In Vitro Differentiation of Human Skin-Derived Cells into Functional Sensory Neurons-Like.

Adeline Bataille,Jean-Luc Carré,Thierry Oddos,Emmanuelle Plée-Gautier,Killian L’Hérondelle,Mehdi Sakka,Laurent Misery,Raphael Leschiera,Nicolas Lebonvallet,Cecilia Brun,Jean-Pierre Pennec,Nelig Le Goux,Olivier Mignen

doi:10.3390/cells9041000

Abstract

Skin-derived precursor cells (SKPs) are neural crest stem cells that persist in certain adult tissues, particularly in the skin. They can generate a large type of cell in vitro, including neurons. SKPs were induced to differentiate into sensory neurons (SNs) by molecules that were previously shown to be important for the generation of SNs: purmorphamine, CHIR99021, BMP4, GDNF, BDNF, and NGF. We showed that the differentiation of SKPs induced the upregulation of neurogenins. At the end of the differentiation protocol, transcriptional analysis was performed on BRN3A and a marker of pain-sensing nerve cell PRDM12 genes: 1000 times higher for PRDM12 and 2500 times higher for BRN3A in differentiated cells than they were in undifferentiated SKPs. Using immunostaining, we showed that 65% and 80% of cells expressed peripheral neuron markers BRN3A and PERIPHERIN, respectively. Furthermore, differentiated cells expressed TRPV1, PAR2, TRPA1, substance P, CGRP, HR1. Using calcium imaging, we observed that a proportion of cells responded to histamine, SLIGKV (a specific agonist of PAR2), polygodial (a specific agonist of TRPA1), and capsaicin (a specific agonist of TRPV1). In conclusion, SKPs are able to differentiate directly into functional SNs. These differentiated cells will be very useful for further in vitro studies.

Highlights

Many studies have described the possibility of producing in vitro sensory neurons (SNs) from stem cells
We provided many arguments showing the differentiation of skin-derived precursor cells (SKPs) into neuronal cells, such as the expression of canonical peripheral markers, such as BRN3A, PERIPHERIN, and PRDM12, in SNs derived from SKPs
Our results show a greater increase in NGN1 and 2 in neural crest stem cells (NCSCs) differentiation, which suggests the possibility of SKP to express NGN after neuronal induction

Summary

Introduction

Many studies have described the possibility of producing in vitro sensory neurons (SNs) from stem cells. Several studies have demonstrated that a wide variety of cell types could be derived from SKPs, such as cells expressing neuronal, glial, osteoblastic, smooth muscle, chondrocytic, and melanocytic lineage markers [12,13,14,15,16,17]. They express some markers common to neural crest stem cells (NCSCs), such as the progenitor neural NESTIN [14], the low-affinity neurotrophin receptor p75 (P75NTR) and transcription factors, such as SOX9, SOX10, PAX3, SLUG, and SNAIL [14,15]. The undifferentiated state of NCSCs is maintained by the control of two pathways involving Wnt and BMP in an endogenous niche, which is the hair follicle [18]

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