Neural differentiation, selection and transcriptomic profiling of human neuromesodermal progenitor-like cells in vitro

Robust protocols for directed differentiation of human pluripotent cells are required to determine whether mechanisms operating in model organisms are relevant to our own development. Recent work in vertebrate embryos has identified neuromesodermal progenitors as a bipotent cell population that cont...

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Veröffentlicht in:Development (Cambridge) 2018-07, Vol.145 (16)
Hauptverfasser: Verrier, Laure, Davidson, Lindsay, Gierliński, Marek, Dady, Alwyn, Storey, Kate G
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container_issue 16
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container_title Development (Cambridge)
container_volume 145
creator Verrier, Laure
Davidson, Lindsay
Gierliński, Marek
Dady, Alwyn
Storey, Kate G
description Robust protocols for directed differentiation of human pluripotent cells are required to determine whether mechanisms operating in model organisms are relevant to our own development. Recent work in vertebrate embryos has identified neuromesodermal progenitors as a bipotent cell population that contributes to paraxial mesoderm and spinal cord. However, precise protocols for differentiation of human spinal cord progenitors are lacking. Informed by signalling in amniote embryos, we show here that transient dual-SMAD inhibition, together with retinoic acid (dSMADi-RA), provides rapid and reproducible induction of human spinal cord progenitors from neuromesodermal progenitor-like cells. Using CRISPR-Cas9 to engineer human embryonic stem cells with a GFP-reporter for neuromesodermal progenitor-associated gene we facilitate selection of this cell population. RNA-sequencing was then used to identify human and conserved neuromesodermal progenitor transcriptional signatures, to validate this differentiation protocol and to reveal new pathways/processes in human neural differentiation. This optimised protocol, novel reporter line and transcriptomic data are useful resources with which to dissect molecular mechanisms regulating human spinal cord generation and allow the scaling-up of distinct cell populations for global analyses, including proteomic, biochemical and chromatin interrogation.
doi_str_mv 10.1242/dev.166215
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source MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Alma/SFX Local Collection; Company of Biologists
subjects Animals
Body Patterning - genetics
Cell Differentiation - genetics
Cell Lineage - genetics
Cells, Cultured
Embryo, Mammalian
Gene Expression Profiling
Gene Expression Regulation, Developmental
Human Development
Humans
Mesoderm - cytology
Mesoderm - embryology
Mesoderm - physiology
Mice
Neural Stem Cells - cytology
Neural Stem Cells - physiology
Neurogenesis - genetics
Neurogenesis - physiology
Spinal Cord - cytology
Spinal Cord - physiology
Stem Cells - cytology
Stem Cells - physiology
title Neural differentiation, selection and transcriptomic profiling of human neuromesodermal progenitor-like cells in vitro
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