Human embryonic stem cell differentiation toward regional specific neural precursors

Abstract

Human embryonic stem cells (hESCs) are self-renewing pluripotent cells that have the capacity to differentiate into a wide variety of cell types. This potentiality represents a promising source to overcome many human diseases by providing an unlimited supply of all cell types, including cells with neural characteristics. Therefore, this review summarizes early neural development and the potential of hESCs to differentiate under in vitro conditions, examining at the same time the potential use of differentiated hESCs for therapeutic applications for neural tissue and cell regeneration.

Figure 1

Steps of different protocols to induce neural differentiation of hESCs. hESCs can be differentiated into neuronal lineages using EBs. Prolonged treatment of EBs with EGF and bFGF in appropriate culture conditions results in the derivation of OPCs that can be used to repair injured spinal cord. Treatment of EBs with RA generates neuroectodermal cellular formations called rosettes. Cells from rosettes differentiate into motoneuron progenitors if triggered with RA and SHH, and can be a very useful tool in the future treatment of spinal cord injuries. Differentiation toward DA progenitors can be induced with FGF8 and SHH in the early rosette stage. Coculture of hESCs with mesenchymal PA6 cells results in a high yield of DA neurons. Abbreviations: AA, ascorbic acid; BDNF, brain-derived neurotrophic factor; bFGF, basic fibroblast growth factor; DA, dopaminergic; EB, embryoid body; EGF, epidermal growth factor; FGF, fibroblast growth factor; GDNF, glial-derived neurotrophic factor; hESC, human embryonic stem cell; IGF, insulin-like growth factor; OPC, oligodendrocyte progenitor cell; RA, retinoic acid; SHH, sonic hedgehog.

Figure 2

hESC-derived neural progenitors treated with retinoic acid display a spinal cord phenotype. The cells are mostly TUJ1⁺ (green, (A)) and HB9⁺ (red, (B)). Almost all ChAT⁺ cells (green, (D)) are also HB9⁺ cells (red, (E)). Blue indicates DAPI. Scale bars: 50 μm (A–C) and 25 μm (D–F). Abbreviations: ChAT, choline acetyl transferase; DAPI, 4′,6-diamidino-2-phenylindole; hESC, human embryonic stem cell.

Figure 3

Differentiation of hESCs into DA neurons. (A): Early rosettes (marked by asterisks, bright light) and specific staining of regional specific neural precursors generated from hESCs in chemically defined medium conditions. The hESC-derived neural progenitors display a rostral phenotype if they are treated with bFGF only. The cells are TUJ1⁺ (green, a) coexpressed with OTX2⁺ (red, b), and a majority of the TH⁺ cells (green, d) are GABA⁺ (red, e). Scale bars: 25 μm (A) and 50 μm (a–f). Abbreviations: bFGF, basic fibroblast growth factor; DA, dopaminergic; GABA, γ-aminobutyric acid; hESC, human embryonic stem cell; TH, tyrosine hydroxylase.

Figure 4

Potential cell therapy with differentiated human embryonic stem cells: retinal neurons for retinitis pigmentosa, dopaminergic neurons for Parkinson's disease, and motoneurons and oligodendrocytes for spinal cord injury.