Biology subjects

Oldak, B.

Publications and source records attributed to Oldak, B..

3 recordsLinked to original sources

bioRxiv2026

Reconstituting Mouse Embryogenesis Ex Utero from Gastrulation to Fetal Development Reveals Maternally Independent Metabolic Programs

Mammalian development takes place inside the maternal uterus, creating technological constraints that make difficult the study of embryogenesis in live developing embryos. A central challenge for understanding the role of metabolism in mammalian development is discriminating placental and uterine-regulated signals from embryo-intrinsic processes independent of maternal influence, a process that until now has remained inseparable during gastrulation and organogenesis1-3. Ex utero culture systems allowing continuous growth of embryos during pre-gastrulation to organogenesis4,5 offer a promising solution to this challenge. Here, we present optimized ex utero culture platforms that support faithful development of mouse embryos from gastrulation (embryonic day 6.5/7.5) through the fetal period (embryonic day [~]12.5) and harnessed these platforms for dissecting metabolic transitions in vivo during embryogenesis independently of uterus and placenta. We characterized the metabolome of in utero and ex utero whole embryos, fetal organs and culture medium between embryonic days E6.5 and E12.5 by liquid chromatography mass-spectrometry (LC-MS) metabolomics, isotope tracing, and single cell transcriptomics. These datasets present a comprehensive overview of the dynamic embryonic metabolism during gastrulation and organogenesis in utero and ex utero. This analysis revealed that the midgestational metabolic switch occurring at E10.5-E11.5 is faithfully recapitulated ex utero, indicating that this transition is intrinsically programmed in embryonic tissues and does not require direct maternal or placental cues. Notably, oxygen availability modulated the extent of this transition, but elevated oxygen was insufficient to induce it prematurely, demonstrating that the switch is developmentally timed and only partially environmental-responsive. We further harnessed the ex utero platform for identifying and perturbing a mitochondrial redox shift at E7.5-E8.5 that is critical for developmental progress after gastrulation. These findings uncover the remarkable metabolic plasticity of the mammalian embryo, demonstrating its capacity to sustain growth independently of maternal inputs from the establishment of the body plan through the onset of the fetal period. Moreover, they highlight the use of long-term ex utero culture as a unique framework for dissecting the mechanisms that shape embryogenesis under physiological and experimentally perturbed conditions, while functionally uncoupling embryonic programs from maternal and placental influences.

developmental biology↗

bioRxiv2023

Transgene-Free Ex Utero Derivation of A Human Post-Implantation Embryo Model Solely from Genetically Unmodified Naive PSCs

Our ability to study early human post-implantation development remains highly limited due to the ethical and technical challenges associated with intrauterine development of the human embryo after implantation. Despite the great progress made on human gastruloids, axioloids and in vitro cultured blastoids, such elegant models do not constitute an integrated Stem cell-derived Embryo Models (SEMs) that includes all the key extra-embryonic tissues of the early post-implantation human conceptus (e.g., hypoblast, yolk-sac, trophoblasts, amnion, and extraembryonic mesoderm), and thus, do not recapitulate post-implantation epiblast development within the context of these extra-embryonic compartments. Mouse naive pluripotent stem cells (PSCs) have recently been shown to give rise to embryonic and extra-embryonic stem cells capable of self-assembling into post-gastrulation mouse SEMs, while bypassing the blastocyst-like stage, and eventually initiating organogenesis ex utero. Here, we implement critical adaptations to extend these finding to humans, while using only genetically unmodified human naive PSCs, thus circumventing the need for ectopic expression of lineage promoting transgenes. Such integrated human SEMs recapitulate the organization of all known compartments of early post-implantation stage human embryos, including epiblast, hypoblast, extra-embryonic mesoderm, and trophoblast surrounding the latter layers. The organized human SEMs recapitulate key hallmarks of post-implantation stage embryogenesis up to 13-14 days post-fertilization (dpf, Carnegie stage 6a), such as bilaminar disk formation, epiblast lumenogenesis, amniogenesis, anterior-posterior symmetry breaking, PGC specification, primary and secondary yolk sac formation, and extra-embryonic mesoderm expansion that defines a chorionic cavity and a connective stalk. This new platform constitutes a tractable stem cell-based model for experimentally interrogating previously inaccessible windows of human peri- and early post-implantation development.

developmental biology↗

bioRxiv2020

Tripartite Inhibition of SRC-WNT-PKC Signalling Consolidates Human Naive Pluripotency

Different conditions have been devised to isolate MEK/ERK signalling independent human naive pluripotent stem cells (PSCs) that are distinct from conventional primed PSCs and better correspond to pre-implantation developmental stages. While the naive conditions described thus far endow human PSCs with different extents of naivety features, isolating human pluripotent cells that retain characteristics of ground state pluripotency while maintaining differentiation potential and genetic integrity, remains a major challenge. Here we engineer reporter systems that allow functional screening for conditions that can endow both the molecular and functional features expected from human naive pluripotency. We establish that simultaneous inhibition of SRC-NF{kappa}B, WNT/{beta}CATENIN and PKC signalling pathways is essential for enabling expansion of teratoma competent fully naive human PSCs in defined or xeno-free conditions. Divergent signalling and transcriptional requirements for maintaining naive pluripotency were found between mouse and human. Finally, we establish alternative naive conditions in which MEK/ERK inhibition is substituted with inhibition for NOTCH/RBPj signalling, which allow obtaining alternative human naive PSCs with diminished risk for loss of imprinting and deleterious global DNA hypomethylation. Our findings set a framework for the signalling foundations of human naive pluripotency and may advance its utilization in future translational applications. Highlights of key findingsO_LICombined inhibition of SRC, WNT and PKC signaling consolidates human naive pluripotency C_LIO_LIStable expansion of DNA/RNA methylation-independent and TGF/ACTIVIN-independent human naive PSCs C_LIO_LIOpposing roles for ACTIVIN and WNT/{beta}CATENIN signaling on mouse vs. human naive pluripotency C_LIO_LI2i and MEK/ERKi independent alternative human naive PSC conditions via inhibiting NOTCH/RBPj signaling C_LI

developmental biology↗