Abstract
BackgroundHuman pluripotent stem cells (hPSCs) provide powerful models for studying cellular differentiations and unlimited sources of cells for regenerative medicine. However, a comprehensive single-cell level differentiation roadmap for hPSCs has not been achieved.ResultsWe use high throughput single-cell RNA-sequencing (scRNA-seq), based on optimized microfluidic circuits, to profile early differentiation lineages in the human embryoid body system. We present a cellular-state landscape for hPSC early differentiation that covers multiple cellular lineages, including neural, muscle, endothelial, stromal, liver, and epithelial cells. Through pseudotime analysis, we construct the developmental trajectories of these progenitor cells and reveal the gene expression dynamics in the process of cell differentiation. We further reprogram primed H9 cells into naïve-like H9 cells to study the cellular-state transition process. We find that genes related to hemogenic endothelium development are enriched in naïve-like H9. Functionally, naïve-like H9 show higher potency for differentiation into hematopoietic lineages than primed cells.ConclusionsOur single-cell analysis reveals the cellular-state landscape of hPSC early differentiation, offering new insights that can be harnessed for optimization of differentiation protocols.
Highlights
Human pluripotent stem cells provide powerful models for studying cellular differentiations and unlimited sources of cells for regenerative medicine
The pre-implantation mouse epiblasts obtained from blastocysts have the ground-state naïve pluripotency that can be recapitulated in vitro in the form of embryonic stem cells (ESCs) [8, 9]
Results scRNA-seq analysis of Human pluripotent stem cells (hPSCs) and embryoid body (EB) In order to systematically map hPSCs early differentiation pathways, Naïve-like H9, Primed H9, and EBs were prepared as single-cell samples for sequencing using Fluidigm C1 system with high-throughput integrated fluidics circuits (HT IFCs) (Fig. 1a)
Summary
Human pluripotent stem cells (hPSCs) provide powerful models for studying cellular differentiations and unlimited sources of cells for regenerative medicine. HPSCs have the capacity of self-renewal and multilineage differentiation both in vitro and in vivo. These features of hPSCs have provided remarkable promise in developmental biology and regenerative medicine [2]. The pre-implantation mouse epiblasts obtained from blastocysts have the ground-state naïve pluripotency that can be recapitulated in vitro in the form of embryonic stem cells (ESCs) [8, 9]. These two states of pluripotent stem cells (i.e. ESCs and EpiSCs) are interchangeable under specific conditions [9] The study of this cellularstate transition process will contribute to the understanding of early development from pre-implantation epiblasts to post-implantation epiblasts.
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