Mass cytometry-based single-cell analysis of human stem cell reprogramming uncovers differential regulation of specific pluripotency markers
Im, I., Son, Y.S, Jung, K.B. et al.Human induced pluripotent stem cells (hiPSCs) are reprogrammed from somatic cells and are regarded as promising cell sources for regenerative medicine and disease research. Recently, techniques for analyses of individual cells, such as single-cell RNA-sequencing and mass cytometry, have been used to understand the stem-cell reprogramming process in the mouse. However, the reprogramming process in hiPSCs remains poorly understood. Here, we used mass cytometry to analyze the expression of pluripotency and cell cycle markers in the reprogramming of human stem cells. We confirmed that during the reprogramming, the main cell population was shifted to intermediate population consisting of neither fibroblasts nor hiPSCs. Detailed population analyses, using computational approaches, including dimensional reduction by spanning-tree progression analysis of density-normalized events (SPADE), PhenoGraph, and diffusion mapping, revealed several distinct cell clusters representing the cells along the reprogramming route. Interestingly, correlation analysis of various markers in hiPSCs revealed that the pluripotency marker TRA-1-60 behaves in different pattern from other pluripotency markers. Furthermore, we found that the expression pattern of another pluripotency marker, octamer-binding protein 4 (OCT4), was distinctive in the pHistone-H3high population (M phase) of the cell cycle. To the best of our knowledge, this is the first mass cytometry-based investigation into human reprogramming and pluripotency. Our analysis elucidates several aspects of hiPSC reprogramming, including several intermediate cell clusters active during the process of reprogramming and distinctive marker expression patterns in hiPSCs.
Im, I., Son, Y.S, Jung, K.B. et al. "Mass cytometry-based single-cell analysis of human stem cell reprogramming uncovers differential regulation of specific pluripotency markers" Journal of Biological Chemistry (2019): doi: 10.1074/jbc.RA119.009061