RNA-seq of sorted cells at day 7.5 of cardiac differentiation from human embyronic stem cells

The experiment aimed to resolve cellular heterogeneity in cardiac differentiation through the purification of different cell populations by lineage markers and analysis of their transcriptomes and chromatin accessibility. The differentiation protocol was designed to promote cell diversity. The addition of SB at day 2, inhibited SMAD2/3 phosphorylation and created a high BMP signalling bias to restrict cardiac differentiation in favour of other mesodermal lineages, whereas the addition of DMH1 at day 2, inhibited SMAD1/5/8 phosphorylation to create a high Activin signaling bias to promote the co-differentiation of endoderm. Cells were sorted based on SOX17-tomato and NKX2-5-GFP knock-in reporters into their major classes: Pop1 = SB_G0_T0 Pop2 = SB_G1_T0 Pop3 = CTRL_G0_T0 Pop4 = CTRL_G1_T0 Pop5 = DM_G0_T0 Pop6 = DM_G1_T0 Pop7 = DM_G1_T1 Pop8 = DM_G0_T1

本实验旨在通过谱系标志物（lineage markers）纯化不同细胞群，并分析其转录组（transcriptomes）与染色质可及性（chromatin accessibility），以解析心脏分化（cardiac differentiation）过程中的细胞异质性（cellular heterogeneity）。该分化方案旨在促进细胞多样性。于第2日添加SB可抑制SMAD2/3磷酸化，并形成高BMP信号通路偏向性，从而限制心脏分化，偏向其他中胚层谱系（mesodermal lineages）；而于第2日添加DMH1则可抑制SMAD1/5/8磷酸化，形成高Activin信号通路偏向性，以促进内胚层（endoderm）的共分化（co-differentiation）。研究人员基于SOX17-tomato与NKX2-5-GFP敲入报告基因（knock-in reporters），将细胞分为以下主要类别：Pop1 = SB_G0_T0、Pop2 = SB_G1_T0、Pop3 = CTRL_G0_T0、Pop4 = CTRL_G1_T0、Pop5 = DM_G0_T0、Pop6 = DM_G1_T0、Pop7 = DM_G1_T1、Pop8 = DM_G0_T1