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The placenta is crucial for fetal development, yet the impact of environmental stressors such as arsenic exposure remains poorly understood. We apply single-cell RNA sequencing to analyze the response of the mouse placenta to arsenic, revealing cell-type-specific gene expression, function, and pathological changes. Notably, the Prap1 gene, which encodes proline-rich acidic protein 1 (PRAP1), is significantly upregulated in 26 placental cell types including various trophoblast cells. Our study shows a female-biased increase in PRAP1 in response to arsenic and localizes it in the placenta. In vitro and ex vivo experiments confirm PRAP1 upregulation following arsenic treatment and demonstrate that recombinant PRAP1 protein reduces arsenic-induced cytotoxicity and downregulates cell cycle pathways in human trophoblast cells. Moreover, PRAP1 knockdown differentially affects cell cycle processes, proliferation, and cell death depending on the presence of arsenic. Our findings provide insights into the placental response to environmental stress, offering potential preventative and therapeutic approaches for environment-related adverse outcomes in mothers and children. Environmental stressors have been associated with placental dysfunction and pregnancy complications. Here, the authors reveal gene expression changes in the mouse placenta exposed to arsenic at single-cell resolution and identify a potential therapeutic target to mitigate its harmful effects on pregnancy and fetal development. [ABSTRACT FROM AUTHOR]

In contrast to rodents, the mechanisms underlying human trophectoderm and early placenta specification are understudied due to ethical barriers and the scarcity of embryos. Recent reports have shown that human pluripotent stem cells (PSCs) can differentiate into trophectoderm (TE)-like cells (TELCs) and trophoblast stem cells (TSCs), offering a valuable in vitro model to study early placenta specification. Here, we demonstrate that the VGLL1 (vestigial-like family member 1), which is highly expressed during human and non-human primate TE specification in vivo but is negligibly expressed in mouse, is a critical regulator of cell fate determination and self-renewal in human TELCs and TSCs derived from naïve PSCs. Mechanistically, VGLL1 partners with the transcription factor TEAD4 (TEA domain transcription factor 4) to regulate chromatin accessibility at target gene loci through histone acetylation and acts in cooperation with GATA3 and TFAP2C. Our work is relevant to understand primate early embryogenesis and how it differs from other mammalian species. Authors report that VGLL1 regulates cell fate determination and self-renewal of human pluripotent stem cell-derived trophectoderm-like cells and trophoblast stem cells via modulation of chromatin accessibility in cooperation with TEAD4. [ABSTRACT FROM AUTHOR]

Cattle traits like average daily weight gain (ADG) greatly impact profitability. Selecting based on ADG considering genetic variability can lead to economic and genetic advancements in cattle breeding. This study aimed to unravel genetic influences on ADG variation in Hanwoo cattle at the skeletal muscle transcriptomic level. RNA sequencing was conducted on longissimus dorsi (LD), semimembranosus (SB), and psoas major (PM) muscles of 14 steers assigned to same feed, grouped by low (≤ 0.71 kg) and high (≥ 0.77 kg) ADG. At P ≤ 0.05 and log2fold > 1.5, the distinct pattern of gene expression was identified with 184, 172, and 210 differentially expressed genes in LD, SB, and PM muscles, respectively. Tissue-specific responses to ADG variation were evident, with myogenesis and differentiation associated JAK-STAT signaling pathway and prolactin signaling pathways enriched in LD and SB muscles, while adipogenesis-related PPAR signaling pathways were enriched in PM muscle. Key hub genes (AXIN2, CDKN1A, MYC, PTGS2, FZD5, SPP1) were upregulated and functionally significant in muscle growth and differentiation. Notably, DPP6, CDKN1A, and FZD5 emerged as possible candidate genes linked to ADG variation. These findings enhance our understanding of genetic factors behind ADG variation in Hanwoo cattle, illuminating skeletal muscle mechanisms influencing ADG. [ABSTRACT FROM AUTHOR]

Reproductive isolation occurs when the genomes of two populations accumulate genetic incompatibilities that prevent interbreeding1,2. Understanding of hybrid incompatibility at the cell biology level is limited, particularly in the case of hybrid female sterility3. Here we find that species divergence in condensin regulation and centromere organization between two mouse species, Mus musculus domesticus and Mus spretus, drives chromosome decondensation and mis-segregation in their F1 hybrid oocytes, reducing female fertility. The decondensation in hybrid oocytes was especially prominent at pericentromeric major satellites, which are highly abundant at M. m.domesticus centromeres4–6, leading to species-specific chromosome mis-segregation and egg aneuploidy. Consistent with the condensation defects, a chromosome structure protein complex, condensin II7,8, was reduced on hybrid oocyte chromosomes. We find that the condensin II subunit NCAPG2 was specifically reduced in the nucleus in prophase and that overexpressing NCAPG2 rescued both the decondensation and egg aneuploidy phenotypes. In addition to the overall reduction in condensin II on chromosomes, major satellites further reduced condensin II levels locally, explaining why this region is particularly prone to decondensation. Together, this study provides cell biological insights into hybrid incompatibility in female meiosis and demonstrates that condensin misregulation and pericentromeric satellite expansion can establish a reproductive isolating barrier in mammals.Species divergence in condensin regulation and centromere organization between the mice Mus musculus domesticus and Mus spretus drives chromosome decondensation and mis-segregation in their F1 hybrid oocytes, reducing female fertility. [ABSTRACT FROM AUTHOR]

The SWI/SNF ATP-dependent chromatin remodeler is a master regulator of the epigenome, controlling pluripotency and differentiation. Towards the C-terminus of the catalytic subunit of SWI/SNF is a motif called the AT-hook that is evolutionary conserved. The AT-hook is present in many chromatin modifiers and generally thought to help anchor them to DNA. We observe however that the AT-hook regulates the intrinsic DNA-stimulated ATPase activity aside from promoting SWI/SNF recruitment to DNA or nucleosomes by increasing the reaction velocity a factor of 13 with no accompanying change in substrate affinity (KM). The changes in ATP hydrolysis causes an equivalent change in nucleosome movement, confirming they are tightly coupled. The catalytic subunit's AT-hook is required in vivo for SWI/SNF remodeling activity in yeast and mouse embryonic stem cells. The AT-hook in SWI/SNF is required for transcription regulation and activation of stage-specific enhancers critical in cell lineage priming. Similarly, growth assays suggest the AT-hook is required in yeast SWI/SNF for activation of genes involved in amino acid biosynthesis and metabolizing ethanol. Our findings highlight the importance of studying SWI/SNF attenuation versus eliminating the catalytic subunit or completely shutting down its enzymatic activity. This study demonstrates that an evolutionary conserved, autoregulatory 'AT-hook' domain of SWI/SNF regulates gene transcription and enhancer activation by modulating SWI/SNF intrinsic catalytic activity and is critical for cell lineage priming. [ABSTRACT FROM AUTHOR]

Purpose: Despite the success of ICSI in treating severe male factor infertile patients, total fertilization failure (FF) still occurs in around 1–3% of ICSI cycles. To overcome FF, the use of calcium ionophores has been proposed to induce oocyte activation and restore fertilization rates. However, assisted oocyte activation (AOA) protocols and ionophores vary between laboratories, and the morphokinetic development underlying AOA remains understudied. Methods: A prospective single-center cohort study involving 81 in vitro matured metaphase-II oocytes from 66 oocyte donation cycles artificially activated by A23187 (GM508 CultActive, Gynemed) (n=42) or ionomycin (n=39). Parthenogenesis was induced, and morphokinetic parameters (tPNa, tPNf, t2-t8, tSB, and tB) were compared between the 2 study groups and a control group comprising 39 2PN-zygotes from standard ICSI cycles. Results: Ionomycin treatment resulted in higher activation rates compared to A23187 (38.5% vs 23.8%, p=0.15). Importantly, none of the A23187-activated parthenotes formed blastocysts. When evaluating the morphokinetic dynamics between the two ionophores, we found that tPNa and tPNf were significantly delayed in the group treated by A23187 (11.84 vs 5.31, p=0.002 and 50.15 vs 29.69, p=0.005, respectively). t2 was significantly delayed in A23187-activated parthenotes when compared to the double heterologous control embryo group. In contrast, the morphokinetic development of ionomycin-activated parthenotes was comparable to control embryos (p>0.05). Conclusion: Our results suggest that A23187 leads to lower oocyte activation rates and profoundly affects morphokinetic timings and preimplantation development in parthenotes. Despite our limited sample size and low parthenote competence, standardization and further optimization of AOA protocols may allow wider use and improved outcomes for FF cycles. [ABSTRACT FROM AUTHOR]

Purpose: Events in the uterus during the peri-implantation period include embryo development, acquisition of uterine receptivity, implantation and decidualization. Hippo signaling pathway regulates cell proliferation, apoptosis and differentiation. We aimed to determine localization and expressions of pYAP (Phospho Yes-associated protein), YAP (Yes-associated protein), TEAD1 (TEA domain family member 1) and CTGF (Connective tissue growth factor), members of the Hippo signaling pathway, in the mouse uterus during the peri-implantation period. Methods: Pregnant mice were randomly separated into 5 groups: 1st, 4th, 5th, 6th, and 8th days of pregnancy groups. Non-pregnant female mice in estrous phase were included in the estrous group. Uteri and implantation sites were collected. Also, inter-implantation sites were collected from the 5th day of pregnancy group. pYAP, YAP, TEAD-1 and CTGF were detected by immunohistochemistry and Western blotting. Results: We observed that the expressions of YAP, TEAD-1 and CTGF were increased in the luminal and glandular epithelium on the 1st and 4th days of pregnancy when epithelial proliferation occurred. pYAP expression was high, and YAP and CTGF expressions were low in the luminal epithelium of the implantation sites on the 5th day of pregnancy, when epithelial differentiation occurred. pYAP expression was low, YAP and CTGF expressions were high at implantation sites on the 6th and 8th days of pregnancy, where decidua was formed. Conclusion: Our findings suggest that the Hippo signaling pathway might be involved in implantation and decidualization. Our findings will guide further studies and may help to elucidate underlying causes of implantation failure and pregnancy loss. [ABSTRACT FROM AUTHOR]

Purpose: This study is to develop a new protocol that combines the use of epigenetic cues and mechanical stimuli to assemble 3D spherical structures, arbitrarily defined "epiBlastoids," whose phenotype is remarkably similar to natural embryos. Methods: A 3-step approach is used to generate epiBlastoids. In the first step, adult dermal fibroblasts are converted into trophoblast (TR)-like cells, combining the use of 5-azacytidine, to erase the original phenotype, with an ad hoc induction protocol, to drive cells towards TR lineage. In the second step, epigenetic erasing is applied once again, in combination with mechanosensing-related cues, to generate inner cell mass (ICM)-like organoids. Specifically, erased cells are encapsulated into micro-bioreactors to promote 3D cell rearrangement and boost pluripotency. In the third step, TR-like cells are co-cultured with ICM-like spheroids in the same micro-bioreactors. Subsequently, the newly generated embryoids are transferred to microwells to favor epiBlastoid formation. Results: Adult dermal fibroblasts are successfully readdressed towards TR lineage. Cells subjected to epigenetic erasing and encapsulated into micro-bioreactors rearrange in 3D ICM-like structures. Co-culture of TR-like cells and ICM-like spheroids into micro-bioreactors and microwells induces the formation of single structures with uniform shape reminiscent in vivo embryos. CDX2+ cells localized in the out layer of the spheroids, while OCT4+ cells in the inner of the structures. TROP2+ cells display YAP nuclear accumulation and actively transcribed for mature TR markers, while TROP2− cells showed YAP cytoplasmic compartmentalization and expressed pluripotency-related genes. Conclusion: We describe the generation of epiBlastoids that may find useful application in the assisted reproduction field. [ABSTRACT FROM AUTHOR]

Protein synthesis is an essential step in gene expression during the development of mammalian preimplantation embryos. This is a complex and highly regulated process. The accuracy of the translation initiation codon is important in various gene expression programs. However, the mechanisms that regulate AUG and non-AUG codon initiation in early embryos remain poorly understood. BZW1 is a key factor in determining the mRNA translation start codon. Here, we show that BZW1 is essential for early embryonic development in mice. Bzw1-knockdown embryos fail to undergo compaction, and show decreased blastocyst formation rates. We also observe defects in the differentiation capacity and implantation potential after Bzw1 interference. Further investigation revealed that Bzw1 knockdown causes the levels of translation initiation with CUG as the start codon to increase. The decline in BZW1 levels result in a decrease in protein synthesis in preimplantation embryos, whereas the total mRNA levels are not altered. Therefore, we concluded that BZW1 contributes to protein synthesis during early embryonic development by restricting non-AUG translational initiation. It is unclear how translational efficiency and codon stringency affect zygotic genome activation and early embryonic development. Here they show that the zygotic gene BZW1 increases start codon stringency, which is important for preimplantation development. [ABSTRACT FROM AUTHOR]

Pluripotent character is described as the potency of cells to differentiate into all three germ layers. The best example to reinstate the term lies in the context of embryonic stem cells (ESCs). Pluripotent ESC describes the in vitro status of those cells that originate during the complex process of embryogenesis. Pre-implantation to post-implantation development of embryo embrace cells with different levels of stemness. Currently, four states of pluripotency have been recognized, in the progressing order of "naïve," "poised," "formative," and "primed." Epigenetics act as the "conductor" in this "orchestra" of transition in pluripotent states. With a distinguishable gene expression profile, these four states associate with different epigenetic signatures, sometimes distinct while otherwise overlapping. The present review focuses on how epigenetic factors, including DNA methylation, bivalent chromatin, chromatin remodelers, chromatin/nuclear architecture, and microRNA, could dictate pluripotent states and their transition among themselves. [ABSTRACT FROM AUTHOR]

Physiologic laminar shear stress (LSS) induces an endothelial gene expression profile that is vasculo-protective. In this report, we delineate how LSS mediates changes in the epigenetic landscape to promote this beneficial response. We show that under LSS, KLF4 interacts with the SWI/SNF nucleosome remodeling complex to increase accessibility at enhancer sites that promote the expression of homeostatic endothelial genes. By combining molecular and computational approaches we discover enhancers that loop to promoters of KLF4- and LSS-responsive genes that stabilize endothelial cells and suppress inflammation, such as BMPR2, SMAD5, and DUSP5. By linking enhancers to genes that they regulate under physiologic LSS, our work establishes a foundation for interpreting how non-coding DNA variants in these regions might disrupt protective gene expression to influence vascular disease. Here the authors studied pulmonary arterial endothelial cells (PAEC) under laminar shear stress and show that this physiologic condition markedly changes chromatin accessibility at regulatory regions, when compared to cells grown in a static state. They find that KLF4 organizes chromatin by interacting with the SWI/SNF nucleosome remodeling complex to regulate vasculo-protective gene expression. [ABSTRACT FROM AUTHOR]

The placenta sustains embryonic development and is critical for a successful pregnancy outcome. It provides the site of exchange between the mother and the embryo, has immunological functions and is a vital endocrine organ. To perform these diverse roles, the placenta comprises highly specialized trophoblast cell types, including syncytiotrophoblast and extravillous trophoblast. The coordinated actions of transcription factors (TFs) regulate their emergence during development, subsequent specialization, and identity. These TFs integrate diverse signaling cues, form TF networks, associate with chromatin remodeling and modifying factors, and collectively determine the cell type-specific characteristics. Here, we summarize the general properties of TFs, provide an overview of TFs involved in the development and function of the human trophoblast, and address similarities and differences to their murine orthologs. In addition, we discuss how the recent establishment of human in vitro models combined with -omics approaches propel our knowledge and transform the human trophoblast field. [ABSTRACT FROM AUTHOR]

Canine mesenchymal stem cells (cMSCs) have potential applications for regenerative therapy, including the generation of insulin-producing cells (IPCs) for studying and treating diabetes. In this study, we established a useful protocol for generating IPCs from canine adipose mesenchymal stem cells (cAD-MSCs). Subsequently, in vitro preservation of pluronic F127-coated alginate (ALGPA)-encapsulated cAD-MSC-derived IPCs was performed to verify ready-to-use IPCs. IPCs were induced from cAD-MSCs with the modulated three-stepwise protocol. The first step of definitive endoderm (DE) induction showed that the cooperation of Chir99021 and Activin A created the effective production of Sox17-expressed DE cells. The second step for pancreatic endocrine (PE) progenitor induction from DE indicated that the treatment with taurine, retinoic acid, FGF2, EGF, TGFβ inhibitor, dorsomorphin, nicotinamide, and DAPT showed the significant upregulation of the pancreatic endocrine precursor markers Pdx1 and Ngn3. The last step of IPC production, the combination of taurine, nicotinamide, Glp-1, forskolin, PI3K inhibitor, and TGFβ inhibitor, yielded efficiently functional IPCs from PE precursors. Afterward, the maintenance of ALGPA-encapsulated cAD-MSC-derived IPCs with VSCBIC-1, a specialized medium, enhanced IPC properties. Conclusion, the modulated three-stepwise protocol generates the functional IPCs. Together, the encapsulation of cAD-MSC-derived IPCs and the cultivation with VSCBIC-1 enrich the maturation of generated IPCs. [ABSTRACT FROM AUTHOR]

Lipids are high energy, complex biomolecular compounds essential for cellular and organellar membrane formation. Accumulation of circulatory lipids is however associated with pathophysiological conditions including cardiometabolic disorders, diabetic dyslipidemia and obesity. Epidemiological studies have correlated heavy-metal exposure with dyslipidemias and metabolic syndrome. Here, we review the role of cadmium, lead, mercury and arsenic on inducing dyslipidemias through lipid metabolism dysregulation, with focus on metal effects on lipogenic genes, gut microbiome and endocrine secretion. The main gene transcription factors impaired by heavy metals are CCAAT-enhancer binding protein, peroxisome proliferative-activated receptor, sterol regulatory element binding protein, carbohydrate responsive element binding protein and liver × receptor. These factors regulate genes responsible for β-oxidation, de novo lipogenesis, and the synthesis and transport of fatty acids, cholesterol, phospholipids and triglycerides. Dysregulated lipid profiles in organisms exposed to metals show higher cholesterol and triglycerides, very low-density lipoprotein and non-high density lipoprotein cholesterol levels, with a corresponding low high-density lipoprotein cholesterol. Hormones and gut microbiome are also impaired by heavy-metal exposure. [ABSTRACT FROM AUTHOR]

The process of epigenetic silencing, while fundamentally important, is not yet completely understood. Here we report a replenishable female mouse embryonic stem cell (mESC) system, Xmas, that allows rapid assessment of X chromosome inactivation (XCI), the epigenetic silencing mechanism of one of the two X chromosomes that enables dosage compensation in female mammals. Through a targeted genetic screen in differentiating Xmas mESCs, we reveal that the BAF complex is required to create nucleosome-depleted regions at promoters on the inactive X chromosome during the earliest stages of establishment of XCI. Without this action gene silencing fails. Xmas mESCs provide a tractable model for screen-based approaches that enable the discovery of unknown facets of the female-specific process of XCI and epigenetic silencing more broadly. Female embryonic stem cells (ESCs) are the ideal model to study X chromosome inactivation (XCI) establishment; however, these cells are challenging to keep in culture. Here the authors create fluorescent 'Xmas' reporter mice as a renewable source of ESCs and show nucleosome remodelers Smarcc1 and Smarca4 create a nucleosome-free promoter region prior to the establishment of silencing. [ABSTRACT FROM AUTHOR]

First cell fate determination plays crucial roles in cell specification during early phases of embryonic development. Three classical concepts have been proposed to explain the lineage specification mechanism of the preimplantation embryo: inside-outside, pre-patterning, and polarity models. Transcriptional effectors of the Hippo signal pathway are YAP and TAZ activators that can create a shuttle between the cytoplasm and the nucleus. Despite different localizations of YAP in the cell, it determines the fate of ICM and TE. How the decisive cue driving factors that determine YAP localization are coordinated remains a central unanswered question. How can an embryonic cell find its position? The objective of this review is to summarize the molecular and mechanical aspects in cell fate decision during mouse preimplantation embryonic development. The findings will reveal the relationship between cell–cell adhesion, cell polarity, and determination of cell fate during early embryonic development in mice and elucidate the inducing/inhibiting mechanisms that are involved in cell specification following zygotic genome activation and compaction processes. With future studies, new biophysical and chemical cues in the cell fate determination will impart significant spatiotemporal effects on early embryonic development. The achieved knowledge will provide important information to the development of new approaches to be used in infertility treatment and increase the success of pregnancy. [ABSTRACT FROM AUTHOR]