Generation of cynomolgus embryonic stem cell line (KP-CmESC-C1) from blastocysts produced by intracellular sperm injection.

Camizestrant-induced heart rate reduction is mediated via a sinoatrial node pacemaker channel mechanism.

Enhancing the utility of Sendai virus-based vectors through antiviral compound-mediated removal.

miR-671-5p-enriched exosomes derived from human embryonic stem cells under hypoxia balance oxidative stress homeostasis and macrophage reprogramming to alleviate Legg-Calvé-Perthes disease by targeting NOX2.

The Wnt signaling pathway plays a key role in regulating the pluripotent state of rbESCs and promoting their commitment to the primordial germ cell fate.

Polyazulene is an underrated conductive polymer characterized by its unique building blocks involving a fused-ring structure with an intrinsic dipole moment. It has been studied with a focus on the development of organic electronics, such as photovoltaic cells. However, the biological properties of polyazulene have not been reported to date. This study, the first of its kind, not only describes the properties of electrochemically and chemically synthesised polyazulenes and so-called true polyazulene, but also characterises their cytocompatibility. The physico-chemical properties of polyazulene powders and films were characterized by FTIR, Raman and UV-Vis spectroscopy, profilometry, scanning electron microscopy, voltammetry, van der Pauw conductivity measurement, and thermogravimetric analysis. Their cytocompatibility was explored using NIH/3T3 mouse embryonic fibroblasts, HaCaT human keratinocytes, and ES R1 mouse embryonic stem cells. Our results indicate that the tested polyazulenes exhibit an overall favourable short-term in vitro cytocompatibility with limitations varying according to the synthesis method and conditions. Electrochemically synthesized polyazulene formed continuous films with moderate conductivity under optimized deposition conditions, whereas chemically prepared films were inhomogeneous for reliable conductivity measurements. These differences in conductivity therefore primarily reflect variation in film morphology and deposition conditions rather than intrinsic bulk electronic properties of the respective polyazulene types. Is polyazulene cytocompatible? It depends This study presents the first biological evaluation of polyazulenes synthesized by various methods. It examines their physicochemical characteristics and cytocompatibility with multiple cell lines. The findings demonstrate that synthesis conditions influence both conductivity and biocompatibility. The work highlights polyazulene’s potential in biomedical applications and lays the groundwork for future exploration of its role in bioelectronic materials. • Azulene electrochemical polymerization gives conductive polyazulene films. • Chemical polymerization produces polyazulene types with distinct structures. • Films´ physico-chemical properties strongly depend on synthesis conditions. • Cytocompatibility of polyazulenes varies with synthesis method and processing conditions.

While current pluripotency models capture discrete embryonic stages, they inadequately resolve transitional states during peri-implantation development. Here, we establish rosette-formative intermediate stem cells (rfISCs) from mouse embryonic stem cells using the MEK inhibitor PD0325901, the Wnt inhibitor IWR1, and the PKA activator Forskolin. These cells exhibit transcriptomic/epigenetic profiles mirroring those of E5.0‒5.5 epiblasts, bridging rosette-stage and formative pluripotency. rfISCs demonstrate developmental bipotency, retaining in vitro germline differentiation capacity while generating germline-competent chimeras in vivo. Mechanistically, we identified opposing signaling axes that govern rfISC identification through the regulation of lineage priming: IWR1 stabilizes Tcf7l1 to drive Otx2-mediated rfISC specification and neural priming, whereas Forskolin activates PKA to induce Id1-dependent neural suppression. This creates a bistable regulatory circuit in which Otx2/Id1 synergy maintains pluripotency plasticity under MEK inhibition. Notably, rfISCs can be directly derived from E5.25 epiblasts, confirming their physiological relevance. Our work bridges a fundamental gap in pluripotency modeling by capturing the RSC-to-FSC transition through dynamic signaling equilibria.

Time- and dose-dependent toxicity profiles and molecular correspondence of nanoplastics in human embryonic stem cells (H9).

Nickel exposure disrupts epigenetic repression of developmental genes in mouse embryonic stem cells.

F-53B (6:2 Cl-PFESA), a major replacement for perfluorooctanesulfonate (PFOS), is frequently detected in human cord blood, yet its developmental neurotoxicity risks remain poorly characterized. This study establishes a quantitative testing strategy coupling in vitro phenotypic profiling, transcriptomics, and a probabilistic adverse outcome pathway-Bayesian network (AOP-BN) with pregnancy physiologically based pharmacokinetic (PBPK) modeling. Using human embryonic neural stem cells, we found that F-53B induced dose-dependent oxidative stress and mitochondrial dysfunction, resulting in compromised neurogenesis. Transcriptomics supported these phenotypic results. We derived a benchmark dose of 3.26 μmol/g of protein for learning and memory impairment and utilized AOP-BN to quantify the probability of adverse outcomes across exposure gradients. By coupling this framework with a pregnancy PBPK model, we estimated fetal brain concentrations of 0.09-14.66 ng/mL (Q5-Q95) based on human biomonitoring data. While these levels remain below the point of departure for downstream neurogenic defects, the narrow margins of exposure for early molecular events, specifically ROS elevation, indicate potential safety concerns. Consequently, this study identifies oxidative stress as a sensitive trigger for F-53B toxicity and demonstrates a robust, mechanistically anchored framework for human-relevant risk assessment of emerging PFAS.

Stable maintenance of MERVL-positive embryonic stem cells reveals sustained transcriptional programs and enhancer remodeling.

Stem cell pluripotency is shaped by culture microenvironment. Growth factors, cytokines, and oxygen tension are key regulators of self-renewal and pluripotency state. Here we evaluated the adaptation of bovine embryonic stem cells (bESC) derived on mouse embryonic fibroblasts (MEF) to feeder-free conditions and the effects of low oxygen (5% O2) on their molecular profile. Primed bESC established on MEF and 21% O2 (high oxygen) were adapted to 5% O2 (low oxygen) and to feeder-free conditions in vitronectin to generate four culture conditions: high and low O2 tension in MEF and feeder-free. Adaptation to feeder-free upregulated transcripts related to cell differentiation in bESC cultured in high compared to low oxygen. Transition of bESC on MEF from normoxia to 5% O2 did not alter cell morphology or growth. Interestingly, colony morphology was maintained when transitioning bESC from MEF into feeder-free in low oxygen. Furthermore, there were fewer transcriptomic changes in bESC grown on MEF compared to feeder-free in low compared to high oxygen. While low oxygen tension did not alter the pluripotency profile of bESC, it promoted transcriptional changes related to energetic metabolism and increased mitochondrial membrane potential. These findings emphasize the importance of oxygen tension for derivation, maintenance, and performance of ESC.

Developmental dynamics of skeletal muscle can be recapitulated in vitro from pig embryonic stem cells.

Mechanical responses are critical to elucidating the early-stage differentiation of human embryonic stem cells (hESCs), as these cells typically form cohesive clones and exhibit heterogeneous differentiation states. But mechano-regulatory mechanisms underlying the spatial heterogeneity of hESC clones during early differentiation remain poorly understood. Here, we investigated the role of cell-substrate adhesion and associated mechanotransductive pathways in governing the regional differentiation of hESCs into definitive endoderm (DE). At this early differentiation stage, H1 hESCs displayed spatial heterogeneity, with elevated DE marker expression at the periphery of individual clones compared to the interior. This pattern aligned well with a differential distribution of cellular traction forces, which was peaked at the periphery. Correlative spatial distributions of β1-integrin, phosphorylated FAK (p-FAK) and vinculin were observed, supporting the mechanical dominance of peripheral regions. Inhibition of β1-integrin or disruption of F-actin suppressed both traction force and DE differentiation capacity, primarily by inhibiting YAP nuclear translocation and thereby reducing spatial heterogeneity between peripheral and interior regions. The positive correlation between traction force and differentiation capacity was further validated by increasing cellular traction forces via culture on stiff substrates. This work highlights the pivotal role of biomechanical cues in early fate decisions of hESCs and provides insights into optimizing early differentiation through mechanical modulation. STATEMENT OF SIGNIFICANCE: This study defines a mechano-regulatory mechanism underlying spatial heterogeneity during early definitive endoderm (DE) differentiation of human embryonic stem cells (hESCs). Within self-organized H1 colonies, peripheral cells exhibit enhanced DE commitment driven by elevated traction forces transmitted through β1-integrin-F-actin-YAP axis. This spatial bias emerges intrinsically, even on mechanically uniform substrates, and is further modulated by substrate stiffness. By integrating single-cell transcriptomics with traction force microscopy and high-resolution imaging, we demonstrate that colony-scale mechanical compartmentalization governs early lineage specification. These findings advance understanding of integrin-mediated mechanotransduction in stem cell early differentiation and provide a mechanobiological framework for optimizing directed differentiation through controlled mechanical microenvironments.

Functional characterization of UHRF1 variants in facilitating DNA methylation.

Nonsense-mediated RNA decay (NMD) was originally discovered by virtue of its "quality control" function of degrading aberrant mRNAs with premature termination codons (PTCs). NMD was subsequently found to be a highly selective and conserved RNA turnover pathway that also degrades subsets of normal mRNAs harboring stop codons in specific contexts. The discovery that many normal mRNAs encoding full-length normal proteins are degraded by NMD has led to a search for biological functions for NMD. In this review, we focus on the evidence for NMD's roles in early embryonic development, nervous system development, spermatogenesis, thymic development, and other developmental processes in mice. NMD also has roles in stem cells, including dictating self-renewal vs. differentiation decisions in embryonic and neural stem cells. We also discuss evidence for NMD's roles in some adult functions, such as circadian rhythm and neuronal activities. Finally, we highlight NMD's causative roles in some human diseases and how therapeutic intervention of this critical pathway can be modeled in mice.

The three-dimensional (3D) organization of the genome is strongly influenced by interactions between chromatin and lamin proteins at the nuclear envelope. Here, we investigate the role of lamina-associated domains (LADs) in shaping genome architecture using coarse-grained polymer models of mouse embryonic fibroblasts and embryonic stem cells. By integrating genome-wide LAD maps from DamID assays, we simulate chromatin conformations with and without LAD attachment. Incorporating LAD-lamina interactions reproduces the experimentally observed radial chromatin distribution and reveals that LADs induce extensive long-range (70-120 Mbp) chromatin contacts beyond typical loops and topologically associating domains (TADs). We describe these contacts in terms of two limiting geometric scenarios: LAD crowding, in which peripheral tethering increases the proximity of nearby non-LAD regions to LADs, and LAD anchoring, in which lamina-bound LADs constrain neighboring chromatin positions. LAD-induced interactions were especially prominent in chromatin regions lacking architectural proteins such as CTCF, and were associated with lower gene density and reduced transcriptional activity. Together, these results suggest that LAD-lamina tethering reshapes long-range chromatin contact probabilities through boundary-driven effects and is associated with gene-poor, less transcriptionally active chromatin regions.

Oncogenic fusion transcription factors (TFs) frequently drive hematopoietic malignancies by altering gene expression in key developmental programs. TCF3::HLF is a fusion TF that characterizes a rare, treatment-resistant subtype of B cell acute lymphoblastic leukemia [t(17;19) TCF3::HLF-positive B-ALL]. Despite its clinical significance, the mechanisms by which TCF3::HLF induces leukemia are unclear. We used HiChIP mapping and genetic interference to analyze TCF3::HLF at the 3D genome level, revealing enhancer-promoter interactions that control gene activation or repression. Notably, TCF3::HLF directly regulates MEF2C expression through its enhancer, as interference disrupted MEF2C transcription and inhibited leukemia propagation. This disruption also diminished embryonal hematopoietic stem cell (HSC) gene signatures and restored mature HSC and B-lymphoid markers. These findings highlight MEF2C as a critical component of the transcriptional network reprogrammed by TCF3::HLF. Our study provides insight into how TCF3::HLF rewires the 3D genome to drive leukemia and serves as a resource for further exploration of the TCF3::HLF regulome.

The skin is densely innervated by peripheral sensory neurons that detect various stimuli through specialized nerve endings in the dermis and epidermis. In recessive dystrophic epidermolysis bullosa (RDEB), repeated skin injury disrupts epidermal nerve fibers, leading to neuropathic pain and reduced thermal sensitivity. Normally, keratinocyte-derived neurotrophic signals guide sensory fiber re-entry into healed epidermis. We hypothesized that impaired neurotrophic support underlies failed reinnervation in RDEB. To investigate the mechanisms behind failed reinnervation, we assessed neurotrophic factor expression in a human skin wound model. We analyzed the secretome of primary keratinocytes from healthy donors and patients with RDEB and tested its effects on neurite outgrowth in sensory neurons derived from embryonic rodents and human induced pluripotent stem cells. We also evaluated the regenerative potential of tropomyosin receptor kinase A (TrkA) and glial cell derived neurotrophic factor (GDNF) receptor agonists (gambogic amide and XIB4035) in vitro and in a mouse model of RDEB. In healthy skin, injury triggered robust neurotrophic factor secretion, whereas RDEB skin did not. Secretomes from healthy keratinocytes promoted neurite outgrowth, whereas those from RDEB keratinocytes failed to do so. Receptor agonist treatment restored neurite growth in vitro, enhanced intraepidermal innervation, and reversed thermal hyposensitivity in RDEB mice.These findings suggest that impaired neurotrophic support from RDEB keratinocytes contributes to defective epidermal reinnervation. Pharmacological activation of TrkA and GDNF receptors may offer a therapeutic strategy to restore sensory function and relieve neuropathic pain in RDEB.

MSI1-FTHL17C-iron circuit couples metabolic and epigenetic control of pluripotency in mouse embryonic stem cells.