Background:
The cellular diversity of the lung endothelium has not been systematically characterized in humans. Here, we provide a reference atlas of human lung endothelial cells (ECs) to facilitate a better understanding of the phenotypic diversity and composition of cells comprising the lung endothelium.
Methods:
We reprocessed human control single cell RNA sequencing (scRNAseq) data from six datasets. EC populations were characterized through iterative clustering with subsequent differential expression analysis. Marker genes were validated by fluorescent microscopy and
in situ
hybridization. scRNAseq of primary lung ECs cultured
in-vitro
was performed. The signaling network between different lung cell types was studied. For cross species analysis or disease relevance, we applied the same methods to scRNAseq data obtained from mouse lungs or from human lungs with pulmonary hypertension.
Results:
Six lung scRNAseq datasets were reanalyzed and annotated to identify over 15,000 vascular EC cells from 73 individuals. Differential expression analysis of EC revealed signatures corresponding to endothelial lineage, including pan-endothelial, pan-vascular and subpopulation-specific marker gene sets. Beyond the broad cellular categories of lymphatic, capillary, arterial and venous ECs, we found previously indistinguishable subpopulations: among venous EC, we identified two previously indistinguishable populations, pulmonary-venous ECs (COL15A1neg) localized to the lung parenchyma and systemic-venous ECs (COL15A1pos) localized to the airways and the visceral pleura; among capillary EC, we confirmed their subclassification into recently discovered aerocytes characterized by EDNRB, SOSTDC1 and TBX2 and general capillary EC. We confirmed that all six endothelial cell types, including the systemic-venous EC and aerocytes, are present in mice and identified endothelial marker genes conserved in humans and mice. Ligand-receptor connectome analysis revealed important homeostatic crosstalk of EC with other lung resident cell types. scRNAseq of commercially available primary lung ECs demonstrated a loss of their native lung phenotype in culture. scRNAseq revealed that the endothelial diversity is maintained in pulmonary hypertension. Our manuscript is accompanied
by an online data mining tool (www.LungEndothelialCellAtlas.com).
Conclusions:
Our integrated analysis provides the comprehensive and well-crafted reference atlas of lung endothelial cells in the normal lung and confirms and describes in detail previously unrecognized endothelial populations across a large number of humans and mice.
Blood outgrowth endothelial cells overexpressing eNOS mitigate pulmonary hypertension in rats: a unique carrier cell enabling autologous cell-based gene therapy
