Single cell analysis of murine fibroblasts identifies neonatal to adult switching that regulates cardiomyocyte maturation

Cardiac maturation lays the foundation for postnatal heart development and disease, yet little is known about the contributions of the microenvironment to cardiomyocyte maturation. By integrating single-cell RNA-sequencing data of mouse hearts at multiple postnatal stages, we construct cellular interactomes and regulatory signaling networks. Here we report switching of fibroblast subtypes from a neonatal to adult state and this drives cardiomyocyte maturation. Molecular and functional maturation of neonatal mouse cardiomyocytes and human embryonic stem cell-derived cardiomyocytes are considerably enhanced upon coculture with corresponding adult cardiac fibroblasts. Further, single-cell analysis of in vivo and in vitro cardiomyocyte maturation trajectories identify highly conserved signaling pathways, pharmacological targeting of which substantially delays cardiomyocyte maturation in postnatal hearts, and markedly enhances cardiomyocyte proliferation and improves cardiac function in infarcted hearts. Together, we identify cardiac fibroblasts as a key constituent in the microenvironment promoting cardiomyocyte maturation, providing insights into how the manipulation of cardiomyocyte maturity may impact on disease development and regeneration. Single cell RNASeq at multiple postnatal stages.

心脏成熟是出生后心脏发育与疾病发生的核心基础，但目前对于微环境（microenvironment）在心肌细胞（cardiomyocyte）成熟过程中的调控作用仍知之甚少。本研究整合多个出生后发育阶段小鼠心脏的单细胞RNA测序（single-cell RNA-sequencing, scRNA-seq）数据，构建了细胞互作组与调控信号网络。本研究揭示成纤维细胞（fibroblast）亚型从新生态向成年态的转换可驱动心肌细胞成熟。将新生小鼠心肌细胞与人类胚胎干细胞诱导心肌细胞（human embryonic stem cell-derived cardiomyocytes, hESC-CMs）与对应成年心脏成纤维细胞共培养后，二者的分子与功能成熟水平均得到显著提升。此外，通过对体内（in vivo）、体外（in vitro）心肌细胞成熟轨迹的单细胞分析，本研究鉴定出高度保守的信号通路；对该通路进行药理学靶向干预，可显著延缓出生后小鼠心脏的心肌细胞成熟过程，并在梗死心脏中大幅促进心肌细胞增殖、改善心脏功能。综上，本研究证实心脏成纤维细胞是促进心肌细胞成熟的微环境关键组成成分，为调控心肌细胞成熟度如何影响疾病发生与组织再生提供了全新视角。本数据集涵盖多个出生后发育阶段的单细胞RNA测序数据。