Abstract
We used single cell sequencing technology to characterize the transcriptomes of 1,174 human embryonic stem cell-derived retinal ganglion cells (RGCs) at the single cell level. The human embryonic stem cell line BRN3B-mCherry (A81-H7), was differentiated to RGCs using a guided differentiation approach. Cells were harvested at day 36 and prepared for single cell RNA sequencing. Our data indicates the presence of three distinct subpopulations of cells, with various degrees of maturity. One cluster of 288 cells showed increased expression of genes involved in axon guidance together with semaphorin interactions, cell-extracellular matrix interactions and ECM proteoglycans, suggestive of a more mature RGC phenotype.
Highlights
Since the isolation of embryonic stem cells (ESCs)[1,2,3,4] and generation of induced pluripotent stem cells[5,6], pluripotent stem cells (PSCs) have made a tremendous contribution towards improving our understanding of mechanisms involved in development and disease
PSC-derived progeny allow the investigation of diseaseaffected cell types that are not readily accessible due to their anatomical location, such as retinal cells[7,8,9]
By sequencing large number of single cells from an individual ‘sample’ it is possible to dissect the cellular composition of apparently homogenous tissues or cell cultures11–13. sc-RNAseq opens the possibility of examining rare cell populations that could not otherwise be resolved using bulk RNA sequencing (RNA-seq), and further characterising well-known cell types, for example oligodendrocytes[14] or sensory neurons[15]
Summary
Since the isolation of embryonic stem cells (ESCs)[1,2,3,4] and generation of induced pluripotent stem cells (iPSCs)[5,6], pluripotent stem cells (PSCs) have made a tremendous contribution towards improving our understanding of mechanisms involved in development and disease. PSC-derived progeny allow the investigation of diseaseaffected cell types that are not readily accessible due to their anatomical location, such as retinal cells[7,8,9] Utilising such disease-affected cells will significantly improve the drug development pipeline through efficacy profiling and side effect or toxicity assessment[10]. The development of RNA sequencing (RNA-seq) technology has allowed for the rapid quantification of individual gene transcripts Integrating this high-throughput data with computational and statistical methods provides a toolbox to study the molecular functions of human tissues. Stages across a cell lineage can be distinguished by their unique transcriptional signature[16] This technology has been used in cell culture, in particular with PSCs, their differentiated progeny and organoids, including of the nervous system[17,18], as a way to distinctively characterize cellular subpopulations. We present a dataset of scRNA-seq to characterize the transcriptome of RGCs derived from human ESCs (hESCs) at a single cell level
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