Lineage tracing reveals the fate of lung endothelial cells in the progression of pulmonary fibrosis

<bold>Background:</bold> Progressive pulmonary fibrosis is a rapidly advancing, deadly disease caused by uncontrolled scarring. The parenchymal destruction is accompanied by heterogeneous capillarization and vascular remodeling. Yet it remains elusive, whether this is associated with local endothelial cell (EC) adaptations, endothelial to mesenchymal transition and altered proliferation/apoptosis. <bold>Objective:</bold> We aimed to systematically trace EC proliferation, apoptosis and mesenchymal transition in the course of lung fibrosis. <bold>Methods:</bold> Proliferation, apoptosis and mesenchymal transition were investigated in genetically labeled ECs in the bleomycin-induced (3d, 14d) and the Fra-2 overexpressing (8w, 16w) mouse models by FACS, IF and scRNAseq. Results were compared to human samples. <bold>Results:</bold> The proportion of pulmonary CD31⁺ cells was consistent in the mice over time. There was no increase of CD31⁺SMA⁺ cells (%) in fibrotic mouse lungs as compared to controls. The proportion of CD31⁺Ki67⁺ cells was significantly increased 14d post-bleomycin, while CD31⁺Casp3⁺ cells were decreased by trend. No changes of EC proliferation/apoptosis were apparent in Fra-2 transgenic mice. In human fibrotic lungs, the proportion of CD31+ cells was reduced by trend. No changes were observed in CD31⁺SMA⁺, CD31⁺Ki67⁺ and CD31⁺Casp3⁺ populations (%). <bold>Conclusion:</bold> We conclude that EC proliferation might contribute to the rapid fibrosis development in the bleomycin model. In contrast, an impaired regenerative capacity of the endothelium in Fra-2 transgenic mice and in patients may fuel the chronic nature of the disease. Modulation of EC proliferation in-vivo will uncover whether increased angiogenesis ameliorates lung fibrosis.