAT1 provides the proper context for temporal and tissue‐specific expression of AtDGAT1 in pollen. Furthermore, we discuss possible mechanisms of intron mediated regulation and how regulatory elements can be used as genetic tools to functionally replace TAG biosynthetic enzymes in Arabidopsis. [ABSTRACT FROM AUTHOR]

Background The evolutionary success of flowering plants is associated with the vast diversity of their reproductive structures. Despite recent progress in understanding angiosperm-wide trends in floral structure and evolution, a synthetic view of the diversity in seed form and function across angiosperms is lacking. Scope Here we present a roadmap to synthesize the diversity of seed forms in extant angiosperms, relying on the morphospace concept, i.e. a mathematical representation which relates multiple traits and describes the realized morphologies. We provide recommendations on how to broaden the range of measurable traits beyond mass, by using key morphological traits representative of the embryo, endosperm and seed coat but also fruit attributes (e.g. dehiscence, fleshiness). These key traits were used to construct and analyse a morphospace to detect evolutionary trends and gain insight into how morphological traits relate to seed functions. Finally, we outline challenges and future research directions, combining the morphospace with macroevolutionary comparative methods to underline the drivers that gave rise to the diversity of observed seed forms. Conclusions We conclude that this multidimensional approach has the potential, although still untapped, to improve our understanding of covariation among reproductive traits, and further elucidate angiosperm reproductive biology as a whole. [ABSTRACT FROM AUTHOR]

The permeation of cryoprotectants into insect embryos is critical for successful cryopreservation. However, the permeability of silkworm embryos to cryoprotectants and the effects of cryopreservation remain poorly studied. In this study, we evaluated the permeability and toxicity of four cryoprotective agents (CPAs) as well as the vitrification effect on the viability of silkworm embryos. Among the four CPAs, propylene glycol (PG) showed the best permeability. Ethylene glycol (EG) and PG were the least toxic CPAs, but glycerol (GLY) and dimethyl sulfoxide (DMSO) were more toxic. Moreover, we examined several factors including the kind and the concentration of CPAs, exposure time, embryonic stage, and silkworm strains. Embryos at the earlier phases of stage 25 were more tolerant to vitrification using EG. We found that over 21% of embryos treated with EG at the early 2 phase of stage 25: 163 h after egg laying (AEL) developed and progressed to serosa ingestion after vitrification and rewarming. The result was the same in other strains as well. Our results are valuable for the development of new cryopreservation protocols of silkworm embryos. [ABSTRACT FROM AUTHOR]

Background: Like the coronavirus disease 2019, the hepatitis B virus is also wreaking havoc worldwide, which has infected over 2 billion people globally. Using an experimental animal model, our previous research observed that the hepatitis B virus genes integrated into human spermatozoa can replicate and express after being transmitted to embryos. However, as of now, this phenomenon has not been confirmed in clinical data from patients. Objectives: To explore the integration of the hepatitis B virus into patients' sperm genome and its potential clinical risks. Materials and methods: Forty‐eight patients with chronic hepatitis B virus infection were categorized into two groups: Test Group‐1 comprised 23 patients without integration of hepatitis B virus DNA within the sperm genome. Test Group‐2 comprised 25 patients with integration of hepatitis B virus DNA within the sperm genome. Forty‐eight healthy male donors were included as control. The standard semen parameter analysis, real‐time polymerase chain reaction, quantitative real‐time polymerase chain reaction, sperm chromatin structure assay, fluorescence in situ hybridization, and immunofluorescence assays were utilized. Results: The difference in the median copy number of hepatitis B virus DNA per mL of sera between Test Group‐1 and Group‐2 was not statistically significant. In Test Group‐2, the integration rate of hepatitis B virus DNA was 0.109%, which showed a significant correlation with the median copy number of hepatitis B virus DNA in motile spermatozoa (1.18 × 103/mL). Abnormal semen parameters were found in almost all these 25 patients. The integrated hepatitis B virus S, C, X, and P genes were detected to be introduced into sperm‐derived embryos through fertilization and retained their function in replication, transcription, and translation. Conclusion: Our findings suggest that hepatitis B virus infection can lead to sperm quality deterioration and reduced fertilization capacity. Furthermore, viral integration causes instability in the sperm genome, increasing the potential risk of termination, miscarriage, and stillbirth. This study identified an unconventional mode of hepatitis B virus transmission through genes rather than virions. The presence of viral sequences in the embryonic genome poses a risk of liver inflammation and cancer. [ABSTRACT FROM AUTHOR]

STUDY QUESTION Can novel genetic factors contributing to early embryonic arrest in infertile patients be identified, along with the underlying mechanisms of the pathogenic variant? SUMMARY ANSWER We identified a heterozygous variant in the SPRY4 (sprouty RTK signaling antagonist 4) in infertile patients and conducted in vitro and in vivo studies to investigate the effects of the variant/deletion, highlighting its critical role in female reproductive health. WHAT IS KNOWN ALREADY SPRY4 acts as a negative regulator of receptor tyrosine kinases (RTKs) and functions as a tumor suppressor. Its abnormal expression can lead to recurrent miscarriage by affecting trophoblast function. In mice, Spry4 knockout (KO) leads to craniofacial anomalies and growth defects. A human study links the SPRY4 variant to a male patient with isolated hypogonadotropic hypogonadism (IHH), hypothetically impacting gonadotropin-releasing hormone (GnRH) neurons, and causing reproductive dysfunctions. SPRY4 is thus potentially integral in regulating endocrine homeostasis and reproductive function. To date, no study has reported SPRY4 variants associated with female fertility, and a causal relationship has not been established with functional evidence. STUDY DESIGN, SIZE, DURATION Whole-exome sequencing (WES) was performed in 392 infertile women who suffered from primary infertility of unknown reason, and the heterozygous SPRY4 variant were identified in one independent family. The infertile patients presenting were recruited from July 2017 to November 2023. PARTICIPANTS/MATERIALS, SETTING, METHODS Women diagnosed with primary infertility were recruited from the Reproduction Center of Zhongshan Hospital, Fudan University. Genomic DNA was extracted from peripheral blood for WES analysis. The SPRY4 variant were identified through WES, in silico analysis, and variant screening. All variants were confirmed by Sanger sequencing. The effects of the variants were investigated in human embryonic kidney (HEK) 293T (HEK293T) cells via western blotting, and in mouse oocytes and embryos through complementary RNA (cRNA) injection, RNA sequencing, fluorescence, absorbance, and RT-qPCR assays. Gene function was further examined in Spry4 KO mice via histology, western blotting, ELISA, and RT-qPCR assays. MAIN RESULTS AND THE ROLE OF CHANCE We identified a missense heterozygous pathogenic variant in SPRY4 (GRCh38, GenBank: NM_030964.5 , c.157C>T p.(Arg53Trp), rs200531302) that reduces SPRY4 protein levels in HEK293T cells and disrupts the redox system and mitochondrial function in mouse oocyte, and perturbs developmental potential in mouse embryos. These phenotypes could be partially reversed by the exogenous addition of Nrf1 cRNA. Additionally, Spry4−/ − mice exhibit ovarian oxidative stress and decreased ovarian function. LIMITATIONS, REASONS FOR CAUTION Due to the limited WES data and population, we identified only one family with a SPRY4 mutation. The deeper mechanism and therapeutic strategy should be further investigated through mutant mice and recovery experiment. WIDER IMPLICATIONS OF THE FINDINGS Our study has identified a pathogenic variant in SPRY4 associated with early embryonic arrest in humans. These findings enhance our understanding of the role of SPRY4 in early embryonic development and present a new genetic marker for female infertility. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by the National Natural Science Foundation of China (82071643 and 82171655) and Natural Science Foundation of Shanghai (22ZR1456200). None of the authors have any competing interests. TRIAL REGISTRATION NUMBER N/A [ABSTRACT FROM AUTHOR]

STUDY QUESTION Is the degree of maternal vulnerability positively associated with stress biomarkers (stress hormones, C-reactive protein, tryptophan metabolites, and one-carbon metabolites), and does long-term exposure to stress hormones reduce first-trimester growth? SUMMARY ANSWER The maternal vulnerability risk score is positively associated with concentrations of hair cortisol and cortisone and negatively with tryptophan, while higher hair cortisol concentrations are associated with reduced first-trimester growth without mediation of tryptophan. WHAT IS KNOWN ALREADY A high degree of maternal vulnerability during the periconception period is associated with impaired first-trimester growth and pregnancy complications, with consequences for long-term health of the child and future life course. However, due to the challenges of early identification of vulnerable women, the uptake of periconception care is low in this target group. STUDY DESIGN, SIZE, DURATION Between June 2022 and June 2023, this study was conducted in a sub-cohort of 160 pregnant women participating in the Rotterdam Periconceptional Cohort (Predict Study), an ongoing prospective tertiary hospital-based cohort. PARTICIPANTS/MATERIALS, SETTING, METHODS One hundred and thirty-two women with ongoing pregnancies and available stress biomarker data were included in the analysis. Data on periconceptional social, lifestyle, and medical risk factors were collected via self-administered questionnaires, and these factors were used for the development of a composite maternal vulnerability risk score. Stress biomarkers, including stress hormones (hair cortisol and cortisone) and inflammatory and oxidative stress biomarkers (C-reactive protein, total homocysteine, and tryptophan metabolites) were determined in the first trimester of pregnancy. First-trimester growth was assessed by crown–rump length (CRL) and embryonic volume (EV) measurements at 7, 9, and 11 weeks gestation by making use of an artificial intelligence algorithm and virtual reality techniques using 3D ultrasound data sets. The associations between the maternal vulnerability risk score and stress biomarkers were identified using linear regression models, and between stress hormones and CRL- and EV-trajectories using mixed models. A mediation analysis was performed to assess the contribution of tryptophan. All associations were adjusted for potential confounders, which were identified using a data-driven approach. Several sensitivity analyses were performed to check the robustness of the findings. MAIN RESULTS AND THE ROLE OF CHANCE The maternal vulnerability risk score was positively associated with concentrations of hair cortisol and cortisone (pg/mg) (β = 0.366, 95% CI = 0.010–0.722; β = 0.897, 95% CI = 0.102–1.691, respectively), and negatively with tryptophan concentrations (µmol/L) (β = –1.637, 95% CI = –2.693 to –0.582). No associations revealed for C-reactive protein and total homocysteine. Higher hair cortisol concentrations were associated with reduced EV-trajectories (3√EV: β = –0.010, 95% CI = –0.017 to –0.002), while no associations were found with CRL-trajectories. Mediation by tryptophan was not shown. LIMITATIONS, REASONS FOR CAUTION Residual confounding cannot be ruled out, and the external validity may be limited due to the study's single-center observational design in a tertiary hospital. WIDER IMPLICATIONS OF THE FINDINGS There is mounting evidence that a high degree of maternal vulnerability negatively affects maternal and perinatal health, and that of the future life course. The results of our study emphasize the need to identify highly vulnerable women as early as possible, at least before conception. Our findings suggest that the chronic stress response and alterations of the maternal tryptophan metabolism are involved in maternal vulnerability, affecting first-trimester growth, with potential impact on the long-term health of the offspring. STUDY FUNDING/COMPETING INTEREST(S) This study was funded by the Departments of Obstetrics and Gynecology and Clinical Chemistry of the Erasmus MC, University Medical Center, Rotterdam, the Netherlands, and the Junior Award granted by the De Snoo—van 't Hoogerhuijs Foundation in March 2022. There are no conflicts of interest. TRIAL REGISTRATION NUMBER N/A. [ABSTRACT FROM AUTHOR]

Cell cycle regulation is crucial to assure expansion of a cell population, while preserving genome integrity. This notion is especially relevant to fertilization and early embryo development, a time when the cell cycle transforms from meiotic into mitotic cycles. Zygote-to-embryo transition is acutely error-prone, causing major developmental perturbations, including cleavage delays, tri- and multi-chotomous cleavages, and cell fragmentation. Another such alteration is bi- and multinucleation, consisting of the simultaneous formation of two or more nuclei at interphase. Indeed, multinucleation affects a large proportion of early human embryos, typically at the two-cell stage. Mechanistically, several factors, including spindle dysfunction, failed cleavage, and cell fusion, may generate this cell anomaly. In assisted reproduction treatment, multinucleation is associated with reduced developmental rates and lower implantation rates in Days 2–3 embryo transfers. However, many multinucleated embryos can develop to the blastocyst stage. In blastocyst transfers, the current evidence does not suggest a major impact of a previous history of multinucleation on the odds of euploidy or successful treatment outcomes. Human embryo multinucleation remains a not-fully-understood but developmentally relevant and intriguing phenomenon which requires further research of its generative mechanisms and clinical implications. [ABSTRACT FROM AUTHOR]

The toxicity of methylmercury (MeHg) during embryonic development is a relevant issue that remains unclear and deserves investigation. In this sense, there is evidence that links the intake of contaminated food with cardiovascular pathologies in human adults and children. Thus, this study aimed to verify the impact of MeHg on the structure and integrity of extraembryonic and cardiac blood vessels and the contractile function of cardiomyocytes, also evaluating embryonic weight and the cardiosomatic index (CSI). Thus, chicken embryos, used as an experimental model, were exposed to a single dose of 0.1 μg MeHg/50 μl saline at E1.5 and analyzed at E10. After exposure, an increase in the number of extraembryonic blood vessels and the veins of the cardiac tissue was observed. These increases were accompanied by a reduction in the content of VEGF and VCAM proteins related to vessel growth and adhesiveness. Together, these results were related to reduced nitrite (NOx) levels. Furthermore, MeHg reduces the number of sarcomeres and increases the content of cardiac troponin I (cTnI), a protein that regulates contraction. In general, exposure to MeHg affected the integrity of extraembryonic and cardiac vessels and the contractile function of cardiomyocytes, which had a systemic impact evidenced by the reduction in embryonic weight gain and CSI. Ingestion of MeHg through contaminated food is associated with cardiovascular problems. This study assessed MeHg effects on extraembryonic and cardiac blood vessels, cardiomyocyte function, embryonic weight, and cardiosomatic index (CSI). Gallus domesticus embryos received a single dose 0.1 μg MeHg/50 μl saline at E1.5, analyzed at E10. MeHg exposure increased extraembryonic and cardiac vessel numbers, reduced VEGF and VCAM proteins, and lowered NOx levels. MeHg also decreased sarcomeres and increased cTnI, impacting cardiomyocyte function, embryonic weight, and CSI. [ABSTRACT FROM AUTHOR]

An understanding of the dynamics of bone growth is key to interpreting life‐history parameters of vertebrates. In this study, we used fluorochrome labels in captive leopard geckos (Eublepharis macularius) to track bone growth and intraskeletal variability from embryonic to adult growth stages. Thirteen individuals were administered fluorochromes from pre‐hatching to 4 years of age. The left tibia, fibula, femur, humerus, radius, and ulna were examined histologically and compared for differences in the number of labels within and between individuals at each sampled growth stage, and the amount of bone growth between labels was calculated. Results suggest that limb elements had differing growth rates; the fibula grew the fastest per day on average and the femur grew the slowest per day on average. All labels administered in ovo were still present in all limb elements in adults except for the tibia, suggesting growth marks are not lost in most elements and accurate calculations of growth rates could be performed in individuals up to 3 years old. All ex ovo labels were accounted for; however, when two fluorochromes were administered 3 weeks apart, the labels could not be differentiated from each other due to the new bone not being deposited at a quantifiable level. Overall, the tibia in leopard geckos is the least reliable limb bone to use for skeletochronology and the humerus, radius, and fibula preserve the longest growth record. This research highlights that, as in other extinct and extant animals, patterns of bone growth are not consistent across reptiles. This study adds to the growing body of knowledge on growth variability in reptiles. [ABSTRACT FROM AUTHOR]

SUMMARY Somatic embryogenesis (SE) is a powerful model system for studying embryo development and an important method for scaling up availability of elite and climate‐adapted genetic material of Norway spruce (Picea abies L. Karst). However, there are several steps during the development of the somatic embryo (Sem) that are suboptimal compared to zygotic embryo (Zem) development. These differences are poorly understood and result in substantial yield losses during plant production, which limits cost‐effective large‐scale production of SE plants. This study presents a comprehensive data resource profiling gene expression during zygotic and somatic embryo development to support studies aiming to advance understanding of gene regulatory programmes controlling embryo development. Transcriptome expression patterns were analysed during zygotic embryogenesis (ZE) in Norway spruce, including separated samples of the female gametophytes and Zem, and at multiple stages during SE. Expression data from eight developmental stages of SE, starting with pro‐embryogenic masses (PEMs) up until germination, revealed extensive modulation of the transcriptome between the early and mid‐stage maturing embryos and at the transition of desiccated embryos to germination. Comparative analysis of gene expression changes during ZE and SE identified differences in the pattern of gene expression changes and functional enrichment of these provided insight into the associated biological processes. Orthologs of transcription factors known to regulate embryo development in angiosperms were differentially regulated during Zem and Sem development and in the different zygotic embryo tissues, providing clues to the differences in development observed between Zem and Sem. This resource represents the most comprehensive dataset available for exploring embryo development in conifers. [ABSTRACT FROM AUTHOR]

Background: Cockayne syndrome (CS) is a rare, multisystem, autosomal recessive disorder characterized by cachectic dwarfism, nervous system abnormalities, and premature aging. Mutations in the ERCC6 and ERCC8 genes are the predominant causes of Cockayne syndrome, with ERCC6 gene mutations present in approximately 75% of cases. Methods: Trio-based whole-exome sequencing (trio-WES) was employed to identify potential pathogenic variants associated with CS. Preimplantation genetic testing for monogenic disorders (PGT-M) was conducted to prevent the transmission of the pathogenic variant. Results: Two compound heterozygous mutations were identified in ERCC6--c.1297G>T (p. Glu433*) and c.1607T>G (p. Leu536Trp)--with c.1297G>T representing a novel mutation. Four blastocysts resulting from intracytoplasmic sperm injection were subjected to biopsy. Genetic analyses revealed that E1 harbored maternal mutations in diploid embryos, E2 and E3 carried both paternal and maternal mutations in non-diploid embryos, and E4 did not carry paternal or maternal mutations in diploid embryos. Following the transfer of the E4 embryos, a single successful pregnancy was achieved. Conclusion: The successful application of PGT-M in this family offers a potential approach for addressing other monogenic diseases. The findings of this study broaden the variant spectrum of ERCC6 and will contribute to the molecular diagnosis and genetic counseling of CS. This case highlights the feasibility and effectiveness of PGT-M in preventing CS and provides valuable insights for similarly affected families. [ABSTRACT FROM AUTHOR]

AbstractZinc oxide quantum dots, also known as ZnO QDs, are highly desirable due to their numerous favorable characteristics, such as their beneficial photoluminescence, solubility in water, along with sunlight absorption. They are well-suited for use in biomedical applications, drugs, and bioimaging. However, study on the in-vivo toxicology of these QDs is needed before they can be used in humans. Zebrafish (Danio rerio) are cheap, fast-growing, and similar to humans, which makes them ideal as in vivo model for studying the toxicity of nanomaterials. The toxicity investigations involving zinc oxide QDs (ZnO QDs) and zinc oxide bionanocomposite (ZnO BC) in zebrafish that were concentration-dependent are evaluated, and the Box-Behnken design (BBD) was utilized to optimize the results. To determine the proper dosage, a study on cell line as well as hemocompatibility was carried out prior to testing the toxic effects of ZnO QDs along with ZnO BC upon zebrafish. When administered at 2.5 μg/l of ZnO BC and 2 μg/l of ZnO QDs, neither ZnO BC nor ZnO QDs appeared to be toxic to embryos during hatching and development. The testing of larval behavior in visible light revealed a dose-dependent decrease in both the total diving distance as well as speed. Nevertheless, at ZnO BC and ZnO QDs levels >250 μg/l and >200 μg/l, respectively, notable effects were seen in zebrafish embryos. Hence, ZnO QDs and BC at low concentrations were notably nontoxic. In order to guarantee the safety of nanomaterials in bio applications, this research supports upcoming in-vivo imaging investigations on their harmful effects. [ABSTRACT FROM AUTHOR]

Background: In vitro embryo production is increasingly used for genetic improvement in cattle but bypasses the oviduct environment and exposes the embryos to oxidative stress with deleterious effects on further development. Here we aimed to examine the effect of oviduct epithelial spheroids (OES) on embryo development and quality in terms of morphology and gene expression during two co-culture times (4 days: up to embryonic genome activation at 8–16 cell stage vs. 7 days: up to blastocyst stage) and under two oxygen levels (5% vs. 20%). Methods: Bovine presumptive zygotes produced by in vitro fertilization (day 0) using in-vitro matured oocytes were cultured in droplets of synthetic oviductal fluid (SOF) medium with or without (controls) OES for 4 or 7 days under 5% or 20% oxygen (4 treated and 2 control groups). Cleavage rates were evaluated on day 2 and blastocyst rates on days 7–8. Expanded blastocysts on days 7–8 were evaluated for total cell numbers and gene expression analysis by RNA-sequencing. Results: Under 20% oxygen, blastocyst rates and total cell numbers were significantly higher in the presence of OES for 4 and 7 days compared to controls (P < 0.05), with no difference according to the co-culture time. Under 5% oxygen, the presence of OES did not affect blastocyst rates but increased the number of cells per blastocyst after 7 days of co-culture (P < 0.05). Both oxygen level and OES co-culture had a significant impact on the embryonic transcriptome. The highest number of differentially expressed genes (DEGs) was identified after 7 days of co-culture under 20% oxygen. DEGs were involved in a wide range of functions, including lipid metabolism, membrane organization, response to external signals, early embryo development, and transport of small molecules among the most significantly impacted. Conclusion: OES had beneficial effects on embryo development and quality under both 5% and 20% oxygen, mitigating oxidative stress. Stronger effects on embryo quality and transcriptome were obtained after 7 than 4 days of co-culture. This study shows the impact of OES on embryo development and reveals potential molecular targets of OES-embryo dialog involved in response to stress and early embryonic development. [ABSTRACT FROM AUTHOR]

The initial contraction of the heart during the embryonic stage necessitates a substantial energy supply, predominantly derived from mitochondrial function. However, during embryonic heart development, mitochondria influence beyond energy supplementation. Increasing evidence suggests that mitochondrial permeability transition pore opening and closing, mitochondrial fusion and fission, mitophagy, reactive oxygen species production, apoptosis regulation, Ca2+ homeostasis, and cellular redox state also play critical roles in early cardiac development. Therefore, this review aims to describe the essential roles of mitochondrial non-energetic function embryonic cardiac development. [ABSTRACT FROM AUTHOR]

Aim Design Results Conclusions To compare clinical outcomes using short and long co‐incubation protocols in sibling oocytes based on embryo morphokinetic outcomes measured by time‐lapse incubator with stratification based on a woman's age and sperm quality.Our study included 72 cycles with >6 oocytes retrieved. Sibling oocytes were distributed for two parallel protocols: short (3 h; n = 421) or long (16–20 h; n = 434) insemination, using the same amount of spermatozoa from the same prepared sample. Oocytes were then washed and incubated for 5 days. Time‐lapse annotations of embryos were performed by experienced embryologists and artificial intelligence‐based Known Implantation Data scores for day 3 and day 5 were calculated with EmbryoScope software.Short‐insemination group exhibited a higher blastulation rate, better morphokinetic indicators, and higher Known Implantation Data scores on day 3 and day 5 of the utilized embryos. However, the fertilization rate and clinical pregnancy rate per embryo transfer did not differ between experimental groups. A higher rate of abnormal fertilization (>2 pronuclei) after long insemination was recorded in women under 35 years old or with a total motile sperm count above 5 million and above 40% motility after preparation. A higher rate of usable embryos was observed after short insemination with a total motile sperm count above 30 million before preparation or 5 million and over 40% motility after preparation.Our results suggest that a short insemination protocol results in better embryo quality and should be considered as a favorable protocol, especially in young female patients or male patients with high sperm quality. [ABSTRACT FROM AUTHOR]

The lipidomic secretions of embryos provide a unique opportunity to examine the cellular processes of the early conceptus. In this study we profiled lipids released by the early equine conceptus, using high-resolution mass spectrometry to detect individual lipid species. This study examined the lipidomic profile in embryo-conditioned media from in vivo-produced, 8-9 day-old equine embryos (n = 3) cultured in vitro for 36 h, analyzed over 3 timepoints. A total of 1,077 lipid IDs were recorded across all samples, containing predominantly glycerolipids. Seventy-nine of these were significantly altered in embryo conditioned-media versus media only control (p < 0.05, fold-change >2 or < 0.5). Fifty-five lipids were found to be released into the embryo-conditioned media, of which 54.5% were triacylglycerols and 23.6% were ceramides. The sterol lipid, cholesterol, was also identified and secreted in significant amounts as embryos developed. Further, 24 lipids were found to be depleted from the media during culture, of which 70.8% were diacylglycerols, 16.7% were triacylglycerols and 12.5% were ceramides. As lipid-free media contained consistently detectable lipid peaks, a further profile analysis of the various components of non-embryo-conditioned media consistently showed the presence of 137 lipids. Lipid peaks in non-embryo-conditioned media increased in response to incubation under mineral oil, and contained ceramides, diacylglycerols and triacylglycerols. These results emphasize the importance of a defined embryo culture medium and a need to identify the lipid requirements of the embryo precisely. This study sheds light on early embryo lipid metabolism and the transfer of lipids during in vitro culture. [ABSTRACT FROM AUTHOR]

After cryopreservation, reactive oxygen species (ROS) can damage sperm. Antioxidants are the primary defense against oxidative damage. Berberine is a bioactive alkaloid found in Berberis vulgaris, Curcuma longa, and Ergon grape, and is a potent antioxidant. Due to the negative effects of free radicals in oxidative stress processes, antioxidant chemicals are required to protect sperm. However, berberine has low bioavailability, making it less effective. Loading techniques on nanoparticles and nanotechnology can help overcome this limitation. Selenium nanoparticles were synthesized with barberry extract, and berberine was loaded on them. Berberine nanoparticles were then synthesized using anti-solvent precipitation with a syringe pump technique. The synthesis of nanoparticles was confirmed by EDX, UV-visible, FE-SEM, Zeta-Potential, and FTIR tests. In this experiment, we aim to investigate the impact of nano-berberine and berberine loaded on Se-NPs on goat sperm parameters after freeze-thawing. We assessed the generation of reactive oxygen species (ROS), in vitro fertility, and the subsequent embryo development of zygote with treated sperm after determining the optimal concentration of various chemicals on sperm parameters. The study found that all treatments had significant differences from the control group in terms of motility, viability, DNA and membrane integrity, ROS level, lipid peroxidation, in vitro fertility ability, and the capacity to develop inseminated oocytes (p < 0.05). The most significant outcomes were observed with berberine loaded on Se-NPs and the combination of selenium nanoparticles with berberine nanoparticles. [ABSTRACT FROM AUTHOR]

Mustard is a commercial oilseed crop worldwide infected by a highly infectious turnip mosaic virus (TuMV). In the experimental field at ICAR-IARI, New Delhi, in 2022, a 100% incidence of TuMV infection was observed in brown, black and yellow mustard. A very low aphid population suggested the possibility of seed transmission. Earlier, the virus genome was characterized by high throughput sequencing and it was a recombinant of World-B and Asian-BR isolates. The presence of TuMV in immature seeds was confirmed in eight field-grown genotypes via RT-PCR using CP-specific primers designed from the same genome sequence. TuMV was found to be localized in embryo and cotyledon, indicating its true seed-borne nature. Presence of TuMV was also confirmed by RT-PCR in the grow out plants from seeds of field grown eight infected genotypes and 9 genotypes collected from seed stock, that were grown in an aphid-free growth chamber. Further, out of 24 seedlings of Pusa Gold (seed stock) and Pusa Karishma (seeds from field grown plants), 20 and 17 seedlings were found infected with TuMV, respectively. The internally seed-borne nature of the virus leads to its early establishment at the seedling stage, leading to stunting and leaf-puckering symptoms in the progeny plants. This study is the first evidence of seed embryo infection and seedling transmission of TuMV of all the three species of mustard plants (brown, black and yellow mustard). Seed transmission of TuMV in mustard genotypes have implications for the seed exchange programme of mustard seeds. [ABSTRACT FROM AUTHOR]

As interest in deep space travel grows exponentially, understanding human adaptation in becoming an interplanetary species is crucial. This includes the prospect of reproduction. This review summarizes recent updates and innovations in assisted reproductive technologies (ART) on Earth, while also discussing current challenges and areas for improvement in adapting ART studies to the space environment. We discuss the critical components of ART - gamete handling and preparation, fertilization, embryo culture, and cryopreservation - from the daily practice perspective of clinical embryologists and reproductive endocrinologists and lay out the complicated path ahead. In vitro embryo development in low Earth orbit and beyond remains questionable due to synergetic effects of microgravity and radiation-induced damage observed in simulated and actual in-space mammalian studies. Cryopreservation and long-term storage of frozen samples face substantial obstacles - temperature limitations, lack of trained personnel, and absence of adapted cosmic engineering options. We touch on recent innovations, which may offer potential solutions, such as microfluidic devices and automated systems. Lastly, we stress the necessity for intensive studies and the importance of an interdisciplinary approach to address numerous practical challenges in advancing reproductive medicine in space, with possible implications for both space exploration and terrestrial fertility treatments. [ABSTRACT FROM AUTHOR]

Lipids constitute one of the most abundant components within cells and play a fundamental role as an energy source during embryonic development, as well as in many physiological processes. Understanding their composition is crucial to understand the sequence of events required for successful embryogenesis. This study aimed to assess the variation in the content and composition of lipids and their role during the embryonic developmental stages of the coral Galaxea fascicularis. The major lipid classes, including sterol ester (SE), wax ester (WE), triacylglycerol (TAG), cholesterol (CHO), phosphatidylethanolamine (PE), phosphatidylcholine (PC), lysophosphatidylcholine, and fatty acids, were evaluated. The gametes and embryos were homogenized using a combination of chloroform and methanol, resulting in a two-phase system where the lipids were found in the chloroform layer. The lipid and fatty acid content was determined using thin layer chromatography, transesterification to fatty acid methyl ester, and gas chromatography mass spectrometry. The findings indicated that the content of SE, WE, and TAG changed across several stages of development. CHO and PC were the most prevalent lipids in all stages, while the levels of TAG, PE, and SE were considerably lower compared to the aforementioned lipid classes. Unlike the lipid profile of the oocyte, the sperm had a significantly smaller amount of WE and did not include any SE. However, there were no significant differences in the quantity of the other lipid classes. Regarding the fatty acid composition, there was a notable increase in concentration throughout the early stages of embryonic development, primarily consisting of saturated fatty acids. Conversely, the amount of fatty acids in sperm was relatively low. The study demonstrated that the lipid content and composition can vary during different phases of embryonic development. This discovery will surely contribute to future lipid studies in other species of coral. [ABSTRACT FROM AUTHOR]

Rabbits have played a significant role in both livestock production and the advancement of reproductive scientific research. Their unique biological traits, including induced ovulation and a reproductive process that closely mirrors that of humans, have been pivotal in their use as a model. Moreover, their body size is perfectly aligned with the 3Rs principles: Replacement, Reduction, and Refinement. Consequently, techniques for gamete collection and embryo recovery, followed by their use in artificial insemination or embryo transfer, are characterized by being minimally invasive. However, refining in vitro fertilization and embryo culture techniques continues to present challenges. The incorporation of cutting‐edge genomic editing tools, such as CRISPR/Cas9, has reestablished rabbits as essential models in genetic and biomedical research, driving scientific progress. This review aims to describe the most effective reproductive biotechnologies for both male and female rabbits and how these methodologies are in line with the 3Rs principles—Replacement, Reduction, and Refinement—highlighting their significance in conducting ethical research. [ABSTRACT FROM AUTHOR]

Summary Fungal endometritis and placentitis have been reported as a rare but important cause of infertility in the mare. There are no reported clinical cases of fungal endometritis causing embryonic loss in the mare. This case report aims to highlight the potential of fungal endometritis as an aetiological agent in embryonic loss in the mare; an incidental diagnosis of a case of fungal endometritis and embryonic loss while undertaking routine pregnancy assessments in a Standardbred mare. After a positive pregnancy diagnosis after 13 days post‐ovulation, a mare was diagnosed with endometritis and pregnancy loss associated with the growth of an unusual fungus, likely of the Acremonium spp., 28 days post‐ovulation. While fungal embryonic loss is rare, it should be considered a potential reason for pregnancy wastage in the mare. [ABSTRACT FROM AUTHOR]

Introduction: The posterior meniscofemoral ligament (pMFL) of knee joint is a ligament that runs posterior to the posterior cruciate ligament and it is known that the height of the pMFL attachment site causes meniscus avulsion. Therefore, understanding the three-dimensional (3D) structure of the pMFL attachment site is essential to better understand the pathogenesis of meniscus disorders. However, the developmental process of pMFL has not been well investigated. The purpose of this study was to analyze pMFL development in rat knee joints using 3D reconstructed images produced from episcopic fluorescence image capture (EFIC) images and examine its relationship with other knee joint components. Methods: Knee joints of Wistar rat embryos between embryonic day (E) 16 and E21 were observed with HE-stained tissues. Serial EFIC images of the hind limbs of E17–E21 were, respectively, captured from which 3D images were reconstructed and the features of pMFL structure: length and angle were measured. Besides, the chronological volume changes and the volume ratio of the knee joint components compared to E17 were calculated to identify the differences in growth by components. Results: pMFL was observed from E17 and was attached to the medial femoral condyle and lateral meniscus at all developmental stages, as in mature rats. The lack of marked variation in the attachment site and angle of the pMFL with the developmental stage indicates that the pMFL and surrounding knee joint components developed while maintaining their positional relationship from the onset of development. Conclusion: Current results may support to congenital etiology of meniscus disorder. [ABSTRACT FROM AUTHOR]

Developmental biology has greatly profited from genetic and reverse genetic approaches to indirectly studying protein function. More recently, nanobodies and other protein binders derived from different synthetic scaffolds have been used to directly dissect protein function. Protein binders have been fused to functional domains, such as to lead to protein degradation, relocalization, visualization, or posttranslational modification of the target protein upon binding. The use of such functionalized protein binders has allowed the study of the proteome during development in an unprecedented manner. In the coming years, the advent of the computational design of protein binders, together with further advances in scaffold engineering and synthetic biology, will fuel the development of novel protein binder–based technologies. Studying the proteome with increased precision will contribute to a better understanding of the immense molecular complexities hidden in each step along the way to generate form and function during development. [ABSTRACT FROM AUTHOR]

Mitochondria are essential organelles with specialized functions, which play crucial roles in energy production, calcium homeostasis, and programmed cell death. In oocytes, mitochondrial populations are inherited maternally and are vital for developmental competence. Dysfunction in mitochondrial quality control mechanisms can lead to reproductive failure. Due to their central role in oocyte and embryo development, mitochondria have been investigated as potential diagnostic and therapeutic targets in assisted reproduction. Pharmacological agents that target mitochondrial function and show promise in improving assisted reproduction outcomes include antioxidant coenzyme Q10 and mitoquinone, mammalian target of rapamycin signaling pathway inhibitor rapamycin, and nicotinamide mononucleotide. Mitochondrial replacement therapies (MRTs) offer solutions for infertility and mitochondrial disorders. Autologous germline mitochondrial energy transfer initially showed promise but failed to demonstrate significant benefits in clinical trials. Maternal spindle transfer (MST) and pronuclear transfer hold potential for preventing mitochondrial disease transmission and improving oocyte quality. Clinical trials of MST have shown promising outcomes, but larger studies are needed to confirm safety and efficacy. However, ethical and legislative challenges complicate the widespread implementation of MRTs. [ABSTRACT FROM AUTHOR]

Mitochondria are commonly recognized as the powerhouses of the cell, primarily responsible for energy production through oxidative phosphorylation. Alongside this vital function, they also play crucial roles in regulating calcium signaling, maintaining membrane potential, and modulating apoptosis. Their involvement in various cellular pathways becomes particularly evident during oogenesis and embryogenesis, where mitochondrial quantity, morphology, and distribution are tightly controlled. The efficiency of the mitochondrial network is maintained through multiple quality control mechanisms that are essential for reproductive success. These include mitochondrial unfolded protein response, mitochondrial dynamics, and mitophagy. Not surprisingly, mitochondrial dysfunction has been implicated in infertility and ovarian aging, prompting investigation into mitochondria as diagnostic and therapeutic targets in assisted reproduction. To date, mitochondrial DNA copy number in oocytes, cumulus cells, and trophectoderm biopsies, and fluorescent lifetime imaging microscopy-based assessment of NADH and flavin adenine dinucleotide content have been explored as potential predictors of embryo competence, yielding limited success. Despite challenges in the clinical application of mitochondrial diagnostic strategies, these enigmatic organelles have a significant impact on reproduction, and their potential role as diagnostic targets in assisted reproduction is likely to remain an active area of investigation in the foreseeable future. [ABSTRACT FROM AUTHOR]

The field of reproductive biology has made significant progress in recent years, identifying specific molecular players that influence oocyte development and function. Among them, sirtuin 3 (SIRT3) has attracted particular attention for its central role in mediating mitochondrial function and cellular stress responses in oocytes. So far, studies have demonstrated that the knockdown of SIRT3 leads to a decrease in blastocyst formation and an increase in oxidative stress within an embryo, underscoring the importance of SIRT3 in maintaining the cellular redox balance critical for embryonic survival and growth. Furthermore, the literature reveals specific signaling pathways, such as the SIRT3- Glycogen synthase kinase-3 beta (GSK3β) deacetylation pathway, crucial for mitigating oxidative stress-related anomalies in oocyte meiosis, particularly under conditions like maternal diabetes. Overall, the emerging role of SIRT3 in regulating oocyte mitochondrial function and development highlights the critical importance of understanding the intricate connections between cellular metabolism, stress response pathways, and overall reproductive health and function. This knowledge could lead to the development of novel strategies to support oocyte quality and fertility, with far-reaching implications for assisted reproductive technologies and women's healthcare. This commentary aims to provide an overview of the importance of SIRT3 in oocytes by synthesizing results from a multitude of studies. The aim is to elucidate the role of SIRT3 in oocyte development, maturation, and aging and to identify areas where further research is needed. [ABSTRACT FROM AUTHOR]

Dynamic changes in maternal‒zygotic transition (MZT) require complex regulation of zygote formation, maternal transcript decay, embryonic genome activation (EGA), and cell cycle progression. Although these changes are well described, some key regulatory factors are still elusive. Sirtuin-1 (SIRT1), an NAD+-dependent histone deacetylase, is a versatile driver of MZT via its epigenetic and nonepigenetic substrates. This study focused on the dynamics of SIRT1 in early embryos and its contribution to MZT. A conditional SIRT1-deficient knockout mouse model was used, accompanied by porcine and human embryos. Embryos across mammalian species showed the prominent localization of SIRT1 in the nucleus throughout early embryonic development. Accordingly, SIRT1 interacts with histone H4 on lysine K16 (H4K16) in both mouse and human blastocysts. While maternal SIRT1 is dispensable for MZT, at least one allele of embryonic Sirt1 is required for early embryonic development around the time of EGA. This role of SIRT1 is surprisingly mediated via a transcription-independent mode of action. [ABSTRACT FROM AUTHOR]

The bovine embryo production industry has seen significant growth over the past two decades, particularly in the production of in vitro produced embryos. This growth, driven by advancements in cryopreservation, in vitro culture mediums, ovum pick-up (OPU) procedures, ultrasonography devices, and embryo transfer (ET) has been notable. Particularly, ET is crucial for disseminating high genetic merit and amplifying foreign breeds by importing frozen embryos. This retrospective study aimed to assess factors affecting conception per embryo transfer (CPET) in Holstein-Friesian cattle in South Korea from October 2008 to July 2022. We evaluated type of embryo breed, type of embryo production (fresh and frozen; in vitro and in vivo production), recipient conditions including estrus type, corpus luteum quality, parity (nulliparous heifers, primiparous, and multiparous cows), and the daily mean temperature-humidity index (THI) as an index for heat stress. Type of embryo breed and estrus had no significant impact on CPET. However, we observed higher CPET in recipients with good quality corpus luteum, nulliparous heifers, and surrogates receiving fresh in vitro and frozen in vivo embryos. Importantly, CPET was not adversely affected by mild heat stress conditions (up to daily mean THI 76), indicating that using frozen in vivo embryos produced by multiple ovulation embryo transfer and fresh in vitro embryos by OPU-ET can help alleviate the subfertility issues in dairy cattle caused by global warming in Korea. • Recipient parity and corpus luteum were significant determinants of conception per embryo transfer (CPET) in Holstein cattle • The temperature-humidity index, a measure of heat stress, did not significantly affect CPET. • Embryo type significantly influenced CPET, • Selecting recipients by parity and CL quality is crucial for ET success, offering an effective AI alternative in heat-stressed environments. [ABSTRACT FROM AUTHOR]

METTL3-mediated N6-methyladenosine (m6A) modification is critical for gametogenesis and early embryonic development. However, the function of METTL3-mediated m6A modification in the early development of somatic nuclear transfer embryos (SCNT) remains unclear. Here, we found that METTL3 mRNA and protein levels exhibit dynamic changes during the early development of porcine SCNT embryos. The levels of METTL3 mRNA and protein in SCNT embryos at specific developmental stages differ from those in parthenogenetic activation (PA) counterparts. SiRNA injection effectively reduced the levels of METTL3 mRNA and protein in 4-cell embryos and blastocysts. METTL3 knockdown significantly reduced the cleavage and blastocyst rates of SCNT embryos. METTL3 knockdown significantly reduced the number of total cells and trophectoderm (TE) cells in the resulting blastocysts and perturbed cell lineage allocation. In addition, METTL3 knockdown reduced the levels of m6A modification in 4-cell embryos and blastocysts. Importantly, METTL3 knockdown decreased the expression levels of CDX2 , GATA3 , NANOG and YAP , and increased the expression levels of SOX2 and OCT4. Taken together, these results demonstrate that METTL3-mediated m6A modification regulates early development and lineage differentiation of porcine SCNT embryos. [ABSTRACT FROM AUTHOR]

This work explored whether a well-characterized recombinant human interleukin-6 (hIL6) protein will influence in vitro produced (IVP) bovine embryo development and survival after cryopreservation. Cumulus oocyte complexes were collected from abattoir derived ovaries, matured for 24 h, and fertilized using pooled semen from Holstein bulls. Embryos were treated with 0, 25, 50, or 100 ng/mL hIL6 on day 5 post-fertilization. An increase in ICM cell numbers was observed in each hIL6 treatment, with the lowest hIL6 treatment having the same magnitude of response as the middle and highest hIL6 concentration. No effects on TE cell numbers were observed. The second study involved cryopreserving (via slow freezing) of hIL6-treated blastocysts, then examining post-thaw blastocyst survival by incubating for 24 h in the absence of hIL6 treatments. Blastocyst re-expansion and hatching rates were unaffected by any of the IL6 treatments, however, increases in both ICM and TE cell numbers were detected at 24 h post-thawing in blastocysts exposed to 100 ng/mL hIL6 but not lower concentrations before freezing. A reduction in the percentage of TUNEL-positive TE cells was observed after thawing in blastocysts exposed to 25, 50 and 100 ng/mL hIL6 before cryopreservation. No treatment-dependent changes in TUNEL-positive ICM cells were observed. In summary, hIL6 supplementation improves ICM cell numbers in bovine blastocysts to a degree that is commensurate with what has been observed when using bovine recombinant IL6. This positive effect of hIL6 on ICM cell numbers is maintained after freezing and thawing, and a novel improvement in post-thaw TE cell numbers occur in hIL6 treated embryos. This positive effect on TE cell numbers is attributed, at least in part, to an hIL6-dependent reduction in TE cell apoptosis. [Display omitted] • Increases in inner cell mass cell numbers are achieved by supplementing recombinant human interleukin-6. • Positive effects of human interleukin-6 supplementation on both the inner cell mass and trophectoderm were detected after embryo cryopreservation. • Human interleukin-6 may is effective in bovine embryos. [ABSTRACT FROM AUTHOR]

This study explored the alteration in kisspeptin and reproductive hormones during different superovulation protocols (SOP) in dromedary camel. The kisspeptin and reproductive hormonal profile, ovarian response, and the quality and quantity of embryos in dromedary camel donors were evaluated. A total of thirty donor camels were divided into two groups: the 5dSOP group, which received diluent containing 400 mg pFSH dissolved in 20 ml and administered two times daily for 5 days at decreasing doses (2.5, 2, 1.5, and 1 ml); and the 3dSOP group, which received diluent containing 400 mg pFSH dissolved in 12 ml and administered two times daily for 3 days at decreasing doses (3 ml, 2 ml, and 1 ml). Ultrasonography was used to monitor the ovarian environment, recording daily follicle count and dimensions and the time taken for follicles to mature. On the sixth day after mating, a corpus luteum (CL) count was conducted. On the 8th day after mating, records of the quantity and quality of embryos collected were kept. Blood samples from the jugular vein were collected at the commencement of the superovulation protocol and at 8:00 a.m. for the following 48 h to measure the concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2), kisspeptin (KP), and progesterone (P4). The findings indicated that the 3dSOP yielded superior results compared to the 5dSOP in terms of follicle quantity and size, as well as the quantity of CL and embryos. This improvement was attributed to significantly higher concentrations of reproductive hormones, including FSH, LH, E2, kisspeptin, and P4 (P ≤ 0.05), in the 3dSOP than in the 5dSOP. In conclusion, reducing the duration of superovulation protocols contributed to the proliferation of follicles with improved dimensions and counts, ultimately resulting in a greater quantity of embryos of superior quality. The levels of FSH, LH, E2, KP, and P4 were affected significantly by SOP and time of evaluation. • This is the first experiment to study the alteration in kisspeptin, FSH, LH, E2, and P4 during superovulation protocols in camel. • This study recommended reducing the doses of pFSH to three days instead of five days. • There were significantly higher concentrations of FSH, LH, E2, kisspeptin, and P4 in the 3dSOP than in the 5dSOP. [ABSTRACT FROM AUTHOR]

Previous studies indicate a positive correlation between the duration of estrus prior to ovulation and likelihood of pregnancy in embryo recipient mares. However, the mechanisms by which the duration of estrus before may affect fertility remains unclear. This study aimed to determine the effect of different durations of estradiol exposure, prior to progesterone administration, on embryo viability in anestrous recipient mares, and endometrial expression of genes thought to influence embryo survival. Three groups of anestrous recipient mares treated with different duration of estradiol were used: long (LE), short (SE) and no treatment (NE). Day 8 embryos were transferred into recipient mares four days after long-acting progesterone administration and recovered 48h later to examine embryo growth and viability. The endometrial gene expression profile of selected genes was also investigated. The likelihood of recovering an embryo 48h after transfer was 46.1% (6/13), 62.5% (5/8) and 85.7% (6/7) for recipient mares from the NE, SE and LE groups, respectively (P =.09). Embryos recovered from the different groups of recipients did not, however, differ in size, morphology or the proportion of nuclei undergoing mitosis (P >.05). Abundance of mRNA for uterocalin (P19) and insulin-like growth factor 1 (IGF1) were increased in the LE compared to the NE group, while fibroblast growth factor 2 (FGF2), progesterone receptor (PGR) and insulin-like growth factor 1 receptor (IGF1R) transcript abundances were increased (P < 0.05) in the NE group compared to both SE and LE groups. In conclusion, a longer exposure of the endometrium to estradiol before progesterone tended to improve embryo survival within 48h of transfer. However, the grade, growth rate, and proportion of mitotic cells in surviving embryos did not differ among groups. If embryos are destined to fail in a suboptimal endometrial environment, they die and disappear quickly. Moreover, a more adequately estradiol-primed uterus, before the progesterone rise, seems to create a uterine environment, in terms of P19 , IGF1 , FGF2 and PGR gene expression, more conducive to embryo survival and further development. • Embryo survival after transfer tended to be influenced by the duration of estradiol treatment. • Viability of surviving embryos within 48h of transfer was not affected by estradiol-primed uterus. • Endometrial expression of selected genes was influenced by estradiol-primed uterus. [ABSTRACT FROM AUTHOR]

Objective: This study investigated the metabolic status of the spent culture media from embryos of patients with repeated implantation failure (RIF) undergoing in vitro fertilization–intracytoplasmic sperm injection cycles in comparison with the embryos from healthy fertile women. Methods: Metabolite levels in spent culture media were assessed and compared between embryos from RIF patients (n=35) and oocyte donors as controls (n=15). Protein levels of insulin-like growth factor 1 (IGF-1) were determined using Western blotting. Concentrations of glucose, pyruvate, and lactate were measured using spectrophotometry. Ionic colorimetric assay kits were utilized to analyze the concentrations of sodium, chloride, calcium, and magnesium ions. High-performance liquid chromatography was employed to measure the concentrations of glutamic acid, aspartic acid, methionine, phenylalanine, and histidine. Results: Glucose consumption and lactate secretion were higher in the control group than in the RIF group. The magnesium concentration was significantly higher in the control group than in the RIF group, but glutamic acid and aspartic acid concentrations were lower in the control group than in the RIF patients (p<0.05). The levels of IGF-1, sodium, calcium, chloride, methionine, histidine, and phenylalanine did not show statistically significant differences between the two groups. Conclusion: The metabolic profile of the culture medium of the embryos in the RIF group differed from that of the control group. These findings suggest potential factors that may affect implantation capacity in RIF patients and provide a new perspective on embryo selection. [ABSTRACT FROM AUTHOR]

Introduction: Co-administration of clomiphene citrate during the early phase in in vitro fertilisation (IVF) cycles has been used by clinicians for several years to improve outcomes. Materials and methods: This is a single centre retrospective study involving 222 patients with 81 patients underwent IVF using co-administration of Clomiphene Citrate (CC) with gonadotropins (study group - group A) and 141 patients used only gonadotropins (control group - group B) were compared. The outcome data of the IVF cycles were focused on four major aspects of which are the total gonadotropin usage, number of oocytes obtained, fertilisation rate and top-quality embryos produced. Results: The Mean ± Standard Deviation (SD) of patients age is (34.72 ± 3.25) and (34.39 ± 3.39) years old from group A and B respectively. Mean ± (SD) of total usage of gonadotropins (1834.57 ± 477.34iu) vs (2645.04 ± 669.66iu), (p< 0.0001), mean number of oocytes ± (SD) retrieved (8.54 ± 5.95) vs (10.32 ± 6.67), (p = 0.048), mean ± (SD) fertilisation rate (39.82 ± 30.46) vs (49.04 ± 27.62), (p=0.027) and mean percentage ± (SD) of the top-quality embryos are (17.96 ± 25.68) vs (28.02 ± 32.19), (p=0.02) from group A and B respectively. Conclusion: Co-administration of CC with gonadotropins in IVF reduces the total amount of gonadotropins required in the IVF cycle however it also reduces the number of oocytes retrieved, fertilization rate and top quality embryos. [ABSTRACT FROM AUTHOR]

This study evaluates the utilization of in vitro embryo production (IVEP) technology for the conservation and breeding of the Auliekol cattle breed, a primary beef breed in Kazakhstan facing population decline due to the cessation of breeding programs and the incursion of transboundary diseases. We assessed the effect of consecutive ovum pick-up (OPU) procedures on oocyte yield and embryo production in Auliekol and Aberdeen Angus cows. A total of 2232 and 3659 oocytes were aspirated from Auliekol and Aberdeen Angus donors, respectively, with significantly higher yields and embryo production observed in Aberdeen Angus cows. The application of a meiotic block using Butyrolactone I (BLI) and subsequent in vitro fertilization (IVF) protocols was employed, with embryo development monitored up to the morula/blastocyst stage. Results indicated that Auliekol cows exhibited lower oocyte recovery, cleavage, and blastocyst rates compared to Aberdeen Angus cows, likely due to genetic characteristics. Despite the challenges, IVEP presents a valuable tool for the preservation and future propagation of the Auliekol breed, highlighting the need for further research to enhance reproductive outcomes and conservation strategies. [ABSTRACT FROM AUTHOR]

ICSI is one of the most commonly used techniques to treat infertility. The sperm selection for the procedure is done 'randomly' by the embryologist according to the motility and morphology parameters which is known not to reflect the potential of a sperm for fertilization, pregnancy and a healthy childbearing. Since the apoptosis rate is higher in sperm cells of infertile patients, it is more likely to choose an apoptotic sperm by the 'random selection method'. We recently introduced a novel sperm selection technique namely 'Annexin-V coated polystrene bead technique'(APB-Tech), for the selection of non-apoptotic sperm cells. The principal of the technique is based on the binding affinity of an apoptotic sperm to 'Annexin-V covered beads' enabling to distinguish a viable and a healthy sperm by light microscopy. The aim of the present study was to observe the effects of this technique on ICSI outcomes in mice. Sibling-oocyte trial was conducted and the outcome measures were compared with the results of traditional sperm selection method. Embryo and blastocyst qualities and blastocyst development rates were significantly increased in APB-Tech group, while the other parameters were not affected. Promising results obtained from the technique reflect its promising potential as a new and powerful tool for sperm selection and thus infertility techniques. [ABSTRACT FROM AUTHOR]

Key message: In Cyrtanthus mackenii, development of embryo and endosperm were differentially affected by fertilization of male gametes with DNA damage and mutations. Pollen irradiation with ionizing radiations has been applied in plant breeding and genetic research, and haploid plant induction has mainly been performed by male inactivation with high-dose irradiation. However, the fertilization process of irradiated male gametes and the early development of embryo and endosperm have not received much attention. Heavy-ion beams, a type of radiation, have been widely applied as effective mutagens for plants and show a high mutation rate even at low-dose irradiation. In this study, we analyzed the effects of male gametes of Cyrtanthus mackenii irradiated with a carbon-ion beam at low doses on fertilization. In immature seeds derived from the pollination of irradiated pollen grains, two types of embryo sacs were observed: embryo sac with a normally developed embryo and endosperm and embryo sac with an egg cell or an undivided zygote and an endosperm. Abnormalities in chromosome segregation, such as chromosomal bridges, were observed only in the endosperm nuclei, irrespective of the presence or absence of embryogenesis. Therefore, in Cyrtanthus, embryogenesis is strongly affected by DNA damage or mutations in male gametes. Moreover, various DNA contents were detected in the embryo and endosperm nuclei, and endoreduplication may have occurred in the endosperm nuclei. As carbon-ion irradiation causes chromosomal rearrangements even at low doses, pollen irradiation can be an interesting tool for studying double fertilization and mutation heritability. [ABSTRACT FROM AUTHOR]

STUDY QUESTION Is periconceptional multiple-micronutrient supplement (MMS) use including folic acid (FA) compared to FA use only associated with increased embryonic growth, development, and birth weight in a high-risk population? SUMMARY ANSWER Women with MMS intake show no significant differences in first-trimester morphological embryo development, but increased first-trimester embryonic growth trajectories and fewer neonates born small for gestational age (SGA), less than the 3rd percentile (