Changbin Sun1,2,3, Hailun Wang4, Qiwang Ma1,3, Chao Chen1,3, Jianhui Yue1,3,5, Bo Li6,7, Xi Zhang8,9. 1. BGI-Shenzhen, Shenzhen, 518083, China. 2. BGI Education Center, University of Chinese Academy of Sciences, Shenzhen, 518083, China. 3. China National GeneBank, BGI-Shenzhen, Shenzhen, 518082, China. 4. Department of Radiation Oncology, School of Medicine, Johns Hopkins University, Baltimore, MD, 21218, USA. 5. Section of Cell Biology and Physiology, Department of Biology, University of Copenhagen, Copenhagen, Denmark. 6. BGI-Shenzhen, Shenzhen, 518083, China. libo@genomics.cn. 7. China National GeneBank, BGI-Shenzhen, Shenzhen, 518082, China. libo@genomics.cn. 8. BGI-Shenzhen, Shenzhen, 518083, China. zhangxi1@genomics.cn. 9. China National GeneBank, BGI-Shenzhen, Shenzhen, 518082, China. zhangxi1@genomics.cn.
Abstract
BACKGROUND: Human pluripotent stem cell-derived limbal stem cells (hPSC-derived LSCs) provide a promising cell source for corneal transplants and ocular surface reconstruction. Although recent efforts in the identification of LSC markers have increased our understanding of the biology of LSCs, much more remains to be characterized in the developmental origin, cell fate determination, and identity of human LSCs. The lack of knowledge hindered the establishment of efficient differentiation protocols for generating hPSC-derived LSCs and held back their clinical application. RESULTS: Here, we performed a time-course single-cell RNA-seq to investigate transcriptional heterogeneity and expression changes of LSCs derived from human embryonic stem cells (hESCs). Based on current protocol, expression heterogeneity of reported LSC markers were identified in subpopulations of differentiated cells. EMT has been shown to occur during differentiation process, which could possibly result in generation of untargeted cells. Pseudotime trajectory analysis revealed transcriptional changes and signatures of commitment of hESCs-derived LSCs and their progeny-the transit amplifying cells. CONCLUSION: Single-cell RNA-seq revealed time-course expression changes and significant transcriptional heterogeneity during hESC-derived LSC differentiation in vitro. Our results demonstrated candidate developmental trajectory and several new candidate markers for LSCs, which could facilitate elucidating the identity and developmental origin of human LSCs in vivo.
BACKGROUND:Human pluripotent stem cell-derived limbal stem cells (hPSC-derived LSCs) provide a promising cell source for corneal transplants and ocular surface reconstruction. Although recent efforts in the identification of LSC markers have increased our understanding of the biology of LSCs, much more remains to be characterized in the developmental origin, cell fate determination, and identity of human LSCs. The lack of knowledge hindered the establishment of efficient differentiation protocols for generating hPSC-derived LSCs and held back their clinical application. RESULTS: Here, we performed a time-course single-cell RNA-seq to investigate transcriptional heterogeneity and expression changes of LSCs derived from human embryonic stem cells (hESCs). Based on current protocol, expression heterogeneity of reported LSC markers were identified in subpopulations of differentiated cells. EMT has been shown to occur during differentiation process, which could possibly result in generation of untargeted cells. Pseudotime trajectory analysis revealed transcriptional changes and signatures of commitment of hESCs-derived LSCs and their progeny-the transit amplifying cells. CONCLUSION: Single-cell RNA-seq revealed time-course expression changes and significant transcriptional heterogeneity during hESC-derived LSC differentiation in vitro. Our results demonstrated candidate developmental trajectory and several new candidate markers for LSCs, which could facilitate elucidating the identity and developmental origin of human LSCs in vivo.
Authors: Feng Zhen Chen; Li Jin You; Fan Yang; Li Na Wang; Xue Qin Guo; Fei Gao; Cong Hua; Cong Tan; Lin Fang; Ri Qiang Shan; Wen Jun Zeng; Bo Wang; Ren Wang; Xun Xu; Xiao Feng Wei Journal: Yi Chuan Date: 2020-08-20