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1.
Cell Stem Cell ; 31(8): 1113-1126.e6, 2024 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-38981471

RESUMEN

Emerging human pluripotent stem cell (hPSC)-based embryo models are useful for studying human embryogenesis. Particularly, there are hPSC-based somitogenesis models using free-floating culture that recapitulate somite formation. Somitogenesis in vivo involves intricately orchestrated biochemical and biomechanical events. However, none of the current somitogenesis models controls biochemical gradients or biomechanical signals in the culture, limiting their applicability to untangle complex biochemical-biomechanical interactions that drive somitogenesis. Herein, we develop a human somitogenesis model by confining hPSC-derived presomitic mesoderm (PSM) tissues in microfabricated trenches. Exogenous microfluidic morphogen gradients imposed on the PSM tissues cause axial patterning and trigger spontaneous rostral-to-caudal somite formation. A mechanical theory is developed to explain the size dependency between somites and the PSM. The microfluidic somitogenesis model is further exploited to reveal regulatory roles of cellular and tissue biomechanics in somite formation. This study presents a useful microengineered, hPSC-based model for understanding the biochemical and biomechanical events that guide somite formation.


Asunto(s)
Microfluídica , Modelos Biológicos , Células Madre Pluripotentes , Somitos , Humanos , Células Madre Pluripotentes/citología , Células Madre Pluripotentes/metabolismo , Somitos/citología , Somitos/metabolismo , Microfluídica/métodos , Desarrollo Embrionario , Mesodermo/citología , Diferenciación Celular
2.
Nat Commun ; 15(1): 5055, 2024 Jun 13.
Artículo en Inglés | MEDLINE | ID: mdl-38871742

RESUMEN

The anterior-posterior axis of the mammalian embryo is laid down by the anterior visceral endoderm (AVE), an extraembryonic signaling center that is specified within the visceral endoderm. Current models posit that AVE differentiation is promoted globally by epiblast-derived Nodal signals, and spatially restricted by a BMP gradient established by the extraembryonic ectoderm. Here, we report spatially restricted AVE differentiation in bilayered embryo-like aggregates made from mouse embryonic stem cells that lack an extraembryonic ectoderm. Notably, clusters of AVE cells also form in pure visceral endoderm cultures upon activation of Nodal signaling, indicating that tissue-intrinsic factors can restrict AVE differentiation. We identify ß-catenin activity as a tissue-intrinsic factor that antagonizes AVE-inducing Nodal signals. Together, our results show how an AVE-like population can arise through interactions between epiblast and visceral endoderm alone. This mechanism may be a flexible solution for axis patterning in a wide range of embryo geometries, and provide robustness to axis patterning when coupled with signal gradients.


Asunto(s)
Tipificación del Cuerpo , Diferenciación Celular , Endodermo , Proteína Nodal , Transducción de Señal , beta Catenina , Animales , Endodermo/citología , Endodermo/metabolismo , Endodermo/embriología , beta Catenina/metabolismo , Ratones , Proteína Nodal/metabolismo , Proteína Nodal/genética , Estratos Germinativos/metabolismo , Estratos Germinativos/citología , Células Madre Embrionarias de Ratones/metabolismo , Células Madre Embrionarias de Ratones/citología , Regulación del Desarrollo de la Expresión Génica , Embrión de Mamíferos/citología
3.
Nature ; 628(8007): 391-399, 2024 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-38408487

RESUMEN

The human nervous system is a highly complex but organized organ. The foundation of its complexity and organization is laid down during regional patterning of the neural tube, the embryonic precursor to the human nervous system. Historically, studies of neural tube patterning have relied on animal models to uncover underlying principles. Recently, models of neurodevelopment based on human pluripotent stem cells, including neural organoids1-5 and bioengineered neural tube development models6-10, have emerged. However, such models fail to recapitulate neural patterning along both rostral-caudal and dorsal-ventral axes in a three-dimensional tubular geometry, a hallmark of neural tube development. Here we report a human pluripotent stem cell-based, microfluidic neural tube-like structure, the development of which recapitulates several crucial aspects of neural patterning in brain and spinal cord regions and along rostral-caudal and dorsal-ventral axes. This structure was utilized for studying neuronal lineage development, which revealed pre-patterning of axial identities of neural crest progenitors and functional roles of neuromesodermal progenitors and the caudal gene CDX2 in spinal cord and trunk neural crest development. We further developed dorsal-ventral patterned microfluidic forebrain-like structures with spatially segregated dorsal and ventral regions and layered apicobasal cellular organizations that mimic development of the human forebrain pallium and subpallium, respectively. Together, these microfluidics-based neurodevelopment models provide three-dimensional lumenal tissue architectures with in vivo-like spatiotemporal cell differentiation and organization, which will facilitate the study of human neurodevelopment and disease.


Asunto(s)
Tipificación del Cuerpo , Microfluídica , Tubo Neural , Humanos , Técnicas de Cultivo Tridimensional de Células , Diferenciación Celular , Cresta Neural/citología , Cresta Neural/embriología , Tubo Neural/citología , Tubo Neural/embriología , Células Madre Pluripotentes/citología , Prosencéfalo/citología , Prosencéfalo/embriología , Médula Espinal/citología , Médula Espinal/embriología
4.
bioRxiv ; 2023 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-37961125

RESUMEN

Emerging human pluripotent stem cell (hPSC)-based embryo models are useful for studying human embryogenesis. Particularly, there are hPSC-based somitogenesis models using free-floating culture that recapitulate somite formation. Somitogenesis in vivo involves intricately orchestrated bio-chemical and -mechanical events. However, none of the current somitogenesis models controls biochemical gradients or biomechanical signals in the culture, limiting their applicability to untangle complex biochemical-biomechanical interactions that drive somitogenesis. Here we report a new human somitogenesis model by confining hPSC-derived presomitic mesoderm (PSM) tissues in microfabricated trenches. Exogenous microfluidic morphogen gradients imposed on PSM cause axial patterning and trigger spontaneous rostral-to-caudal somite formation. A mechanical theory is developed to explain the size dependency between somites and PSM. The microfluidic somitogenesis model is further exploited to reveal regulatory roles of cellular and tissue biomechanics in somite formation. This study presents a useful microengineered, hPSC-based model for understanding the bio-chemical and -mechanical events that guide somite formation.

5.
Sci Adv ; 8(44): eabl9583, 2022 Nov 04.
Artículo en Inglés | MEDLINE | ID: mdl-36332016

RESUMEN

Two fundamental elements of pre-implantation embryogenesis are cells' intrinsic self-organization program and their developmental plasticity, which allows embryos to compensate for alterations in cell position and number; yet, these elements are still poorly understood. To be able to decipher these features, we established culture conditions that enable the two fates of blastocysts' extraembryonic lineages-the primitive endoderm and the trophectoderm-to coexist. This plasticity emerges following the mechanisms of the first lineage segregation in the mouse embryo, and it manifests as an extended potential for extraembryonic chimerism during the pre-implantation embryogenesis. Moreover, this shared state enables robust assembly into higher-order blastocyst-like structures, thus combining both the cell fate plasticity and self-organization features of the early extraembryonic lineages.

6.
Dev Cell ; 57(16): 1937-1956.e8, 2022 08 22.
Artículo en Inglés | MEDLINE | ID: mdl-35998584

RESUMEN

The complex architecture of the murine fetus originates from a simple ball of pluripotent epiblast cells, which initiate morphogenesis upon implantation. In turn, this establishes an intermediate state of tissue-scale organization of the embryonic lineage in the form of an epithelial monolayer, where patterning signals delineate the body plan. However, how this major morphogenetic process is orchestrated on a cellular level and synchronized with the developmental progression of the epiblast is still obscure. Here, we identified that the small GTPase Rap1 plays a critical role in reshaping the pluripotent lineage. We found that Rap1 activity is controlled via Oct4/Esrrb input and is required for the transmission of polarization cues, which enables the de novo epithelialization and formation of tricellular junctions in the epiblast. Thus, Rap1 acts as a molecular switch that coordinates the morphogenetic program in the embryonic lineage, in sync with the cellular states of pluripotency.


Asunto(s)
Implantación del Embrión , Estratos Germinativos , Animales , Diferenciación Celular , Regulación del Desarrollo de la Expresión Génica , Ratones , Morfogénesis
7.
Semin Cell Dev Biol ; 131: 110-116, 2022 11.
Artículo en Inglés | MEDLINE | ID: mdl-35513973

RESUMEN

As the early mouse embryo develops, fundamental steps include the sequential formation of the first lumens in the murine conceptus. The first cavity established in the pre-implantation embryo is the blastocoel, followed by the emergence of the proamniotic cavity during the peri-implantation stages. The mouse embryo is a dynamic system which switches its modes of lumenogenesis before and after implantation. The blastocoel emerges in between the basolateral membranes, whereas the proamniotic cavity is formed on the apical interface. Defects in the sculpting of these luminal spaces are associated with developmental abnormalities and embryonic lethality. Here, we review the mechanisms by which these early embryonic cavities are formed and discuss the cavities in terms of their common and stage-specific principles of lumenogenesis and their functions.


Asunto(s)
Blastocisto , Endodermo , Animales , Implantación del Embrión , Embrión de Mamíferos , Desarrollo Embrionario , Ratones
8.
Sci Adv ; 7(11)2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33692105

RESUMEN

During the peri-implantation stages, the mouse embryo radically changes its appearance, transforming from a hollow-shaped blastocyst to an egg cylinder. At the same time, the epiblast gets reorganized from a simple ball of cells to a cup-shaped epithelial monolayer enclosing the proamniotic cavity. However, the cavity's function and mechanism of formation have so far been obscure. Through investigating the cavity formation, we found that in the epiblast, the process of lumenogenesis is driven by reorganization of intercellular adhesion, vectoral fluid transport, and mitotic paracellular water influx from the blastocoel into the emerging proamniotic cavity. By experimentally blocking lumenogenesis, we found that the proamniotic cavity functions as a hub for communication between the early lineages, enabling proper growth and patterning of the postimplantation embryo.

9.
Nat Commun ; 11(1): 5499, 2020 10 30.
Artículo en Inglés | MEDLINE | ID: mdl-33127892

RESUMEN

The epiblast, which provides the foundation of the future body, is actively reshaped during early embryogenesis, but the reshaping mechanisms are poorly understood. Here, using a 3D in vitro model of early epiblast development, we identify the canonical Wnt/ß-catenin pathway and its central downstream factor Esrrb as the key signalling cascade regulating the tissue-scale organization of the murine pluripotent lineage. Although in vivo the Wnt/ß-catenin/Esrrb circuit is dispensable for embryonic development before implantation, autocrine Wnt activity controls the morphogenesis and long-term maintenance of the epiblast when development is put on hold during diapause. During this phase, the progressive changes in the epiblast architecture and Wnt signalling response show that diapause is not a stasis but instead is a dynamic process with underlying mechanisms that can appear redundant during transient embryogenesis.


Asunto(s)
Diapausa/fisiología , Células Madre Embrionarias/metabolismo , Receptores de Estrógenos/metabolismo , Vía de Señalización Wnt/fisiología , Animales , Desarrollo Embrionario , Femenino , Estratos Germinativos/metabolismo , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Morfogénesis , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores de Estrógenos/genética , beta Catenina/genética
10.
Stem Cells Dev ; 25(1): 1-12, 2016 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-26456050

RESUMEN

Müller glia are responsible for the retina regeneration observed in zebrafish. Although the human retina harbors Müller glia with stem cell characteristics, there is no evidence that they regenerate the retina after disease or injury. Transforming growth factor-ß (TGFß) and Wnt signaling regulate retinal neurogenesis and inflammation, but their roles in the neural differentiation of human Müller stem cells (hMSC) are not known. We examined hMSC lines in vitro for the expression of various Wnt signaling components and for their modulation by TGFß1, as well as the effect of this cytokine on the photoreceptor differentiation of these cells. Culture of hMSC with a combination of factors that induce photoreceptor differentiation of hMSC (FGF2, taurine, retinoic acid, and insulin-like growth factor type1; FTRI), markedly upregulated the expression of components of the canonical Wnt signaling pathway, including WNT2B, DKK1, and active ß-CATENIN. Although FTRI did not modify mRNA expression of WNT5B, a component of the noncanonical/planar cell polarity Wnt pathway, it upregulated its secretion. Furthermore, TGFß1 not only decreased WNT2B expression, but also inhibited FTRI-induced photoreceptor differentiation of hMSC, as determined by expression of the photoreceptor markers NR2E3, RHODOPSIN, and RECOVERIN. Inhibition of TGFß1 signaling by an ALK5 inhibitor prevented TGFß1-induced changes in the expression of the two Wnt ligands examined. More importantly, inhibition of the canonical WNT signaling by XAV-939 prevented FTRI-induced photoreceptor differentiation. These observations suggest that TGFß may play a key role in preventing neural differentiation of hMSC and may constitute a potential target for induction of endogenous regeneration of the human retina.


Asunto(s)
Diferenciación Celular/efectos de los fármacos , Células Ependimogliales/efectos de los fármacos , Células Fotorreceptoras/efectos de los fármacos , Factor de Crecimiento Transformador beta1/farmacología , Vía de Señalización Wnt/efectos de los fármacos , Adulto , Diferenciación Celular/genética , Línea Celular , Regulación hacia Abajo/efectos de los fármacos , Células Ependimogliales/citología , Células Ependimogliales/fisiología , Regulación de la Expresión Génica/efectos de los fármacos , Glicoproteínas/genética , Glicoproteínas/metabolismo , Humanos , Neurogénesis/efectos de los fármacos , Neurogénesis/genética , Células Fotorreceptoras/fisiología , Células Madre/citología , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Vía de Señalización Wnt/genética
11.
PLoS One ; 10(11): e0142049, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26561861

RESUMEN

Substance dependence is a frequently observed comorbid disorder in schizophrenia, but little is known about genetic factors possibly shared between the two psychotic disorders. GABRB2, a schizophrenia candidate gene coding for GABAA receptor ß2 subunit, is examined for possible association with heroin dependence in Han Chinese population. Four single nucleotide polymorphisms (SNPs) in GABRB2, namely rs6556547 (S1), rs1816071 (S3), rs18016072 (S5), and rs187269 (S29), previously associated with schizophrenia, were examined for their association with heroin dependence. Two additional SNPs, rs10051667 (S31) and rs967771 (S32), previously associated with alcohol dependence and bipolar disorder respectively, were also analyzed. The six SNPs were genotyped by direct sequencing of PCR amplicons of target regions for 564 heroin dependent individuals and 498 controls of Han Chinese origin. Interestingly, it was found that recombination between the haplotypes of all-derived-allele (H1; OR = 1.00) and all-ancestral-allele (H2; OR = 0.74) at S5-S29 junction generated two recombinants H3 (OR = 8.51) and H4 (OR = 5.58), both conferring high susceptibility to heroin dependence. Additional recombination between H2 and H3 haplotypes at S1-S3 junction resulted in a risk-conferring haplotype H5 (OR = 1.94x109). In contrast, recombination between H1 and H2 haplotypes at S3-S5 junction rescued the risk-conferring effect of recombination at S5-S29 junction, giving rise to the protective haplotype H6 (OR = 0.68). Risk-conferring effects of S1-S3 and S5-S29 crossovers and protective effects of S3-S5 crossover were seen in both pure heroin dependent and multiple substance dependence subgroups. In conclusion, significant association was found with haplotypes of the S1-S29 segment in GABRB2 for heroin dependence in Han Chinese population. Local recombination was an important determining factor for switching haplotypes between risk-conferring and protective statuses. The present study provide evidence for the schizophrenia candidate gene GABRB2 to play a role in heroin dependence, but replication of these findings is required.


Asunto(s)
Predisposición Genética a la Enfermedad/genética , Haplotipos , Dependencia de Heroína/genética , Polimorfismo de Nucleótido Simple , Receptores de GABA-A/genética , Adulto , Pueblo Asiatico/genética , China , Femenino , Frecuencia de los Genes , Predisposición Genética a la Enfermedad/etnología , Genotipo , Dependencia de Heroína/etnología , Humanos , Desequilibrio de Ligamiento , Masculino , Oportunidad Relativa , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Adulto Joven
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