Cardiac differentiation roadmap for analysis of plasticity and balanced lineage commitment

Stem cell-based models of human heart tissue and cardiac differentiation employ monolayer and 3D organoid cultures with different properties, cell type composition, and maturity. Here we show how cardiac monolayer, embryoid body, and engineered heart tissue trajectories compare in a single-cell roadmap of atrial and ventricular differentiation conditions. Using a multiomic approach and gene-regulatory network inference, we identified regulators of the epicardial, atrial and ventricular cardiomyocyte lineages. We identified ZNF711 as a regulatory switch and safeguard for cardiomyocyte commitment. We show that ZNF711 ablation prevents cardiomyocyte differentiation in the absence of retinoic acid, causing progenitors to be diverted more prominently to epicardial and other lineages. Retinoic acid rescues this shift in lineage commitment and promotes atrial cardiomyocyte differentiation by regulation of shared and complementary target genes, showing an interplay between ZNF711 and retinoic acid in cardiac lineage commitment. Overall design: scRNA-seq experiments used to build an integrated scRoadmap of in vitro cardiac differentiation.

基于干细胞的人类心脏组织模型及心脏分化模型，通常采用单层培养与三维类器官（organoid）培养体系，两类体系在生物学特性、细胞类型组成及成熟度层面均存在差异。本研究基于心房与心室分化条件下的单细胞路线图，对比了心脏单层培养、拟胚体（embryoid body）及工程化心脏组织（engineered heart tissue）的分化轨迹。本研究采用多组学方法与基因调控网络推断技术，鉴定得到心外膜、心房及心室心肌细胞谱系的调控因子。研究发现ZNF711可作为心肌细胞谱系定向的调控开关与保障因子。实验表明，在缺乏视黄酸（retinoic acid）的情况下，敲除ZNF711会阻断心肌细胞分化，使祖细胞更显著地转向心外膜及其他谱系。视黄酸可挽救这一谱系定向偏移，并通过调控共有与互补靶基因，促进心房心肌细胞分化，揭示了ZNF711与视黄酸在心脏谱系定向过程中的相互调控关系。整体实验设计：通过单细胞RNA测序（scRNA-seq）实验，构建体外心脏分化的整合式scRoadmap（单细胞路线图）。