468 - Remarkable lung endothelial cell heterogeneity at single-cell resolution in neonatal hyperoxic lung injury

Poster Number: 468
Publication Number: 468.432

Krithika Lingappan, Children's Hospital of Philadelphia, Philadelphia, PA, United States; Manuel E. Cantu Gutierrez, Baylor College of Medicine, Houston, TX, United States; Abiud Cantu Gutierrez, Childrens Hospital of Philadelphia, Houston, TX, United States

Background: Lung endothelial cells are particularly susceptible to the effects of hyperoxia exposure and play a crucial role both during lung development and in the pathogenesis of bronchopulmonary dysplasia (BPD). The impact of neonatal hyperoxic injury during the saccular stage of lung development on the lung endothelium at single-cell resolution was the focus of this study.

Objective: To understand the heterogeneity in the lung endothelial cell subpopulations and their response to neonatal hyperoxic injury during the saccular stage of lung development.

Design/Methods: Lungs from three mice of either sex were analyzed by scRNAseq on PND7 after postnatal hyperoxia exposure (95% FiO2) from PND 1-5 (saccular stage of lung development) with the 10X Genomics platform. After rigorous quality control of the data, computational analysis was performed using Seurat.

Results: Analysis of 35934 single-cell transcriptomes identified 27 distinct epithelial, endothelial, immune and mesenchymal populations. Within the endothelial cells, 7 distinct sub-populations; venous, lymphatic, arterial, gCaP, proliferating gCaP, aCaP, and reactive aCaP were identified (Fig 1A). The most significant impact was noticed with the aCaP (or the aerocyte) capillary endothelial cells which undergo remarkable expansion upon exposure to hyperoxia to form the reactive aCaP cells (Fig 1B) characterized by the increased expression of genes such as Cd93 (localized to filopodia and expressed in tip cells), Ptprb (Vascular endothelial protein tyrosine phosphatase) which regulates vascular integrity by dephosphorylating substrates that are key players in endothelial junction stability, such as Tie-2, VE-cadherin and VEGFR2 and Endothelin 1. We also noticed upregulation of biological pathways related to oxidative phosphorylation, fatty acid metabolism and mTORC1 signaling, whereas pathways related to VEGF signaling and TGF-beta activated receptor activity were downregulated. All the other endothelial cells including the gCaP cells which serve as progenitor endothelial cells in the lung are decreased except for the arterial and lymphatic endothelial cells which are relatively stable (Fig 1C). Conclusion(s): scRNAseq analysis in the murine lung exposed to hyperoxia during the saccular stage of lung development reveals marked heterogeneity in the lung endothelial cells. The reactive aCaP cell population shows a significant expansion after hyperoxia exposure. This may represent the response of the developing lung to injury and may be imperative towards the repair or a step towards the pathogenesis of the disease.
Remarkable heterogeneity in lung endothelial cell response to neonatal hyperoxia exposureFigure 1A: Distinct endothelial cell subpopulations were identified in the neonatal lung at PND 7. Figure 1B: The effect of hyperoxia exposure during the saccular stage of lung development on the lung endothelium at single-cell resolution in the male (M) and female (F) lung. Figure 1C: The impact of hyperoxia on the relative composition of different endothelial cell subpopulations