Newly Emergent Apelin Expressing Endothelial Stem-like Cells Orchestrate Lung Microvascular Repair

Godoy, Rafael Soares; Cook, David P; Cober, Nicholas D; Deng, Yupu; Wang, Liyuan; Chakravarti, Ananya; Rowe, Katelynn; Stewart, Duncan J

Rational Endothelial damage plays a central role in acute lung injury, and regeneration of lung vascular endothelium is required for its resolution in preclinical models. Objectives We sought to define the cellular and molecular mechanisms underlying lung microvascular regeneration in acute lung injury induced by lung endothelial cell ablation. Methods Transgenic mice were created expressing endothelial-targeted human diphtheria toxin receptor. Changes in lung cell populations and gene expression profiles were determined using single-cell RNA sequencing of dissociated lung cells (10x Genomics) at baseline (day 0) and days 3, 5 and 7 days after lung endothelial cell ablation. Measurements and Main Results Intratracheal instillation of diphtheria toxin resulted in ablation of ∼70% of lung endothelial cells, producing severe acute lung injury, with complete resolution by 7 days. Single cell analysis revealed 8 distinct endothelial cell clusters, including type-A capillary endothelial cells which were characterized by the unique expression of apelin at baseline. Diphtheria toxin-induced ablation resulted in the emergence of novel stem-like endothelial cells in the transitional ‘general’ capillary type-B endothelial population at day 3, characterized by the de novo expression of apelin. This was followed by the appearance of proliferative endothelial cells at day 5 expressing apelin receptor and Forkhead box M1 which were responsible for replenishment of all depleted endothelial cell populations. Treatment with an apelin receptor antagonist prevented recovery post DT resulting in excessive mortality. Conclusions Targeted endothelial cell ablation revealed a remarkable regenerative capacity of the lung microvasculature orchestrated by newly emergent apelin-expressing endothelial stem-like cells primed for endothelial repair.