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. 2020 Apr 29;11:2082. doi: 10.1038/s41467-020-16017-x

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Schematic of the human embryonic stem cell (hESC) differentiation protocol to the endocrine cell stage (EN) and experimental plan for LSD1 inhibition. b Immunofluorescent staining for pancreatic hormones insulin (INS), glucagon (GCG) and somatostatin (SST) or PDX1 and NKX6.1 in control EN cells compared to EN cells with early (LSD1i^early) and late (LSD1i^late) LSD1 inhibition (representative images, n = 10 independent differentiations). Scale bar, 50 µm. c qRT-PCR analysis for INS, GCG and SST in control, LSD1i^early and LSD1i^late EN cells. Data are shown as mean ± S.E.M. (n = 3 replicates from independent differentiations with n = 3 technical replicates per sample; source data are provided as a Source Data file). P = 7.93 e−4, 1.42 e−2, 2.32 e−4, 8.71 e−4, 3.5 e−2, and 1.52 e−3, respectively, Student’s t-test, 2 sided. d Flow cytometry analysis at EN stage for NKX6.1, PDX1 and INS comparing control, LSD1i^early and LSD1i^late cells. Isotype control for each antibody is shown in red and target protein staining in green. Percentage of cells expressing each protein is indicated (representative experiment, n = 2 independent differentiations). D, day; AA, activin A; ITS, insulin-transferrin-selenium; TGFBi, TGFβ R1 kinase inhibitor; KC, KAAD-cyclopamine; KGF, keratinocyte growth factor; RA, retinoic acid; EGF, epidermal growth factor; ES, human embryonic stem cells; DE, definitive endoderm; GT, primitive gut tube; PP1, early pancreatic progenitors; PP2, late pancreatic progenitors; EN, endocrine cell stage; FSC-A, forward scatter area.