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Background: MicroRNAs (miRNAs) are presented in the uterine lumen of many mammals, and in vitro experiments have determined that several miRNAs are important for the regulation of endometrial and trophoblast functions. Our aim was to identify and contrast the miRNAs present in extracellular vesicles (EVs) in the uterine lumen fluid (ULF) at the onset of attachment in cattle pregnancies (gestation d 18) initiated by artificial insemination (AI) or by the transfer of an in vitro-produced blastocyst (IVP-ET). A third group had no conceptus after the transfer of an IVP embryo. Results: The abundance of 263 annotated miRNAs was quantified in the EVs collected from ULF. There was an increase in the transcript abundance of 20 miRNAs in the ULF EVs from the AI pregnant group, while 4 miRNAs had a lower abundance relative to the group not containing a conceptus. Additionally, 4 miRNAs were more abundant in ULF EVs in the AI pregnant group relative to IVP-ET group (bta-mir-17, bta-mir-7-3, MIR7-1, MIR18A). Specific miRNAs in the ULF EVs were co-expressed with messenger RNAs expressed in extra-embryonic tissues and endometrium, including genes that are known to be their targets. Conclusions: The results provide biological insights into the participation of miRNAs in the regulation of trophoblast proliferation and differentiation, as well as in endometrium receptivity. The knowledge that in vitro cultured embryos can contribute to the altered abundance of specific miRNAs in the uterine lumen can lead to the development of corrective approaches to reduce conceptus losses during the first month of pregnancy in cattle. [ABSTRACT FROM AUTHOR]

Background: Appropriate regulation of genes expressed in oocytes and embryos is essential for acquisition of developmental competence in mammals. Here, we hypothesized that several genes expressed in oocytes and pre-implantation embryos remain unknown. Our goal was to reconstruct the transcriptome of oocytes (germinal vesicle and metaphase II) and pre-implantation cattle embryos (blastocysts) using short-read and long-read sequences to identify putative new genes. Results: We identified 274,342 transcript sequences and 3,033 of those loci do not match a gene present in official annotations and thus are potential new genes. Notably, 63.67% (1,931/3,033) of potential novel genes exhibited coding potential. Also noteworthy, 97.92% of the putative novel genes overlapped annotation with transposable elements. Comparative analysis of transcript abundance identified that 1,840 novel genes (recently added to the annotation) or potential new genes were differentially expressed between developmental stages (FDR < 0.01). We also determined that 522 novel or potential new genes (448 and 34, respectively) were upregulated at eight-cell embryos compared to oocytes (FDR < 0.01). In eight-cell embryos, 102 novel or putative new genes were co-expressed (|r|> 0.85, P < 1 × 10–8) with several genes annotated with gene ontology biological processes related to pluripotency maintenance and embryo development. CRISPR-Cas9 genome editing confirmed that the disruption of one of the novel genes highly expressed in eight-cell embryos reduced blastocyst development (ENSBTAG00000068261, P = 1.55 × 10–7). Conclusions: Our results revealed several putative new genes that need careful annotation. Many of the putative new genes have dynamic regulation during pre-implantation development and are important components of gene regulatory networks involved in pluripotency and blastocyst formation. [ABSTRACT FROM AUTHOR]

Mammalian oocytes accumulate more than ten thousand mRNAs, of which three to four thousand mRNAs are translationally repressed. The timings and sites of translational activation of these dormant mRNAs are crucial for promoting oocyte maturation and embryonic development. How these mRNAs are accumulated and distributed in oocytes is therefore a fundamental issue to be explored. A method that enables visualization of mRNA molecules with high resolution in a simple manner would be valuable for understanding how oocytes accumulate and regulate the dormant mRNAs. We have developed a highly sensitive whole-mount in situ hybridization method using in vitro-synthesized RNA probes and the tyramide signal amplification (TSA) system optimized for mouse oocytes and embryos. By using this method, Pou5f1/Oct4, Emi2, and cyclin B1 mRNAs were detected in immature oocytes and 2-cell stage embryos. Confocal microscopy showed that these mRNAs formed granular structures in the oocyte cytoplasm. The structures of Pou5f1/Oct4 and cyclin B1 mRNAs persisted in 2-cell stage embryos. Pou5f1/Oct4 RNA granules exhibited a solid-like property in immature oocytes and became liquid-like droplets in 2-cell stage embryos. Double-staining of cyclin B1 mRNA with Emi2 or Pou5f1/Oct4 mRNA revealed that these mRNAs were distributed as different RNA granules without overlapping each other and that the size of cyclin B1 RNA granules tended to be larger than that of Emi2 RNA granules. The structures and distribution patterns of these mRNAs were further analyzed by N-SIM super-resolution microscopy. This analysis revealed that the large-sized RNA granules consist of many small-sized granules, suggesting the accumulation and regulation of dormant mRNAs as basal-sized RNA granules. The method established in this study can easily visualize the structure and distribution of mRNAs accumulated in mammalian oocytes and embryos with high sensitivity and super-resolution. This method is useful for investigating the cellular and molecular mechanisms of translational control of mRNAs by which maturation and early developmental processes are promoted. [ABSTRACT FROM AUTHOR]

Objective: To explore the optimal models for predicting the formation of high-quality embryos in Poor Ovarian Response (POR) Patients with Progestin-Primed Ovarian Stimulation (PPOS) using machine learning algorithms. Methods: A retrospective analysis was conducted on the clinical data of 4,216 POR cycles who underwent in vitro fertilization (IVF) / intracytoplasmic sperm injection (ICSI) at Sichuan Jinxin Xinan Women and Children's Hospital from January 2015 to December 2021. Based on the presence of high-quality cleavage embryos 72 h post-fertilization, the samples were divided into the high-quality cleavage embryo group (N = 1950) and the non-high-quality cleavage embryo group (N = 2266). Additionally, based on whether high-quality blastocysts were observed following full blastocyst culture, the samples were categorized into the high-quality blastocyst group (N = 124) and the non-high-quality blastocyst group (N = 1800). The factors influencing the formation of high-quality embryos were analyzed using logistic regression. The predictive models based on machine learning methods were constructed and evaluated accordingly. Results: Differential analysis revealed that there are statistically significant differences in 14 factors between high-quality and non-high-quality cleavage embryos. Logistic regression analysis identified 14 factors as influential in forming high-quality cleavage embryos. In models excluding three variables (retrieved oocytes, MII oocytes, and 2PN fertilized oocytes), the XGBoost model performed slightly better (AUC = 0.672, 95% CI = 0.636–0.708). Conversely, in models including these three variables, the Random Forest model exhibited the best performance (AUC = 0.788, 95% CI = 0.759–0.818). In the analysis of high-quality blastocysts, significant differences were found in 17 factors. Logistic regression analysis indicated that 13 factors influence the formation of high-quality blastocysts. Including these variables in the predictive model, the XGBoost model showed the highest performance (AUC = 0.813, 95% CI = 0.741–0.884). Conclusion: We developed a predictive model for the formation of high-quality embryos using machine learning methods for patients with POR undergoing treatment with the PPOS protocol. This model can help infertility patients better understand the likelihood of forming high-quality embryos following treatment and help clinicians better understand and predict treatment outcomes, thus facilitating more targeted and effective interventions. [ABSTRACT FROM AUTHOR]

Background: In recent years, with benefits from the continuous improvement of clinical technology and the advantage of fertility preservation, the application of embryo cryopreservation has been growing rapidly worldwide. However, amidst this growth, concerns about its safety persist. Numerous studies have highlighted the elevated risk of perinatal complications linked to frozen embryo transfer (FET), such as large for gestational age (LGA) and hypertensive disorders during pregnancy. Thus, it is imperative to explore the potential risk of embryo cryopreservation and its related mechanisms. Methods: Given the strict ethical constraints on clinical samples, we employed mouse models in this study. Three experimental groups were established: the naturally conceived (NC) group, the fresh embryo transfer (Fresh-ET) group, and the FET group. Blastocyst formation rates and implantation rates were calculated post-embryo cryopreservation. The impact of FET on fetal growth was evaluated upon fetal and placental weight. Placental RNA-seq was conducted, encompassing comprehensive analyses of various comparisons (Fresh-ET vs. NC, FET vs. NC, and FET vs. Fresh-ET). Results: Reduced rates of blastocyst formation and implantation were observed post-embryo cryopreservation. Fresh-ET resulted in a significant decrease in fetal weight compared to NC group, whereas FET reversed this decline. RNA-seq analysis indicated that the majority of the expression changes in FET were inherited from Fresh-ET, and alterations solely attributed to embryo cryopreservation were moderate. Unexpectedly, certain genes that showed alterations in Fresh-ET tended to be restored in FET. Further analysis suggested that this regression may underlie the improvement of fetal growth restriction in FET. The expression of imprinted genes was disrupted in both FET and Fresh-ET groups. Conclusion: Based on our experimental data on mouse models, the impact of embryo cryopreservation is less pronounced than other in vitro manipulations in Fresh-ET. However, the impairment of the embryonic developmental potential and the gene alterations in placenta still suggested it to be a risky operation. [ABSTRACT FROM AUTHOR]

Background: Although, oocytes from prepubertal donors are known to be less developmentally competent than those from adult donors it does not restrain their ability to produce full-term pregnancies. The transcriptomic profile of embryos could be used as a predictor for embryo's individual developmental competence. The aim of the study was to compare transcriptomic profile of blastocysts derived from prepubertal and pubertal heifers oocytes. Bovine cumulus-oocyte complexes (COCs) were obtained by ovum pick- up method from prepubertal and pubertal heifers. After in vitro maturation COCs were fertilized and cultured to the blastocyst stage. Total RNA was isolated from both groups of blastocysts and RNA-seq was performed. Gene ontology analysis was performed by DAVID (Database for Annotation, Visualization and Integrated Discovery). Results: A higher average blastocyst rate was obtained in the pubertal than in the pre-pubertal group. There were no differences in the quality of blastocysts between the examined groups. We identified 436 differentially expressed genes (DEGs) between blastocysts derived from researched groups, of which 247 DEGs were downregulated in blastocysts derived from pubertal compared to prepubertal heifers oocytes, and 189 DEGs were upregulated. The genes involved in mitochondrial function, including oxidative phosphorylation (OXPHOS) were found to be different in studied groups using Kyoto Encyclopedia of Genes (KEGG) pathway analysis and 8 of those DEGs were upregulated and 1 was downregulated in blastocysts derived from pubertal compared to prepubertal heifers oocytes. DEGs associated with mitochondrial function were found: ATP synthases (ATP5MF-ATP synthase membrane subunit f, ATP5PD- ATP synthase peripheral stalk subunit d, ATP12A- ATPase H+/K + transporting non-gastric alpha2 subunit), NADH dehydrogenases (NDUFS3- NADH: ubiquinone oxidoreductase subunit core subunit S3, NDUFA13- NADH: ubiquinone oxidoreductase subunit A13, NDUFA3- NADH: ubiquinone oxidoreductase subunit A3), cytochrome c oxidase (COX17), cytochrome c somatic (CYCS) and ubiquinol cytochrome c reductase core protein 1 (UQCRC1). We found lower number of apoptotic cells in blastocysts derived from oocytes collected from prepubertal than those obtained from pubertal donors. Conclusions: Despite decreased expression of genes associated with OXPHOS pathway in blastocysts from prepubertal heifers oocytes, the increased level of ATP12A together with the lower number of apoptotic cells in these blastocysts might support their survival after transfer. [ABSTRACT FROM AUTHOR]

Background: Parental care benefits offspring but comes with costs. To optimize the trade-off of costs and benefits, parents should adjust care based on intrinsic and/or extrinsic conditions. The harm to offspring hypothesis suggests that parents should invest more in younger offspring than older offspring because younger offspring are more vulnerable. However, this hypothesis has rarely been comprehensively tested, as many studies only reveal an inverse correlation between parental care and offspring age, without directly testing the effects of offspring age on their vulnerability. To test this hypothesis, we studied Kurixalus eiffingeri, an arboreal treefrog with paternal care. We first performed a field survey by monitoring paternal care during embryonic development. Subsequently, we conducted a field experiment to assess the prevalence of egg predators (a semi-slug, Parmarion martensi) and the plasticity of male care. Finally, we conducted a laboratory experiment to assess how embryo age affects predation by P. martensi. Results: Our results showed that (1) male attendance and brooding frequency affected embryo survival, and (2) males attended and brooded eggs more frequently in the early stage than in the late stage. The experimental results showed that (3) males increased attendance frequency when the predators were present, and (4) the embryonic predation by the semi-slug during the early was significantly higher than in the late stage. Conclusions: Our findings highlight the importance of paternal care to embryo survival, and the care behavior is plastic. Moreover, our results provide evidence consistent with the predictions of the harm to offspring hypothesis, as males tend to care more for younger offspring which are more vulnerable. [ABSTRACT FROM AUTHOR]

Background: Previous studies have shown that the vitrification of metaphase II (MII) oocytes significantly represses their developmental potential. Abnormally increased oxidative stress is the probable factor; however, the underlying mechanism remains unclear. The walnut-derived peptide TW-7 was initially isolated and purified from walnut protein hydrolysate. Accumulating evidences implied that TW-7 was a powerful antioxidant, while its prospective application in oocyte cryopreservation has not been reported. Result: Here, we found that parthenogenetic activation (PA) zygotes derived from vitrified MII oocytes showed elevated ROS level and delayed progression of pronucleus formation. Addition of 25 μmol/L TW-7 in warming, recovery, PA, and embryo culture medium could alleviate oxidative stress in PA zygotes from vitrified mouse MII oocytes, furtherly increase proteins related to histone lactylation such as LDHA, LDHB, and EP300 and finally improve histone lactylation in PA zygotes. The elevated histone lactylation facilitated the expression of minor zygotic genome activation (ZGA) genes and preimplantation embryo development. Conclusions: Our findings revealed the mechanism of oxidative stress inducing repressed development of PA embryos from vitrified mouse MII oocytes and found a potent and easy-obtained short peptide that could significantly rescue the decreased developmental potential of vitrified oocytes, which would potentially contribute to reproductive medicine, animal protection, and breeding. [ABSTRACT FROM AUTHOR]

Background: Increasing evidence points to an active role of oviductal extracellular vesicles (oEVs) in the early embryo-maternal dialogue. However, it remains unclear whether oEVs contribute to the recognition of the presence of embryos and their quality in the oviduct. Hence, we examined whether the molecular cargo of oEVs secreted by bovine oviduct epithelial cells (BOEC) differs depending on the presence of good (≥ 8 cells, G) or poor (< 8 cells, P) quality embryos. In addition, differences in RNA profiles between G and P embryos were analyzed in attempt to distinguish oEVs and embryonic EVs cargos. Methods: For this purpose, primary BOEC were co-cultured with in vitro produced embryos (IVP) 53 h post fertilization as follows: BOEC with G embryos (BGE); BOEC with P embryos (BPE); G embryos alone (GE); P embryos alone (PE); BOEC alone (B) and medium control (M). After 24 h of co-culture, conditioned media were collected from all groups and EVs were isolated and characterized. MicroRNA profiling of EVs and embryos was performed by small RNA-sequencing. Results: In EVs, 84 miRNAs were identified, with 8 differentially abundant (DA) miRNAs for BGE vs. B and 4 for BPE vs. B (P-value < 0.01). In embryos, 187 miRNAs were identified, with 12 DA miRNAs for BGE vs. BPE, 3 for G vs. P, 8 for BGE vs. GE, and 11 for BPE vs. PE (P-value < 0.01). Conclusions: These results indicated that oEVs are involved in the oviductal-embryo recognition and pointed to specific miRNAs with signaling and supporting roles during early embryo development. [ABSTRACT FROM AUTHOR]

Background: The best method for selecting embryos ploidy is preimplantation genetic testing for aneuploidies (PGT-A). However, it takes more labour, money, and experience. As such, more approachable, non- invasive techniques were still needed. Analyses driven by artificial intelligence have been presented recently to automate and objectify picture assessments. Methods: In present retrospective study, a total of 3448 biopsied blastocysts from 979 Time-lapse (TL)-PGT cycles were retrospectively analyzed. The "intelligent data analysis (iDA) Score" as a deep learning algorithm was used in TL incubators and assigned each blastocyst with a score between 1.0 and 9.9. Results: Significant differences were observed in iDAScore among blastocysts with different ploidy. Additionally, multivariate logistic regression analysis showed that higher scores were significantly correlated with euploidy (p < 0.001). The Area Under the Curve (AUC) of iDAScore alone for predicting euploidy embryo is 0.612, but rose to 0.688 by adding clinical and embryonic characteristics. Conclusions: This study provided additional information to strengthen the clinical applicability of iDAScore. This may provide a non-invasive and inexpensive alternative for patients who have no available blastocyst for biopsy or who are economically disadvantaged. However, the accuracy of embryo ploidy is still dependent on the results of next-generation sequencing technology (NGS) analysis. [ABSTRACT FROM AUTHOR]

The role of cytoplasmic fragmentation in human embryo development and reproductive potential is widely recognized, albeit without standard definition nor agreed upon implication. While fragmentation is best understood to be a natural process across species, the origin of fragmentation remains incompletely understood and likely multifactorial. Several factors including embryo culture condition, gamete quality, aneuploidy, and abnormal cytokinesis seem to have important role in the etiology of cytoplasmic fragmentation. Fragmentation reduces the volume of cytoplasm and depletes embryo of essential organelles and regulatory proteins, compromising the developmental potential of the embryo. While it has been shown that degree of fragmentation and embryo implantation potential are inversely proportional, the degree, pattern, and distribution of fragmentation as it relates to pregnancy outcome is debated in the literature. This review highlights some of the challenges in analysis of fragmentation, while revealing trends in our evolving knowledge of how fragmentation may relate to functional development of the human embryos, implantation, and pregnancy outcome. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to compare the per OPU clinical outcomes for transfer of Day 3 double cleavage-stage embryos (DET) and Day 5 single blastocyst-stage (SBT) in patients with five or fewer good quality embryos on day 3 per occyte pick-up cycle (OPU) in antagonist cycles with consideration of blastocyst formation failure. Methods: This was a retrospective, observational cohort study of 2,116 cases of OPU treated with antagonist protocol in the affiliated Chenggong Hospital of Xiamen University between January 2013 and December 2020. DET was performed in 1,811cycles and SBT was performed in 305 cycles. The DET group was matched to the SBT group by propensity score (PS) matching according to multiple maternal baseline covariates. After PS matching, there were 303 ET cycles in each group. The primary outcomes were the cumulative live birth rate (CLBR), cumulative multiple pregnancy rate(CMPR)per OPU and the number of ET to achieve live birth per OPU. Secondary outcomes were the percentage of clinical pregnancy(CPR), live birth rate(LBR), multiple pregnancy rate(MPR). Results: Following PS mating, the CLBR was slightly higher (48.8% versus 40.3% ; P = 0.041) and the CMPR was significantly higher in the DET group compared to SBT group(44.2% versus 7.9%, P < 0.001). The CPR, LBR and MPR per fresh transfer were higher in DET group compared to SBT group(50.2% versus 28.7%; 41.3% versus 21.5%;29.6% versus 0%, P < 0.001). The number of ET to achieve live birth per OPU in SBT group was obiviously more than in DET group(1.48 ± 0.578 versus 1.22 ± 0.557 ,P < 0.001). Conclusion: With a marginal difference cumulative live birth rate, the lower live birth rate per fresh transfer and higher number of ET per OPU in the SBT group suggested that it might take longer time to achieve a live birth with single blastocyst strategy. A trade-off decision should be made between efficiency and safety. [ABSTRACT FROM AUTHOR]

Background: Rose is recognized as an important ornamental plant worldwide, and it is also one of the most widely used flowers in gardens. At present, the improvement of rose traits is still difficult and uncertain, and molecular breeding can provide new ideas for the improvement of modern rose varieties. Somatic embryos are quite good receptors for genetic transformation. However, little is known about the molecular mechanisms underlying during the regeneration process of rose somatic embryos. To elucidate the molecular regulation mechanism of somatic embryo plantlet regeneration, the relationship between the differences in traits of the two different regenerated materials and the significantly differentially expressed genes (DEGs) related to phytohormone pathways in the process of regeneration were be investigated. Results: These representative two regenerated samples from single-piece cotyledonary somatic embryo (SPC) culture of Rosa hybrida 'John F. Kennedy', were harvested for transcriptome analysis, with the SPC explants at the initial culture (Day 0) as the control. The differentially expressed genes (DEGs) in the materials from two different types for regeneration approach (SBF type: the regeneration approach type of single bud formed from SPC explants; MBF type: the regeneration approach type of multiple buds formed from SPC explants) were be screened by means of the transcriptome sequencing technology. In this study, a total of about 396.24 million clean reads were obtained, of which 78.95–82.92% were localized to the reference genome, compared with the initial material (CK sample), there were 5594 specific genes in the material of SBF type and 6142 specific genes in the MBF type. The DEGs from the SBF type material were mainly concentrated in the biological processes of GO terms such as phytohormones, substance transport, cell differentiation, and redox reaction. The KEGG enrichment analysis revealed these DEGs were more active in ubiquinone and other terpenoid-quinone biosynthesis, fatty acid elongation, steroid biosynthesis, and glycosphingolipid biosynthesis-globo and isoglobo series. In contrast, the DEGs induced by the MBF type material were mainly associated with the biological processes such as phytohormones, phosphorylation, photosynthesis and signal transduction. According to KEGG analysis, these DEGs of MBF type were significantly enriched in the porphyrin and chlorophyll metabolism, brassinosteroid biosynthesis, carotenoid biosynthesis, and peroxisome. Furthermore, the results from the phytohormone pathways analysis showed that the auxin-responsive factor SAUR and the cell wall modifying enzyme gene XTH were upregulated for expression but the protein phosphatase gene PP2C was downregulated for expression in SBF type; the higher expression of the ethylene receptor ETR, the ethylene transduction genes EBF1/2, the transcription factor EIN3, and the ethylene-responsive transcription factor ERF1/2 were induced by MBF type. Conclusions: According to the GO and KEGG analysis, it indicated the DEGs between two different regenerated materials from somatic embryos were significantly different which might be causing morphological differences. That was somatic embryos from Rosa hybrida 'John F. Kennedy' could regenerate plantlet via both classic somatic embryogenesis (seed-like germination) and organogenesis, cotyledonary somatic embryos should be considered as one kind of intermediate materials similiar to callus, rather than the indicator materials for somatic embryogenesis. [ABSTRACT FROM AUTHOR]

Congenital infection caused by vertical transmission of microsporidia N. bombycis can result in severe economic losses in the silkworm-rearing industry. Whole-transcriptome analyses have revealed non-coding RNAs and their regulatory networks in N. bombycis infected embryos and larvae. However, transcriptomic changes in the microsporidia proliferation and host responses in congenitally infected embryos and larvae remains unclear. Here, we simultaneously compared the transcriptomes of N. bombycis and its host B. mori embryos of 5-day and larvae of 1-, 5- and 10-day during congenital infection. For the transcriptome of N. bombycis, a comparison of parasite expression patterns between congenital-infected embryos and larva showed most genes related to parasite central carbon metabolism were down-regulated in larvae during infection, whereas the majority of genes involved in parasite proliferation and growth were up-regulated. Interestingly, a large number of distinct or shared differentially expressed genes (DEGs) were revealed by the Venn diagram and heat map, many of them were connected to infection related factors such as Ricin B lectin, spore wall protein, polar tube protein, and polysaccharide deacetylase. For the transcriptome of B. mori infected with N. bombycis, beyond numerous DEGs related to DNA replication and repair, mRNA surveillance pathway, RNA transport, protein biosynthesis, and proteolysis, with the progression of infection, a large number of DEGs related to immune and infection pathways, including phagocytosis, apoptosis, TNF, Toll-like receptor, NF-kappa B, Fc epsilon RI, and some diseases, were successively identified. In contrast, most genes associated with the insulin signaling pathway, 2-oxacarboxylic acid metabolism, amino acid biosynthesis, and lipid metabolisms were up-regulated in larvae compared to those in embryos. Furthermore, dozens of distinct and three shared DEGs that were involved in the epigenetic regulations, such as polycomb, histone-lysine-specific demethylases, and histone-lysine-N-methyltransferases, were identified via the Venn diagram and heat maps. Notably, many DEGs of host and parasite associated with lipid-related metabolisms were verified by RT-qPCR. Taken together, simultaneous transcriptomic analyses of both host and parasite genes lead to a better understanding of changes in the microsporidia proliferation and host responses in embryos and larvae in N. bombycis congenital infection. [ABSTRACT FROM AUTHOR]

Background: Extracellular vesicles (EVs) and their cargoes, including MicroRNAs (miRNAs) play a crucial role in cell-to-cell communication. We previously demonstrated the upregulation of bta-mir-148b in EVs from oviductal fluid of cyclic cows. This miRNA is linked to the TGF-β pathway in the cell proliferation. Our aim was to verify whether miR-148b is taken up by embryos through gymnosis, validate its target genes, and investigate the effect of miR-148b supplementation on early embryo development and quality. Methods: Zygotes were cultured in SOF + 0.3% BSA (Control) or supplemented with: 1 µM miR-148b mimics during: D1-D7 (miR148b) or D1-D4 (miR148b-OV: representing miRNA effect in the oviduct) or D4-D7 (miR148b-UT: representing miRNA effect in the uterus) or 1 µM control mimics was used during: D1-D7 (CMimic). Embryos at ≥ 16-cells and D7 blastocysts (BD7) were collected to examine the mRNA abundance of transcripts linked to the TGF-β pathway (TGFBR2, SMAD1, SMAD2, SMAD3, SMAD5, BMPR2, RPS6KB1, POU5F1, NANOG), total cell number (TC), trophectoderm (TE), and inner cell mass (ICM) were also evaluated. One-way ANOVA was used for all analyses. Results: We demonstrated that miR-148b can be taken up in both 16-cell embryos and BD7 by gymnosis, and we observed a decrease in SMAD5 mRNA, suggesting it's a potential target of miR-148b. Cleavage and blastocysts rates were not affected in any groups; however, supplementation of miR-148b mimics had a positive effect on TC, TE and ICM, with values of 136.4 ± 1.6, 92.5 ± 0.9, 43.9 ± 1.3 for miR148b and 135.3 ± 1.5, 92.6 ± 1.2, 42.7 ± 0.8, for miR148b-OV group. Furthermore, mRNA transcripts of SMAD1 and SMAD5 were decreased (P ≤ 0.001) in 16-cell embryos and BD7 from miR148b and miR148b-OV groups, while POU5F1 and NANOG were upregulated (P ≤ 0.001) in BD7 and TGFBR2 was only downregulated in 16-cell embryos. pSMAD1/5 levels were higher in the miR148b and miR148b-OV groups. Conclusions: Our findings suggest that supplementation of bta-miR-148b mimics during the entire culture period (D1 - D7) or from D1 - D4 improves embryo quality and influences the TGF-β signaling pathway by altering the transcription of genes associated with cellular differentiation and proliferation. This highlights the importance of miR-148b on embryo quality and development. [ABSTRACT FROM AUTHOR]

Background: Artificial Intelligence entails the application of computer algorithms to the huge and heterogeneous amount of morphodynamic data produced by Time-Lapse Technology. In this context, Machine Learning (ML) methods were developed in order to assist embryologists with automatized and objective predictive models able to standardize human embryo assessment. In this study, we aimed at developing a novel ML-based strategy to identify relevant patterns associated with the prediction of blastocyst development stage on day 5. Methods: We retrospectively analysed the morphokinetics of 575 embryos obtained from 80 women who underwent IVF at our Unit. Embryo morphokinetics was registered using the Geri plus® time-lapse system. Overall, 30 clinical, morphological and morphokinetic variables related to women and embryos were recorded and combined. Some embryos reached the expanded blastocyst stage on day 5 (BL Group, n = 210), some others did not (nBL Group, n = 365). Results: The novel EmbryoMLSelection framework was developed following four-steps: Feature Selection, Rules Extraction, Rules Selection and Rules Evaluation. Six rules composed by a combination of 8 variables were finally selected, and provided a predictive power described by an AUC of 0.84 and an accuracy of 81%. Conclusions: We provided herein a new feature-signature able to identify with an high performance embryos with the best developmental competence to reach the expanded blastocyst stage on day 5. Clear and clinically relevant cut-offs were identified for each considered variable, providing an objective tool for early embryo developmental assessment. [ABSTRACT FROM AUTHOR]

Tobacco pollutants are prevalent in the environment, leading to inadvertent exposure of pregnant females. Studies of these pollutants' toxic effects on embryonic development have not fully elucidated the potential underlying mechanisms. Therefore, in this study, we aimed to investigate the developmental toxicity induced by cigarette smoke extract (CSE) at concentrations of 0.25, 1, and 2.5% using a zebrafish embryo toxicity test and integrated transcriptomic analysis of microRNA (miRNA) and messenger RNA (mRNA). The findings revealed that CSE caused developmental toxicity, including increased mortality and decreased incubation rate, in a dose-dependent manner. Moreover, CSE induced malformations and apoptosis, specifically in the head and heart of zebrafish larvae. We used mRNA and miRNA sequencing analyses to compare changes in the expression of genes and miRNAs in zebrafish larvae. The bioinformatics analysis indicates that the mechanism underlying CSE-induced developmental toxicity was associated with compromised genetic material damage repair, deregulated apoptosis, and disturbed lipid metabolism. The enrichment analysis and RT-qPCR show that the ctsba gene plays a crucial function in embryo developmental apoptosis, and the fads2 gene mainly regulates lipid metabolic toxicity. The results of this study improve the understanding of CSE-induced developmental toxicity in zebrafish embryos and contribute insights into the formulation of novel preventive strategies against tobacco pollutants during early embryonic development. [ABSTRACT FROM AUTHOR]

Background: Recent studies about the effect of gonadotropin (Gn) dose on the clinical outcomes of IVF are still controversial, and no studies have analyzed the relationship between Gn dose and embryo quality. Since AMH is a strong predictor of oocyte quality, we aim to evaluate the relationship between total Gn dose and embryo quality and clinical outcomes at different AMH levels in IVF cycles. Methods: A total of 12,588 patients were enrolled in the retrospective study. The included cycles were categorized by serum AMH levels (AMH ≤ 1 ng/ml, 1 ng/ml < AMH ≤ 3 ng/ml, 3 ng/ml < AMH ≤ 5 ng/ml, AMH > 5 ng/ml), total Gn dosage (< 1875 IU, 1875–3750 IU and ≥ 3750 IU) and female age (< 35 years and 35–42 years). The embryo quality and clinical outcomes were the measure outcomes. Results: The top-day3 embryos rate decreased with the increase of total Gn dose in nearly all age and AMH subgroups, but this trend was not obvious in the AMH > 5 ng/ml group and AMH ≤ 1 ng/ml group. The blastocyst formation rate and high-quality blastulation rate had a negative relationship with Gn does for women aged < 35 years in the AMH ≤ 5 ng/ml groups, except for the AMH > 5 ng/ml group (P < 0.001). However, when women were 35–42 years old, regardless of AMH levels, the blastocyst formation rate and high-quality blastulation rate decreased as Gn dose increased. Clinical outcomes (implantation rate, clinical pregnancy rate and live birth rate) decreased with the increase of Gn dose in all ages and AMH stratifications. Conclusions: The total dose of Gn may have different effects on embryo quality at different serum AMH levels, and the negative effects of total dose of Gn on clinical outcomes may be realized by impairing both embryo quality and endometrium. [ABSTRACT FROM AUTHOR]

Purpose: The introduction of the time-lapse monitoring system (TMS) and the development of predictive algorithms could contribute to the optimal embryos selection for transfer. Therefore, the present study aims at investigating the efficiency of KIDScore and iDAScore systems for blastocyst stage embryos in predicting live birth events. Methods: The present retrospective study was conducted in a private IVF Unit setting throughout a 10-month period from October 2021 to July 2022, and included the analysis of 429 embryos deriving from 91 IVF/ICSI cycles conducted due to infertility of various etiologies. Embryos incubated at the Embryoscope+ timelapse incubator were analyzed through the established scoring systems: KIDScore and iDAScore®. The main outcome measure was the comparison of the two scoring systems in terms of live birth prediction. Embryos with the higher scores at day 5 (KID5 score/iDA5 score) were transferred or cryopreserved for later use. Results: Embryos with high KID5 and iDA5 scores positively correlated with the probability of successful live birth, with KID5 score yielding a higher efficiency in predicting a successful reproductive outcome compared to a proportionally high iDA5 score. KID5 demonstrated conservative performance in successfully predicting live birth compared to iDA5 score, indicating that an efficient prediction can be either provided by a relatively lower KID5 score or a relatively higher iDA5 score. Conclusion: The developed artificial intelligence tools should be implemented in clinical practice in conjunction with the conventional morphological assessment for the conduction of optimized embryo transfer in terms of a successful live birth. [ABSTRACT FROM AUTHOR]

Background: Evidence from the Istanbul consensus workshop suggests correlations between morphological parameters and embryo developments. 8-cell embryos are the best blastomere stage on day 3. No good quality evidence exists to support high-quality embryonic selection following blastulation and clinical outcomes. This study aimed to investigate the factors that affect blastocyst formation, blastocyst quality, and clinical outcomes of high-quality cleavage-stage embryos in fresh cycles. Methods: This study was a retrospective analysis of 9608 high-quality cleavage-stage embryos from 2987 couples between January 2017 to June 2021, namely 1520 embryos categorized as "812" (8-cell, grade 2, mild fragmentation), 2961 as "821" (8-cell, grade 2, mild asymmetry), 896 as "711" (7-cell, grade 1), and 517 as "911" (9-cell, grade 1) compared with 3714 embryos categorized as "811" (8-cell, grade 1). The primary outcomes were clinical pregnancy rate (CPR) and live birth rate (LBR). Blastulation rate (BR), available late blastocyst rate (ABR) and high-quality late blastocyst rate (HBR) were secondary outcome measures. Results: BR, ABR, and HBR had significant differences among the five groups (P < 0.001), while CPR and LBR were also significantly different in cleavage-stage fresh transfer (P < 0.01). The multivariable multilevel logistic regression analysis revealed a significant association between cell number, cell size, blastocyst development and clinical outcomes. For 7 to 9-cell highest-quality embryo, mild fragmentation and more blastomeres were more conducive to blastocyst formation and clinical outcomes. While cleavage-stage embryos developed into blastocysts, the negative impact of their initial morphology on clinical outcomes would be erased. Conclusions: Our study firstly evaluated blastocyst development and clinical outcomes of high-quality cleavage-stage embryos in fresh cycles, with rankings of 811, 812, 911, 821, and 711. We found the initial morphological characteristics of the high-quality cleavage-stage embryos did not adversely impact clinical outcomes, even as they progressed to the blastocyst stage. [ABSTRACT FROM AUTHOR]

Pachyzoidae is a little-known family of deep-sea ctenostome Bryozoa that until now was monospecific for Pachyzoon atlanticum. Originally described from the Atlantic Ocean, the genus was also found off southeastern New Caledonia in deep waters of the geological continent of Zealandia. Pachyzoon atlanticum forms globular to flat round colonies, living on soft, muddy to sandy bottoms with a few rhizoidal cystid appendages extending from the basal, substrate-oriented side. In this study, we investigate additional pachyzoids, collected between 1965 and 2015 from over 40 sites around New Zealand, by means of detailed morphological and histological investigations. In total, several hundred colonies were encountered in the NIWA Invertebrate Collection, comprising two new species of the genus Pachyzoon, P. grischenkoi sp. nov. and P. pulvinaris sp. nov., and the new genus and species Jeanloupia zealandica gen. et sp. nov.. The genus Jeanloupia is characterized by small disc-shaped colonies with highly elongated peristomes and a quadrangular aperture, distinct from the round apertures of the genus Pachyzoon. Pachyzoid species differ in colony structure and shape, apertural papillae and polypide features such as tentacle number or digestive-tract details. Cystid appendages are non-kenozooidal, but may originate from laterally flanking kenozooids. Based on published images, alleged P. atlanticum from New Caledonia is re-interpreted as P. pulvinaris n. sp.. Morphological characters support alcyonidioidean relationships, as previously suggested. First observations on pachyzoid reproduction show macrolecithal oocytes and brooding of embryos, which seems to be the general pattern for this family. The occurrence of three new Zealandian species in a comparatively small geographical area far from the Atlantic indicates a high possibility of more species to discovered. [ABSTRACT FROM AUTHOR]

During embryo implantation, blastocyst interacts with the receptivity endometrium and the endometrial epithelium secretes nurturing fluid to support embryonic development. Interferon-λ (IFN-λ) is a novel, non-redundant regulator that participates in the fetal–maternal interaction; however, the precise molecular mechanism underlying its impact on uterine receptivity remains elusive. Here, microarray profiling revealed that 149 specific miRNAs were differentially expressed in the human endometrial cells following IFN-λ treatment. In particular, miR-124-3p expression was significantly reduced after IFN-λ treatment (p < 0.05). An in vivo mouse pregnancy model showed that miR-124-3p overexpression notably decreased embryo implantation rate and led to an aberrant epithelial phenotype. Furthermore, miR-124-3p negatively impacted the migration and proliferation of endometrial cells, and hindered embryonic developmental competence in terms of blastocyst formation and global DNA re-methylation. Downstream analysis showed that LIF, MUC1 and BCL2 are potential target genes for miR-124-3p, which was confirmed using western blotting and immunofluorescence assays. In conclusion, IFN-λ-driven downregulation of miR-124-3p during embryo implantation modulates uterine receptivity. The dual functional role of miR-124-3p suggests a cross-talk model wherein, maternal endometrial miRNA acts as a transcriptomic modifier of the peri-implantation endometrium and embryo development. [ABSTRACT FROM AUTHOR]

Background: Basal energetic metabolism in sperm, particularly oxidative phosphorylation, is known to condition not only their oocyte fertilising ability, but also the subsequent embryo development. While the molecular pathways underlying these events still need to be elucidated, reactive oxygen species (ROS) could have a relevant role. We, therefore, aimed to describe the mechanisms through which mitochondrial activity can influence the first stages of embryo development. Results: We first show that embryo development is tightly influenced by both intracellular ROS and mitochondrial activity. In addition, we depict that the inhibition of mitochondrial activity dramatically decreases intracellular ROS levels. Finally, we also demonstrate that the inhibition of mitochondrial respiration positively influences sperm DNA integrity, most likely because of the depletion of intracellular ROS formation. Conclusion: Collectively, the data presented in this work reveals that impairment of early embryo development may result from the accumulation of sperm DNA damage caused by mitochondrial-derived ROS. [ABSTRACT FROM AUTHOR]

Background: Organisms from many distinct evolutionary lineages acquired the capacity to enter a dormant state in response to environmental conditions incompatible with maintaining normal life activities. Most studied organisms exhibit seasonal or annual episodes of dormancy, but numerous less studied organisms enter long-term dormancy, lasting decades or even centuries. Intriguingly, many planktonic animals produce encased embryos known as resting eggs or cysts that, like plant seeds, may remain dormant for decades. Herein, we studied a rotifer Brachionus plicatilis as a model planktonic species that forms encased dormant embryos via sexual reproduction and non-dormant embryos via asexual reproduction and raised the following questions: Which genes are expressed at which time points during embryogenesis? How do temporal transcript abundance profiles differ between the two types of embryos? When does the cell cycle arrest? How do dormant embryos manage energy? Results: As the molecular developmental kinetics of encased embryos remain unknown, we employed single embryo RNA sequencing (CEL-seq) of samples collected during dormant and non-dormant embryogenesis. We identified comprehensive and temporal transcript abundance patterns of genes and their associated enriched functional pathways. Striking differences were uncovered between dormant and non-dormant embryos. In early development, the cell cycle-associated pathways were enriched in both embryo types but terminated with fewer nuclei in dormant embryos. As development progressed, the gene transcript abundance profiles became increasingly divergent between dormant and non-dormant embryos. Organogenesis was suspended in dormant embryos, concomitant with low transcript abundance of homeobox genes, and was replaced with an ATP-poor preparatory phase characterized by very high transcript abundance of genes encoding for hallmark dormancy proteins (e.g., LEA proteins, sHSP, and anti-ROS proteins, also found in plant seeds) and proteins involved in dormancy exit. Surprisingly, this period appeared analogous to the late maturation phase of plant seeds. Conclusions: The study highlights novel divergent temporal transcript abundance patterns between dormant and non-dormant embryos. Remarkably, several convergent functional solutions appear during the development of resting eggs and plant seeds, suggesting a similar preparatory phase for long-term dormancy. This study accentuated the broad novel molecular features of long-term dormancy in encased animal embryos that behave like "animal seeds". [ABSTRACT FROM AUTHOR]

Introduction: Assisted Reproductive Technology utilizes human sperm, eggs, or embryos in vitro to produce pregnancy. However, there is no evidence of the acceptance of these technologies by the community. Objective: This study aimed to determine the pooled prevalence of positive attitudes toward the acceptance of donor eggs, embryos, and sperm. Methods: The protocol was registered in PROSPERO (number: CRD42022348036). The Condition, Context and Population (CoCoPop) protocol of the systematic review was used to address the relevant questions regarding the objective of the study. Data were extracted into Excel and pooled estimates were calculated using STATA Version 16. Results: The pooled prevalence of positive attitudes toward accepting donor eggs, embryos, and sperms was 38.63%, 33.20%, and 31.34%, respectively. Subgroup analysis revealed that the pooled prevalence of positive attitudes toward accepting donor eggs was high in non-Asian countries (47.78%) and among infertile men (38.60%). Similarly, the pooled prevalence of positive attitudes toward accepting donor eggs was high in non-Asian countries (47.78%) and among infertile men (28.67%). However, the pooled prevalence of positive attitudes toward accepting donor sperm was high in non-Asian countries (37.6%) and among infertile women (28.19%). Conclusion: The pooled estimate of the prevalence of positive attitudes toward accepting donor eggs was higher than the prevalence of positive attitudes toward accepting donor embryos and sperm. Infertile men and non-Asian countries have a higher prevalence of positive attitudes toward accepting eggs and embryos, whereas non-Asian countries and infertile women present a higher prevalence of positive attitudes toward accepting donor sperm. Therefore, regulatory bodies and policymakers should modify their rules and regulations to ensure the availability of minimum standards for the ethical and safe practice of donor conception as a treatment for infertility at national and international levels. Plain language summary: Assisted Reproductive Technology (ART) utilizes human sperm, eggs, or embryos in vitro to induce pregnancy; however, there is no evidence of community acceptance of these technologies. A systematic review and meta-analysis found that 38.63% of infertile couples had positive attitudes toward donor eggs, while 33.20% and 31.34% had negative attitudes. Females are more amenable to accepting donor gametes, embryos, or eggs than males, and females are more amenable to accepting donor eggs than donor sperm. To improve attitudes toward donor conception, infertile couples must understand the medical and obstetric risks associated with donor-assisted conception. This review recommends strengthening counseling for infertile couples and offering support to those with negative attitudes toward donor conception. Regulatory bodies and policymakers should consider the needs of infertile couples and modify their rules to ensure minimum standards for ethical and safe practices of donor conception as a treatment for infertility. [ABSTRACT FROM AUTHOR]

Background: High environmental temperatures induce heat stress in broiler chickens, affecting their health and production performance. Several dietary, managerial, and genetics strategies have been tested with some success in mitigating heat stress (HS) in broilers. Developing novel HS mitigation strategies for sustaining broiler production is critically needed. This study investigated the effects of pre-hatch thermal manipulation (TM) and post-hatch baicalein supplementation on growth performance and health parameters in heat-stressed broilers. Results: Six hundred fertile Cobb 500 eggs were incubated for 21 d. After candling on embryonic day (ED) 10, 238 eggs were thermally manipulated at 38.5 °C with 55% relative humidity (RH) from ED 12 to 18, then transferred to the hatcher (ED 19 to 21, standard temperature) and 236 eggs were incubated at a controlled temperature (37.5 °C) till hatch. After hatch, 180-day-old chicks from both groups were raised in 36 pens (n = 10 birds/pen, 6 replicates per treatment). The treatments were: 1) Control, 2) TM, 3) control heat stress (CHS), 4) thermal manipulation heat stress (TMHS), 5) control heat stress supplement (CHSS), and 6) thermal manipulation heat stress supplement (TMHSS). All birds were raised under the standard environment for 21 d, followed by chronic heat stress from d 22 to 35 (32–33 °C for 8 h) in the CHS, TMHS, CHSS, and TMHSS groups. A thermoneutral (22–24 °C) environment was maintained in the Control and TM groups. RH was constant (50% ± 5%) throughout the trial. All the data were analyzed using one-way ANOVA in R and GraphPad software at P < 0.05 and are presented as mean ± SEM. Heat stress significantly decreased (P < 0.05) the final body weight and ADG in CHS and TMHS groups compared to the other groups. Embryonic TM significantly increased (P < 0.05) the expression of heat shock protein-related genes (HSP70, HSP90, and HSPH1) and antioxidant-related genes (GPX1 and TXN). TMHS birds showed a significant increment (P < 0.05) in total cecal volatile fatty acid (VFA) concentration compared to the CHS birds. The cecal microbial analysis showed significant enrichment (P < 0.05) in alpha and beta diversity and Coprococcus in the TMHSS group. Conclusions: Pre-hatch TM and post-hatch baicalein supplementation in heat-stressed birds mitigate the detrimental effects of heat stress on chickens' growth performance, upregulate favorable gene expression, increase VFA production, and promote gut health by increasing beneficial microbial communities. [ABSTRACT FROM AUTHOR]

Background: Erythropoietin (Epo) is a potent vascular growth factor that induces angiogenesis and antiapoptotic signalling. We investigated whether the development of numerous follicles and corpora lutea during in vitro fertilization (IVF) cycle affects circulating Epo levels and further, if Epo could be used as a novel marker for ovarian hyperstimulation syndrome (OHSS). Methods: 24 women were included in the uncomplicated IVF group and 35 women in the OHSS group. Repeated blood samples from both groups were analysed for Epo, progesterone, blood haemoglobin, and creatinine. Follicular fluid from the IVF group was analysed for Epo and progesterone. Repeated measure analysis was performed for the variables and circulating Epo levels were compared between the IVF group and early OHSS. Furthermore, related growth factors, vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1 (HIF-1) were analysed from subgroup of women to test for correlation with Epo. Results: During IVF, circulating Epo increased from natural mid-luteal phase to stimulated mid-luteal phase (median 9.5; 95% CI 7.2–13.4 IU/L and 12.5; 10.3–13.4 IU/L; p = 0.003). In cycles resulting in pregnancy, Epo level decreased 14 days after oocyte pick-up (OPU) and remained low thereafter. In cycles not resulting in pregnancy, Epo level increased again 35 days after OPU. Follicle fluid Epo concentration was 1.5 times higher than the serum concentration (median 15.4; 95% CI 10.4–19.2 IU/L vs. 10.2; 8.8–12.7; p = 0.006). There was no difference in circulating Epo concentration between early OHSS and uncomplicated IVF. Circulating Epo did not correlate with VEGF or HIF-1. Conclusions: Circulating Epo levels fluctuate during IVF cycle. We hypothesise this may suggest Epo's involvement in ovarian physiology and angiogenesis. However, Epo was not a clinical marker for OHSS. [ABSTRACT FROM AUTHOR]

Background: We previously reported a modification of the CUT&Tag method (NTU-CAT) that allows genome-wide histone modification analysis in individual preimplantation embryos. In the present study, NTU-CAT was further simplified by taking advantage of the Well-of-the-Well (WOW) system, which enables the processing of multiple embryos in a shorter time with less reagent and cell loss during the procedure (WOW-CUT&Tag, WOW-CAT). Results: WOW-CAT allowed histone modification profiling from not only a single blastocyst but also from a portion of it. WOW-CAT generated similar H3K4me3 profiles as NTU-CAT, but they were closer to the profiles produced by chromatin immunoprecipitation-sequencing, such as a valley-like trend and relatively lower false positive rates, indicating that WOW-CAT may attenuate the bias of Tn5 transposase to cut open chromatin regions. Simultaneous WOW-CAT of two halves of single blastocysts was conducted to analyze two different histone modifications (H3K4me3 and H3K27ac) within the same embryo. Furthermore, trophectoderm cells were biopsied and subjected to WOW-CAT in anticipation of preimplantation diagnosis of histone modifications. WOW-CAT allowed the monitoring of epigenetic modifications in the main body of the embryo. For example, analysis of H3K4me3 modifications of XIST and DDX3Y in trophectoderm biopsies could be used to sex embryos in combination with quantitative PCR, but without the need for deep sequencing. Conclusions: These results suggest the applicability of WOW-CAT for flexible epigenetic analysis of individual embryos in preimplantation epigenetic diagnosis. [ABSTRACT FROM AUTHOR]

Background: Assisted reproduction technology (ART) has advanced significantly, raising concerns regarding its impact on the secondary sex ratio (SSR), which is the sex ratio at birth in offspring. This study aimed to explore factors affecting SSR in singletons, singletons from twin gestation, and twins from twin gestation within the context of ART. Methods: A retrospective analysis was conducted on data from 8335 births involving 6,223 couples undergoing ART. Binary logistic regression assessed relationships between parental and embryonic factors and SSR in singletons and singletons from twin gestation. Multinomial logistic regression models were utilized to identify factors influencing SSR in twins from twin gestation. Results: Secondary infertility (OR = 1.164, 95% CI: 1.009–1.342), advanced paternal age (OR = 1.261, 95% CI: 1.038–1.534), and blastocyst embryo transfer (OR = 1.339, 95% CI: 1.030–1.742) were associated with an increased SSR, while frozen embryo transfer (FET) showed a negative association with SSR (OR = 0.738, 95% CI: 0.597–0.912) in singletons. A longer duration of gonadotropin (Gn) usage reduced SSR in singletons (OR = 0.961, 95% CI: 0.932–0.990) and singletons from twin gestation (OR = 0.906, 95% CI: 0.838–0.980). In singletons from twin gestation, male-induced infertility (OR = 2.208, 95% CI: 1.120–4.348) and higher Gn dosage (OR = 1.250, 95% CI: 1.010–1.548) were significantly associated with an increased SSR. Women aged > 35 years and intracytoplasmic sperm injection (ICSI) were associated with lower SSR (OR = 0.539, 95% CI: 0.293–0.990 and OR = 0.331, 95% CI: 0.158–0.690, respectively). In twins from twin gestation, paternal age exceeded maternal age (OR = 0.682, 95% CI: 0.492–0.945) and higher Gn dosage (OR = 0.837, 95% CI: 0.715–0.980) were associated with a higher proportion of male twins. Cleavage stage transfer (OR = 1.754, 95% CI: 1.133–2.716) resulted in a higher percentage of boy-girl twins compared to blastocyst transfer. Conclusion: This study demonstrates the complex interplay of various factors in determining the SSR in ART, highlighting the importance of considering infertility type, paternal age, fertilization method, embryo transfer stage, and Gn use duration when assessing SSR. Nevertheless, further research with a large sample size is necessary to confirm and expand upon the findings of this study. [ABSTRACT FROM AUTHOR]

Background: Various methods for ex utero culture systems have been explored. However, limitations remain regarding the in vitro culture platforms used before implanting mouse embryos and the normal development of mouse blastocysts in vitro. Furthermore, vascular niche support during mouse embryo development from embryonic day (E) 3.5 to E7.5 is unknown in vitro. Methods: This study established a three-dimensional (3D) "sandwich" vascular niche culture system with in vitro culture medium (IVCM) using human placenta perivascular stem cells (hPPSCs) and human umbilical vein endothelial cells (hUVECs) as supportive cells (which were seeded into the bottom layer of Matrigel) to test mouse embryos from E3.5 to E7.5 in vitro. The development rates and greatest diameters of mouse embryos from E3.5 to E7.5 were quantitatively determined using SPSS software statistics. Pluripotent markers and embryo transplantation were used to monitor mouse embryo quality and function in vivo. Results: Embryos in the IVCM + Cells (hPPSCs + hUVECs) group showed higher development rates and greater diameters at each stage than those in the IVCM group. Embryos in the IVCM + Cells group cultured to E5.5 morphologically resembled natural egg cylinders and expressed specific embryonic cell markers, including Oct4 and Nanog. These features were similar to those of embryos developed in vivo. After transplantation, the embryos were re-implanted in the internal uterus and continued to develop to a particular stage. Conclusions: The 3D in vitro culture system enabled embryo development from E3.5 to E7.5, and the vascularization microenvironment constructed by Matrigel, hPPSCs, and hUVECs significantly promoted the development of implanted embryos. This system allowed us to further study the physical and molecular mechanisms of embryo implantation in vitro. [ABSTRACT FROM AUTHOR]

Background: Embryo aneuploidy is a main of principal reason of pregnancy loss, in vitro fertilization (IVF) failure and birth defects in offspring. Previous researchs have demonstrated that Y chromosome AZFc microdeletion was associated with reproduction outcomes, however, the relationship between Y chromosome AZFc microdeletion and embryo aneuploidy remains unexplored. Methods: This retrospective cohort study enrolled 513 patients with 603 cycles in the reproductive center of Nanjing Maternity and Child Health Care Hospital from January 1, 2016 to June 30, 2022. The study cohort was divided into two groups: the AZFc microdeletion group, comprising 53 patients and 58 cycles, and the control group, comprising 460 patients and 545 cycles. Statistical methods including restricted cubic spline and generalized estimating equation (GEE) were employed to evaluate the relationship between Y chromosome AZFc microdeletion and embryo euploidy. Results: 294 and 2833 blastocysts were selected as AZFc microdeletion group and control group, respectively. Patients with Y chromosome AZFc microdeletion had significantly higher embryo aneuploid rate (33.0% vs. 27.3%, P < 0.05), lower rate of normal fertilization rate (81.5% vs. 90.3%, P < 0.05) and lower blastocysts formation rate (47.0% vs. 57.8%, P < 0.05) compared with the control group. However, no significant differences in pregnancy outcomes after euploid embryos transfer were observed between these two groups. Conclusions: Our study underscored the association between Y chromosome AZFc microdeletion and an elevated risk of embryo aneuploidy. Before the conventional intracytoplasmic sperm injection (ICSI) treatment, couples with Y chromosome AZFc microdeletion should be apprised of the heightened susceptibility to embryo aneuploidy. Preimplantation genetic testing for aneuploidy (PGT-A) should be introduced for selection. [ABSTRACT FROM AUTHOR]

Objective: Prior studies have shown controversial results on the vertical transmission of BK virus (BKV). The present study aimed to assess the possibility of BKV vertical transmission from mother to fetus in the product of conception (embryo, fetuses, and/or placentas) over the three stages of pregnancy. Results: Of the 26 placental studied tissues, 6 were in the first trimester, and none of which were positive. Only one out of the 13 (7.7%) placental materials in the second trimester was positive. Only one out of 7 (14%) placental materials of the third trimester was positive. There were cases that no virus was detected in their placental but BKV was detected in their other tissues. Among 26 conceptuses, 17 (65%) were negative for BKV and 9 (34.6%) were positive, 7/13 (54%) were positive in the second, and 2/7 (29%) were positive in the third trimester fetuses. BKV was most frequently detected in the liver (eight cases), heart (three cases), and placenta (2 cases). There were cases that no virus was detected in their placental but BKV was detected in their other tissues. [ABSTRACT FROM AUTHOR]

Background: Metabolic homeostasis is closely related to early impairment of cell fate determination and embryo development. The protein kinase mechanistic target of rapamycin (mTOR) is a key regulator of cellular metabolism in the body. Inhibition of mTOR signaling in early embryo causes postimplantation development failure, yet the mechanisms are still poorly understood. Methods: Pregnancy mice and preimplantation mouse embryo were treated with mTOR inhibitor in vivo and in vitro respectively, and subsequently examined the blastocyst formation, implantation, and post-implantation development. We used immunofluorescence staining, RNA-Seq smart2, and genome-wide bisulfite sequencing technologies to investigate the impact of mTOR inhibitors on the quality, cell fate determination, and molecular alterations in developing embryos. Results: We showed mTOR suppression during preimplantation decreases the rate of blastocyst formation and the competency of implantation, impairs the post implantation embryonic development. We discovered that blocking mTOR signaling negatively affected the transformation of 8-cell embryos into blastocysts and caused various deficiencies in blastocyst quality. These included problems with compromised trophectoderm cell differentiation, as well as disruptions in cell fate specification. mTOR suppression significantly affected the transcription and DNA methylation of embryos. Treatment with mTOR inhibitors increase lysosomal activation and disrupts the organization and dynamics of the actin cytoskeleton in blastocysts. Conclusions: These results demonstrate that mTOR plays a crucial role in 8-cell to blastocyst transition and safeguards embryo quality during early embryo development. [ABSTRACT FROM AUTHOR]

Background: Gene knock‐in (KI) in animal cells via homology‐directed repair (HDR) is an inefficient process, requiring a laborious work for screening from few modified cells. HDR tends to occur in the S and G2/M phases of cell cycle; therefore, strategies that enhance the proportion of cells in these specific phases could improve HDR efficiency. Results: We used various types of cell cycle inhibitors to synchronize the cell cycle in S and G2/M phases in order to investigate their effect on regulating CRISPR/Cas9-mediated HDR. Our results indicated that the four small molecules—docetaxel, irinotecan, nocodazole and mitomycin C—promoted CRISPR/Cas9-mediated KI with different homologous donor types in various animal cells. Moreover, the small molecule inhibitors enhanced KI in animal embryos. Molecular analysis identified common signal pathways activated during crosstalk between cell cycle and DNA repair. Synchronization of the cell cycle in the S and G2/M phases results in CDK1/CCNB1 protein accumulation, which can initiate the HDR process by activating HDR factors to facilitate effective end resection of CRISPR-cleaved double-strand breaks. We have demonstrated that augmenting protein levels of factors associated with the cell cycle via overexpression can facilitate KI in animal cells, consistent with the effect of small molecules. Conclusion: Small molecules that induce cell cycle synchronization in S and G2/M phases promote CRISPR/Cas9-mediated HDR efficiency in animal cells and embryos. Our research reveals the common molecular mechanisms that bridge cell cycle progression and HDR activity, which will inform further work to use HDR as an effective tool for preparing genetically modified animals or for gene therapy. [ABSTRACT FROM AUTHOR]

The article discusses a recent ruling by the Alabama Supreme Court that declared frozen embryos to be legally considered children. This ruling has raised concerns and created uncertainty in the field of in vitro fertilization (IVF), as it potentially exposes healthcare professionals and facilities to liability and criminal prosecution if embryos are damaged or destroyed during the IVF process. The ruling is seen as an overreach of authority and a threat to secular democracy, as it combines theocracy and law without separation of church and state. However, it is predicted that this ruling will not significantly impact the practice of IVF in the long run, as it is an essential therapy for infertility and an important means of family building. The article emphasizes the need for advocacy and political engagement to protect reproductive rights in the face of future challenges. [Extracted from the article]

CRISPR/Cas9, a highly versatile genome-editing tool, has garnered significant attention in recent years. Despite the unique characteristics of oocytes and early embryos compared to other cell types, this technology has been increasing used in mammalian reproduction. In this comprehensive review, we elucidate the fundamental principles of CRISPR/Cas9-related methodologies and explore their wide-ranging applications in deciphering molecular intricacies during oocyte and early embryo development as well as in addressing associated diseases. However, it is imperative to acknowledge the limitations inherent to these technologies, including the potential for off-target effects, as well as the ethical concerns surrounding the manipulation of human embryos. Thus, a judicious and thoughtful approach is warranted. Regardless of these challenges, CRISPR/Cas9 technology undeniably represents a formidable tool for genome and epigenome manipulation within oocytes and early embryos. Continuous refinements in this field are poised to fortify its future prospects and applications. [ABSTRACT FROM AUTHOR]

Key message: We have examined thermal control for embryo growth and seed germination in European ash. Using historical data (span of 36 years) of embryo index in matured seeds, we showed that maturation is not influenced by the annual average temperature. Secondary dormancy in ash's seeds is easily induced, and the thermal window for both embryo growth and seed germination was between 10 and 20 °C. Context: Seed germination plays a crucial role in establishing plant communities. Ash seeds are characterized by deep morphophysiological dormancy, usually overcome by 32 weeks of stratification. Observed climate changes might disrupt ash's thermal window for embryo growth and seed germination. Thus, proper examination of seed adaptability can be important for forest management. Aims: We investigated the effect of temperature on the embryo index (ratio between embryo and seed length) as well as embryo growth and germination of European ash (Fraxinus excelsior L.) seeds under different thermal conditions. Methods: We used seeds collected at the same site in Poland between 1985 and 2021 to first analyze the influence of the growing season on the initial embryo index in mature seeds. Subsequently, we examined embryo growth and germination of seeds at constant temperatures of 3°, 10°, 15°, 20°, 25°, and 30 °C from seeds collected in 2021. Using the modeled rate of embryo growth and germination, we predicted the cardinal temperatures for those processes in ash seeds. Results: The embryo index of ash seeds varied; however, there was no relationship between the index and year of seed collection. The base temperature for embryo growth was 1.4 ± 1.1 °C, with a ceiling temperature of 29.9 ± 2.0 °C and an optimal temperature of 15.7 ± 0.3 °C. The base temperature for germination was at − 1.8 ± 0.3 °C. Due to the induction of secondary dormancy, the ceiling temperature of germination was impossible to establish. Conclusions: The initial embryo index of mature ash seeds seems to be an individual trait and was not affected by local average annual temperatures. The most effective temperature ranges for embryo growth and germination were similar, however narrow (between 10 and 15 °C). Temperatures above 20 °C inducted secondary dormancy, hampering embryo growth as well as germination. Such a narrow thermal window for embryo growth can potentially act as a limiting factor for the natural regeneration of ash seeds. [ABSTRACT FROM AUTHOR]

Background: In the current context of global warming, thermal manipulation of avian embryos has received increasing attention as a strategy to promote heat tolerance in avian species by simply increasing the egg incubation temperature. However, because of their likely epigenetic origin, thermal manipulation effects may last more than one generation with consequences for the poultry industry. In this work, a multigenerational and transgenerational analysis of thermal manipulation during embryogenesis was performed to uncover the long-term effects of such procedure. Results: Thermal manipulation repeated during 4 generations had an effect on hatchability, body weight, and weight of eggs laid in Japanese quails, with some effects increasing in importance over generations. Moreover, the effects on body weight and egg weight could be transmitted transgenerationally, suggesting non-genetic inheritance mechanisms. This hypothesis is reinforced by the observed reversion of the effect on growth after five unexposed generations. Interestingly, a beneficial effect of thermal manipulation on heat tolerance was observed a few days after hatching, but this effect was not transgenerational. Conclusions: Our multigenerational study showed that thermal conditioning of quail embryos has a beneficial effect on post-hatch heat tolerance hampered by transgenerational but reversible defects on growth. Assuming that no genetic variability underlies these changes, this study provides the first demonstration of epigenetic inheritance of traits induced by environmental temperature modification associated with long-term impacts in an avian species. [ABSTRACT FROM AUTHOR]

Background: Previous studies looked into the connections between pregnancy and the Zona Pellucida (ZP) thickness and Zona Pellucida Thickness Variation (ZPTV), as well as the embryo's radius, circumference, perimeter and global symmetry. However, no research has linked embryo implantation and pregnancy to the percentage of ZP thinning, the reduction in ooplasm volume, and the increase in perivitelline space (PVS) volume. Our objective is to correlate the percentage of ZP thinning, the percentage of ooplasm volume shrinkage and the percentage of PVS increase to the implantation. These data will be used for embryo selection as well as it can be put into a software that will assist embryo selection. Materials and methods: Retrospective study included 281 patients, all of them had 2 embryos transferred, 149 patients got pregnant with two gestation sacs and 132 patients did not get pregnant. All of the transferred embryos had the ZP thickness measured several times from time of ICSI till Embryo Transfer (ET), the ooplasm volume was calculated from time of ICSI till two Pronuclei (2PN) fading and the PVS was calculated from the ICSI time till the 2PN fading. Results: The first characteristic is the change in the average ZP thickness that decreased by 32.7% + 5.3% at 70 h for the implanted embryos (Group 1) versus 23.6% + 4.8% for non-implanted embryos (Group 2) p = 0.000. The second characteristic is the average reduction in the volume of the ooplasm which is 20.5% + 4.3% in Group 1 versus 15.1% + 5.2% in Group 2, p = 0.000. The third characteristic is the increase in the volume of the PVS which was 38.1% + 7.6% in Group 1 versus 31.6% + 9.7% in Group 2 p = 0.000. Conclusion: The implanted embryos showed higher percent of ZP thinning, higher percent of ooplasm reduction and higher percent of PVS increase. [ABSTRACT FROM AUTHOR]

Background: Studies show conflicting results on neonatal outcomes following embryo biopsy for PGT, primarily due to small sample sizes and/or heterogeneity in the timing of embryo biopsy (day 3; EBD3 or day 5/6; EBD5) and type of embryo transfer. Even fewer data exist on the impact on children's health beyond the neonatal period. This study aimed to explore outcomes in children born after EBD3 or EBD5 followed by fresh (FRESH) or frozen-thawed embryo transfer (FET). Methods: This single-centre cohort study compared birth data of 630 children after EBD3, of 222 EBD5 and of 1532 after non-biopsied embryo transfers performed between 2014 and 2018. Follow-up data on growth were available for 426, 131 and 662 children, respectively. Results: Embryo biopsy, either at EBD3 or EBD5 in FET and FRESH cycles did not negatively affect anthropometry at birth, infancy or childhood compared to outcomes in non-biopsied FET and FRESH cycles. While there was no adverse effect of the timing of embryo biopsy (EBD3 versus EBD5), children born after EBD3 followed by FET had larger sizes at birth, but not thereafter, than children born after EBD3 followed by FRESH. Reassuringly, weight and height gain, proportions of major congenital malformations, developmental problems, hospital admissions and surgical interventions were similar between comparison groups. Conclusion: Our study indicated that neither EBD3 nor EBD5 followed by FRESH or FET had a negative impact on anthropometry and on health outcomes up to 2 years of age. [ABSTRACT FROM AUTHOR]

Background: Previous studies have reported that after laparoscopic cystectomy of ovarial endometrioma, the ovarian response to gonadotropin (Gn) significantly decreased. However, for patients with diminished ovarian reserve (DOR) after ovarian surgery, how to choose the most appropriate controlled ovarian hyperstimulation protocol has not been concluded. Compared with the traditional agonist regimen, the gonadotropin (Gn)-releasing hormone (GnRH) antagonist, microstimulation, and progestin-primed ovarian stimulation (PPOS) protocols are simple to operate and have a shorter cycle, which are often used in patients with DOR. So the purpose of our study is to compare the assisted reproductive outcomes of these three controlled ovarian hyperstimulation protocols in patients with DOR following laparoscopic cystectomy of ovarial endometrioma. Methods: In this retrospective cohort study, 89 patients with DOR who had undergone in vitro fertilisation/intracytoplasmic sperm injection at the Department of Reproductive Medicine at the Third Affiliated Hospital of Zhengzhou University from 1 to 2018 to 31 December 2020 were included. According to the controlled ovarian hyperstimulation protocols employed, the patients were divided into GnRH antagonist (38 patients), PPOS (27 patients), and microstimulation (24 patients) groups. The basic data and clinical outcomes of the three groups were compared. The main outcome measure was the cumulative live birth rate. Results: No significant differences in the age of the female patients and their spouses and female patients' body mass index and basal endocrine levels (follicle-stimulating hormone and oestradiol) were noted among the three groups (P > 0.05). The GnRH antagonist group had higher antral follicle counts, greater endometrial thickness on the human chorionic Gn injection day, greater number of oocytes retrieved, and higher two pronuclear embryo counts than did the other two groups. However, the starting dosage of Gn was lower in the GnRH antagonist group than in the other two groups. The microstimulation group had a significantly higher oocyte output rate and high-quality embryo rate than did the other two groups (P < 0.05). No significant differences in the total dosage of Gn, cumulative pregnancy rate, cumulative live birth rate, viable embryo rate, and blastocyst formation rate were observed among the three groups (P > 0.05). Conclusion: In conclusion, for patients aged under 40 years who experienced DOR after laparoscopic cystectomy of ovarial endometrioma, GnRH antagonist protocol and PPOS protocol can obtain better ovulation induction outcomes and cumulative live birth rate than microstimulation protocol, and are more suitable ovulation induction protocols. [ABSTRACT FROM AUTHOR]

Recent advances in methods to culture pluripotent stem cells to model human development have resulted in entities that increasingly have recapitulated advanced stages of early embryo development. These entities, referred to by numerous terms such as embryoids, are becoming more sophisticated and could resemble human embryos ever more closely as research progresses. This paper reports a systematic review of the ethical, legal, regulatory, and policy questions and concerns found in the literature concerning human embryoid research published from 2016 to 2022. We identified 56 papers that use 53 distinct names or terms to refer to embryoids and four broad categories of ethical, legal, regulatory, or policy considerations in the literature: research justifications/benefits, ethical significance or moral status, permissible use, and regulatory and oversight challenges. Analyzing the full range of issues is a critical step toward fostering more robust ethical, legal, and social implications research in this emerging area and toward developing appropriate oversight. [ABSTRACT FROM AUTHOR]

Background: Cryopreservation of embryos plays a major role in the in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatment. However, the storage condition of the cryopreserved embryo can change temporarily due to repeated retrieval of the embryo from the liquid nitrogen (LN2) tank during the practical application during cryopreservation. Whether the implantation potential of a cryopreserved embryo will be damaged when the cane containing it is temporarily exposed to air due to the transfer between the LN2 tank and LN2 container is yet to be elucidated. Also, whether the exposed-to-air frequency (EAF) of cryopreserved embryos influences the clinical outcomes is unclear. Objective: To investigate whether the EAF of cryopreserved embryo affects the clinical outcomes of vitrified-warmed embryo transfer. Methods: A total of 9200 vitrified-warmed embryo transfer cycles were included in this study. All cycles were divided into five groups according to different EAFs (2, 4, 6, 8, or ≥ 10). Post-warming survival rates and clinical outcomes, including implantation, clinical pregnancy and live birth rates were investigated. Kruskal–Wallis test and Pearson's chi-squared tests were used to compare the patient characteristics and clinical outcomes among the five groups. Furthermore, multivariate logistic regression analyses were conducted to investigate the association between EAF and clinical outcomes. Results: No significant differences were observed in the positive HCG rate, implantation rate and live birth rate (P > 0.05) among five EAF groups with respect to D3 embryo, D5 blastocyst and D6 blastocyst. Post-warmed survival rate of D3 embryos (P = 0.015) differed significantly among the five EAF groups, but it was not EAF-dependent. Although clinical pregnancy was different among the five groups with respect to D5 blastocyst (P = 0.042), multivariate logistic regression analysis adjusted for confounding variables suggested that EAF did not adversely affect clinical pregnancy or live birth. Conclusion: These findings indicated that human vitrified embryos in the open system could be repeatedly retrieved from the LN2 tank without affecting the implantation potential of the embryo. [ABSTRACT FROM AUTHOR]

Background: During mammalian pre-implantation embryonic development (PED), the process of maternal-to-zygote transition (MZT) is well orchestrated by epigenetic modification and gene sequential expression, and it is related to the embryonic genome activation (EGA). During MZT, the embryos are sensitive to the environment and easy to arrest at this stage in vitro. However, the timing and regulation mechanism of EGA in buffaloes remain obscure. Results: Buffalo pre-implantation embryos were subjected to trace cell based RNA-seq and whole-genome bisulfite sequencing (WGBS) to draw landscapes of transcription and DNA-methylation. Four typical developmental steps were classified during buffalo PED. Buffalo major EGA was identified at the 16-cell stage by the comprehensive analysis of gene expression and DNA methylation dynamics. By weighted gene co-expression network analysis, stage-specific modules were identified during buffalo maternal-to-zygotic transition, and key signaling pathways and biological process events were further revealed. Programmed and continuous activation of these pathways was necessary for success of buffalo EGA. In addition, the hub gene, CDK1, was identified to play a critical role in buffalo EGA. Conclusions: Our study provides a landscape of transcription and DNA methylation in buffalo PED and reveals deeply the molecular mechanism of the buffalo EGA and genetic programming during buffalo MZT. It will lay a foundation for improving the in vitro development of buffalo embryos. [ABSTRACT FROM AUTHOR]

Telomeres are repetitive DNA sequences at eukaryotic chromosome ends and function in maintaining genome integrity and stability. These unique structures undergo shortening due to various factors including biological aging, consecutive DNA replication, oxidative stress, and genotoxic agents. Shortened telomeres can be lengthened by the enzyme telomerase and alternative lengthening of telomeres in germ cells, early embryos, stem cells, and activated lymphocytes. If telomeres reach to critical length, it may lead to genomic instability, chromosome segregation defects, aneuploidy, and apoptosis. These phenotypes also occur in the oocytes and early embryos, produced using assisted reproductive technologies (ARTs). Thus, a number of studies have examined the potential effects of ART applications such as ovarian stimulation, culture conditions, and cryopreservation procedures on telomeres. Herein, we comprehensively reviewed impacts of these applications on telomere length and telomerase activity in ART-derived oocytes and embryos. Further, we discussed use of these parameters in ART centers as a biomarker in determining oocyte and embryo quality. [ABSTRACT FROM AUTHOR]

Background: Conserved polycomb repressive complex 2 (PRC2) mediates H3K27me3 to direct transcriptional repression and has a key role in cell fate determination and cell differentiation in both animals and plants. PRC2 subunits have undergone independent multiplication and functional divergence in higher plants. However, relevant information is still absent in gymnosperms. Results: To launch gymnosperm PRC2 research, we identified and cloned the PRC2 core component genes in the conifer model species Picea abies, including one Esc/FIE homolog PaFIE, two p55/MSI homologs PaMSI1a and PaMSI1b, two E(z) homologs PaKMT6A2 and PaKMT6A4, a Su(z)12 homolog PaEMF2 and a PaEMF2-like fragment. Phylogenetic and protein domain analyses were conducted. The Esc/FIE homologs were highly conserved in the land plant, except the monocots. The other gymnospermous PRC2 subunits underwent independent evolution with angiospermous species to different extents. The relative transcript levels of these genes were measured in endosperm and zygotic and somatic embryos at different developmental stages. The obtained results proposed the involvement of PaMSI1b and PaKMT6A4 in embryogenesis and PaKMT6A2 and PaEMF2 in the transition from embryos to seedlings. The PaEMF2-like fragment was predominantly expressed in the endosperm but not in the embryo. In addition, immunohistochemistry assay showed that H3K27me3 deposits were generally enriched at meristem regions during seed development in P. abies. Conclusions: This study reports the first characterization of the PRC2 core component genes in the coniferous species P. abies. Our work may enable a deeper understanding of the cell reprogramming process during seed and embryo development and may guide further research on embryonic potential and development in conifers. [ABSTRACT FROM AUTHOR]

Background: The Caribbean spiny lobster Panulirus argus is heavily fished throughout its Greater Caribbean and Gulf of Mexico distribution, suggesting a heightened susceptibility to a fisheries collapse. In 2017, a nemertean worm, Carcinonemertes conanobrieni was described from ovigerous females of P. argus in Florida, USA. A year later, the presence of the same egg predator was recorded along the southern Caribbean coast (Colombia). The effect of this egg predator on the reproductive performance, including fecundity, embryo mortality, and reproductive output, of its host is unknown. This study tested whether C. conanobrieni affects embryo mortality, fecundity, and reproductive output in brooding females of P. argus. Results: Artisan fishers caught 90 ovigerous lobsters near Pueblo Viejo, Magdalena, Colombia. Each ovigerous female was examined for the presence/absence of the egg predator. Lobster egg mortality (%), fecundity (nº eggs female−1), and reproductive output (%) were estimated. Prevalence of C. conanobrieni in the studied population was 87.78%. The mean intensity of C. conanobrieni (all life stages) in the population was 11.68 (± 1.98) egg predators per brood mass sample. Infected females brooding late-stage embryos exhibited lower fecundity, lower reproductive performance values, and higher embryo mortality compared to infected females brooding early-stage embryos. Embryo stage and worm infection level negatively impacted fecundity and reproductive output. Worm infection level and the number of adult nemertean worms also negatively affected embryo mortality. Conclusions: These results demonstrate an adverse effect of C. conanobrieni on the reproductive performance of P. argus. The interactive impact of this egg predator, natural stressors, and anthropogenic stressors on individual P. argus reproductive performance could facilitate losses at large-scale fisheries levels. [ABSTRACT FROM AUTHOR]

Background: Currently, optimal method of ovarian stimulation (OS) to in-vitro fertilization (IVF) in the patients with polycystic ovarian syndrome (PCOS) is unknown. The present research aims to study the efficiency of minimal-OS method in treatment of infertile patients with PCOS and also the effect of gonadotropin type (recombinant FSH (r-FSH) vs. urinary Human menopausal gonadotropin (u-HMG)) on treatment cycles with GnRH-antagonist. Methods: In this randomized controlled trial, a total of 120 eligible patients were randomly allocated into four groups of OS to IVF: minimal-OS with r-FSH, minimal-OS with u-HMG, mild-OS with r-FSH and mild-OS with u-HMG. IVF outcomes of groups were analyzed statically. Results: The statistical analysis showed that there were significant differences among groups regarding stimulation duration (p < 0.0001), number of retrieved oocytes (p < 0.0001), number of obtained embryos (p < 0.0001). There were no statistically significant differences in fertilization rate (p = 0.289) and implantation rate (p = 0.757) among our participants. There were also significant differences among these four groups in terms of clinical pregnancy rate (/ET and /cycles) (p < 0.0001, p = 0.021, respectively) and live birth rate/cycles (p < 0.0001). Also cases of freeze all embryos due to prevention of ovarian hyper stimulation syndrome (OHSS) (p = 0.004). Conclusions: On the basis of present results the minimal-OS with u-HMG may be one of optimal methods of control OS in the patients with PCOS in respect to serum levels of estradiol on the day of triggering final oocyte maturation, total dose of prescribed gonadotropin, the optimal number of oocytes and embryos obtained, rate of clinical pregnancy and the incidence of OHSS risk. Trial registration: NCT, NCT03876145. Registered 15/03/2019. Retrospectively registered, http://www.clinicaltrial.gov/ NCT03876145. [ABSTRACT FROM AUTHOR]

Background: Metabolites in spent embryo culture medium correlate with the embryo's viability. However, there is no widely accepted method using metabolite dada to predict successful implantation. We sought to combine metabolomic profiling of spent embryo culture medium and clinical variables to create an implantation prediction model as an adjunct to morphological screening of day 3 embryos. Methods: This investigation was a prospective, nested case-control study. Forty-two day 3 embryos from 34 patients were transferred, and the spent embryo culture medium was collected. Twenty-two embryos implanted successfully, and the others failed. Metabolites in the medium relevant to implantation were detected and measured by Liquid Chromatography-Mass Spectrometry. Clinical signatures relevant to embryo implantation were subjected to univariate analysis to select candidates for a prediction model. Multivariate logistical regression of the clinical and metabolomic candidates was used to construct a prediction model for embryo implantation potential. Results: The levels of 13 metabolites were significantly different between the successful and failed groups, among which five were most relevant and interpretable selected by Least Absolute Shrinkage and Selection Operator regression analysis. None of the clinical variables significantly affected day 3 embryo implantation. The most relevant and interpretable set of metabolites was used to construct a prediction model for day 3 embryo implantation potential with an accuracy of 0.88. Conclusions: Day 3 embryos'implantation potential could be noninvasively predicted by the spent embryo culture medium's metabolites measured by LC-MS. This approach may become a useful adjunct to morphological evaluation of day 3 embryos. [ABSTRACT FROM AUTHOR]

Background: In the course of animal developmental processes, various tissues are differentiated through complex interactions within the gene regulatory network. As a general concept, differentiation has been considered to be the endpoint of specification processes. Previous works followed this view and provided a genetic control scheme of differentiation in sea urchin embryos: early specification genes generate distinct regulatory territories in an embryo to express a small set of differentiation driver genes; these genes eventually stimulate the expression of tissue-specific effector genes, which provide biological identity to differentiated cells, in each region. However, some tissue-specific effector genes begin to be expressed in parallel with the expression onset of early specification genes, raising questions about the simplistic regulatory scheme of tissue-specific effector gene expression and the current concept of differentiation itself. Results: Here, we examined the dynamics of effector gene expression patterns during sea urchin embryogenesis. Our transcriptome-based analysis indicated that many tissue-specific effector genes begin to be expressed and accumulated along with the advancing specification GRN in the distinct cell lineages of embryos. Moreover, we found that the expression of some of the tissue-specific effector genes commences before cell lineage segregation occurs. Conclusions: Based on this finding, we propose that the expression onset of tissue-specific effector genes is controlled more dynamically than suggested in the previously proposed simplistic regulation scheme. Thus, we suggest that differentiation should be conceptualized as a seamless process of accumulation of effector expression along with the advancing specification GRN. This pattern of effector gene expression may have interesting implications for the evolution of novel cell types. [ABSTRACT FROM AUTHOR]