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1.
Nat Commun ; 14(1): 349, 2023 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-36681697

RESUMO

The processes of primitive streak formation and fate specification in the mammalian epiblast rely on complex interactions between morphogens and tissue organization. Little is known about how these instructive cues functionally interact to regulate gastrulation. We interrogated the interplay between tissue organization and morphogens by using human induced pluripotent stem cells (hiPSCs) downregulated for the morphogen regulator GLYPICAN-4, in which defects in tight junctions result in areas of disrupted epithelial integrity. Remarkably, this phenotype does not affect hiPSC stemness, but impacts on cell fate acquisition. Strikingly, cells within disrupted areas become competent to perceive the gastrulation signals BMP4 and ACTIVIN A, an in vitro surrogate for NODAL, and thus differentiate into mesendoderm. Yet, disruption of epithelial integrity sustains activation of BMP4 and ACTIVIN A downstream effectors and correlates with enhanced hiPSC endoderm/mesoderm differentiation. Altogether, our results disclose epithelial integrity as a key determinant of TGF-ß activity and highlight an additional mechanism guiding morphogen sensing and spatial cell fate change within an epithelium.


Assuntos
Células-Tronco Pluripotentes Induzidas , Células-Tronco Pluripotentes , Animais , Humanos , Fator de Crescimento Transformador beta/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Diferenciação Celular/fisiologia , Camadas Germinativas/metabolismo , Mesoderma/metabolismo , Endoderma/metabolismo , Mamíferos/metabolismo
2.
Nat Commun ; 14(1): 1, 2023 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-36596776

RESUMO

Pancreatic cancer is characterized by abundant desmoplasia, a dense stroma composed of extra-cellular and cellular components, with cancer associated fibroblasts (CAFs) being the major cellular component. However, the tissue(s) of origin for CAFs remains controversial. Here we determine the tissue origin of pancreatic CAFs through comprehensive lineage tracing studies in mice. We find that the splanchnic mesenchyme, the fetal cell layer surrounding the endoderm from which the pancreatic epithelium originates, gives rise to the majority of resident fibroblasts in the normal pancreas. In a genetic mouse model of pancreatic cancer, resident fibroblasts expand and constitute the bulk of CAFs. Single cell RNA profiling identifies gene expression signatures that are shared among the fetal splanchnic mesenchyme, adult fibroblasts and CAFs, suggesting a persistent transcriptional program underlies splanchnic lineage differentiation. Together, this study defines the phylogeny of the mesenchymal component of the pancreas and provides insights into pancreatic morphogenesis and tumorigenesis.


Assuntos
Pâncreas , Neoplasias Pancreáticas , Camundongos , Animais , Pâncreas/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Fibroblastos/metabolismo , Carcinogênese/genética , Carcinogênese/metabolismo , Transformação Celular Neoplásica/metabolismo , Mesoderma/metabolismo , Homeostase
3.
Nat Mater ; 22(1): 135-143, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36577855

RESUMO

Tissue morphogenesis, homoeostasis and repair require cells to constantly monitor their three-dimensional microenvironment and adapt their behaviours in response to local biochemical and mechanical cues. Yet the mechanical parameters of the cellular microenvironment probed by cells in vivo remain unclear. Here, we report the mechanics of the cellular microenvironment that cells probe in vivo and in situ during zebrafish presomitic mesoderm differentiation. By quantifying both endogenous cell-generated strains and tissue mechanics, we show that individual cells probe the stiffness associated with deformations of the supracellular, foam-like tissue architecture. Stress relaxation leads to a perceived microenvironment stiffness that decreases over time, with cells probing the softest regime. We find that most mechanical parameters, including those probed by cells, vary along the anteroposterior axis as mesodermal progenitors differentiate. These findings expand our understanding of in vivo mechanosensation and might aid the design of advanced scaffolds for tissue engineering applications.


Assuntos
Mesoderma , Peixe-Zebra , Animais , Mesoderma/fisiologia , Diferenciação Celular/fisiologia , Morfogênese , Microambiente Celular
4.
Curr Biol ; 33(1): 197-205.e2, 2023 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-36563692

RESUMO

Evolutionary transitions are frequently associated with novel anatomical structures,1 but the origins of the structures themselves are often poorly known. We use developmental, genetic, and paleontological data to demonstrate that the therian sternum was assembled from pre-existing elements. Imaging of the perinatal mouse reveals two paired sternal elements, both composed primarily of cells with lateral plate mesoderm origin. Location, articulations, and development identify them as homologs of the interclavicle and the sternal bands of synapsid outgroups. The interclavicle, not previously recognized in therians,2 articulates with the clavicle and differs from the sternal bands in both embryonic HOX expression and pattern of skeletal maturation. The sternal bands articulate with the ribs in two styles, most clearly differentiated by their association with sternebrae. Evolutionary trait mapping indicates that the interclavicle and sternal bands were independent elements throughout most of synapsid history. The differentiation of rib articulation styles and the subdivision of the sternal bands into sternebrae were key innovations likely associated with transitions in locomotor and respiratory mechanics.3,4 Fusion of the interclavicle and the anterior sternal bands to form a presternum anterior to the first sternebra was a historically recent innovation unique to therians. Subsequent disassembly of the radically reduced sternum of mysticete cetaceans was element specific, reflecting the constraints that conserved developmental programs exert on composite structures.


Assuntos
Evolução Biológica , Esterno , Animais , Camundongos , Mamíferos , Mesoderma , Costelas , Cetáceos
5.
Dev Dyn ; 251(3): 536-550, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-34494340

RESUMO

BACKGROUND: Tissue organoids derived from primary cells have high potential for studying organ development and diseases in numerous organs. They recreate the morphological structure and mimic the functions of given organ while being compact in size, easy to produce, and suitable for use in various experimental setups. RESULTS: In this study we established the number of cells that form mouse kidney rudiments at E11.5, and generated renal organoids of various sizes from the mouse primary cells of the metanephric mesenchyme (MM). We investigated the ability of renal organoids to undergo nephrogenesis upon Wnt/ ß-catenin pathway-mediated tubule induction with a GSK-3 inhibitor (BIO) or by initiation through the ureteric bud (UB). We found that 5000 cells of MM cells are necessary to successfully form renal organoids with well-structured nephrons as judged by fluorescent microscopy, transmission electron microscopy (TEM), and quantitative Polymerase Chain Reaction (qPCR). These mouse organoids also recapitulated renal secretion function in the proximal tubules. CONCLUSIONS: We show that a significant decrease of cells used to generate renal mouse organoids in a dissociation/re-aggregation assay, does not interfere with development, and goes toward 3Rs. This enables generation of more experimental samples with one mouse litter, limiting the number of animals used for studies.


Assuntos
Quinase 3 da Glicogênio Sintase , Organogênese , Animais , Rim , Mesoderma , Camundongos , Néfrons
6.
Drug Des Devel Ther ; 16: 4079-4089, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36465266

RESUMO

Background: Colorectal cancer (CRC) is a multifactorial disease and one of the most common malignancies worldwide. Salidroside (Sal) is a plant with a wide range of pharmacological effects and plays an important role in the treatment of many diseases, and is considered a new hope for the treatment of tumors. The purpose of this study was to investigate the effect of the combination of Sal and paclitaxel (Pac) on colorectal cancer cells and its mechanism of action. Methods: The effects of different mass concentrations of Sal, Pac, and the combination intervened in the cells for 48 h were examined using the CCK8 method. The inhibition rate was obtained, and the optimal concentration of the respective drug group was screened. The proliferative capacity of the respective group was obtained. Subsequently, the results of apoptosis, cloning, migration, invasion, and angiogenesis were observed through cell morphological analysis (shape observation and Hoechst staining), colony formation assay, cell scratching assay, Transwell, angiogenesis assay, and protein immunoblotting (Western blotting) to detect the expression of epithelial-mesenchymal transition (EMT)-associated proteins and PI3K pathway-associated proteins. Results: Different concentrations of Sal, Pac, and the combined application had significant effects in inhibiting cells in a concentration-dependent manner. Compared with the control group, the Sal group, the Pac group, and the combination group significantly inhibited the clonal number, migration, invasion, and tube-forming ability of colorectal cancer cells. Besides, the combined application had a better effect than the Sal and Pac groups. The apoptosis level was up-regulated in all drug groups, and the up-regulation was more significant in the combination group. The expression of E-cad protein was up-regulated, the expression of N-cad and Vim protein was down-regulated, and the expression of PI3K and AKT phosphorylation was down-regulated in the respective group, and the difference was more significant in the combination group compared with the group of individual drugs. Conclusion: The combined application of Sal and Pac significantly can decrease the survival rate of colorectal cancer cells, and the mechanism may be correlated with the blocking of the PI3K/AKT pathway, thus inhibiting EMT.


Assuntos
Neoplasias Colorretais , Paclitaxel , Humanos , Paclitaxel/farmacologia , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Mesoderma , Neoplasias Colorretais/tratamento farmacológico , Proliferação de Células
7.
Nat Commun ; 13(1): 7766, 2022 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-36522318

RESUMO

The vertebrate main-body axis is laid down during embryonic stages in an anterior-to-posterior (head-to-tail) direction, driven and supplied by posteriorly located progenitors. Whilst posterior expansion and segmentation appears broadly uniform along the axis, there is developmental and evolutionary support for at least two discrete modules controlling processes within different axial regions: a trunk and a tail module. Here, we identify Nuclear receptor subfamily 6 group A member 1 (Nr6a1) as a master regulator of trunk development in the mouse. Specifically, Nr6a1 was found to control vertebral number and segmentation of the trunk region, autonomously from other axial regions. Moreover, Nr6a1 was essential for the timely progression of Hox signatures, and neural versus mesodermal cell fate choice, within axial progenitors. Collectively, Nr6a1 has an axially-restricted role in all major cellular and tissue-level events required for vertebral column formation, supporting the view that changes in Nr6a1 levels may underlie evolutionary changes in axial formulae.


Assuntos
Mesoderma , Vertebrados , Animais , Camundongos , Vertebrados/genética , Coluna Vertebral , Regulação da Expressão Gênica no Desenvolvimento , Padronização Corporal/genética
8.
Nat Commun ; 13(1): 7934, 2022 12 24.
Artigo em Inglês | MEDLINE | ID: mdl-36566327

RESUMO

Classic microsurgical techniques, such as those used in the early 1900s by Mangold and Spemann, have been instrumental in advancing our understanding of embryonic development. However, these techniques are highly specialized, leading to issues of inter-operator variability. Here we introduce a user-friendly robotic microsurgery platform that allows precise mechanical manipulation of soft tissues in zebrafish embryos. Using our platform, we reproducibly targeted precise regions of tail explants, and quantified the response in real-time by following notochord and presomitic mesoderm (PSM) morphogenesis and segmentation clock dynamics during vertebrate anteroposterior axis elongation. We find an extension force generated through the posterior notochord that is strong enough to buckle the structure. Our data suggest that this force generates a unidirectional notochord extension towards the tailbud because PSM tissue around the posterior notochord does not let it slide anteriorly. These results complement existing biomechanical models of axis elongation, revealing a critical coupling between the posterior notochord, the tailbud, and the PSM, and show that somite patterning is robust against structural perturbations.


Assuntos
Robótica , Peixe-Zebra , Animais , Morfogênese , Somitos , Mesoderma , Notocorda/fisiologia , Micromanipulação , Padronização Corporal/fisiologia
9.
Nature ; 612(7941): 732-738, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36517595

RESUMO

Our understanding of human early development is severely hampered by limited access to embryonic tissues. Due to their close evolutionary relationship with humans, nonhuman primates are often used as surrogates to understand human development but currently suffer from a lack of in vivo datasets, especially from gastrulation to early organogenesis during which the major embryonic cell types are dynamically specified. To fill this gap, we collected six Carnegie stage 8-11 cynomolgus monkey (Macaca fascicularis) embryos and performed in-depth transcriptomic analyses of 56,636 single cells. Our analyses show transcriptomic features of major perigastrulation cell types, which help shed light on morphogenetic events including primitive streak development, somitogenesis, gut tube formation, neural tube patterning and neural crest differentiation in primates. In addition, comparative analyses with mouse embryos and human embryoids uncovered conserved and divergent features of perigastrulation development across species-for example, species-specific dependency on Hippo signalling during presomitic mesoderm differentiation-and provide an initial assessment of relevant stem cell models of human early organogenesis. This comprehensive single-cell transcriptome atlas not only fills the knowledge gap in the nonhuman primate research field but also serves as an invaluable resource for understanding human embryogenesis and developmental disorders.


Assuntos
Gastrulação , Macaca fascicularis , Organogênese , Análise de Célula Única , Animais , Humanos , Camundongos , Gastrulação/genética , Macaca fascicularis/embriologia , Macaca fascicularis/genética , Organogênese/genética , Corpos Embrioides , Perfilação da Expressão Gênica , Linha Primitiva/citologia , Linha Primitiva/embriologia , Tubo Neural/citologia , Tubo Neural/embriologia , Crista Neural/citologia , Crista Neural/embriologia , Via de Sinalização Hippo , Mesoderma/citologia , Mesoderma/embriologia , Células-Tronco
10.
Front Cell Infect Microbiol ; 12: 887278, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36389136

RESUMO

BCL6A is a transcriptional repressor implicated in the development and survival of B and T lymphoctyes, which is also highly expressed in many non-Hodgkin's lymphomas, such as diffuse large B cell lymphoma and follicular lymphoma. Roles in other cell types, including macrophages and non-hematopoietic cells, have also been suggested but require further investigation. This study sought to identify and characterize zebrafish BCL6A and investigate its role in immune cell development and function, with a focus on early macrophages. Bioinformatics analysis identified a homologue for BCL6A (bcl6aa), as well as an additional fish-specific duplicate (bcl6ab) and a homologue for the closely-related BCL6B (bcl6b). The human BCL6A and zebrafish Bcl6aa proteins were highly conserved across the constituent BTB/POZ, PEST and zinc finger domains. Expression of bcl6aa during early zebrafish embryogenesis was observed in the lateral plate mesoderm, a site of early myeloid cell development, with later expression seen in the brain, eye and thymus. Homozygous bcl6aa mutants developed normally until around 14 days post fertilization (dpf), after which their subsequent growth and maturation was severely impacted along with their relative survival, with heterozygous bcl6aa mutants showing an intermediate phenotype. Analysis of immune cell development revealed significantly decreased lymphoid and macrophage cells in both homozygous and heterozygous bcl6aa mutants, being exacerbated in homozygous mutants. In contrast, the number of neutrophils was unaffected. Only the homozygous bcl6aa mutants showed decreased macrophage mobility in response to wounding and reduced ability to contain bacterial infection. Collectively, this suggests strong conservation of BCL6A across evolution, including a role in macrophage biology.


Assuntos
Linfoma de Células B , Peixe-Zebra , Animais , Humanos , Macrófagos , Mesoderma , Fatores de Transcrição/metabolismo , Peixe-Zebra/genética
11.
Nat Commun ; 13(1): 7137, 2022 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-36414616

RESUMO

The lung's gas exchange surface is comprised of alveolar AT1 and AT2 cells that are corrupted in several common and deadly diseases. They arise from a bipotent progenitor whose differentiation is thought to be dictated by differential mechanical forces. Here we show the critical determinant is FGF signaling. Fgfr2 is expressed in the developing progenitors in mouse then restricts to nascent AT2 cells and remains on throughout life. Its ligands are expressed in surrounding mesenchyme and can, in the absence of exogenous mechanical cues, induce progenitors to form alveolospheres with intermingled AT2 and AT1 cells. FGF signaling directly and cell autonomously specifies AT2 fate; progenitors lacking Fgfr2 in vitro and in vivo exclusively acquire AT1 fate. Fgfr2 loss in AT2 cells perinatally results in reprogramming to AT1 identity, whereas loss or inhibition later in life triggers AT2 apoptosis and compensatory regeneration. We propose that Fgfr2 signaling selects AT2 fate during development, induces a cell non-autonomous AT1 differentiation signal, then continuously maintains AT2 identity and survival throughout life.


Assuntos
Células Epiteliais Alveolares , Mesoderma , Animais , Camundongos , Diferenciação Celular , Transdução de Sinais , Apoptose
12.
Tissue Cell ; 79: 101970, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36370625

RESUMO

Mesenchymal stem cells (MSCs) are widely present in the interstitial tissue of embryos. Although the existence of the metanephros stromal stem cell population has been demonstrated, the focus has been on understanding the process of nephrogenesis, but the biological characterization of stromal stem cell population is less precise. Metanephric mesenchymal stem cells (MMSCs) have a vast potential in kidney tissue engineering and represent excellent candidates in cellular replacement therapy for human disease and veterinary research. Here, we aimed to isolate, culture, and characterize bovine MMSCs. We have successfully obtained a new population of stem cells using renal tissue isolated from three-month-old bovine embryo. MMSCs were isolated by collagenase digestion. The spindle-shaped cells adhered to plastic and exhibited extensive proliferation for more than 26 passages and good clonogenic ability in vitro. Moreover, the metanephric mesenchymal stem cells could be induced to differentiate into mesoderm-derived cells (such as osteoblasts, chondrocytes, and adipocytes) and endoderm-derived hepatocyte-like cells in vitro. These results indicated the multiple differentiations potential of MMSCs. Aside from colony-forming, self-renewal, and multilineage differentiation capabilities, the experiments of immunofluorescence and RT-PCR showed that spindle-shaped cells were positive for MSCs-related markers (CD29, CD44, CD73, CD90, CD106), nestin and vimentin, while the hematopoietic cell surface markers CD34 and pan-leukocyte marker CD45 were undetectable. This study provides a technical platform for the preservation of valuable bovine genetic resources and offers a new source for tissue engineering and cell transplantation therapy.


Assuntos
Células-Tronco Mesenquimais , Mesoderma , Humanos , Bovinos , Animais , Lactente , Terapia Baseada em Transplante de Células e Tecidos , Diferenciação Celular , Engenharia Tecidual
13.
Ann Plast Surg ; 89(6): 684-693, 2022 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-36416704

RESUMO

BACKGROUND: This study evaluated the potential of Wharton's jelly mesenchymal stem cells with high tolerogenic properties in reducing immunosuppressive dosage and related adverse effects. METHODS: A 4- to 6-week-old, 30-40 g weight, male inbred CD57BL/6 mice were used as skin allograft donors, whereas Balb/c mice with similar characteristics were used as recipients. Wharton's jelly stem cells were obtained from a commercial kit sourced from human umbilical cord. Skin allografts were performed from CD57Bl6 to Balb/c mice (day 0). Group 1 (control) received no treatment. Group 2 received 15 mg/kg cyclosporin A on days 0 to 30. Group 3 received 5.7 × 10 6 and 10.3 × 10 6 cell/kg Wharton's jelly stem cells on days 0 and 3, respectively. Groups 4, 5, and 6 received a combination of 15, 10, and 5 mg/kg per day cyclosporine A (days 0 to 30) with the same stem cell dose with group 3, respectively. Graft rejection was evaluated with digital photography and thermal imaging, histopathology (Banff grading, epithelialization scores, dermoepidermal dissociation), immunochemistry (Ki-67 and Bcl-2), and biochemical methods (interleukin 10, interleukin 2, interferon γ, tumor necrosis factor α) (day 10). Cumulative adverse effects of cyclosporin A occurring in the groups were revealed by histopathological evaluation of kidney and liver (a modified semiquantitative method of infiltration of inflammatory cells around the portal area and lobular region in liver; modification of the Banff rating of proximal tubules and hypertrophia of juxtaglomerular apparatus cells in kidney) (day 30). RESULTS: There was no rejection in groups 2, 4, and 5 until the end of study. These were statistically different versus groups 1 (day 10 ± 0.71), 3 (day 11 ± 0.82), and 6 (day 11 ± 0.58) (all P 's < 0.05). Groups 4 and 5 have exhibited statistically similar findings in histopathological (4 epithelization score: 3.7 ± 1.3; 5 epithelization score: 3.5 ± 0.5; 4 Banff grading score: 0.8 ± 0.6; 5 Banff grading score: 1.0 ± 0.5; both P 's = 1.00), immunohistochemical (4 Bcl-2 score: 3.5 ± 0.5, P = 0.618; 5 Bcl-2 score: 3.4 ± 0.5, P = 1.00; 4 Ki-67 score: 3.7 ± 0.4, P = 1.00; 5 Ki-67 score: 3.5 ± 0.5, both P 's = 1.00), and levels of cytokines (both P 's = 1.00) versus group 2. Adverse effects on kidneys and liver were lowest and statistically similar in groups 3, 5, and 6 (all P 's = 00) versus group 1. CONCLUSIONS: Wharton's jelly mesenchymal stem cells alter bioavailability of cyclosporine, albeit at much lower doses and with fewer systemic adverse effects.


Assuntos
Ciclosporina , Células-Tronco Mesenquimais , Masculino , Humanos , Camundongos , Animais , Ciclosporina/farmacologia , Antígeno Ki-67 , Cordão Umbilical , Mesoderma , Imunossupressores/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2
14.
Cell Rep ; 41(8): 111701, 2022 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-36417876

RESUMO

The mouse digit tip regenerates following amputation. How the regenerate is patterned is unknown, but a long-standing hypothesis proposes developmental patterning mechanisms are re-used during regeneration. The digit tip bone exhibits dorsal-ventral (DV) polarity, so we focus on En1 and Lmx1b, two factors necessary for DV patterning during limb development. We investigate whether they are re-expressed during regeneration in a developmental-like pattern and whether they direct DV morphology of the regenerate. We find that both En1 and Lmx1b are expressed in the regenerating digit tip epithelium and mesenchyme, respectively, but without DV polarity. Conditional genetics and quantitative analysis of digit tip bone morphology determine that genetic deletion of En1 or Lmx1b in adult digit tip regeneration modestly reduces bone regeneration but does not affect DV patterning. Collectively, our data suggest that, while En1 and Lmx1b are re-expressed during mouse digit tip regeneration, they do not define the DV axis during regeneration.


Assuntos
Mesoderma , Camundongos , Animais , Regeneração Óssea , Osso e Ossos
15.
Nat Ecol Evol ; 6(12): 1921-1939, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36396969

RESUMO

Transcription factors are crucial drivers of cellular differentiation during animal development and often share ancient evolutionary origins. The T-box transcription factor Brachyury plays a pivotal role as an early mesoderm determinant and neural repressor in vertebrates; yet, the ancestral function and key evolutionary transitions of the role of this transcription factor remain obscure. Here, we present a genome-wide target-gene screen using chromatin immunoprecipitation sequencing in the sea anemone Nematostella vectensis, an early branching non-bilaterian, and the sea urchin Strongylocentrotus purpuratus, a representative of the sister lineage of chordates. Our analysis reveals an ancestral gene regulatory feedback loop connecting Brachyury, FoxA and canonical Wnt signalling involved in axial patterning that predates the cnidarian-bilaterian split about 700 million years ago. Surprisingly, we also found that part of the gene regulatory network controlling the fate of neuromesodermal progenitors in vertebrates was already present in the common ancestor of cnidarians and bilaterians. However, while several endodermal and neuronal Brachyury target genes are ancestrally shared, hardly any of the key mesodermal downstream targets in vertebrates are found in the sea anemone or the sea urchin. Our study suggests that a limited number of target genes involved in mesoderm formation were newly acquired in the vertebrate lineage, leading to a dramatic shift in the function of this ancestral developmental regulator.


Assuntos
Mesoderma , Anêmonas-do-Mar , Animais , Retroalimentação , Fatores de Transcrição , Anêmonas-do-Mar/genética
16.
Development ; 149(23)2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36367707

RESUMO

Certain cranial neural crest cells are uniquely endowed with the ability to make skeletal cell types otherwise only derived from mesoderm. As these cells migrate into the pharyngeal arches, they downregulate neural crest specifier genes and upregulate so-called ectomesenchyme genes that are characteristic of skeletal progenitors. Although both external and intrinsic factors have been proposed as triggers of this transition, the details remain obscure. Here, we report the Nr2f nuclear receptors as intrinsic activators of the ectomesenchyme program: zebrafish nr2f5 single and nr2f2;nr2f5 double mutants show marked delays in upregulation of ectomesenchyme genes, such as dlx2a, prrx1a, prrx1b, sox9a, twist1a and fli1a, and in downregulation of sox10, which is normally restricted to early neural crest and non-ectomesenchyme lineages. Mutation of sox10 fully rescued skeletal development in nr2f5 single but not nr2f2;nr2f5 double mutants, but the initial ectomesenchyme delay persisted in both. Sox10 perdurance thus antagonizes the recovery but does not explain the impaired ectomesenchyme transition. Unraveling the mechanisms of Nr2f function will help solve the enduring puzzle of how cranial neural crest cells transition to the skeletal progenitor state.


Assuntos
Placa Neural , Peixe-Zebra , Animais , Peixe-Zebra/genética , Crista Neural , Mesoderma , Receptores Citoplasmáticos e Nucleares/genética , Regulação da Expressão Gênica no Desenvolvimento
17.
Dev Cell ; 57(21): 2450-2468.e7, 2022 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-36347239

RESUMO

The mammalian genome encodes thousands of long non-coding RNAs (lncRNAs), many of which are developmentally regulated and differentially expressed across tissues, suggesting their potential roles in cellular differentiation. Despite this expression pattern, little is known about how lncRNAs influence lineage commitment at the molecular level. Here, we demonstrate that perturbation of an embryonic stem cell/early embryonic lncRNA, pluripotency-associated transcript 4 (Platr4), directly influences the specification of cardiac-mesoderm-lineage differentiation. We show that Platr4 acts as a molecular scaffold or chaperone interacting with the Hippo-signaling pathway molecules Yap and Tead4 to regulate the expression of a downstream target gene, Ctgf, which is crucial to the cardiac-lineage program. Importantly, Platr4 knockout mice exhibit myocardial atrophy and valve mucinous degeneration, which are both associated with reduced cardiac output and sudden heart failure. Together, our findings provide evidence that Platr4 is required in cardiac-lineage specification and adult heart function in mice.


Assuntos
RNA Longo não Codificante , Camundongos , Animais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Células-Tronco Embrionárias , Mesoderma/metabolismo , Diferenciação Celular/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Linhagem da Célula/genética , Mamíferos/metabolismo
18.
EMBO J ; 41(23): e110928, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36245268

RESUMO

Each vertebrate species appears to have a unique timing mechanism for forming somites along the vertebral column, and the process in human remains poorly understood at the molecular level due to technical and ethical limitations. Here, we report the reconstitution of human segmentation clock by direct reprogramming. We first reprogrammed human urine epithelial cells to a presomitic mesoderm (PSM) state capable of long-term self-renewal and formation of somitoids with an anterior-to-posterior axis. By inserting the RNA reporter Pepper into HES7 and MESP2 loci of these iPSM cells, we show that both transcripts oscillate in the resulting somitoids at ~5 h/cycle. GFP-tagged endogenous HES7 protein moves along the anterior-to-posterior axis during somitoid formation. The geo-sequencing analysis further confirmed anterior-to-posterior polarity and revealed the localized expression of WNT, BMP, FGF, and RA signaling molecules and HOXA-D family members. Our study demonstrates the direct reconstitution of human segmentation clock from somatic cells, which may allow future dissection of the mechanism and components of such a clock and aid regenerative medicine.


Assuntos
Mesoderma , Somitos , Humanos , Somitos/metabolismo , Mesoderma/metabolismo , Transdução de Sinais , Regulação da Expressão Gênica no Desenvolvimento , Padronização Corporal/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo
19.
Biomed Mater ; 17(6)2022 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-36270422

RESUMO

Three-dimensional cell constructs comprising only tissue-specific cells and extracellular matrix secreted by them would be ideal transplants, but their fabrication in a cell aggregation manner without cell scaffolds relies on random cell self-aggregation, making the control of their size and shape difficult. In this study, we propose a method to fabricate band-shaped tissues by inducing the self-aggregation of cell sheets using the developed cell self-aggregation technique (CAT). Acting as cell aggregation stoppers, silicone semicircular pillars were attached to two positions equidistant from both short ends of the rounded rectangular culture groove and coated with a specifically charged biomimetic polymer as a CAT-inducing surface. Mesenchymal stem cells, chondrocytes, and skeletal myoblast cells seeded on the surface of the culture grooves formed band-shaped aggregates between the two aggregation stoppers following spontaneous detachment with aggregation of the cell sheet from the outer edge of the grooves during day one of culture. The aggregated chondrocyte band matured into a cartilage-like plate with an abundant cartilage matrix while retaining its band shape after two weeks of chondrogenic cultivation. Additionally, the aggregates of mesenchymal stem cells and myoblast cell bands could patch the induced collagen membrane derived from rat subcutaneous tissue like a bandage immediately after their formation and successfully mature into fat and muscle tissues, respectively. These results indicate that, depending on the cell type, scaffold-free band-shaped cell aggregates produced by CAT have the potential to achieve tissue regeneration that follows the shape of the defect viain vitromaturation culture orin vivoorganization.


Assuntos
Condrogênese , Células-Tronco Mesenquimais , Ratos , Animais , Cartilagem/fisiologia , Condrócitos , Mesoderma
20.
BMC Genomics ; 23(1): 723, 2022 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-36273135

RESUMO

BACKGROUND: During embryogenesis, the developmental potential of initially pluripotent cells becomes progressively restricted as they transit to lineage restricted states. The pluripotent cells of Xenopus blastula-stage embryos are an ideal system in which to study cell state transitions during developmental decision-making, as gene expression dynamics can be followed at high temporal resolution. RESULTS: Here we use transcriptomics to interrogate the process by which pluripotent cells transit to four different lineage-restricted states: neural progenitors, epidermis, endoderm and ventral mesoderm, providing quantitative insights into the dynamics of Waddington's landscape. Our findings provide novel insights into why the neural progenitor state is the default lineage state for pluripotent cells and uncover novel components of lineage-specific gene regulation. These data reveal an unexpected overlap in the transcriptional responses to BMP4/7 and Activin signaling and provide mechanistic insight into how the timing of signaling inputs such as BMP are temporally controlled to ensure correct lineage decisions. CONCLUSIONS: Together these analyses provide quantitative insights into the logic and dynamics of developmental decision making in early embryos. They also provide valuable lineage-specific time series data following the acquisition of specific lineage states during development.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Transcriptoma , Mesoderma , Endoderma/metabolismo , Ativinas/genética , Ativinas/metabolismo , Diferenciação Celular/genética , Proteínas de Xenopus/genética , Proteínas de Xenopus/metabolismo
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