Single-cell transcriptomics reveals correct developmental dynamics and high-quality midbrain cell types by improved hESC differentiation

• Published in: Stem cell reports Vol. 18; no. 1; pp. 337 - 353
• Main Authors: Nishimura, Kaneyasu, Yang, Shanzheng, Lee, Ka Wai, Ásgrímsdóttir, Emilía Sif, Nikouei, Kasra, Paslawski, Wojciech, Gnodde, Sabine, Lyu, Guochang, Hu, Lijuan, Saltó, Carmen, Svenningsson, Per, Hjerling-Leffler, Jens, Linnarsson, Sten, Arenas, Ernest
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 10-01-2023; Elsevier
• Subjects: Cell Differentiation - genetics; development; dopaminergic neuron; Dopaminergic Neurons - metabolism; embryonic stem cell; function; human; Human Embryonic Stem Cells; Humans; Medicin och hälsovetenskap; Mesencephalon; midbrain; Parkinson's disease; Resource; single-cell RNA sequencing; Transcriptome; single-cell RNA sequencing; development; midbrain; Parkinson's disease; embryonic stem cell; function; dopaminergic neuron; human
• ISSN: 2213-6711; 2213-6711
• DOI: 10.1016/j.stemcr.2022.10.016

Stem cell technologies provide new opportunities for modeling cells in health and disease and for regenerative medicine. In both cases, developmental knowledge and defining the molecular properties and quality of the cell types is essential. In this study, we identify developmental factors important for the differentiation of human embryonic stem cells (hESCs) into functional midbrain dopaminergic (mDA) neurons. We found that laminin-511, and dual canonical and non-canonical WNT activation followed by GSK3β inhibition plus FGF8b, improved midbrain patterning. In addition, neurogenesis and differentiation were enhanced by activation of liver X receptors and inhibition of fibroblast growth factor signaling. Moreover, single-cell RNA-sequencing analysis revealed a developmental dynamics similar to that of the endogenous human ventral midbrain and the emergence of high-quality molecularly defined midbrain cell types, including mDA neurons. Our study identifies novel factors important for human midbrain development and opens the door for a future application of molecularly defined hESC-derived cell types in Parkinson disease.
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•A human midbrain-based protocol improves the generation of functional hESC-mDA neurons•Laminin 511, dual WNT5A-CHIR, and FGF8b enhance midbrain patterning of hESCs•LXR activation and FGF inhibition improve midbrain neurogenesis and differentiation•scRNA-seq revealed high-quality hESC-derived cell types, resembling the human midbrain
In this article, Arenas and colleagues present a new hESC differentiation protocol to generate functional mDA neurons that recapitulates key features of human ventral midbrain development. The function of developmental factors is examined, and the quality of hESC-derived cell types is compared with human midbrain standards by scRNA-seq. Implications for future PD cell replacement therapy are discussed.