Directed differentiation of human pluripotent stem cells into epidermal stem and progenitor cells.
Mol Biol Rep
; 48(8): 6213-6222, 2021 Aug.
Article
en En
| MEDLINE
| ID: mdl-34350551
ABSTRACT
BACKGROUND:
Pluripotent stem cells (PSCs) produced by somatic cell reprogramming self-renew in culture and can differentiate into any cell type, representing a powerful tool for disease modeling, drug screening, regenerative medicine, and the discovery of personalized therapies to treat tissue-specific pathologies. We previously reported the directed differentiation of human PSCs into epidermal stem and progenitor cells (ESPCs) and 3D epidermis to model the inherited syndrome Fanconi anemia (FA), wherein epidermal cell-junctional defects discovered using this system were validated in patient populations. Here, we describe in detail the corresponding protocol for generating PSC-derived keratinocytes using a distinct, normal PSC line (209.2 PSC). METHODS ANDRESULTS:
Our approach modifies previous protocols to minimize spontaneous cell death and terminal differentiation, eliminate cell stress-inducing keratinocyte selection steps, and reduce total protocol duration and cost. Independent donor-derived PSC lines were converted into ESPCs through the addition of relevant morphogens and a ROCK inhibitor. Results for the 209.2 PSC line highlight consistencies in 2D and also variable features in 3D epidermis compared to the previously published FA-PSC lines. 209.2 PSC-derived ESPCs exhibited a basal cell phenotype while maintaining the capacity to form epidermal organotypic rafts with morphology consistent with fetal epidermis. Transcriptional analyses demonstrated 209.2 ESPCs express epidermis-selective markers and not early endoderm markers, thus supporting an immature stage of p63+ epidermal development.CONCLUSIONS:
This protocol provides an accelerated path for the generation of human ESPCs and 3D epidermal models to study normal epidermal development and homeostasis, elucidate mechanisms of epidermal disease pathogenesis, and provides a platform for developing personalized therapies.Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Queratinocitos
/
Técnicas de Cultivo de Célula
/
Células Madre Pluripotentes
Tipo de estudio:
Guideline
/
Prognostic_studies
Límite:
Humans
Idioma:
En
Revista:
Mol Biol Rep
Año:
2021
Tipo del documento:
Article
País de afiliación:
Estados Unidos