Project description:Neuromuscular networks assemble during early human embryonic development and are essential for the control of body movement. Previous neuromuscular junction modeling efforts using human pluripotent stem cells (hPSCs) generated either spinal cord neurons or skeletal muscles in monolayer culture. Here, we use hPSC-derived axial stem cells, the building blocks of the posterior body, to simultaneously generate spinal cord neurons and skeletal muscle cells that self-organize to generate human neuromuscular organoids (NMOs) that can be maintained in 3D for several months. Single-cell RNA-sequencing of individual organoids revealed reproducibility across experiments and enabled the tracking of the neural and mesodermal differentiation trajectories as organoids developed and matured. NMOs contain functional neuromuscular junctions supported by terminal Schwann cells. They contract and develop central pattern generator-like neuronal circuits. Finally, we successfully use NMOs to recapitulate key aspects of myasthenia gravis pathology, thus highlighting the significant potential of NMOs for modeling neuromuscular diseases in the future.
Project description:Gastruloids have recently emerged as an efficient four-dimensional model for studying some aspects of post-implantation embryonic patterning. Their ability to undergo gastrulation-like processes leading to the self-organization into highly reproducible biological objects, has opened new avenues to investigate early embryonic patterning. Here, we sought to uncover the molecular and cellular mechanism underlying this remarkable property. We report that self-organization competence is associated with a cell-specific coordination of a Cadherin switch. We find that N-Cadherin hinders gastruloids morphogenetic competence, for its inactivation leads to the formation of trunk-like structures without relying on the otherwise requested extra-cellular matrix analogues such as Matrigel. In contrast, the repression of E-Cadherin by Snai1 is critical for self-organization: Snai1 establishes a cell-specific repressive pace by triggering the repression of a pluripotency-associated transcription program and its chromatin landscape, thus allowing a proper transition from E- to N-Cadherin to occur. Altogether, this work establishes a molecular mechanism that integrates the exit from pluripotency and the pace of cell differentiation, leading to the observed self-organizing potential of gastruloids.
Project description:Gastruloids have recently emerged as an efficient four-dimensional model for studying some aspects of post-implantation embryonic patterning. Their ability to undergo gastrulation-like processes leading to the self-organization into highly reproducible biological objects, has opened new avenues to investigate early embryonic patterning. Here, we sought to uncover the molecular and cellular mechanism underlying this remarkable property. We report that self-organization competence is associated with a cell-specific coordination of a Cadherin switch. We find that N-Cadherin hinders gastruloids morphogenetic competence, for its inactivation leads to the formation of trunk-like structures without relying on the otherwise requested extra-cellular matrix analogues such as Matrigel. In contrast, the repression of E-Cadherin by Snai1 is critical for self-organization: Snai1 establishes a cell-specific repressive pace by triggering the repression of a pluripotency-associated transcription program and its chromatin landscape, thus allowing a proper transition from E- to N-Cadherin to occur. Altogether, this work establishes a molecular mechanism that integrates the exit from pluripotency and the pace of cell differentiation, leading to the observed self-organizing potential of gastruloids.
Project description:Gastruloids have recently emerged as an efficient four-dimensional model for studying some aspects of post-implantation embryonic patterning. Their ability to undergo gastrulation-like processes leading to the self-organization into highly reproducible biological objects, has opened new avenues to investigate early embryonic patterning. Here, we sought to uncover the molecular and cellular mechanism underlying this remarkable property. We report that self-organization competence is associated with a cell-specific coordination of a Cadherin switch. We find that N-Cadherin hinders gastruloids morphogenetic competence, for its inactivation leads to the formation of trunk-like structures without relying on the otherwise requested extra-cellular matrix analogues such as Matrigel. In contrast, the repression of E-Cadherin by Snai1 is critical for self-organization: Snai1 establishes a cell-specific repressive pace by triggering the repression of a pluripotency-associated transcription program and its chromatin landscape, thus allowing a proper transition from E- to N-Cadherin to occur. Altogether, this work establishes a molecular mechanism that integrates the exit from pluripotency and the pace of cell differentiation, leading to the observed self-organizing potential of gastruloids.
Project description:Gastruloids have recently emerged as an efficient four-dimensional model for studying some aspects of post-implantation embryonic patterning. Their ability to undergo gastrulation-like processes leading to the self-organization into highly reproducible biological objects, has opened new avenues to investigate early embryonic patterning. Here, we sought to uncover the molecular and cellular mechanism underlying this remarkable property. We report that self-organization competence is associated with a cell-specific coordination of a Cadherin switch. We find that N-Cadherin hinders gastruloids morphogenetic competence, for its inactivation leads to the formation of trunk-like structures without relying on the otherwise requested extra-cellular matrix analogues such as Matrigel. In contrast, the repression of E-Cadherin by Snai1 is critical for self-organization: Snai1 establishes a cell-specific repressive pace by triggering the repression of a pluripotency-associated transcription program and its chromatin landscape, thus allowing a proper transition from E- to N-Cadherin to occur. Altogether, this work establishes a molecular mechanism that integrates the exit from pluripotency and the pace of cell differentiation, leading to the observed self-organizing potential of gastruloids.
Project description:Integrated in vitro models of human organogenesis are needed to elucidate the multi-systemic events underlying development and disease. We report the generation of human trunk-like structures that model the co-morphogenesis, patterning, and differentiation of the human spine and spinal cord. We identified differentiation conditions for human pluripotent stem cells favoring the formation of an embryo-like extending antero-posterior (AP) axis. Single cell and spatial transcriptomics show that somitic and spinal cord differentiation trajectories organize along this axis and can self-assemble into neural tubes surrounded by somites upon extracellular matrix addition. Morphogenesis is coupled with AP patterning mechanisms which results, at later stages of organogenesis, in in vivo-like arrays of neural subtypes along a neural tube surrounded by spine and muscle progenitors contacted by neuronal projections. This integrated system of trunk development indicates that in vivo-like multi-tissue morphogenesis and topographic organization of terminal cell types can be achieved in human organoids, opening windows for the development of more complex models of organogenesis.
Project description:Integrated in vitro models of human organogenesis are needed to elucidate the multi-systemic events underlying development and disease. We report the generation of human trunk-like structures that model the co-morphogenesis, patterning, and differentiation of the human spine and spinal cord. We identified differentiation conditions for human pluripotent stem cells favoring the formation of an embryo-like extending antero-posterior (AP) axis. Single cell and spatial transcriptomics show that somitic and spinal cord differentiation trajectories organize along this axis and can self-assemble into neural tubes surrounded by somites upon extracellular matrix addition. Morphogenesis is coupled with AP patterning mechanisms which results, at later stages of organogenesis, in in vivo-like arrays of neural subtypes along a neural tube surrounded by spine and muscle progenitors contacted by neuronal projections. This integrated system of trunk development indicates that in vivo-like multi-tissue morphogenesis and topographic organization of terminal cell types can be achieved in human organoids, opening windows for the development of more complex models of organogenesis.