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BioRxiv : the Preprint Server For... Apr 2024As cross-disciplinary approaches drawing from physics and mechanics have increasingly influenced our understanding of morphogenesis, the tools available to measure and...
As cross-disciplinary approaches drawing from physics and mechanics have increasingly influenced our understanding of morphogenesis, the tools available to measure and perturb physical aspects of embryonic development have expanded as well. However, it remains a challenge to measure mechanical properties and apply exogenous tissue-scale forces , particularly for epithelia. Exploiting the size and accessibility of the developing chick embryo, here we describe a simple technique to quantitatively apply exogenous forces on the order of 1-100 N to the endodermal epithelium. To demonstrate the utility of this approach, we performed a series of proof-of-concept experiments that reveal fundamental and unexpected mechanical behaviors in the early chick embryo, including mechanotype heterogeneity among cells of the midgut endoderm, complex non-cell autonomous effects of actin disruption, and a high degree of mechanical coupling between the endoderm and adjacent paraxial mesoderm. To illustrate the broader utility of this method, we determined that forces on the order of 10 N are sufficient to unzip the neural tube during primary neurulation. Together, these findings provide basic insights into the mechanics of embryonic epithelia in the early avian embryo, and provide a useful tool for future investigations of how morphogenesis is influenced by mechanical factors.
PubMed: 38617324
DOI: 10.1101/2024.04.04.588089 -
Toxicology in Vitro : An International... Apr 2024Human induced pluripotent stem cells (iPSC) have the potential to produce desired target cell types in vitro and allow for the high-throughput screening of...
Human induced pluripotent stem cells (iPSC) have the potential to produce desired target cell types in vitro and allow for the high-throughput screening of drugs/chemicals at population level thereby minimising the cost of drug discovery and drug withdrawals after clinical trials. There is a substantial need for the characterisation of the iPSC derived models to better understand and utilise them for toxicological relevant applications. In our study, iPSC (SBAD2 or SBAD3 lines obtained from StemBANCC project) were differentiated towards toxicologically relevant cell types: alveolar macrophages, brain capillary endothelial cells, brain cells, endothelial cells, hepatocytes, lung airway epithelium, monocytes, podocytes and renal proximal tubular cells. A targeted transcriptomic approach was employed to understand the effects of differentiation protocols on these cell types. Pearson correlation and principal component analysis (PCA) separated most of the intended target cell types and undifferentiated iPSC models as distinct groups with a high correlation among replicates from the same model. Based on PCA, the intended target cell types could also be separated into the three germ layer groups (ectoderm, endoderm and mesoderm). Differential expression analysis (DESeq2) presented the upregulated genes in each intended target cell types that allowed the evaluation of the differentiation to certain degree and the selection of key differentiation markers. In conclusion, these data confirm the versatile use of iPSC differentiated cell types as standardizable and relevant model systems for in vitro toxicology.
PubMed: 38615723
DOI: 10.1016/j.tiv.2024.105826 -
Ear, Nose, & Throat Journal Apr 2024Hairy polyps, considered a highly unusual congenital anomaly of the pharynx, are believed by many scholars to arise from the ectoderm and mesoderm during the embryonic...
Hairy polyps, considered a highly unusual congenital anomaly of the pharynx, are believed by many scholars to arise from the ectoderm and mesoderm during the embryonic stage. These growths often have a pear or sausage shape, are pedunculated, and their size ranges between 0.5 and 6 cm. They are typically grayish white or pink in color. This article discusses a 12-year-old female who had a growth at the Eustachian tube's entrance on the left side of the nasopharyngeal wall, as identified by a computed tomography scan of the neck soft tissue; it was suspected to be a hairy polyp originating from the left Eustachian tube. The diagnosis of a hairy polyp was confirmed through pathology. The hairy polyp at the Eustachian tube, in this case, showed an irregular form with a wide base, making it look similar to an adenoid; thus, increasing the risk of it being misdiagnosed as residual adenoid tissue.
PubMed: 38605528
DOI: 10.1177/01455613241244667 -
BioRxiv : the Preprint Server For... Mar 2024Micropatterned human pluripotent stem cells (hPSCs) treated with BMP4 (2D gastruloids) are among the most widely used stem cell models for human gastrulation. Due to its...
Micropatterned human pluripotent stem cells (hPSCs) treated with BMP4 (2D gastruloids) are among the most widely used stem cell models for human gastrulation. Due to its simplicity and reproducibility, this system is ideal for high throughput quantitative studies of tissue patterning and has led to many insights into the mechanisms of mammalian gastrulation. However, 2D gastruloids have only been studied up to 48h. Here we extended this system to 96h. We discovered a phase of highly reproducible morphogenesis during which directed migration from the primitive streak-like region gives rise to a mesodermal layer beneath an epiblast-like layer. Multiple types of mesoderm arise with striking spatial organization including lateral mesoderm-like cells on the colony border and paraxial mesoderm-like further inside the colony. Single cell transcriptomics showed strong similarity of these cells to mesoderm in human and non-human primate embryos. However, our data suggest that the annotation of the reference human embryo may need to be revised. This illustrates that extended culture of 2D gastruloids provides a powerful model for human mesoderm differentiation and morphogenesis.
PubMed: 38585971
DOI: 10.1101/2024.03.21.585753 -
BioRxiv : the Preprint Server For... Mar 2024Transposable elements (TEs) make up the bulk of eukaryotic genomes and examples abound of TE-derived sequences repurposed for organismal function. The process by which...
Transposable elements (TEs) make up the bulk of eukaryotic genomes and examples abound of TE-derived sequences repurposed for organismal function. The process by which TEs become coopted remains obscure because most cases involve ancient, transpositionally inactive elements. Reports of active TEs serving beneficial functions are scarce and often contentious due to difficulties in manipulating repetitive sequences. Here we show that recently active TEs in zebrafish encode products critical for embryonic development. Knockdown and rescue experiments demonstrate that the endogenous retrovirus family BHIKHARI-1 (Bik-1) encodes a Gag protein essential for mesoderm development. Mechanistically, Bik-1 Gag associates with the cell membrane and its ectopic expression in chicken embryos alters cell migration. Similarly, depletion of BHIKHARI-2 Gag, a relative of Bik-1, causes defects in neural crest development in zebrafish. We propose an "addiction" model to explain how active TEs can be integrated into conserved developmental processes.
PubMed: 38585793
DOI: 10.1101/2024.03.25.586437 -
BioRxiv : the Preprint Server For... Mar 2024Cultured beef holds promising potential as an alternative to traditional meat options. While adult stem cells are commonly used as the cell source for cultured beef,...
Cultured beef holds promising potential as an alternative to traditional meat options. While adult stem cells are commonly used as the cell source for cultured beef, their proliferation and differentiation capacities are limited. To produce cultured beef steaks, current manufacturing plans often require the separate preparation of multiple cell types and intricate engineering for assembling them into structured tissues. In this study, we propose and report the co-induction of skeletal muscle, neuronal, and endothelial cells from bovine embryonic stem cells (ESCs) and the self-organization of tissue structures in 2- and 3-dimensional cultures. Bovine myocytes were induced in a stepwise manner through the induction of presomitic mesoderm (PSM) from bovine ESCs. Muscle fibers with sarcomeres appeared within 15 days, displaying calcium oscillations responsive to inputs from co-induced bovine spinal neurons. Bovine endothelial cells were also co-induced via PSM, forming uniform vessel networks inside tissues. Our serum-free, rapid co-induction protocols represent a milestone toward self-organizing beef steaks with integrated vasculature and innervation.
PubMed: 38585777
DOI: 10.1101/2024.03.22.586252 -
Cytotherapy Mar 2024Substrate elasticity may direct cell-fate decisions of stem cells. However, it is largely unclear how matrix stiffness affects the differentiation of induced pluripotent...
BACKGROUND AIMS
Substrate elasticity may direct cell-fate decisions of stem cells. However, it is largely unclear how matrix stiffness affects the differentiation of induced pluripotent stem cells (iPSCs) and whether this is also reflected by epigenetic modifications.
METHODS
We cultured iPSCs on tissue culture plastic (TCP) and polydimethylsiloxane (PDMS) with different Young's modulus (0.2 kPa, 16 kPa or 64 kPa) to investigate the sequel on growth and differentiation toward endoderm, mesoderm and ectoderm.
RESULTS
Immunofluorescence and gene expression of canonical differentiation markers were hardly affected by the substrates. Notably, when we analyzed DNA methylation profiles of undifferentiated iPSCs or after three-lineage differentiation, we did not see any significant differences on the three different PDMS elasticities. Only when we compared DNA methylation profiles on PDMS-substrates versus TCP we did observe epigenetic differences, particularly on mesodermal differentiation.
CONCLUSIONS
Stiffness of PDMS substrates did not affect directed differentiation of iPSCs, whereas the moderate epigenetic differences on TCP might also be attributed to other chemical parameters.
PubMed: 38583169
DOI: 10.1016/j.jcyt.2024.03.485 -
Communications Biology Apr 2024The ten-eleven-translocation family of proteins (TET1/2/3) are epigenetic regulators of gene expression. They regulate genes by promoting DNA demethylation (i.e.,...
The ten-eleven-translocation family of proteins (TET1/2/3) are epigenetic regulators of gene expression. They regulate genes by promoting DNA demethylation (i.e., catalytic activity) and by partnering with regulatory proteins (i.e., non-catalytic functions). Unlike Tet1 and Tet2, Tet3 is not expressed in mouse embryonic stem cells (ESCs) but is induced upon ESC differentiation. However, the significance of its dual roles in lineage specification is less defined. By generating TET3 catalytic-mutant (Tet3) and knockout (Tet3) mouse ESCs and differentiating them to neuroectoderm (NE), we identify distinct catalytic-dependent and independent roles of TET3 in NE specification. We find that the catalytic activity of TET3 is important for activation of neural genes while its non-catalytic functions are involved in suppressing mesodermal programs. Interestingly, the vast majority of differentially methylated regions (DMRs) in Tet3 and Tet3 NE cells are hypomethylated. The hypo-DMRs are associated to aberrantly upregulated genes while the hyper-DMRs are linked to downregulated neural genes. We find the maintenance methyltransferase Dnmt1 as a direct target of TET3, which is downregulated in TET3-deficient NE cells and may contribute to the increased DNA hypomethylation. Our findings establish that the catalytic-dependent and -independent roles of TET3 have distinct contributions to NE specification with potential implications in development.
Topics: Animals; Mice; Cell Differentiation; Dioxygenases; DNA Methylation; DNA-Binding Proteins; Neural Plate
PubMed: 38580843
DOI: 10.1038/s42003-024-06120-w -
Developmental Cell Apr 2024Embryogenesis requires substantial coordination to translate genetic programs to the collective behavior of differentiating cells, but understanding how cellular...
Embryogenesis requires substantial coordination to translate genetic programs to the collective behavior of differentiating cells, but understanding how cellular decisions control tissue morphology remains conceptually and technically challenging. Here, we combine continuous Cas9-based molecular recording with a mouse embryonic stem cell-based model of the embryonic trunk to build single-cell phylogenies that describe the behavior of transient, multipotent neuro-mesodermal progenitors (NMPs) as they commit into neural and somitic cell types. We find that NMPs show subtle transcriptional signatures related to their recent differentiation and contribute to downstream lineages through a surprisingly broad distribution of individual fate outcomes. Although decision-making can be heavily influenced by environmental cues to induce morphological phenotypes, axial progenitors intrinsically mature over developmental time to favor the neural lineage. Using these data, we present an experimental and analytical framework for exploring the non-homeostatic dynamics of transient progenitor populations as they shape complex tissues during critical developmental windows.
PubMed: 38579718
DOI: 10.1016/j.devcel.2024.03.024 -
Stem Cell Reports May 2024SOX2 is a transcription factor involved in the regulatory network maintaining the pluripotency of embryonic stem cells in culture as well as in early embryos. In...
SOX2 is a transcription factor involved in the regulatory network maintaining the pluripotency of embryonic stem cells in culture as well as in early embryos. In addition, SOX2 plays a pivotal role in neural stem cell formation and neurogenesis. How SOX2 can serve both processes has remained elusive. Here, we identified a set of SOX2-dependent neural-associated enhancers required for neural lineage priming. They form a distinct subgroup (1,898) among 8,531 OCT4/SOX2/NANOG-bound enhancers characterized by enhanced SOX2 binding and chromatin accessibility. Activation of these enhancers is triggered by neural induction of wild-type cells or by default in Smad4-ablated cells resistant to mesoderm induction and is antagonized by mesodermal transcription factors via Sox2 repression. Our data provide mechanistic insight into the transition from the pluripotency state to the early neural fate and into the regulation of early neural versus mesodermal specification in embryonic stem cells and embryos.
Topics: SOXB1 Transcription Factors; Animals; Enhancer Elements, Genetic; Mice; Neural Stem Cells; Mesoderm; Neurogenesis; Gene Expression Regulation, Developmental; Octamer Transcription Factor-3; Cell Differentiation; Nanog Homeobox Protein; Cell Lineage; Smad4 Protein; Embryonic Stem Cells; Mouse Embryonic Stem Cells; Chromatin; Protein Binding
PubMed: 38579708
DOI: 10.1016/j.stemcr.2024.03.003