Human heart-forming organoids recapitulate early heart and foregut development - single-cell RNA sequencing data

Organoid models of early tissue development have been produced for the intestine, brain, kidney and other organs, but similar approaches for the heart have been lacking. Here we generate complex, highly structured, three-dimensional heart-forming organoids (HFOs) by embedding human pluripotent stem cell aggregates in Matrigel followed by directed cardiac differentiation via biphasic WNT pathway modulation with small molecules. HFOs are composed of a myocardial layer lined by endocardial-like cells and surrounded by septum-transversum-like anlagen; they further contain spatially and molecularly distinct anterior versus posterior foregut endoderm tissues and a vascular network. The architecture of HFOs closely resembles aspects of early native heart anlagen before heart tube formation, which is known to require an interplay with foregut endoderm development. We apply HFOs to study genetic defects in vitro by demonstrating that NKX2.5-knockout HFOs show a phenotype reminiscent of cardiac malformations previously observed in transgenic mice. Overall design: Single-cell RNA sequencing of two HFOs derived from the HES3 NKX2.5-eGFP reporter line on day 13 of differentiation.

目前已针对肠道、大脑、肾脏等器官构建了早期组织发育类器官模型，但心脏领域的同类研究方法仍存在空白。本研究通过将人多能干细胞聚集体包埋于基质胶（Matrigel）中，并借助小分子化合物进行双相WNT通路调控以实现定向心脏分化，成功构建出结构复杂、高度有序的三维心脏形成类器官（heart-forming organoids，以下简称HFOs）。
HFOs由被心内膜样细胞包被的心肌层构成，外层被横隔样原基包裹；此外，其内部还包含空间分布与分子特征均存在显著差异的前、后前肠内胚层组织，以及血管网络。HFOs的组织结构与心脏管形成前的早期天然心脏原基高度相似，而已知心脏管形成过程需要与前肠内胚层发育协同互作。
本研究通过证实NKX2.5基因敲除的HFOs可呈现出与此前在转基因小鼠中观察到的心脏畸形表型类似的特征，将HFOs应用于体外遗传缺陷的研究。
实验整体设计：对分化第13天、源自HES3 NKX2.5-eGFP报告细胞系的2个HFOs开展单细胞RNA测序。