Single-cell transcriptomic reveals molecular diversity and developmental heterogeneity of human stem cell-derived oligodendrocyte lineage cells

Xitiz Chamling,Cynthia A Berlinicke,Hongkai Ji,Katharine A Whartenby,Tza-Huei Wang,Itzy E Morales Pantoja,Liben Chen,Aniruddha Kaushik,Prajwal Devkota,Matthew D Smith,Peter A Calabresi,Zhicheng Ji,Weixiang Fang,Alyssa Kallman,Joseph L Mertz,Donald J Zack,Hai-Quan Mao,Calvin Chang

doi:10.1038/s41467-021-20892-3

Xitiz Chamling, Cynthia A Berlinicke + Show 16 more

Open Access

https://doi.org/10.1038/s41467-021-20892-3

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Abstract

Injury and loss of oligodendrocytes can cause demyelinating diseases such as multiple sclerosis. To improve our understanding of human oligodendrocyte development, which could facilitate development of remyelination-based treatment strategies, here we describe time-course single-cell-transcriptomic analysis of developing human stem cell-derived oligodendrocyte-lineage-cells (hOLLCs). The study includes hOLLCs derived from both genome engineered embryonic stem cell (ESC) reporter cells containing an Identification-and-Purification tag driven by the endogenous PDGFRα promoter and from unmodified induced pluripotent (iPS) cells. Our analysis uncovers substantial transcriptional heterogeneity of PDGFRα-lineage hOLLCs. We discover sub-populations of human oligodendrocyte progenitor cells (hOPCs) including a potential cytokine-responsive hOPC subset, and identify candidate regulatory genes/networks that define the identity of these sub-populations. Pseudotime trajectory analysis defines developmental pathways of oligodendrocytes vs astrocytes from PDGFRα-expressing hOPCs and predicts differentially expressed genes between the two lineages. In addition, pathway enrichment analysis followed by pharmacological intervention of these pathways confirm that mTOR and cholesterol biosynthesis signaling pathways are involved in maturation of oligodendrocytes from hOPCs.

Highlights

Injury and loss of oligodendrocytes can cause demyelinating diseases such as multiple sclerosis
One of the bottlenecks limiting the use of human stem cell-derived oligodendrocyte-lineage-cells (hOLLCs) in developmental, transcriptomic, and drug-discovery studies is the challenge of obtaining sufficient numbers of purified cells—primary human OPCs are rare, difficult to isolate, and cannot be expanded following isolation[11]
The IAP tag consists of a tdTomato fluorescent marker and a mouse cellsurface protein, Thy1.2, separated from each other and from the endogenous PDGFRα gene product by the “ribosome-skipping” 2A peptide[24] (P2A-tdTomato-P2A-Thy1.2) (Fig. 1a)

Summary

Introduction

Injury and loss of oligodendrocytes can cause demyelinating diseases such as multiple sclerosis. Over two hundred human OPC genes are not expressed by mouse OPCs7–9, and expression of Ascl[1], which is essential for OL fate induction from mouse neural precursor cells, is dispensable in human cells[8,10] For both improved disease modeling and to support myelination-based drug-discovery efforts, a more detailed transcriptomic analysis using human oligodendrocyte-lineage cells (hOLLCs) would be helpful. Taking advantage of the bipotential nature of the reporter cells, we performed pseudotime analysis[19] to track the differentiation trajectories of the subsets of OLs and astrocytes This analysis identified genetic factors that are enriched in hOLs or astrocytes, and are potentially involved in regulating human OL vs astrocyte lineage specification

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