A Novel Human Pluripotent Stem Cell-Derived Neural Crest Model of Treacher Collins Syndrome Shows Defects in Cell Death and Migration.

Serrano, Felipe; Bernard, William George; Granata, Alessandra; Iyer, Dharini; Steventon, Ben; Kim, Matthew; Vallier, Ludovic; Gambardella, Laure; Sinha, Sanjay

Serrano, Felipe; Bernard, William George; Granata, Alessandra; Iyer, Dharini; Steventon, Ben; Kim, Matthew; Vallier, Ludovic; Gambardella, Laure; Sinha, Sanjay.

Afiliación

Serrano F; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.
Bernard WG; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.
Granata A; 2 Division of Clinical Neurosciences, Clifford Allbutt Building, Cambridge Biomedical Campus, Cambridge, United Kingdom.
Iyer D; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.
Steventon B; 3 Department of Genetics, University of Cambridge, Cambridge, United Kingdom.
Kim M; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.
Vallier L; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.
Gambardella L; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.
Sinha S; 1 Anne McLaren Laboratory, Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Department of Medicine, University of Cambridge, Cambridge, United Kingdom.

Stem Cells Dev ; 28(2): 81-100, 2019 01 15.

Article en En | MEDLINE | ID: mdl-30375284

RESUMEN

The neural crest (NC) is a transient multipotent cell population present during embryonic development. The NC can give rise to multiple cell types and is involved in a number of different diseases. Therefore, the development of new strategies to model NC in vitro enables investigations into the mechanisms involved in NC development and disease. In this study, we report a simple and efficient protocol to differentiate human pluripotent stem cells (HPSC) into NC using a chemically defined media, with basic fibroblast growth factor 2 (FGF2) and the transforming growth factor-ß inhibitor SB-431542. The cell population generated expresses a range of NC markers, including P75, TWIST1, SOX10, and TFAP2A. NC purification was achieved in vitro through serial passaging of the population, recreating the developmental stages of NC differentiation. The generated NC cells are highly proliferative, capable of differentiating to their derivatives in vitro and engraft in vivo to NC specific locations. In addition, these cells could be frozen for storage and thawed with no loss of NC properties, nor the ability to generate cellular derivatives. We assessed the potential of the derived NC population to model the neurocristopathy, Treacher Collins Syndrome (TCS), using small interfering RNA (siRNA) knockdown of TCOF1 and by creating different TCOF1+/- HPSC lines through CRISPR/Cas9 technology. The NC cells derived from TCOF1+/- HPSC recapitulate the phenotype of the reported TCS murine model. We also report for the first time an impairment of migration in TCOF1+/- NC and mesenchymal stem cells. In conclusion, the developed protocol permits the generation of the large number of NC cells required for developmental studies, disease modeling, and for drug discovery platforms in vitro.

Asunto(s)

Diferenciación Celular; Técnicas de Reprogramación Celular/métodos; Disostosis Mandibulofacial/genética; Cresta Neural/citología; Células Madre Pluripotentes/citología; Proteínas Adaptadoras Transductoras de Señales/genética; Proteínas Adaptadoras Transductoras de Señales/metabolismo; Animales; Benzamidas/farmacología; Muerte Celular; Movimiento Celular; Embrión de Pollo; Dioxoles/farmacología; Factor 2 de Crecimiento de Fibroblastos/farmacología; Humanos; Disostosis Mandibulofacial/patología; Cresta Neural/metabolismo; Células-Madre Neurales/citología; Células-Madre Neurales/metabolismo; Proteínas Nucleares/genética; Proteínas Nucleares/metabolismo; Fosfoproteínas/genética; Células Madre Pluripotentes/efectos de los fármacos; Células Madre Pluripotentes/metabolismo; Factores de Transcripción SOXE/genética; Factores de Transcripción SOXE/metabolismo; Factor de Transcripción AP-2/genética; Factor de Transcripción AP-2/metabolismo; Factores de Transcripción/genética; Factores de Transcripción/metabolismo; Proteína 1 Relacionada con Twist/genética; Proteína 1 Relacionada con Twist/metabolismo

Palabras clave

Treacher Collins syndrome; differentiation; disease modeling; human pluripotent stem cells; neural crest

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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Diferenciación Celular / Células Madre Pluripotentes / Técnicas de Reprogramación Celular / Disostosis Mandibulofacial / Cresta Neural Tipo de estudio: Prognostic_studies Límite: Animals / Humans Idioma: En Revista: Stem Cells Dev Asunto de la revista: HEMATOLOGIA Año: 2019 Tipo del documento: Article País de afiliación: Reino Unido Pais de publicación: Estados Unidos

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