bims-mazytr 2025-08-03 papers

bims-mazytr

Biomed News

on Maternal‐to‐zygotic transition

Issue of 2025–08–03
ten papers selected by
川一刀

Nuclear Tracking During Early Embryonic Divisions.
Highly Multiplexed Immunofluorescence Imaging of Mouse Oocytes.
RNA-Fluorescence In Situ Hybridization for the Detection of RNA in Mammalian Oocytes and Embryos.
Optical Coherence Microscopy for Oocyte Imaging.
Micro magnetic resonance spectroscopy for noninvasive metabolic screening of mammalian embryos and oocytes.
Live imaging endogenous transcription factor dynamics reveals mechanisms of epiblast and primitive endoderm fate segregation.
Electron Microscopy of Human Oocytes.
Ca2+-driven cytoplasmic backflow ensures spindle anchoring in fertilized mouse eggs.
PAL-AI reveals genetic determinants that control poly(A)-tail length during oocyte maturation, with relevance to human fertility.
Deep-learning model for embryo selection using time-lapse imaging of matched high-quality embryos.

Methods Mol Biol. 2025 ;2946 151-161

Nuclear Tracking During Early Embryonic Divisions.

Yunan Ye, Hayden A Homer.

  Positioning of the largest organelle, the nucleus, provides important cues for the geometry of cell divisions and cell fate determination. A primary function of nuclear positioning is to determine the spindle position and cleavage plane localization during cell division. Thus, centrally located nuclei lead to symmetrical divisions typical of mitosis, and off-center spindles culminate in symmetry-breaking as in mammalian oocytes. More recently, nuclear positioning has also been implicated in regulating mechanotransduction and gene expression in mouse oocytes. In human embryos, equal blastomere size arising from symmetrical cleavage-stage divisions is associated with favorable developmental outcomes, whereas off-center nuclear positioning affects division symmetry that can derail development. Therefore, investigating the mechanisms involved in nuclear positioning is critical for understanding embryo biology and the basis for clinical infertility. Here, we describe a method involving confocal imaging for performing nuclear tracking during early embryonic divisions in mouse embryos.

Keywords:  Embryo; Nuclear positioning; Nuclear tracking; Oocyte; Time-lapse imaging; TrackMate

DOI:  https://doi.org/10.1007/978-1-0716-4658-8_12
Methods Mol Biol. 2025 ;2946 163-173

Highly Multiplexed Immunofluorescence Imaging of Mouse Oocytes.

Cyprien Noble, Typhaine Esteves, Adel Al Jord.

  Highly multiplexed immunofluorescence imaging methods advanced our understanding of biology across scales, from tissues down to molecules. By enabling the visualization of tens of proteins in a single sample, highly multiplexed imaging is especially relevant for scarce and challenging biospecimens like mammalian oocytes. However, most methods remain relatively costly and complex to implement. Here, we provide a cost-effective simple protocol, based on iterative indirect immunofluorescence imaging (4i), allowing to capture the distribution and abundance of tens of proteins in a single mouse oocyte. Our approach is adaptable to other mammalian oocytes or analogously large non-adherent cells like the early embryo.

Keywords:  4i; Germ cells; Iterative indirect immunofluorescence imaging; Mammalian oocyte; Meiosis I; Multiplex immunofluorescence; Multiplexed imaging; Non-adherent cells; Oocyte

DOI:  https://doi.org/10.1007/978-1-0716-4658-8_13
Methods Mol Biol. 2025 ;2946 57-68

RNA-Fluorescence In Situ Hybridization for the Detection of RNA in Mammalian Oocytes and Embryos.

Denisa Jansova, Fatima J Berro, Andrej Susor.

  RNA metabolism plays an essential role in the development of both oocytes and embryos. Their dependence on stored maternal transcripts is due to a long period of silenced transcription in which the oocyte undergoes meiotic maturation and fertilization. Although maternal RNAs are unusually stable, they should be replaced by "zygotic" transcripts during the transition from oocyte to zygote. Analysis of ncRNA and mRNA distribution in the oocyte can provide clues to the fate of RNA in the single-cell environment.Our work focuses on the visualization of the subcellular distribution of specific RNAs in mammalian oocytes and early embryos. The localization of many RNAs in the oocyte and embryo is still not fully understood. In this chapter, we describe an optimized protocol for RNAscope, a type of RNA fluorescence in situ hybridization (RNA FISH), which is a valuable technique for exploring mammalian oocytes and embryos. The method is based on a specific probe design strategy that enables signal amplification together with simultaneous background suppression. Additionally, it is suitable for quantitative spatio-temporal analysis of specific RNA transcripts. RNAscope, a simple and a reliable protocol, contributes to advancing our understanding of RNA biology in the context of germ cell development.

Keywords:  Embryo; Oocyte; RNA FISH; Single-molecule imaging; mRNA

DOI:  https://doi.org/10.1007/978-1-0716-4658-8_5
Methods Mol Biol. 2025 ;2946 87-102

Optical Coherence Microscopy for Oocyte Imaging.

Anna Ajduk, Szymon Tamborski, Maciej Szkulmowski.

  Optical coherence microscopy (OCM) is a novel approach to fluorophore-free 3D live imaging of cells, particularly mammalian oocytes and embryos. It allows for 3D high-resolution visualization of the intracellular architecture: nuclei with nucleoli, metaphase spindles, and networks of membranous structures. Moreover, as it is compatible with time-lapse imaging, it enables monitoring and quantitative analysis of the dynamic behavior of these organelles over time. Importantly, OCM, when imaging settings are properly optimized, is safe for oocytes and embryos and does not negatively affect their developmental capabilities. Therefore, OCM is an interesting alternative to currently used imaging techniques, not only in basic research but also in clinical applications. In the present chapter, we describe the main principles of spectral OCM and show how this technique can be applied to visualize mouse oocytes.

Keywords:  Embryo; Image processing; Imaging; Mouse; Nucleolus; Nucleus; Oocyte; Optical coherence microscopy; Spindle

DOI:  https://doi.org/10.1007/978-1-0716-4658-8_7
Proc Natl Acad Sci U S A. 2025 Aug 05. 122(31): e2424459122

Micro magnetic resonance spectroscopy for noninvasive metabolic screening of mammalian embryos and oocytes.

Giulia Sivelli, Arthur Barakat, Kathryn B Marable, Guillaume Gruet, Serena L Bitetti, Barry Behr, Valentina Lodde, Alberto Maria Luciano, Carolina Herrera, Michelle Blom, Marco Grisi.

  Analyzing cellular health and metabolism without compromising cell integrity is a major challenge. We present a noninvasive technique using micro magnetic resonance spectroscopy (micro MRS) for nondestructive metabolic fingerprinting at the single-cell scale. This is an application of micro MRS to bovine preimplantation embryos (~8 cells) and oocytes (single cell), with measurements performed on a total of over 150 samples. Among various applications, this method holds significant potential for assisted reproductive technologies (ART), where metabolic assessments of preimplantation embryos could improve treatment outcomes. Early results indicate that classification models using micro MRS data effectively distinguish embryos with high developmental potential and show correlation with oocytes maturity. Furthermore, a multigenerational safety study in a mouse model revealed no adverse effects from embryo exposure to static magnetic field. These findings indicate that micro MRS is a promising, safe tool for assessing embryo metabolism, potentially improving the efficiency and outcomes of ART.

Keywords:  Magnetic Resonance Spectroscopy; metabolic fingerprinting; micro MRS; non-invasive embryo screening; single-cell MRS

DOI:  https://doi.org/10.1073/pnas.2424459122
Curr Biol. 2025 Jul 26. pii: S0960-9822(25)00897-8. [Epub ahead of print]

Live imaging endogenous transcription factor dynamics reveals mechanisms of epiblast and primitive endoderm fate segregation.

Rebecca P Kim-Yip, David Denberg, Denis F Faerberg, Hayden Nunley, Isabella Leite, Madeleine Chalifoux, Bradley Joyce, Jared Toettcher, Bin Gu, Eszter Posfai.

  The segregation of the epiblast (EPI) and primitive endoderm (PE) cell types in the preimplantation mouse embryo is not only a crucial decision that sets aside the precursors of the embryo proper from extraembryonic cells, respectively, but also has served as a central model to study a key concept in mammalian development: how much of developmental patterning is predetermined vs. stochastically emergent. Here, we address this question by quantitative live imaging of multiple endogenously tagged transcription factors key to this fate decision and trace their dynamics at a single-cell resolution through the formation of EPI and PE cell fates. Strikingly, we reveal an initial symmetry breaking event, the formation of a primary EPI cell lineage, and show that this is linked to the dynamics of the prior inner cell mass/trophectoderm fate decision through the expression of SOX2. This primary EPI lineage, through fibroblast growth factor (FGF) signaling, induces an increase in the transcription factor GATA6 in other inner cell mass cells, setting them on the course toward PE differentiation. Interestingly, this trajectory can switch during a defined developmental window, leading to the emergence of secondary EPI cells. Finally, we show that early expression levels of NANOG, which are seemingly stochastic, can bias whether a cell's trajectory switches to secondary EPI or continues as PE. Our data give unique insight into how fate patterning is initiated and propagated during unperturbed embryonic development through the interplay of lineage-history-biased and stochastic cell-intrinsic molecular features, unifying previous models of EPI/PE segregation.

Keywords:  GATA6; ICM; NANOG; SOX2; blastocyst; epiblast; live imaging; mouse; preimplantation embryo; primitive endoderm

DOI:  https://doi.org/10.1016/j.cub.2025.07.031
Methods Mol Biol. 2025 ;2946 103-114

Electron Microscopy of Human Oocytes.

Zuzana Holubcová, Barbora Baďurová, Zuzana Trebichalská.

  Electron microscopy represents a powerful visualizing technique, capable of a million times magnification. Prior to imaging, biological samples must undergo complex preparation to withstand the exposition to electrons in the vacuum inside the electron microscope. Here, we describe a preparation technique allowing preservation of scarce and delicate human oocytes for ultrastructural investigation.

Keywords:  Electron microscopy; Focused ion beam scanning electron microscopy; Human oocytes; Transmission electron microscopy; Ultrastructure

DOI:  https://doi.org/10.1007/978-1-0716-4658-8_8
Curr Biol. 2025 Jul 23. pii: S0960-9822(25)00859-0. [Epub ahead of print]

Ca2+-driven cytoplasmic backflow ensures spindle anchoring in fertilized mouse eggs.

Takaya Totsuka, Miho Ohsugi, Takashi Akera.

  Female meiosis is highly asymmetric, producing a large egg and a small polar body to preserve maternal storage essential for embryogenesis. To achieve asymmetric division, the egg spindle must maintain its cortical position until fertilization completes meiosis. In mice, fertilization triggers chromosome segregation, followed by spindle rotation to achieve the perpendicular orientation relative to the cortex, leading to the extrusion of one set of chromosomes. However, it was unknown how the spindle maintains its cortical position while rotating. Here, we developed a high-resolution live-imaging method to investigate spindle dynamics during fertilization. Our results indicate that Ca2+ oscillations put the brakes on spindle rotation by transiently reversing cytoplasmic streaming and that this cytoplasmic backflow secures the spindle localization at the cortex. Mechanistically, Ca2+ oscillations drive cortical actomyosin contraction to induce the cytoplasmic backflow. Altogether, this work revealed a previously unknown role of Ca2+ oscillations in maintaining spindle position, ensuring the highly asymmetric divisions inherent to female meiosis.

Keywords:  Ca(2+) oscillations; actomyosin contraction; cytoplasmic streaming; fertilization; live imaging; meiosis II; mouse egg; second polar body extrusion; spindle dynamics

DOI:  https://doi.org/10.1016/j.cub.2025.06.073
Nat Commun. 2025 Aug 01. 16(1): 7079

PAL-AI reveals genetic determinants that control poly(A)-tail length during oocyte maturation, with relevance to human fertility.

Kehui Xiang, David P Bartel.

In oocytes of mammals and other animals, gene regulation is mediated primarily through changes in poly(A)-tail length. Here, we introduce PAL-AI, an integrated neural network machine-learning model that accurately predicts tail-length changes in maturing oocytes of frogs and mammals. We show that PAL-AI learned known and previously unknown sequence elements and their contextual features that control poly(A)-tail length, enabling it to predict tail-length changes resulting from 3'-untranslated region single-nucleotide substitutions. It also predicted tail-length-mediated translational changes, allowing us to nominate genes important for oocyte maturation. When comparing predicted tail-length changes in human oocytes with genomic datasets of the All of Us Research Program and gnomAD, we found that genetic variants predicted to disrupt tail lengthening have been under negative selection in the human population, thereby linking mRNA tail lengthening to human female fertility.

DOI: https://doi.org/10.1038/s41467-025-62171-5
Sci Rep. 2025 Aug 01. 15(1): 28068

Deep-learning model for embryo selection using time-lapse imaging of matched high-quality embryos.

Lisa Boucret, Floris Chabrun, Magalie Boguenet, Pascal Reynier, Pierre-Emmanuel Bouet, Pascale May-Panloup.

  Time-lapse imaging and deep-learning algorithms are promising tools to assess the most viable embryos and improve embryo selection in IVF laboratories. Here, we developed and validated a deep learning model based on self-supervised contrastive learning. The model was developed with a new approach based on matched KID (Known Implantation Data) embryos derived from the same cohort of a stimulation cycle, both judged to be of good quality according to classical morphological criteria and morphokinetics, transferred fresh or frozen, but with a different implantation fate (clinical pregnancy vs. failure of implantation). We used self-supervised contrastive learning to train convolutional neural networks to ensure an unbiased and comprehensive learning of the morphokinetics features of the embryos, followed by a Siamese neural network fine-tuning and an XGBoost final prediction model to prevent overfitting. 1580 embryo videos of 460 patients were included between January 2020 and February 2023. With the knowledge of the implantation outcome of a previous transfer of an embryo derived from the same stimulation cycle, this model could predict the pregnancy outcome of the subsequent transfer with an AUC of 0.57. Without any knowledge of transfer history, the model achieved a satisfactory performance in predicting implantation (AUC = 0.64). This model could be considered as an adjunct tool for biologists to better select embryos and reduce the number of useless transfers per patient, when a cohort with several embryos classified as good quality by classical criteria is obtained.

Keywords:  Artificial intelligence; Deep learning; Embryo morphokinetics; Implantation; Machine learning; Time-lapse

DOI:  https://doi.org/10.1038/s41598-025-10531-y