Identification of a Nkx2-5 dosage-dependent progenitor cell population contributing to the Purkinje fiber network

Abstract

Introduction

Cardiovascular diseases (CVDs) remain the leading cause of death worldwide, particularly in developed countries. Notably, approximately 50% of these deaths occur suddenly, often due to ventricular arrhythmias resulting from defects in the Purkinje fiber (PF) network. PFs are crucial for the rapid conduction of electrical impulses through the ventricles, ensuring the synchronized contraction of the heart. Despite their clinical relevance, the molecular mechanisms governing PF development and maturation are not fully understood. Our group has previously shown that PFs arise from embryonic trabecular myocardium, with their commitment to the PF fate being regulated by high levels of the transcription factor Nkx2-5. Genetic deletion of one copy of Nkx2-5 in mice results in a hypoplastic PF network, likely due to defective PF commitment during ventricular morphogenesis.

Objective

Building on this model, we aim to define the transcriptional landscape that governs PF commitment and maturation, both under normal and pathological conditions.

Method

1) Single cell transcriptomic analysis (scRNA-seq) from embryonic (E)11.5 WT and Nkx2–5 ± hearts. 2) Expression pattern and genetic lineage tracing of Lysozyme M (Lyz2), a novel PF marker.

Results

ScRNA-seq cluster and DEG analysis identifies a trabecular cardiomyocyte (CM) population expressing high levels of Lyz2 in WT but not Nkx2–5 ± E11.5 hearts. Intriguingly, Lyz2 expressing CMs (high Lyz2) co-express a subset of genes required for conductive CM commitment. Time-course lineage tracing analysis using a Lyz2-Cre driver reveals that Lyz2-derived CMs contribute to the VCS in adult WT hearts. In contrast, in Nkx2–5 ± hearts, Lyz2-derived cells only minimally contribute to the hypoplastic PF network.

Conclusion

Our results identify a progenitor cell population within the developing trabeculae that expresses high levels of Lyz2 and contributes to the formation of the VCS in an Nkx2–5-dependent manner.